BioMed Research International: Biochemistry The latest articles from Hindawi Publishing Corporation © 2015 , Hindawi Publishing Corporation . All rights reserved. ω-3 PUFAs in the Prevention and Cure of Inflammatory, Degenerative, and Neoplastic Diseases 2014 Wed, 05 Aug 2015 06:45:44 +0000 Karsten H. Weylandt, Yong Q. Chen, Kyu Lim, Hui-Min Su, Achille Cittadini, and Gabriella Calviello Copyright © 2015 Karsten H. Weylandt et al. All rights reserved. Heterologous Reconstitution of Omega-3 Polyunsaturated Fatty Acids in Arabidopsis Mon, 03 Aug 2015 11:35:05 +0000 Reconstitution of nonnative, very-long-chain polyunsaturated fatty acid (VLC-PUFA) biosynthetic pathways in Arabidopsis thaliana was undertaken. The introduction of three primary biosynthetic activities to cells requires the stable coexpression of multiple proteins within the same cell. Herein, we report that C22 VLC-PUFAs were synthesized from C18 precursors by reactions catalyzed by -desaturase, an ELOVL5-like enzyme involved in VLC-PUFA elongation, and -desaturase. Coexpression of the corresponding genes (McD6DES, AsELOVL5, and PtD5DES) under the control of the seed-specific vicilin promoter resulted in production of docosapentaenoic acid (22:5 n-3) and docosatetraenoic acid (22:4 n-6) as well as eicosapentaenoic acid (20:5 n-3) and arachidonic acid (20:4 n-6) in Arabidopsis seeds. The contributions of the transgenic enzymes and endogenous fatty acid metabolism were determined. Specifically, the reasonable synthesis of omega-3 stearidonic acid (18:4 n-3) could be a useful tool to obtain a sustainable system for the production of omega-3 fatty acids in seeds of a transgenic T3 line 63-1. The results indicated that coexpression of the three proteins was stable. Therefore, this study suggests that metabolic engineering of oilseed crops to produce VLC-PUFAs is feasible. Sun Hee Kim, Kyung Hee Roh, Jong-Sug Park, Kwang-Soo Kim, Hyun Uk Kim, Kyeong-Ryeol Lee, Han-Chul Kang, and Jong-Bum Kim Copyright © 2015 Sun Hee Kim et al. All rights reserved. Combination of Antiestrogens and Omega-3 Fatty Acids for Breast Cancer Prevention Mon, 03 Aug 2015 11:30:55 +0000 The molecular and biological heterogeneity of human breast cancer emphasizes the importance of a multitargeted approach for effective chemoprevention. Targeting the estrogen receptor pathway alone with the antiestrogens, Tamoxifen and Raloxifene reduces the incidence of estrogen receptor positive tumors but is ineffective against the development of hormone independent cancers. Our preclinical data indicate that the administration of omega-3 fatty acids potentiates the antitumor effects of Tamoxifen by inhibiting multiple proliferative and antiapoptotic pathways, several of which interact with estrogen receptor signaling. The complementarity in the mechanism of antitumor action of Tamoxifen and omega-3 fatty acids is well supported by our signaling, genomic, and proteomic studies. Furthermore, administration of omega-3 fatty acids allows the use of lower and, hence, likely less toxic doses of Tamoxifen. If these findings are supported in the clinical setting, the combination of omega-3 fatty acids and anteistrogens may emerge as a promising, effective, and safe chemopreventive strategy to be tested in a large multi-institutional trial using breast cancer incidence as the primary endpoint. Andrea Manni, Karam El-Bayoumy, Christine G. Skibinski, Henry J. Thompson, Julia Santucci-Pereira, Lucas Tadeu Bidinotto, and Jose Russo Copyright © 2015 Andrea Manni et al. All rights reserved. Omega-3 Polyunsaturated Fatty Acids Intake to Regulate Helicobacter pylori-Associated Gastric Diseases as Nonantimicrobial Dietary Approach Mon, 03 Aug 2015 11:01:42 +0000 Omega-3 polyunsaturated fatty acids (n-3 PUFAs), commonly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been acknowledged as essential long-chain fatty acids imposing either optimal health promotion or the rescuing from chronic inflammatory diseases such as atherosclerosis, fatty liver, and various inflammatory gastrointestinal diseases. Recent studies dealing with EPA and DHA have sparked highest interests because detailed molecular mechanisms had been documented with the identification of its receptor, G protein coupled receptor, and GPR120. In this review article, we have described clear evidences showing that n-3 PUFAs could reduce various Helicobacter pylori- (H. pylori-) associated gastric diseases and extended to play even cancer preventive outcomes including H. pylori-associated gastric cancer by influencing multiple targets, including proliferation, survival, angiogenesis, inflammation, and metastasis. Since our previous studies strongly concluded that nonantimicrobial dietary approach for reducing inflammation, for instance, application of phytoceuticals, probiotics, natural products including Korean red ginseng, and walnut plentiful of n-3 PUFAs, might be prerequisite step for preventing H. pylori-associated gastric cancer as well as facilitating the rejuvenation of precancerous atrophic gastritis, these beneficial lipids can restore or modify inflammation-associated lipid distortion and correction of altered lipid rafts to send right signaling to maintain healthy stomach even after chronic H. pylori infection. Jong-Min Park, Migyeong Jeong, Eun-Hee Kim, Young-Min Han, Sung Hun Kwon, and Ki-Baik Hahm Copyright © 2015 Jong-Min Park et al. All rights reserved. Docosahexaenoic Acid Induces Cell Death in Human Non-Small Cell Lung Cancer Cells by Repressing mTOR via AMPK Activation and PI3K/Akt Inhibition Mon, 03 Aug 2015 09:31:40 +0000 The anticancer properties and mechanism of action of omega-3 polyunsaturated fatty acids (ω3-PUFAs) have been demonstrated in several cancers; however, the mechanism in lung cancer remains unclear. Here, we show that docosahexaenoic acid (DHA), a ω3-PUFA, induced apoptosis and autophagy in non-small cell lung cancer (NSCLC) cells. DHA-induced cell death was accompanied by AMP-activated protein kinase (AMPK) activation and inactivated phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling. Knocking down AMPK and overexpressing Akt increased mTOR activity and attenuated DHA-induced cell death, suggesting that DHA induces cell death via AMPK- and Akt-regulated mTOR inactivation. This was confirmed in Fat-1 transgenic mice, which produce ω3-PUFAs. Lewis lung cancer (LLC) tumor cells implanted into Fat-1 mice showed slower growth, lower phospho-Akt levels, and higher levels of apoptosis and autophagy than cells implanted into wild-type mice. Taken together, these data suggest that DHA-induced apoptosis and autophagy in NSCLC cells are associated with AMPK activation and PI3K/Akt inhibition, which in turn lead to suppression of mTOR; thus ω3-PUFAs may be utilized as potential therapeutic agents for NSCLC treatment. Nayeong Kim, Soyeon Jeong, Kaipeng Jing, Soyeon Shin, Soyeon Kim, Jun-Young Heo, Gi-Ryang Kweon, Seung-Kiel Park, Tong Wu, Jong-Il Park, and Kyu Lim Copyright © 2015 Nayeong Kim et al. All rights reserved. The Pharmacokinetic Profile of a New Gastroresistant Capsule Preparation of Eicosapentaenoic Acid as the Free Fatty Acid Mon, 03 Aug 2015 08:25:42 +0000 Supplementation with n-3 polyunsaturated fatty acids (n-3 PUFAs) may be beneficial for patients with inflammatory bowel diseases (IBD). In this study we analyzed the pharmacokinetic profile of eicosapentaenoic acid (EPA), as the free fatty acid (FFA), in an enteric-coated preparation, in 10 ulcerative colitis (UC) and 10 Crohn’s disease (CD) patients and 15 healthy volunteers (HV). Subjects received 2 g daily of EPA-FFA for 8 weeks. Plasma phospholipid and red blood cell (RBC) membrane fatty acid content were measured by gas chromatography-mass spectrometry. There was a rapid incorporation of EPA into plasma phospholipids by 2 weeks and a slower, but highly consistent, incorporation into RBC membranes (4% total fatty acid content; coefficient of variation 10–16%). There was a concomitant reduction in relative n-6 PUFA content. Elongation and desaturation of EPA into docosahexaenoic acid (DHA) via docosapentaenoic acid (DPA) were apparent and DHA content also increased in membranes. EPA-FFA is well tolerated and no difference in the pharmacokinetic profile of n-3 PUFA incorporation was detected between IBD patients and HV. Our data support the concept that EPA can be considered the “universal donor” with respect to key n-3 PUFAs and that this enteric-coated formulation allows long term treatment with a high level of compliance. Eleonora Scaioli, Carla Cardamone, Elisa Liverani, Alessandra Munarini, Mark A. Hull, and Andrea Belluzzi Copyright © 2015 Eleonora Scaioli et al. All rights reserved. Effect of Dietary ω-3 Polyunsaturated Fatty Acid DHA on Glycolytic Enzymes and Warburg Phenotypes in Cancer Mon, 03 Aug 2015 08:08:13 +0000 The omega-3 polyunsaturated fatty acids (ω-3 PUFAs) are a class of lipids that has been shown to have beneficial effects on some chronic degenerative diseases such as cardiovascular diseases, rheumatoid arthritis, inflammatory disorders, diabetes, and cancer. Among ω-3 polyunsaturated fatty acids (PUFAs), docosahexaenoic acid (DHA) has received particular attention for its antiproliferative, proapoptotic, antiangiogenetic, anti-invasion, and antimetastatic properties, even though the involved molecular mechanisms are not well understood. Recently, some in vitro studies showed that DHA promotes the inhibition of glycolytic enzymes and the Warburg phenotype. For example, it was shown that in breast cancer cell lines the modulation of bioenergetic functions is due to the capacity of DHA to activate the AMPK signalling and negatively regulate the HIF-1α functions. Taking into account these considerations, this review is focused on current knowledge concerning the role of DHA in interfering with cancer cell metabolism; this could be considered a further mechanism by which DHA inhibits cancer cell survival and progression. Laura Manzi, Lara Costantini, Romina Molinari, and Nicolò Merendino Copyright © 2015 Laura Manzi et al. All rights reserved. Biological Roles of Resolvins and Related Substances in the Resolution of Pain Mon, 03 Aug 2015 08:04:48 +0000 Endogenous pain-inhibitory substances have rarely been found. A group of powerful pain suppressor molecules that are endogenously generated are now emerging: resolvins and related compounds including neuroprotectins and maresins. These molecules began to be unveiled in a series of inflammation studies more than a decade ago, rapidly shifting the paradigm that explains the mechanism for the inflammatory phase switch. The resolution phase was considered a passive process as proinflammatory mediators disappeared; it is now understood to be actively drawn by the actions of resolvins. Surprisingly, these substances potently affect the pain state. Although this research area is not fully matured, consistently beneficial outcomes have been observed in a various in vivo and in vitro pain models. Furthermore, multiple hypotheses on the neuronal and molecular mechanisms for alleviating pain are being tested, deriving inspiration from existing inflammation and pain studies. This paper serves as a brief summary of the proresolving roles of resolvins and related lipid mediators in inflammation and also as a review for accumulated information of their painkilling actions. This also includes potential receptor-mediated mechanisms and discusses future scientific perspectives. Further diverse approaches will help to construct a hidden axis of natural protection principles and establish proofs of concept for pain relief. Ji Yeon Lim, Chul-Kyu Park, and Sun Wook Hwang Copyright © 2015 Ji Yeon Lim et al. All rights reserved. The Role of Omega-3 Polyunsaturated Fatty Acids in the Treatment of Patients with Acute Respiratory Distress Syndrome: A Clinical Review Mon, 03 Aug 2015 08:03:38 +0000 Acute respiratory distress syndrome (ARDS) is defined as the acute onset of noncardiogenic edema and subsequent gas-exchange impairment due to a severe inflammatory process. Recent report on the prognostic value of eicosanoids in patients with ARDS suggests that modulating the inflammatory response through the use of polyunsaturated fatty acids may be a useful strategy for ARDS treatment. The use of enteral diets enriched with eicosapentaenoic acid (EPA) and gamma-linolenic acid (GLA) has reported promising results, showing an improvement in respiratory variables and haemodynamics. However, the interpretation of the studies is limited by their heterogeneity and methodology and the effect of ω-3 fatty acid-enriched lipid emulsion or enteral diets on patients with ARDS remains unclear. Therefore, the routine use of ω-3 fatty acid-enriched nutrition cannot be recommended and further large, homogeneous, and high-quality clinical trials need to be conducted to clarify the effectiveness of ω-3 polyunsaturated fatty acids. M. García de Acilu, S. Leal, B. Caralt, O. Roca, J. Sabater, and J. R. Masclans Copyright © 2015 M. García de Acilu et al. All rights reserved. Endogenous Generation and Signaling Actions of Omega-3 Fatty Acid Electrophilic Derivatives Mon, 03 Aug 2015 06:54:10 +0000 Dietary omega-3 polyunsaturated fatty acids (PUFAs) are beneficial for a number of conditions ranging from cardiovascular disease to chronic airways disorders, neurodegeneration, and cancer. Growing evidence has shown that bioactive oxygenated derivatives are responsible for transducing these salutary effects. Electrophilic oxo-derivatives of omega-3 PUFAs represent a class of oxidized derivatives that can be generated via enzymatic and nonenzymatic pathways. Inflammation and oxidative stress favor the formation of these signaling species to promote the resolution of inflammation within a fine autoregulatory loop. Endogenous generation of electrophilic oxo-derivatives of omega-3 PUFAs has been observed in in vitro and ex vivo human models and dietary supplementation of omega-3 PUFAs has been reported to increase their formation. Due to the presence of an α,β-unsaturated ketone moiety, these compounds covalently and reversibly react with nucleophilic residues on target proteins triggering the activation of cytoprotective pathways, including the Nrf2 antioxidant response, the heat shock response, and the peroxisome proliferator activated receptor γ (PPARγ) and suppressing the NF-κB proinflammatory pathway. The endogenous nature of electrophilic oxo-derivatives of omega-3 PUFAs combined with their ability to simultaneously activate multiple cytoprotective pathways has made these compounds attractive for the development of new therapies for the treatment of chronic disorders and acute events characterized by inflammation and oxidative stress. Chiara Cipollina Copyright © 2015 Chiara Cipollina. All rights reserved. N-3 Polyunsaturated Fatty Acids and Inflammation in Obesity: Local Effect and Systemic Benefit Mon, 03 Aug 2015 06:40:54 +0000 Overwhelming consensus emerges among countless evidences that obesity is characterized by a chronic low-grade inflammation in the adipose tissue (AT), which subsequently develops into a systemic inflammatory state contributing to obesity-associated diseases. N-3 Polyunsaturated fatty acids (n-3 PUFA), known as important modulators participating in inflammatory process, turn out to be an effective mitigating strategy dealing with local and systemic inflammation observed in obesity. Some of the effects of n-3 PUFA are brought about by regulation of gene expression through interacting with nuclear receptors and transcription factors; other effects are elicited by modulation of the amount and type of mediator derived from PUFAs. The metabolic effects of n-3 PUFA mainly result from their interactions with several organ systems, not limited to AT. Notably, the attenuation of inflammation in hard-hit AT, in turn, contributes to reducing circulating concentrations of proinflammatory cytokines and detrimental metabolic derivatives, which is beneficial for the function of other involved organs. The present review highlights a bridging mechanism between n-3 PUFA-mediated inflammation relief in AT and systemic benefits. Yue Wang and Feiruo Huang Copyright © 2015 Yue Wang and Feiruo Huang. All rights reserved. Effect of Docosahexaenoic Acid on Apoptosis and Proliferation in the Placenta: Preliminary Report Mon, 03 Aug 2015 06:19:15 +0000 Introduction. Observational studies confirm a higher incidence of preeclampsia in patients with low erythrocyte concentrations of omega-3 fatty acids. Observations point to an association of disorders of pregnancy, such as intrauterine growth restriction (IUGR) and preeclampsia, with excessive apoptosis. One potential mechanism of action of docosahexaenoic acid (DHA) promoting a reduction in the risk of pathological pregnancy may be by influencing these processes in the placenta. Materials and Methods. We investigated 28 pregnant women supplemented with a fish oil product containing 300 mg DHA starting from pregnancy week 20 until delivery (DHA group). The control group consisted of 50 women who did not receive such supplementation (control group). We determined the expression of Ki-67 and p21 as markers of proliferation and caspase 3 activity as a marker of apoptosis and DHA levels in umbilical cord blood. Results. Caspase 3 activity was significantly lower in the DHA group in comparison to the control group. Umbilical cord blood DHA concentration was higher in the DHA group. The expression of the proteins p21 and Ki-67 did not differ significantly between the groups. Conclusions. We observed an association between DHA supplementation and inhibition of placental apoptosis. We did not find an association between DHA and proliferation process in the placenta. Ewa Wietrak, Krzysztof Kamiński, Bożena Leszczyńska-Gorzelak, and Jan Oleszczuk Copyright © 2015 Ewa Wietrak et al. All rights reserved. Krill Oil Ameliorates Mitochondrial Dysfunctions in Rats Treated with High-Fat Diet Sun, 02 Aug 2015 14:03:54 +0000 In recent years, several studies focused their attention on the role of dietary fats in the pathogenesis of hepatic steatosis. It has been demonstrated that a high-fat diet is able to induce hyperglycemia, hyperinsulinemia, obesity, and nonalcoholic fatty liver disease. On the other hand, krill oil, a novel dietary supplement of n-3 PUFAs, has the ability to improve lipid and glucose metabolism, exerting possible protective effects against hepatic steatosis. In this study we have investigated the effects of krill oil on mitochondrial energetic metabolism in animals fed a high-fat diet. To this end, male Sprague-Dawley rats were divided into three groups and fed for 4 weeks with a standard diet (control group), a diet with 35% fat (HF group), or a high-fat diet supplemented with 2.5% krill oil (HF+KO group). The obtained results suggest that krill oil promotes the burning of fat excess introduced by the high-fat diet. This effect is obtained by stimulating mitochondrial metabolic pathways such as fatty acid oxidation, Krebs cycle, and respiratory chain complexes activity. Modulation of the expression of carrier proteins involved in mitochondrial uncoupling was also observed. Overall, krill oil counteracts the negative effects of a high-fat diet on mitochondrial energetic metabolism. Alessandra Ferramosca, Annalea Conte, and Vincenzo Zara Copyright © 2015 Alessandra Ferramosca et al. All rights reserved. Omega-3 Fatty Acids in Early Prevention of Inflammatory Neurodegenerative Disease: A Focus on Alzheimer’s Disease Sun, 02 Aug 2015 13:32:40 +0000 Alzheimer’s disease (AD) is the leading cause of dementia and the most common neurodegenerative disease in the elderly. Furthermore, AD has provided the most positive indication to support the fact that inflammation contributes to neurodegenerative disease. The exact etiology of AD is unknown, but environmental and genetic factors are thought to contribute, such as advancing age, family history, presence of chronic diseases such as cardiovascular disease (CVD) and diabetes, and poor diet and lifestyle. It is hypothesised that early prevention or management of inflammation could delay the onset or reduce the symptoms of AD. Normal physiological changes to the brain with ageing include depletion of long chain omega-3 fatty acids and brains of AD patients have lower docosahexaenoic acid (DHA) levels. DHA supplementation can reduce markers of inflammation. This review specifically focusses on the evidence in humans from epidemiological, dietary intervention, and supplementation studies, which supports the role of long chain omega-3 fatty acids in the prevention or delay of cognitive decline in AD in its early stages. Longer term trials with long chain omega-3 supplementation in early stage AD are warranted. We also highlight the importance of overall quality and composition of the diet to protect against AD and dementia. J. Thomas, C. J. Thomas, J. Radcliffe, and C. Itsiopoulos Copyright © 2015 J. Thomas et al. All rights reserved. Moderate Dietary Supplementation with Omega-3 Fatty Acids Does Not Impact Plasma Von Willebrand Factor Profile in Mildly Hypertensive Subjects Sun, 02 Aug 2015 13:23:39 +0000 Long chain omega-3 polyunsaturated fatty acids (LC n-3 PUFAs) have blood pressure lowering and antithrombotic effects, which may benefit hypertensive patients. Increased plasma concentration of von Willebrand factor (vWF), a procoagulant glycoprotein, has been identified in patients with severe hypertension, with some, but not all studies showing an increase with mild hypertension. In this study, we determined the plasma concentration, multimer distribution, and collagen binding activity of vWF in subjects with mild hypertension and determined whether these parameters might improve after dietary supplementation with moderate amounts of LC n-3 PUFAs. Hypertensive and normotensive subjects were randomized to 12-week treatment with LC n-3 PUFAs (2.52 g/day) or placebo (canola oil). Home blood pressure measurements were recorded daily, and blood samples were collected every 3 weeks. LC n-3 PUFAs increased the n-3 index to cardioprotective levels (>8%). Plasma concentration, multimer distribution, and collagen binding activity of vWF were not reduced by LC n-3 PUFA treatment. We conclude that, at the concentration and duration used in this study, benefits of LC n-3 PUFAs in subjects with mild hypertension are not associated with a direct effect on vWF concentration or function. This trial is registered with the Australian New Zealand Clinical Trials Registry ACTRN12610000713099. Corinna S. Bürgin-Maunder, Peter R. Brooks, Deborah Hitchen-Holmes, and Fraser D. Russell Copyright © 2015 Corinna S. Bürgin-Maunder et al. All rights reserved. Omega-3 PUFAs Lower the Propensity for Arachidonic Acid Cascade Overreactions Sun, 02 Aug 2015 12:55:54 +0000 A productive view of the benefits from omega-3 (n-3) nutrients is that the dietary essential omega-6 (n-6) linoleic acid has a very narrow therapeutic window which is widened by n-3 nutrients. The benefit from moderate physiological actions of the arachidonic acid cascade can easily shift to harm from excessive pathophysiological actions. Recognizing the factors that predispose the cascade to an unwanted overactivity gives a rational approach for arranging beneficial interactions between the n-3 and n-6 essential nutrients that are initial components of the cascade. Much detailed evidence for harmful cascade actions was collected by pharmaceutical companies as they developed drugs to decrease those actions. A remaining challenge is to understand the factors that predispose the cascade toward unwanted outcomes and create the need for therapeutic interventions. Such understanding involves recognizing the similar dynamics for dietary n-3 and n-6 nutrients in forming the immediate precursors of the cascade plus the more vigorous actions of the n-6 precursor, arachidonic acid, in forming potent mediators that amplify unwanted cascade outcomes. Tools have been developed to aid deliberate day-to-day quantitative management of the propensity for cascade overactivity in ways that can decrease the need for drug treatments. Bill Lands Copyright © 2015 Bill Lands. All rights reserved. ω-3 PUFAs and Resveratrol Differently Modulate Acute and Chronic Inflammatory Processes Sun, 02 Aug 2015 12:37:18 +0000 -3 PUFAs and polyphenols have multiple effects on inflammation in vivo and in vitro. The effects of eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and resveratrol (RV) were investigated in LPS-stimulated peripheral blood leukocytes (PBLs) (i.e., acute inflammation) and IL-1 activated human chondrocytes (i.e., chronic inflammation). Inflammatory mediators including chemokines, cytokines, interleukins, and PGE2 were measured by multiplex analysis and gene expression was quantified by RT-PCR. In PBLs, RV decreased the secretion of PGE2, CCL5/RANTES, and CXCL8/IL-8 but increased IL-1, IL-6, and IL-10. In contrast to RV, -3 PUFAs augmented the production of PGE2 and CXCL8/IL-8. EPA and DHA similarly affected the pattern of inflammatory mediators. Combination of RV and -3 PUFAs exerted synergistic effects on CCL5/RANTES and had additive effects on IL-6 or CXCL8/IL-8. Both -3 PUFAs and RV reduced catabolic gene expression (e.g., MMPs, ADAMTS-4, IL-1, and IL-6) in activated chondrocytes. The data suggest that -3 PUFAs and RV differ in the regulation of acute inflammation of peripheral blood leukocytes but have common properties in modulating features related to chronic inflammation of chondrocytes. Joseph Schwager, Nathalie Richard, Christoph Riegger, and Norman Salem Jr. Copyright © 2015 Joseph Schwager et al. All rights reserved. Omega-3 Polyunsaturated Fatty Acids: Structural and Functional Effects on the Vascular Wall Sun, 02 Aug 2015 12:29:47 +0000 Omega-3 polyunsaturated fatty acids (n-3 PUFA) consumption is associated with reduced cardiovascular disease risk. Increasing evidence demonstrating a beneficial effect of n-3 PUFA on arterial wall properties is progressively emerging. We reviewed the recent available evidence for the cardiovascular effects of n-3 PUFA focusing on structural and functional properties of the vascular wall. In experimental studies and clinical trials n-3 PUFA have shown the ability to improve arterial hemodynamics by reducing arterial stiffness, thus explaining some of its cardioprotective properties. Recent studies suggest beneficial effects of n-3 PUFA on endothelial activation, which are likely to improve vascular function. Several molecular, cellular, and physiological pathways influenced by n-3 PUFA can affect arterial wall properties and therefore interfere with the atherosclerotic process. Although the relative weight of different physiological and molecular mechanisms and the dose-response on arterial wall properties have yet to be determined, n-3 PUFA have the potential to beneficially impact arterial wall remodeling and cardiovascular outcomes by targeting arterial wall stiffening and endothelial dysfunction. Michela Zanetti, Andrea Grillo, Pasquale Losurdo, Emiliano Panizon, Filippo Mearelli, Luigi Cattin, Rocco Barazzoni, and Renzo Carretta Copyright © 2015 Michela Zanetti et al. All rights reserved. Omega-3 Polyunsaturated Fatty Acids: The Way Forward in Times of Mixed Evidence Sun, 02 Aug 2015 12:22:15 +0000 Almost forty years ago, it was first hypothesized that an increased dietary intake of omega-3 polyunsaturated fatty acids (PUFA) from fish fat could exert protective effects against several pathologies. Decades of intense preclinical investigation have supported this hypothesis in a variety of model systems. Several clinical cardiovascular studies demonstrated the beneficial health effects of omega-3 PUFA, leading medical institutions worldwide to publish recommendations for their increased intake. However, particularly in recent years, contradictory results have been obtained in human studies focusing on cardiovascular disease and the clinical evidence in other diseases, particularly chronic inflammatory and neoplastic diseases, was never established to a degree that led to clear approval of treatment with omega-3 PUFA. Recent data not in line with the previous findings have sparked a debate on the health efficacy of omega-3 PUFA and the usefulness of increasing their intake for the prevention of a number of pathologies. In this review, we aim to examine the controversies on the possible use of these fatty acids as preventive/curative tools against the development of cardiovascular, metabolic, and inflammatory diseases, as well as several kinds of cancer. Karsten H. Weylandt, Simona Serini, Yong Q. Chen, Hui-Min Su, Kyu Lim, Achille Cittadini, and Gabriella Calviello Copyright © 2015 Karsten H. Weylandt et al. All rights reserved. The Omega-3 Fatty Acid Docosahexaenoic Acid Modulates Inflammatory Mediator Release in Human Alveolar Cells Exposed to Bronchoalveolar Lavage Fluid of ARDS Patients Sun, 02 Aug 2015 12:20:32 +0000 Background. This study investigated whether the 1 : 2 ω-3/ω-6 ratio may reduce proinflammatory response in human alveolar cells (A549) exposed to an ex vivo inflammatory stimulus (bronchoalveolar lavage fluid (BALF) of acute respiratory distress syndrome (ARDS) patients). Methods. We exposed A549 cells to the BALF collected from 12 ARDS patients. After 18 hours, fatty acids (FA) were added as docosahexaenoic acid (DHA, ω-3) and arachidonic acid (AA, ω-6) in two ratios (1 : 2 or 1 : 7). 24 hours later, in culture supernatants were evaluated cytokines (TNF-α, IL-6, IL-8, and IL-10) and prostaglandins (PGE2 and PGE3) release. The FA percentage content in A549 membrane phospholipids, content of COX-2, level of PPARγ, and NF-κB binding activity were determined. Results. The 1 : 2 DHA/AA ratio reversed the baseline predominance of ω-6 over ω-3 in the cell membranes (P < 0.001). The proinflammatory cytokine release was reduced by the 1 : 2 ratio (P < 0.01 to <0.001) but was increased by the 1 : 7 ratio (P < 0.01). The 1 : 2 ratio reduced COX-2 and PGE2 (P < 0.001) as well as NF-κB translocation into the nucleus (P < 0.01), while it increased activation of PPARγ and IL-10 release (P < 0.001). Conclusion. This study demonstrated that shifting the FA supply from ω-6 to ω-3 decreased proinflammatory mediator release in human alveolar cells exposed to BALF of ARDS patients. Paolo Cotogni, Antonella Trombetta, Giuliana Muzio, Marina Maggiora, and Rosa Angela Canuto Copyright © 2015 Paolo Cotogni et al. All rights reserved. Determination of Elemental Composition of Malabar spinach, Lettuce, Spinach, Hyacinth Bean, and Cauliflower Vegetables Using Proton Induced X-Ray Emission Technique at Savar Subdistrict in Bangladesh Thu, 02 Jul 2015 06:42:58 +0000 The concentrations of 18 different elements (K, Ca, Fe, Cl, P, Zn, S, Mn, Ti, Cr, Rb, Co, Br, Sr, Ru, Si, Ni, and Cu) were analyzed in five selected vegetables through Proton Induced X-ray Emission (PIXE) technique. The objective of this study was to provide updated information on concentrations of elements in vegetables available in the local markets at Savar subdistrict in Bangladesh. These elements were found in varying concentrations in the studied vegetables. The results also indicated that P, Cl, K, Ca, Mn, Fe, and Zn were found in all vegetables. Overall, K and Ca exhibited the highest concentrations. Cu and Ni exhibited the lowest concentrations in vegetables. The necessity of these elements was also evaluated, based on the established limits of regulatory standards. The findings of this study suggest that the consumption of these vegetables is not completely free of health risks. S. M. Fahad, A. F. M. Mahmudul Islam, Mahiuddin Ahmed, Nizam Uddin, Md. Rezaul Alam, Md. Ferdous Alam, Md. Farhan Khalik, Md. Sazzad Hossain, Md. Lokman Hossain, and Md. Joynal Abedin Copyright © 2015 S. M. Fahad et al. All rights reserved. Mass Spectrometry Applications in Biomedical Research Thu, 21 May 2015 09:56:37 +0000 Fa-Yun Che, Hai-Teng Deng, and Shi-Jian Ding Copyright © 2015 Fa-Yun Che et al. All rights reserved. Molecular Mechanisms of NAFLD in Metabolic Syndrome Wed, 20 May 2015 07:00:27 +0000 Maria João Martins, António Ascensão, José Magalhães, Maria Carmen Collado, and Piero Portincasa Copyright © 2015 Maria João Martins et al. All rights reserved. Gut Microbiota: Association with NAFLD and Metabolic Disturbances Tue, 19 May 2015 12:35:34 +0000 Nonalcoholic fatty liver disease is the hepatic expression of metabolic syndrome, being frequently associated with obesity, insulin resistance, and dyslipidemia. Recent lines of evidence have demonstrated a role of gut microbiota in insulin resistance, obesity, and associated metabolic disturbances, raising the interest in its relationship with NAFLD pathogenesis. Therefore, intestinal microbiota has emerged as a potential factor involved in NAFLD, through different pathways, including its influence in energy storage, lipid and choline metabolism, ethanol production, immune balance, and inflammation. The main objective of this review is to address the pathogenic association of gut microbiota to NAFLD. This comprehension may allow the development of integrated strategies to modulate intestinal microbiota in order to treat NAFLD. E. Lau, D. Carvalho, and P. Freitas Copyright © 2015 E. Lau et al. All rights reserved. Metabolomics for Biomarker Discovery: Moving to the Clinic Tue, 19 May 2015 11:46:55 +0000 To improve the clinical course of diseases, more accurate diagnostic and assessment methods are required as early as possible. In order to achieve this, metabolomics offers new opportunities for biomarker discovery in complex diseases and may provide pathological understanding of diseases beyond traditional technologies. It is the systematic analysis of low-molecular-weight metabolites in biological samples and has become an important tool in clinical research and the diagnosis of human disease and has been applied to discovery and identification of the perturbed pathways. It provides a powerful approach to discover biomarkers in biological systems and offers a holistic approach with the promise to clinically enhance diagnostics. When carried out properly, it could provide insight into the understanding of the underlying mechanisms of diseases, help to identify patients at risk of disease, and predict the response to specific treatments. Currently, metabolomics has become an important tool in clinical research and the diagnosis of human disease and becomes a hot topic. This review will highlight the importance and benefit of metabolomics for identifying biomarkers that accurately screen potential biomarkers of diseases. Aihua Zhang, Hui Sun, Guangli Yan, Ping Wang, and Xijun Wang Copyright © 2015 Aihua Zhang et al. All rights reserved. MALDI-TOF Mass Array Analysis of Nell-1 Promoter Methylation Patterns in Human Gastric Cancer Tue, 19 May 2015 11:38:06 +0000 Mass spectrometry (MS) enables rapid and sensitive qualitative and quantitative analyses of biomolecules (proteins, peptides, oligosaccharides, lipids, DNA, and RNA), drugs, and metabolites. MS has become an essential tool in modern biomedical research, including the analysis of DNA methylation. DNA methylation has been reported in many cancers, suggesting that it can be utilized as an early biomarker to improve the early diagnosis rate. Using matrix-assisted laser desorption/ionization time-of-flight MS and MassCLEAVE reagent, we compared Nell-1 hypermethylation levels among tumor tissues, paracarcinoma tissues, and normal tissues from gastric cancer patients. Almost 80% of the CpG sites in the amplicons produced were covered by the analysis. Our results indicate a significant difference in methylation status between gastric cancer tissue (a higher level) and normal tissue. The same trend was identified in gastric cancer tissue versus paracarcinoma tissue. We also detected lower relative expression of Nell-1 by real-time PCR. Furthermore, immunohistochemical analyses revealed that Nell-1 staining was less intense in cancer tissue relative to normal tissue and that the tumor cells had spread to the muscle layer. These findings may serve as a guide for the early diagnosis of gastric cancer. Changlu Gao, Qian Zhang, Deyang Kong, Di Wu, Changlei Su, Jinxue Tong, Feng Chen, and Qifan Zhang Copyright © 2015 Changlu Gao et al. All rights reserved. Mass Spectrometry in Pharmacokinetic Studies of a Synthetic Compound for Spinal Cord Injury Treatment Tue, 19 May 2015 07:39:17 +0000 The studies of drugs that could constitute a palliative to spinal cord injury (SCI) are a continuous and increasing demand in biomedicine field from developed societies. Recently we described the chemical synthesis and antiglioma activity of synthetic glycosides. A synthetic sulfated glycolipid (here IG20) has shown chemical stability, solubility in polar solvents, and high inhibitory capacity over glioma growth. We have used mass spectrometry (MS) to monitor IG20 in cells and tissues of the central nervous system (CNS) that are involved in SCI recovery. IG20 was detected by MS in serum and homogenates from CNS tissue of rats, though in the latter a previous deproteinization step was required. The pharmacokinetic parameters of serum clearance at 24 h and half-life at 4 h were determined for synthetic glycoside in the adult rat using MS. A local administration of the drug near of spinal lesion site is proposed. María Sánchez-Sierra, Isabel García-Álvarez, Alfonso Fernández-Mayoralas, Sandra Moreno-Lillo, Gemma Barroso García, Verónica Moral Dardé, and Ernesto Doncel-Pérez Copyright © 2015 María Sánchez-Sierra et al. All rights reserved. Proteomic Analysis of Estrogen-Mediated Signal Transduction in Osteoclasts Formation Mon, 18 May 2015 14:25:07 +0000 Estrogen plays an important role in inhibiting osteoclast differentiation and protecting against bone loss from osteoporosis, especially in postmenopausal women. However, the precise mechanisms underlying the effect of estrogen on osteoclasts are not well known. In the present study, we performed proteomics analysis and bioinformatics analysis to comprehensively compare the differential expression of proteins in receptor activator of nuclear factor-κB ligand RANKL-induced osteoclasts in the presence and absence of estrogen. We identified 6403 proteins, of which 124 were upregulated and 231 were downregulated by estrogen. Bioinformatics analysis showed that estrogen treatment interfered with 77 intracellular pathways, including both confirmed canonical and unconfirmed pathways of osteoclast formation. Our findings validate the inhibitory effect of estrogen on osteoclasts via the promotion of apoptosis and suppression of differentiation and polarization and suggest that estrogen might inhibit osteoclast formation via other pathways, which requires further investigation and verification. Qi Xiong, Peifu Tang, Yanpan Gao, Lihai Zhang, and Wei Ge Copyright © 2015 Qi Xiong et al. All rights reserved. Global Methylation and Hydroxymethylation in DNA from Blood and Saliva in Healthy Volunteers Mon, 18 May 2015 13:24:03 +0000 Aims. We describe a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify and compare simultaneously global methylation and hydroxymethylation in human DNA of different tissues. Materials and Methods. Blood and saliva DNA from fourteen volunteers was processed for epigenetic endpoints using LC-MS/MS and PCR-pyrosequencing technology. Results. Global DNA methylation was significantly lower in saliva (mean 4.61% ±  0.80%), compared to blood samples (5.70% ± 0.22%). In contrast, saliva (0.036% ± 0.011%) revealed significantly higher hydroxymethylation compared to blood samples (mean 0.027% ± 0.004%). Whereas we did not find significant correlations for both epigenetic measures between the tissues, a significant association was observed between global methylation and global hydroxymethylation in saliva DNA. Neither LINE-1 nor Alu elements of blood and saliva correlated, nor were they correlated with the DNA hydroxymethylation of blood or saliva, respectively. Conclusion. Global DNA methylation and hydroxymethylation of cytosine can be quantified simultaneously by LC-MS/MS. Saliva DNA cannot be considered as a surrogate for blood DNA to study epigenetic endpoints. Lode Godderis, Caroline Schouteden, Ali Tabish, Katrien Poels, Peter Hoet, Andrea A. Baccarelli, and Kirsten Van Landuyt Copyright © 2015 Lode Godderis et al. All rights reserved. Mass Spectral Profile for Rapid Differentiating Beta-Lactams from Their Ring-Opened Impurities Mon, 18 May 2015 12:21:15 +0000 High performance liquid chromatography tandem mass spectrometry (HPLC MS) has been widely used for β-lactam antibiotics determination. However, its application to identify impurities of these frequently used drugs is not sufficient at present. In this job, characteristic profiles of the collision induced dissociation (CID) spectra of both β-lactams and ring-opened β-lactams were extracted from the MS data of six β-lactam antibiotics and their forty-five impurities, and were confirmed by the MS data reported in the literature. These characteristics have been successfully applied to rapid differentiation of β-lactam and ring-opened β-lactam impurities in cefixime, cefdinir, and cefaclor. However, these characteristic profiles can only be obtained under low activating voltage. They did not display in the high energy activated CID spectra. Diagnostic fragmentations for determining the localization of double bond and substituents on the thiazine ring and the side chain were also observed. In addition, several characteristic fragmentations are hopeful to be used to differentiate the configurations of C-2 on the thiazine ring of ring-opened impurities, which is generally disadvantageous of mass spectrometry. Taken together, forty-five impurities were identified from the capsules of cefixime, cefdinir, and cefaclor. Hecheng Wang, Haiwei Huang, Jin Cao, Dehua Chui, and Shengyuan Xiao Copyright © 2015 Hecheng Wang et al. All rights reserved. Developmental Programming of Nonalcoholic Fatty Liver Disease: The Effect of Early Life Nutrition on Susceptibility and Disease Severity in Later Life Mon, 18 May 2015 12:14:20 +0000 Nonalcoholic fatty liver disease (NAFLD) is fast becoming the most common liver disease globally and parallels rising obesity rates. The developmental origins of health and disease hypothesis have linked alterations in the early life environment to an increased risk of metabolic disorders in later life. Altered early life nutrition, in addition to increasing risk for the development of obesity, type 2 diabetes, and cardiovascular disease in offspring, is now associated with an increased risk for the development of NAFLD. This review summarizes emerging research on the developmental programming of NAFLD by both maternal obesity and undernutrition with a particular focus on the possible mechanisms underlying the development of hepatic dysfunction and potential strategies for intervention. Minglan Li, Clare M. Reynolds, Stephanie A. Segovia, Clint Gray, and Mark H. Vickers Copyright © 2015 Minglan Li et al. All rights reserved. The Dual Role of Nrf2 in Nonalcoholic Fatty Liver Disease: Regulation of Antioxidant Defenses and Hepatic Lipid Metabolism Mon, 18 May 2015 10:04:26 +0000 Nonalcoholic fatty liver disease (NAFLD) is a progressive liver disease with ever-growing incidence in the industrialized world. It starts with the simple accumulation of lipids in the hepatocyte and can progress to the more severe nonalcoholic steatohepatitis (NASH), which is associated with inflammation, fibrosis, and cirrhosis. There is increasing awareness that reactive oxygen species and electrophiles are implicated in the pathogenesis of NASH. Transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) is a positive regulator of the expression of a battery of genes involved in the protection against oxidative/electrophilic stress. In rodents, Nrf2 is also known to participate in hepatic fatty acid metabolism, as a negative regulator of genes that promote hepatosteatosis. We review relevant evidence in the literature that these two mechanisms may contribute to the protective role of Nrf2 in the development of hepatic steatosis and in the progression to steatohepatitis, particularly in young animals. We propose that age may be a key to explain contradictory findings in the literature. In summary, Nrf2 mediates the crosstalk between lipid metabolism and antioxidant defense mechanisms in experimental models of NAFLD, and the nutritional or pharmacological induction of Nrf2 represents a promising potential new strategy for its prevention and treatment. Sílvia S. Chambel, Andreia Santos-Gonçalves, and Tiago L. Duarte Copyright © 2015 Sílvia S. Chambel et al. All rights reserved. Mass Spectrometry-Based Proteomic Study Makes High-Density Lipoprotein a Biomarker for Atherosclerotic Vascular Disease Mon, 18 May 2015 09:59:21 +0000 High-density lipoprotein (HDL) is a lipid and protein complex that consists of apolipoproteins and lower level HDL-associated enzymes. HDL dysfunction is a factor in atherosclerosis and decreases patient survival. Mass spectrometry- (MS-) based proteomics provides a high throughput approach for analyzing the composition and modifications of complex HDL proteins in diseases. HDL can be separated according to size, surface charge, electronegativity, or apoprotein composition. MS-based proteomics on subfractionated HDL then allows investigation of lipoprotein roles in diseases. Herein, we review recent developments in MS-based quantitative proteomic techniques, HDL proteomics and lipoprotein modifications in diseases, and HDL subfractionation studies. We also discuss future directions and perspectives in MS-based proteomics on HDL. Chiz-Tzung Chang, Chao-Yuh Yang, Fuu-Jen Tsai, Shih-Yi Lin, and Chao-Jung Chen Copyright © 2015 Chiz-Tzung Chang et al. All rights reserved. Identification of Glioblastoma Phosphotyrosine-Containing Proteins with Two-Dimensional Western Blotting and Tandem Mass Spectrometry Mon, 18 May 2015 09:55:13 +0000 To investigate the presence of, and the potential biological roles of, protein tyrosine phosphorylation in the glioblastoma pathogenesis, two-dimensional gel electrophoresis- (2DGE-) based Western blotting coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis was used to detect and identify the phosphotyrosine immunoreaction-positive proteins in a glioblastoma tissue. MS/MS and Mascot analyses were used to determine the phosphotyrosine sites of each phosphopeptide. Protein domain and motif analysis and systems pathway analysis were used to determine the protein domains/motifs that contained phosphotyrosine residue and signal pathway networks to clarify the potential biological functions of protein tyrosine phosphorylation. A total of 24 phosphotyrosine-containing proteins were identified. Each phosphotyrosine-containing protein contained at least one tyrosine kinase phosphorylation motif and a certain structural and functional domains. Those phosphotyrosine-containing proteins were involved in the multiple signal pathway systems such as oxidative stress, stress response, and cell migration. Those data show 2DGE-based Western blotting, MS/MS, and bioinformatics are a set of effective approaches to detect and identify glioblastoma tyrosine-phosphorylated proteome and to effectively rationalize the biological roles of tyrosine phosphorylation in the glioblastoma biological systems. It provides novel insights regarding tyrosine phosphorylation and its potential role in the molecular mechanism of a glioblastoma. Tianyao Guo, Xiaowei Wang, Maoyu Li, Haiyan Yang, Ling Li, Fang Peng, and Xianquan Zhan Copyright © 2015 Tianyao Guo et al. All rights reserved. How Inflammation Impinges on NAFLD: A Role for Kupffer Cells Mon, 18 May 2015 09:39:20 +0000 Nonalcoholic fatty liver disease (NAFLD) is rapidly becoming the most prevalent cause of liver disease worldwide and afflicts adults and children as currently associated with obesity and insulin resistance. Even though lately some advances have been made to elucidate the mechanism and causes of the disease much remains unknown about NAFLD. The aim of this paper is to discuss the present knowledge regarding the pathogenesis of the disease aiming at the initial steps of NAFLD development, when inflammation impinges on fat liver deposition. At this stage, the Kupffer cells attain a prominent role. This knowledge becomes subsequently relevant for the development of future diagnostic, prevention, and therapeutic options for the management of NAFLD. Nádia Duarte, Inês C. Coelho, Rita S. Patarrão, Joana I. Almeida, Carlos Penha-Gonçalves, and M. Paula Macedo Copyright © 2015 Nádia Duarte et al. All rights reserved. Individual CLA Isomers, c9t11 and t10c12, Prevent Excess Liver Glycogen Storage and Inhibit Lipogenic Genes Expression Induced by High-Fructose Diet in Rats Mon, 18 May 2015 09:35:04 +0000 This study assessed the effects of individual conjugated linoleic acid isomers, c9t11-CLA and t10c12-CLA, on nonalcoholic fatty liver disease (NAFLD) and systemic endothelial dysfunction in rats fed for four weeks with control or high-fructose diet. The high-fructose diet hampered body weight gain (without influencing food intake), increased liver weight and glycogen storage in hepatocytes, upregulated expression of fatty acid synthase (FAS) and stearoyl-CoA desaturase-1 (SCD-1), and increased saturated fatty acid (SFA) content in the liver. Both CLA isomers prevented excessive accumulation of glycogen in the liver. Specifically, t10c12-CLA decreased concentration of serum triacylglycerols and LDL + VLDL cholesterol, increased HDL cholesterol, and affected liver lipid content and fatty acid composition by downregulation of liver SCD-1 and FAS expression. In turn, the c9t11-CLA decreased LDL+VLDL cholesterol in the control group and downregulated liver expression of FAS without significant effects on liver weight, lipid content, and fatty acid composition. In summary, feeding rats with a high-fructose diet resulted in increased liver glycogen storage, indicating the induction of gluconeogenesis despite simultaneous upregulation of genes involved in de novo lipogenesis. Although both CLA isomers (c9t11 and t10c12) display hepatoprotective activity, the hypolipemic action of the t10c12-CLA isomer proved to be more pronounced than that of c9t11-CLA. Edyta Maslak, Elzbieta Buczek, Antoni Szumny, Wojciech Szczepnski, Magdalena Franczyk-Zarow, Aneta Kopec, Stefan Chlopicki, Teresa Leszczynska, and Renata B. Kostogrys Copyright © 2015 Edyta Maslak et al. All rights reserved. Potential Peripartum Markers of Infectious-Inflammatory Complications in Spontaneous Preterm Birth Mon, 18 May 2015 09:11:45 +0000 Spontaneous preterm birth significantly contributes to the overall neonatal morbidity associated with preterm deliveries. Nearly 50% of cases are associated with microbial invasion of the amniotic cavity followed by an inflammatory response. Robust diagnostic tools for neonates jeopardized by infection and inflammation may thus decrease the overall neonatal morbidity substantially. Amniotic fluid retrieved during labor retains fetal and pregnancy-related protein fingerprint and its sampling does not place any unwanted stress on women. Using exploratory and targeted methods we analyzed proteomes of amniotic fluid sampled at the end of spontaneous preterm labor prior to delivery from women with and without infection and inflammation. Exploratory data indicated several amniotic fluid proteins to be associated with infectious-inflammatory complications in spontaneous preterm birth. LC-SRM analysis subsequently verified statistically significant changes in lipocalin-1 ( and AUC = 0.67, ), glycodelin ( and AUC = 0.73, ), and nicotinamide phosphoribosyltransferase ( and AUC = 0.71, ). Vojtech Tambor, Marie Vajrychova, Marian Kacerovsky, Marek Link, Petra Domasinska, Ramkumar Menon, and Juraj Lenco Copyright © 2015 Vojtech Tambor et al. All rights reserved. Magnetic Bead-Based Serum Peptidome Profiling in Patients with Gestational Diabetes Mellitus Mon, 18 May 2015 08:56:03 +0000 Gestational diabetes mellitus (GDM) is a frequent medical condition during pregnancy. Early diagnosis and treatment of GDM are crucial for both the mother and the baby. In the present study, we aimed to identify specific biomarkers to assist in the early detection of GDM and give some clues to the possible causes of GDM by comparing serum peptide profile differences between GDM patients and healthy controls. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used in combination with weak cation exchange magnetic bead (WCX-MB). Levels of four peptides (4418.9, 2219.7, 2211.5, and 1533.4 Da) were significantly different. Interestingly, three of them (4418.9, 2211.5, and 1533.4 Da) were identified when GDM patients with two degrees of glucose intolerance were compared. Additionally, peptides 2211.5 and 1533.4 Da showed a decreasing trend as glucose intolerance increased, while peptide 4418.9 Da exhibited the reverse tendency. In conclusion, our study provides novel insights into the altered serum peptide profile of GDM patients. The specific candidate biomarkers may contribute to the development of GDM. Tingting Ai, Feng Chen, Shaonan Zhou, Jieni Zhang, Hui Zheng, Yanheng Zhou, Wei Hu, Xiaofei Liu, Li Li, and Jiuxiang Lin Copyright © 2015 Tingting Ai et al. All rights reserved. Nonalcoholic Fatty Liver Disease Relationship with Metabolic Syndrome in Class III Obesity Individuals Mon, 18 May 2015 08:43:45 +0000 Introduction. Obesity is represented mainly by abdominal obesity and insulin resistance (IR), both present in most individuals diagnosed with metabolic syndrome (MS). IR is the key risk factor in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Objective. To relate NAFLD to MS in class III obese individuals. Methodology. A descriptive cross-sectional study with class III obese individuals, aged ≥ 20–60 years. Blood pressure measurement, weight, height, body mass index (BMI), waist circumference (WC) and blood glucose, insulin, high-density lipoprotein cholesterol (HDL-c), and triglycerides data were obtained. HOMA-IR (homeostatic model assessment insulin resistance) calculation was carried out with a cutoff value of 2.71 for IR evaluation. The diagnosis of NAFLD was performed by liver biopsy and the diagnosis of MS was performed in accordance with the National Cholesterol Education Program/Adult Treatment Panel III (NCEPATP III). Results. Of the 50 individuals evaluated, 86% were women and BMI means were 45.4 ± 3.6 Kg/m2. The overall individuals had NAFLD, 70% steatosis, and 30% steatohepatitis. The diagnosis of MS occurred in 56% but showed no significant association with NAFLD (). Triglycerides (178 ± 65.5 mg/dL) and insulin (28.2 ± 22.6 mcU/mL) mean values were significantly higher in steatohepatitis ( and , resp.) compared to individuals with steatosis. IR was confirmed in 76% and showed a relationship with NAFLD severity. Conclusion. NAFLD was not related to MS; however, MS components, evaluated in isolation, as well as IR, were related to the presence and severity of NAFLD. A. Cordeiro, S. E. Pereira, C. J. Saboya, and A. Ramalho Copyright © 2015 A. Cordeiro et al. All rights reserved. Polyuridylation in Eukaryotes: A 3′-End Modification Regulating RNA Life Mon, 11 May 2015 09:03:19 +0000 In eukaryotes, mRNA polyadenylation is a well-known modification that is essential for many aspects of the protein-coding RNAs life cycle. However, modification of the 3′ terminal nucleotide within various RNA molecules is a general and conserved process that broadly modulates RNA function in all kingdoms of life. Numerous types of modifications have been characterized, which are generally specific for a given type of RNA such as the CCA addition found in tRNAs. In recent years, the addition of nontemplated uridine nucleotides or uridylation has been shown to occur in various types of RNA molecules and in various cellular compartments with significantly different outcomes. Indeed, uridylation is able to alter RNA half-life both in positive and in negative ways, highlighting the importance of the enzymes in charge of performing this modification. The present review aims at summarizing the current knowledge on the various processes leading to RNA 3′-end uridylation and on their potential impacts in various diseases. Paola Munoz-Tello, Lional Rajappa, Sandrine Coquille, and Stéphane Thore Copyright © 2015 Paola Munoz-Tello et al. All rights reserved. Drug Resistance in Hematologic Malignancies: Induction Mechanisms, Genetics, and Therapeutics Mon, 27 Apr 2015 11:15:56 +0000 Fenghuang Zhan, Maurizio Zangari, and Lugui Qiu Copyright © 2015 Fenghuang Zhan et al. All rights reserved. Synergistic Activity of Carfilzomib and Panobinostat in Multiple Myeloma Cells via Modulation of ROS Generation and ERK1/2 Mon, 27 Apr 2015 07:15:46 +0000 Relapse of disease and subsequent resistance to established therapies remain as major challenges in the treatment of multiple myeloma (MM). New therapeutic options are needed for these extensively pretreated patients. To explore an optimized combinational therapy, interactions between the irreversible proteasome inhibitor carfilzomib exhibiting a well-tolerated side-effect profile and histone deacetylase inhibitor (HDACi) panobinostat (LBH589) were examined in MM cells. Coadministration of carfilzomib and LBH589 led to a synergistic inhibition of proliferation in MM cells. Further studies showed that the combined treatment synergistically increased mitochondrial injury, caspase activation, and apoptosis in MM cells. Lethality of the carfilzomib/LBH589 combination was associated with the reactive oxygen species (ROS) generation and ERK1/2 inactivation. In addition, the free radical scavenger N-acetylcysteine (NAC) could block carfilzomib and LBH589-induced oxidative stress and the subsequent apoptosis. Together, these findings argue that the strategy of combining carfilzomib and LBH589 warrants attention in MM. Lu Gao, Minjie Gao, Guang Yang, Yi Tao, Yuanyuan Kong, Ruixue Yang, Xiuqin Meng, Gongwen Ai, Rong Wei, Huiqun Wu, Xiaosong Wu, and Jumei Shi Copyright © 2015 Lu Gao et al. All rights reserved. Smoldering Multiple Myeloma Mon, 27 Apr 2015 07:09:39 +0000 Smoldering multiple myeloma (SMM) is an asymptomatic precursor stage of multiple myeloma (MM) characterized by clonal bone marrow plasma cells (BMPC) ≥ 10% and/or M protein level ≥ 30 g/L in the absence of end organ damage. It represents an intermediate stage between monoclonal gammopathy of undetermined significance (MGUS) and symptomatic MM. The risk of progression to symptomatic MM is not uniform, and several parameters have been reported to predict the risk of progression. These include the level of M protein and the percentage of BMPC, the proportion of immunophenotypically aberrant plasma cells, and the presence of immunoparesis, free light-chain (FLC) ratio, peripheral blood plasma cells (PBPC), pattern of serum M protein evolution, abnormal magnetic resonance imaging (MRI), cytogenetic abnormalities, IgA isotype, and Bence Jones proteinuria. So far treatment is still not recommended for SMM, because several trials suggested that patients with SMM do not benefit from early treatment. However, the Mateos et al. trial showed a survival benefit after early treatment with lenalidomide plus dexamethasone in patients with high-risk SMM. This trial has prompted a reevaluation of early treatment in an asymptomatic patient population. Minjie Gao, Guang Yang, Yuanyuan Kong, Xiaosong Wu, and Jumei Shi Copyright © 2015 Minjie Gao et al. All rights reserved. Constitutive NF-κB Activation Underlines Major Mechanism of Drug Resistance in Relapsed Refractory Diffuse Large B Cell Lymphoma Thu, 23 Apr 2015 14:32:04 +0000 Diffuse large B cell lymphoma (DLBCL) is the most common subtype of B cell non-Hodgkin’s lymphoma (NHL), encompassing 30–40% of the estimated 70,000 cases of NHL in 2014 in the USA. Despite major improvements with immune-chemotherapy, the fraction of patients who still succumb to a refractory or relapsed disease remains high. This review addresses whether the better understanding of the biology of DLBCL defines new therapeutic avenues that may overcome the emerging resistance of this disease to traditional immune-chemotherapy, such as rituximab in combination with traditional chemotherapy agents. Emerging targeted therapy for relapsed refractory DLBCL encompasses more complex molecular abnormalities involving signaling pathways other than NF-κB as mechanism of resistance to immune-chemotherapy. Our review suggests that NF-κB pathway is an important crossroad where other pathways converge as phenotype of resistance that emerges in patients who fail frontline and salvage immune-chemotherapy. Future efforts should aim at targeting the role of NF-κB resistance in clinical trials, where novel agents like lenalidomide and proteasome inhibitors with established activity in this perspective will be an important component in combination therapy, along with new monoclonal antibody, BTK-inhibitors, and other novel therapy agents. Francesco Turturro Copyright © 2015 Francesco Turturro. All rights reserved. Extramedullary Manifestation in Multiple Myeloma Bears High Incidence of Poor Cytogenetic Aberration and Novel Agents Resistance Thu, 23 Apr 2015 13:30:00 +0000 Extramedullary disease (EMD) in multiple myeloma (MM) patients is an uncommon event and more attention was directed toward the feature of these patients. Cytogenetic aberration is an important characteristic of MM and is associated with patients’ outcome. In this study, we aimed to compare the cytogenetic abnormality of patients with and without extramedullary manifestation, and to analyze the clinical outcomes of novel agents in EMD patients. We retrospectively investigated data from 41 MM patients. Our analyses showed del(17p13) in 31% of EMD versus 13% of medullary disease () and amp(1q21) in 55% versus 32% (). No differences were shown in del(13q14) and t(4;14). 24/27 patients with EMD at diagnosis responded to the novel agents-containing regimens. However, when relapsed, 70% of patients did not benefit from the sequential use of novel agents as salvage therapy. In 14 patients who developed EMD at relapse phase, only 2 patients responded to novel agents therapy. Median overall survival of patients with extramedullary manifestations was 30 months, in comparison to 104 months for patients without EMD (). Patients with extramedullary manifestation bore high incidence of poor cytogenetic aberration and novel agents resistance. Xiaoyan Qu, Lijuan Chen, Hairong Qiu, Hua Lu, Hanxin Wu, Hongxia Qiu, Peng Liu, Rui Guo, and Jianyong Li Copyright © 2015 Xiaoyan Qu et al. All rights reserved. Neurotensin Branched Peptide as a Tumor-Targeting Agent for Human Bladder Cancer Thu, 23 Apr 2015 06:44:40 +0000 Despite recent advances in multimodal therapy, bladder cancer still ranks ninth in worldwide cancer incidence. New molecules which might improve early diagnosis and therapeutic efficiency for tumors of such high epidemiological impact therefore have very high priority. In the present study, the tetrabranched neurotensin peptide NT4 was conjugated with functional units for cancer-cell imaging or therapy and was tested on bladder cancer cell lines and specimens from bladder cancer surgical resections, in order to evaluate its potential for targeted personalized therapy of bladder cancer. Fluorophore-conjugated NT4 distinguished healthy and cancer tissues with good statistical significance . NT4 conjugated to methotrexate or gemcitabine was cytotoxic for human bladder cancer cell lines at micromolar concentrations. Their selectivity for bladder cancer tissue and capacity to carry tracers or drugs make NT4 peptides candidate tumor targeting agents for tracing cancer cells and for personalized therapy of human bladder cancer. Jlenia Brunetti, Chiara Falciani, Barbara Lelli, Andrea Minervini, Niccolò Ravenni, Lorenzo Depau, Giampaolo Siena, Eleonora Tenori, Stefano Menichetti, Alessandro Pini, Marco Carini, and Luisa Bracci Copyright © 2015 Jlenia Brunetti et al. All rights reserved. Functional Polymorphisms in COX-2 Gene Are Correlated with the Risk of Oral Cancer Tue, 21 Apr 2015 07:53:52 +0000 Background. This meta-analysis investigated the association between functional COX-2 gene polymorphisms and the risk of oral cancer. Methods. Several electronic databases were searched for published studies using combinations of keywords related to COX-2 gene polymorphisms and oral cancer. After selection of relevant studies, following strict inclusion and exclusion criteria, data was performed using STATA 12.0 software. Results. We retrieved 83 studies from database search using specific search terms. After multiple rounds of selection and elimination, 7 studies were finally identified as suitable to be included in our present meta-analysis, based on their relevance and data integrity. These 7 studies contained a combined total of 2,296 oral cancer patients and 3,647 healthy controls. Our findings demonstrated that +837 T > C (rs5275) polymorphism in COX-2 showed statistically significant differences in gene frequencies in case and control groups in allele model and dominant model. Similar results were obtained with COX-2 gene polymorphism 765 G > C (rs20417). On the other hand, 1195 A > G (rs689466) polymorphism in COX-2 did not confer susceptibility to oral cancers. Conclusion. Based on our results, COX-2 gene polymorphisms, +837 T > C (rs5275) and −765G > C (rs20417), showed clear links with oral cancer susceptibility, and the 1195A > G (rs689466) polymorphism did not show such a correlation. Dong Li, Shu-Hong Hao, Yan Sun, Chun-Mei Hu, Zhi-Hua Ma, Zhi Ming Wang, Jie Liu, Hong Bo Liu, Ming Ye, Yu Fei Zhang, Dong Sheng Yang, and Guang Shi Copyright © 2015 Dong Li et al. All rights reserved. Nutrients and Muscle Disease Thu, 19 Mar 2015 06:38:01 +0000 Enzo Nisoli, Robert W. Grange, and Giuseppe D’Antona Copyright © 2015 Enzo Nisoli et al. All rights reserved. Improvement of Endurance of DMD Animal Model Using Natural Polyphenols Sun, 15 Mar 2015 13:46:24 +0000 Duchenne muscular dystrophy (DMD), the most common form of muscular dystrophy, is characterized by muscular wasting caused by dystrophin deficiency that ultimately ends in force reduction and premature death. In addition to primary genetic defect, several mechanisms contribute to DMD pathogenesis. Recently, antioxidant supplementation was shown to be effective in the treatment of multiple diseases including muscular dystrophy. Different mechanisms were hypothesized such as reduced hydroxyl radicals, nuclear factor-κB deactivation, and NO protection from inactivation. Following these promising evidences, we investigated the effect of the administration of a mix of dietary natural polyphenols (ProAbe) on dystrophic mdx mice in terms of muscular architecture and functionality. We observed a reduction of muscle fibrosis deposition and myofiber necrosis together with an amelioration of vascularization. More importantly, the recovery of the morphological features of dystrophic muscle leads to an improvement of the endurance of treated dystrophic mice. Our data confirmed that ProAbe-based diet may represent a strategy to coadjuvate the treatment of DMD. Clementina Sitzia, Andrea Farini, Federica Colleoni, Francesco Fortunato, Paola Razini, Silvia Erratico, Alessandro Tavelli, Francesco Fabrizi, Marzia Belicchi, Mirella Meregalli, Giacomo Comi, and Yvan Torrente Copyright © 2015 Clementina Sitzia et al. All rights reserved. Effect of Different Germination Conditions on Antioxidative Properties and Bioactive Compounds of Germinated Brown Rice Thu, 12 Mar 2015 08:09:37 +0000 This study investigates antioxidative activity and bioactive compounds of ungerminated brown rice (UBR) and germinated brown rice (GBR). We used two rice cultivars (Oryza sativa L.), Taiwan Japonica 9 (TJ-9) and Taichung Indica 10 (TCI-10), as the materials in our experiments. The conditions for inducing germination are soaking time in water 24, 48, or 72 h; temperature 26 or 36°C; incubation in light or darkness; and open or closed vessels, in which the antioxidative activities and bioactive compounds of GBR were determined. We found that, in order to maximize antioxidative activity and bioactive compounds, germination should be under higher temperature (36°C), long soaking time (72 h), darkness, and closed vessel. GBR contains much higher levels of antioxidative activity and bioactive compounds than ungerminated brown rice (UBR). We found a strong correlation between antioxidative activities (DPPH radical scavenging ability, reducing power, and Trolox equivalent antioxidant capacity) and bioactive compounds (γ-oryzanols, tocopherol, and tocotrienol). Higher temperature (36°C) is also conducive to the production of GABA in GBR. These results are considered very useful research references for the development of future functional foods and additives. You-Tung Lin, Cheng-Cheng Pao, Shwu-Tzy Wu, and Chi-Yue Chang Copyright © 2015 You-Tung Lin et al. All rights reserved. Antioxidant Strategies in the Management of Diabetic Neuropathy Mon, 02 Mar 2015 15:04:39 +0000 Chronic hyperglycaemia (an abnormally high glucose concentration in the blood) resulting from defects in insulin secretion/action, or both, is the major hallmark of diabetes in which it is known to be involved in the progression of the condition to different complications that include diabetic neuropathy. Diabetic neuropathy (diabetes-induced nerve damage) is the most common diabetic complication and can be devastating because it can lead to disability. There is an increasing body of evidence associating diabetic neuropathy with oxidative stress. Oxidative stress results from the production of oxygen free radicals in the body in excess of its ability to eliminate them by antioxidant activity. Antioxidants have different mechanisms and sites of actions by which they exert their biochemical effects and ameliorate nerve dysfunction in diabetes by acting directly against oxidative damage. This review will examine different strategies for managing diabetic neuropathy which rely on exogenous antioxidants. Ayodeji Babatunde Oyenihi, Ademola Olabode Ayeleso, Emmanuel Mukwevho, and Bubuya Masola Copyright © 2015 Ayodeji Babatunde Oyenihi et al. All rights reserved. Recent Developments of Carbonic Anhydrase Inhibitors as Potential Drugs Mon, 02 Mar 2015 12:21:01 +0000 Jamshed Iqbal, Mariya Al-Rashida, Serdar Durdagi, Vincenzo Alterio, and Anna Di Fiore Copyright © 2015 Jamshed Iqbal et al. All rights reserved. Genetic and Metabolic Intraspecific Biodiversity of Ganoderma lucidum Sun, 01 Mar 2015 09:27:30 +0000 Fourteen Ganoderma lucidum strains from different geographic regions were identified using ITS region sequencing. Based on the sequences obtained, the genomic relationship between the analyzed strains was determined. All G. lucidum strains were also genetically characterized using the AFLP technique. G. lucidum strains included in the analysis displayed an AFLP profile similarity level in the range from 9.6 to 33.9%. Biolog FF MicroPlates were applied to obtain data on utilization of 95 carbon sources and mitochondrial activity. The analysis allowed comparison of functional diversity of the fungal strains. The substrate utilization profiles for the isolates tested revealed a broad variability within the analyzed G. lucidum species and proved to be a good profiling technology for studying the diversity in fungi. Significant differences have been demonstrated in substrate richness values. Interestingly, the analysis of growth and biomass production also differentiated the strains based on the growth rate on the agar and sawdust substrate. In general, the mycelial growth on the sawdust substrate was more balanced and the fastest fungal growth was observed for GRE3 and FCL192. Anna Pawlik, Grzegorz Janusz, Iwona Dębska, Marek Siwulski, Magdalena Frąc, and Jerzy Rogalski Copyright © 2015 Anna Pawlik et al. All rights reserved. Probing the Surface of Human Carbonic Anhydrase for Clues towards the Design of Isoform Specific Inhibitors Tue, 24 Feb 2015 08:40:36 +0000 The alpha carbonic anhydrases (α-CAs) are a group of structurally related zinc metalloenzymes that catalyze the reversible hydration of CO2 to . Humans have 15 different α-CAs with numerous physiological roles and expression patterns. Of these, 12 are catalytically active, and abnormal expression and activities are linked with various diseases, including glaucoma and cancer. Hence there is a need for CA isoform specific inhibitors to avoid off-target CA inhibition, but due to the high amino acid conservation of the active site and surrounding regions between each enzyme, this has proven difficult. However, residues towards the exit of the active site are variable and can be exploited to design isoform selective inhibitors. Here we discuss and characterize this region of “selective drug targetability” and how these observations can be utilized to develop isoform selective CA inhibitors. Melissa A. Pinard, Brian Mahon, and Robert McKenna Copyright © 2015 Melissa A. Pinard et al. All rights reserved. Competing Role of Bioactive Constituents in Moringa oleifera Extract and Conventional Nutrition Feed on the Performance of Cobb 500 Broilers Sun, 22 Feb 2015 14:12:34 +0000 The influence of Moringa oleifera (MO) leaf extract as a dietary supplement on the growth performance and antioxidant parameters was evaluated on broiler meat and the compounds responsible for the corresponding antioxidant activity were identified. 0.5%, 1.0%, and 1.5% w/v of MO leaf aqueous extracts (MOLE) were prepared, and nutritional feed supplemented with 0%, 0.5%, 1.0%, and 1.5% w/w of MO leaf meal (MOLM) extracts were also prepared and analysed for their in vitro antioxidant potential. Furthermore, the treated broiler groups (control (T1) and treatment (T2, T3, and T4)) were evaluated for performance, meat quality, and antioxidant status. The results of this study revealed that, among the broilers fed MOLM, the broilers fed 0.5% w/w MOLM (T2) exhibited enhanced meat quality and antioxidant status (). However, the antioxidant activity of the MOLE is greater than that of the MOLM. The LC-MS/MS analysis of MOLM showed high expression of isoflavones and fatty acids from soy and corn source, which antagonistically inhibit the expression of the flavonoids/phenols in the MO leaves thereby masking its antioxidant effects. Thus, altering the soy and corn gradients in conventional nutrition feed with 0.5% w/w MO leaves supplement would provide an efficient and cost-effective feed supplement. Govindarajan Karthivashan, Palanisamy Arulselvan, Abd. Razak Alimon, Intan Safinar Ismail, and Sharida Fakurazi Copyright © 2015 Govindarajan Karthivashan et al. All rights reserved. Binding of Carbonic Anhydrase IX to 45S rDNA Genes Is Prevented by Exportin-1 in Hypoxic Cells Sun, 22 Feb 2015 08:54:21 +0000 Carbonic anhydrase IX (CA IX) is a surrogate marker of hypoxia, involved in survival and pH regulation in hypoxic cells. We have recently characterized its interactome, describing a set of proteins interacting with CA IX, mainly in hypoxic cells, including several members of the nucleocytoplasmic shuttling apparatuses. Accordingly, we described complex subcellular localization for this enzyme in human cells, as well as the redistribution of a carbonic anhydrase IX pool to nucleoli during hypoxia. Starting from this evidence, we analyzed the possible contribution of carbonic anhydrase IX to transcription of the 45S rDNA genes, a process occurring in nucleoli. We highlighted the binding of carbonic anhydrase IX to nucleolar chromatin, which is regulated by oxygen levels. In fact, CA IX was found on 45S rDNA gene promoters in normoxic cells and less represented on these sites, in hypoxic cells and in cells subjected to acetazolamide-induced acidosis. Both conditions were associated with increased representation of carbonic anhydrase IX/exportin-1 complexes in nucleoli. 45S rRNA transcript levels were accordingly downrepresented. Inhibition of nuclear export by leptomycin B suggests a model in which exportin-1 acts as a decoy, in hypoxic cells, preventing carbonic anhydrase IX association with 45S rDNA gene promoters. Emanuele Sasso, Monica Vitale, Francesca Monteleone, Francesca Ludovica Boffo, Margherita Santoriello, Daniela Sarnataro, Corrado Garbi, Mariangela Sabatella, Bianca Crifò, Luca Alfredo Paolella, Giuseppina Minopoli, Jean-Yves Winum, and Nicola Zambrano Copyright © 2015 Emanuele Sasso et al. All rights reserved. Food Bioactive Compounds against Diseases of the 21st Century Sun, 22 Feb 2015 08:23:24 +0000 Chia-Chien Hsieh, Juliana Maria Leite Nobrega de Moura Bell, and Blanca Hernández-Ledesma Copyright © 2015 Chia-Chien Hsieh et al. All rights reserved. Alpha-Linolenic Acid: An Omega-3 Fatty Acid with Neuroprotective Properties—Ready for Use in the Stroke Clinic? Thu, 19 Feb 2015 09:30:14 +0000 Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must be obtained through the diet. This could explain in part why the severe deficiency in omega-3 intake pointed by numerous epidemiologic studies may increase the brain’s vulnerability representing an important risk factor in the development and/or deterioration of certain cardio- and neuropathologies. The roles of ALA in neurological disorders remain unclear, especially in stroke that is a leading cause of death. We and others have identified ALA as a potential nutraceutical to protect the brain from stroke, characterized by its pleiotropic effects in neuroprotection, vasodilation of brain arteries, and neuroplasticity. This review highlights how chronic administration of ALA protects against rodent models of hypoxic-ischemic injury and exerts an anti-depressant-like activity, effects that likely involve multiple mechanisms in brain, and may be applied in stroke prevention. One major effect may be through an increase in mature brain-derived neurotrophic factor (BDNF), a widely expressed protein in brain that plays critical roles in neuronal maintenance, and learning and memory. Understanding the precise roles of ALA in neurological disorders will provide the underpinnings for the development of new therapies for patients and families who could be devastated by these disorders. Nicolas Blondeau, Robert H. Lipsky, Miled Bourourou, Mark W. Duncan, Philip B. Gorelick, and Ann M. Marini Copyright © 2015 Nicolas Blondeau et al. All rights reserved. Milk Proteins, Peptides, and Oligosaccharides: Effects against the 21st Century Disorders Thu, 19 Feb 2015 08:08:03 +0000 Milk is the most complete food for mammals, as it supplies all the energy and nutrients needed for the proper growth and development of the neonate. Milk is a source of many bioactive components, which not only help meeting the nutritional requirements of the consumers, but also play a relevant role in preventing various disorders. Milk-derived proteins and peptides have the potential to act as coadjuvants in conventional therapies, addressing cardiovascular diseases, metabolic disorders, intestinal health, and chemopreventive properties. In addition to being a source of proteins and peptides, milk contains complex oligosaccharides that possess important functions related to the newborn’s development and health. Some of the health benefits attributed to milk oligosaccharides include prebiotic probifidogenic effects, antiadherence of pathogenic bacteria, and immunomodulation. This review focuses on recent findings demonstrating the biological activities of milk peptides, proteins, and oligosaccharides towards the prevention of diseases of the 21st century. Processing challenges hindering large-scale production and commercialization of those bioactive compounds have been also addressed. Chia-Chien Hsieh, Blanca Hernández-Ledesma, Samuel Fernández-Tomé, Valerie Weinborn, Daniela Barile, and Juliana María Leite Nobrega de Moura Bell Copyright © 2015 Chia-Chien Hsieh et al. All rights reserved. Genome-Wide Methylome Analyses Reveal Novel Epigenetic Regulation Patterns in Schizophrenia and Bipolar Disorder Wed, 04 Feb 2015 08:59:12 +0000 Schizophrenia (SZ) and bipolar disorder (BP) are complex genetic disorders. Their appearance is also likely informed by as yet only partially described epigenetic contributions. Using a sequencing-based method for genome-wide analysis, we quantitatively compared the blood DNA methylation landscapes in SZ and BP subjects to control, both in an understudied population, Hispanics along the US-Mexico border. Remarkably, we identified thousands of differentially methylated regions for SZ and BP preferentially located in promoters 3′-UTRs and 5′-UTRs of genes. Distinct patterns of aberrant methylation of promoter sequences were located surrounding transcription start sites. In these instances, aberrant methylation occurred in CpG islands (CGIs) as well as in flanking regions as well as in CGI sparse promoters. Pathway analysis of genes displaying these distinct aberrant promoter methylation patterns showed enhancement of epigenetic changes in numerous genes previously related to psychiatric disorders and neurodevelopment. Integration of gene expression data further suggests that in SZ aberrant promoter methylation is significantly associated with altered gene transcription. In particular, we found significant associations between (1) promoter CGIs hypermethylation with gene repression and (2) CGI 3′-shore hypomethylation with increased gene expression. Finally, we constructed a specific methylation analysis platform that facilitates viewing and comparing aberrant genome methylation in human neuropsychiatric disorders. Yongsheng Li, Cynthia Camarillo, Juan Xu, Tania Bedard Arana, Yun Xiao, Zheng Zhao, Hong Chen, Mercedes Ramirez, Juan Zavala, Michael A. Escamilla, Regina Armas, Ricardo Mendoza, Alfonso Ontiveros, Humberto Nicolini, Alvaro Antonio Jerez Magaña, Lewis P. Rubin, Xia Li, and Chun Xu Copyright © 2015 Yongsheng Li et al. All rights reserved. Bioenergetics and the Role of Soluble Cytochromes c for Alkaline Adaptation in Gram-Negative Alkaliphilic Pseudomonas Mon, 02 Feb 2015 07:29:37 +0000 Very few studies have been conducted on alkaline adaptation of Gram-negative alkaliphiles. The reversed difference of H+ concentration across the membrane will make energy production considerably difficult for Gram-negative as well as Gram-positive bacteria. Cells of the alkaliphilic Gram-negative bacterium Pseudomonas alcaliphila AL15-21T grown at pH 10 under low-aeration intensity have a soluble cytochrome c content that is 3.6-fold higher than that of the cells grown at pH 7 under high-aeration intensity. Cytochrome c-552 content was higher (64% in all soluble cytochromes c) than those of cytochrome c-554 and cytochrome c-551. In the cytochrome c-552-dificient mutant grown at pH 10 under low-aeration intensity showed a marked decrease in [h−1] (40%) and maximum cell turbidity (25%) relative to those of the wild type. Considering the high electron-retaining abilities of the three soluble cytochromes c, the deteriorations in the growth of the cytochrome c-552-deficient mutant could be caused by the soluble cytochromes c acting as electron storages in the periplasmic space of the bacterium. These electron-retaining cytochromes c may play a role as electron and H+ condenser, which facilitate terminal oxidation at high pH under air-limited conditions, which is difficult to respire owing to less oxygen and less H+. T. Matsuno and I. Yumoto Copyright © 2015 T. Matsuno and I. Yumoto. All rights reserved. The Effect of Walterinnesia aegyptia Venom Proteins on TCA Cycle Activity and Mitochondrial NAD+-Redox State in Cultured Human Fibroblasts Sun, 01 Feb 2015 12:19:37 +0000 Fibroblast cultures were used to study the effects of crude Walterinnesia aegyptia venom and its F1–F7 protein fractions on TCA cycle enzyme activities and mitochondrial NAD-redox state. Confluent cells were incubated with 10 μg of venom proteins for 4 hours at 37°C. The activities of all studied TCA enzymes and the non-TCA mitochondrial NADP+-dependent isocitrate dehydrogenase underwent significant reductions of similar magnitude (50–60% of control activity) upon incubation of cells with the crude venom and fractions F4, F5, and F7 and 60–70% for fractions F3 and F6. In addition, the crude and fractions F3–F7 venom proteins caused a drop in mitochondrial NAD+ and NADP+ levels equivalent to around 25% of control values. Whereas the crude and fractions F4, F5, and F7 venom proteins caused similar magnitude drops in NADH and NADPH (around 55% of control levels), fractions F3 and F6 caused a more drastic drop (60–70% of control levels) of both reduced coenzymes. Results indicate that the effects of venom proteins could be directed at the mitochondrial level and/or the rates of NAD+ and NADP+ biosynthesis. Hazem K. Ghneim, Yazeed A. Al-Sheikh, and Mourad A. M. Aboul-Soud Copyright © 2015 Hazem K. Ghneim et al. All rights reserved. Effects of Partial Substitution of Fish Meal by Soybean Meal with or without Heat-Killed Lactobacillus plantarum (LP20) on Growth Performance, Digestibility, and Immune Response of Amberjack, Seriola dumerili Juveniles Sun, 01 Feb 2015 12:17:03 +0000 A 56-day feeding trial was conducted to evaluate the effects of supplemented diets with heat-killed Lactobacillus plantarum (HK-LP) with graded levels of soybean meal (SBM) on growth, digestibility, blood parameters, and immune response of Seriola dumerili (initial weight, 25.05 ± 0.1 g). Seven isonitrogenous and isolipidic practical diets were formulated to contain 0%, 15%, 30%, and 45% SBM, and each SBM level was supplemented with HK-LP at 0.0 and 0.1%. Fish fed diet which contains 30% SBM with HK-LP grew significantly faster than the other groups with notable feed intake and protein retention. Further, protein gain, whole body protein content, protease activity, protein, and lipid digestibility were significantly increased for all fish groups except for fish fed diet which contains 45% SBM with or without HK-LP. Interestingly, lysozyme activity was significantly enhanced in fish fed diets that contain 15% and 30% SBM with HK-LP. Hematocrit, peroxidase, and bactericidal activities revealed a significant increase in 30% SBM with HK-LP group. In addition, fish fed diets which contain 0% and 30% SBM with HK-LP showed higher tolerance against low-salinity stress compared with other groups. In conclusion, the addition of HK-LP to amberjack diets appeared to improve SBM utilization, immune response, and stress resistance. Mahmoud A. O. Dawood, Shunsuke Koshio, Manabu Ishikawa, and Saichiro Yokoyama Copyright © 2015 Mahmoud A. O. Dawood et al. All rights reserved. Novel Insights on Nutrient Management of Sarcopenia in Elderly Thu, 29 Jan 2015 07:06:54 +0000 Sarcopenia is defined as a syndrome characterized by progressive and generalized loss of muscle mass and strength. The more rationale approach to delay the progression of sarcopenia is based on the combination of proper nutrition, possibly associated with the use of dietary supplements and a regular exercise program. We performed a narrative literature review to evaluate the till-now evidence regarding (1) the metabolic and nutritional correlates of sarcopenia; (2) the optimum diet therapy for the treatment of these abnormalities. This review included 67 eligible studies. In addition to the well recognized link between adequate intake of proteins/amino acids and sarcopenia, the recent literature underlines that in sarcopenic elderly subjects there is an unbalance in vitamin D synthesis and in omega-6/omega-3 PUFA ratio. Given the detrimental effect of these metabolic abnormalities, a change in the lifestyle must be the cornerstone in the treatment of sarcopenia. The optimum diet therapy for the sarcopenia treatment must aim at achieving specific metabolic goals, which must be reached through accession of the elderly to specific personalized dietary program aimed at achieving and/or maintaining muscle mass; increasing their intake of fish (4 times/week) or taking omega-3 PUFA supplements; taking vitamin D supplementation, if there are low serum levels. Mariangela Rondanelli, Milena Faliva, Francesca Monteferrario, Gabriella Peroni, Erica Repaci, Francesca Allieri, and Simone Perna Copyright © 2015 Mariangela Rondanelli et al. All rights reserved. Comparison between the Traditional and a Rapid Screening Test for Cryoimmunoglobulins Detection Mon, 26 Jan 2015 14:27:13 +0000 Objectives. A new rapid, automatic, and sensitive screening test useful to detect cryoglobulins in serum samples is proposed. Design and Methods. The increase of turbidity during the cryoglobulin aggregation was monitored spectrophotometrically in sera from 400 patients with clinical evidence of cryoglobulinemia related disorders and 100 controls. Results were correlated to those obtained by the traditional method. Results. Kinetics of the aggregation curves were described by their maximum turbidity increase, lag time, and slope. Despite a partial correspondence between the traditional and the rapid test, patients with symptomatic cryoglobulinemia showed turbidity values significantly higher than the determined cutoff. Moreover, a functional classification of cryoglobulins is proposed. Conclusions. Due to its high reproducibility, operator independence, low cost, and results obtained within 2 hours, the rapid test can be used as a “real time” monitoring of cryoglobulinemia related diseases and for the evaluation of plasmapheresis efficacy. Federica Romitelli, Leopoldo Paolo Pucillo, Umberto Basile, and Enrico Di Stasio Copyright © 2015 Federica Romitelli et al. All rights reserved. Synthetic Covalently Linked Dimeric Form of H2 Relaxin Retains Native RXFP1 Activity and Has Improved In Vitro Serum Stability Thu, 22 Jan 2015 12:31:29 +0000 Human (H2) relaxin is a two-chain peptide member of the insulin superfamily and possesses potent pleiotropic roles including regulation of connective tissue remodeling and systemic and renal vasodilation. These effects are mediated through interaction with its cognate G-protein-coupled receptor, RXFP1. H2 relaxin recently passed Phase III clinical trials for the treatment of congestive heart failure. However, its in vivo half-life is short due to its susceptibility to proteolytic degradation and renal clearance. To increase its residence time, a covalent dimer of H2 relaxin was designed and assembled through solid phase synthesis of the two chains, including a judiciously monoalkyne sited B-chain, followed by their combination through regioselective disulfide bond formation. Use of a bisazido PEG7 linker and “click” chemistry afforded a dimeric H2 relaxin with its active site structurally unhindered. The resulting peptide possessed a similar secondary structure to the native monomeric H2 relaxin and bound to and activated RXFP1 equally well. It had fewer propensities to activate RXFP2, the receptor for the related insulin-like peptide 3. In human serum, the dimer had a modestly increased half-life compared to the monomeric H2 relaxin suggesting that additional oligomerization may be a viable strategy for producing longer acting variants of H2 relaxin. Vinojini B. Nair, Ross A. D. Bathgate, Frances Separovic, Chrishan S. Samuel, Mohammed Akhter Hossain, and John D. Wade Copyright © 2015 Vinojini B. Nair et al. All rights reserved. Receptor Binding Domain Based HIV Vaccines Thu, 15 Jan 2015 10:14:37 +0000 This paper analyzes the main trend of the development of acquired immunodeficiency syndrome (AIDS) vaccines in recent years. Designing an HIV-1 vaccine that provides robust protection from HIV-1 infection remains a challenge despite many years of effort. Therefore, we describe the receptor binding domain of gp120 as a target for developing AIDS vaccines. And we recommend some measures that could induce efficiently and produce cross-reactive neutralizing antibodies with high binding affinity. Those measures may offer a new way of the research and development of the potent and broad AIDS vaccines. Huan Liu, Wenwen Bi, Qian Wang, Lu Lu, and Shibo Jiang Copyright © 2015 Huan Liu et al. All rights reserved. In Vitro Bactericidal Activity of 4- and 5-Chloro-2-hydroxy-N-[1-oxo-1-(phenylamino)alkan-2-yl]benzamides against MRSA Thu, 15 Jan 2015 06:27:15 +0000 A series of nine substituted 2-hydroxy-N-[1-oxo-1-(phenylamino)alkan-2-yl]benzamides was assessed as prospective bactericidal agents against three clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus ATCC 29213 as the reference and quality control strain. The minimum bactericidal concentration was determined by subculturing aliquots from MIC determination onto substance-free agar plates. The bactericidal kinetics of compounds 5-chloro-2-hydroxy-N-[(2S)-3-methyl-1-oxo-1-[4-(trifluoromethyl)phenyl]aminobutan-2-yl]benzamide (1f), N-(2S)-1-[(4-bromophenyl)amino]-3-methyl-1-oxobutan-2-yl-4-chloro-2-hydroxybenzamide (1g), and 4-chloro-N-(2S)-1-[(3,4-dichlorophenyl)amino]-3-methyl-1-oxobutan-2-yl-2-hydroxybenzamide (1h) was established by time-kill assay with a final concentration of the compound equal to 1x, 2x, and 4x MIC; aliquots were removed at 0, 4, 6, 8, and 24 h time points. The most potent bactericidal agent was compound 1f exhibiting remarkable rapid concentration-dependent bactericidal effect even at 2x MIC at 4, 6, and 8 h (with a reduction in bacterial count ranging from 3.08 to 3.75  CFU/mL) and at 4x MIC at 4, 6, 8, and 24 h (5.30  CFU/mL reduction in bacterial count) after incubation against MRSA 63718. Reliable bactericidal effect against other strains was maintained at 4x MIC at 24 h. Iveta Zadrazilova, Sarka Pospisilova, Karel Pauk, Ales Imramovsky, Jarmila Vinsova, Alois Cizek, and Josef Jampilek Copyright © 2015 Iveta Zadrazilova et al. All rights reserved. Beneficial Effect of Sugar Osmolytes on the Refolding of Guanidine Hydrochloride-Denatured Trehalose-6-phosphate Hydrolase from Bacillus licheniformis Tue, 13 Jan 2015 06:49:07 +0000 The influence of three sugar osmolytes on the refolding of guanidine hydrochloride- (GdnHCl-) denatured trehalose-6-phosphate hydrolase of Bacillus licheniformis (BlTreA) was studied by circular dichroism (CD) spectra, fluorescence emission spectra, and the recovery of enzymatic activity. These experimental results clearly indicated that sorbitol, sucrose, and trehalose at a concentration of 0.75 M improved the refolding yields of GdnHCl-denatured  BlTreA, probably due to the fact that these sugars favored the formation of tertiary architectures. Far-UV CD measurements demonstrated the ability of sugar osmolytes to shift the secondary structure of GdnHCl-denatured enzyme towards near-native conformations. ANS fluorescence intensity measurements revealed a reduction of exposed hydrophobic surfaces upon the treatment of denatured enzyme with sugar osmolytes. These observations suggest that sugar osmolytes possibly play a chaperone role in the refolding of chemically denatured BlTreA. Jiau-Hua Chen, Meng-Chun Chi, Min-Guan Lin, Long-Liu Lin, and Tzu-Fan Wang Copyright © 2015 Jiau-Hua Chen et al. All rights reserved. Relationship between Hyperuricemia and Lipid Profiles in US Adults Mon, 05 Jan 2015 13:59:29 +0000 Background. Although the link between hyperuricemia and metabolic syndrome had been recognized, the association of the dyslipidemia among individuals with hyperuricemia remains not comprehensively assessed. Methods. Using NHANES III study, we examined the relation between serum lipid profiles and different serum uric acid levels, including serum total cholesterol, LDL cholesterol, triglycerides, HDL cholesterol, apolipoprotein-B, lipoprotein (a), apolipoprotein AI, ratio of triglycerides to HDL cholesterol, and ratio of apolipoprotein-B to AI. Results. After adjusting for potential confounders, average differences (95% confidence interval) comparing the top to the bottom (reference) serum uric acid were 0.29 (0.19, 0.39) mmol/L for total cholesterol, 0.33 (0.26, 0.41) mmol/L for triglycerides, 0.14 (0.01, 0.27) mmol/L for LDL cholesterol, −0.08 (−0.11, −0.05) mmol/L for HDL, and 0.09 (0.05, 0.12) g/L for serum apolipoprotein-B. Notably, ratios of triglycerides to HDL cholesterol and apolipoprotein-B to AI were also linearly associated with uric acid levels (P for trend < 0.001). Conclusions. This study suggested that serum LDL cholesterol, triglycerides, total cholesterol, apolipoprotein-B levels, ratio of triglycerides to HDL cholesterol, and ratio of apolipoprotein-B to AI are strongly associated with serum uric acid levels, whereas serum HDL cholesterol levels are significantly inversely associated. In the clinical practice, the more comprehensive strategic management to deal with dyslipidemia and hyperuricemia deserves further investigation. Tao-Chun Peng, Chung-Ching Wang, Tung-Wei Kao, James Yi-Hsin Chan, Ya-Hui Yang, Yaw-Wen Chang, and Wei-Liang Chen Copyright © 2015 Tao-Chun Peng et al. All rights reserved. Aberrant Hypermethylation of Aldehyde Dehydrogenase 2 Promoter Upstream Sequence in Rats with Experimental Myocardial Infarction Mon, 05 Jan 2015 13:07:55 +0000 Background. Aldehyde dehydrogenase 2 (ALDH2) plays a crucial role in myocardial protection against ischemia. Downregulation of ALDH2 was evidenced after myocardial infarction and the underlying mechanism is not fully understood. DNA methylation can regulate gene transcription in epigenetic level. We thus hypothesized that DNA methylation may affect ALDH2 expression in myocardial infarction (MI). Methods. MI was induced in Sprague-Dawley rats. MI border zone tissues were harvested at 1st week, 2nd week, and 3rd week after MI. Bisulfite sequencing PCR (BSP) was performed to detect the methylation levels of ALDH2 core promoter. Sequenom MassARRAY platform (MassARRAY) was used to examine the methylation levels of ALDH2 promoter upstream sequence. ALDH2 protein and mRNA expression were assayed by Western blot and real-time PCR, respectively. Results. Compared with Sham group, ALDH2 protein and mRNA expression of MI groups was significantly downregulated. Compared with Sham group, DNA methylation level of CpG sites in ALDH2 promoter upstream sequence was significantly higher in MI groups in a time-dependent manner (CpG1, CpG2, and CpG7, ). DNA methylation did not affect ALDH2 core promoter sequence during the progress of MI. No significant difference was detected in DNA methylation level of ALDH2 promoter upstream sequence among MI groups. Conclusion. Aberrant hypermethylation of CpG sites in ALDH2 promoter upstream sequence is associated with myocardial ischemia injury and may partly result in ALDH2 downregulation after MI. Peng Wang, Cheng Shen, Lei Diao, Zhiyin Yang, Fan Fan, Cong Wang, Xiangwei Liu, Xiaolei Sun, Zhen Dong, Hong Zhu, Xin Ma, Quan Cao, Xiaona Zhao, Duan Ma, Yunzeng Zou, Kai Hu, Aijun Sun, and Junbo Ge Copyright © 2015 Peng Wang et al. All rights reserved. The Antidiabetic Effect of Low Doses of Moringa oleifera Lam. Seeds on Streptozotocin Induced Diabetes and Diabetic Nephropathy in Male Rats Mon, 05 Jan 2015 06:50:54 +0000 The antidiabetic activity of two low doses of Moringa seed powder (50 and 100 mg/kg body weight, in the diet) on streptozotocin (STZ) induced diabetes male rats was investigated. Forty rats were divided into four groups. The diabetic positive control (STZ treated) group showed increased lipid peroxide, increased IL-6, and decreased antioxidant enzyme in the serum and kidney tissue homogenate compared with that of the negative control group. Immunoglobulins (IgA, IgG), fasting blood sugar, and glycosylated hemoglobin were also increased as a result of diabetes in G2 rats. Moreover albumin was decreased, and liver enzymes and α-amylase were not affected. In addition, the renal functions and potassium and sodium levels in G2 were increased as a sign of diabetic nephropathy. Urine analysis showed also glucosuria and increased potassium, sodium, creatinine, uric acid, and albumin levels. Kidney and pancreas tissues showed also pathological alteration compared to the negative control group. Treating the diabetic rats with 50 or 100 mg Moringa seeds powder/kg body weight in G3 and G4, respectively, ameliorated the levels of all these parameters approaching the negative control values and restored the normal histology of both kidney and pancreas compared with that of the diabetic positive control group. Abdulrahman L. Al-Malki and Haddad A. El Rabey Copyright © 2015 Abdulrahman L. Al-Malki and Haddad A. El Rabey. All rights reserved. Association Analysis between g.18873C>T and g.27522G>A Genetic Polymorphisms of OPG and Bone Mineral Density in Chinese Postmenopausal Women Mon, 15 Dec 2014 00:10:59 +0000 Several studies report that the OPG is an important candidate gene in the pathogenesis of osteoporosis. This study aimed to detect the potential association of OPG gene polymorphisms with osteoporosis in postmenopausal women. We recruited 928 subjects containing 463 with primary postmenopausal osteoporosis and 465 healthy volunteers as controls. The BMD of neck hip, lumbar spine (), and total hip were assessed by dual-energy X-ray absorptiometry (DEXA). Through the created restriction site-polymerase chain reaction (CRS-PCR), PCR-restriction fragment length polymorphism (PCR-RFLP), and DNA sequencing methods, the g.18873C>T and g.27522G>A have been investigated. As for g.18873C>T, our data indicated that subjects with CC genotype have significantly higher BMD value than those of CT and TT genotypes (all values < 0.05). As for g.27522G>A, the BMD values of subjects with GG genotype were significantly higher than those of GA and AA genotypes (all values < 0.05). Our findings suggest that the OPG g.18873C>T and g.27522G>A genetic polymorphisms are associated with the decreased risk for osteoporosis in Chinese postmenopausal women. Fei Wang, Yi Cao, Fang Li, Jianlin Shan, and Tianlin Wen Copyright © 2014 Fei Wang et al. All rights reserved. Erratum to “A Comparative Study for the Evaluation of Two Doses of Ellagic Acid on Hepatic Drug Metabolizing and Antioxidant Enzymes in the Rat” Mon, 01 Dec 2014 07:59:58 +0000 Gurbet Celik, Aslı Semiz, Serdar Karakurt, Sevki Arslan, Orhan Adali, and Alaattin Sen Copyright © 2014 Gurbet Celik et al. All rights reserved. Carborane-Based Carbonic Anhydrase Inhibitors: Insight into CAII/CAIX Specificity from a High-Resolution Crystal Structure, Modeling, and Quantum Chemical Calculations Thu, 18 Sep 2014 08:08:35 +0000 Carborane-based compounds are promising lead structures for development of inhibitors of carbonic anhydrases (CAs). Here, we report structural and computational analysis applicable to structure-based design of carborane compounds with selectivity toward the cancer-specific CAIX isoenzyme. We determined the crystal structure of CAII in complex with 1-methylenesulfamide-1,2-dicarba-closo-dodecaborane at 1.0 Å resolution and used this structure to model the 1-methylenesulfamide-1,2-dicarba-closo-dodecaborane interactions with CAIX. A virtual glycine scan revealed the contributions of individual residues to the energy of binding of 1-methylenesulfamide-1,2-dicarba-closo-dodecaborane to CAII and CAIX, respectively. Pavel Mader, Adam Pecina, Petr Cígler, Martin Lepšík, Václav Šícha, Pavel Hobza, Bohumír Grüner, Jindřich Fanfrlík, Jiří Brynda, and Pavlína Řezáčová Copyright © 2014 Pavel Mader et al. All rights reserved. Inhibition of Nek2 by Small Molecules Affects Proteasome Activity Wed, 17 Sep 2014 00:00:00 +0000 Background. Nek2 is a serine/threonine kinase localized to the centrosome. It promotes cell cycle progression from G2 to M by inducing centrosome separation. Recent studies have shown that high Nek2 expression is correlated with drug resistance in multiple myeloma patients. Materials and Methods. To investigate the role of Nek2 in bortezomib resistance, we ectopically overexpressed Nek2 in several cancer cell lines, including multiple myeloma lines. Small-molecule inhibitors of Nek2 were discovered using an in-house library of compounds. We tested the inhibitors on proteasome and cell cycle activity in several cell lines. Results. Proteasome activity was elevated in Nek2-overexpressing cell lines. The Nek2 inhibitors inhibited proteasome activity in these cancer cell lines. Treatment with these inhibitors resulted in inhibition of proteasome-mediated degradation of several cell cycle regulators in HeLa cells, leaving them arrested in G2/M. Combining these Nek2 inhibitors with bortezomib increased the efficacy of bortezomib in decreasing proteasome activity in vitro. Treatment with these novel Nek2 inhibitors successfully mitigated drug resistance in bortezomib-resistant multiple myeloma. Conclusion. Nek2 plays a central role in proteasome-mediated cell cycle regulation and in conferring resistance to bortezomib in cancer cells. Taken together, our results introduce Nek2 as a therapeutic target in bortezomib-resistant multiple myeloma. Lingyao Meng, Kent Carpenter, Alexis Mollard, Hariprasad Vankayalapati, Steven L. Warner, Sunil Sharma, Guido Tricot, Fenghuang Zhan, and David J. Bearss Copyright © 2014 Lingyao Meng et al. All rights reserved. Antioxidant/Prooxidant and Antibacterial/Probacterial Effects of a Grape Seed Extract in Complex with Lipoxygenase Mon, 15 Sep 2014 05:36:09 +0000 In an attempt to determine the antioxidant/prooxidant, antibacterial/probacterial action of flavan-3-ols and procyanidins from grape seeds, pure catechin (CS), and an aqueous grape seed extract (PE), were applied in the absence and presence of pure lipoxygenase (LS) or in extract (LE) to leucocyte culture, Escherichia coli B41 and Brevibacterium linens, and observed whether there was any effect on lipid peroxidation, cytotoxicity, or growth rate. Short time periods of coincubation of cells with the polyphenols, followed by the exposure to LS and LE, revealed a high level of lipid peroxidation and a prooxidative effect. Longer coincubation and addition of LS and LE resulted in the reversal of the prooxidant action either to antioxidant activity for CS + LS and PE + LS or to the control level for CS + LE and PE + LE. Lipid peroxidation was significantly reduced when cells were exposed to polyphenols over a longer period. Longer exposure of E. coli to CS or PE followed by addition of LS for 3 h resulted in bactericidal activity. Significant stimulatory effect on microbial growth was observed for PE + LS and PE + LE treatments in B. linens, illustrating the potential probacterial activity in B. linens cultures. Lipoxygenase-polyphenols complex formation was found to be responsible for the observed effects. Veronica Sanda Chedea, Cornelia Braicu, Flore Chirilă, Henry Joseph Oduor Ogola, Rodica Ştefania Pelmuş, Loredana Georgeta Călin, and Carmen Socaciu Copyright © 2014 Veronica Sanda Chedea et al. All rights reserved. Structural Basis of Binding and Rationale for the Potent Urease Inhibitory Activity of Biscoumarins Tue, 09 Sep 2014 11:39:30 +0000 Urease belongs to a family of highly conserved urea-hydrolyzing enzymes. A common feature of these enzymes is the presence of two Lewis acid nickel ions and reactive cysteine residue in the active sites. In the current study we examined a series of biscoumarins 1–10 for their mechanisms of inhibition with the nickel containing active sites of Jack bean and Bacillus pasteurii ureases. All these compounds competitively inhibited Jack bean urease through interaction with the nickel metallocentre, as deduced from Michaelis-Menten kinetics, UV-visible absorbance spectroscopic, and molecular docking simulation studies. Some of the compounds behaved differently in case of Bacillus pasteurii urease. We conducted the enzyme kinetics, UV-visible spectroscopy, and molecular docking results in terms of the known protein structure of the enzyme. We also evaluated possible molecular interpretations for the site of biscoumarins binding and found that phenyl ring is the major active pharmacophore. The excellent in vitro potency and selectivity profile of the several compounds described combined with their nontoxicity against the human cells and plants suggest that these compounds may represent a viable lead series for the treatment of urease associated problems. Muhammad Arif Lodhi, Sulaiman Shams, Muhammad Iqbal Choudhary, Atif Lodhi, Zaheer Ul-Haq, Saima Jalil, Sarfraz Ahmad Nawaz, Khalid Mohammed Khan, Sajid Iqbal, and Atta-ur Rahman Copyright © 2014 Muhammad Arif Lodhi et al. All rights reserved. Sulfa Drugs as Inhibitors of Carbonic Anhydrase: New Targets for the Old Drugs Mon, 08 Sep 2014 11:03:09 +0000 Sulfa drugs are well-known antibacterial agents containing N-substituted sulfonamide group on para position of aniline ring (NH2RSO2NHR′). In this study 2,4-dichloro-1,3,5-triazine derivatives of sulfa drugs, sulfamerazine (1b), sulfaquinoxaline (2b), sulfadiazine (3b), sulfadimidine (4b), and sulfachloropyrazine (5b) (1a–5a) were synthesized and characterized. Their carbonic anhydrase inhibition activity was evaluated against bovine cytosolic carbonic anhydrase isozyme II (bCA II). For the sake of comparison the CA inhibition activity of the parent sulfa drugs (1b–5b) was also evaluated. A significant increase in CA inhibition activity of sulfa drugs was observed upon substitution with 2,4-dichloro-1,3,5-triazine moiety. Molecular docking studies were carried out to highlight binding site interactions. ADME properties were calculated to evaluate drug likeness of the compounds. Mariya al-Rashida, Sajad Hussain, Mehwish Hamayoun, Aisha Altaf, and Jamshed Iqbal Copyright © 2014 Mariya al-Rashida et al. All rights reserved. Saliva-Based Biosensors: Noninvasive Monitoring Tool for Clinical Diagnostics Mon, 08 Sep 2014 09:37:12 +0000 Saliva is increasingly recognised as an attractive diagnostic fluid. The presence of various disease signalling salivary biomarkers that accurately reflect normal and disease states in humans and the sampling benefits compared to blood sampling are some of the reasons for this recognition. This explains the burgeoning research field in assay developments and technological advancements for the detection of various salivary biomarkers to improve clinical diagnosis, management, and treatment. This paper reviews the significance of salivary biomarkers for clinical diagnosis and therapeutic applications, with focus on the technologies and biosensing platforms that have been reported for screening these biomarkers. Radha S. P. Malon, Sahba Sadir, Malarvili Balakrishnan, and Emma P. Córcoles Copyright © 2014 Radha S. P. Malon et al. All rights reserved. Saccharin Sulfonamides as Inhibitors of Carbonic Anhydrases I, II, VII, XII, and XIII Wed, 03 Sep 2014 10:55:10 +0000 A series of modified saccharin sulfonamides have been designed as carbonic anhydrase (CA) inhibitors and synthesized. Their binding to CA isoforms I, II, VII, XII, and XIII was measured by the fluorescent thermal shift assay (FTSA) and isothermal titration calorimetry (ITC). Saccharin bound the CAs weakly, exhibiting the affinities of 1–10 mM for four CAs except CA I where binding could not be detected. Several sulfonamide-bearing saccharines exhibited strong affinities of 1–10 nM towards particular CA isoforms. The functional group binding Gibbs free energy additivity maps are presented which may provide insights into the design of compounds with increased affinity towards selected CAs. Vaida Morkūnaitė, Lina Baranauskienė, Asta Zubrienė, Visvaldas Kairys, Jekaterina Ivanova, Pēteris Trapencieris, and Daumantas Matulis Copyright © 2014 Vaida Morkūnaitė et al. All rights reserved. Hydrophobic Substituents of the Phenylmethylsulfamide Moiety Can Be Used for the Development of New Selective Carbonic Anhydrase Inhibitors Tue, 02 Sep 2014 13:18:41 +0000 A new series of compounds containing a sulfamide moiety as zinc-binding group (ZBG) has been synthesized and tested for determining inhibitory properties against four human carbonic anhydrase (hCA) isoforms, namely, CAs I, II, IX, and XII. The X-ray structure of the cytosolic dominant isoform hCA II in complex with the best inhibitor of the series has also been determined providing further insights into sulfamide binding mechanism and confirming that such zinc-binding group, if opportunely derivatized, can be usefully exploited for obtaining new potent and selective CAIs. The analysis of the structure also suggests that for drug design purposes the but-2-yn-1-yloxy moiety tail emerges as a very interesting substituent of the phenylmethylsulfamide moiety due to its capability to establish strong van der Waals interactions with a hydrophobic cleft on the hCA II surface, delimited by residues Phe131, Val135, Pro202, and Leu204. Indeed, the complementarity of this tail with the cleft suggests that different substituents could be used to discriminate between isoforms having clefts with different sizes. Giuseppina De Simone, Ginta Pizika, Simona Maria Monti, Anna Di Fiore, Jekaterina Ivanova, Igor Vozny, Peteris Trapencieris, Raivis Zalubovskis, Claudiu T. Supuran, and Vincenzo Alterio Copyright © 2014 Giuseppina De Simone et al. All rights reserved. Role of Feed Forward Neural Networks Coupled with Genetic Algorithm in Capitalizing of Intracellular Alpha-Galactosidase Production by Acinetobacter sp. Sun, 31 Aug 2014 08:07:46 +0000 Alpha-galactosidase production in submerged fermentation by Acinetobacter sp. was optimized using feed forward neural networks and genetic algorithm (FFNN-GA). Six different parameters, pH, temperature, agitation speed, carbon source (raffinose), nitrogen source (tryptone), and K2HPO4, were chosen and used to construct 6-10-1 topology of feed forward neural network to study interactions between fermentation parameters and enzyme yield. The predicted values were further optimized by genetic algorithm (GA). The predictability of neural networks was further analysed by using mean squared error (MSE), root mean squared error (RMSE), mean absolute error (MAE), mean absolute percentage error (MAPE), and R2-value for training and testing data. Using hybrid neural networks and genetic algorithm, alpha-galactosidase production was improved from 7.5 U/mL to 10.2 U/mL. Sirisha Edupuganti, Ravichandra Potumarthi, Thadikamala Sathish, and Lakshmi Narasu Mangamoori Copyright © 2014 Sirisha Edupuganti et al. All rights reserved. Proteomics Analysis for Finding Serum Markers of Ovarian Cancer Sun, 31 Aug 2014 07:50:14 +0000 A combination of peptide ligand library beads (PLLB) and 1D gel liquid chromatography-mass spectrometry/mass spectrometry (1DGel-LC-MS/MS) was employed to analyze serum samples from patients with ovarian cancer and from healthy controls. Proteomic analysis identified 1200 serum proteins, among which 57 proteins were upregulated and 10 were downregulated in the sera from cancer patients. Retinol binding protein 4 (RBP4) is highly upregulated in the ovarian cancer serum samples. ELISA was employed to measure plasma concentrations of RBP4 in 80 samples from ovarian cancer patients, healthy individuals, myoma patients, and patients with benign ovarian tumor, respectively. The plasma concentrations of RBP4 ranging from 76.91 to 120.08 ng/mL with the mean value  ng/mL in ovarian cancer patients are significantly higher than those in healthy individuals ( ng/mL). Results were further confirmed with immunohistochemistry, demonstrating that RBP4 expression levels in normal ovarian tissue were lower than those in ovarian cancer tissues. Our results suggested that RBP4 is a potential biomarker for diagnostic of screening ovarian cancer. Yushan Cheng, Chongdong Liu, Nawei Zhang, Shengdian Wang, and Zhenyu Zhang Copyright © 2014 Yushan Cheng et al. All rights reserved. Destabilization of Akt Promotes the Death of Myeloma Cell Lines Sun, 31 Aug 2014 00:00:00 +0000 Constitutive activation of Akt is believed to be an oncogenic signal in multiple myeloma and is associated with poor patient prognosis and resistance to available treatment. The stability of Akt proteins is regulated by phosphorylating the highly conserved turn motif (TM) of these proteins and the chaperone protein HSP90. In this study we investigate the antitumor effects of inhibiting mTORC2 plus HSP90 in myeloma cell lines. We show that chronic exposure of cells to rapamycin can inhibit mTORC2 pathway, and AKT will be destabilized by administration of the HSP90 inhibitor 17-allylamino-geldanamycin (17-AAG). Finally, we show that the rapamycin synergizes with 17-AAG and inhibits myeloma cells growth and promotes cell death to a greater extent than either drug alone. Our studies provide a clinical rationale of use mTOR inhibitors and chaperone protein inhibitors in combination regimens for the treatment of human blood cancers. Yanan Zhang, Yunfeng Fu, Fan Zhang, and Jing Liu Copyright © 2014 Yanan Zhang et al. All rights reserved. A Novel Laccase with Potent Antiproliferative and HIV-1 Reverse Transcriptase Inhibitory Activities from Mycelia of Mushroom Coprinus comatus Thu, 28 Aug 2014 15:26:20 +0000 A novel laccase was isolated and purified from fermentation mycelia of mushroom Coprinus comatus with an isolation procedure including three ion-exchange chromatography steps on DEAE-cellulose, CM-cellulose, and Q-Sepharose and one gel-filtration step by fast protein liquid chromatography on Superdex 75. The purified enzyme was a monomeric protein with a molecular weight of 64 kDa. It possessed a unique N-terminal amino acid sequence of AIGPVADLKV, which has considerably high sequence similarity with that of other fungal laccases, but is different from that of C. comatus laccases reported. The enzyme manifested an optimal pH value of 2.0 and an optimal temperature of 60°C using 2,2′-azinobis(3-ethylbenzothiazolone-6-sulfonic acid) diammonium salt (ABTS) as the substrate. The laccase displayed, at pH 2.0 and 37°C, Km values of 1.59 mM towards ABTS. It potently suppressed proliferation of tumor cell lines HepG2 and MCF7, and inhibited human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) with an IC50 value of 3.46 μM, 4.95 μM, and 5.85 μM, respectively, signifying that it is an antipathogenic protein. Shuang Zhao, Cheng-Bo Rong, Chang Kong, Yu Liu, Feng Xu, Qian-Jiang Miao, Shou-Xian Wang, He-Xiang Wang, and Guo-Qing Zhang Copyright © 2014 Shuang Zhao et al. All rights reserved. Creatine, L-Carnitine, and ω3 Polyunsaturated Fatty Acid Supplementation from Healthy to Diseased Skeletal Muscle Thu, 28 Aug 2014 15:19:19 +0000 Myopathies are chronic degenerative pathologies that induce the deterioration of the structure and function of skeletal muscle. So far a definitive therapy has not yet been developed and the main aim of myopathy treatment is to slow the progression of the disease. Current nonpharmacological therapies include rehabilitation, ventilator assistance, and nutritional supplements, all of which aim to delay the onset of the disease and relieve its symptoms. Besides an adequate diet, nutritional supplements could play an important role in the treatment of myopathic patients. Here we review the most recent in vitro and in vivo studies investigating the role supplementation with creatine, L-carnitine, and ω3 PUFAs plays in myopathy treatment. Our results suggest that these dietary supplements could have beneficial effects; nevertheless continued studies are required before they could be recommended as a routine treatment in muscle diseases. Giuseppe D’Antona, Seyed Mohammad Nabavi, Piero Micheletti, Arianna Di Lorenzo, Roberto Aquilani, Enzo Nisoli, Mariangela Rondanelli, and Maria Daglia Copyright © 2014 Giuseppe D’Antona et al. All rights reserved. Symbiotic Plant Peptides Eliminate Candida albicans Both In Vitro and in an Epithelial Infection Model and Inhibit the Proliferation of Immortalized Human Cells Thu, 28 Aug 2014 09:43:50 +0000 The increasing number of multidrug-resistant microbes now emerging necessitates the identification of novel antimicrobial agents. Plants produce a great variety of antimicrobial peptides including hundreds of small, nodule-specific cysteine-rich NCR peptides that, in the legume Medicago truncatula, govern the differentiation of endosymbiotic nitrogen fixing bacteria and, in vitro, can display potent antibacterial activities. In this study, the potential candidacidal activity of 19 NCR peptides was investigated. Cationic NCR peptides having an isoelectric point above 9 were efficient in killing Candida albicans, one of the most common fungal pathogens of humans. None of the tested NCR peptides were toxic for immortalized human epithelial cells at concentrations that effectively killed the fungus; however, at higher concentrations, some of them inhibited the division of the cells. Furthermore, the cationic peptides successfully inhibited C. albicans induced human epithelial cell death in an in vitro coculture model. These results highlight the therapeutic potential of cationic NCR peptides in the treatment of candidiasis. Lilla Ördögh, Andrea Vörös, István Nagy, Éva Kondorosi, and Attila Kereszt Copyright © 2014 Lilla Ördögh et al. All rights reserved. Angiotensin I Converting Enzyme Inhibitory Peptides Obtained after In Vitro Hydrolysis of Pea (Pisum sativum var. Bajka) Globulins Thu, 28 Aug 2014 00:00:00 +0000 Pea seeds represent a valuable source of active compounds that may positively influence health. In this study, the pea globulins were digested in vitro under gastrointestinal condition and potentially bioaccessible angiotensin I converting enzyme (ACE) inhibitory peptides were identified. The degree of hydrolysis after pepsin, 14.42%, and pancreatin, 30.65%, were noted. The peptides with the highest ACE inhibitory properties were separated using ion exchange chromatography on DEAE-cellulose. Thirteen peptides fractions were obtained but only four showed potential antihypertensive properties. The highest inhibitory activity was determined for the fraction F8 (IC50 = 0.0014 mg/mL). This fraction was separated on Sephadex G10 and two peptide fractions were obtained. The peptides fraction (B) with the highest ACE inhibitory activity (IC50 = 0.073 mg/mL) was identified by ESI-MS/MS. The sequences of ACE inhibitory peptides were GGSGNY, DLKLP, GSSDNR, MRDLK, and HNTPSR. Based on Lineweaver-Burk plots for the fraction B, the kinetic parameters as , , and and mode of inhibition were determined. This fraction belongs to uncompetitive inhibitor of ACE activity. The seeds of pea are the source of precursor protein, which releases the ACE inhibitory peptides as a result of enzymatic hydrolysis. Anna Jakubczyk and Barbara Baraniak Copyright © 2014 Anna Jakubczyk and Barbara Baraniak. All rights reserved. Molecular Cloning and Biochemical Characterization of a Recombinant Sterol 3-O-Glucosyltransferase from Gymnema sylvestre R.Br. Catalyzing Biosynthesis of Steryl Glucosides Wed, 27 Aug 2014 07:17:06 +0000 Gymnema sylvestre R.Br., a pharmacologically important herb vernacularly called Gur-Mar (sugar eliminator), is widely known for its antidiabetic action. This property of the herb has been attributed to the presence of bioactive triterpene glycosides. Although some information regarding pharmacology and phytochemical profiles of the plant are available, no attempts have been made so far to decipher the biosynthetic pathway and key enzymes involved in biosynthesis of steryl glucosides. The present report deals with the identification and catalytic characterization of a glucosyltransferase, catalyzing biosynthesis of steryl glycosides. The full length cDNA (2572 bp) contained an open reading frame of 2106 nucleotides that encoded a 701 amino acid protein, falling into GT-B subfamily of glycosyltransferases. The GsSGT was expressed in Escherichia coli and biochemical characterization of the recombinant enzyme suggested its key role in the biosynthesis of steryl glucosides with catalytic preference for C-3 hydroxyl group of sterols. To our knowledge, this pertains to be the first report on cloning and biochemical characterization of a sterol metabolism gene from G. sylvestre R.Br. catalyzing glucosylation of a variety of sterols of biological origin from diverse organisms such as bacteria, fungi, and plants. Pragya Tiwari, Rajender Singh Sangwan, Asha, B. N. Mishra, Farzana Sabir, and Neelam S. Sangwan Copyright © 2014 Pragya Tiwari et al. All rights reserved. A Time-Dose Model to Quantify the Antioxidant Responses of the Oxidative Hemolysis Inhibition Assay (OxHLIA) and Its Extension to Evaluate Other Hemolytic Effectors Wed, 27 Aug 2014 00:00:00 +0000 The development of a convenient mathematical application for testing the antioxidant potential of standard and novel therapeutic agents is essential for the research community to perform evaluations in a more precise form. The in vitro oxidative hemolysis inhibition assay, despite its relevance for in vivo responses, lacks a proper mathematical model to quantify the responses. In this work, a simple nonlinear time-dose tool to test the effectiveness of antioxidant compounds is presented. The model was verified with available experimental data from the bibliography. The model helps to describe accurately the antioxidant response as a function of time and dose allowing comparisons between compounds. Its advantages are a simple application, provision of parametric estimates that characterize the response, simplification of the protocol, economization of experimental effort, and facilitation of rigorous comparisons among the effects of different compounds and experimental approaches. Finally, other effectors that may obstruct or be of interest for the antioxidant determination are also modeled in similar principles. Thus, the basis of more complex multivariable models is provided. In all experimental data fitted, the calculated parameters were always statistically significant, the equations prove to be consistent, and the correlation coefficient of determination was in all cases higher than 0.98. M. A. Prieto and J. A. Vázquez Copyright © 2014 M. A. Prieto and J. A. Vázquez. All rights reserved. Anti-Inflammatory Effects of Siegesbeckia orientalis Ethanol Extract in In Vitro and In Vivo Models Tue, 26 Aug 2014 13:01:25 +0000 This study aims to investigate the anti-inflammatory responses and mechanisms of Siegesbeckia orientalis ethanol extract (SOE). In cell culture experiments, RAW264.7 cells were pretreated with SOE and stimulated with lipopolysaccharide (LPS) for inflammatory mediators assay. In animal experiments, mice were tube-fed with SOE for 1 week, and s.c. injected with λ-carrageenan or i.p. injected with LPS to simulate inflammation. The degree of paw edema was assessed, and cytokine profile in sera and mouse survival were recorded. Data showed that SOE significantly reduced NO, IL-6, and TNF-α production in LPS-stimulated RAW264.7 cells. In vivo studies demonstrated that mice supplemented with 32 mg SOE/kg BW/day significantly lowered sera IL-6 level and resulted a higher survival rate compared to the control group (). Furthermore, SOE inhibited LPS-induced NF-κB activation by blocking the degradation of IκB-α. The SOE also reduced significantly the phosphorylation of ERK1/2, p38, and JNK in a dose-dependent manner. In summary, the in vitro and in vivo evidence indicate that SOE can attenuate acute inflammation by inhibiting inflammatory mediators via suppression of MAPKs- and NF-κB-dependent pathways. Yong-Han Hong, Li-Wen Weng, Chi-Chang Chang, Hsia-Fen Hsu, Chao-Ping Wang, Shih-Wei Wang, and Jer-Yiing Houng Copyright © 2014 Yong-Han Hong et al. All rights reserved. Evaluation of the Antioxidant Activity and Antiproliferative Effect of the Jaboticaba (Myrciaria cauliflora) Seed Extracts in Oral Carcinoma Cells Mon, 18 Aug 2014 00:00:00 +0000 It is becoming increasingly evident that certain phytochemicals possess cancer chemopreventive properties. In this study, the antiproliferative activity of extracts from different parts of the jaboticaba (Myrciaria cauliflora) plant was evaluated for its effect on human oral carcinoma cell lines. The cytotoxicities of various plant extract concentrations were examined and the 50% maximal inhibitory concentration (IC50) was determined. Water extracts of jaboticaba seeds showed concentration-dependent antiproliferative effects. Annexin V/propidium iodide positivity with active caspase-3 induction indicated that the treated cells underwent apoptosis. Several important regulatory proteins (Bcl-2, Bcl-xL, Bid, and survivin) involved in apoptosis were also evaluated. The antioxidant activity of jaboticaba was investigated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assays, and the drug concentration eliciting 50% maximum stimulation (SC50) was determined. The present findings suggest that water extracts of jaboticaba seeds exhibit an antiproliferative effect against oral cancer cells by inducing apoptosis through downregulating survivin expression and thereby activating caspase-mediated Bid cleavage. Wen-Hung Wang, Yu-Chang Tyan, Zong-Shiow Chen, Ching-Gong Lin, Ming-Hui Yang, Shyng-Shiou Yuan, and Wan-Chi Tsai Copyright © 2014 Wen-Hung Wang et al. All rights reserved. Plasma Total Homocysteine Levels in Diabetic Retinopathy Thu, 14 Aug 2014 11:37:17 +0000 Huseyin Kayadibi, Erdim Sertoglu, and Metin Uyanik Copyright © 2014 Huseyin Kayadibi et al. All rights reserved. Proteomic Analysis of Gossypol Induces Necrosis in Multiple Myeloma Cells Thu, 14 Aug 2014 11:22:30 +0000 Gossypol is a phenolic aldehyde extracted from plants and is known to be an antitumor agent to induce cancer cell apoptosis. In the present study, multiple myeloma cells were treated with gossypol, which resulted in an increase of cellular reactive oxygen species (ROS) and cell necrosis. Quantitative proteomic analysis was carried out to identify differentially expressed proteins between untreated and gossypol-treated cells. Proteomic analysis identified 4330 proteins, in which 202 proteins are upregulated and 383 proteins are downregulated in gossypol-treated cells as compared to the untreated cells. Importantly, proteomic and western blot analysis showed that apoptosis regulators BAK and Bax were upregulated in gossypol-treated cells, indicating that Bcl-2 associated death pathway was activated. Similarly, gossypol also induced upregulations of DNA mismatch repair proteins and DNA replication licensing factor, suggesting that gossypol caused significant DNA damage. Furthermore, upregulations of HLA class I and class II histocompatibility antigens and beta-2-microglobulin were observed in gossypol-treated cells, indicating that gossypol has a novel function to activate cellular immune responses. Our data demonstrate that the execution of necrosis is a complex process involving ROS, DNA damage, and Bcl-2 family proteins. Gossypol-activated immune responses are a potential new approach for multiple myeloma chemotherapy. Renhua Xu, Enbing Tian, Haiping Tang, Chongdong Liu, and Qingtao Wang Copyright © 2014 Renhua Xu et al. All rights reserved. Variability in Myosteatosis and Insulin Resistance Induced by High-Fat Diet in Mouse Skeletal Muscles Thu, 14 Aug 2014 00:00:00 +0000 Nutrient overload leads to impaired muscle oxidative capacity and insulin sensitivity. However, comparative analyses of the effects of dietary manipulation on skeletal muscles with different fiber composition are lacking. This study aimed to investigate the selective adaptations in the soleus and tibialis anterior muscles evoked by administration of high-fat diet for 12 weeks in 10 mice (HFD mice) compared to 10 animals fed with a normal chow diet (control mice). Mice fed with the HFD diet exhibited hyperlipidemia, hyperinsulinemia, hyperglycemia, and lower exercise capacity in comparison to control mice. In control mice, soleus fibers showed higher lipid content than tibialis anterior fibers. In contrast, the lipid content was similar between the two muscles in HFD mice. Significant differences in markers of muscle mitochondrial production and/or activity as well as of lipid synthesis were detected between HFD mice and control mice, especially in the tibialis anterior. Moreover, translocation of GLUT-4 transporter to the plasma membrane and activation of the insulin signaling pathway were markedly inhibited in the tibialis and slightly reduced in the soleus of HFD mice compared to control mice. Overall, these results show that adaptive responses to dietary manipulation occur in a muscle-specific pattern. Massimo Collino, Raffaella Mastrocola, Debora Nigro, Fausto Chiazza, Manuela Aragno, Giuseppe D’Antona, and Marco A. Minetto Copyright © 2014 Massimo Collino et al. All rights reserved. Phosphoproteomic Analysis of Gossypol-Induced Apoptosis in Ovarian Cancer Cell Line, HOC1a Tue, 12 Aug 2014 00:00:00 +0000 Ovarian cancer is a major cause for death of gynecological cancer patients. The efficacy of traditional surgery and chemotherapy is rather compromised and platinum-resistant cancer recurs. Finding new therapeutic targets is urgently needed to increase the survival rate and to improve life quality of patients with ovarian cancer. In the present work, phosphoproteomic analysis was carried out on untreated and gossypol-treated ovarian cancer cell line, HOC1a. We identified approximately 9750 phosphopeptides from 3030 phosphoproteins, which are involved in diverse cellular processes including cytoskeletal organization, RNA and nucleotide binding, and cell cycle regulation. Upon gossypol treatment, changes in phosphorylation of twenty-nine proteins including YAP1 and AKAP12 were characterized. Western blotting and qPCR analysis were used to determine expression levels of proteins in YAP1-related Hippo pathway showing that gossypol induced upregulation of LATS1, which phosphorylates YAP1 at Ser 61. Furthermore, our data showed that gossypol targets the actin cytoskeletal organization through mediating phosphorylation states of actin-binding proteins. Taken together, our data provide valuable information to understand effects of gossypol on protein phosphorylation and apoptosis of ovarian cancer cells. Lixu Jin, Yuling Chen, Xinlin Mu, Qingquan Lian, Haiyun Deng, and Renshan Ge Copyright © 2014 Lixu Jin et al. All rights reserved. Laboratory Medicine 2014 Mon, 11 Aug 2014 08:22:10 +0000 Mina Hur, Patrizia Cardelli, and Giulio Mengozzi Copyright © 2014 Mina Hur et al. All rights reserved. Natural Product Polyamines That Inhibit Human Carbonic Anhydrases Tue, 05 Aug 2014 09:48:15 +0000 Natural product compound collections have proven an effective way to access chemical diversity and recent findings have identified phenolic, coumarin, and polyamine natural products as atypical chemotypes that inhibit carbonic anhydrases (CAs). CA enzymes are implicated as targets of variable drug therapeutic classes and the discovery of selective, drug-like CA inhibitors is essential. Just two natural product polyamines, spermine and spermidine, have until now been investigated as CA inhibitors. In this study, five more complex natural product polyamines 1–5, derived from either marine sponge or fungi, were considered for inhibition of six different human CA isozymes of interest in therapeutic drug development. All compounds share a simple polyamine core fragment, either spermine or spermidine, yet display substantially different structure activity relationships for CA inhibition. Notably, polyamines 1–5 were submicromolar inhibitors of the cancer drug target CA IX, this is more potent than either spermine or spermidine. Rohan A. Davis, Daniela Vullo, Claudiu T. Supuran, and Sally-Ann Poulsen Copyright © 2014 Rohan A. Davis et al. All rights reserved. Decoding Amyotrophic Lateral Sclerosis: Discovery of Novel Disease-Related Biomarkers and Future Perspectives in Neurodegeneration Mon, 04 Aug 2014 09:37:49 +0000 Ana Cristina Calvo, Pierre-François Pradat, Deise M. F. Mendonça, and Raquel Manzano Copyright © 2014 Ana Cristina Calvo et al. All rights reserved. Synthesis and In Vitro Inhibition Effect of New Pyrido[2,3-d]pyrimidine Derivatives on Erythrocyte Carbonic Anhydrase I and II Mon, 04 Aug 2014 08:18:45 +0000 In vitro inhibition effects of indolylchalcones and new pyrido[2,3-d]pyrimidine derivatives on purified human carbonic anhydrase I and II (hCA I and II) were investigated by using CO2 as a substrate. The results showed that all compounds inhibited the hCA I and hCA II enzyme activities. Among all the synthesized compounds, 7e ( µM) was found to be the most active compound for hCA I inhibitory activity and 5g ( µM) showed the highest hCA II inhibitory activity. Structure-activity relationships study showed that indolylchalcone derivatives have higher inhibitory activities than pyrido[2,3-d]pyrimidine derivatives on hCA I and hCA II. Additionally, methyl group bonded to uracil ring increases inhibitory activities on both hCA I and hCA II. Hilal Kuday, Fatih Sonmez, Cigdem Bilen, Emre Yavuz, Nahit Gençer, and Mustafa Kucukislamoglu Copyright © 2014 Hilal Kuday et al. All rights reserved. Amyotrophic Lateral Sclerosis: A Focus on Disease Progression Sun, 03 Aug 2014 09:31:32 +0000 Since amyotrophic lateral sclerosis (ALS) was discovered and described in 1869 as a neurodegenerative disease in which motor neuron death is induced, a wide range of biomarkers have been selected to identify therapeutic targets. ALS shares altered molecular pathways with other neurodegenerative diseases, such as Alzheimer’s, Huntington’s, and Parkinson’s diseases. However, the molecular targets that directly influence its aggressive nature remain unknown. What is the first link in the neurodegenerative chain of ALS that makes this disease so peculiar? In this review, we will discuss the progression of the disease from the viewpoint of the potential biomarkers described to date in human and animal model samples. Finally, we will consider potential therapeutic strategies for ALS treatment and future, innovative perspectives. Ana C. Calvo, Raquel Manzano, Deise M. F. Mendonça, María J. Muñoz, Pilar Zaragoza, and Rosario Osta Copyright © 2014 Ana C. Calvo et al. All rights reserved. Peptide-Ligand Binding Modeling of siRNA with Cell-Penetrating Peptides Thu, 24 Jul 2014 11:26:49 +0000 The binding affinity of a series of cell-penetrating peptides (CPP) was modeled through docking and making use of the number of intermolecular hydrogen bonds, lipophilic contacts, and the number of sp3 molecular orbital hybridization carbons. The new ranking of the peptides is consistent with the experimentally determined efficiency in the downregulation of luciferase activity, which includes the peptides’ ability to bind and deliver the siRNA into the cell. The predicted structures of the complexes of peptides to siRNA were stable throughout 10 ns long, explicit water molecular dynamics simulations. The stability and binding affinity of peptide-siRNA complexes was related to the sidechains and modifications of the CPPs, with the stearyl and quinoline groups improving affinity and stability. The reranking of the peptides docked to siRNA, together with explicit water molecular dynamics simulations, appears to be well suited to describe and predict the interaction of CPPs with siRNA. Alfonso T. García-Sosa, Indrek Tulp, Kent Langel, and Ülo Langel Copyright © 2014 Alfonso T. García-Sosa et al. All rights reserved. Aaptamines from the Marine Sponge Aaptos sp. Display Anticancer Activities in Human Cancer Cell Lines and Modulate AP-1-, NF-κB-, and p53-Dependent Transcriptional Activity in Mouse JB6 Cl41 Cells Wed, 23 Jul 2014 00:00:00 +0000 Aaptamine (8,9-dimethoxy-1H-benzo[de][1,6]naphthyridine) is a marine natural compound possessing antioxidative, antimicrobial, antifungal, and antiretroviral activity. Earlier, we have found that aaptamine and its derivatives demonstrate equal anticancer effects against the human germ cell cancer cell lines NT2 and NT2-R and cause some changes in the proteome of these cells. In order to explore further the mechanism of action of aaptamine and its derivatives, we studied the effects of aaptamine (1), demethyl(oxy)aaptamine (2), and isoaaptamine (3) on human cancer cell lines and on AP-1-, NF-κB-, and p53-dependent transcriptional activity in murine JB6 Cl41 cells. We showed that compounds 1–3 demonstrate anticancer activity in THP-1, HeLa, SNU-C4, SK-MEL-28, and MDA-MB-231 human cancer cell lines. Additionally, all compounds were found to prevent EGF-induced neoplastic transformation of murine JB6 Cl41 cells. Nuclear factors AP-1, NF-κB, and p53 are involved in the cellular response to high and nontoxic concentrations of aaptamine alkaloids 1–3. Furthermore, inhibition of EGF-induced JB6 cell transformation, which is exerted by the compounds 1–3 at low nontoxic concentrations of 0.7–2.1 μM, cannot be explained by activation of AP-1 and NF-κB. Sergey A. Dyshlovoy, Sergey N. Fedorov, Larisa K. Shubina, Alexandra S. Kuzmich, Carsten Bokemeyer, Gunhild Keller-von Amsberg, and Friedemann Honecker Copyright © 2014 Sergey A. Dyshlovoy et al. All rights reserved. Changes in Biochemical Characteristics and Activities of Ripening Associated Enzymes in Mango Fruit during the Storage at Different Temperatures Tue, 22 Jul 2014 00:00:00 +0000 As a part of the study to explore the possible strategy for enhancing the shelf life of mango fruits, we investigated the changes in biochemical parameters and activities of ripening associated enzymes of Ashwina hybrid mangoes at 4-day regular intervals during storage at −10°C, 4°C, and °C. Titratable acidity, vitamin C, starch content, and reducing sugar were higher at unripe state and gradually decreased with the increasing of storage time at all storage temperatures while phenol content, total soluble solid, total sugar, and nonreducing sugar contents gradually increased. The activities of amylase, -mannosidase, -glucosidase, and invertase increased sharply within first few days and decreased significantly in the later stage of ripening at °C. Meanwhile polyphenol oxidase, -galactosidase, and -hexosaminidase predominantly increased significantly with the increasing days of storage till later stage of ripening. At −10°C and 4°C, the enzymes as well as carbohydrate contents of storage mango changed slightly up to 4 days and thereafter the enzyme became fully dormant. The results indicated that increase in storage temperature and time correlated with changes in biochemical parameters and activities of glycosidases suggested the suppression of -galactosidase and -hexosaminidase might enhance the shelf life of mango fruits. Md. Anowar Hossain, Md. Masud Rana, Yoshinobu Kimura, and Hairul Azman Roslan Copyright © 2014 Md. Anowar Hossain et al. All rights reserved. Protective Effect of Ethanolic Extract of Tabernaemontana divaricata (L.) R. Br. against DEN and Fe NTA Induced Liver Necrosis in Wistar Albino Rats Thu, 17 Jul 2014 00:00:00 +0000 This study is an attempt to evaluate the hepatoprotective activity of Tabernaemontana divaricata against DEN and Fe NTA induced liver necrosis in rats. Ethanolic extract of the whole plant of Tabernaemontana divaricata at doses of 200 and 400 mg/kg body weight and 5-fluorouracil (standard drug) was orally administered to male Wistar Albino rats once daily for 24 weeks, simultaneously treated with the carcinogen DEN and Fe NTA. In simultaneously treated animals, the plant extract significantly decreased the levels of uric acid, bilirubin, AST, ALT, and ALP in serum and increased the levels of liver marker enzymes in liver. Treatment with the extracts resulted in a significant increase in the levels of antioxidants accompanied by a marked reduction in the levels of malondialdehyde when compared to DEN and Fe NTA treated group. When compared with 200 mg/kg bw rats, 400 mg/kg bw rats and 5-fluorouracil treated rats showed better results in all the parameters. The histopathological studies confirmed the protective effects of extract against DEN and Fe NTA induced liver necrosis. Thus, it could be concluded that the use of Tabernaemontana divaricata extract in the treatment of carcinogen induced hepatic necrosis. Kannappan Poornima, Palanisamy Chella Perumal, and Velliyur Kanniappan Gopalakrishnan Copyright © 2014 Kannappan Poornima et al. All rights reserved. Effect of Cocoa Butter and Sunflower Oil Supplementation on Performance, Immunoglobulin, and Antioxidant Vitamin Status of Rats Wed, 16 Jul 2014 15:51:28 +0000 This study investigated the effects of cocoa butter and sunflower oil alone and in combination on performance, some biochemical parameters, immunoglobulin, and antioxidant vitamin status in Wistar rats. Forty-eight male rats were assigned to four groups, consisting of 12 rats with 3 replicates. Control received balanced rat diet without oil, cocoa butter group received 3.5% cocoa butter, sunflower oil group received 3.5% sunflower oil, the last group received 1.75% sunflower oil + 1.75% cocoa butter supplementation in the rat diet for 8 weeks. The total feed consumption in sunflower oil group was statistically lower than in the other groups. The serum creatinine level was decreased in cocoa butter group compared to control. Triglyceride and VLDL cholesterol levels were decreased in only sunflower oil and only cocoa butter groups as compared to control. The level of Ig M was statistically lower in cocoa butter and cocoa butter + sunflower oil groups than in control and sunflower oil groups. There were no statistically important difference in vitamin concentrations among trial groups. It was concluded that the supplementation of cocoa butter in diet decreased Ig M level, while the supplementation of cocoa butter and sunflower oil alone decreased the triglyceride and VLDL cholesterol levels. Ebru Yıldırım, Miyase Çınar, İlkay Yalçınkaya, Hüsamettin Ekici, Nurgül Atmaca, and Enes Güncüm Copyright © 2014 Ebru Yıldırım et al. All rights reserved. Physiological and Biochemical Changes in Brassica juncea Plants under Cd-Induced Stress Tue, 15 Jul 2014 12:56:40 +0000 Plants of Brassica juncea L. var. RLC-1 were exposed for 30 days to different concentrations (0, 0.2, 0.4, and 0.6 mM) of cadmium (Cd) to analyze the Cd uptake, H2O2 content, hormonal profiling, level of photosynthetic pigments (chlorophyll, carotenoid, and flavonoid), gaseous exchange parameters (photosynthetic rate, vapour pressure deficit, intercellular CO2 concentration, and intrinsic mesophyll rate), antioxidative enzymes (superoxide dismutase, polyphenol oxidase, glutathione-S transferase, and glutathione peroxidase), antioxidant assays (DPPH, ABTS, and total phenolic content), and polyphenols. Results of the present study revealed the increased H2O2 content and Cd uptake with increasing metal doses. UPLC analysis of plants showed the presence of various polyphenols. Gaseous exchange measurements were done by infrared gas analyzer (IRGA), which was negatively affected by metal treatment. In addition, LC/MS study showed the variation in the expression of plant hormones. Level of photosynthetic pigments and activities of antioxidative enzymes were altered significantly in response to metal treatment. In conclusion, the antioxidative defence system of plants got activated due to heavy metal stress, which protects the plants by scavenging free radicals. Dhriti Kapoor, Satwinderjeet Kaur, and Renu Bhardwaj Copyright © 2014 Dhriti Kapoor et al. All rights reserved. Depletion of Arginine by Recombinant Arginine Deiminase Induces nNOS-Activated Neurotoxicity in Neuroblastoma Cells Mon, 14 Jul 2014 09:25:45 +0000 The abnormal regulation of inducible nitric oxide synthase (iNOS) and neuronal nitric oxide synthase (nNOS) is associated with neurodegenerative disorders. Recombinant arginine deiminase (rADI) is a selective NO modulator of iNOS and eNOS in endothelial cells, and it also exhibits neuroprotective activity in an iNOS-induced neuron-microglia coculture system. However, the effect of rADI on nNOS remains unknown. Addressing this issue is important for evaluating the potential application of rADI in neurodegenerative diseases. SH-SY5Y cells were treated with N-methyl-D-aspartic acid (NMDA) to activate nNOS. NMDA increased NO production by 39.7 ± 3.9% via nNOS under arginine-containing conditions, but there was no significant increase in both arginine-free and rADI pretreated arginine-containing (citrulline) buffer. Subsequently, neither NMDA nor rADI alone caused cytotoxicity, whereas cotreatment with NMDA and rADI resulted in dissipation of the cell mitochondrial membrane potential and decreased cell viability. The mechanism of rADI cytotoxicity in the presence of NMDA is caused by the inhibition of NO production via nNOS mediated by the NMDA receptor, which was abolished when extracellular arginine was absent, even in the presence of citrulline. rADI not only reduced NO production but also caused cellular toxicity in nNOS-activated SH-SY5Y cells, suggesting a dual role for rADI in NOS-mediated neurotoxicity. Shan-Erh Lin, Fe-Lin Lin Wu, Ming-Feng Wei, and Li-Jiuan Shen Copyright © 2014 Shan-Erh Lin et al. All rights reserved. Profiling of Biomarkers for the Exposure of Polycyclic Aromatic Hydrocarbons: Lamin-A/C Isoform 3, Poly[ADP-ribose] Polymerase 1, and Mitochondria Copy Number Are Identified as Universal Biomarkers Thu, 10 Jul 2014 00:00:00 +0000 This study investigated the profiling of polycyclic aromatic hydrocarbon- (PAH-) induced genotoxicity in cell lines and zebrafish. Each type of cells displayed different proportionality of apoptosis. Mitochondrial DNA (mtDNA) copy number was dramatically elevated after 5-day treatment of fluoranthene and pyrene. The notable deregulated proteins for PAHs exposure were displayed as follows: lamin-A/C isoform 3 and annexin A1 for benzopyrene; lamin-A/C isoform 3 and DNA topoisomerase 2-alpha for pentacene; poly[ADP-ribose] polymerase 1 (PARP-1) for fluoranthene; and talin-1 and DNA topoisomerase 2-alpha for pyrene. Among them, lamin-A/C isoform 3 and PARP-1 were further confirmed using mRNA and protein expression study. Obvious morphological abnormalities including curved backbone and cardiomegaly in zebrafish were observed in the 54 hpf with more than 400 nM of benzopyrene. In conclusion, the change of mitochondrial genome (increased mtDNA copy number) was closely associated with PAH exposure in cell lines and mesenchymal stem cells. Lamin-A/C isoform 3, talin-1, and annexin A1 were identified as universal biomarkers for PAHs exposure. Zebrafish, specifically at embryo stage, showed suitable in vivo model for monitoring PAHs exposure to hematopoietic tissue and other organs. Hwan-Young Kim, Hye-Ran Kim, Min-Gu Kang, Nguyen Thi Dai Trang, Hee-Jo Baek, Jae-Dong Moon, Jong-Hee Shin, Soon-Pal Suh, Dong-Wook Ryang, Hoon Kook, and Myung-Geun Shin Copyright © 2014 Hwan-Young Kim et al. All rights reserved. Ketogenic Diet in Neuromuscular and Neurodegenerative Diseases Thu, 03 Jul 2014 00:00:00 +0000 An increasing number of data demonstrate the utility of ketogenic diets in a variety of metabolic diseases as obesity, metabolic syndrome, and diabetes. In regard to neurological disorders, ketogenic diet is recognized as an effective treatment for pharmacoresistant epilepsy but emerging data suggests that ketogenic diet could be also useful in amyotrophic lateral sclerosis, Alzheimer, Parkinson’s disease, and some mitochondriopathies. Although these diseases have different pathogenesis and features, there are some common mechanisms that could explain the effects of ketogenic diets. These mechanisms are to provide an efficient source of energy for the treatment of certain types of neurodegenerative diseases characterized by focal brain hypometabolism; to decrease the oxidative damage associated with various kinds of metabolic stress; to increase the mitochondrial biogenesis pathways; and to take advantage of the capacity of ketones to bypass the defect in complex I activity implicated in some neurological diseases. These mechanisms will be discussed in this review. Antonio Paoli, Antonino Bianco, Ernesto Damiani, and Gerardo Bosco Copyright © 2014 Antonio Paoli et al. All rights reserved. Optimization of Alkaline Extraction and Bioactivities of Polysaccharides from Rhizome of Polygonatum odoratum Tue, 01 Jul 2014 12:03:01 +0000 The present study is to explore the optimal extraction parameters, antioxidant activity, and antimicrobial activity of alkaline soluble polysaccharides from rhizome of Polygonatum odoratum. The optimal extraction parameters were determined as the following: NaOH concentration (A) 0.3 M, temperature (B) , ratio of NaOH to solid (C) 10-fold, and extraction time (D) 4 h, in which ratio of NaOH to solid was a key factor. The order of the factors was ratio of NaOH to solid (fold, C) > extraction temperature (, B) > NaOH concentration (M, A) > extraction time (h, D). The monosaccharide compositions of polysaccharides from P. odoratum were rhamnose, mannose, xylose, and arabinose with the molecular ratio of 31.78, 31.89, 11.11, and 1.00, respectively. The reducing power, the 1, 1-diphenyl-2-picryl-hydrazil (DPPH) radical scavenging rate, the hydroxyl radicals scavenging rate, and the inhibition rate to polyunsaturated fatty acid (PUFA) peroxidation of the alkaline soluble polysaccharides from P. odoratum at 1 mg/mL were 9.81%, 52.84%, 19.22%, and 19.42% of ascorbic acid at the same concentration, respectively. They also showed antimicrobial activity against pathogenic bacteria Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus subtilis, and Escherichia coli. Yong Chen, Luoyi Yin, Xuejiao Zhang, Yan Wang, Qiuzhi Chen, Chenzhong Jin, Yihong Hu, and Jihua Wang Copyright © 2014 Yong Chen et al. All rights reserved. Biocatalytic Synthesis of Flavor Ester “Pentyl Valerate” Using Candida rugosa Lipase Immobilized in Microemulsion Based Organogels: Effect of Parameters and Reusability Tue, 01 Jul 2014 11:13:23 +0000 Pentyl valerate was synthesized biocatalytically using Candida rugosa lipase (CRL) immobilized in microemulsion based organogels (MBGs). The optimum conditions were found to be pH 7.0, temperature of 37°C, ratio of concentration of water to surfactant (Wo) of 60, and the surfactant sodium bis-2-(ethylhexyl)sulfosuccinate (AOT) for MBG preparation. Although kinetic studies revealed that the enzyme in free form had high affinity towards substrates ( = 23.2 mM for pentanol and 76.92 mM for valeric acid) whereas, after immobilization, the values increased considerably (74.07 mM for pentanol and 83.3 mM for valeric acid) resulting in a slower reaction rate, the maximum conversion was much higher in case of immobilized enzyme (~99%) as compared to free enzyme (~19%). Simultaneous effects of important parameters were studied using response surface methodology (RSM) conjugated with Box-Behnken design (BBD) with five variables (process parameters), namely, enzyme concentration, initial water content (Wo), solvent used for MBG preparation, substrate ratio and time, and response as the final product formation, that is, pentyl valerate (%). The MBGs were reused for 10 consecutive cycles for ester synthesis. Efficacy of AOT/isooctane as dehydrating agent for extracting excess water from MBGs was found to exert a positive effect on the esterification reaction. Tripti Raghavendra, Nilam Panchal, Jyoti Divecha, Amita Shah, and Datta Madamwar Copyright © 2014 Tripti Raghavendra et al. All rights reserved. Expression of IMPDH mRNA after Mycophenolate Administration in Male Volunteers Tue, 01 Jul 2014 00:00:00 +0000 Background. Mycophenolic acid (MPA) is the first-line antimetabolic immunosuppressants used in solid organ transplantation. Here, in vivo expressions of the pharmacodynamic marker IMPDH mRNA were analyzed to investigate its usefulness in assessing drug effects. Materials and Methods. Six healthy male volunteers who had the same genotype for genes known to be associated with drug metabolism and effects were selected to remove the confounding effect of these genotypes. Mycophenolate mofetil (MMF, 1 g) was administered once to each subject, and blood samples were collected with certain interval before and after MMF administration to measure lymphocyte expression levels of IMPDH1 and IMPDH2 mRNA. One week later, the experiment was repeated. Results. Whereas IMPDH1 mRNA expression was stable, IMPDH2 mRNA expression showed 2 peaks in the first week. Both IMPDH1 and IMPDH2 mRNA expression in the second week remarkably decreased from the first week. Conclusion. The temporary increase in IMPDH2 mRNA expression in the first week might be due to a reactive reaction against the plasma MPA concentration. In the second week, the intracellular guanosine monophosphate might be depleted, rendering IMPDH2 mRNA synthesis inactive. When MPA is regularly administered to reach a steady state, the IMPDH2 mRNA expression may be kept low and may effectively reflect biological responses regardless of drug intake. Sollip Kim, Woochang Lee, Sail Chun, Tae Hyun Um, and Won-Ki Min Copyright © 2014 Sollip Kim et al. All rights reserved. Vitamin D Signaling in Myogenesis: Potential for Treatment of Sarcopenia Mon, 30 Jun 2014 10:58:47 +0000 Muscle mass and strength progressively decrease with age, which results in a condition known as sarcopenia. Sarcopenia would lead to physical disability, poor quality of life, and death. Therefore, much is expected of an effective intervention for sarcopenia. Epidemiologic, clinical, and laboratory evidence suggest an effect of vitamin D on muscle function. However, the precise molecular and cellular mechanisms remain to be elucidated. Recent studies suggest that vitamin D receptor (VDR) might be expressed in muscle fibers and vitamin D signaling via VDR plays a role in the regulation of myoblast proliferation and differentiation. Understanding how vitamin D signaling contributes to myogenesis will provide a valuable insight into an effective nutritional strategy to moderate sarcopenia. Here we will summarize the current knowledge about the effect of vitamin D on skeletal muscle and myogenic cells and discuss the potential for treatment of sarcopenia. Akira Wagatsuma and Kunihiro Sakuma Copyright © 2014 Akira Wagatsuma and Kunihiro Sakuma. All rights reserved. Usefulness of Combining Galectin-3 and BIVA Assessments in Predicting Short- and Long-Term Events in Patients Admitted for Acute Heart Failure Mon, 30 Jun 2014 10:14:13 +0000 Introduction. Acute heart failure (AHF) is associated with a higher risk for the occurrence of rehospitalization and death. Galectin-3 (GAL3) is elevated in AHF patients and is an indicator in predicting short-term mortality. The total body water using bioimpedance vector analysis (BIVA) is able to identify mortality within AHF patients. The aim of this study was to evaluate the short- and long-term predictive value of GAL3, BIVA, and the combination of both in AHF patients in Emergency Department (ED). Methods. 205 ED patients with AHF were evaluated by testing for B type natriuretic peptide (BNP) and GAL3. The primary endpoint was death and rehospitalization at 30, 60, 90, and 180 days and 12 and 18 months. AHF patients were evaluated at the moment of ED arrival with clinical judgment and GAL3 and BIVA measurement. Results. GAL3 level was significantly higher in patients >71 years old, and with  cc/min. The area under the curve (AUC) of , GAL3 and BIVA for death and rehospitalization both when considered in total and when considered serially for the follow-up period showed that the combination has a better prognostic value. Kaplan-Meier survival curve for GAL3 values >17.8 ng/mL shows significant survival difference. At multivariate Cox regression analysis GAL3 is an independent variable to predict death + rehospitalization with a value of 32.24 ng/mL at 30 days (). Conclusion. In patients admitted for AHF an early assessment of GAL3 and BIVA seems to be useful in identifying patients at high risk for death and rehospitalization at short and long term. Combining the biomarker and the device could be of great utility since they monitor the severity of two pathophysiological different mechanisms: heart fibrosis and fluid overload. Benedetta De Berardinis, Laura Magrini, Giorgio Zampini, Benedetta Zancla, Gerardo Salerno, Patrizia Cardelli, Enrico Di Stasio, Hanna K. Gaggin, Arianna Belcher, Blair A. Parry, John T. Nagurney, James L. Januzzi Jr., and Salvatore Di Somma Copyright © 2014 Benedetta De Berardinis et al. All rights reserved. Synthesis and Evaluation of Novel Pyrroles and Pyrrolopyrimidines as Anti-Hyperglycemic Agents Thu, 26 Jun 2014 10:20:31 +0000 A series of pyrrole and pyrrolopyrimidine derivatives were examined for their in vivo antihyperglycemic activity. Compounds Ia–c,e, and IVg showed promising antihyperglycemic activity equivalent to a well-known standard antihyperglycemic drug, Glimepiride (Amaryl, 4 mg/kg). In this paper, we examine and discuss the structure-activity relationships and antihyperglycemic activity of these compounds. M. S. Mohamed, S. A. Ali, D. H. A. Abdelaziz, and Samar S. Fathallah Copyright © 2014 M. S. Mohamed et al. All rights reserved. The Landscape of Protein Biomarkers Proposed for Periodontal Disease: Markers with Functional Meaning Thu, 26 Jun 2014 08:06:01 +0000 Periodontal disease (PD) is characterized by a deregulated inflammatory response which fails to resolve, activating bone resorption. The identification of the proteomes associated with PD has fuelled biomarker proposals; nevertheless, many questions remain. Biomarker selection should favour molecules representing an event which occurs throughout the disease progress. The analysis of proteome results and the information available for each protein, including its functional role, was accomplished using the OralOme database. The integrated analysis of this information ascertains if the suggested proteins reflect the cell and/or molecular mechanisms underlying the different forms of periodontal disease. The evaluation of the proteins present/absent or with very different concentrations in the proteome of each disease state was used for the identification of the mechanisms shared by different PD variants or specific to such state. The information presented is relevant for the adequate design of biomarker panels for PD. Furthermore, it will open new perspectives and help envisage future studies targeted to unveil the functional role of specific proteins and help clarify the deregulation process in the PD inflammatory response. Nuno Rosa, Maria José Correia, Joel P. Arrais, Nuno Costa, José Luís Oliveira, and Marlene Barros Copyright © 2014 Nuno Rosa et al. All rights reserved. Redox Signaling in Degenerative Diseases: From Molecular Mechanisms to Health Implications Wed, 25 Jun 2014 09:17:23 +0000 Cristina Angeloni, Tullia Maraldi, and David Vauzour Copyright © 2014 Cristina Angeloni et al. All rights reserved. Comparison of Commercial Genetic-Testing Services in Korea with 23andMe Service Wed, 25 Jun 2014 05:47:25 +0000 Introduction. Genetic testing services for disease prediction, drug responses, and traits are commercially available by several companies in Korea. However, there has been no evaluation study for the accuracy and usefulness of these services. We aimed to compare two genetic testing services popular in Korea with 23andMe service in the United States. Materials and Methods. We compared the results of two persons (one man and one woman) serviced by Hellogene Platinum (Theragen Bio Institute), DNAGPS Optimus (DNAlink), and 23andMe service. Results. Among 3 services, there were differences in the estimation of relative risks for the same disease. For lung cancer, the range of relative risk was from 0.9 to 2.09. These differences were thought to be due to the differences of applied single nucleotide polymorphisms (SNPs) in each service for the calculation of risk. Also, the algorithm and population database would have influence on the estimation of relative disease risks. The concordance rate of SNP calls between DNAGPS Optimus and 23andMe services was 100% (30/30). Conclusions. Our study showed differences in disease risk estimations among three services, although they gave good concordance rate for SNP calls. We realized that the genetic services need further evaluation and standardization, especially in disease risk estimation algorithm. Sollip Kim, Ki-Won Eom, Chong-Rae Cho, and Tae Hyun Um Copyright © 2014 Sollip Kim et al. All rights reserved. Anticancer and Antioxidant Activity of Bread Enriched with Broccoli Sprouts Tue, 24 Jun 2014 07:11:40 +0000 This study is focused on antioxidant and anticancer capacity of bread enriched with broccoli sprouts (BS) in the light of their potential bioaccessibility and bioavailability. Generally, bread supplementation elevated antioxidant potential of product (both nonenzymatic and enzymatic antioxidant capacities); however, the increase was not correlated with the percent of BS. A replacement up to 2% of BS gives satisfactory overall consumers acceptability and desirable elevation of antioxidant potential. High activity was especially found for extracts obtained after simulated digestion, which allows assuming their protective effect for upper gastrointestinal tract; thus, the anticancer activity against human stomach cancer cells (AGS) was evaluated. A prominent cytostatic response paralleled by the inhibition of AGS motility in the presence of potentially mastication-extractable phytochemicals indicates that phenolic compounds of BS retain their biological activity in bread. Importantly, the efficient phenolics concentration was about 12 μM for buffer extract, 13 μM for extracts after digestion in vitro, and 7 μM for extract after absorption in vitro. Our data confirm chemopreventive potential of bread enriched with BS and indicate that BS comprise valuable food supplement for stomach cancer chemoprevention. Urszula Gawlik-Dziki, Michał Świeca, Dariusz Dziki, Łukasz Sęczyk, Urszula Złotek, Renata Różyło, Kinga Kaszuba, Damian Ryszawy, and Jarosław Czyż Copyright © 2014 Urszula Gawlik-Dziki et al. All rights reserved. Identification of Phenolic Acids and Flavonoids in Monofloral Honey from Bangladesh by High Performance Liquid Chromatography: Determination of Antioxidant Capacity Tue, 24 Jun 2014 00:00:00 +0000 The aim of the present study was to characterize the phenolic acids, flavonoids, and antioxidant properties of monofloral honey collected from five different districts in Bangladesh. A new high performance liquid chromatography (HPLC) equipped with a UV detector method was developed for the identification of the phenolic acids and flavonoids. A total of five different phenolic acids were identified, with the most abundant being caffeic acid, benzoic acid, gallic acid, followed by chlorogenic acid and trans-cinnamic acid. The flavonoids, kaempferol, and catechin were most abundant, followed by myricetin and naringenin. The mean moisture content, total sugar content, and color characteristics of the honey samples were 18.36 ± 0.95%, 67.40 ± 5.63 g/100 g, and 129.27 ± 34.66 mm Pfund, respectively. The mean total phenolic acids, total flavonoid content, and proline content were 199.20 ± 135.23, 46.73 ± 34.16, and 556.40 ± 376.86 mg/kg, respectively, while the mean FRAP values and DPPH radical scavenging activity were 327.30 ± 231.87 μM Fe (II)/100 g and 36.95 ± 20.53%, respectively. Among the different types of honey, kalijira exhibited the highest phenolics and antioxidant properties. Overall, our study confirms that all the investigated honey samples are good sources of phenolic acids and flavonoids with good antioxidant properties. Mohammed Moniruzzaman, Chua Yung An, Pasupuleti Visweswara Rao, Mohammad Nurul Islam Hawlader, Siti Amirah Binti Mohd Azlan, Siti Amrah Sulaiman, and Siew Hua Gan Copyright © 2014 Mohammed Moniruzzaman et al. All rights reserved. Oxygen Radicals Elicit Paralysis and Collapse of Spinal Cord Neuron Growth Cones upon Exposure to Proinflammatory Cytokines Mon, 23 Jun 2014 08:11:50 +0000 A persistent inflammatory and oxidative stress is a hallmark of most chronic CNS pathologies (Alzheimer’s (ALS)) as well as the aging CNS orchestrated by the proinflammatory cytokines tumor necrosis factor alpha (TNFα) and interleukin-1 beta (IL-1β). Loss of the integrity and plasticity of neuronal morphology and connectivity comprises an early step in neuronal degeneration and ultimate decline of cognitive function. We examined in vitro whether TNFα or IL-1β impaired morphology and motility of growth cones in spinal cord neuron cultures. TNFα and IL-1β paralyzed growth cone motility and induced growth cone collapse in a dose-dependent manner reflected by complete attenuation of neurite outgrowth. Scavenging reactive oxygen species (ROS) or inhibiting NADPH oxidase activity rescued loss of neuronal motility and morphology. TNFα and IL-1β provoked rapid, NOX-mediated generation of ROS in advancing growth cones, which preceded paralysis of motility and collapse of morphology. Increases in ROS intermediates were accompanied by an aberrant, nonproductive reorganization of actin filaments. These findings suggest that NADPH oxidase serves as a pivotal source of oxidative stress in neurons and together with disruption of actin filament reorganization contributes to the progressive degeneration of neuronal morphology in the diseased or aging CNS. Thomas B. Kuhn Copyright © 2014 Thomas B. Kuhn. All rights reserved. Hematological and Biochemical Markers of Iron Status in a Male, Young, Physically Active Population Sun, 22 Jun 2014 08:12:07 +0000 The aim of this study was to establish reference intervals (RIs) for the hemogram and iron status biomarkers in a physically active population. The study population included male volunteers () with an average age of 19 ± 1 years who had participated in a regular and controlled exercise program for four months. Blood samples were collected to determine hematological parameters using a Sysmex XE-5000 analyzer (Sysmex, Kobe, Japan). Iron, total iron-binding capacity (TIBC), transferrin saturation and ferritin, and high-sensitivity C-reactive protein (CRP) concentrations in serum samples were measured using commercial kits (Roche Diagnostics, GmbH, Mannheim, Germany) and a Roche/Hitachi 902 analyzer. The RIs were established using the RefVal program 4.1b. The leucocyte count, TIBC, and CRP and ferritin concentrations exhibited higher RIs compared with those in a nonphysically active population. Thirty volunteers (outliers) were removed from the reference population due to blood abnormalities. Among the outliers, 46% exhibited higher CRP concentrations and lower concentrations of iron and reticulocyte hemoglobin compared with the nonphysically active population (). Our results showed that it is important to establish RIs for certain laboratory parameters in a physically active population, especially for tests related to the inflammatory response and iron metabolism. Lázaro Alessandro Soares Nunes, Helena Zerlotti W. Grotto, René Brenzikofer, and Denise Vaz Macedo Copyright © 2014 Lázaro Alessandro Soares Nunes et al. All rights reserved. Current Approaches for Predicting a Lack of Response to Anti-EGFR Therapy in KRAS Wild-Type Patients Wed, 18 Jun 2014 11:27:27 +0000 Targeting epidermal growth factor receptor (EGFR) has been one of the most effective colorectal cancer strategies. Anti-EGFR antibodies function by binding to the extracellular domain of EGFR, preventing its activation, and ultimately providing clinical benefit. KRAS mutations in codons 12 and 13 are recognized prognostic and predictive biomarkers that should be analyzed at the clinic prior to the administration of anti-EGFR therapy. However, still an important fraction of KRAS wild-type patients do not respond to the treatment. The identification of additional genetic determinants of primary or secondary resistance to EGFR targeted therapy for further improving the selection of patients is urgent. Herein, we review the latest published literature highlighting the most important genes that may predict resistance to anti-EGFR monoclonal antibodies in colorectal cancer patients. According to the available findings, the evaluation of BRAF, NRAS, PIK3CA, and PTEN status could be the right strategy to select patients who are likely to respond to anti-EGFR therapies. In the future, the combination of those biomarkers will help establish consensus that can be introduced into clinical practice. Tze-Kiong Er, Chih-Chieh Chen, Luis Bujanda, and Marta Herreros-Villanueva Copyright © 2014 Tze-Kiong Er et al. All rights reserved. Purification and Characterization of Catalase from Marine Bacterium Acinetobacter sp. YS0810 Wed, 18 Jun 2014 09:11:27 +0000 The catalase from marine bacterium Acinetobacter sp. YS0810 (YS0810CAT) was purified and characterized. Consecutive steps were used to achieve the purified enzyme as follows: ethanol precipitation, DEAE Sepharose ion exchange, Superdex 200 gel filtration, and Resource Q ion exchange. The active enzyme consisted of four identical subunits of 57.256 kDa. It showed a Soret peak at 405 nm, indicating the presence of iron protoporphyrin IX. The catalase was not apparently reduced by sodium dithionite but was inhibited by 3-amino-1,2,4-triazole, hydroxylamine hydrochloride, and sodium azide. Peroxidase-like activity was not found with the substrate o-phenylenediamine. So the catalase was determined to be a monofunctional catalase. N-terminal amino acid of the catalase analysis gave the sequence SQDPKKCPVTHLTTE, which showed high degree of homology with those of known catalases from bacteria. The analysis of amino acid sequence of the purified catalase by matrix-assisted laser desorption ionization time-of-flight mass spectrometry showed that it was a new catalase, in spite of its high homology with those of known catalases from other bacteria. The catalase showed high alkali stability and thermostability. Xinhua Fu, Wei Wang, Jianhua Hao, Xianglin Zhu, and Mi Sun Copyright © 2014 Xinhua Fu et al. All rights reserved. Biopolymeric Formulations for Biocatalysis and Biomedical Applications Tue, 17 Jun 2014 11:15:55 +0000 Three gel disks formulations prepared using chitosan (Chito) or polyethylenimine (PEI) followed by glutaraldehyde were prepared for biocatalysis and biomedical applications. The carriers have been used to immobilize lactase covalently and it was evaluated in terms of enzyme loading capacity and enzyme kinetics (km and Vmax). The Km of the Chito formulation was almost half that of the PEI formulations, which is favored in industries. On the other hand, the gel disks were evaluated in terms of their swelling kinetics and the gels’ morphology using SEM. The mechanism of the three gels’ swelling was also studied and it was found to be non-Fickian, where the mechanism of transport depends on both the diffusion and polymer relaxation, which are controlling the overall rate of water uptake. The Chito formulation was 2–5 folds and PEI formulations were 33–62 folds in terms of the swelling rate constant and the diffusion rate, respectively. These results were highly supported by the SEM. This study will help scientists to design the right polymer network for enzymes immobilization as well as control the surface area and the swelling power of the polymers for different applications such as drug delivery systems and tissue engineering. Magdy M. M. Elnashar and Tarek Kahil Copyright © 2014 Magdy M. M. Elnashar and Tarek Kahil. All rights reserved. The Study of Interactions between Active Compounds of Coffee and Willow (Salix sp.) Bark Water Extract Wed, 11 Jun 2014 14:13:04 +0000 Coffee and willow are known as valuable sources of biologically active phytochemicals such as chlorogenic acid, caffeine, and salicin. The aim of the study was to determine the interactions between the active compounds contained in water extracts from coffee and bark of willow (Salix purpurea and Salix myrsinifolia). Raw materials and their mixtures were characterized by multidirectional antioxidant activities; however, bioactive constituents interacted with each other. Synergism was observed for ability of inhibition of lipid peroxidation and reducing power, whereas compounds able to scavenge ABTS radical cation acted antagonistically. Additionally, phytochemicals from willow bark possessed hydrophilic character and thermostability which justifies their potential use as an ingredient in coffee beverages. Proposed mixtures may be used in the prophylaxis or treatment of some civilization diseases linked with oxidative stress. Most importantly, strong synergism observed for phytochemicals able to prevent lipids against oxidation may suggest protective effect for cell membrane phospholipids. Obtained results indicate that extracts from bark tested Salix genotypes as an ingredient in coffee beverages can provide health promoting benefits to the consumers; however, this issue requires further study. Agata Durak and Urszula Gawlik-Dziki Copyright © 2014 Agata Durak and Urszula Gawlik-Dziki. All rights reserved. Novel Epoxy Activated Hydrogels for Solving Lactose Intolerance Wed, 11 Jun 2014 09:25:34 +0000 “Lactose intolerance” is a medical problem for almost 70% of the world population. Milk and dairy products contain 5–10% w/v lactose. Hydrolysis of lactose by immobilized lactase is an industrial solution. In this work, we succeeded to increase the lactase loading capacity to more than 3-fold to 36.3 U/g gel using epoxy activated hydrogels compared to 11 U/g gel using aldehyde activated carrageenan. The hydrogel’s mode of interaction was proven by FTIR, DSC, and TGA. The high activity of the epoxy group was regarded to its ability to attach to the enzyme’s –SH, –NH, and –OH groups, whereas the aldehyde group could only bind to the enzyme’s –NH2 group. The optimum conditions for immobilization such as epoxy chain length and enzyme concentration have been studied. Furthermore, the optimum enzyme conditions were also deliberated and showed better stability for the immobilized enzyme and the Michaelis constants, and , were doubled. Results revealed also that both free and immobilized enzymes reached their maximum rate of lactose conversion after 2 h, albeit, the aldehyde activated hydrogel could only reach 63% of the free enzyme. In brief, the epoxy activated hydrogels are more efficient in immobilizing more enzymes than the aldehyde activated hydrogel. Magdy M. M. Elnashar and Mohamed E. Hassan Copyright © 2014 Magdy M. M. Elnashar and Mohamed E. Hassan. All rights reserved. Essential Amino Acids and Exercise Tolerance in Elderly Muscle-Depleted Subjects with Chronic Diseases: A Rehabilitation without Rehabilitation? Mon, 09 Jun 2014 07:43:21 +0000 Exercise intolerance remains problematic in subjects with chronic heart failure (CHF) and/or chronic obstructive pulmonary disease (COPD). Recent studies show that supplemented essential amino acids (EAAs) may exert beneficial effects on CHF/COPD physical capacity. The results from 3 investigations (2 conducted on CHF and 1 on COPD subjects) served as the basis for this paper. The 3 studies consistently showed that elderly CHF and COPD improved exercise intolerance after 1–3 months of EAA supplementation (8 g/d). In CHF exercise capacity increased 18.7% to 23% (watts; bicycle test), and 12% to 22% (meters) in 6 min walking test. Moreover, patients reduced their resting plasma lactate levels (by 25%) and improved tissue insulin sensitivity by 16% (HOMA index). COPD subjects enjoyed similar benefits as CHF ones. They increased physical autonomy by 78.6% steps/day and decreased resting plasma lactate concentrations by 23%. EAA mechanisms explaining improved exercise intolerance could be increases in muscle aerobic metabolism, mass and function, and improvement of tissue insulin sensitivity (the latter only for the CHF population). These mechanisms could be accounted for by EAA’s intrinsic physiological activity which increases myofibrils and mitochondria genesis in skeletal muscle and myocardium and glucose control. Supplemented EAAs can improve the physical autonomy of subjects with CHF/COPD. Roberto Aquilani, Giuseppe D’Antona, Paola Baiardi, Arianna Gambino, Paolo Iadarola, Simona Viglio, Evasio Pasini, Manuela Verri, Annalisa Barbieri, and Federica Boschi Copyright © 2014 Roberto Aquilani et al. All rights reserved. Feleucins: Novel Bombinin Precursor-Encoded Nonapeptide Amides from the Skin Secretion of Bombina variegata Mon, 09 Jun 2014 07:33:11 +0000 The first amphibian skin antimicrobial peptide (AMP) to be identified was named bombinin, reflecting its origin from the skin of the European yellow-bellied toad (Bombina variegata). Bombinins and their related peptides, the bombinin Hs, were subsequently reported from other bombinid toads. Molecular cloning of bombinin-encoding cDNAs from skin found that bombinins and bombinin Hs were coencoded on the same precursor proteins. Here, we report the molecular cloning of two novel cDNAs from a skin secretion-derived cDNA library of B. variegata whose open-reading frames each encode a novel bombinin (GIGGALLNVGKVALKGLAKGLAEHFANamide) and a C-terminally located single copy of a novel nonapeptide (FLGLLGGLLamide or FLGLIGSLLamide). These novel nonapeptides were named feleucin-BV1 and feleucin-BV2, respectively. The novel bombinin exhibited 89% identity to homologues from the toads, B. microdeladigitora and B. maxima. The feleucins exhibited no identity with any amphibian AMP archived in databases. Synthetic feleucins exhibited a weak activity against Staphylococcus aureus (128–256 mg/L) but feleucin-BV1 exhibited a synergistic action with the novel bombinin. The present report clearly demonstrates that the skin secretions of bombinid toads continue to represent a source of peptides of novel structure that could provide templates for the design of therapeutics. Bing Bai, Xiaojuan Hou, Lei Wang, Lilin Ge, Yu Luo, Chengbang Ma, Mei Zhou, Jinao Duan, Tianbao Chen, and Chris Shaw Copyright © 2014 Bing Bai et al. All rights reserved. Investigating hsp Gene Expression in Liver of Channa striatus under Heat Stress for Understanding the Upper Thermal Acclimation Mon, 09 Jun 2014 06:12:34 +0000 Changes in hsp gene expression profiles in murrel Channa striatus experimentally exposed to temperature stress (36°C) for 4, 15, and 30 days were investigated; fish collected from aquaculture ponds and maintained in laboratory at the pond temperature (25 ± 1°C) served as control. Channa collected from a hot spring runoff (36°C) was included in the study to examine the hsp profiles beyond 30 days of exposure. Gene expression analyses of a battery of hsps in liver tissues were carried out by quantitative RT-PCR and protein expressions were analyzed by immunoblotting. hsps could be grouped into three clusters based on similarity in response to heat stress: hsp70, hsp78, and hsp60, whose transcript level continued to increase with duration of exposure; hsp90 and hsp110 that increased to a much higher level and then decreased; hsp27 and hsp47 that did not significantly vary as compared to control. The results suggest that Hsp70, Hsp78, and Hsp60 are involved in thermal acclimation and long term survival at high temperature. Fish living in the hot spring runoff appears to continuously express hsps that can be approximated by long term induction of hsps in farmed fish if temperature of their environment is raised to 36°C. Gopal Krishna Purohit, Arabinda Mahanty, Mrutyunjay Suar, Anil Prakash Sharma, Bimal Prasanna Mohanty, and Sasmita Mohanty Copyright © 2014 Gopal Krishna Purohit et al. All rights reserved. Comparison of Quasispecies Diversity of HCV between Chronic Hepatitis C and Hepatocellular Carcinoma by Ultradeep Pyrosequencing Thu, 05 Jun 2014 11:59:27 +0000 Backgrounds. Hepatitis C virus (HCV) exists as population of closely related genetic variants known as quasispecies. HCV quasispecies diversity is strongly influenced by host immune pressure on virus. Quasispecies diversity is expected to decline as host immune response to HCV decreases over natural course of progressing from chronic hepatitis C (CHC) to hepatocellular carcinoma (HCC). Methods. Ultradeep pyrosequencing (UDPS) was used to evaluate degree of quasispecies diversity in 49 patients infected with HCV including 26 with CHC and 23 with HCC. Whole structural protein of HCV genome was subjected to UDPS. Results. Shannon’s indices for quasispecies diversity in HCV E1 were significantly lower in patients with HCC than in those with CHC. 14 amino acid positions differed significantly between two groups. Area under curve of ROC analysis for differentiating HCC from CHC was >0.8 for all of 14 amino acid positions. Conclusion. HCV quasispecies diversity as indicator of declining host immune functions was easily assessed by UDPS technology. Shannon’s indices in 14 amino acid positions were found to differentiate between patients with CHC and those with HCC. Our data propose that degree of HCV quasispecies measured by UDPS might be useful to predict progression of HCC in chronic HCV patients. Chang-Wook Park, Min-Chul Cho, Keumrock Hwang, Sun-Young Ko, Heung-Bum Oh, and Han Chu Lee Copyright © 2014 Chang-Wook Park et al. All rights reserved. Biomarkers of Metabolic Disorders: Diagnostic and Prognostic Values, and Insights into the Pathogenesis Thu, 05 Jun 2014 07:50:54 +0000 Cheng Hu, Jiarui Wu, and Wei Jia Copyright © 2014 Cheng Hu et al. All rights reserved. The Cytotoxicity of Benzaldehyde Nitrogen Mustard-2-Pyridine Carboxylic Acid Hydrazone Being Involved in Topoisomerase IIα Inhibition Thu, 05 Jun 2014 07:39:04 +0000 The antitumor property of iron chelators and aromatic nitrogen mustard derivatives has been well documented. Combination of the two pharmacophores in one molecule in drug designation is worth to be explored. We reported previously the syntheses and preliminary cytotoxicity evaluation of benzaldehyde nitrogen mustard pyridine carboxyl acid hydrazones (BNMPH) as extended study, more tumor cell lines (IC50 for HepG2: 26.1 ± 3.5 μM , HCT-116: 57.5 ± 5.3 μM, K562: 48.2 ± 4.0 μM, and PC-12: 19.4 ± 2.2 μM) were used to investigate its cytotoxicity and potential mechanism. In vitro experimental data showed that the BNMPH chelating Fe2+ caused a large number of ROS formations which led to DNA cleavage, and this was further supported by comet assay, implying that ROS might be involved in the cytotoxicity of BNMPH. The ROS induced changes of apoptosis related genes, but the TFR1 and NDRG1 metastatic genes were not obviously regulated, prompting that BNMPH might not be able to deprive Fe2+ of ribonucleotide reductase. The BNMPH induced S phase arrest was different from that of iron chelators (G1) and alkylating agents (G2). BNMPH also exhibited its inhibition of human topoisomerase IIα. Those revealed that the cytotoxic mechanism of the BNMPH could stem from both the topoisomerase II inhibition, ROS generation and DNA alkylation. Yun Fu, Sufeng Zhou, Youxun Liu, Yingli Yang, Xingzhi Sun, and Changzheng Li Copyright © 2014 Yun Fu et al. All rights reserved. Cartilage Dysfunction in ALS Patients as Side Effect of Motion Loss: 3D Mechano-Electrochemical Computational Model Tue, 03 Jun 2014 07:21:21 +0000 Amyotrophic lateral sclerosis (ALS) is a debilitating motor neuron disease characterized by progressive weakness, muscle atrophy, and fasciculation. This fact results in a continuous degeneration and dysfunction of articular soft tissues. Specifically, cartilage is an avascular and nonneural connective tissue that allows smooth motion in diarthrodial joints. Due to the avascular nature of cartilage tissue, cells nutrition and by-product exchange are intermittently occurring during joint motions. Reduced mobility results in a change of proteoglycan density, osmotic pressure, and permeability of the tissue. This work aims to demonstrate the abnormal cartilage deformation in progressive immobilized articular cartilage for ALS patients. For this aim a novel 3D mechano-electrochemical model based on the triphasic theory for charged hydrated soft tissues is developed. ALS patient parameters such as tissue porosity, osmotic coefficient, and fixed anions were incorporated. Considering different mobility reduction of each phase of the disease, results predicted the degree of tissue degeneration and the reduction of its capacity for deformation. The present model can be a useful tool to predict the evolution of joints in ALS patients and the necessity of including specific cartilage protectors, drugs, or maintenance physical activities as part of the symptomatic treatment in amyotrophic lateral sclerosis. Sara Manzano, Eamonn A. Gaffney, Manuel Doblaré, and Mohamed Hamdy Doweidar Copyright © 2014 Sara Manzano et al. All rights reserved. Blood Biomarkers for Amyotrophic Lateral Sclerosis: Myth or Reality? Mon, 02 Jun 2014 08:02:37 +0000 Amyotrophic lateral sclerosis (ALS) is a fatal condition primarily characterized by the selective loss of upper and lower motor neurons. At present, the diagnosis and monitoring of ALS is based on clinical examination, electrophysiological findings, medical history, and exclusion of confounding disorders. There is therefore an undeniable need for molecular biomarkers that could give reliable information on the onset and progression of ALS in clinical practice and therapeutic trials. From a practical point of view, blood offers a series of advantages, including easy handling and multiple testing at a low cost, that make it an ideal source of biomarkers. In this review, we revisited the findings of many studies that investigated the presence of systemic changes at the molecular and cellular level in patients with ALS. The results of these studies reflect the diversity in the pathological mechanisms contributing to disease (e.g., excitotoxicity, oxidative stress, neuroinflammation, metabolic dysfunction, and neurodegeneration, among others) and provide relatively successful evidence of the usefulness of a wide-ranging panel of molecules as potential biomarkers. More studies, hopefully internationally coordinated, would be needed, however, to translate the application of these biomarkers into benefit for patients. Laura Robelin and Jose Luis Gonzalez De Aguilar Copyright © 2014 Laura Robelin and Jose Luis Gonzalez De Aguilar. All rights reserved. Effects of the Angiotensin Receptor Blocker Olmesartan on Adipocyte Hypertrophy and Function in Mice with Metabolic Disorders Mon, 02 Jun 2014 05:45:39 +0000 In the present study, we examined the therapeutic effects of olmesartan, an angiotensin II (Ang II) type 1 receptor (AT1R)-specific blocker, in genetically obese diabetic KKAy mice, a model of human metabolic disorders with visceral obesity, with a focus on an olmesartan effect on the adipose tissue. Olmesartan treatment (3 mg/kg per day) for 4 weeks significantly lowered systolic blood pressure but did not affect body weight during the study period in KKAy mice. However, there were three interesting findings possibly related to the pleiotropic effects of olmesartan on adipose tissue in KKAy mice: (1) an inhibitory effect on adipocyte hypertrophy, (2) a suppressive effect on IL-6 gene expression, and (3) an ameliorating effect on oxidative stress. On the other hand, olmesartan exerted no evident influence on the adipose tissue expression of AT1R-associated protein (ATRAP), which is a molecule interacting with AT1R so as to inhibit pathological AT1R activation and is suggested to be an emerging molecular target in metabolic disorders with visceral obesity. Collectively, these results suggest that the blood pressure lowering effect of olmesartan in KKAy mice is associated with an improvement in adipocyte, including suppression of adipocyte hypertrophy and inhibition of the adipose IL-6-oxidative stress axis. Further study is needed to clarify the functional role of adipose ATRAP in the pleiotropic effects of olmesartan. Akinobu Maeda, Kouichi Tamura, Hiromichi Wakui, Masato Ohsawa, Kengo Azushima, Kazushi Uneda, Tomohiko Kanaoka, Ryu Kobayashi, Kohji Ohki, Miyuki Matsuda, Yuko Tsurumi-Ikeya, Akio Yamashita, Yasuo Tokita, and Satoshi Umemura Copyright © 2014 Akinobu Maeda et al. All rights reserved. Biosorption of Fe(II) and Mn(II) Ions from Aqueous Solution by Rice Husk Ash Sun, 01 Jun 2014 11:06:32 +0000 Rice husk ash (RHA), an agricultural waste, was used as biosorbent for the removal of Iron(II) and Manganese(II) ions from aqueous solutions. The structural and morphological characteristics of RHA and its elemental compositions before and after adsorption of Fe(II) and Mn(II) were determined by scanning electron microscopic (SEM) and X-ray fluorescence (XRF) analyses. Batch experiments were carried out to determine the influence of initial pH, contact time, adsorbent dosage, and initial concentration on the removal of Fe(II) and Mn(II) ions. Langmuir, Freundlich, and Dubinin-Radushkevich (D-R) models were applied to describe the biosorption isotherm of the metal ions by RHA. The correlation coefficient () of Langmuir and Freundlich isotherm models equals 0.995 and 0.901 for Fe(II), 0.9862 and 0.8924 for Mn(II), respectively, so the Langmuir model fitted the equilibrium data better than the Freundlich isotherm model. The mean free energy values evaluated from the D-R model indicated that the biosorption of Fe(II) and Mn(II) onto RHA was physical in nature. Experimental data also showed that the biosorption processes of both metal ions complied with the pseudo-second-order kinetics. Ying Zhang, Jiaying Zhao, Zhao Jiang, Dexin Shan, and Yan Lu Copyright © 2014 Ying Zhang et al. All rights reserved. A-FABP Concentration Is More Strongly Associated with Cardiometabolic Risk Factors and the Occurrence of Metabolic Syndrome in Premenopausal Than in Postmenopausal Middle-Aged Women Thu, 29 May 2014 11:54:06 +0000 We aimed at the evaluation of the relationship between adipocyte fatty acid binding protein (A-FABP) and cardiometabolic risk factors in premenopausal and postmenopausal women. Additionally, we compared A-FABP with adipokines related to metabolic syndrome (MetS) such as leptin and adiponectin. 94 premenopausal and 90 early postmenopausal middle-aged Caucasian women were subject to examinations. Postmenopausal women had higher A-FABP than premenopausal; this difference became insignificant after controlling for age. We found significantly higher correlation coefficients between A-FABP and TC/HDL-C ratio and number of MetS components in premenopausal women, compared to postmenopausal. Each 1 ng/dL increase in A-FABP concentration significantly increased the probability of occurrence of atherogenic lipid profile in premenopausal women, even after multivariate adjustment. All odds ratios became insignificant after controlling for BMI in postmenopausal women. A-FABP was more strongly associated with MetS than leptin and adiponectin in premenopausal women. Adiponectin concentration was a better biomarker for MetS after menopause. Our results suggest that the A-FABP is more strongly associated with some cardiometabolic risk factors in premenopausal than in postmenopausal women. Higher values of A-FABP after menopause are mainly explained by the fact that postmenopausal women are older. Because of the limitation of study, these results should be interpreted with caution. Anna Stefanska, Irena Ponikowska, and Grazyna Sypniewska Copyright © 2014 Anna Stefanska et al. All rights reserved. Misidentification of Candida guilliermondii as C. famata among Strains Isolated from Blood Cultures by the VITEK 2 System Thu, 29 May 2014 11:31:13 +0000 Introduction. The aim of this study was to differentiate between Candida famata and Candida guilliermondii correctly by using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and gene sequencing. Methods. Twenty-eight Candida strains from blood cultures that had been identified as C. famata (), C. famata/C. guilliermondii (), and C. guilliermondii () by the VITEK 2 system using the YST ID card were included. We identified these strains by MALDI-TOF MS and gene sequencing using the 28S rRNA and ITS genes and compared the results with those obtained by the VITEK 2 system. Results. All 28 isolates were finally identified as C. guilliermondii. Sequencing analysis of the 28S rRNA gene showed 99.80%–100% similarity with C. guilliermondii for all 28 strains. The ITS gene sequencing of the strains showed 98.34%–100% homology with C. guilliermondii. By MALDI-TOF, we could correctly identify 21 (75%) of 28 C. guilliermondii isolates. Conclusion. We should suspect misidentification when C. famata is reported by the VITEK 2 system, and we always should keep in mind the possibility of misidentification of any organism when an uncommon species is reported. Si Hyun Kim, Jeong Hwan Shin, Jeong Ha Mok, Shine Young Kim, Sae Am Song, Hye Ran Kim, Joong-Ki Kook, Young-Hyo Chang, Il Kwon Bae, and Kwangha Lee Copyright © 2014 Si Hyun Kim et al. All rights reserved. A Novel Hemagglutinin with Antiproliferative Activity against Tumor Cells from the Hallucinogenic Mushroom Boletus speciosus Thu, 29 May 2014 09:31:59 +0000 Little was known about bioactive compounds from the hallucinogenic mushroom Boletus speciosus. In the present study, a hemagglutinin (BSH, B. speciosus hemagglutinin) was isolated from its fruiting bodies and enzymatic properties were also tested. The chromatographic procedure utilized comprised anion exchange chromatography on Q-Sepharose, cation exchange chromatography on CM-Cellulose, cation exchange chromatography on SP-Sepharose, and gel filtration by FPLC on Superdex 75. The hemagglutinin was a homodimer which was estimated to be approximately 31 kDa in size. The activity of BSH was stable up to 60°C, while there was a precipitous drop in activity when the temperature was elevated to 70°C. BSH retained 25% hemagglutinating activity when exposed to 100 mM NaOH and 25 mM HCl. The activity was potently inhibited by 1.25 mM Hg2+ and slightly inhibited by Fe2+, Ca2+, and Pb2+. None of the sugars tested showed inhibition towards BSH. Its hemagglutinating activity towards human erythrocytes type A, type B, and type AB was higher than type O. The hemagglutinin showed antiproliferative activity towards hepatoma Hep G2 cells and mouse lymphocytic leukemia cells (L1210) in vitro, with IC50 of 4.7 μM and 7.0 μM, respectively. It also exhibited HIV-1 reverse transcriptase inhibitory activity with an IC50 of 7.1 μM. Jian Sun, Tzi-Bun Ng, Hexiang Wang, and Guoqing Zhang Copyright © 2014 Jian Sun et al. All rights reserved. Novel Medicines and Strategies in Cancer Treatment and Prevention Tue, 27 May 2014 06:55:09 +0000 Chih-Hsin Tang, Gautam Sethi, and Po-Lin Kuo Copyright © 2014 Chih-Hsin Tang et al. All rights reserved. Laboratory Markers of Ventricular Arrhythmia Risk in Renal Failure Mon, 26 May 2014 13:45:05 +0000 Sudden cardiac death continues to be a major public health problem. Ventricular arrhythmia is a main cause of sudden cardiac death. The present review addresses the links between renal function tests, several laboratory markers, and ventricular arrhythmia risk in patients with renal disease, undergoing or not hemodialysis or renal transplant, focusing on recent clinical studies. Therapy of hypokalemia, hypocalcemia, and hypomagnesemia should be an emergency and performed simultaneously under electrocardiographic monitoring in patients with renal failure. Serum phosphates and iron, PTH level, renal function, hemoglobin and hematocrit, pH, inflammatory markers, proteinuria and microalbuminuria, and osmolarity should be monitored, besides standard 12-lead ECG, in order to prevent ventricular arrhythmia and sudden cardiac death. Ioana Mozos Copyright © 2014 Ioana Mozos. All rights reserved. Pfetin as a Risk Factor of Recurrence in Gastrointestinal Stromal Tumors Mon, 26 May 2014 00:00:00 +0000 Background. Despite complete resection of gastrointestinal stromal tumors (GIST), recurrent and/or metastatic disease occurs, often depending on the grade of malignancy. As such, markers are needed that accurately predict patients at high risk for recurrence. Previously our group reported Pfetin as a prognostic biomarker for GIST. In order to create an approach for predicting risk of recurrence, we incorporated Pfetin expression with clinicopathological data to produce a predictive model. Object. Forty-five patients with localized primary GIST were treated with complete gross surgical resection surgically at our institution between 1995 and 2010 were included. The majority of tumors originated in the stomach (38 cases), as well as small intestine (6 cases) and rectum (1 case). Method. (1) We performed retrospective analysis of the connection between Pfetin expression, clinicopathological data, and incidences of recurrence, using bivariate and multivariate analyses. (2) The reactivity of the monoclonal antibody against Pfetin was examined by immunohistochemistry. Pfetin. We have reported Pfetin, identified microarray technology, and compared between statistically different GISTs for good and poor prognoses and for prognostic marker. Results. There were 7 cases of recurrences. (1) By univariate analysis, tumor size, mitoses, exposure to abdominal cavity, and complete tumor removal predicted risk of recurrence. (2) Pfetin-negative cases were significantly related to recurrence (P = 0.002). Conclusions. This analysis demonstrates that lack of Pfetin expression is an additional predictor of recurrence in resected GIST. Further study may determine the role of this variable added to the current predictive model for selection of adjuvant therapy. Hajime Orita, Tomoaki Ito, Tomoyuki Kushida, Mutsumi Sakurada, Hiroshi Maekawa, Ryo Wada, Yoshiyuki Suehara, Daisuke Kubota, and Koichi Sato Copyright © 2014 Hajime Orita et al. All rights reserved. Synthesis of Rosin Acid Starch Catalyzed by Lipase Sun, 25 May 2014 05:06:11 +0000 Rosin, an abundant raw material from pine trees, was used as a starting material directly for the synthesis of rosin acid starch. The esterification reaction was catalyzed by lipase (Novozym 435) under mild conditions. Based on single factor experimentation, the optimal esterification conditions were obtained as follows: rosin acid/anhydrous glucose unit in the molar ratio 2 : 1, reaction time 4 h at 45°C, and 15% of lipase dosage. The degree of substitution (DS) reaches 0.098. Product from esterification of cassava starch with rosin acid was confirmed by FTIR spectroscopy and iodine coloration analysis. Scanning electron microscopy and X-ray diffraction analysis showed that the morphology and crystallinity of the cassava starch were largely destroyed. Thermogravimetric analysis indicated that thermal stability of rosin acid starch decreased compared with native starch. Rihui Lin, He Li, Han Long, Jiating Su, and Wenqin Huang Copyright © 2014 Rihui Lin et al. All rights reserved. A Modified Fluorimetric Method for Determination of Hydrogen Peroxide Using Homovanillic Acid Oxidation Principle Mon, 19 May 2014 13:12:08 +0000 Hydrogen peroxide (H2O2) level in biological samples is used as an important index in various studies. Quantification of H2O2 level in tissue fractions in presence of H2O2 metabolizing enzymes may always provide an incorrect result. A modification is proposed for the spectrofluorimetric determination of H2O2 in homovanillic acid (HVA) oxidation method. The modification was included to precipitate biological samples with cold trichloroacetic acid (TCA, 5% w/v) followed by its neutralization with K2HPO4 before the fluorimetric estimation of H2O2 is performed. TCA was used to precipitate the protein portions contained in the tissue fractions. After employing the above modification, it was observed that H2O2 content in tissue samples was ≥2 fold higher than the content observed in unmodified method. Minimum 2 h incubation of samples in reaction mixture was required for completion of the reaction. The stability of the HVA dimer as reaction product was found to be >12 h. The method was validated by using known concentrations of H2O2 and catalase enzyme that quenches H2O2 as substrate. This method can be used efficiently to determine more accurate tissue H2O2 level without using internal standard and multiple samples can be processed at a time with additional low cost reagents such as TCA and K2HPO4. Biswaranjan Paital Copyright © 2014 Biswaranjan Paital. All rights reserved. Evaluation of Assays for Measurement of Serum (Anti)oxidants in Hemodialysis Patients Mon, 19 May 2014 11:20:22 +0000 Background. Various biomarkers and assays have been used for assessment of (anti)oxidant status in hemodialysis patients, including those intended for measurement of serum total (anti)oxidants, most often as a part of panel biomarkers. Methods. Serum (anti)oxidant status was measured in 32 chronically hemodialyzed patients and in 47 healthy persons, using two oxidations and three antioxidant assays. Results. The patients before the hemodialysis session have had higher values of total oxidants in comparison to the healthy persons, with a further increase during the hemodialysis. These findings were confirmed with both oxidation assays, but they differ in the percentage of increase and the statistical significance. All three antioxidant assays showed significantly higher values of the total serum antioxidants in the patients before the hemodialysis session in comparison to the healthy persons, and their significant decrease during the hemodialysis. However, the assays differ in the percentage of decrease, its statistical significance, and the correlations with uric acid. Conclusion. The variability of results of total (anti)oxidants which are obtained using different assays should be taken into account when interpreting data from clinical studies of oxidative stress, especially in complex pathologies such as chronic hemodialysis. Tatjana Ruskovska, Eugene H. J. M. Jansen, and Risto Antarorov Copyright © 2014 Tatjana Ruskovska et al. All rights reserved. Total and Free Serum Sialic Acid Concentration in Liver Diseases Sun, 18 May 2014 14:51:30 +0000 Background. The objective of this study was to compare the levels of total (TSA) and free (FSA) sialic acid in acute and chronic liver diseases. Materials and Methods. The serum TSA and FSA levels were determined in 278 patients suffering from acute and chronic liver diseases of different etiologies. TSA was estimated by enzymatic method and FSA by the thiobarbituric method modified by Skoza and Mohos. Results. There were no significant differences in the serum TSA concentration between liver diseases of different etiologies, although in most of the liver diseases the mean TSA level was significantly lower than that in the control group. In contrast to TSA, the concentration of FSA appears to differ between liver diseases. In toxic hepatitis it was higher than that in nonalcoholic cirrhosis. However, neither of them differs between alcoholic and nonalcoholic cirrhosis or between liver tumors and tumors with cirrhosis. Conclusions. We conclude that the changes in concentrations of TSA and FSA during the same liver diseases indicate significant disturbances in sialylation of serum glycoproteins. Ewa Gruszewska, Bogdan Cylwik, Anatol Panasiuk, Maciej Szmitkowski, Robert Flisiak, and Lech Chrostek Copyright © 2014 Ewa Gruszewska et al. All rights reserved. Gla-Rich Protein Is a Potential New Vitamin K Target in Cancer: Evidences for a Direct GRP-Mineral Interaction Sun, 18 May 2014 11:15:35 +0000 Gla-rich protein (GRP) was described in sturgeon as a new vitamin-K-dependent protein (VKDP) with a high density of Gla residues and associated with ectopic calcifications in humans. Although VKDPs function has been related with γ-carboxylation, the Gla status of GRP in humans is still unknown. Here, we investigated the expression of recently identified GRP spliced transcripts, the γ-carboxylation status, and its association with ectopic calcifications, in skin basal cell and breast carcinomas. GRP-F1 was identified as the predominant splice variant expressed in healthy and cancer tissues. Patterns of γ-carboxylated GRP (cGRP)/undercarboxylated GRP (ucGRP) accumulation in healthy and cancer tissues were determined by immunohistochemistry, using newly developed conformation-specific antibodies. Both GRP protein forms were found colocalized in healthy tissues, while ucGRP was the predominant form associated with tumor cells. Both cGRP and ucGRP found at sites of microcalcifications were shown to have in vitro calcium mineral-binding capacity. The decreased levels of cGRP and predominance of ucGRP in tumor cells suggest that GRP may represent a new target for the anticancer potential of vitamin K. Also, the direct interaction of cGRP and ucGRP with BCP crystals provides a possible mechanism explaining GRP association with pathological mineralization. Carla S. B. Viegas, Marjolein Herfs, Marta S. Rafael, José L. Enriquez, Alexandra Teixeira, Inês M. Luís, Cynthia M. R. van ‘t Hoofd, Alexandre João, Vera L. Maria, Sofia Cavaco, Ana Ferreira, Manuel Serra, Elke Theuwissen, Cees Vermeer, and Dina C. Simes Copyright © 2014 Carla S. B. Viegas et al. All rights reserved. Relationship between Serum Total Cholesterol Level and Serum Biochemical Bone Turnover Markers in Healthy Pre- and Postmenopausal Women Thu, 15 May 2014 12:41:05 +0000 Background. The presence of common risk factors suggests that there is a relationship between osteoporosis and cardiovascular disease, possibly via dyslipidemia and inflammation. We investigated the relationships among the lipid profile, the inflammation marker high-sensitivity C-reactive protein (hsCRP), bone turnover markers, and bone mineral density (BMD) to assess the correlation between osteoporosis and cardiovascular disease and identify factors predicting osteoporosis. Methods. The study included 759 Korean women older than 20 years of age. The BMD, serum lipid profile, and levels of hsCRP, cross-linked C-terminal peptide (CTX), and osteocalcin were measured. We compared the serum biomarkers between groups with normal and low BMD and assessed the correlations between the levels of bone turnover markers and the lipid profile and hsCRP level. Results. The concentrations of CTX, osteocalcin, and total cholesterol were significantly higher in the low BMD group than in the normal BMD group in premenopausal women group. However, hsCRP was not correlated with these parameters. Multivariate logistic regression analysis revealed that TC (OR, 1.647; 95% CI, 1.190–2.279) and osteocalcin (OR, 1.044; 95% CI, 1.002–1.088) had an increased risk of low BMD in premenopausal women. Conclusions. These results indicate that total cholesterol concentration is correlated with the levels of bone turnover markers, suggesting that it might predict osteoporosis in premenopausal women. Tae-Dong Jeong, Woochang Lee, Sung-Eun Choi, Jae Seung Kim, Hong-Kyu Kim, Sung Jin Bae, Sail Chun, and Won-Ki Min Copyright © 2014 Tae-Dong Jeong et al. All rights reserved. Breath Ketone Testing: A New Biomarker for Diagnosis and Therapeutic Monitoring of Diabetic Ketosis Sun, 11 May 2014 13:46:44 +0000 Background. Acetone, β-hydroxybutyric acid, and acetoacetic acid are three types of ketone body that may be found in the breath, blood, and urine. Detecting altered concentrations of ketones in the breath, blood, and urine is crucial for the diagnosis and treatment of diabetic ketosis. The aim of this study was to evaluate the advantages of different detection methods for ketones, and to establish whether detection of the concentration of ketones in the breath is an effective and practical technique. Methods. We measured the concentrations of acetone in the breath using gas chromatography-mass spectrometry and β-hydroxybutyrate in fingertip blood collected from 99 patients with diabetes assigned to groups 1 (−), 2 (±), 3 (+), 4 (++), or 5 (+++) according to urinary ketone concentrations. Results. There were strong relationships between fasting blood glucose, age, and diabetic ketosis. Exhaled acetone concentration significantly correlated with concentrations of fasting blood glucose, ketones in the blood and urine, LDL-C, creatinine, and blood urea nitrogen. Conclusions. Breath testing for ketones has a high sensitivity and specificity and appears to be a noninvasive, convenient, and repeatable method for the diagnosis and therapeutic monitoring of diabetic ketosis. Yue Qiao, Zhaohua Gao, Yong Liu, Yan Cheng, Mengxiao Yu, Lingling Zhao, Yixiang Duan, and Yu Liu Copyright © 2014 Yue Qiao et al. All rights reserved. Mutation Analysis of JAK2V617F, FLT3-ITD, NPM1, and DNMT3A in Chinese Patients with Myeloproliferative Neoplasms Sun, 11 May 2014 12:22:45 +0000 Since the discovery of JAK2V617F tyrosine kinase-activating mutation, several genes have been found mutated in myeloproliferative neoplasms (MPNs). FLT3-ITD, NPM1, and DNMT3A mutations frequently occurred in AML patients and have been found conferred with myeloproliferative neoplasms in mouse model. Therefore, we sought to search for mutations in JAK2V617F, FLT3-ITD, NPM1, and DNMT3A in 129 cases including 120 classic MPN cases and 9 MDS/MPN cases. JAK2V617F mutation was found in 60% of the 120 classic MPNs. However, none of the patients displayed FLT3-ITD and NPM1 mutations; only 2 patients harbored DNMT3A R882 mutation. Further studies including whole-genome sequence will be conducted to investigate the possible involvement of these genes in MPN. Min Wang, Na He, Tian Tian, Lu Liu, Shuang Yu, and Daoxin Ma Copyright © 2014 Min Wang et al. All rights reserved. Novel Molecular Biomarkers at the Blood-Brain Barrier in ALS Sun, 11 May 2014 07:09:17 +0000 Recently neuroinflammation has gained a particular focus as a key mechanism of ALS. Several studies in vivo as well as in vitro have nominated immunoglobulin G (IgG) isolated from ALS patients as an active contributor to disease onset and progression. We have shown that ALS IgG affects astroglial Ca2+ excitability and induces downstream activation of phosphatidylinositol 3-kinase. These studies were hampered by a lack of knowledge of the pathway of entry of immune factors in the CNS. Our MRI data revealed the blood-brain barrier BBB leakage and T cell infiltration into brain parenchyma in ALS G93A rats. Since astrocyte ensheathes blood vessel wall contributing to BBB stability and plays an important role in ALS pathogenesis, we have studied astrocytic membrane proteins water channel aquaporin-4 and the inwardly rectifying potassium channel. In this review, we will summarize data related to BBB disruption with particular emphasis on impaired function of astrocytes in ALS. We will discuss implication of membrane proteins expressed on astrocytic endfeet, aquaporin-4, and inwardly rectifying potassium channel in the pathology of ALS. In addition to ALS-specific IgGs, these membrane proteins are proposed as novel biomarkers of the disease. Danijela Bataveljic, Milena Milosevic, Lidija Radenovic, and Pavle Andjus Copyright © 2014 Danijela Bataveljic et al. All rights reserved. Retracted: Stable Plastid Transformation for High-Level Recombinant Protein Expression: Promises and Challenges Thu, 08 May 2014 16:21:26 +0000 BioMed Research International Copyright © 2014 BioMed Research International. All rights reserved. Association between the Delta Estimated Glomerular Filtration Rate and the Prevalence of Monoclonal Gammopathy of Undetermined Significance in Korean Males Thu, 08 May 2014 12:35:34 +0000 Background. We investigated the association between the reduction in the estimated glomerular filtration rate (eGFR) and the prevalence of monoclonal gammopathy of undetermined significance (MGUS) in Korean males. Methods. We enrolled 723 healthy Korean males. Serum creatinine concentration, serum electrophoresis, serum immunofixation, and the serum free light chain assay were performed. We calculated delta eGFR per year (ΔeGFR/yr). The prevalence of MGUS was compared based on the ΔeGFR/yr and age group. Results. Thirteen (1.8%) of 723 participants exhibited the monoclonal band on serum immunofixation. Prevalence of MGUS by age group was 0.00% (0/172 for 40 years), 1.63% (6/367 for 60 years), and 3.80% (7/184 for >60 years). The median decrease in ΔeGFR/yr was 5.3%. The prevalence of MGUS in participants in their 50s with >5.3% decline in ΔeGFR/yr was significantly higher than those with <5.3% decrease in ΔeGFR/yr (3.16% versus 0.00%; ). The prevalence of MGUS in participants in their 50s with >5.3% decrease in ΔeGFR/yr was similar to that of healthy males in their 60s. Conclusion. Using the rate of reduction in ΔeGFR/yr in healthy Korean males who had their serum creatinine level checked regularly may increase the MGUS detection rate in clinical practice. Tae-Dong Jeong, Woochang Lee, Sail Chun, and Won-Ki Min Copyright © 2014 Tae-Dong Jeong et al. All rights reserved. Platelet Function Tests: A Review of Progresses in Clinical Application Thu, 08 May 2014 12:34:03 +0000 The major goal of traditional platelet function tests has been to screen and diagnose patients who present with bleeding problems. However, as the central role of platelets implicated in the etiology of arterial thrombotic diseases such as myocardial infarction and stroke became widely known, platelet function tests are now being promoted to monitor the efficacy of antiplatelet drugs and also to potentially identify patients at increased risk of thrombosis. Beyond hemostasis and thrombosis, an increasing number of studies indicate that platelets play an integral role in intercellular communication, are mediators of inflammation, and have immunomodulatory activity. As new potential biomarkers and technologies arrive at the horizon, platelet functions testing appears to take on a new aspect. This review article discusses currently available clinical application of platelet function tests, placing emphasis on essential characteristics. Jae-Lim Choi, Shuhua Li, and Jin-Yeong Han Copyright © 2014 Jae-Lim Choi et al. All rights reserved. Erythropoietic Potential of CD34+ Hematopoietic Stem Cells from Human Cord Blood and G-CSF-Mobilized Peripheral Blood Mon, 05 May 2014 13:59:44 +0000 Red blood cell (RBC) supply for transfusion has been severely constrained by the limited availability of donor blood and the emergence of infection and contamination issues. Alternatively, hematopoietic stem cells (HSCs) from human organs have been increasingly considered as safe and effective blood source. Several methods have been studied to obtain mature RBCs from CD34+ hematopoietic stem cells via in vitro culture. Among them, human cord blood (CB) and granulocyte colony-stimulating factor-mobilized adult peripheral blood (mPB) are common adult stem cells used for allogeneic transplantation. Our present study focuses on comparing CB- and mPB-derived stem cells in differentiation from CD34+ cells into mature RBCs. By using CD34+ cells from cord blood and G-CSF mobilized peripheral blood, we showed in vitro RBC generation of artificial red blood cells. Our results demonstrate that CB- and mPB-derived CD34+ hematopoietic stem cells have similar characteristics when cultured under the same conditions, but differ considerably with respect to expression levels of various genes and hemoglobin development. This study is the first to compare the characteristics of CB- and mPB-derived erythrocytes. The results support the idea that CB and mPB, despite some similarities, possess different erythropoietic potentials in in vitro culture systems. Honglian Jin, Han-Soo Kim, Sinyoung Kim, and Hyun Ok Kim Copyright © 2014 Honglian Jin et al. All rights reserved. Mass Spectrometric Analysis of Differentially Expressed Proteins in an Endangered Medicinal Herb, Picrorhiza kurroa Sun, 04 May 2014 00:00:00 +0000 Picrorhiza kurroa grown in the Northwestern Himalayan region is used in various herbal formulations but extensive harvesting of this plant has led it to near extinction. The active constituents responsible for the medicinal properties of P. kurroa have been identified as picroside-I and picroside-II which are present in a particular ratio (1 : 1.5) in herbal formulations like Picroliv. The biosynthetic pathway of picrosides has been partially deciphered till date and needs to be elucidated completely. Review of literature revealed that no information is available as of today on the proteome analysis of Picrorhiza kurroa w.r.t. picroside-II biosynthesis. Therefore, with the aim of identifying proteins associated with picroside biosynthesis in Picrorhiza kurroa, differential protein expression was studied under picroside accumulating versus nonaccumulating conditions using SDS-PAGE. A total of 19 differentially expressed proteins were identified using MALDI-TOF/TOF MS followed by MASCOT search. Proteins involved in diverse functions were identified amongst which the most important proteins were glyceraldehyde-3-phosphate dehydrogenase, 1-aminocyclopropane-1-carboxylate oxidase, photosystem I reaction centre subunit V, 2-oxoglutarate ferrous-dependent oxygenase and putative cytochrome P450 superfamily protein because of their role in picroside biosynthesis. These identified proteins provide an insight and a basic platform for thorough understanding of biosynthesis of secondary metabolites and various other physiological processes of P. kurroa. Amit Sud, Rajinder Singh Chauhan, and Chanderdeep Tandon Copyright © 2014 Amit Sud et al. All rights reserved. CCAAT/Enhancer-Binding Protein α Is a Crucial Regulator of Human Fat Mass and Obesity Associated Gene Transcription and Expression Tue, 29 Apr 2014 14:17:03 +0000 Several susceptibility loci have been reported associated with obesity and T2DM in GWAS. Fat mass and obesity associated gene (FTO) is the first gene associated with body mass index (BMI) and risk for diabetes in diverse patient populations. FTO is highly expressed in the brain and pancreas, and is involved in regulating dietary intake and energy expenditure. While much is known about the epigenetic mutations contributing to obesity and T2DM, less is certain with the expression regulation of FTO gene. In this study, a highly conserved canonical C/EBPα binding site was located around position −45~−54 bp relative to the human FTO gene transcriptional start site. Site-directed mutagenesis of the putative C/EBPα binding sites decreased FTO promoter activity. Overexpression and RNAi studies also indicated that C/EBPα was required for the expression of FTO. Chromatin immunoprecipitation (ChIP) experiment was carried out and the result shows direct binding of C/EBPα to the putative binding regions in the FTO promoter. Collectively, our data suggest that C/EBPα may act as a positive regulator binding to FTO promoter and consequently, activates the gene transcription. Wei Ren, Jianjin Guo, Feng Jiang, Jun Lu, Ying Ding, Aimei Li, Xiubin Liang, and Weiping Jia Copyright © 2014 Wei Ren et al. All rights reserved. Roles of Vascular Endothelial Growth Factor in Amyotrophic Lateral Sclerosis Tue, 29 Apr 2014 13:37:14 +0000 Amyotrophic lateral sclerosis (ALS) is a fatal devastating neurodegenerative disorder, involving progressive degeneration of motor neurons in spinal cord, brainstem, and motor cortex. Riluzole is the only drug approved in ALS but it only confers a modest improvement in survival. In spite of a high number of clinical trials no other drug has proved effectiveness. Recent studies support that vascular endothelial growth factor (VEGF), originally described as a key angiogenic factor, also plays a key role in the nervous system, including neurogenesis, neuronal survival, neuronal migration, and axon guidance. VEGF has been used in exploratory clinical studies with promising results in ALS and other neurological disorders. Although VEGF is a very promising compound, translating the basic science breakthroughs into clinical practice is the major challenge ahead. VEGF-B, presenting a single safety profile, protects motor neurons from degeneration in ALS animal models and, therefore, it will be particularly interesting to test its effects in ALS patients. In the present paper the authors make a brief description of the molecular properties of VEGF and its receptors and review its different features and therapeutic potential in the nervous system/neurodegenerative disease, particularly in ALS. Ana Catarina Pronto-Laborinho, Susana Pinto, and Mamede de Carvalho Copyright © 2014 Ana Catarina Pronto-Laborinho et al. All rights reserved. Evaluation of Three Automated Nucleic Acid Extraction Systems for Identification of Respiratory Viruses in Clinical Specimens by Multiplex Real-Time PCR Mon, 28 Apr 2014 11:52:36 +0000 A total of 84 nasopharyngeal swab specimens were collected from 84 patients. Viral nucleic acid was extracted by three automated extraction systems: QIAcube (Qiagen, Germany), EZ1 Advanced XL (Qiagen), and MICROLAB Nimbus IVD (Hamilton, USA). Fourteen RNA viruses and two DNA viruses were detected using the Anyplex II RV16 Detection kit (Seegene, Republic of Korea). The EZ1 Advanced XL system demonstrated the best analytical sensitivity for all the three viral strains. The nucleic acids extracted by EZ1 Advanced XL showed higher positive rates for virus detection than the others. Meanwhile, the MICROLAB Nimbus IVD system was comprised of fully automated steps from nucleic extraction to PCR setup function that could reduce human errors. For the nucleic acids recovered from nasopharyngeal swab specimens, the QIAcube system showed the fewest false negative results and the best concordance rate, and it may be more suitable for detecting various viruses including RNA and DNA virus strains. Each system showed different sensitivity and specificity for detection of certain viral pathogens and demonstrated different characteristics such as turnaround time and sample capacity. Therefore, these factors should be considered when new nucleic acid extraction systems are introduced to the laboratory. Yoonjung Kim, Mi-Soon Han, Juwon Kim, Aerin Kwon, and Kyung-A Lee Copyright © 2014 Yoonjung Kim et al. All rights reserved. Homocysteine Serum Levels in Diabetic Patients with Non Proliferative, Proliferative and without Retinopathy Mon, 28 Apr 2014 11:47:26 +0000 Homocysteine has been associated with extracellular matrix changes. The diabetic retinopathy is a neurovascular complication of diabetes mellitus and it is the leading cause of vision loss among working adults worldwide. In this study, we evaluate the role of homocysteine in diabetic retinopathy analyzing the plasma levels of homocysteine in 63 diabetic type 2 patients with nonproliferative retinopathy (NPDR), 62 patients with proliferative diabetic retinopathy (PDR), 50 healthy subjects used as control group, and 75 randomly selected patients. Giulia Malaguarnera, Caterina Gagliano, Maria Giordano, Salvatore Salomone, Marco Vacante, Claudio Bucolo, Filippo Caraci, Michele Reibaldi, Filippo Drago, Teresio Avitabile, and Massimo Motta Copyright © 2014 Giulia Malaguarnera et al. All rights reserved. New Insights into c-Ret Signalling Pathway in the Enteric Nervous System and Its Relationship with ALS Mon, 28 Apr 2014 08:49:33 +0000 The receptor tyrosine kinase Ret (c-Ret) transduces the glial cell line-derived neurotrophic factor (GDNF) signal, one of the neurotrophic factors related to the degeneration process or the regeneration activity of motor neurons in amyotrophic lateral sclerosis (ALS). The phosphorylation of several tyrosine residues of c-Ret seems to be altered in ALS. c-Ret is expressed in motor neurons and in the enteric nervous system (ENS) during the embryonic period. The characteristics of the ENS allow using it as model for central nervous system (CNS) study and being potentially useful for the research of human neurological diseases such as ALS. The aim of the present study was to investigate the cellular localization and quantitative evaluation of marker c-Ret in the adult human gut. To assess the nature of c-Ret positive cells, we performed colocalization with specific markers of cells that typically are located in the enteric ganglia. The colocalization of PGP9.5 and c-Ret was preferentially intense in enteric neurons with oval morphology and mostly peripherally localized in the ganglion, so we concluded that the c-Ret receptor is expressed by a specific subtype of enteric neurons in the mature human ENS of the gut. The functional significance of these c-Ret positive neurons is discussed. M. J. Luesma, I. Cantarero, J. M. Álvarez-Dotu, S. Santander, and C. Junquera Copyright © 2014 M. J. Luesma et al. All rights reserved. Naja naja karachiensis Envenomation: Biochemical Parameters for Cardiac, Liver, and Renal Damage along with Their Neutralization by Medicinal Plants Sun, 27 Apr 2014 13:02:37 +0000 Naja naja karachiensis envenomation was found to hit more drastically heart, liver, and kidneys. 400 μg/kg of venom-raised moderate serum levels of ALT ( U/L, ), AST ( U/L, ), urea ( mg/dL, ), creatinine ( mg/dL, ), CK-MB ( U/L, ), and LDH ( U/L, ) were injected in experimental rabbits. However, lethality was enhanced with 800 μg/kg of venom in terms of significant release of ALT ( U/L, ), AST ( U/L, ), urea ( mg/dL, ), creatinine ( mg/dL, ), CK-MB ( U/L, ), and LDH ( U/L, ). Among twenty-eight tested medicinal plant extracts, only Stenolobium stans (L.) Seem was found the best antivenom () compared to the efficacy of standard antidote (ALT  U/L, AST  U/L, urea  mg/dL, creatinine  mg/dL, CK-MB  U/L, and LDH  U/L). Other plant extracts were proved less beneficial and partly neutralized the toxicities posed by cobra venom. However, it is essential in future to isolate and characterize bioactive compound(s) from Stenolobium stans (L.) Seem extract to overcome the complications of snake bite. Muhammad Hassham Hassan Bin Asad, Ghulam Murtaza, Muhammad Ubaid, Durr-e-Sabih, Ashif Sajjad, Rubada Mehmood, Qaisar Mahmood, Muhammad Muzzmil Ansari, Sabiha Karim, Zahid Mehmood, and Izhar Hussain Copyright © 2014 Muhammad Hassham Hassan Bin Asad et al. All rights reserved. Accumulation of Misfolded SOD1 in Dorsal Root Ganglion Degenerating Proprioceptive Sensory Neurons of Transgenic Mice with Amyotrophic Lateral Sclerosis Sun, 27 Apr 2014 10:05:01 +0000 Amyotrophic lateral sclerosis (ALS) is an adult-onset progressive neurodegenerative disease affecting upper and lower motoneurons (MNs). Although the motor phenotype is a hallmark for ALS, there is increasing evidence that systems other than the efferent MN system can be involved. Mutations of superoxide dismutase 1 (SOD1) gene cause a proportion of familial forms of this disease. Misfolding and aggregation of mutant SOD1 exert neurotoxicity in a noncell autonomous manner, as evidenced in studies using transgenic mouse models. Here, we used the mouse model for ALS to detect, by means of conformational-specific anti-SOD1 antibodies, whether misfolded SOD1-mediated neurotoxicity extended to neuronal types other than MNs. We report that large dorsal root ganglion (DRG) proprioceptive neurons accumulate misfolded SOD1 and suffer a degenerative process involving the inflammatory recruitment of macrophagic cells. Degenerating sensory axons were also detected in association with activated microglial cells in the spinal cord dorsal horn of diseased animals. As large proprioceptive DRG neurons project monosynaptically to ventral horn MNs, we hypothesise that a prion-like mechanism may be responsible for the transsynaptic propagation of SOD1 misfolding from ventral horn MNs to DRG sensory neurons. Javier Sábado, Anna Casanovas, Olga Tarabal, Marta Hereu, Lídia Piedrafita, Jordi Calderó, and Josep E. Esquerda Copyright © 2014 Javier Sábado et al. All rights reserved. The E3 Ligase CHIP: Insights into Its Structure and Regulation Thu, 24 Apr 2014 12:37:31 +0000 The carboxy-terminus of Hsc70 interacting protein (CHIP) is a cochaperone E3 ligase containing three tandem repeats of tetratricopeptide (TPR) motifs and a C-terminal U-box domain separated by a charged coiled-coil region. CHIP is known to function as a central quality control E3 ligase and regulates several proteins involved in a myriad of physiological and pathological processes. Recent studies have highlighted varied regulatory mechanisms operating on the activity of CHIP which is crucial for cellular homeostasis. In this review article, we give a concise account of our current knowledge on the biochemistry and regulation of CHIP. Indranil Paul and Mrinal K. Ghosh Copyright © 2014 Indranil Paul and Mrinal K. Ghosh. All rights reserved. Oral and Intraperitoneal Administration of Quercetin Decreased Lymphocyte DNA Damage and Plasma Lipid Peroxidation Induced by TSA In Vivo Wed, 23 Apr 2014 13:42:27 +0000 Our previous study showed that quercetin enhances the anticancer effect of trichostatin A (TSA) in xenograft mice given quercetin intraperitoneally (10 mg/kg, 3 times/week). Herein, we investigate whether quercetin administered orally exerts such an effect and prevents the cytotoxic side effects of TSA. We found that quercetin given orally (20 and 100 mg/kg, 3 times/week) failed to enhance the antitumor effect of TSA although it increased the total quercetin concentration more than quercetin administered intraperitoneally in the plasma. The compound quercetin-3-glucuronide (Q3G) increased the most. However, quercetin administered intraperitoneally increased the total quercetin level in tumor tissues more than oral quercetin. Oral and intraperitoneal administration of quercetin similarly decreased lymphocyte DNA damage and plasma lipid peroxidation level induced by TSA. Furthermore, we found that the enhancing effect of Q3G on the antitumor effect of TSA and the incorporation of Q3G was less than that of quercetin in A549 cells. However, we found that A549 cells possessed the ability to convert Q3G to quercetin. In conclusion, different from quercetin administered intraperitoneally, quercetin administered orally failed to enhance the antitumor effect of TSA because of its metabolic conversion. However, it prevented TSA-induced DNA damage and lipid peroxidation. Shu-Ting Chan, Yi-Chin Lin, Cheng-Hung Chuang, Rong-Jen Shiau, Jiunn-Wang Liao, and Shu-Lan Yeh Copyright © 2014 Shu-Ting Chan et al. All rights reserved. Neuroimaging to Investigate Multisystem Involvement and Provide Biomarkers in Amyotrophic Lateral Sclerosis Thu, 17 Apr 2014 13:29:05 +0000 Neuroimaging allows investigating the extent of neurological systems degeneration in amyotrophic lateral sclerosis (ALS). Advanced MRI methods can detect changes related to the degeneration of upper motor neurons but have also demonstrated the participation of other systems such as the sensory system or basal ganglia, demonstrating in vivo that ALS is a multisystem disorder. Structural and functional imaging also allows studying dysfunction of brain areas associated with cognitive signs. From a biomarker perspective, numerous studies using diffusion tensor imaging showed a decrease of fractional anisotropy in the intracranial portion of the corticospinal tract but its diagnostic value at the individual level remains limited. A multiparametric approach will be required to use MRI in the diagnostic workup of ALS. A promising avenue is the new methodological developments of spinal cord imaging that has the advantage to investigate the two motor system components that are involved in ALS, that is, the lower and upper motor neuron. For all neuroimaging modalities, due to the intrinsic heterogeneity of ALS, larger pooled banks of images with standardized image acquisition and analysis procedures are needed. In this paper, we will review the main findings obtained with MRI, PET, SPECT, and nuclear magnetic resonance spectroscopy in ALS. Pierre-François Pradat and Mohamed-Mounir El Mendili Copyright © 2014 Pierre-François Pradat and Mohamed-Mounir El Mendili. All rights reserved. Genetics of Type 2 Diabetes: Insights into the Pathogenesis and Its Clinical Application Thu, 17 Apr 2014 06:30:37 +0000 With rapidly increasing prevalence, diabetes has become one of the major causes of mortality worldwide. According to the latest studies, genetic information makes substantial contributions towards the prediction of diabetes risk and individualized antidiabetic treatment. To date, approximately 70 susceptibility genes have been identified as being associated with type 2 diabetes (T2D) at a genome-wide significant level (). However, all the genetic loci identified so far account for only about 10% of the overall heritability of T2D. In addition, how these novel susceptibility loci correlate with the pathophysiology of the disease remains largely unknown. This review covers the major genetic studies on the risk of T2D based on ethnicity and briefly discusses the potential mechanisms and clinical utility of the genetic information underlying T2D. Xue Sun, Weihui Yu, and Cheng Hu Copyright © 2014 Xue Sun et al. All rights reserved. Detection of Herpes Simplex and Varicella-Zoster Virus in Clinical Specimens by Multiplex Real-Time PCR and Melting Curve Analysis Wed, 16 Apr 2014 15:54:38 +0000 Herpes simplex viruses types 1 and 2 (HSV-1 and HSV-2), and varicella-zoster virus (VZV) are common agents resulting in various forms of clinical manifestation from skin vesicle to disseminated viral infection. The aim of the present study was to develop a real-time PCR and melting curve analysis which detect and differentiate HSV-1, HSV-2, and VZV, to compare with PCR-RFLP using clinical specimens, and to introduce the 4-year experience in the clinical laboratory. Three pairs of primers for HSV-1, HSV-2, and VZV were designed. Primers for human endogenous retrovirus-3 (HERV-3), an internal control, were adopted. A hundred selected specimens and many clinical specimens were tested for methods comparison and assay validation. Increased sensitivity and specificity were obtained from real-time PCR. In review of results of clinical specimens submitted to clinical laboratory, a total of 46 of 3,513 specimens were positive in cerebrospinal fluids, blood, skin vesicles, genital swabs, aqueous humor, and ear discharge. Thus, this method could be a rapid and accurate alternative to virus culture and other molecular tests for detection and typing of HSV-1, HSV-2, and VZV. Yun Ji Hong, Mi Suk Lim, Sang Mee Hwang, Taek Soo Kim, Kyoung Un Park, Junghan Song, and Eui Chong Kim Copyright © 2014 Yun Ji Hong et al. All rights reserved. Effect of the Combination of Gelam Honey and Ginger on Oxidative Stress and Metabolic Profile in Streptozotocin-Induced Diabetic Sprague-Dawley Rats Wed, 16 Apr 2014 13:58:52 +0000 Diabetic complications occur as a result of increased reactive oxygen species (ROS) due to long term hyperglycaemia. Honey and ginger have been shown to exhibit antioxidant activity which can scavenge ROS. The main aim of this study was to evaluate the antioxidant and antidiabetic effects of gelam honey, ginger, and their combination. Sprague-Dawley rats were divided into 2 major groups which consisted of diabetic and nondiabetic rats. Diabetes was induced with streptozotocin intramuscularly (55 mg/kg body weight). Each group was further divided into 4 smaller groups according to the supplements administered: distilled water, honey (2 g/kg body weight), ginger (60 mg/kg body weight), and honey + ginger. Body weight and glucose levels were recorded weekly, while blood from the orbital sinus was obtained after 3 weeks of supplementation for the estimation of metabolic profile: glucose, triglyceride (TG), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), reduced glutathione (GSH): oxidized glutathione (GSSG), and malondialdehyde (MDA). The combination of gelam honey and ginger did not show hypoglycaemic potential; however, the combination treatment reduced significantly () SOD and CAT activities as well as MDA level, while GSH level and GSH/GSSG ratio were significantly elevated () in STZ-induced diabetic rats compared to diabetic control rats. Nur Fathiah Abdul Sani, Levin Kesu Belani, Chong Pui Sin, Siti Nor Amilah Abdul Rahman, Srijit Das, Thent Zar Chi, Suzana Makpol, and Yasmin Anum Mohd Yusof Copyright © 2014 Nur Fathiah Abdul Sani et al. All rights reserved. Plastic Changes in the Spinal Cord in Motor Neuron Disease Wed, 16 Apr 2014 08:56:03 +0000 In the present paper, we analyze the cell number within lamina X at the end stage of disease in a G93A mouse model of ALS; the effects induced by lithium; the stem-cell like phenotype of lamina X cells during ALS; the differentiation of these cells towards either a glial or neuronal phenotype. In summary we found that G93A mouse model of ALS produces an increase in lamina X cells which is further augmented by lithium administration. In the absence of lithium these nestin positive stem-like cells preferentially differentiate into glia (GFAP positive), while in the presence of lithium these cells differentiate towards a neuron-like phenotype (βIII-tubulin, NeuN, and calbindin-D28K positive). These effects of lithium are observed concomitantly with attenuation in disease progression and are reminiscent of neurogenetic effects induced by lithium in the subependymal ventricular zone of the hippocampus. Francesco Fornai, Michela Ferrucci, Paola Lenzi, Alessandra Falleni, Francesca Biagioni, Marina Flaibani, Gabriele Siciliano, Francesco Giannessi, and Antonio Paparelli Copyright © 2014 Francesco Fornai et al. All rights reserved. Metabolomic Prediction of Pregnancy Viability in Superovulated Cattle Embryos and Recipients with Fourier Transform Infrared Spectroscopy Tue, 15 Apr 2014 00:00:00 +0000 We analyzed embryo culture medium (CM) and recipient blood plasma using Fourier transform infrared spectroscopy (FTIR) metabolomics to identify spectral models predictive of pregnancy outcome. Embryos collected on Day 6 from superovulated cows in 2 countries were individually cultured in synthetic oviduct fluid medium with BSA for 24 h before embryo transfer. Spent CM, blank controls, and plasma samples (Day 0 and Day 7) were evaluated using FTIR. The spectra obtained were analyzed. The discrimination capability of the classifiers was assessed for accuracy, sensitivity (pregnancy), specificity (nonpregnancy), and area under the ROC curve (AUC). Endpoints considered were Day 60 pregnancy and birth. High AUC was obtained for Day 60 pregnancy in CM within individual laboratories (France , Spain ), while cumulative data decreased the AUC (). Predictions for CM at birth were lower than Day 60 pregnancy. Predictions with plasma at birth improved cumulative over individual results (Day 0: France ; Spain ; cumulative ). Plasma generally predicted pregnancy and birth better than CM. These first results show that FTIR metabolomics could allow the identification of embryos and recipients with improved pregnancy viability, which may contribute to increasing the efficiency of selection schemes based on ET. Marta Muñoz, Asli Uyar, Eva Correia, Claire Ponsart, Catherine Guyader-Joly, Daniel Martínez-Bello, Brigitte Marquant-Le Guienne, Alfonso Fernandez-Gonzalez, Carmen Díez, Jose Nestor Caamaño, Beatriz Trigal, Patrice Humblot, Susana Carrocera, David Martin, Emre Seli, and Enrique Gomez Copyright © 2014 Marta Muñoz et al. All rights reserved. Regulation of p63 Protein Stability via Ubiquitin-Proteasome Pathway Tue, 15 Apr 2014 00:00:00 +0000 The p53-related p63 gene encodes multiple protein isoforms, which are involved in a variety of biological activities. p63 protein stability is mainly regulated by the ubiquitin-dependent proteasomal degradation pathway. Several ubiquitin E3 ligases have been identified and some protein kinases as well as other kinds of proteins are involved in regulation of p63 protein stability. These regulators are responsive to diverse extracellular signaling, resulting in changes of the p63 protein levels and impacting different biological processes. Chenghua Li and Zhi-Xiong Xiao Copyright © 2014 Chenghua Li and Zhi-Xiong Xiao. All rights reserved. Castanea sativa Mill. Flowers amongst the Most Powerful Antioxidant Matrices: A Phytochemical Approach in Decoctions and Infusions Mon, 14 Apr 2014 17:22:12 +0000 Infusions and decoction of chestnut tree flowers have been used for different medical purposes, but their phytochemical profile and antioxidant activity are still mostly unknown. Herein, decoctions and infusions of flowers from the two most appreciated chestnut cultivars (longal and judia) in Trás-os-Montes, Portugal, were prepared and characterized with regard to their content in free sugars, organic acids, and phenolic compounds, such as flavonoids and hydrolyzable tannins, and their antioxidant activity. Overall, the decoction of the cultivar judia was the sample with both the highest quantity of flavonoids and antioxidant activity. The phenolic compound with the highest abundance in all samples was trigalloyl-HHDP-glucoside, followed by pentagalloyl glucoside. The sample with the highest quantity of total phenolic compounds was judia infusion, closely followed by longal decoction, which also gave the highest quantities of ellagitannins. Regarding sugars and organic acids, the profiles were more similar. These results corroborate ancestral claims of the health benefits of infusions and decoctions of chestnut flowers. Márcio Carocho, Lillian Barros, Albino Bento, Celestino Santos-Buelga, Patricia Morales, and Isabel C. F. R. Ferreira Copyright © 2014 Márcio Carocho et al. All rights reserved. Preliminary Phytochemical Screening and In Vitro Antioxidant Activities of Parkinsonia aculeata Linn. Sun, 13 Apr 2014 11:12:47 +0000 Butanol and hexane leaves extracts of Parkinsonia aculeata L. (Fabaceae) were assessed for its antioxidant potential by in vitro methods. Phytochemical analysis and antioxidant activity of plant extracts were studied using different in vitro assays. UPLC analysis of extracts was carried out for the identification of chemical constituents. The total phenolic contents of the butanol and hexane leaf extract were 42 mgGAE/g and 34 mgGAE/g whereas flavonoid contents of these extracts were found to be 0.044 mgRE/g and 0.005 mgRE/g, respectively. Among both extracts, butanol extract shows maximum inhibition (%) of 93.88%, 80.02%, 52.06%, 94.68%, and 69.37% in DPPH, non-site-specific and site-specific, FTC, and TBA assays and absorbance of 0.852 and 0.522 in reducing power and CUPRAC assay at the highest concentration tested. The FRAP and TAC values of butanol extract were found to be 678 μM Fe(II)/g and 36 mgAAE/100 mg. UPLC analysis of extracts revealed the presence of various polyphenols. The tested plant extracts were found to possess potent antioxidant and free radical scavenging activity which may be due to the presence of flavonoids and polyphenols. Sonia Sharma and Adarsh Pal Vig Copyright © 2014 Sonia Sharma and Adarsh Pal Vig. All rights reserved. A Paradoxical Chemoresistance and Tumor Suppressive Role of Antioxidant in Solid Cancer Cells: A Strange Case of Dr. Jekyll and Mr. Hyde Thu, 03 Apr 2014 08:23:03 +0000 Modulation of intracellular antioxidant concentration is a double-edged sword, with both sides exploited for potential therapeutic benefits. While antioxidants may hamper the efficacy of chemotherapy by scavenging reactive oxygen species and free radicals, it is also possible that antioxidants alleviate unwanted chemotherapy-induced toxicity, thus allowing for increased chemotherapy doses. Under normoxic environment, antioxidants neutralize toxic oxidants, such as reactive oxygen species (ROS), maintaining them within narrow boundaries level. This redox balance is achieved by various scavenging systems such as enzymatic system (e.g., superoxide dismutases, catalase, and peroxiredoxins), nonenzymatic systems (e.g., glutathione, cysteine, and thioredoxin), and metal-binding proteins (e.g., ferritin, metallothionein, and ceruloplasmin) that sequester prooxidant metals inhibiting their participation in redox reactions. On the other hand, therapeutic strategies that promote oxidative stress and eventually tumor cells apoptosis have been explored based on availability of chemotherapy agents that inhibit ROS-scavenging systems. These contradictory assertions suggest that antioxidant supplementation during chemotherapy treatment can have varied outcomes depending on the tumor cellular context. Therefore, understanding the antioxidant-driven molecular pathways might be crucial to design new therapeutic strategies to fight cancer progression. Jolie Kiemlian Kwee Copyright © 2014 Jolie Kiemlian Kwee. All rights reserved. Effects of Chlorophenoxy Herbicides and Their Main Transformation Products on DNA Damage and Acetylcholinesterase Activity Thu, 27 Mar 2014 14:17:26 +0000 Persistent pesticide transformation products (TPs) are increasingly being detected among different environmental compartments, including groundwater and surface water. However, there is no sufficient experimental data on their toxicological potential to assess the risk associated with TPs, even if their occurrence is known. In this study, the interaction of chlorophenoxy herbicides (MCPA, mecoprop, 2,4-D and dichlorprop) and their main transformation products with calf thymus DNA by UV-visible absorption spectroscopy has been assessed. Additionally, the toxicity of the chlorophenoxy herbicides and TPs was also assessed evaluating the inhibition of acetylcholinesterase activity. On the basis of the results found, it seems that AChE is not the main target of chlorophenoxy herbicides and their TPs. However, the results found showed that the transformation products displayed a higher inhibitory activity when compared with the parent herbicides. The results obtained in the DNA interaction studies showed, in general, a slight effect on the stability of the double helix. However, the data found for 4-chloro-2-methyl-6-nitrophenol suggest that this transformation product can interact with DNA through a noncovalent mode. Sofia Benfeito, Tiago Silva, Jorge Garrido, Paula B. Andrade, M. J. Sottomayor, Fernanda Borges, and E. Manuela Garrido Copyright © 2014 Sofia Benfeito et al. All rights reserved. Significance of Lewis Phenotyping Using Saliva and Gastric Tissue: Comparison with the Lewis Phenotype Inferred from Lewis and Secretor Genotypes Mon, 24 Mar 2014 12:11:12 +0000 Lewis phenotypes using various types of specimen were compared with the Lewis phenotype predicted from Lewis and Secretor genotypes. This is the first logical step in explaining the association between the Lewis expression and Helicobacter pylori. We performed a study of the followings on 209 patients who underwent routine gastroscopy: erythrocyte and saliva Lewis phenotyping, gastric Lewis phenotyping by the tissue array, and the Lewis and Secretor genes genotyping. The results of phenotyping were as follows [Le(a−b−), Le(a+b−), Le(a−b+), and Le(a+b+), respectively, in order]: erythrocyte (12.4%, 25.8%, 61.2%, and 0.5%); saliva (2.4%, 27.3%, 70.3%, and 0.0%); gastric mucosa (8.1%, 6.7%, 45.5%, and 39.7%). The frequency of Le, , , and alleles was 74.6%, 21.3%, 3.1%, and 1.0%, respectively, among 418 alleles. The saliva Lewis phenotype was completely consistent with the Lewis phenotype inferred from Lewis and Secretor genotypes, but that of gastric mucosa could not be predicted from genotypes. Lewis phenotyping using erythrocytes is only adequate for transfusion needs. Saliva testing for the Lewis phenotype is a more reliable method for determining the peripheral Lewis phenotype of an individual and the gastric Lewis phenotype must be used for the study on the association between Helicobacter pylori and the Lewis phenotype. Yun Ji Hong, Sang Mee Hwang, Taek Soo Kim, Eun Young Song, Kyoung Un Park, Junghan Song, and Kyou-Sup Han Copyright © 2014 Yun Ji Hong et al. All rights reserved. Lack of Association between TLR4 Genetic Polymorphisms and Diabetic Nephropathy in a Chinese Population Sun, 23 Mar 2014 12:37:36 +0000 Objective. Toll-like receptor 4 (TLR4) plays a central role in innate immunity. Activation of innate immune response and subsequent chronic low-grade inflammation are thought to be involved in the pathogenesis of diabetic nephropathy. In this study, we aimed to investigate whether TLR4 variants are associated with diabetic nephropathy in the Chinese population. Methods. Seven tagging single nucleotide polymorphisms (SNPs) of TLR4 based on HapMap Chinese data were genotyped in 1,455 Chinese type 2 diabetic patients. Of these patients, 622 were diagnosed with diabetic nephropathy and 833 were patients with diabetes for over 5 years but without diabetic nephropathy. Results. None of the SNPs and haplotypes showed significant association to diabetic nephropathy in our study. No association between the SNPs and quantitative traits was observed either. Conclusion. We concluded that common variants within TLR4 genes were not associated with diabetic nephropathy in the Chinese type 2 diabetes patients. Danfeng Peng, Jie Wang, Jiemin Pan, Rong Zhang, Shanshan Tang, Feng Jiang, Miao Chen, Jing Yan, Xue Sun, Tao Wang, Shiyun Wang, Yuqian Bao, and Weiping Jia Copyright © 2014 Danfeng Peng et al. All rights reserved. A Short-Term Incubation with High Glucose Impairs VASP Phosphorylation at Serine 239 in response to the Nitric Oxide/cGMP Pathway in Vascular Smooth Muscle Cells: Role of Oxidative Stress Sun, 23 Mar 2014 08:57:26 +0000 A reduction of the nitric oxide (NO) action in vascular smooth muscle cells (VSMC) could play a role in the vascular damage induced by the glycaemic excursions occurring in diabetic patients; in this study, we aimed to clarify whether a short-term incubation of cultured VSMC with high glucose reduces the NO ability to increase cGMP and the cGMP ability to phosphorylate VASP at Ser-239. We observed that a 180 min incubation of rat VSMC with 25 mmol/L glucose does not impair the NO-induced cGMP increase but reduces VASP phosphorylation in response to both NO and cGMP with a mechanism blunted by antioxidants. We further demonstrated that high glucose increases radical oxygen species (ROS) production and that this phenomenon is prevented by the PKC inhibitor chelerythrine and the NADPH oxidase inhibitor apocynin. The following sequence of events is supported by these results: (i) in VSMC high glucose activates PKC; (ii) PKC activates NADPH oxidase; (iii) NADPH oxidase induces oxidative stress; (iv) ROS impair the signalling of cGMP, which is involved in the antiatherogenic actions of NO. Thus, high glucose, via oxidative stress, can reduce the cardiovascular protection conferred by the NO/cGMP pathway via phosphorylation of the cytoskeleton protein VASP in VSMC. Isabella Russo, Michela Viretto, Gabriella Doronzo, Cristina Barale, Luigi Mattiello, Giovanni Anfossi, and Mariella Trovati Copyright © 2014 Isabella Russo et al. All rights reserved. Role of Plasma Membrane Caveolae/Lipid Rafts in VEGF-Induced Redox Signaling in Human Leukemia Cells Tue, 11 Mar 2014 09:24:57 +0000 Caveolae/lipid rafts are membrane-rich cholesterol domains endowed with several functions in signal transduction and caveolin-1 (Cav-1) has been reported to be implicated in regulating multiple cancer-associated processes, ranging from tumor growth to multidrug resistance and angiogenesis. Vascular endothelial growth factor receptor-2 (VEGFR-2) and Cav-1 are frequently colocalized, suggesting an important role played by this interaction on cancer cell survival and proliferation. Thus, our attention was directed to a leukemia cell line (B1647) that constitutively produces VEGF and expresses the tyrosine-kinase receptor VEGFR-2. We investigated the presence of VEGFR-2 in caveolae/lipid rafts, focusing on the correlation between reactive oxygen species (ROS) production and glucose transport modulation induced by VEGF, peculiar features of tumor proliferation. In order to better understand the involvement of VEGF/VEGFR-2 in the redox signal transduction, we evaluated the effect of different compounds able to inhibit VEGF interaction with its receptor by different mechanisms, corroborating the obtained results by immunoprecipitation and fluorescence techniques. Results here reported showed that, in B1647 leukemia cells, VEGFR-2 is present in caveolae through association with Cav-1, demonstrating that caveolae/lipid rafts act as platforms for negative modulation of VEGF redox signal transduction cascades leading to glucose uptake and cell proliferation, suggesting therefore novel potential targets. Cristiana Caliceti, Laura Zambonin, Benedetta Rizzo, Diana Fiorentini, Francesco Vieceli Dalla Sega, Silvana Hrelia, and Cecilia Prata Copyright © 2014 Cristiana Caliceti et al. All rights reserved. Role of G Protein-Coupled Receptors in Control of Dendritic Cell Migration Mon, 10 Mar 2014 13:55:50 +0000 Dendritic cells (DCs) are highly efficient antigen-presenting cells. The migratory properties of DCs give them the capacity to be a sentinel of the body and the vital role in the induction and regulation of adaptive immune responses. Therefore, it is important to understand the mechanisms in control of migration of DCs to lymphoid and nonlymphoid tissues. This may provide us novel insight into the clinical treatment of diseases such as autoimmune disease, infectious disease, and tumor. The chemotactic G protein-coupled receptors (GPCR) play a vital role in control of DCs migration. Here, we reviewed the recent advances regarding the role of GPCR in control of migration of subsets of DCs, with a focus on the chemokine receptors. Understanding subsets of DCs migration could provide a rational basis for the design of novel therapies in various clinical conditions. Yuan Liu and Guixiu Shi Copyright © 2014 Yuan Liu and Guixiu Shi. All rights reserved. Role of Methylglyoxal in Alzheimer’s Disease Sun, 09 Mar 2014 11:29:36 +0000 Alzheimer’s disease is the most common and lethal neurodegenerative disorder. The major hallmarks of Alzheimer’s disease are extracellular aggregation of amyloid β peptides and, the presence of intracellular neurofibrillary tangles formed by precipitation/aggregation of hyperphosphorylated tau protein. The etiology of Alzheimer’s disease is multifactorial and a full understanding of its pathogenesis remains elusive. Some years ago, it has been suggested that glycation may contribute to both extensive protein cross-linking and oxidative stress in Alzheimer’s disease. Glycation is an endogenous process that leads to the production of a class of compounds known as advanced glycation end products (AGEs). Interestingly, increased levels of AGEs have been observed in brains of Alzheimer’s disease patients. Methylglyoxal, a reactive intermediate of cellular metabolism, is the most potent precursor of AGEs and is strictly correlated with an increase of oxidative stress in Alzheimer’s disease. Many studies are showing that methylglyoxal and methylglyoxal-derived AGEs play a key role in the etiopathogenesis of Alzheimer's disease. Cristina Angeloni, Laura Zambonin, and Silvana Hrelia Copyright © 2014 Cristina Angeloni et al. All rights reserved. PPAR-γ Impairment Alters Peroxisome Functionality in Primary Astrocyte Cell Cultures Tue, 04 Mar 2014 10:03:46 +0000 Peroxisomes provide glial cells with protective functions against the harmful effects of H2O2 on neurons and peroxisome impairment results in nervous lesions. Agonists of the γ-subtype of the Peroxisome-Proliferator-Activated-Receptors (PPAR) have been proposed as neuroprotective agents in neurodegenerative disorders. Nevertheless, the role of PPAR-γ alterations in pathophysiological mechanisms and the relevance of peroxisome functions in the PPAR-γ effects are not yet clear. In a primary cell culture of rat astrocytes, the irreversible PPAR-γ antagonist GW9662 concentration-dependently decreased the activity of catalase, the most important antioxidant defense enzyme in peroxisomes. Catalase functionality recovered in a few days and the PPAR-γ agonist rosiglitazone promoted reversal of enzymatic damage. The reversible antagonist G3335 reduced both the activity and expression of catalase in a rosiglitazone-prevented manner. G3335 reduced also the glutathione reductase expression, indicating that enzyme involved in glutathione regeneration was compromised. Neither the PPAR-α target gene Acyl-Coenzyme-A-oxidase-1 nor the mitochondrial detoxifying enzyme NADH:ubiquinone-oxidoreductase (NDFUS3) was altered by PPAR-γ inhibition. In conclusion, PPAR-γ inhibition induced impairment of catalase in astrocytes. A general decrease of the antioxidant defenses of the cell suggests that a PPAR-γ hypofunction could participate in neurodegenerative mechanisms through peroxisomal damage. This series of experiments could be a useful model for studying compounds able to restore peroxisome functionality. Lorenzo Di Cesare Mannelli, Matteo Zanardelli, Laura Micheli, and Carla Ghelardini Copyright © 2014 Lorenzo Di Cesare Mannelli et al. All rights reserved. Hepatitis B Virus X Upregulates HuR Protein Level to Stabilize HER2 Expression in Hepatocellular Carcinoma Cells Thu, 27 Feb 2014 13:48:52 +0000 Hepatitis B virus- (HBV-) associated hepatocellular carcinoma (HCC) is the most common type of liver cancer. However, the underlying mechanism of HCC tumorigenesis is very complicated and HBV-encoded X protein (HBx) has been reported to play the most important role in this process. Activation of downstream signal pathways of epidermal growth factor receptor (EGFR) family is known to mediate HBx-dependent HCC tumor progression. Interestingly, HER2 (also known as ErbB2/Neu/EGFR2) is frequently overexpressed in HBx-expressing HCC patients and is associated with their poor prognosis. However, it remains unclear whether and how HBx regulates HER2 expression. In this study, our data showed that HBx expression increased HER2 protein level via enhancing its mRNA stability. The induction of RNA-binding protein HuR expression by HBx mediated the HER2 mRNA stabilization. Finally, the upregulated HER2 expression promoted the migration ability of HBx-expressing HCC cells. These findings deciphered the molecular mechanism of HBx-mediated HER2 upregulation in HBV-associated HCC. Chao-Ming Hung, Wei-Chien Huang, Hsiao-Lin Pan, Pei-Hsuan Chien, Chih-Wen Lin, Lei-Chin Chen, Yu-Fong Chien, Ching-Chiao Lin, Kar-Hee Leow, Wen-Shu Chen, Jhen-Yu Chen, Chien-Yi Ho, Pao-Sheng Hou, and Yun-Ju Chen Copyright © 2014 Chao-Ming Hung et al. All rights reserved. Nuclear Nox4-Derived Reactive Oxygen Species in Myelodysplastic Syndromes Wed, 26 Feb 2014 07:31:48 +0000 A role for intracellular ROS production has been recently implicated in the pathogenesis and progression of a wide variety of neoplasias. ROS sources, such as NAD(P)H oxidase (Nox) complexes, are frequently activated in AML (acute myeloid leukemia) blasts and strongly contribute to their proliferation, survival, and drug resistance. Myelodysplastic syndromes (MDS) comprise a heterogeneous group of disorders characterized by ineffective hematopoiesis, with an increased propensity to develop AML. The molecular basis for MDS progression is unknown, but a key element in MDS disease progression is the genomic instability. NADPH oxidases are now recognized to have specific subcellular localizations, this targeting to specific compartments for localized ROS production. Local Nox-dependent ROS production in the nucleus may contribute to the regulation of redox-dependent cell growth, differentiation, senescence, DNA damage, and apoptosis. We observed that Nox1, 2, and 4 isoforms and p22phox and Rac1 subunits are expressed in MDS/AML cell lines and MDS samples, also in the nuclear fractions. Interestingly, Nox4 interacts with ERK and Akt1 within nuclear speckle domain, suggesting that Nox4 could be involved in regulating gene expression and splicing factor activity. These data contribute to the elucidation of the molecular mechanisms used by nuclear ROS to drive MDS evolution to AML. Marianna Guida, Tullia Maraldi, Francesca Beretti, Matilde Y. Follo, Lucia Manzoli, and Anto De Pol Copyright © 2014 Marianna Guida et al. All rights reserved. Altered Nitrogen Balance and Decreased Urea Excretion in Male Rats Fed Cafeteria Diet Are Related to Arginine Availability Mon, 24 Feb 2014 09:02:07 +0000 Hyperlipidic diets limit glucose oxidation and favor amino acid preservation, hampering the elimination of excess dietary nitrogen and the catabolic utilization of amino acids. We analyzed whether reduced urea excretion was a consequence of higher ; (nitrite, nitrate, and other derivatives) availability caused by increased nitric oxide production in metabolic syndrome. Rats fed a cafeteria diet for 30 days had a higher intake and accumulation of amino acid nitrogen and lower urea excretion. There were no differences in plasma nitrate or nitrite. and creatinine excretion accounted for only a small part of total nitrogen excretion. Rats fed a cafeteria diet had higher plasma levels of glutamine, serine, threonine, glycine, and ornithine when compared with controls, whereas arginine was lower. Liver carbamoyl-phosphate synthetase I activity was higher in cafeteria diet-fed rats, but arginase I was lower. The high carbamoyl-phosphate synthetase activity and ornithine levels suggest activation of the urea cycle in cafeteria diet-fed rats, but low arginine levels point to a block in the urea cycle between ornithine and arginine, thereby preventing the elimination of excess nitrogen as urea. The ultimate consequence of this paradoxical block in the urea cycle seems to be the limitation of arginine production and/or availability. David Sabater, Silvia Agnelli, Sofía Arriarán, José-Antonio Fernández-López, María del Mar Romero, Marià Alemany, and Xavier Remesar Copyright © 2014 David Sabater et al. All rights reserved. A Novel Mutation in Leptin Gene Is Associated with Severe Obesity in Chinese Individuals Sun, 23 Feb 2014 11:30:33 +0000 Obesity is a clinical syndrome which is driven by interactions between multiple genetic and environmental factors. Monogenic obesity is a rare type of obesity which is caused by a mutation in a single gene. Patients with monogenic obesity may develop early onset of obesity and severe metabolic abnormalities. In this study, we screened mutations of LEP in a total of 135 Chinese individuals including 35 obese patients whose BMI ≥32 kg/m2 and 100 controls with BMI <25 kg/m2. Moreover, detailed information and clinical measurements of the participants were also collected. Finally, we identified a novel nonsynonymous mutation H118L in exon 3 of LEP in one patient with BMI 46.0 kg/m2. This mutation was not identified in the controls. We speculated that the mutation H118L in LEP might be associated with severe obesity in Chinese subjects. However, the substantial mechanism should be further investigated. Yue Zhao, Nanchao Hong, Xiao Liu, Beibei Wu, Shanshan Tang, Jianjun Yang, Cheng Hu, and Weiping Jia Copyright © 2014 Yue Zhao et al. All rights reserved. Trichostatin A Suppresses EGFR Expression through Induction of MicroRNA-7 in an HDAC-Independent Manner in Lapatinib-Treated Cells Sun, 23 Feb 2014 00:00:00 +0000 Lapatinib, a dual EGFR/HER2 tyrosine kinase inhibitor, has been shown to improve the survival rate of patients with advanced HER2-positive breast cancers. However, the off-target activity of lapatinib in inducing EGFR expression without tyrosine kinase activity was demonstrated to render HER2-negative breast cancer cells more metastatic, suggesting a limitation to the therapeutic effectiveness of this dual inhibitor in HER2-heterogeneous tumors. Therefore, targeting EGFR expression may be a feasible approach to improve the anticancer efficiency of lapatinib-based therapy. Inhibition of HDAC has been previously reported to epigenetically suppress EGFR protein expression. In this study, however, our data indicated that treatment with HDAC inhibitors trichostatin A (TSA), but not suberoylanilide hydroxamic acid (SAHA) or HDAC siRNA, can attenuate both protein and mRNA expressions of EGFR in lapatinib-treated triple-negative breast cancer cells, suggesting that TSA may suppress EGFR expression independently of HDAC inhibition. Nevertheless, TSA reduced EGFR 3′UTR activity and induced the gene expression of microRNA-7, a known EGFR-targeting microRNA. Furthermore, treatment with microRNA-7 inhibitor attenuated TSA-mediated EGFR suppression. These results suggest that TSA induced microRNA-7 expression to downregulate EGFR expression in an HDAC-independent manner. Chih-Yen Tu, Chia-Hung Chen, Te-Chun Hsia, Min-Hsiang Hsu, Ya-Ling Wei, Meng-Chieh Yu, Wen-Shu Chen, Ke-Wei Hsu, Ming-Hsin Yeh, Liang-Chih Liu, Yun-Ju Chen, and Wei-Chien Huang Copyright © 2014 Chih-Yen Tu et al. All rights reserved. Redox Signaling as a Therapeutic Target to Inhibit Myofibroblast Activation in Degenerative Fibrotic Disease Thu, 20 Feb 2014 10:53:29 +0000 Degenerative fibrotic diseases encompass numerous systemic and organ-specific disorders. Despite their associated significant morbidity and mortality, there is currently no effective antifibrotic treatment. Fibrosis is characterized by the development and persistence of myofibroblasts, whose unregulated deposition of extracellular matrix components disrupts signaling cascades and normal tissue architecture leading to organ failure and death. The profibrotic cytokine transforming growth factor beta (TGFβ) is considered the foremost inducer of fibrosis, driving myofibroblast differentiation in diverse tissues. This review summarizes recent in vitro and in vivo data demonstrating that TGFβ-induced myofibroblast differentiation is driven by a prooxidant shift in redox homeostasis. Elevated NADPH oxidase 4 (NOX4)-derived hydrogen peroxide (H2O2) supported by concomitant decreases in nitric oxide (NO) signaling and reactive oxygen species scavengers are central factors in the molecular pathogenesis of fibrosis in numerous tissues and organs. Moreover, complex interplay between NOX4-derived H2O2 and NO signaling regulates myofibroblast differentiation. Restoring redox homeostasis via antioxidants or NOX4 inactivation as well as by enhancing NO signaling via activation of soluble guanylyl cyclases or inhibition of phosphodiesterases can inhibit and reverse myofibroblast differentiation. Thus, dysregulated redox signaling represents a potential therapeutic target for the treatment of wide variety of different degenerative fibrotic disorders. Natalie Sampson, Peter Berger, and Christoph Zenzmaier Copyright © 2014 Natalie Sampson et al. All rights reserved. Effects of Dietary Cholesterol and Its Oxidation Products on Pathological Lesions and Cholesterol and Lipid Oxidation in the Rabbit Liver Thu, 20 Feb 2014 09:28:10 +0000 This study was conducted to determine the effects of dietary cholesterol (CHO) and cholesterol oxidation products (COPs) on the induction of pathological lesions in rabbit liver tissues. Liver lesions were induced only when the levels of CHO and COPs in the diet were very high. The amount of CHO measured in the liver increased when dietary CHO was increased; by comparison, dietary COPs affected liver CHO amounts to a lesser extent. The TBARS (thiobarbituric acid reactive substances) value measured for the liver samples also increased when dietary CHO and COP levels were elevated, and the TBARS value was more strongly affected by the amount of COPs in the diet than by the amount of CHO. At 6 and 12 weeks, COP levels were the highest in the group that received 1.2 g CHO + 0.8 g COPs, followed by the 0.5 g CHO + 0.5 g COPs and 1.6 g CHO + 0.4 g COPs groups; the control (0 g) group showed the lowest COP levels among all groups. In this study, we found that not only dietary CHO but also COPs were involved in hypercholesterolemia induced liver lesions when the amount of CHO and COPs was high. Sun Jin Hur, Ki Chang Nam, Byungrok Min, Min Du, Kwon Il Seo, and Dong Uk Ahn Copyright © 2014 Sun Jin Hur et al. All rights reserved. Characterization of -Treated Rice Husk Adsorbent and Adsorption of Copper(II) from Aqueous Solution Tue, 11 Feb 2014 09:38:39 +0000 Rice husk, a surplus agricultural byproduct, was applied to the sorption of copper from aqueous solutions. Chemical modifications by treating rice husk with H3PO4 increased the sorption ability of rice husk for Cu(II). This work investigated the sorption characteristics for Cu(II) and examined the optimum conditions of the sorption processes. The elemental compositions of native rice husk and H3PO4-treated rice husk were determined by X-ray fluorescence (XRF) analysis. The scanning electron microscopic (SEM) analysis was carried out for structural and morphological characteristics of H3PO4-treated rice husk. The surface functional groups (i.e., carbonyl, carboxyl, and hydroxyl) of adsorbent were examined by Fourier Transform Infrared Technique (FT-IR) and contributed to the adsorption for Cu(II). Adsorption isotherm experiments were carried out at room temperature and the data obtained from batch studies fitted well with the Langmuir and Freundlich models with of 0.999 and 0.9303, respectively. The maximum sorption amount was 17.0358 mg/g at a dosage of 2 g/L after 180 min. The results showed that optimum pH was attained at pH 4.0. The equilibrium data was well represented by the pseudo-second-order kinetics. The percentage removal for Cu(II) approached equilibrium at 180 min with 88.9% removal. Ying Zhang, Ru Zheng, Jiaying Zhao, Fang Ma, Yingchao Zhang, and Qingjuan Meng Copyright © 2014 Ying Zhang et al. All rights reserved. ROS, Notch, and Wnt Signaling Pathways: Crosstalk between Three Major Regulators of Cardiovascular Biology Tue, 04 Feb 2014 09:41:44 +0000 Reactive oxygen species (ROS), traditionally viewed as toxic by-products that cause damage to biomolecules, now are clearly recognized as key modulators in a variety of biological processes and pathological states. The development and regulation of the cardiovascular system require orchestrated activities; Notch and Wnt/β-catenin signaling pathways are implicated in many aspects of them, including cardiomyocytes and smooth muscle cells survival, angiogenesis, progenitor cells recruitment and differentiation, arteriovenous specification, vascular cell migration, and cardiac remodelling. Several novel findings regarding the role of ROS in Notch and Wnt/β-catenin modulation prompted us to review their emerging function in the cardiovascular system during embryogenesis and postnatally. C. Caliceti, P. Nigro, P. Rizzo, and R. Ferrari Copyright © 2014 C. Caliceti et al. All rights reserved. Evaluation of Buspirone on Streptozotocin Induced Type 1 Diabetes and Its Associated Complications Mon, 20 Jan 2014 11:27:01 +0000 We have evaluated the effect of buspirone (1.5 mg/kg/day, p.o.) type 1 diabetes induced cardiovascular complications induced by streptozotocin (STZ, 45 mg/kg, i.v.) in Wistar rats. Various biochemical, cardiovascular, and hemodynamic parameters were measured at the end of 8 weeks of treatment. STZ produced significant hyperglycaemia, hypoinsulinemia, and dyslipidemia, which was prevented by buspirone treatment. STZ produced increase in serum creatinine, urea, lactate dehydrogenase, creatinine kinase, and C-reactive protein levels and treatment with buspirone produced reduction in these levels. STZ produced increase in cardiac and LV hypertrophy index, LV/RV ratio, and LV collagen, which were decreased by buspirone treatment. Buspirone also prevented STZ induced hemodynamic alterations and oxidative stress. These results were further supported by histopathological studies in which buspirone showed marked reduction in fibrosis and cardiac fiber disarray. In conclusion, our data suggests that buspirone is beneficial as an antidiabetic agent in type 1 diabetes mellitus and also prevents its cardiac complications. Suchi Raghunathan, Pratik Tank, Shraddha Bhadada, and Bhoomika Patel Copyright © 2014 Suchi Raghunathan et al. All rights reserved. PLP-Dependent Enzymes Wed, 15 Jan 2014 09:53:51 +0000 Alessandro Paiardini, Roberto Contestabile, Ashley M. Buckle, and Barbara Cellini Copyright © 2014 Alessandro Paiardini et al. All rights reserved. The CCN Family Proteins: Modulators of Bone Development and Novel Targets in Bone-Associated Tumors Tue, 14 Jan 2014 14:20:27 +0000 The CCN family of proteins is composed of six extracellular matrix-associated proteins that play crucial roles in skeletal development, wound healing, fibrosis, and cancer. Members of the CCN family share four conserved cysteine-rich modular domains that trigger signal transduction in cell adhesion, migration, proliferation, differentiation, and survival through direct binding to specific integrin receptors and heparan sulfate proteoglycans. In the present review, we discuss the roles of the CCN family proteins in regulating resident cells of the bone microenvironment. In vertebrate development, the CCN family plays a critical role in osteo/chondrogenesis and vasculo/angiogenesis. These effects are regulated through signaling via integrins, bone morphogenetic protein, vascular endothelial growth factor, Wnt, and Notch via direct binding to CCN family proteins. Due to the important roles of CCN family proteins in skeletal development, abnormal expression of CCN proteins is related to the tumorigenesis of primary bone tumors such as osteosarcoma, Ewing sarcoma, and chondrosarcoma. Additionally, emerging studies have suggested that CCN proteins may affect progression of secondary metastatic bone tumors by moderating the bone microenvironment. CCN proteins could therefore serve as potential therapeutic targets for drug development against primary and metastatic bone tumors. Po-Chun Chen, Hsu-Chen Cheng, Shun-Fa Yang, Chiao-Wen Lin, and Chih-Hsin Tang Copyright © 2014 Po-Chun Chen et al. All rights reserved. Oxidative Stress and Bone Resorption Interplay as a Possible Trigger for Postmenopausal Osteoporosis Sun, 12 Jan 2014 10:15:40 +0000 The underlying mechanism in postmenopausal osteoporosis (PO) is an imbalance between bone resorption and formation. This study was conducted to investigate whether oxidative stress (OxS) might have a role in this derangement of bone homeostasis. In a sample of 167 postmenopausal women, we found that increased serum levels of a lipid peroxidation marker, hydroperoxides, were negatively and independently associated with decreased bone mineral density (BMD) in total body (, ), lumbar spine (, ), and total hip (, ), as well as with increased bone resorption rate (, ), as assessed by the serum concentration of C-terminal telopeptide of type I collagen (CTX-1). On the contrary, the OxS marker failed to be correlated with the serum levels of bone-specific alkaline phosphatase (BAP), that is, elective marker of bone formation. Importantly, multiple regression analysis revealed that hydroperoxides is a determinant factor for the statistical association between lumbar spine BMD and CTX-1 levels. Taken together, our data suggest that OxS might mediate, by enhancing bone resorption, the uncoupling of bone turnover that underlies PO development. Carlo Cervellati, Gloria Bonaccorsi, Eleonora Cremonini, Arianna Romani, Enrica Fila, Maria Cristina Castaldini, Stefania Ferrazzini, Melchiorre Giganti, and Leo Massari Copyright © 2014 Carlo Cervellati et al. All rights reserved. Systemic Oxidative Stress and Conversion to Dementia of Elderly Patients with Mild Cognitive Impairment Sun, 12 Jan 2014 10:14:06 +0000 Mild cognitive impairment (MCI) is regarded as a prodromal phase of late onset Alzheimer’s disease (LOAD). It has been proposed that oxidative stress (OxS) might be implicated in the pathogenesis of LOAD. The aim of this study was to investigate whether a redox imbalance measured as serum level of hydroperoxides (i.e., by-products of lipid peroxidation) and/or serum antioxidant capacity might be predictive of the clinical progression of MCI to LOAD. The levels of these two markers were measured in 111 patients with MCI (follow-up: years), 105 patients with LOAD, and 118 nondemented healthy controls. Multivariate analysis adjusted for potential confounding factors, including age, gender, smoking, and comorbidities, showed a significant increase () in baseline levels of OxS in MCI and LOAD as compared to cognitive healthy controls. No differences in either of OxS markers were found by comparing MCI patients who converted () or not converted () to LOAD. Overall, these results suggest that systemic OxS might be a precocious feature of MCI and LOAD. However, the role of OxS as an early prognostic marker of progression to LOAD needs further investigations. Carlo Cervellati, Arianna Romani, Davide Seripa, Eleonora Cremonini, Cristina Bosi, Stefania Magon, Carlo M. Bergamini, Giuseppe Valacchi, Alberto Pilotto, and Giovanni Zuliani Copyright © 2014 Carlo Cervellati et al. All rights reserved.