Stem Cells International The latest articles from Hindawi © 2017 , Hindawi Limited . All rights reserved. Roles of Mesenchymal Stem Cells in Spinal Cord Injury Sun, 28 May 2017 00:00:00 +0000 Spinal cord injury (SCI) represents one of the most complicated and heterogeneous pathological processes of central nervous system (CNS) impairments, which is still beyond functional regeneration. Transplantation of mesenchymal stem cells (MSCs) has been shown to promote the repair of the injured spinal cord tissues in animal models, and therefore, there is much interest in the clinical use of these cells. However, many questions which are essential to improve the therapy effects remain unanswered. For instance, the functional roles and related molecular regulatory mechanisms of MSCs in vivo are not yet completely determined. It is important for transplanted cells to migrate into the injured tissue, to survive and undergo neural differentiation, or to play neural protection roles by various mechanisms after SCI. In this review, we will focus on some of the recent knowledge about the biological behavior and function of MSCs in SCI. Meanwhile, we highlight the function of biomaterials to direct the behavior of MSCs based on our series of work on silk fibroin biomaterials and attempt to emphasize combinational strategies such as tissue engineering for functional improvement of SCI. Jing Qu and Huanxiang Zhang Copyright © 2017 Jing Qu and Huanxiang Zhang. All rights reserved. PGE2 Promotes the Migration of Mesenchymal Stem Cells through the Activation of FAK and ERK1/2 Pathway Sun, 28 May 2017 00:00:00 +0000 A critical step of MSCs therapy is dependent on its ability to migrate into the sites of injury, so various approaches have been introduced to boost the migratory ability of MSCs. PGE2 is the major prostaglandin generated by COX enzymes and has been implicated in inflammatory response. Evidence indicates that PGE2 can facilitate MSCs migration. Further exploration of the underlying molecular mechanism participating in the promigratory ability of PGE2 may provide a novel strategy to improve MSC transplantation efficacy. In this study, our findings suggested that EP2 prostanoid receptor promotes MSCs migration through activation of FAK and ERK1/2 pathways. Furthermore, MSCs migration induced by PGE2 was blunted by FAK or ERK1/2 inhibitors. EP2-mediated MSCs migration depends on the activation of FAK and ERK1/2. However, the current study did not investigate the migration of MSCs over a blood vessel endothelial barrier. In conclusion, our findings reveal EP2-mediated FAK and ERK1/2 activation was essential for MSCs migration induced by PGE2, indicating that activation of EP2 receptor and FAK/ERK pathways may be a promising strategy to accelerate homing efficiency of MSCs, which in turn enhances therapeutic potential of MSCs transplantation. Xiaomin Lu, Jibin Han, Xiupin Xu, Jingyuan Xu, Ling Liu, Yingzi Huang, Yi Yang, and Haibo Qiu Copyright © 2017 Xiaomin Lu et al. All rights reserved. Cellular Responses in Human Dental Pulp Stem Cells Treated with Three Endodontic Materials Wed, 24 May 2017 05:22:51 +0000 Human dental pulp stem cells (HDPSCs) are of special relevance in future regenerative dental therapies. Characterizing cytotoxicity and genotoxicity produced by endodontic materials is required to evaluate the potential for regeneration of injured tissues in future strategies combining regenerative and root canal therapies. This study explores the cytotoxicity and genotoxicity mediated by oxidative stress of three endodontic materials that are widely used on HDPSCs: a mineral trioxide aggregate (MTA-Angelus white), an epoxy resin sealant (AH-Plus cement), and an MTA-based cement sealer (MTA-Fillapex). Cell viability and cell death rate were assessed by flow cytometry. Oxidative stress was measured by OxyBlot. Levels of antioxidant enzymes were evaluated by Western blot. Genotoxicity was studied by quantifying the expression levels of DNA damage sensors such as ATM and RAD53 genes and DNA damage repair sensors such as RAD51 and PARP-1. Results indicate that AH-Plus increased apoptosis, oxidative stress, and genotoxicity markers in HDPSCs. MTA-Fillapex was the most cytotoxic oxidative stress inductor and genotoxic material for HDPSCs at longer times in preincubated cell culture medium, and MTA-Angelus was less cytotoxic and genotoxic than AH-Plus and MTA-Fillapex at all times assayed. Alejandro Victoria-Escandell, José Santiago Ibañez-Cabellos, Sergio Bañuls-Sánchez de Cutanda, Ester Berenguer-Pascual, Jesús Beltrán-García, Eva García-López, Federico V. Pallardó, José Luis García-Giménez, Antonio Pallardés-Sabater, Ignacio Zarzosa-López, and Manuel Monterde Copyright © 2017 Alejandro Victoria-Escandell et al. All rights reserved. Safety of Allogeneic Canine Adipose Tissue-Derived Mesenchymal Stem Cell Intraspinal Transplantation in Dogs with Chronic Spinal Cord Injury Mon, 22 May 2017 08:32:51 +0000 This is a pilot clinical study primarily designed to assess the feasibility and safety of X-ray-guided percutaneous intraspinal injection of allogeneic canine adipose tissue-derived mesenchymal stem cells in dogs with chronic spinal cord injury. Six dogs with chronic paraplegia (≥six months) were intraparenchymally injected with allogeneic cells in the site of lesion. Cells were obtained from subcutaneous adipose tissue of a healthy dog, cultured to passage 3, labeled with 99mTechnetium, and transplanted into the lesion by percutaneous X-ray-guided injection. Digital X-ray efficiently guided cell injection as 99mTechnetium-labeled cells remained in the injection site for at least 24 hours after transplantation. No adverse effects or complications (infection, neuropathic pain, or worsening of neurological function) were observed during the 16-week follow-up period after transplantation. Three animals improved locomotion as assessed by the Olby scale. One animal walked without support, but no changes in deep pain perception were observed. We conclude that X-ray-guided percutaneous intraspinal transplantation of allogeneic cells in dogs with chronic spinal cord injury is feasible and safe. The efficacy of the treatment will be assessed in a new study involving a larger number of animals. Cláudia Cardoso Maciel Escalhão, Isalira Peroba Ramos, Camila Hochman-Mendez, Tais Hanae Kasai Brunswick, Sergio Augusto Lopes Souza, Bianca Gutfilen, Regina Coeli dos Santos Goldenberg, and Tatiana Coelho-Sampaio Copyright © 2017 Cláudia Cardoso Maciel Escalhão et al. All rights reserved. The Transcription Factor Nrf2 Protects Angiogenic Capacity of Endothelial Colony-Forming Cells in High-Oxygen Radical Stress Conditions Thu, 18 May 2017 09:24:26 +0000 Background. Endothelial colony forming cells (ECFCs) have shown a promise in tissue engineering of vascular constructs, where they act as endothelial progenitor cells. After implantation, ECFCs are likely to be subjected to elevated reactive oxygen species (ROS). The transcription factor Nrf2 regulates the expression of antioxidant enzymes in response to ROS. Methods. Stable knockdown of Nrf2 and Keap1 was achieved by transduction with lentiviral shRNAs; activation of Nrf2 was induced by incubation with sulforaphane (SFN). Expression of Nrf2 target genes was assessed by qPCR, oxidative stress was assessed using CM-DCFDA, and angiogenesis was quantified by scratch-wound and tubule-formation assays. Results. Nrf2 knockdown led to a reduction of antioxidant gene expression and increased ROS. Angiogenesis was disturbed after Nrf2 knockdown even in the absence of ROS. Conversely, angiogenesis was preserved in high ROS conditions after knockdown of Keap1. Preincubation of ECFCs with SFN reduced intracellular ROS in the presence of H2O2 and preserved scratch-wound closure and tubule-formation. Conclusion. The results of this study indicate that Nrf2 plays an important role in the angiogenic capacity of ECFCs, particularly under conditions of increased oxidative stress. Pretreatment of ECFCs with SFN prior to implantation may be a protective strategy for tissue-engineered constructs or cell therapies. Hendrik Gremmels, Olivier G. de Jong, Diënty H. Hazenbrink, Joost O. Fledderus, and Marianne C. Verhaar Copyright © 2017 Hendrik Gremmels et al. All rights reserved. Characterization of Mesenchymal Stem Cell-Like Cells Derived From Human iPSCs via Neural Crest Development and Their Application for Osteochondral Repair Thu, 18 May 2017 07:50:33 +0000 Mesenchymal stem cells (MSCs) derived from induced pluripotent stem cells (iPSCs) are a promising cell source for the repair of skeletal disorders. Recently, neural crest cells (NCCs) were reported to be effective for inducing mesenchymal progenitors, which have potential to differentiate into osteochondral lineages. Our aim was to investigate the feasibility of MSC-like cells originated from iPSCs via NCCs for osteochondral repair. Initially, MSC-like cells derived from iPSC-NCCs (iNCCs) were generated and characterized in vitro. These iNCC-derived MSC-like cells (iNCMSCs) exhibited a homogenous population and potential for osteochondral differentiation. No upregulation of pluripotent markers was detected during culture. Second, we implanted iNCMSC-derived tissue-engineered constructs into rat osteochondral defects without any preinduction for specific differentiation lineages. The implanted cells remained alive at the implanted site, whereas they failed to repair the defects, with only scarce development of osteochondral tissue in vivo. With regard to tumorigenesis, the implanted cells gradually disappeared and no malignant cells were detected throughout the 2-month follow-up. While this study did not show that iNCMSCs have efficacy for repair of osteochondral defects when implanted under undifferentiated conditions, iNCMSCs exhibited good chondrogenic potential in vitro under appropriate conditions. With further optimization, iNCMSCs may be a new source for tissue engineering of cartilage. Ryota Chijimatsu, Makoto Ikeya, Yukihiko Yasui, Yasutoshi Ikeda, Kosuke Ebina, Yu Moriguchi, Kazunori Shimomura, David A Hart, Yoshikawa Hideki, and Nakamura Norimasa Copyright © 2017 Ryota Chijimatsu et al. All rights reserved. Cellular Reprogramming, Genome Editing, and Alternative CRISPR Cas9 Technologies for Precise Gene Therapy of Duchenne Muscular Dystrophy Thu, 18 May 2017 02:45:25 +0000 In the past decade, the development of two innovative technologies, namely, induced pluripotent stem cells (iPSCs) and the CRISPR Cas9 system, has enabled researchers to model diseases derived from patient cells and precisely edit DNA sequences of interest, respectively. In particular, Duchenne muscular dystrophy (DMD) has been an exemplary monogenic disease model for combining these technologies to demonstrate that genome editing can correct genetic mutations in DMD patient-derived iPSCs. DMD is an X-linked genetic disorder caused by mutations that disrupt the open reading frame of the dystrophin gene, which plays a critical role in stabilizing muscle cells during contraction and relaxation. The CRISPR Cas9 system has been shown to be capable of targeting the dystrophin gene and rescuing its expression in in vitro patient-derived iPSCs and in vivo DMD mouse models. In this review, we highlight recent advances made using the CRISPR Cas9 system to correct genetic mutations and discuss how emerging CRISPR technologies and iPSCs in a combined platform can play a role in bringing a therapy for DMD closer to the clinic. Peter Gee, Huaigeng Xu, and Akitsu Hotta Copyright © 2017 Peter Gee et al. All rights reserved. Bone Marrow Aspirate Concentrate-Enhanced Marrow Stimulation of Chondral Defects Thu, 18 May 2017 00:00:00 +0000 Mesenchymal stem cells (MSCs) from bone marrow play a critical role in osteochondral repair. A bone marrow clot forms within the cartilage defect either as a result of marrow stimulation or during the course of the spontaneous repair of osteochondral defects. Mobilized pluripotent MSCs from the subchondral bone migrate into the defect filled with the clot, differentiate into chondrocytes and osteoblasts, and form a repair tissue over time. The additional application of a bone marrow aspirate (BMA) to the procedure of marrow stimulation is thought to enhance cartilage repair as it may provide both an additional cell population capable of chondrogenesis and a source of growth factors stimulating cartilage repair. Moreover, the BMA clot provides a three-dimensional environment, possibly further supporting chondrogenesis and protecting the subchondral bone from structural alterations. The purpose of this review is to bridge the gap in our understanding between the basic science knowledge on MSCs and BMA and the clinical and technical aspects of marrow stimulation-based cartilage repair by examining available data on the role and mechanisms of MSCs and BMA in osteochondral repair. Implications of findings from both translational and clinical studies using BMA concentrate-enhanced marrow stimulation are discussed. Henning Madry, Liang Gao, Hermann Eichler, Patrick Orth, and Magali Cucchiarini Copyright © 2017 Henning Madry et al. All rights reserved. In Utero Stem Cell Transplantation: Potential Therapeutic Application for Muscle Diseases Wed, 17 May 2017 07:35:04 +0000 Muscular dystrophies, myopathies, and traumatic muscle injury and loss encompass a large group of conditions that currently have no cure. Myoblast transplantations have been investigated as potential cures for these conditions for decades. However, current techniques lack the ability to generate cell numbers required to produce any therapeutic benefit. In utero stem cell transplantation into embryos has been studied for many years mainly in the context of hematopoietic cells and has shown to have experimental advantages and therapeutic applications. Moreover, patient-derived cells can be used for experimental transplantation into nonhuman animal embryos via in utero injection as the immune response is absent at such early stages of development. We therefore propose in utero transplantation as a potential method to generate patient-derived humanized skeletal muscle as well as muscle stem cells in animals for therapeutic purposes as well as patient-specific drug screening. Neeladri Chowdhury and Atsushi Asakura Copyright © 2017 Neeladri Chowdhury and Atsushi Asakura. All rights reserved. Environmental Factors That Influence Stem Cell Migration: An “Electric Field” Mon, 15 May 2017 00:00:00 +0000 Environmental Stimulus of Electric Fields on Stem Cell Migration. The movement of cells in response to electric potential gradients is called galvanotaxis. In vivo galvanotaxis, powered by endogenous electric fields (EFs), plays a critical role during development and wound healing. This review aims to provide a perspective on how stem cells transduce EFs into directed migration and an understanding of the current literature relating to the mechanisms by which cells sense and transduce EFs. We will comment on potential EF-based regenerative medicine therapeutics. Stephanie N. Iwasa, Robart Babona-Pilipos, and Cindi M. Morshead Copyright © 2017 Stephanie N. Iwasa et al. All rights reserved. Angiogenic Capacity of Dental Pulp Stem Cell Regulated by SDF-1α-CXCR4 Axis Mon, 15 May 2017 00:00:00 +0000 Previously, the perivascular characteristics of dental pulp stem cells (DPSCs) were reported, which suggested the potential application of DPSCs as perivascular cell source. In this study, we investigated whether DPSCs had angiogenic capacity by coinjection with human umbilical vein endothelial cells (HUVECs) in vivo; in addition, we determined the role of stromal cell-derived factor 1-α (SDF-1α) and C-X-C chemokine receptor type 4 (CXCR4) axis in the mutual interaction between DPSCs and HUVECs. Primarily isolated DPSCs showed mesenchymal stem cell- (MSC-) like characteristics. Moreover, DPSCs expressed perivascular markers such as NG2, α-smooth muscle actin (α-SMA), platelet-derived growth factor receptor β (PDGFRβ), and CD146. In vivo angiogenic capacity of DPSCs was demonstrated by in vivo Matrigel plug assay. We could observe microvessel-like structures in the coinjection of DPSCs and HUVECs at 7 days postinjection. To block SDF-1α and CXCR4 axis between DPSCs and HUVECs, AMD3100, a CXCR4 antagonist, was added into Matrigel plug. No significant microvessel-like structures were observed at 7 days postinjection. In conclusion, DPSCs have perivascular characteristics that contribute to in vivo angiogenesis. The findings of this study have potential applications in neovascularization of engineered tissues and vascular diseases. Hyun Nam, Gee-Hye Kim, Yoon-Kyung Bae, Da-Eun Jeong, Kyeung-Min Joo, Kyunghoon Lee, and Sun-Ho Lee Copyright © 2017 Hyun Nam et al. All rights reserved. Delayed Mesoderm and Erythroid Differentiation of Murine Embryonic Stem Cells in the Absence of the Transcriptional Regulator FUBP1 Mon, 15 May 2017 00:00:00 +0000 The transcriptional regulator far upstream binding protein 1 (FUBP1) is essential for fetal and adult hematopoietic stem cell (HSC) self-renewal, and the constitutive absence of FUBP1 activity during early development leads to embryonic lethality in homozygous mutant mice. To investigate the role of FUBP1 in murine embryonic stem cells (ESCs) and in particular during differentiation into hematopoietic lineages, we generated Fubp1 knockout (KO) ESC clones using CRISPR/Cas9 technology. Although FUBP1 is expressed in undifferentiated ESCs and during spontaneous differentiation following aggregation into embryoid bodies (EBs), absence of FUBP1 did not affect ESC maintenance. Interestingly, we observed a delayed differentiation of FUBP1-deficient ESCs into the mesoderm germ layer, as indicated by impaired expression of several mesoderm markers including Brachyury at an early time point of ESC differentiation upon aggregation to EBs. Coculture experiments with OP9 cells in the presence of erythropoietin revealed a diminished differentiation capacity of Fubp1 KO ESCs into the erythroid lineage. Our data showed that FUBP1 is important for the onset of mesoderm differentiation and maturation of hematopoietic progenitor cells into the erythroid lineage, a finding that is supported by the phenotype of FUBP1-deficient mice. Josephine Wesely, Marlene Steiner, Frank Schnütgen, Manuel Kaulich, Michael A. Rieger, and Martin Zörnig Copyright © 2017 Josephine Wesely et al. All rights reserved. Human Bone Marrow Mesenchymal Stem/Stromal Cells Preserve Their Immunomodulatory and Chemotactic Properties When Expanded in a Human Plasma Derived Xeno-Free Medium Sun, 14 May 2017 09:54:10 +0000 Due to their immunomodulatory and chemotactic properties, hMSC are being explored to treat immune-related diseases. For their use in human therapies, it is necessary to culture hMSC in xeno-free conditions. In this study, the impact that a xeno-free medium based on a human plasma derivate has on these properties was analysed. Bone marrow-derived hMSC preserved their immunosuppressive and immunostimulatory properties, as observed with in vitro assays with hMSC cocultured with mixed leukocyte reactions or with mitogen-stimulated leukocytes. Moreover, hMSC expanded in xeno-free medium were recruited by macrophages in both migration and invasion assays, which indicates that the cells maintained their chemotactic properties. These data suggest that xeno-free expanded hMSC preserved their immunomodulatory and chemotactic properties, indicating that the described xeno-free medium composition is a potential candidate to culture and expand hMSC for human cell therapies. A. Blázquez-Prunera, C. R. Almeida, and M. A. Barbosa Copyright © 2017 A. Blázquez-Prunera et al. All rights reserved. hTERT-Immortalized Bone Mesenchymal Stromal Cells Expressing Rat Galanin via a Single Tetracycline-Inducible Lentivirus System Thu, 11 May 2017 02:30:33 +0000 The use of human telomerase reverse transcriptase-immortalized bone marrow mesenchymal stromal cells (hTERT-BMSCs) as vehicles to deliver antinociceptive galanin (GAL) molecules into pain-processing centers represents a novel cell therapy strategy for pain management. Here, an hTERT-BMSCs/Tet-on/GAL cell line was constructed using a single Tet-on-inducible lentivirus system, and subsequent experiments demonstrated that the secretion of rat GAL from hTERT-BMSCs/Tet-on/GAL was switched on and off under the control of an inducer in a dose-dependent manner. The construction of this cell line is the first promising step in the regulation of GAL secretion from hTERT-immortalized BMSCs, and the potential application of this system may provide a stem cell-based research platform for pain. Ke An, Hui-ping Liu, Xiao-long Zhong, David Y. B. Deng, Jing-jun Zhang, and Zhi-heng Liu Copyright © 2017 Ke An et al. All rights reserved. Cytokine, Chemokine, and Growth Factor Profile Characterization of Undifferentiated and Osteoinduced Human Adipose-Derived Stem Cells Wed, 10 May 2017 00:00:00 +0000 Bone is the second most manipulated tissue after blood. Adipose-derived stem cells (ASCs) may become a convenient source of MSC for bone regenerative protocols. Surprisingly, little is known about the most significant biomolecules these cells produce and release after being osteoinduced. Therefore, the present study aimed at dosing 13 candidates chosen among the most representative cytokines, chemokines, and growth factors within the conditioned media of osteodifferentiated and undifferentiated ASCs. Two acknowledged osteoblastic cell models, that is, MG-63 and SaOs-2 cells, were compared. Notably, IL-6, IL-8, MCP-1, and VEGF were highly produced and detectable in ASCs. In addition, while IL-6 and IL-8 seemed to be significantly induced by the osteogenic medium, no such effect was seen for MCP-1 and VEGF. Overall SaOS-2 had a poor expression profile, which may be consistent with the more differentiated phenotype of SaOs-2 compared to ASCs and MG-63. Instead, in maintaining medium, MG-63 displayed a very rich production of IL-12, MCP-1, IP-10, and VEGF, which were significantly reduced in osteogenic conditions, with the only exception of MCP-1. The high expression of MCP-1 and VEGF, even after the osteogenic commitment, may support the usage of ASCs in bone regenerative protocols by recruiting both osteoblasts and osteoclasts of the host. F. Mussano, T. Genova, M. Corsalini, G. Schierano, F. Pettini, D. Di Venere, and S. Carossa Copyright © 2017 F. Mussano et al. All rights reserved. Human Adipose-Derived Stem Cells Exhibit Enhanced Proliferative Capacity and Retain Multipotency Longer than Donor-Matched Bone Marrow Mesenchymal Stem Cells during Expansion In Vitro Wed, 03 May 2017 08:53:15 +0000 Bone marrow-derived mesenchymal stem cells (MSCs) and adipose-derived multipotent/mesenchymal stem cells (ASCs) have been proposed as the ideal cell types for a range of musculoskeletal tissue engineering and regenerative medicine therapies. However, extensive in vitro expansion is required to generate sufficient cells for clinical application and previous studies have demonstrated differences in the proliferative capacity and the impact of expansion on differentiation capacity of both MSCs and ASCs. Significantly, these studies routinely use cells from different donors, making direct comparisons difficult. Importantly, this study directly compared the proliferative capacity and multipotency of human MSCs and ASCs from the same donors to determine how each cell type was affected by in vitro expansion. The study identified that ASCs were able to proliferate faster and undergo greater population doublings than donor-matched MSCs and that senescence was primarily driven via telomere shortening and upregulation of p16ink4a. Both donor-matched MSCs and ASCs were capable of trilineage differentiation early in cultures; however, while differentiation capacity diminished with time in culture, ASCs retained enhanced capacity compared to MSCs. These findings suggest that ASCs may be the most appropriate cell type for musculoskeletal tissue engineering and regenerative medicine therapies due to their enhanced in vitro expansion capacity and limited loss of differentiation potential. Kimberley L. Burrow, Judith A. Hoyland, and Stephen M. Richardson Copyright © 2017 Kimberley L. Burrow et al. All rights reserved. Inhibition of Lysyl Oxidases Impairs Migration and Angiogenic Properties of Tumor-Associated Pericytes Wed, 03 May 2017 04:02:08 +0000 Pericytes are important cellular components of the tumor microenviroment with established roles in angiogenesis and metastasis. These two cancer hallmarks are modulated by enzymes of the LOX family, but thus far, information about LOX relevance in tumor-associated pericytes is lacking. Here, we performed a comparative characterization of normal and tumoral pericytes and report for the first time the modulatory effects of LOX enzymes on activated pericyte properties. Tumoral pericytes isolated from childhood ependymoma and neuroblastoma specimens displayed angiogenic properties in vitro and expressed typical markers, including CD146, NG2, and PDGFRβ. Expression of all LOX family members could be detected in both normal and tumor-associated pericytes. In most pericyte samples, LOXL3 was the family member displaying the highest transcript levels. Inhibition of LOX/LOXL activity with the inhibitor β-aminopropionitrile (βAPN) significantly reduced migration of pericytes, while proliferation rates were kept unaltered. Formation of tube-like structures in vitro by pericytes was also significantly impaired upon inhibition of LOX/LOXL activity with βAPN, which induced more prominent effects in tumor-associated pericytes. These findings reveal a novel involvement of the LOX family of enzymes in migration and angiogenic properties of pericytes, with implications in tumor development and in therapeutic targeting tumor microenvironment constituents. Aline Lopes Ribeiro, Carolini Kaid, Patrícia B. G. Silva, Beatriz A. Cortez, and Oswaldo Keith Okamoto Copyright © 2017 Aline Lopes Ribeiro et al. All rights reserved. In Vivo Immunogenic Response to Allogeneic Mesenchymal Stem Cells and the Role of Preactivated Mesenchymal Stem Cells Cotransplanted with Allogeneic Islets Wed, 03 May 2017 00:00:00 +0000 Mesenchymal stem cells (MSCs) are multipotent cells capable of differentiating into cells from the mesenchymal lineage. The hypoimmunogenic characteristic of MSCs has encouraged studies using allogeneic MSCs for the treatment of autoimmune diseases and inflammatory conditions. Promising preclinical results and the safety of allogeneic MSC transplantation have created the possibility of “off-the-shelf” clinical application of allogeneic cells. This study has aimed to evaluate the survival of untreated and IFN-γ- and TNF-α-treated (preactivated) allogeneic MSCs transplanted under the kidney capsule of immunocompetent mice together with the role of preactivated MSCs after cotransplantation with allogeneic islets. The preactivation of MSCs upregulated the gene expression of anti-inflammatory molecules and also enhanced their immunomodulatory capacity in vitro. In vivo, allogeneic MSCs provoked an immunogenic response, with the infiltration of inflammatory cells at the transplant site and full graft rejection in both the untreated and preactivated groups. Allogeneic islets cotransplanted with preactivated MSCs prolonged graft survival for about 6 days, compared with islet alone. The present results corroborate the hypothesis that allogeneic MSCs are not immune-privileged and that after playing their therapeutic role they are rejected. Strategies that reduce allogeneic MSC immunogenicity can potentially prolong their in vivo persistence and improve the therapeutic effects. Régis Linhares Oliveira, Pedro Cesar Chagastelles, Patrícia Sesterheim, and Patricia Pranke Copyright © 2017 Régis Linhares Oliveira et al. All rights reserved. Bone Regeneration Induced by Bone Porcine Block with Bone Marrow Stromal Stem Cells in a Minipig Model of Mandibular “Critical Size” Defect Tue, 02 May 2017 09:12:48 +0000 Introduction. Adding stem cells to biodegradable scaffolds to enhance bone regeneration is a valuable option. Different kinds of stem cells with osteoblastic activity were tested, such as bone marrow stromal stem cells (BMSSCs). Aim. To assess a correct protocol for osteogenic stem cell differentiation, so BMSSCs were seeded on a bone porcine block (BPB). Materials and Methods. Bone marrow from six minipigs was extracted from tibiae and humeri and treated to isolate BMSSCs. After seeding on BPB, critical-size defects were created on each mandible of the minipigs and implanted with BPB and BPB/BMSSCs. After three months, histomorphometric analysis was performed. Results. Histomorphometric analysis provided percentages of the three groups. Tissues present in control defects were 23 ± 2% lamellar bone, 28 ± 1% woven bone, and 56 ± 4% marrow spaces; in BPB defects were 20 ± 5% BPB, 32 ± 2% lamellar bone, 24 ± 1% woven bone, and 28 ± 2% marrow spaces; in BPB/BMSSCs defects were 17 ± 4% BPB/BMSSCs, 42 ± 2% lamellar bone, 12 ± 1% woven bone, and 22 ± 3% marrow spaces. Conclusion. BPB used as a scaffold to induce bone regeneration may benefit from the addition of BDPSCs in the tissue-engineered constructs. Antonio Scarano, Vito Crincoli, Adriana Di Benedetto, Valerio Cozzolino, Felice Lorusso, Michele Podaliri Vulpiani, Maria Grano, Zamira Kalemaj, Giorgio Mori, and Felice Roberto Grassi Copyright © 2017 Antonio Scarano et al. All rights reserved. Quantitative Assessment of Optimal Bone Marrow Site for the Isolation of Porcine Mesenchymal Stem Cells Sun, 30 Apr 2017 00:00:00 +0000 Background. One of the most plentiful sources for MSCs is the bone marrow; however, it is unknown whether MSC yield differs among different bone marrow sites. In this study, we quantified cellular yield and evaluated resident MSC population from five bone marrow sites in the porcine model. In addition, we assessed the feasibility of a commercially available platelet concentrator (Magellan® MAR01™ Arteriocyte Medical Systems, Hopkinton, MA) as a bedside stem cell concentration device. Methods. Analyses of bone marrow aspirate (BMA) and concentrated bone marrow aspirate (cBMA) included bone marrow volume, platelet and nucleated cell yield, colony-forming unit fibroblast (CFU-F) number, flow cytometry, and assessment of differentiation potential. Results. Following processing, the concentration of platelets and nucleated cells significantly increased but was not significantly different between sites. The iliac crest had significantly less bone marrow volume; however, it yielded significantly more CFUs compared to the other bone marrow sites. Culture-expanded cells from all tested sites expressed high levels of MSC surface markers and demonstrated adipogenic and osteogenic differentiation potential. Conclusions. All anatomical bone marrow sites contained MSCs, but the iliac crest was the most abundant source of MSCs. Additionally, the Magellan can function effectively as a bedside stem cell concentrator. J. S. McDaniel, B. Antebi, M. Pilia, B. J. Hurtgen, S. Belenkiy, C. Necsoiu, L. C. Cancio, C. R. Rathbone, and A. I. Batchinsky Copyright © 2017 J. S. McDaniel et al. All rights reserved. Skeletal Muscle Cell Induction from Pluripotent Stem Cells Wed, 26 Apr 2017 00:00:00 +0000 Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have the potential to differentiate into various types of cells including skeletal muscle cells. The approach of converting ESCs/iPSCs into skeletal muscle cells offers hope for patients afflicted with the skeletal muscle diseases such as the Duchenne muscular dystrophy (DMD). Patient-derived iPSCs are an especially ideal cell source to obtain an unlimited number of myogenic cells that escape immune rejection after engraftment. Currently, there are several approaches to induce differentiation of ESCs and iPSCs to skeletal muscle. A key to the generation of skeletal muscle cells from ESCs/iPSCs is the mimicking of embryonic mesodermal induction followed by myogenic induction. Thus, current approaches of skeletal muscle cell induction of ESCs/iPSCs utilize techniques including overexpression of myogenic transcription factors such as MyoD or Pax3, using small molecules to induce mesodermal cells followed by myogenic progenitor cells, and utilizing epigenetic myogenic memory existing in muscle cell-derived iPSCs. This review summarizes the current methods used in myogenic differentiation and highlights areas of recent improvement. Yusaku Kodaka, Gemachu Rabu, and Atsushi Asakura Copyright © 2017 Yusaku Kodaka et al. All rights reserved. Pulsed Electromagnetic Field Regulates MicroRNA 21 Expression to Activate TGF-β Signaling in Human Bone Marrow Stromal Cells to Enhance Osteoblast Differentiation Sun, 23 Apr 2017 00:00:00 +0000 Pulsed electromagnetic fields (PEMFs) have been documented to promote bone fracture healing in nonunions and increase lumbar spinal fusion rates. However, the molecular mechanisms by which PEMF stimulates differentiation of human bone marrow stromal cells (hBMSCs) into osteoblasts are not well understood. In this study the PEMF effects on hBMSCs were studied by microarray analysis. PEMF stimulation of hBMSCs’ cell numbers mainly affected genes of cell cycle regulation, cell structure, and growth receptors or kinase pathways. In the differentiation and mineralization stages, PEMF regulated preosteoblast gene expression and notably, the transforming growth factor-beta (TGF-β) signaling pathway and microRNA 21 (miR21) were most highly regulated. PEMF stimulated activation of Smad2 and miR21-5p expression in differentiated osteoblasts, and TGF-β signaling was essential for PEMF stimulation of alkaline phosphatase mRNA expression. Smad7, an antagonist of the TGF-β signaling pathway, was found to be miR21-5p’s putative target gene and PEMF caused a decrease in Smad7 expression. Expression of Runx2 was increased by PEMF treatment and the miR21-5p inhibitor prevented the PEMF stimulation of Runx2 expression in differentiating cells. Thus, PEMF could mediate its effects on bone metabolism by activation of the TGF-β signaling pathway and stimulation of expression of miR21-5p in hBMSCs. Nagarajan Selvamurugan, Zhiming He, Daniel Rifkin, Branka Dabovic, and Nicola C. Partridge Copyright © 2017 Nagarajan Selvamurugan et al. All rights reserved. Resveratrol Induces Differentiation of Human Umbilical Cord Mesenchymal Stem Cells into Neuron-Like Cells Thu, 20 Apr 2017 09:45:45 +0000 Objective. Human umbilical cord mesenchymal stem cells (hUC-MSCs) potentially differentiate to various types of cells including neuron-like cells. The natural polyphenol resveratrol benefits patients with many diseases including ischemic brain injury. We hypothesize that resveratrol induces differentiation of hUC-MSCs into neuron-like cells. Methods. Flow cytometry was used to determine the surface antigens in different stage of hUC-MSCs (P2, P5, and P10). Nestin, neuron-specific enolase (NSE), and glial fibrillary acidic protein (GFAP) were detected by immunocytochemistry, Western blotting, and real time RT-PCT. The cultured hUC-MSCs were treated with resveratrol at different concentrations (0, 7.5, 15.0, and 30.0 mg/L). Nestin, GFAP, and NSE protein and mRNA were measured at posttreatment time points of 2 h, 4 h, 6 h, 12 h, and 24 h. Results. Neuron-like cells were found in hUC-MSCs treated by resveratrol at concentrations of 15.0 and 30.0 mg/L, but not in hUC-MSCs treated with vehicle and 7.5 mg/L resveratrol. Furthermore, immunocytochemical staining revealed that nestin and NSE immunoreactivities were positive in resveratrol-treated hUC-MSCs at concentrations of 15.0 and 30.0 mg/L. Resveratrol treatment significantly increased nestin and NSE protein and mRNA levels 4 h after the treatment. However, resveratrol treatment did not change GFAP immunoreactivities and protein and mRNA expression levels in cultured hUC-MSCs. Conclusions. Taken together, resveratrol treatment induces a differentiation of hUC-MSCs into neuron-like cells at relatively high concentrations. Li Guo, Liang Wang, Li Wang, Shi Yun-peng, Jing-jing Zhou, Zongmao Zhao, and De-Pei Li Copyright © 2017 Li Guo et al. All rights reserved. Vitamin C in Stem Cell Biology: Impact on Extracellular Matrix Homeostasis and Epigenetics Thu, 20 Apr 2017 06:09:11 +0000 Transcription factors and signaling molecules are well-known regulators of stem cell identity and behavior; however, increasing evidence indicates that environmental cues contribute to this complex network of stimuli, acting as crucial determinants of stem cell fate. L-Ascorbic acid (vitamin C (VitC)) has gained growing interest for its multiple functions and mechanisms of action, contributing to the homeostasis of normal tissues and organs as well as to tissue regeneration. Here, we review the main functions of VitC and its effects on stem cells, focusing on its activity as cofactor of Fe+2/αKG dioxygenases, which regulate the epigenetic signatures, the redox status, and the extracellular matrix (ECM) composition, depending on the enzymes’ subcellular localization. Acting as cofactor of collagen prolyl hydroxylases in the endoplasmic reticulum, VitC regulates ECM/collagen homeostasis and plays a key role in the differentiation of mesenchymal stem cells towards osteoblasts, chondrocytes, and tendons. In the nucleus, VitC enhances the activity of DNA and histone demethylases, improving somatic cell reprogramming and pushing embryonic stem cell towards the naive pluripotent state. The broad spectrum of actions of VitC highlights its relevance for stem cell biology in both physiology and disease. Cristina D'Aniello, Federica Cermola, Eduardo Jorge Patriarca, and Gabriella Minchiotti Copyright © 2017 Cristina D'Aniello et al. All rights reserved. Mitochondrial Dynamics: In Cell Reprogramming as It Is in Cancer Tue, 18 Apr 2017 00:00:00 +0000 Somatic cells can be reprogrammed into a pluripotent cellular state similar to that of embryonic stem cells. Given the significant physiological differences between the somatic and pluripotent cells, cell reprogramming is associated with a profound reorganization of the somatic phenotype at all levels. The remodeling of mitochondrial morphology is one of these dramatic changes that somatic cells have to undertake during cell reprogramming. Somatic cells transform their tubular and interconnected mitochondrial network to the fragmented and isolated organelles found in pluripotent stem cells early during cell reprogramming. Accordingly, mitochondrial fission, the process whereby the mitochondria divide, plays an important role in the cell reprogramming process. Here, we present an overview of the importance of mitochondrial fission in both cell reprogramming and cellular transformation. Javier Prieto and Josema Torres Copyright © 2017 Javier Prieto and Josema Torres. All rights reserved. Insulin Promotes the Proliferation of Human Umbilical Cord Matrix-Derived Mesenchymal Stem Cells by Activating the Akt-Cyclin D1 Axis Tue, 18 Apr 2017 00:00:00 +0000 Background. The functions of insulin in mesenchymal stem cells (MSC) remain poorly understood. Methods. MSC from human umbilical cord matrix (UCM) cultured in serum-free media (SFM) with or without insulin were subjected to various molecular biological analyses to determine their proliferation and growth states, expression levels of Akt-cyclin D1 signaling molecules, and in vitro differentiation capacities. Results. Insulin accelerated the G1-S cell cycle progression of UCM-MSC and significantly stimulated their proliferation and growth in SFM. The pro-proliferative action of insulin was associated with augmented cyclin D1 and phosphorylated Akt expression levels. Akt inactivation remarkably abrogated insulin-induced increases in cyclin D1 expression and cell proliferation, indicating that insulin enhances the proliferation of UCM-MSC via acceleration of the G1-S transition mediated by the Akt-cyclin D1 pathway. Additionally, the UCM-MSC propagated in SFM supplemented with insulin exhibited similar specific surface antigen profiles and differentiation capacities as those generated in conventional media containing fetal bovine serum. Conclusions. These findings suggest that insulin acts solely to promote UCM-MSC proliferation without affecting their immunophenotype and differentiation potentials and thus have important implications for utilizing insulin to expand clinical-grade MSC in vitro. Peng Li, Jinsong Wei, Xiang Gao, Bo Wei, Hao Lin, Rui Huang, Yanru Niu, Kyu Lim, Kaipeng Jing, and Jiaqi Chu Copyright © 2017 Peng Li et al. All rights reserved. Wnt5a Signaling in Normal and Cancer Stem Cells Wed, 12 Apr 2017 00:00:00 +0000 Wnt5a is involved in activating several noncanonical Wnt signaling pathways, which can inhibit or activate canonical Wnt/β-catenin signaling pathway in a receptor context-dependent manner. Wnt5a signaling is critical for regulating normal developmental processes, including stem cell self-renewal, proliferation, differentiation, migration, adhesion, and polarity. Moreover, the aberrant activation or inhibition of Wnt5a signaling is emerging as an important event in cancer progression, exerting both oncogenic and tumor suppressive effects. Recent studies show the involvement of Wnt5a signaling in regulating normal and cancer stem cell self-renewal, cancer cell proliferation, migration, and invasion. In this article, we review recent findings regarding the molecular mechanisms and roles of Wnt5a signaling in stem cells in embryogenesis and in the normal or neoplastic breast or ovary, highlighting that Wnt5a may have different effects on target cells depending on the surface receptors expressed by the target cell. Yan Zhou, Thomas J. Kipps, and Suping Zhang Copyright © 2017 Yan Zhou et al. All rights reserved. Induction of Pluripotent Stem Cells from a Manifesting Carrier of Duchenne Muscular Dystrophy and Characterization of Their X-Inactivation Status Wed, 12 Apr 2017 00:00:00 +0000 Three to eight percent of female carriers of Duchenne muscular dystrophy (DMD) develop dystrophic symptoms ranging from mild muscle weakness to a rapidly progressive DMD-like muscular dystrophy due to skewed inactivation of X chromosomes during early development. Here, we generated human induced pluripotent stem cells (hiPSCs) from a manifesting female carrier using retroviral or Sendai viral (SeV) vectors and determined their X-inactivation status. Although manifesting carrier-derived iPS cells showed normal expression of human embryonic stem cell markers and formed well-differentiated teratomas in vivo, many hiPS clones showed bi-allelic expression of the androgen receptor (AR) gene and loss of X-inactivation-specific transcript and trimethyl-histone H3 (Lys27) signals on X chromosomes, suggesting that both X chromosomes of the hiPS cells are in an active state. Importantly, normal dystrophin was expressed in multinucleated myotubes differentiated from a manifesting carrier of DMD-hiPS cells with XaXa pattern. AR transcripts were also equally transcribed from both alleles in induced myotubes. Our results indicated that the inactivated X chromosome in the patient’s fibroblasts was activated during reprogramming, and XCI occurred randomly during differentiation. Yuko Miyagoe-Suzuki, Takashi Nishiyama, Miho Nakamura, Asako Narita, Fusako Takemura, Satoru Masuda, Narihiro Minami, Kumiko Murayama, Hirofumi Komaki, Yu-ichi Goto, and Shin’ichi Takeda Copyright © 2017 Yuko Miyagoe-Suzuki et al. All rights reserved. Therapeutic Effect and Location of GFP-Labeled Placental Mesenchymal Stem Cells on Hepatic Fibrosis in Rats Wed, 12 Apr 2017 00:00:00 +0000 Background. Liver fibrosis is a chronic progressive liver disease, but no established effective treatment exists except for liver transplantation. The present study was designed to investigate the effect of human placenta mesenchymal stem cells (hPMSCs) expressing green fluorescent protein (GFP) on carbon tetrachloride- (CCl4-) induced liver fibrosis in rats. Methods. Liver fibrosis was induced by subcutaneous injection with CCl4; hPMSCs were directly transplanted into rats through the caudal vein. The therapeutic efficacy of hPMSCs on liver fibrosis was measured by liver function tests, liver elastography, histopathology, Masson’s trichrome and Sirius red staining, and immunohistochemical studies. The expression levels of fibrotic markers, transforming growth factor β1 (TGF-β1) and α-smooth muscle actin (α-SMA), were assessed using real-time polymerase chain reaction. Results. We demonstrated that liver fibrosis was significantly dampened in the hPMSC transplantation group according to the Laennec fibrosis scoring system and histological data. The Sirius red-stained collagen area and the elastography score were significantly reduced in the hPMSC-treated group. Meanwhile, hPMSC administration significantly decreased TGF-β1 and α-SMA expression and enhanced liver functions in CCl4-induced fibrotic rats. Conclusion. This study indicates that transplantation of hPMSCs could repair liver fibrosis induced by CCl4 in rats, which may serve as a valuable therapeutic approach to treat liver diseases. Jiong Yu, Guangshu Hao, Dan Wang, Jingqi Liu, Xiaotian Dong, Yanni Sun, Qiaoling Pan, Yang Li, Xiaowei Shi, Lanjuan Li, and Hongcui Cao Copyright © 2017 Jiong Yu et al. All rights reserved. Immunoprofiling of Adult-Derived Human Liver Stem/Progenitor Cells: Impact of Hepatogenic Differentiation and Inflammation Tue, 11 Apr 2017 00:00:00 +0000 Adult-derived human liver stem/progenitor cells (ADHLSCs) are, nowadays, developed as therapeutic medicinal product for the treatment of liver defects. In this study, the impact of hepatogenic differentiation and inflammation priming on the ADHLSCs’ immune profile was assessed in vitro and compared to that of mature hepatocytes. The constitutive immunological profile of ADHLSCs was greatly different from that of hepatocytes. Differences in the expression of the stromal markers CD90 and CD105, adhesion molecules CD44 and CD49e, immunoregulatory molecules CD73 and HO-1, and NK ligands CD112 and CD155 were noted. While they globally preserved their immunological profile in comparison to undifferentiated counterparts, differentiated ADHLSCs showed a significant downregulation of CD200 expression as in hepatocytes. This was mainly induced by signals issued from EGF and OSM. On the other hand, the impact of inflammation was quite similar for all studied cell populations with an increased expression level of CD54 and CD106 and induction of that of CD40 and CD274. In conclusion, our immune profiling study suggests CD200 as a key factor in regulating the immunobiology of differentiated ADHLSCs. A better understanding of the molecular and physiological events related to such marker could help in designing the optimal conditions for an efficient therapeutic use of ADHLSCs. Hoda El-Kehdy, Camillo Sargiacomo, Mohammad Fayyad-Kazan, Hussein Fayyad-Kazan, Catherine Lombard, Laurence Lagneaux, Etienne Sokal, Mehdi Najar, and Mustapha Najimi Copyright © 2017 Hoda El-Kehdy et al. All rights reserved.