BioMed Research International: Tissue Engineering The latest articles from Hindawi Publishing Corporation © 2014 , Hindawi Publishing Corporation . All rights reserved. The Pilot Study of Fibrin with Temporomandibular Joint Derived Synovial Stem Cells in Repairing TMJ Disc Perforation Tue, 15 Apr 2014 13:12:10 +0000 TMJ disc related diseases are difficult to be cured due to the poor repair ability of the disc. TMJ-SDSCs were ideal cell sources for cartilage tissue engineering which have been widely used in hyaline cartilage regeneration. Fibrin gel has been demonstrated as a potential scaffold for neocartilage formation. The aim of this study was to repair the TMJ disc perforation using fibrin/chitosan hybrid scaffold combined with TMJ-SDSCs. Rat TMJ-SDSCs were cultured on hybrid scaffold or pure chitosan scaffolds. The cell seeding efficiency, distribution, proliferation, and chondrogenic differentiation capacity were investigated. To evaluate the in vivo repair ability of cell/scaffold construct, rat TMJ disc explants were punched with a defect to mimic TMJ disc perforation. Cell seeded scaffolds were inserted into the defect of TMJ disc explants and then were implanted subcutaneously in nude mice for 4 weeks. Results demonstrated that fibrin may improve cell seeding, proliferation, and chondrogenic induction in vitro. The in vivo experiments showed more cartilage ECM deposition in fibrin/chitosan scaffold, which suggested an enhanced reparative ability. This pilot study demonstrated that the regenerative ability of TMJ-SDSCs seeded in fibrin/chitosan scaffold could be applied for repairing TMJ disc perforation. Yang Wu, Zhongcheng Gong, Jian Li, Qinggong Meng, Wei Fang, and Xing Long Copyright © 2014 Yang Wu et al. All rights reserved. Generation of Insulin-Producing Cells from Human Bone Marrow-Derived Mesenchymal Stem Cells: Comparison of Three Differentiation Protocols Thu, 10 Apr 2014 11:14:48 +0000 Introduction. Many protocols were utilized for directed differentiation of mesenchymal stem cells (MSCs) to form insulin-producing cells (IPCs). We compared the relative efficiency of three differentiation protocols. Methods. Human bone marrow-derived MSCs (HBM-MSCs) were obtained from three insulin-dependent type 2 diabetic patients. Differentiation into IPCs was carried out by three protocols: conophylline-based (one-step protocol), trichostatin-A-based (two-step protocol), and β-mercaptoethanol-based (three-step protocol). At the end of differentiation, cells were evaluated by immunolabeling for insulin production, expression of pancreatic endocrine genes, and release of insulin and c-peptide in response to increasing glucose concentrations. Results. By immunolabeling, the proportion of generated IPCs was modest (≃3%) in all the three protocols. All relevant pancreatic endocrine genes, insulin, glucagon, and somatostatin, were expressed. There was a stepwise increase in insulin and c-peptide release in response to glucose challenge, but the released amounts were low when compared with those of pancreatic islets. Conclusion. The yield of functional IPCs following directed differentiation of HBM-MSCs was modest and was comparable among the three tested protocols. Protocols for directed differentiation of MSCs need further optimization in order to be clinically meaningful. To this end, addition of an extracellular matrix and/or a suitable template should be attempted. Mahmoud M. Gabr, Mahmoud M. Zakaria, Ayman F. Refaie, Sherry M. Khater, Sylvia A. Ashamallah, Amani M. Ismail, Nagwa El-Badri, and Mohamed A. Ghoneim Copyright © 2014 Mahmoud M. Gabr et al. All rights reserved. Evaluation of Insulin Medium or Chondrogenic Medium on Proliferation and Chondrogenesis of ATDC5 Cells Thu, 10 Apr 2014 09:09:17 +0000 Background. The ATDC5 cell line is regarded as an excellent cell model for chondrogenesis. In most studies with ATDC5 cells, insulin medium (IM) was used to induce chondrogenesis while chondrogenic medium (CM), which was usually applied in chondrogenesis of mesenchymal stem cells (MSCs), was rarely used for ATDC5 cells. This study was mainly designed to investigate the effect of IM, CM, and growth medium (GM) on chondrogenesis of ATDC5 cells. Methods. ATDC5 cells were, respectively, cultured in IM, CM, and GM for a certain time. Then the proliferation and the chondrogenesis progress of cells in these groups were analyzed. Results. Compared with CM and GM, IM promoted the proliferation of cells significantly. CM was effective for enhancement of cartilage specific markers, while IM induced the cells to express endochondral ossification related genes. Although GAG deposition per cell in CM group was significantly higher than that in IM and GM groups, the total GAG contents in IM group were the most. Conclusion. This study demonstrated that CM focused on induction of chondrogenic differentiation while IM was in favor of promoting proliferation and expression of endochondral ossification related genes. Combinational use of these two media would be more beneficial to bone/cartilage repair. Yongchang Yao, Zhichen Zhai, and Yingjun Wang Copyright © 2014 Yongchang Yao et al. All rights reserved. Enhanced Osteogenicity of Bioactive Composites with Biomimetic Treatment Wed, 09 Apr 2014 07:12:59 +0000 Purpose. This study aimed to explore if initiation of biomimetic apatite nucleation can be used to enhance osteoblast response to biodegradable tissue regeneration composite membranes. Materials and Methods. Bioactive thermoplastic composites consisting of poly(ε-caprolactone/DL-lactide) and bioactive glass (BAG) were prepared at different stages of biomimetic calcium phosphate deposition by immersion in simulated body fluid (SBF). The modulation of the BAG dissolution and the osteogenic response of rat mesenchymal stem cells (MSCs) were analyzed. Results. SBF treatment resulted in a gradual calcium phosphate deposition on the composites and decreased BAG reactivity in the subsequent cell cultures. Untreated composites and composites covered by thick calcium phosphate layer (14 days in SBF) expedited MSC mineralization in comparison to neat polymers without BAG, whereas other osteogenic markers—alkaline phosphatase activity, bone sialoprotein, and osteocalcin expression—were initially decreased. In contrast, surfaces with only small calcium phosphate aggregates (five days in SBF) had similar early response than neat polymers but still demonstrated enhanced mineralization. Conclusion. A short biomimetic treatment enhances osteoblast response to bioactive composite membranes. Ville V. Meretoja, Teemu Tirri, Minna Malin, Jukka V. Seppälä, and Timo O. Närhi Copyright © 2014 Ville V. Meretoja et al. All rights reserved. Effect of Cyclic Mechanical Stimulation on the Expression of Osteogenesis Genes in Human Intraoral Mesenchymal Stromal and Progenitor Cells Mon, 07 Apr 2014 07:12:18 +0000 We evaluated the effects of mechanical stimulation on the osteogenic differentiation of human intraoral mesenchymal stem and progenitor cells (MSPCs) using the Flexcell FX5K Tension System that mediated cyclic tensile stretch on the cells. MSPCs were isolated from human mandibular retromolar bones and characterized using flow cytometry. The positive expression of CD73, CD90, and CD105 and negativity for CD14, CD19, CD34, CD45, and HLA-DR confirmed the MSPC phenotype. Mean MSPC doubling time was  hrs. The percentage of lactate dehydrogenase (LDH) release showed no significant difference between the mechanically stimulated groups and the unstimulated controls. Reverse transcription quantitative real-time PCR revealed that 10% continuous cyclic strain (0.5 Hz) for 7 and 14 days induced a significant increase in the mRNA expression of the osteogenesis-specific markers type-I collagen (Col1A1), osteonectin (SPARC), bone morphogenetic protein 2 (BMP2), osteopontin (SPP1), and osteocalcin (BGLAP) in osteogenic differentiated MSPCs. Furthermore, mechanically stimulated groups produced significantly higher amounts of calcium deposited into the cultures and alkaline phosphatase (ALP). These results will contribute to a better understanding of strain-induced bone remodelling and will form the basis for the correct choice of applied force in oral and maxillofacial surgery. Birgit Lohberger, Heike Kaltenegger, Nicole Stuendl, Michael Payer, Beate Rinner, and Andreas Leithner Copyright © 2014 Birgit Lohberger et al. All rights reserved. Current Trends in Bone Tissue Engineering Sun, 06 Apr 2014 12:21:10 +0000 The development of tissue engineering and regeneration constitutes a new platform for translational medical research. Effective therapies for bone engineering typically employ the coordinated manipulation of cells, biologically active signaling molecules, and biomimetic, biodegradable scaffolds. Bone tissue engineering has become increasingly dependent on the merging of innovations from each of these fields, as they continue to evolve independently. This foreword will highlight some of the most recent advances in bone tissue engineering and regeneration, emphasizing the interconnected fields of stem cell biology, cell signaling biology, and biomaterial research. These include, for example, novel methods for mesenchymal stem cell purification, new methods of Wnt signaling pathway manipulation, and cutting edge computer assisted nanoscale design of bone scaffold materials. In the following special issue, we sought to incorporate these diverse areas of emphasis in order to reflect current trends in the field. Marco Mravic, Bruno Péault, and Aaron W. James Copyright © 2014 Marco Mravic et al. All rights reserved. Expressions of ABCG2, CD133, and Podoplanin in Salivary Adenoid Cystic Carcinoma Sun, 06 Apr 2014 11:28:47 +0000 Adenoid cystic carcinoma (ACC) is one of the most common salivary gland malignant tumors with a high risk of recurrence and metastasis. Current studies on cancer stem cells (CSCs) have verified that CSCs are the driving force behind tumor initiation and progression, suggesting that new cancer therapies may be established by effectively targeting and killing the CSCs. The primary goal of this study is to investigate the expression patterns of ABCG2, CD133, and podoplanin in ACC of minor salivary glands by immunohistochemistry analysis. We found that ABCG2 was weakly expressed in normal looking salivary gland tissues. A significant upregulation of ABCG2 expression in ACC was observed with a similar expression pattern of Ki-67. CD133 was detected in apical membrane of epithelial cells and podoplanin was expressed positively in myoepithelial cells of both normal looking tissue and ACC. However, no significant difference was found of the expression pattern of CD133 and podoplanin between normal looking tissues and ACC. Our observations suggest that CSCs may exist in quiescent cells with ABCG2 positive staining, which are surrounded by cells with positive expression of ABCG2 and Ki-67 in ACC, and costaining with ABCG2 and Ki-67 may help predict the location of CSCs. Wuwei Li, Ryo Tamamura, Bo Wang, Qigui Liu, Han Liu, Tingjiao Liu, Naoki Katase, Jing Xiao, and Hitoshi Nagatsuka Copyright © 2014 Wuwei Li et al. All rights reserved. Deregulation of Bone Forming Cells in Bone Diseases and Anabolic Effects of Strontium-Containing Agents and Biomaterials Mon, 31 Mar 2014 15:17:17 +0000 Age-related bone loss and osteoporosis are associated with bone remodeling changes that are featured with decreased trabecular and periosteal bone formation relative to bone resorption. Current anticatabolic therapies focusing on the inhibition of bone resorption may not be sufficient in the prevention or reversal of age-related bone deterioration and there is a big need in promoting osteoblastogenesis and bone formation. Enhanced understanding of the network formed by key signaling pathways and molecules regulating bone forming cells in health and diseases has therefore become highly significant. The successful development of agonist/antagonist of the PTH and Wnt signaling pathways are profits of the understanding of these key pathways. As the core component of an approved antiosteoporosis agent, strontium takes its effect on osteoblasts at multilevel through multiple pathways, representing a good example in revealing and exploring anabolic mechanisms. The recognition of strontium effects on bone has led to its expected application in a variety of biomaterial scaffolds used in tissue engineering strategies aiming at bone repairing and regeneration. While summarizing the recent progress in these respects, this review also proposes the new approaches such as systems biology in order to reveal new insights in the pathology of osteoporosis as well as possible discovery of new therapies. Shuang Tan, Binbin Zhang, Xiaomei Zhu, Ping Ao, Huajie Guo, Weihong Yi, and Guang-Qian Zhou Copyright © 2014 Shuang Tan et al. All rights reserved. Characterization of Silk Fibroin Modified Surface: A Proteomic View of Cellular Response Proteins Induced by Biomaterials Tue, 25 Mar 2014 13:30:48 +0000 The purpose of this study was to develop the pathway of silk fibroin (SF) biopolymer surface induced cell membrane protein activation. Fibroblasts were used as an experimental model to evaluate the responses of cellular proteins induced by biopolymer material using a mass spectrometry-based profiling system. The surface was covered by multiwalled carbon nanotubes (CNTs) and SF to increase the surface area, enhance the adhesion of biopolymer, and promote the rate of cell proliferation. The amount of adhered fibroblasts on CNTs/SF electrodes of quartz crystal microbalance (QCM) greatly exceeded those on other surfaces. Moreover, analyzing differential protein expressions of adhered fibroblasts on the biopolymer surface by proteomic approaches indicated that CD44 may be a key protein. Through this study, utilization of mass spectrometry-based proteomics in evaluation of cell adhesion on biopolymer was proposed. Ming-Hui Yang, Shyng-Shiou Yuan, Tze-Wen Chung, Shiang-Bin Jong, Chi-Yu Lu, Wan-Chi Tsai, Wen-Cheng Chen, Po-Chiao Lin, Pei-Wen Chiang, and Yu-Chang Tyan Copyright © 2014 Ming-Hui Yang et al. All rights reserved. Adipose-Derived Mesenchymal Stem Cells from Ventral Hernia Repair Patients Demonstrate Decreased Vasculogenesis Mon, 17 Mar 2014 08:40:41 +0000 Introduction. In adipose tissue healing, angiogenesis is stimulated by adipose-derived stromal stem cells (ASCs). Ventral hernia repair (VHR) patients are at high risk for wound infections. We hypothesize that ASCs from VHR patients are less vasculogenic than ASCs from healthy controls. Methods. ASCs were harvested from the subcutaneous fat of patients undergoing VHR by the component separation technique and from matched abdominoplasty patients. RNA and protein were harvested on culture days 0 and 3. Both groups of ASCs were subjected to hypoxic conditions for 12 and 24 hours. RNA was analyzed using qRT-PCR, and protein was used for western blotting. ASCs were also grown in Matrigel under hypoxic conditions and assayed for tubule formation after 24 hours. Results. Hernia patient ASCs demonstrated decreased levels of VEGF-A protein and vasculogenic RNA at 3 days of growth in differentiation media. There were also decreases in VEGF-A protein and vasculogenic RNA after growth in hypoxic conditions compared to control ASCs. After 24 hours in hypoxia, VHR ASCs formed fewer tubules in Matrigel than in control patient ASCs. Conclusion. ASCs derived from VHR patients appear to express fewer vasculogenic markers and form fewer tubules in Matrigel than ASCs from abdominoplasty patients, suggesting decreased vasculogenic activity. Jeffrey Lisiecki, Jacob Rinkinen, Oluwatobi Eboda, Jonathan Peterson, Sara De La Rosa, Shailesh Agarwal, Justin Dimick, Oliver A. Varban, Paul S. Cederna, Stewart C. Wang, and Benjamin Levi Copyright © 2014 Jeffrey Lisiecki et al. All rights reserved. A Standardized Critical Size Defect Model in Normal and Osteoporotic Rats to Evaluate Bone Tissue Engineered Constructs Tue, 11 Mar 2014 09:15:33 +0000 Tissue engineered constructs should be tested for their efficacy not only in normal but also in osteoporotic bone. The rat is an established animal model for osteoporosis and is used often for bone healing studies. In this study a defined and standardized critical size defect model in the rat suitable for screening new tissue engineered constructs in normal and osteoporotic bone is described and validated. Normal and ovariectomised Wistar rats received a unilateral middiaphyseal 5 mm defect in the femur, which was instrumented with a radiolucent PEEK plate fixed with angular stable titanium screws and left untreated. All animals were euthanized eight weeks after defect surgery and the bone healing was evaluated using radiographs, computed tomography measurements, and histology. The developed fixation system provided good stability, even in osteoporotic bone. The implants and ancillary instruments ensured consistent and facile placement of the PEEK plates. The untreated defects did not heal without intervention making the model a well-defined and standardized critical size defect model highly useful for evaluating tissue engineered solutions in normal and osteoporotic bone. Livia Poser, Romano Matthys, Peter Schawalder, Simon Pearce, Mauro Alini, and Stephan Zeiter Copyright © 2014 Livia Poser et al. All rights reserved. Nanosized Mesoporous Bioactive Glass/Poly(lactic-co-glycolic Acid) Composite-Coated CaSiO3 Scaffolds with Multifunctional Properties for Bone Tissue Engineering Sun, 02 Mar 2014 12:09:56 +0000 It is of great importance to prepare multifunctional scaffolds combining good mechanical strength, bioactivity, and drug delivery ability for bone tissue engineering. In this study, nanosized mesoporous bioglass/poly(lactic-co-glycolic acid) composite-coated calcium silicate scaffolds, named NMBG-PLGA/CS, were successfully prepared. The morphology and structure of the prepared scaffolds were characterized by scanning electron microscopy and X-ray diffraction. The effects of NMBG on the apatite mineralization activity and mechanical strength of the scaffolds and the attachment, proliferation, and alkaline phosphatase activity of MC3T3 cells as well as drug ibuprofen delivery properties were systematically studied. Compared to pure CS scaffolds and PLGA/CS scaffolds, the prepared NMBG-PLGA/CS scaffolds had greatly improved apatite mineralization activity in simulated body fluids, much higher mechanical property, and supported the attachment of MC3T3 cells and enhanced the cell proliferation and ALP activity. Furthermore, the prepared NMBG-PLGA/CS scaffolds could be used for delivering ibuprofen with a sustained release profile. Our study suggests that the prepared NMBG-PLGA/CS scaffolds have improved physicochemical, biological, and drug-delivery property as compared to conventional CS scaffolds, indicating that the multifunctional property of the prepared scaffolds for the potential application of bone tissue engineering. Mengchao Shi, Dong Zhai, Lang Zhao, Chengtie Wu, and Jiang Chang Copyright © 2014 Mengchao Shi et al. All rights reserved. RANKL Expression in Periodontal Disease: Where Does RANKL Come from? Thu, 27 Feb 2014 07:02:34 +0000 Periodontitis is an inflammatory disease characterized by periodontal pocket formation and alveolar bone resorption. Periodontal bone resorption is induced by osteoclasts and receptor activator of nuclear factor-κB ligand (RANKL) which is an essential and central regulator of osteoclast development and osteoclast function. Therefore, RANKL plays a critical role in periodontal bone resorption. In this review, we have summarized the sources of RANKL in periodontal disease and explored which factors may regulate RANKL expression in this disease. Bin Chen, Wenlei Wu, Weibin Sun, Qian Zhang, Fuhua Yan, and Yin Xiao Copyright © 2014 Bin Chen et al. All rights reserved. Calcium Phosphate Based Three-Dimensional Cold Plotted Bone Scaffolds for Critical Size Bone Defects Wed, 26 Feb 2014 13:36:35 +0000 Bone substitutes, like calcium phosphate, are implemented more frequently in orthopaedic surgery to reconstruct critical size defects, since autograft often results in donor site morbidity and allograft can transmit diseases. A novel bone cement, based on β-tricalcium phosphate, polyethylene glycol, and trisodium citrate, was developed to allow the rapid manufacturing of scaffolds, by extrusion freeform fabrication, at room temperature. The cement composition exhibits good resorption properties and serves as a basis for customised (e.g., drug or growth factor loaded) scaffolds for critical size bone defects. In vitro toxicity tests confirmed proliferation and differentiation of ATDC5 cells in scaffold-conditioned culture medium. Implantation of scaffolds in the iliac wing of sheep showed bone remodelling throughout the defects, outperforming the empty defects on both mineral volume and density present in the defect after 12 weeks. Both scaffolds outperformed the autograft filled defects on mineral density, while the mineral volume present in the scaffold treated defects was at least equal to the mineral volume present in the autograft treated defects. We conclude that the formulated bone cement composition is suitable for scaffold production at room temperature and that the established scaffold material can serve as a basis for future bone substitutes to enhance de novo bone formation in critical size defects. Christian J. D. Bergmann, Jim C. E. Odekerken, Tim J. M. Welting, Franz Jungwirth, Declan Devine, Ludovic Bouré, Stephan Zeiter, Lodewijk W. van Rhijn, Rainer Telle, Horst Fischer, and Pieter J. Emans Copyright © 2014 Christian J. D. Bergmann et al. All rights reserved. Comparing Microspheres with Different Internal Phase of Polyelectrolyte as Local Drug Delivery System for Bone Tuberculosis Therapy Sun, 23 Feb 2014 00:00:00 +0000 We use hydrophobic poly(lactic-co-glycolic) acid (PLGA) to encapsulate hydrophilic ofloxacin to form drug loading microspheres. Hyaluronic acid (HA) and polylysine (Pls) were used as internal phase additives to see their influences on the drug loading and releasing. Double emulsion (water-in-oil-in-water) solvent extraction/evaporation method was used for the purpose. Particle size analysis display that the polyelectrolytes have low impact on the microsphere average size and distribution. Scanning electron microscope (SEM) pictures show the wrinkled surface resulted by the internal microcavity of the microspheres. Microspheres with HA inside have higher drug loading amounts than microspheres with Pls inside. The loading drug amounts of the microspheres increase with the HA amounts inside, while decreasing with the Pls amounts inside. All the polyelectrolytes adding groups have burst release observed in experiments. The microspheres with Pls internal phase have faster release rate than the HA groups. Among the same polyelectrolyte internal phase groups, the release rate increases with the amounts increasing when Pls is inside, while it decreases with the amounts increasing when HA is inside. Gang Wu, Long Chen, Hong Li, Chun-Ling Deng, and Xiao-Feng Chen Copyright © 2014 Gang Wu et al. All rights reserved. The Effect of Hypoxia on the Stemness and Differentiation Capacity of PDLC and DPC Thu, 20 Feb 2014 09:35:14 +0000 Introduction. Stem cells are regularly cultured under normoxic conditions. However, the physiological oxygen tension in the stem cell niche is known to be as low as 1-2% oxygen, suggesting that hypoxia has a distinct impact on stem cell maintenance. Periodontal ligament cells (PDLCs) and dental pulp cells (DPCs) are attractive candidates in dental tissue regeneration. It is of great interest to know whether hypoxia plays a role in maintaining the stemness and differentiation capacity of PDLCs and DPCs. Methods. PDLCs and DPCs were cultured either in normoxia (20% O2) or hypoxia (2% O2). Cell viability assays were performed and the expressions of pluripotency markers (Oct-4, Sox2, and c-Myc) were detected by qRT-PCR and western blotting. Mineralization, glycosaminoglycan (GAG) deposition, and lipid droplets formation were assessed by Alizarin red S, Safranin O, and Oil red O staining, respectively. Results. Hypoxia did not show negative effects on the proliferation of PDLCs and DPCs. The pluripotency markers and differentiation potentials of PDLCs and DPCs significantly increased in response to hypoxic environment. Conclusions. Our findings suggest that hypoxia plays an important role in maintaining the stemness and differentiation capacity of PDLCs and DPCs. Yinghong Zhou, Wei Fan, and Yin Xiao Copyright © 2014 Yinghong Zhou et al. All rights reserved. Characterization of an Ovine Bilateral Critical Sized Bone Defect Iliac Wing Model to Examine Treatment Modalities Based on Bone Tissue Engineering Sun, 16 Feb 2014 12:35:23 +0000 Critical sized bone defect (CSBD) animal models are used to evaluate and confirm efficacy and potency of new treatment modalities based on bone tissue engineering before the latter can be applied in clinical practice. In this study, a bilateral CSBD model in the iliac wings of sheep is described in detail. To demonstrate that this is a large animal CSBD model in sheep, bone healing within the defect left empty (negative control) or filled with autologous corticocancellous bone graft (clinical gold standard, positive control) was assessed using micro-CT, histology, histomorphometric, and fluorochrome analysis. After three months, new bone into the defect site was formed across the whole defect in the positive controls but limited to the edge of the defects in the negative controls. Bone volume in the positive controls was statistically higher than in the negative controls, with the latter having less than 10% new bone growth. There were no intraoperative or postoperative complications. The model described here represents a reliable and reproducible bilateral CSBD in sheep with low morbidity that can be used for in vivo evaluation of new treatment modalities based on bone tissue engineering. Jennifer L. Lansdowne, Declan Devine, Ursula Eberli, Pieter Emans, Tim J. M. Welting, Jim C. E. Odekerken, Damiano Schiuma, Martin Thalhauser, Ludovic Bouré, and Stephan Zeiter Copyright © 2014 Jennifer L. Lansdowne et al. All rights reserved. Interleukin-10 Inhibits Bone Resorption: A Potential Therapeutic Strategy in Periodontitis and Other Bone Loss Diseases Sun, 16 Feb 2014 12:05:40 +0000 Periodontitis and other bone loss diseases, decreasing bone volume and strength, have a significant impact on millions of people with the risk of tooth loss and bone fracture. The integrity and strength of bone are maintained through the balance between bone resorption and bone formation by osteoclasts and osteoblasts, respectively, so the loss of bone results from the disruption of such balance due to increased resorption or/and decreased formation of bone. The goal of therapies for diseases of bone loss is to reduce bone loss, improve bone formation, and then keep healthy bone density. Current therapies have mostly relied on long-term medication, exercise, anti-inflammatory therapies, and changing of the life style. However there are some limitations for some patients in the effective treatments for bone loss diseases because of the complexity of bone loss. Interleukin-10 (IL-10) is a potent anti-inflammatory cytokine, and recent studies have indicated that IL-10 can contribute to the maintenance of bone mass through inhibition of osteoclastic bone resorption and regulation of osteoblastic bone formation. This paper will provide a brief overview of the role of IL-10 in bone loss diseases and discuss the possibility of IL-10 adoption in therapy of bone loss diseases therapy. Qian Zhang, Bin Chen, Fuhua Yan, Jianbin Guo, Xiaofeng Zhu, Shouzhi Ma, and Wenrong Yang Copyright © 2014 Qian Zhang et al. All rights reserved. Association between Postmenopausal Osteoporosis and Experimental Periodontitis Mon, 10 Feb 2014 13:02:10 +0000 To investigate the correlation between postmenopausal osteoporosis (PMO) and the pathogenesis of periodontitis, ovariectomized rats were generated and the experimental periodontitis was induced using a silk ligature. The inflammatory factors and bone metabolic markers were measured in the serum and periodontal tissues of ovariectomized rats using an automatic chemistry analyzer, enzyme-linked immunosorbent assays, and immunohistochemistry. The bone mineral density of whole body, pelvis, and spine was analyzed using dual-energy X-ray absorptiometry and image analysis. All data were analyzed using SPSS 13.0 statistical software. It was found that ovariectomy could upregulate the expression of interleukin- (IL-)6, the receptor activator of nuclear factor-κB ligand (RANKL), and osteoprotegerin (OPG) and downregulate IL-10 expression in periodontal tissues, which resulted in progressive alveolar bone loss in experimental periodontitis. This study indicates that changes of cytokines and bone turnover markers in the periodontal tissues of ovariectomized rats contribute to the damage of periodontal tissues. Kai Luo, Souzhi Ma, Jianbin Guo, Yongling Huang, Fuhua Yan, and Yin Xiao Copyright © 2014 Kai Luo et al. All rights reserved. Regenerative Repair of Damaged Meniscus with Autologous Adipose Tissue-Derived Stem Cells Thu, 30 Jan 2014 06:35:35 +0000 Mesenchymal stem cells (MSCs) are defined as pluripotent cells found in numerous human tissues, including bone marrow and adipose tissue. Such MSCs, isolated from bone marrow and adipose tissue, have been shown to differentiate into bone and cartilage, along with other types of tissues. Therefore, MSCs represent a promising new therapy in regenerative medicine. The initial treatment of meniscus tear of the knee is managed conservatively with nonsteroidal anti-inflammatory drugs and physical therapy. When such conservative treatment fails, an arthroscopic resection of the meniscus is necessary. However, the major drawback of the meniscectomy is an early onset of osteoarthritis. Therefore, an effective and noninvasive treatment for patients with continuous knee pain due to damaged meniscus has been sought. Here, we present a review, highlighting the possible regenerative mechanisms of damaged meniscus with MSCs (especially adipose tissue-derived stem cells (ASCs)), along with a case of successful repair of torn meniscus with significant reduction of knee pain by percutaneous injection of autologous ASCs into an adult human knee. Jaewoo Pak, Jung Hun Lee, and Sang Hee Lee Copyright © 2014 Jaewoo Pak et al. All rights reserved. rhPDGF-BB Promotes Proliferation and Osteogenic Differentiation of Bone Marrow Stromal Cells from Streptozotocin-Induced Diabetic Rats through ERK Pathway Wed, 29 Jan 2014 00:00:00 +0000 Management of nonunion fracture and massive segmental bone defects in diabetes remains a challenging clinical problem. Bone marrow stromal cells (BMSCs) are crucial for bone remodeling and hold promise for bone regeneration. However, we have showed previously that diabetes can affect the proliferation and osteogenic potential of BMSCs adversely and a strategy to attenuate the impaired functions of BMSCs is required. Platelet-derived growth factor-BB (PDGF-BB) plays an important role in bone formation. However, little information is available about its effect on diabetic BMSCs. In this study, BMSCs were isolated from streptozotocin-induced diabetic rats. After treatment with recombinant human PDGF-BB (rhPDGF-BB), diabetic BMSCs demonstrated enhanced cell proliferation and osteogenic differentiation based on increased expressions of osteogenic genes (Runx2, alkaline phosphatase, and osteocalcin) and Runx2 protein, as well as upregulated alkaline phosphatase activity and mineralization. Furthermore, blocking extracellular signal regulated kinase (ERK) pathway by inhibitor PD98059 repressed the enhanced proliferation and osteogenic differentiation in diabetic BMSCs induced by rhPDGF-BB. Together, these results indicated that rhPDGF-BB stimulates proliferation and osteogenic differentiation partially through ERK pathway in diabetic BMSCs. Therefore, modulation of diabetic BMSCs could augment BMSCs function affected by diabetes and holds significance for future strategies to treat diabetic bone complications. Yanfang Zhao, Songmei Zhang, Deliang Zeng, Lunguo Xia, Ashwini Lamichhane, Xinquan Jiang, and Fuqiang Zhang Copyright © 2014 Yanfang Zhao et al. All rights reserved. Development of a Stereotaxic Device for Low Impact Implantation of Neural Constructs or Pieces of Neural Tissues into the Mammalian Brain Thu, 23 Jan 2014 07:02:33 +0000 Implanting pieces of tissue or scaffolding material into the mammalian central nervous system (CNS) is wrought with difficulties surrounding the size of tools needed to conduct such implants and the ability to maintain the orientation and integrity of the constructs during and after their transplantation. Here, novel technology has been developed that allows for the implantation of neural constructs or intact pieces of neural tissue into the CNS with low trauma. By “laying out” (instead of forcibly expelling) the implantable material from a thin walled glass capillary, this technology has the potential to enhance neural transplantation procedures by reducing trauma to the host brain during implantation and allowing for the implantation of engineered/dissected tissues or constructs in such a way that their orientation and integrity are maintained in the host. Such technology may be useful for treating various CNS disorders which require the reestablishment of point-to-point contacts (e.g., Parkinson’s disease) across the adult CNS, an environment which is not normally permissive to axonal growth. Andrzej Jozwiak, Yiwen Liu, Ying Yang, and Monte A. Gates Copyright © 2014 Andrzej Jozwiak et al. All rights reserved. PHBV/PAM Scaffolds with Local Oriented Structure through UV Polymerization for Tissue Engineering Wed, 22 Jan 2014 11:08:38 +0000 Locally oriented tissue engineering scaffolds can provoke cellular orientation and direct cell spread and migration, offering an exciting potential way for the regeneration of the complex tissue. Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) scaffolds with locally oriented hydrophilic polyacrylamide (PAM) inside the macropores of the scaffolds were achieved through UV graft polymerization. The interpenetrating PAM chains enabled good interconnectivity of PHBV/PAM scaffolds that presented a lower porosity and minor diameter of pores than PHBV scaffolds. The pores with diameter below 100 μm increased to 82.15% of PHBV/PAM scaffolds compared with 31.5% of PHBV scaffolds. PHBV/PAM scaffold showed a much higher compressive elastic modulus than PHBV scaffold due to PAM stuffing. At 5 days of culturing, sheep chondrocytes spread along the similar direction in the macropores of PHBV/PAM scaffolds. The locally oriented PAM chains might guide the attachment and spreading of chondrocytes and direct the formation of microfilaments via contact guidance. Yu Ke, Gang Wu, and Yingjun Wang Copyright © 2014 Yu Ke et al. All rights reserved. Chemical Synthesis, Characterisation, and Biocompatibility of Nanometre Scale Porous Anodic Aluminium Oxide Membranes for Use as a Cell Culture Substrate for the Vero Cell Line: A Preliminary Study Tue, 21 Jan 2014 08:04:31 +0000 In this preliminary study we investigate for the first time the biomedical potential of using porous anodic aluminium oxide (AAO) membranes as a cell substrate for culturing the Cercopithecus aethiops (African green monkey) Kidney (Vero) epithelial cell line. One advantage of using the inorganic AAO membrane is the presence of nanometre scale pore channels that allow the exchange of molecules and nutrients across the membrane. The size of the pore channels can be preselected by adjusting the controlling parameters of a temperature controlled two-step anodization process. The cellular interaction and response of the Vero cell line with an in-house synthesised AAO membrane, a commercially available membrane, and a glass control were assessed by investigating cell adhesion, morphology, and proliferation over a 72 h period. The number of viable cells proliferating over the respective membrane surfaces revealed that the locally produced in-house AAO membrane had cells numbers similar to the glass control. The study revealed evidence of focal adhesion sites over the surface of the nanoporous membranes and the penetration of cellular extensions into the pore structure as well. The outcome of the study has revealed that nanometre scale porous AAO membranes have the potential to become practical cell culture scaffold substrates with the capability to enhance adhesion and proliferation of Vero cells. Gérrard Eddy Jai Poinern, Xuan Thi Le, Mark O'Dea, Thomas Becker, and Derek Fawcett Copyright © 2014 Gérrard Eddy Jai Poinern et al. All rights reserved. A Novel Platelet Concentrate: Titanium-Prepared Platelet-Rich Fibrin Tue, 21 Jan 2014 00:00:00 +0000 We developed a new product called titanium-prepared platelet-rich fibrin (T-PRF). The T-PRF method is based on the hypothesis that titanium may be more effective in activating platelets than the silica activators used with glass tubes in Chouckroun’s leukocyte- and platelet-rich fibrin (L-PRF) method. In this study, we aimed to define the structural characteristics of T-PRF and compare it with L-PRF. Blood samples were collected from 10 healthy male volunteers. The blood samples were drawn using a syringe. Nine milliliters was transferred to a dry glass tube, and 9 mL was transferred to a titanium tube. Half of each clot (i.e., the blood that was clotted using T-PRF or L-PRF) was processed with a scanning electron microscope (SEM). The other half of each clot was processed for fluorescence microscopy analysis and light microscopy analysis. The T-PRF samples seemed to have a highly organized network with continuous integrity compared to the other L-PRF samples. Histomorphometric analysis showed that T-PRF fibrin network covers larger area than L-PRF fibrin network; also fibrin seemed thicker in the T-PRF samples. This is the first human study to define T-PRF as an autogenous leukocyte- and platelet-rich fibrin product. The platelet activation by titanium seems to offer some high characteristics to T-PRF. Mustafa Tunalı, Hakan Özdemir, Zafer Küçükodacı, Serhan Akman, Emre Yaprak, Hülya Toker, and Erhan Fıratlı Copyright © 2014 Mustafa Tunalı et al. All rights reserved. Synergistic Effects of Orbital Shear Stress on In Vitro Growth and Osteogenic Differentiation of Human Alveolar Bone-Derived Mesenchymal Stem Cells Tue, 14 Jan 2014 11:17:00 +0000 Cellular behavior is dependent on a variety of physical cues required for normal tissue function. In order to mimic native tissue environments, human alveolar bone-derived mesenchymal stem cells (hABMSCs) were exposed to orbital shear stress (OSS) in a low-speed orbital shaker. The synergistic effects of OSS on proliferation and differentiation of hABMSCs were investigated. In particular, we induced the osteoblastic differentiation of hABMSCs cultured in the absence of OM by exposing hABMSCs to OSS (0.86–1.51 dyne/cm2). Activation of Cx43 was associated with exposure of hABMSCs to OSS. The viability of cells stimulated for 10, 30, 60, 120, and 180 min/day increased by approximately 10% compared with that of control. The OSS groups with stimulation of 10, 30, and 60 min/day had more intense mineralized nodules compared with the control group. In quantification of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) protein, VEGF protein levels under stimulation for 10, 60, and 180 min/day and BMP-2 levels under stimulation for 60, 120, and 180 min/day were significantly different compared with those of the control. In conclusion, the results indicated that exposing hABMSCs to OSS enhanced their differentiation and maturation. Ki Taek Lim, Jin Hexiu, Jangho Kim, Hoon Seonwoo, Pill-Hoon Choung, and Jong Hoon Chung Copyright © 2014 Ki Taek Lim et al. All rights reserved. Erratum to “Detergent-Enzymatic Decellularization of Swine Blood Vessels: Insight on Mechanical Properties for Vascular Tissue Engineering” Sun, 05 Jan 2014 14:14:16 +0000 Alessandro F. Pellegata, M. Adelaide Asnaghi, Ilaria Stefani, Anna Maestroni, Silvia Maestroni, Tommaso Dominioni, Sandro Zonta, Gianpaolo Zerbini, and Sara Mantero Copyright © 2014 Alessandro F. Pellegata et al. All rights reserved. αVβ5 and CD44 Are Oxygen-Regulated Human Embryonic Stem Cell Attachment Factors Thu, 26 Dec 2013 12:38:40 +0000 Human embryonic stem cells (hESCs) have great potential for clinical therapeutic use. However, relatively little is known of the mechanisms which dictate their specificity of adhesion to substrates through adhesion proteins including integrins. Previous observations demonstrated enhanced clonogenicity in reduced oxygen culture systems. Here, we demonstrated via antibody blocking experiments that αVβ5 and α6 significantly promoted hESC attachment in 2% O2 only, whereas blockage of CD44 inhibited cell attachment in 21% O2 alone. Immunofluorescence confirmed expression of αVβ5 and CD44 in both 2% O2 and 21% O2 cultured hESCs while flow cytometry revealed significantly higher αVβ5 expression in 2% O2 versus 21% O2 cultured hESCs and higher CD44 expression in 21% O2 versus 2% O2 cultured hESCs. Adhered hESCs following blockage of αVβ5 in 2% O2 displayed a reduction in nuclear colocalisation of Oct-4 and Nanog with little effect observed in 21% O2. Blockage of CD44 had the converse effect with dramatic reductions in nuclear colocalisation of Oct-4 and Nanog in 21% O2 cultured hESC which retained adherence, but not in 2% O2 cultured cells. Identification of oxygen-dependent substrate attachment mechanisms in hESCs has the potential to play a role in the development of novel substrates to improve hESC attachment and culture. Deepak Kumar, Saniya Gupta, Ying Yang, and Nicholas R. Forsyth Copyright © 2013 Deepak Kumar et al. All rights reserved. Cellular Performance Comparison of Biomimetic Calcium Phosphate Coating and Alkaline-Treated Titanium Surface Tue, 24 Dec 2013 08:55:32 +0000 The influence of biomimetic calcium phosphate coating on osteoblasts behavior in vitro is not well established yet. In this study, we investigated the behavior of osteoblastic rat osteosarcoma 17/2.8 cells (ROS17/2.8) on two groups of biomaterial surfaces: alkaline-treated titanium surface (ATT) and biomimetic calcium phosphate coated ATT (CaP). The cell attachment, proliferation, differentiation, and morphology on these surfaces were extensively evaluated to reveal the impact of substrate surface on osteoblastic cell responses. It was found that the ROS17/2.8 cells cultured on the ATT surface had higher attachment and proliferation rates compared to those on the CaP surface. Our results also showed that the calcium phosphate coatings generated in this work have an inhibiting effect on osteoblast adhesion and further influenced the proliferation and differentiation of osteoblast compared to the ATT surface in vitro. Cells on the ATT surface also exhibited a higher alkaline phosphatase activity than on the CaP surface after two weeks of culture. Immunofluorescence staining and scanning electron microscopy results showed that the cells adhered and spread faster on the ATT surface than on the CaP surface. These results collectively suggested that substrate surface properties directly influence cell adhesion on different biomaterials, which would result in further influence on the cell proliferation and differentiation. Xiaohua Yu and Mei Wei Copyright © 2013 Xiaohua Yu and Mei Wei. All rights reserved. Enhancing the Migration Ability of Mesenchymal Stromal Cells by Targeting the SDF-1/CXCR4 Axis Thu, 05 Dec 2013 17:44:51 +0000 Mesenchymal stromal cells (MSCs) are currently being investigated in numerous clinical trials of tissue repair and various immunological disorders based on their ability to secrete trophic factors and to modulate inflammatory responses. MSCs have been shown to migrate to sites of injury and inflammation in response to soluble mediators including the chemokine stromal cell-derived factor-(SDF-)1, but during in vitro culture expansion MSCs lose surface expression of key homing receptors particularly of the SDF-1 receptor, CXCR4. Here we review studies on enhancement of SDF-1-directed migration of MSCs with the premise that their improved recruitment could translate to therapeutic benefits. We describe our studies on approaches to increase the CXCR4 expression in in vitro-expanded cord blood-derived MSCs, namely, transfection, using the commercial liposomal reagent IBAfect, chemical treatment with the histone deacetylase inhibitor valproic acid, and exposure to recombinant complement component C1q. These methodologies will be presented in the context of other cell targeting and delivery strategies that exploit pathways involved in MSC migration. Taken together, these findings indicate that MSCs can be manipulated in vitro to enhance their in vivo recruitment and efficacy for tissue repair. Leah A. Marquez-Curtis and Anna Janowska-Wieczorek Copyright © 2013 Leah A. Marquez-Curtis and Anna Janowska-Wieczorek. All rights reserved. The Study on Biocompatibility of Porous nHA/PLGA Composite Scaffolds for Tissue Engineering with Rabbit Chondrocytes In Vitro Wed, 27 Nov 2013 10:37:29 +0000 Objective. To examine the biocompatibility of a novel nanohydroxyapatite/poly[lactic-co-glycolic acid] (nHA/PLGA) composite and evaluate its feasibility as a scaffold for cartilage tissue engineering. Methods. Chondrocytes of fetal rabbit were cultured with nHA/PLGA scaffold in vitro and the cell viability was assessed by MTT assay first. Cells adhering to nHA/PLGA scaffold were then observed by inverted microscope and scanning electron microscope (SEM). The cell cycle profile was analyzed by flow cytometry. Results. The viability of the chondrocytes on the scaffold was not affected by nHA/PLGA comparing with the control group as it was shown by MTT assay. Cells on the surface and in the pores of the scaffold increased in a time-dependent manner. Results obtained from flow cytometry showed that there was no significant difference in cell cycle profiles between the coculture group and control (). Conclusion. The porous nHA/PLGA composite scaffold is a biocompatible and good kind of scaffold for cartilage tissue engineering. Lei Chen, Wei-Min Zhu, Zhi-Qiang Fei, Jie-Lin Chen, Jian-Yi Xiong, Ju-Feng Zhang, Li Duan, Jianghong Huang, Zhiyong Liu, Daping Wang, and Yanjun Zeng Copyright © 2013 Lei Chen et al. All rights reserved. Chitosan Dermal Substitute and Chitosan Skin Substitute Contribute to Accelerated Full-Thickness Wound Healing in Irradiated Rats Wed, 13 Nov 2013 10:38:40 +0000 Wounds with full-thickness skin loss are commonly managed by skin grafting. In the absence of a graft, reepithelialization is imperfect and leads to increased scar formation. Biomaterials can alter wound healing so that it produces more regenerative tissue and fewer scars. This current study use the new chitosan based biomaterial in full-thickness wound with impaired healing on rat model. Wounds were evaluated after being treated with a chitosan dermal substitute, a chitosan skin substitute, or duoderm CGF. Wounds treated with the chitosan skin substitute showed the most re-epithelialization (33.2 ± 2.8%), longest epithelial tongue (1.62 ± 0.13 mm), and shortest migratory tongue distance (7.11 ± 0.25 mm). The scar size of wounds treated with the chitosan dermal substitute (0.13 ± 0.02 cm) and chitosan skin substitute (0.16 ± 0.05 cm) were significantly decreased () compared with duoderm (0.45 ± 0.11 cm). Human leukocyte antigen (HLA) expression on days 7, 14, and 21 revealed the presence of human hair follicle stem cells and fibroblasts that were incorporated into and surviving in the irradiated wound. We have proven that a chitosan dermal substitute and chitosan skin substitute are suitable for wound healing in full-thickness wounds that are impaired due to radiation. Abu Bakar Mohd Hilmi, Ahmad Sukari Halim, Hasnan Jaafar, Abu Bakar Asiah, and Asma Hassan Copyright © 2013 Abu Bakar Mohd Hilmi et al. All rights reserved. Biocompatibility Assessment of Novel Collagen-Sericin Scaffolds Improved with Hyaluronic Acid and Chondroitin Sulfate for Cartilage Regeneration Thu, 07 Nov 2013 15:38:10 +0000 Cartilage tissue engineering (CTE) applications are focused towards the use of implantable biohybrids consisting of biodegradable scaffolds combined with in vitro cultured cells. Hyaluronic acid (HA) and chondroitin sulfate (CS) were identified as the most potent prochondrogenic factors used to design new biomaterials for CTE, while human adipose-derived stem cells (ASCs) were proved to display high chondrogenic potential. In this context, our aim was not only to build novel 3D porous scaffolds based on natural compounds but also to evaluate their in vitro biological performances. Therefore, for prospective CTE, collagen-sericin (Coll-SS) scaffolds improved with HA (5% or 10%) and CS (5% or 10%) were used as temporary physical supports for ASCs and were analyzed in terms of structural, thermal, morphological, and swelling properties and cytotoxic potential. To complete biocompatibility data, ASCs viability and proliferation potential were also assessed. Our studies revealed that Coll-SS hydrogels improved with 10% HA and 5% CS displayed the best biological performances in terms of cell viability, proliferation, morphology, and distribution. Thus, further work will address a novel 3D system including both HA 10% and CS 5% glycoproteins, which will probably be exposed to prochondrogenic conditions in order to assess its potential use in CTE applications. Sorina Dinescu, Bianca Gălăţeanu, Mădălina Albu, Adriana Lungu, Eugen Radu, Anca Hermenean, and Marieta Costache Copyright © 2013 Sorina Dinescu et al. All rights reserved. The Use of Carbon Nanotubes to Reinforce 45S5 Bioglass-Based Scaffolds for Tissue Engineering Applications Mon, 04 Nov 2013 15:02:11 +0000 Bioglass has been used for bone-filling material in bone tissue engineering, but its lean mechanical strength limits its applications in load-bearing positions. Carbon nanotubes (CNTs), with their high aspect ratio and excellent mechanical properties, have the potential to strengthen and toughen bioactive glass material without offsetting its bioactivity. Therefore, in this research, multiwall carbon nanotube (MWCNT)/45S5 Bioglass composite scaffolds have been successfully prepared by means of freeze casting process. 45S5 Bioglass was synthesized by the sol-gel processing method. The obtained material was characterized with X-ray powder diffraction (XRD). The mechanical properties of the scaffolds, such as compression strength and elastic modulus, were measured. Finally, compared with the scaffolds prepared by 100% 45S5 Bioglass powders, the addition of 0.25 wt.% MWCNTs increases the compressive strength and elastic modulus of 45S5 Bioglass scaffolds from 2.08 to 4.56 MPa (a 119% increase) and 111.50 to 266.59 MPa (a 139% increase), respectively. R. Touri, F. Moztarzadeh, Z. Sadeghian, D. Bizari, M. Tahriri, and M. Mozafari Copyright © 2013 R. Touri et al. All rights reserved. Transplantation of Autologous Minced Bladder Mucosa for a One-Step Reconstruction of a Tissue Engineered Bladder Conduit Thu, 31 Oct 2013 18:39:27 +0000 Surgical intervention is sometimes needed to create a conduit from the abdominal wall to the bladder for self-catheterization. We developed a method for tissue engineering a conduit for bladder emptying without in vitro cell culturing as a one-step procedure. In a porcine animal model bladder, wall tissue was excised and the mucosa was minced to small particles. The particles were attached to a tube in a 1 : 3 expansion rate with fibrin glue and transplanted back by attaching the tube to the bladder and through the abdominal wall. Sham served as controls. After 4-5 weeks, conduits were assessed in respect to macroscopic and microscopic appearance in 6 pigs. Two pigs underwent radiology before termination. Gross examination revealed a patent conduit with an opening to the bladder. Histology and immunostaining showed a multilayered transitional uroepithelium in all cases. Up to 89% of the luminal surface area was neoepithelialized but with a loose attachment to the submucosa. No epithelium was found in control animals. CT imaging revealed a patent channel that could be used for filling and emptying the bladder. Animals that experienced surgical complications did not form conduits. Minced autologous bladder mucosa can be transplanted around a tubular mold to create a conduit to the urinary bladder without in vitro culturing. Gisela Reinfeldt Engberg, Johan Lundberg, Clara Ibel Chamorro, Agneta Nordenskjöld, and Magdalena Fossum Copyright © 2013 Gisela Reinfeldt Engberg et al. All rights reserved. In Vitro Effects of Low-Intensity Pulsed Ultrasound Stimulation on the Osteogenic Differentiation of Human Alveolar Bone-Derived Mesenchymal Stem Cells for Tooth Tissue Engineering Mon, 30 Sep 2013 08:54:54 +0000 Ultrasound stimulation produces significant multifunctional effects that are directly relevant to alveolar bone formation, which is necessary for periodontal healing and regeneration. We focused to find out effects of specific duty cycles and the percentage of time that ultrasound is being generated over one on/off pulse period, under ultrasound stimulation. Low-intensity pulsed ultrasound ((LIPUS) 1 MHz) with duty cycles of 20% and 50% was used in this study, and human alveolar bone-derived mesenchymal stem cells (hABMSCs) were treated with an intensity of 50 mW/cm2 and exposure time of 10 min/day. hABMSCs exposed at duty cycles of 20% and 50% had similar cell viability (O.D.), which was higher () than that of control cells. The alkaline phosphatase (ALP) was significantly enhanced at 1 week with LIPUS treatment in osteogenic cultures as compared to control. Gene expressions showed significantly higher expression levels of CD29, CD44, COL1, and OCN in the hABMSCs under LIPUS treatment when compared to control after two weeks of treatment. The effects were partially controlled by LIPUS treatment, indicating that modulation of osteogenesis in hABMSCs was related to the specific stimulation. Furthermore, mineralized nodule formation was markedly increased after LIPUS treatment than that seen in untreated cells. Through simple staining methods such as Alizarin red and von Kossa staining, calcium deposits generated their highest levels at about 3 weeks. These results suggest that LIPUS could enhance the cell viability and osteogenic differentiation of hABMSCs, and could be part of effective treatment methods for clinical applications. KiTaek Lim, Jangho Kim, Hoon Seonwoo, Soo Hyun Park, Pill-Hoon Choung, and Jong Hoon Chung Copyright © 2013 KiTaek Lim et al. All rights reserved. P.R.L. Platelet Rich Lipotransfert: Our Experience and Current State of Art in the Combined Use of Fat and PRP Sat, 28 Sep 2013 11:49:46 +0000 The authors report their experience about the use of P.R.L. PLATELET RICH LIPOTRANSFERT method (platelet rich plasma mixed fat grafting) in 223 patients affected by soft tissue defects (ulcers, Romberg syndrome, Hemifacial atrophy, loss of substance, and signs of aging). This paper introduces the reader to PRP therapy and reviews the current literature on this emerging treatment modality, showing at the current clinical use of PRP in plastic and reconstructive surgery, with description of innovative methods and future prospects. This technique provides a promising alternative to surgery by promoting safe and natural healing. Here recent studies concerning the use of PRP in the treatment of chronic ulcers and soft tissue defect are reviewed. V. Cervelli, I. Bocchini, C. Di Pasquali, B. De Angelis, G. Cervelli, C. B. Curcio, A. Orlandi, M. G. Scioli, E. Tati, P. Delogu, and Pietro Gentile Copyright © 2013 V. Cervelli et al. All rights reserved. Osteogenic Matrix Cell Sheet Transplantation Enhances Early Tendon Graft to Bone Tunnel Healing in Rabbits Wed, 11 Sep 2013 13:59:53 +0000 The objective of this study was to determine whether osteogenic matrix cell sheets (OMCS) could induce bone formation around grafted tendons, thereby enhancing early stage tendon to bone tunnel healing in skeletally mature male Japanese white rabbits. First, the osteogenic potential of rabbit OMCS was evaluated. Then, the OMCS were transplanted into the interface between the grafted tendon and the bone tunnel created at the tibia. Histological assessments and biomechanical tensile testing were performed after 3 weeks. The rabbit OMCS showed high alkaline phosphatase (ALP) activity, positive staining of ALP, and osteogenic potential when transplanted subcutaneously with beta tricalcium phosphate disks. Newly formed bony walls and positive collagen type I staining were seen around the grafted tendon with OMCS transplantation, whereas such bony walls were thinner or less frequent without OMCS transplantation. Micro-computed tomography images showed significantly higher bone volume in the OMCS transplantation group. The pullout strength was significantly higher with OMCS ( N/mm2) than without OMCS ( N/mm2). These results show that OMCS enhance early tendon to bone tunnel healing. This method can be applied to cases requiring early tendon to bone tunnel healing after ligament reconstruction surgery. Yusuke Inagaki, Kota Uematsu, Manabu Akahane, Yusuke Morita, Munehiro Ogawa, Tomoyuki Ueha, Takamasa Shimizu, Tomohiko Kura, Kenji Kawate, and Yasuhito Tanaka Copyright © 2013 Yusuke Inagaki et al. All rights reserved. Myogenic Differentiation of Mesenchymal Stem Cells in a Newly Developed Neurotised AV-Loop Model Tue, 10 Sep 2013 14:31:43 +0000 Generation of axially vascularized muscle tissue constitutes a promising new approach to restoration of damaged muscle tissue. Mesenchymal stemcells (MSC), with their ability to be expanded to large cell numbers without losing their differentiation capacity into the myogenic lineage, could offer a promising cell source to generate neomuscle tissue. In vitro experiments showed that cocultures of primary myoblasts and MSC undergo myogenic differentiation by stimulation with bFGF and dexamethasone. A newly developed AV-Loop model with neurotization was established in this study. It encompasses axial vascularization and the additional implantation of a motor nerve serving as myogenic stimulator. Myoblasts and MSCs were coimplantated in a prevascularized isolation chamber. Cells were differentiated by addition of bFGF and dexamethasone plus implantation of a motor nerve. After 8 weeks, we could observe areas of myogenic differentiation with α-sarcomeric actin and MHC expression in the constructs. Quantitative PCR analysis showed an expression of myogenic markers in all specimens. Thus, neurotization and addition of bFGF and dexamethasone allow myogenic differentiation of MSC in an axially vascularized in vivo model for the first time. These findings are a new step towards clinical applicability of skeletal muscle tissue engineering and display its potential for regenerative medicine. Franz F. Bitto, Dorothee Klumpp, Claudia Lange, Anja M. Boos, Andreas Arkudas, Oliver Bleiziffer, Raymund E. Horch, Ulrich Kneser, and Justus P. Beier Copyright © 2013 Franz F. Bitto et al. All rights reserved. Mesenchymal Stem Cells and Platelet Gel Improve Bone Deposition within CAD-CAM Custom-Made Ceramic HA Scaffolds for Condyle Substitution Sun, 01 Sep 2013 14:04:31 +0000 Purpose. This study evaluated the efficacy of a regenerative approach using mesenchymal stem cells (MSCs) and CAD-CAM customized pure and porous hydroxyapatite (HA) scaffolds to replace the temporomandibular joint (TMJ) condyle. Methods. Pure HA scaffolds with a 70% total porosity volume were prototyped using CAD-CAM technology to replace the two temporomandibular condyles (left and right) of the same animal. MSCs were derived from the aspirated iliac crest bone marrow, and platelets were obtained from the venous blood of the sheep. Custom-made surgical guides were created by direct metal laser sintering and were used to export the virtual planning of the bone cut lines into the surgical environment. Sheep were sacrificed 4 months postoperatively. The HA scaffolds were explanted, histological specimens were prepared, and histomorphometric analysis was performed. Results. Analysis of the porosity reduction for apposition of newly formed bone showed a statistically significant difference in bone formation between condyles loaded with MSC and condyles without (). The bone ingrowth (BI) relative values of split-mouth comparison (right versus left side) showed a significant difference between condyles with and without MSCs (). Analysis of the test and control sides in the same animal using a split-mouth study design was performed; the condyle with MSCs showed greater bone formation. Conclusion. The split-mouth design confirmed an increment of bone regeneration into the HA scaffold of up to 797% upon application of MSCs. L. Ciocca, D. Donati, S. Ragazzini, B. Dozza, F. Rossi, M. Fantini, A. Spadari, N. Romagnoli, E. Landi, A. Tampieri, A. Piattelli, G. Iezzi, and R. Scotti Copyright © 2013 L. Ciocca et al. All rights reserved. Activation of the ERK1/2 Signaling Pathway during the Osteogenic Differentiation of Mesenchymal Stem Cells Cultured on Substrates Modified with Various Chemical Groups Wed, 28 Aug 2013 09:24:26 +0000 The current study examined the influence of culture substrates modified with the functional groups –OH, –COOH, –NH2, and –CH3 using SAMs technology, in conjunction with TAAB control, on the osteogenic differentiation of rabbit BMSCs. The CCK-8 assay revealed that BMSCs exhibited substrate-dependent cell viability. The cells plated on –NH2- and –OH-modified substrates were well spread and homogeneous, but those on the –COOH- and –CH3-modified substrates showed more rounded phenotype. The mRNA expression of BMSCs revealed that –NH2-modified substrate promoted the mRNA expression and osteogenic differentiation of the BMSCs. The contribution of ERK1/2 signaling pathway to the osteogenic differentiation of BMSCs cultured on the –NH2-modified substrate was investigated in vitro. The –NH2-modified substrate promoted the expression of integrins; the activation of FAK and ERK1/2. Inhibition of ERK1/2 activation by PD98059, a specific inhibitor of the ERK signaling pathway, blocked ERK1/2 activation in a dose-dependent manner, as revealed for expression of Cbfα-1 and ALP. Blockade of ERK1/2 phosphorylation in BMSCs by PD98059 suppressed osteogenic differentiation on chemical surfaces. These findings indicate a potential role for ERK in the osteogenic differentiation of BMSCs on surfaces modified by specific chemical functional groups, indicating that the microenvironment affects the differentiation of BMSCs. This observation has important implications for bone tissue engineering. Bing Bai, Jin He, Yan-Shu Li, Xiu-Mei Wang, Hong-Jun Ai, and Fu-Zhai Cui Copyright © 2013 Bing Bai et al. All rights reserved. The Influence of Chain Microstructure of Biodegradable Copolyesters Obtained with Low-Toxic Zirconium Initiator to In Vitro Biocompatibility Sun, 25 Aug 2013 15:02:16 +0000 Because of the wide use of biodegradable materials in tissue engineering, it is necessary to obtain biocompatible polymers with different mechanical and physical properties as well as degradation ratio. Novel co- and terpolymers of various composition and chain microstructure have been developed and applied for cell culture. The aim of this study was to evaluate the adhesion and proliferation of human chondrocytes to four biodegradable copolymers: lactide-coglycolide, lactide-co--caprolactone, lactide-co-trimethylene carbonate, glycolide-co--caprolactone, and one terpolymer glycolide-colactide-co--caprolactone synthesized with the use of zirconium acetylacetonate as a nontoxic initiator. Chain microstructure of the copolymers was analyzed by means of 1H and 13C NMR spectroscopy and surface properties by AFM technique. Cell adhesion and proliferation were determined by CyQUANT Cell Proliferation Assay Kit. After 4 h the chondrocyte adhesion on the surface of studied materials was comparable to standard TCPS. Cell proliferation occurred on all the substrates; however, among the studied polymers poly(L-lactide-coglycolide) 85 : 15 that characterized the most blocky structure best supported cell growth. Chondrocytes retained the cell membrane integrity evaluated by the LDH release assay. As can be summarized from the results of the study, all the studied polymers are well tolerated by the cells that make them appropriate for human chondrocytes growth. Arkadiusz Orchel, Katarzyna Jelonek, Janusz Kasperczyk, Piotr Dobrzynski, Andrzej Marcinkowski, Elzbieta Pamula, Joanna Orchel, Ireneusz Bielecki, and Anna Kulczycka Copyright © 2013 Arkadiusz Orchel et al. All rights reserved. Engineering Parameters in Bioreactor’s Design: A Critical Aspect in Tissue Engineering Mon, 05 Aug 2013 11:38:11 +0000 Bioreactors are important inevitable part of any tissue engineering (TE) strategy as they aid the construction of three-dimensional functional tissues. Since the ultimate aim of a bioreactor is to create a biological product, the engineering parameters, for example, internal and external mass transfer, fluid velocity, shear stress, electrical current distribution, and so forth, are worth to be thoroughly investigated. The effects of such engineering parameters on biological cultures have been addressed in only a few preceding studies. Furthermore, it would be highly inefficient to determine the optimal engineering parameters by trial and error method. A solution is provided by emerging modeling and computational tools and by analyzing oxygen, carbon dioxide, and nutrient and metabolism waste material transports, which can simulate and predict the experimental results. Discovering the optimal engineering parameters is crucial not only to reduce the cost and time of experiments, but also to enhance efficacy and functionality of the tissue construct. This review intends to provide an inclusive package of the engineering parameters together with their calculation procedure in addition to the modeling techniques in TE bioreactors. Nasim Salehi-Nik, Ghassem Amoabediny, Behdad Pouran, Hadi Tabesh, Mohammad Ali Shokrgozar, Nooshin Haghighipour, Nahid Khatibi, Fatemeh Anisi, Khosrow Mottaghy, and Behrouz Zandieh-Doulabi Copyright © 2013 Nasim Salehi-Nik et al. All rights reserved. Development of Composite Scaffolds for Load-Bearing Segmental Bone Defects Mon, 29 Jul 2013 10:00:20 +0000 The need for a suitable tissue-engineered scaffold that can be used to heal load-bearing segmental bone defects (SBDs) is both immediate and increasing. During the past 30 years, various ceramic and polymer scaffolds have been investigated for this application. More recently, while composite scaffolds built using a combination of ceramics and polymeric materials are being investigated in a greater number, very few products have progressed from laboratory benchtop studies to preclinical testing in animals. This review is based on an exhaustive literature search of various composite scaffolds designed to serve as bone regenerative therapies. We analyzed the benefits and drawbacks of different composite scaffold manufacturing techniques, the properties of commonly used ceramics and polymers, and the properties of currently investigated synthetic composite grafts. To follow, a comprehensive review of in vivo models used to test composite scaffolds in SBDs is detailed to serve as a guide to design appropriate translational studies and to identify the challenges that need to be overcome in scaffold design for successful translation. This includes selecting the animal type, determining the anatomical location within the animals, choosing the correct study duration, and finally, an overview of scaffold performance assessment. Marcello Pilia, Teja Guda, and Mark Appleford Copyright © 2013 Marcello Pilia et al. All rights reserved. Isolation and Characterisation of Mesenchymal Stem Cells from Different Regions of the Human Umbilical Cord Thu, 25 Jul 2013 11:35:38 +0000 Umbilical cords as a source of stem cells are of increasing interest for cell therapies as they present little ethical consideration and are reported to contain immune privileged cells which may be suitable for allogeneic based therapies. Mesenchymal stem cells (MSCs) sourced from several different cord regions, including artery, vein, cord lining, and Wharton’s jelly, are described in the literature. However, no one study has yet isolated and characterised MSCs from all regions of the same cord to determine the most suitable cells for cell based therapeutics. Claire Mennan, Karina Wright, Atanu Bhattacharjee, Birender Balain, James Richardson, and Sally Roberts Copyright © 2013 Claire Mennan et al. All rights reserved. In Vivo Biocompatibility of PLGA-Polyhexylthiophene Nanofiber Scaffolds in a Rat Model Tue, 23 Jul 2013 09:03:32 +0000 Electroactive polymers have applications in tissue engineering as a physical template for cell adhesion and carry electrical signals to improve tissue regeneration. Present study demonstrated the biocompatibility and biodegradability of poly(lactide-co-glycolide)-poly(3-hexylthiophene) (PLGA-PHT) blend electrospun scaffolds in a subcutaneous rat model. The biocompatibility of PLGA-undoped PHT, PLGA-doped PHT, and aligned PLGA-doped PHT nanofibers was evaluated and compared with random PLGA fibers. The animals were sacrificed at 2, 4, and 8 weeks; the surrounding tissue along with the implant was removed to evaluate biocompatibility and biodegradability by histologic analysis and GPC, respectively. Histology results demonstrated that all scaffolds except PLGA-undoped PHT showed decrease in inflammation over time. It was observed that the aligned PLGA-doped PHT fibers elicited moderate response at 2 weeks, which further reduced to a mild response over time with well-organized tissue structure and collagen deposition. The degradation of aligned nanofibers was found to be very slow when compared to random fibers. Further, there was no reduction in the molecular weight of undoped form of PHT throughout the study. These experiments revealed the biocompatibility and biodegradability of PLGA-PHT nanofibers that potentiate it to be used as a biomaterial for various applications. Anuradha Subramanian, Uma Maheswari Krishnan, and Swaminathan Sethuraman Copyright © 2013 Anuradha Subramanian et al. All rights reserved. Three-Dimensional Supermacroporous Carrageenan-Gelatin Cryogel Matrix for Tissue Engineering Applications Sun, 07 Jul 2013 16:12:43 +0000 A tissue-engineered polymeric scaffold should provide suitable macroporous structure similar to that of extracellular matrix which can induce cellular activities and guide tissue regeneration. Cryogelation is a technique in which appropriate monomers or polymeric precursors frozen at sub-zero temperature leads to the formation of supermacroporous cryogel matrices. In this study carrageenan-gelatin (natural polymers) cryogels were synthesized by using glutaraldehyde and 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride and N-hydroxysuccinimide (EDC-NHS) as crosslinking agent at optimum concentrations. Matrices showed large and interconnected pores which were in the range of 60–100 μm diameter. Unconfined compression analysis showed elasticity and physical integrity of all cryogels, as these matrices regained their original length after 90% compressing from the original size. Moreover Young’s modulus was found to be in the range of 4–11 kPa for the dry cryogel sections. These cryogels also exhibited good in vitro degradation capacity at 37 °C within 4 weeks of incubation. Supermacroporous carrageenan-gelatin cryogels showed efficient cell adherence and proliferation of Cos-7 cells which was examined by SEM. PI nuclear stain was used to observe cell-matrix interaction. Cytotoxicity of the scaffolds was checked by MTT assay which showed that cryogels are biocompatible and act as a potential material for tissue engineering and regenerative medicine. Archana Sharma, Sumrita Bhat, Tanushree Vishnoi, Vijayashree Nayak, and Ashok Kumar Copyright © 2013 Archana Sharma et al. All rights reserved. Side-by-Side Comparison of the Biological Characteristics of Human Umbilical Cord and Adipose Tissue-Derived Mesenchymal Stem Cells Sun, 07 Jul 2013 08:08:08 +0000 Both human adipose tissue-derived mesenchymal stem cells (ASCs) and umbilical cord-derived mesenchymal stem cells (UC-MSCs) have been explored as attractive mesenchymal stem cells (MSCs) sources, but very few parallel comparative studies of these two cell types have been made. We designed a side-by-side comparative study by isolating MSCs from the adipose tissue and umbilical cords from mothers delivering full-term babies and thus compared the various biological aspects of ASCs and UC-MSCs derived from the same individual, in one study. Both types of cells expressed cell surface markers characteristic of MSCs. ASCs and UC-MSCs both could be efficiently induced into adipocytes, osteoblasts, and neuronal phenotypes. While there were no significant differences in their osteogenic differentiation, the adipogenesis of ASCs was more prominent and efficient than UC-MSCs. In the meanwhile, ASCs responded better to neuronal induction methods, exhibiting the higher differentiation rate in a relatively shorter time. In addition, UC-MSCs exhibited a more prominent secretion profile of cytokines than ASCs. These results indicate that although ASCs and UC-MSCs share considerable similarities in their immunological phenotype and pluripotentiality, certain biological differences do exist, which might have different implications for future cell-based therapy. Li Hu, Jingqiong Hu, Jiajia Zhao, Jiarong Liu, Weixiang Ouyang, Chao Yang, Niya Gong, Luyang Du, Abhilasha Khanal, and Lili Chen Copyright © 2013 Li Hu et al. All rights reserved. Comparative Study of Various Delivery Methods for the Supply of Alpha-Ketoglutarate to the Neural Cells for Tissue Engineering Wed, 26 Jun 2013 11:20:29 +0000 Delivery of growth factors or bioactive molecules plays an important role in tissue engineering, as the duration to which these are supplied can modulate the cell fate. Thus, the delivery method plays an important role, and the same is presented in this work wherein the exogenous supply of alpha-ketoglutarate (α-KG) gave better results for fast proliferating cells as compared to delivery by microspheres or microspheres incorporated scaffolds which can be used while culturing slow growing cells. All these studies were performed in two dimensional (2D) and three dimensional (3D) setups in which chitosan-gelatin-polypyrrole has been used as 3-D scaffolds. Chitosan and gelatin microspheres alone as well as incorporated in the cryogels were characterized. MTT assay done using neuro-2a cell line showed approximately 42% and 70% increment in cellular proliferation when gelatin and chitosan microspheres were added in a 3-D setup, respectively, as compared to the control. Biochemical analysis of ammonia showed 6-fold reductions in ammonia level in a 3-D setup compared to the control. We also studied the synthesis of a neurotransmitter-like glutamate and found that its concentration increased up to 0.25 mg/ml when the microspheres were added exogenously in a 3-D system. Tanushree Vishnoi and Ashok Kumar Copyright © 2013 Tanushree Vishnoi and Ashok Kumar. All rights reserved. Detergent-Enzymatic Decellularization of Swine Blood Vessels: Insight on Mechanical Properties for Vascular Tissue Engineering Thu, 20 Jun 2013 09:07:45 +0000 Small caliber vessels substitutes still remain an unmet clinical need; few autologous substitutes are available, while synthetic grafts show insufficient patency in the long term. Decellularization is the complete removal of all cellular and nuclear matters from a tissue while leaving a preserved extracellular matrix representing a promising tool for the generation of acellular scaffolds for tissue engineering, already used for various tissues with positive outcomes. The aim of this work is to investigate the effect of a detergent-enzymatic decellularization protocol on swine arteries in terms of cell removal, extracellular matrix preservation, and mechanical properties. Furthermore, the effect of storage at −80°C on the mechanical properties of the tissue is evaluated. Swine arteries were harvested, frozen, and decellularized; histological analysis revealed complete cell removal and preserved extracellular matrix. Furthermore, the residual DNA content in decellularized tissues was far low compared to native one. Mechanical testings were performed on native, defrozen, and decellularized tissues; no statistically significant differences were reported for Young’s modulus, ultimate stress, compliance, burst pressure, and suture retention strength, while ultimate strain and stress relaxation of decellularized vessels were significantly different from the native ones. Considering the overall results, the process was confirmed to be suitable for the generation of acellular scaffolds for vascular tissue engineering. Alessandro F. Pellegata, M. Adelaide Asnaghi, Ilaria Stefani, Anna Maestroni, Silvia Maestroni, Tommaso Dominioni, Sandro Zonta, Gianpaolo Zerbini, and Sara Mantero Copyright © 2013 Alessandro F. Pellegata et al. All rights reserved. Effects of Electromagnetic Fields on Osteogenesis of Human Alveolar Bone-Derived Mesenchymal Stem Cells Wed, 19 Jun 2013 14:31:01 +0000 This study was performed to investigate the effects of extremely low frequency pulsed electromagnetic fields (ELF-PEMFs) on the proliferation and differentiation of human alveolar bone-derived mesenchymal stem cells (hABMSCs). Osteogenesis is a complex series of events involving the differentiation of mesenchymal stem cells to generate new bone. In this study, we examined not merely the effect of ELF-PEMFs on cell proliferation, alkaline phosphatase (ALP) activity, and mineralization of the extracellular matrix but vinculin, vimentin, and calmodulin (CaM) expressions in hABMSCs during osteogenic differentiation. Exposure of hABMSCs to ELF-PEMFs increased proliferation by 15% compared to untreated cells at day 5. In addition, exposure to ELF-PEMFs significantly increased ALP expression during the early stages of osteogenesis and substantially enhanced mineralization near the midpoint of osteogenesis within 2 weeks. ELF-PEMFs also increased vinculin, vimentin, and CaM expressions, compared to control. In particular, CaM indicated that ELF-PEMFs significantly altered the expression of osteogenesis-related genes. The results indicated that ELF-PEMFs could enhance early cell proliferation in hABMSCs-mediated osteogenesis and accelerate the osteogenesis. KiTaek Lim, Jin Hexiu, Jangho Kim, Hoon Seonwoo, Woo Jae Cho, Pill-Hoon Choung, and Jong Hoon Chung Copyright © 2013 KiTaek Lim et al. All rights reserved. Modification of Decellularized Goat-Lung Scaffold with Chitosan/Nanohydroxyapatite Composite for Bone Tissue Engineering Applications Thu, 13 Jun 2013 13:02:15 +0000 Decellularized goat-lung scaffold was fabricated by removing cells from cadaver goat-lung tissue, and the scaffold was modified with chitosan/nanohydroxyapatite composite for the purpose of bone tissue engineering applications. MTT assay with osteoblasts, seeded over the chitosan/nanohydroxyapatite-modified decellularized scaffold, demonstrated significantly higher cell growth as compared to the decellularized scaffold without modification. SEM analysis of cell-seeded scaffold, after incubation for 7 days, represented a good cell adhesion, and the cells spread over the chitosan/nanohydroxyapatite-modified decellularized scaffold. Expression of bone-tissue-specific osteocalcin gene in the osteoblast cells grown over the chitosan/nanohydroxyapatite-modified decellularized scaffold clearly signifies that the cells maintained their osteoblastic phenotype with the chitosan/nanohydroxyapatite-modified decellularized scaffold. Therefore, it can be concluded that the decellularized goat-lung scaffold-modified with chitosan/nanohydroxyapatite composite, may provide enhanced osteogenic potential when used as a scaffold for bone tissue engineering. Sweta K. Gupta, Amit K. Dinda, Pravin D. Potdar, and Narayan C. Mishra Copyright © 2013 Sweta K. Gupta et al. All rights reserved. Preparation, Modification, and Characterization of Alginate Hydrogel with Nano-/Microfibers: A New Perspective for Tissue Engineering Wed, 05 Jun 2013 14:43:05 +0000 We aimed to develop an alginate hydrogel (AH) modified with nano-/microfibers of titanium dioxide (nfTD) and hydroxyapatite (nfHY) and evaluated its biological and chemical properties. Nano-/microfibers of nfTD and nfHY were combined with AH, and its chemical properties were evaluated by FTIR spectroscopy, X-ray diffraction, energy dispersive X-Ray analysis, and the cytocompatibility by the WST-1 assay. The results demonstrate that the association of nfTD and nfHY nano-/microfibers to AH did not modified the chemical characteristics of the scaffold and that the association was not cytotoxic. In the first 3 h of culture with NIH/3T3 cells nfHY AH scaffolds showed a slight increase in cell viability when compared to AH alone or associated with nfTD. However, an increase in cell viability was observed in 24 h when nfTD was associated with AH scaffold. In conclusion our study demonstrates that the combination of nfHY and nfTD nano-/microfibers in AH scaffold maintains the chemical characteristics of alginate and that this association is cytocompatible. Additionally the combination of nfHY with AH favored cell viability in a short term, and the addition of nfTD increased cell viability in a long term. Bianca Palma Santana, Fernanda Nedel, Evandro Piva, Rodrigo Varella de Carvalho, Flávio Fernando Demarco, and Neftali Lenin Villarreal Carreño Copyright © 2013 Bianca Palma Santana et al. All rights reserved. Differentiation of Mesenchymal Stem Cells from Human Umbilical Cord Tissue into Odontoblast-Like Cells Using the Conditioned Medium of Tooth Germ Cells In Vitro Mon, 13 May 2013 14:59:16 +0000 The easily accessible mesenchymal stem cells in the Wharton's jelly of human umbilical cord tissue (hUCMSCs) have excellent proliferation and differentiation potential, but it remains unclear whether hUCMSCs can differentiate into odontoblasts. In this study, mesenchymal stem cells were isolated from the Wharton's jelly of human umbilical cord tissue using the simple method of tissue blocks culture attachment. UCMSC surface marker expression was then evaluated for the isolated cells using flow cytometry. The third-passage hUCMSCs induced by conditioned medium from developing tooth germ cells (TGC-CM) displayed high alkaline phosphatase (ALP) levels (), an enhanced ability to proliferate (), and the presence of mineralized nodules. These effects were not observed in cells treated with regular medium. After induction of hUCMSCs, the results of reverse transcriptional polymerase chain reaction (PCR) indicated that the dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP1) genes were significantly tested. Additionally, dentin sialoprotein (DSP) and DMP1 demonstrated significant levels of staining in an immunofluorescence analysis. In contrast, the control cells failed to display the characteristics of odontoblasts. Taken together, these results suggest that hUCMSCs can be induced to differentiate into odontoblast-like cells with TGC-CM and provide a novel strategy for tooth regeneration research. Tian Xia Li, Jie Yuan, Yan Chen, Li Jie Pan, Chun Song, Liang Jia Bi, and Xiao Hui Jiao Copyright © 2013 Tian Xia Li et al. All rights reserved. Late Adherent Human Bone Marrow Stromal Cells Form Bone and Restore the Hematopoietic Microenvironment In Vivo Wed, 24 Apr 2013 10:15:14 +0000 Bone marrow stromal cells (BMSCs) are a valuable resource for skeletal regenerative medicine because of their osteogenic potential. In spite of the very general term “stem cell,” this population of cells is far from homogeneous, and different BMSCs clones have greatly different phenotypic properties and, therefore, potentially different therapeutic potential. Adherence to a culture flask surface is a primary defining characteristic of BMSCs. We hypothesized that based on the adherence time we could obtain an enriched population of cells with a greater therapeutic potential. We characterized two populations of bone marrow-derived cells, those that adhered by three days (R-cells) and those that did not adhere by three days but did by six days (L-cells). Clones derived from L-cells could be induced into adipogenic, chondrogenic, and osteogenic differentiation in vitro. L-cells appeared to have greater proliferative capacity, as manifested by larger colony diameter and clones with higher CD146 expression. Only clones from L-cells developed bone marrow stroma in vivo. We conclude that the use of late adherence of BMSCs is one parameter that can be used to enrich for cells that will constitute a superior final product for cell therapy in orthopedics. Verônica Fernandes Vianna, Danielle Cabral Bonfim, Amanda dos Santos Cavalcanti, Marco Cury Fernandes, Suzana Assad Kahn, Priscila Ladeira Casado, Inayá Correa Lima, Samuel S. Murray, Elsa J. Brochmann Murray, and Maria Eugenia Leite Duarte Copyright © 2013 Verônica Fernandes Vianna et al. All rights reserved. Comparison of Explant-Derived and Enzymatic Digestion-Derived MSCs and the Growth Factors from Wharton’s Jelly Tue, 09 Apr 2013 10:58:21 +0000 Wharton’s jelly is not only one of the most promising tissue sources for mesenchymal stem cells (MSCs) but also a source of natural growth factors. To prove that we can get both natural growth factors and MSCs from Wharton’s jelly, we compared cellular characteristics and the level of basic fibroblast growth factor (bFGF) from samples using the explant culture method to those derived from the traditional enzymatic culture method. The levels of bFGF were  ng/g on day 3,  ng/g on day 6, and decreased to  ng/g on day 14. The total amount of bFGF released was  ng/g on explant culture. Compared with the traditional enzymatic digestion method, the explant culture method showed a tendency to release higher levels of bFGF in supernatant media for the first week of culture, and the higher cellular yield at passage 0 (/g versus /g, ). In addition, the genes related to mitosis were upregulated in the explant-derived MSCs. Jong Hyun Yoon, Eun Youn Roh, Sue Shin, Nam Hee Jung, Eun Young Song, Ju Young Chang, Byoung Jae Kim, and Hye Won Jeon Copyright © 2013 Jong Hyun Yoon et al. All rights reserved. In Vitro Construction of Scaffold-Free Bilayered Tissue-Engineered Skin Containing Capillary Networks Wed, 27 Mar 2013 15:37:31 +0000 Many types of skin substitutes have been constructed using exogenous materials. Angiogenesis is an important factor for tissue-engineered skin constructs. In this study, we constructed a scaffold-free bilayered tissue-engineered skin containing a capillary network. First, we cocultured dermal fibroblasts with dermal microvascular endothelial cells at a ratio of 2 : 1. A fibrous sheet was formed by the interactions between the fibroblasts and the endothelial cells, and capillary-like structures were observed after 20 days of coculture. Epithelial cells were then seeded on the fibrous sheet to assemble the bilayered tissue. HE staining showed that tissue-engineered skin exhibited a stratified epidermis after 7 days. Immunostaining showed that the epithelium promoted the formation of capillary-like structures. Transmission electron microscopy (TEM) analysis showed that the capillary-like structures were typical microblood vessels. ELISA demonstrated that vascularization was promoted by significant upregulation of vascularization associated growth factors due to interactions among the 3 types of cells in the bilayer, as compared to cocultures of fibroblast and endothelial cells and monocultures. Yuan Liu, Hailang Luo, Xinwen Wang, Akimichi Takemura, Yi Ru Fang, Yan Jin, and Fumihiko Suwa Copyright © 2013 Yuan Liu et al. All rights reserved. Stem Cell Therapy for Lower Extremity Diabetic Ulcers: Where Do We Stand? Mon, 18 Mar 2013 09:16:53 +0000 The impairment of wound healing in diabetic patients is an important clinical problem affecting millions of patients worldwide. Various clinical and basic science studies show that stem cell therapy, as a regenerative medical therapy, can be a good solution. In this paper, we begin with an introduction of the cellular mechanism of the diabetic ulcer. We will then discuss the advantages and limitations of various stem cell therapies that have been under extensive recent study. Mei Yang, Lingling Sheng, Tian R. Zhang, and Qingfeng Li Copyright © 2013 Mei Yang et al. All rights reserved. Trends in Tissue Engineering for Blood Vessels Thu, 08 Nov 2012 15:44:19 +0000 Over the years, cardiovascular diseases continue to increase and affect not only human health but also the economic stability worldwide. The advancement in tissue engineering is contributing a lot in dealing with this immediate need of alleviating human health. Blood vessel diseases are considered as major cardiovascular health problems. Although blood vessel transplantation is the most convenient treatment, it has been delimited due to scarcity of donors and the patient’s conditions. However, tissue-engineered blood vessels are promising alternatives as mode of treatment for blood vessel defects. The purpose of this paper is to show the importance of the advancement on biofabrication technology for treatment of soft tissue defects particularly for vascular tissues. This will also provide an overview and update on the current status of tissue reconstruction especially from autologous stem cells, scaffolds, and scaffold-free cellular transplantable constructs. The discussion of this paper will be focused on the historical view of cardiovascular tissue engineering and stem cell biology. The representative studies featured in this paper are limited within the last decade in order to trace the trend and evolution of techniques for blood vessel tissue engineering. Judee Grace Nemeno-Guanzon, Soojung Lee, Johan Robert Berg, Yong Hwa Jo, Jee Eun Yeo, Bo Mi Nam, Yong-Gon Koh, and Jeong Ik Lee Copyright © 2012 Judee Grace Nemeno-Guanzon et al. All rights reserved. Adipose Tissue Regeneration: A State of the Art Wed, 03 Oct 2012 10:27:55 +0000 Adipose tissue pathologies and defects have always represented a reconstructive challenge for plastic surgeons. In more recent years, several allogenic and alloplastic materials have been developed and used as fillers for soft tissue defects. However, their clinical use has been limited by further documented complications, such as foreign-body reactions potentially affecting function, degradation over time, and the risk for immunogenicity. Tissue-engineering strategies are thus being investigated to develop methods for generating adipose tissue. This paper will discuss the current state of the art in adipose tissue engineering techniques, exploring the biomaterials used, stem cells application, culture strategies, and current regulatory framework that are in use are here described and discussed. Alessandro Casadei, Roberta Epis, Letizia Ferroni, Ilaria Tocco, Chiara Gardin, Eriberto Bressan, Stefano Sivolella, Vincenzo Vindigni, Paolo Pinton, Giuseppe Mucci, and Barbara Zavan Copyright © 2012 Alessandro Casadei et al. All rights reserved. Controlling Self-Renewal and Differentiation of Stem Cells via Mechanical Cues Tue, 02 Oct 2012 12:17:54 +0000 The control of stem cell response in vitro, including self-renewal and lineage commitment, has been proved to be directed by mechanical cues, even in the absence of biochemical stimuli. Through integrin-mediated focal adhesions, cells are able to anchor onto the underlying substrate, sense the surrounding microenvironment, and react to its properties. Substrate-cell and cell-cell interactions activate specific mechanotransduction pathways that regulate stem cell fate. Mechanical factors, including substrate stiffness, surface nanotopography, microgeometry, and extracellular forces can all have significant influence on regulating stem cell activities. In this paper, we review all the most recent literature on the effect of purely mechanical cues on stem cell response, and we introduce the concept of “force isotropy” relevant to cytoskeletal forces and relevant to extracellular loads acting on cells, to provide an interpretation of how the effects of insoluble biophysical signals can be used to direct stem cells fate in vitro. Michele M. Nava, Manuela T. Raimondi, and Riccardo Pietrabissa Copyright © 2012 Michele M. Nava et al. All rights reserved. The Osteogenic Study of Tissue Engineering Bone with BMP2 and BMP7 Gene-Modified Rat Adipose-Derived Stem Cell Thu, 21 Jun 2012 08:33:45 +0000 To evaluate the feasibility and advantages of constructing a novel tissue engineering bone, using 𝛽-tricalcium phosphate (𝛽-TCP) and rat adipose-derived stem cells (ADSCs), modified with BMP2 and BMP7 by lentivirus. In the present study, ADSCs transfected with Lv-BMP2 and Lv-BMP7, alone or together, were seeded on 𝛽-TCP scaffold and cultured in vitro. Based on the results of DNA assay, alkaline phosphatase (ALP) activity, alizarin red staining and osteogenic marker genes expression analysis, the BMP2 and BMP7 genes cotransfection group exhibited a higher degree of osteogenic differentiation in vitro. To investigate the in vivo osteogenesis of the tissue engineering bone, the ADSCs/𝛽-TCP constructs were implanted in rat femurs defects for 6 weeks and studied histomorphology and radiography. The results showed that BMP2 and BMP7 genes cotransfection group dramatically enhanced the efficiency of new bone formation than BMP2 group and BMP7 group in vivo. These results demonstrated that it was advantageous to construct tissue engineering bone using ADSCs cotransfected with BMP2 and BMP7 on 𝛽-TCP, providing a potential way for treating bone defects. Wang Qing, Chen Guang-Xing, Guo Lin, and Yang Liu Copyright © 2012 Wang Qing et al. All rights reserved. Physical Properties and Biocompatibility of a Core-Sheath Structure Composite Scaffold for Bone Tissue Engineering In Vitro Thu, 15 Mar 2012 11:07:38 +0000 Scaffolds play a critical role in the practical realization of bone tissue engineering. The purpose of this study was to assess whether a core-sheath structure composite scaffold possesses admirable physical properties and biocompatibility in vitro. A novel scaffold composed of poly(lactic-co-glycolic acid)/β-tricalcium phosphate (PLGA/β-TCP) skeleton wrapped with Type I collagen via low-temperature deposition manufacturing (LDM) was prepared, and bone mesenchymal stem cells (BMSCs) were used to evaluate cell behavior on the scaffold. PLGA/β-TCP skeleton was chosen as the control group. Physical properties were evaluated by pority ratio, compressive strength, and Young’s modulus. Scanning electron microscope (SEM) was used to study morphology of cells. Hydrophilicity was evaluated by water absorption ratio. Cell proliferation was tested by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay (MTT). Osteogenic differentiation of BMSCs was evaluated by alkaline phosphates activity (ALP). The results indicated that physical properties of the novel scaffold were as good as those of the control group, hydrophilicity was observably better (𝑃<0.01) than that of control group, and abilities of proliferation and osteogenic differentiation of BMSCs on novel scaffold were significantly greater (𝑃<0.05) than those of control group, which suggests that the novel scaffold possesses preferable characteristics and have high value in bone tissue engineering. Chuangjian Wang, Guolin Meng, Laquan Zhang, Zuo Xiong, and Jian Liu Copyright © 2012 Chuangjian Wang et al. All rights reserved. Xenogenic Esophagus Scaffolds Fixed with Several Agents: Comparative In Vivo Study of Rejection and Inflammation Thu, 08 Mar 2012 08:38:19 +0000 Most infants with long-gap esophageal atresia receive an esophageal replacement with tissue from stomach or colon, because the native esophagus is too short for true primary repair. Tissue-engineered esophageal conducts could present an attractive alternative. In this paper, circular decellularized porcine esophageal scaffold tissues were implanted subcutaneously into Sprague-Dawley rats. Depending on scaffold cross-linking with genipin, glutaraldehyde, and carbodiimide (untreated scaffolds : positive control; bovine pericardium : gold standard), the number of infiltrating fibroblasts, lymphocytes, macrophages, giant cells, and capillaries was determined to quantify the host response after 1, 9, and 30 days. Decellularized esophagus scaffolds were shown to maintain native matrix morphology and extracellular matrix composition. Typical inflammatory reactions were observed in all implants; however, the cellular infiltration was reduced in the genipin group. We conclude that genipin is the most efficient and best tolerated cross-linking agent to attenuate inflammation and to improve the integration of esophageal scaffolds into its surrounding tissue after implantation. Holger Koch, Cora Graneist, Frank Emmrich, Holger Till, Roman Metzger, Heike Aupperle, Katrin Schierle, Ulrich Sack, and Andreas Boldt Copyright © 2012 Holger Koch et al. All rights reserved. CFP and YFP, but Not GFP, Provide Stable Fluorescent Marking of Rat Hepatic Adult Stem Cells Sun, 30 Mar 2008 00:00:00 +0000 The stable expression of reporter genes in adult stem cells (ASCs) has important applications in stem cell biology. The ability to integrate a noncytotoxic, fluorescent reporter gene into the genome of ASCs with the capability to track ASCs and their progeny is particularly desirable for transplantation studies. The use of fluorescent proteins has greatly aided the investigations of protein and cell function on short-time scales. In contrast, the obtainment of stably expressing cell strains with low variability in expression for studies on longer-time scales is often problematic. We show that this difficulty is partly due to the cytotoxicity of a commonly used reporter, green fluorescent protein (GFP). To avoid GFP-specific toxicity effects during attempts to stably mark a rat hepatic ASC strain and, therefore, obtain stable, long-term fluorescent ASCs, we evaluated cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP), in addition to GFP. Although we were unable to derive stable GFP-expressing strains, stable fluorescent clones (up to 140 doublings) expressing either CFP or YFP were established. When fluorescently marked ASCs were induced to produce differentiated progeny cells, stable fluorescence expression was maintained. This property is essential for studies that track fluorescently marked ASCs and their differentiated progeny in transplantation studies. Rouzbeh R. Taghizadeh and James L. Sherley Copyright © 2008 Rouzbeh R. Taghizadeh and James L. Sherley. All rights reserved.