Cell Transplantation 25(10) Abstracts

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Cell Transplantation, Vol. 25, pp. 1723-1746, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691664
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Review

Bowel Radiation Injury: Complexity of the Pathophysiology and Promises of Cell and Tissue Engineering

Lara Moussa, Benoît Usunier, Christelle Demarquay, Marc Benderitter, Radia Tamarat, Alexandra Sémont, and Noëlle Mathieu

Institut de Radioprotection et de Sûreté Nucléaire (IRSN), PRP-HOM/SRBE/LR2I, Fontenay-aux-Roses, France

Ionizing radiation is effective to treat malignant pelvic cancers, but the toxicity to surrounding healthy tissue remains a substantial limitation. Early and late side effects not only limit the escalation of the radiation dose to the tumor but may also be life-threatening in some patients. Numerous preclinical studies determined specific mechanisms induced after irradiation in different compartments of the intestine. This review outlines the complexity of the pathogenesis, highlighting the roles of the epithelial barrier in the vascular network, and the inflammatory microenvironment, which together lead to chronic fibrosis. Despite the large number of pharmacological molecules available, the studies presented in this review provide encouraging proof of concept regarding the use of mesenchymal stromal cell (MSC) therapy to treat radiation-induced intestinal damage. The therapeutic efficacy of MSCs has been demonstrated in animal models and in patients, but an enormous number of cells and multiple injections are needed due to their poor engraftment capacity. Moreover, it has been observed that although MSCs have pleiotropic effects, some intestinal compartments are less restored after a high dose of irradiation. Future research should seek to optimize the efficacy of the injected cells, particularly with regard to extending their life span in the irradiated tissue. Moreover, improving the host microenvironment, combining MSCs with other specific regenerative cells, or introducing new tissue engineering strategies could be tested as methods to treat the severe side effects of pelvic radiotherapy.

Key words: Mesenchymal stromal cells (MSCs); Bowel; Radiotherapy side effects; Regenerative medicine

Received February 15, 2016; final acceptance August 5, 2016. Online prepub date: May 18, 2016. Address correspondence to Noëlle Mathieu, IRSN/PRP-HOM/SRBE/LR2I, 2262 Fontenay-aux-Roses, France. Tel: +33158357640; Fax: +33158358467; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1747-1761, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691655
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Fully Automated Islet Cell Counter (ICC) for the Assessment of Islet Mass, Purity, and Size Distribution by Digital Image Analysis

Peter Buchwald,*† Andres Bernal,‡ Felipe Echeverri,§ Alejandro Tamayo-Garcia,* Elina Linetsky,* and Camillo Ricordi*

*Diabetes Research Institute, Miller School of Medicine, University of Miami, Miami, FL, USA
†Molecular and Cellular Pharmacology, Miller School of Medicine, University of Miami, Miami, FL, USA
‡Bioniko Consulting LLC, Miami, FL, USA
§Biorep Technologies Inc., Miami, FL, USA

For isolated pancreatic islet cell preparations, it is important to be able to reliably assess their mass and quality, and for clinical applications, it is part of the regulatory requirement. Accurate assessment, however, is difficult because islets are spheroid-like cell aggregates of different sizes (<50 to 500 μm) resulting in possible thousandfold differences between the mass contribution of individual particles. The current standard manual counting method that uses size-based group classification is known to be error prone and operator dependent. Digital image analysis (DIA)-based methods can provide less subjective, more reproducible, and better-documented islet cell mass (IEQ) estimates; however, so far, none has become widely accepted or used. Here we present results obtained using a compact, self-contained islet cell counter (ICC3) that includes both the hardware and software needed for automated islet counting and requires minimal operator training and input; hence, it can be easily adapted at any center and could provide a convenient standardized cGMP-compliant IEQ assessment. Using cross-validated sample counting, we found that for most human islet cell preparations, ICC3 provides islet mass (IEQ) estimates that correlate well with those obtained by trained operators using the current manual SOP method (r2 = 0.78, slope = 1.02). Variability and reproducibility are also improved compared to the manual method, and most of the remaining variability (CV = 8.9%) results from the rearrangement of the islet particles due to movement of the sample between counts. Characterization of the size distribution is also important, and the present digitally collected data allow more detailed analysis and coverage of a wider size range. We found again that for human islet cell preparations, a Weibull distribution function provides good description of the particle size.

Key words: Islets of Langerhans; Islet characterization; Digital image analysis (DIA)

Received February 10, 2016; final acceptance July 22, 2016. Online prepub date: May 13, 2016. Address correspondence to Peter Buchwald, Diabetes Research Institute, Miller School of Medicine, University of Miami, 1450 NW 10 Avenue (R-134), Miami, FL 33136, USA. Tel: 305 243-9657; Fax: 305 243-4404; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 25, pp. 1763-1775, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691475
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Plasticity and Aggregation of Juvenile Porcine Islets in Modified Culture: Preliminary Observations

Bradley P. Weegman,*†1 Michael J. Taylor,†‡§1 Simona C. Baicu,‡¶ Kate Mueller,# Timothy D. O’Brien,** John Wilson,†† and Klearchos K. Papas‡‡

*Department of Radiology, Center for Magnetic Resonance Research, University of Minnesota, Minneapolis, MN, USA
†Sylvatica Biotech, LLC, N. Charleston, SC, USA
‡Tissue Testing Technologies, LLC, N. Charleston, SC, USA
§Department of Mechanical Engineering, Carnegie Mellon University, Pittsburgh, PA, USA
¶LifePoint, Inc., Charleston, SC, USA
#Department of Surgery, University of Minnesota, Minneapolis, MN, USA
**Veterinary Population Medicine Department, University of Minnesota, St. Paul, MN, USA
††Wilson Wolf Manufacturing, New Brighton, MN, USA
‡‡Department of Surgery, University of Arizona, Tucson, AZ, USA

Diabetes is a major health problem worldwide, and there is substantial interest in developing xenogeneic islet transplantation as a potential treatment. The potential to relieve the demand on an inadequate supply of human pancreata is dependent upon the efficiency of techniques for isolating and culturing islets from the source pancreata. Porcine islets are favored for xenotransplantation, but mature pigs (>2 years) present logistic and economic challenges, and young pigs (3–6 months) have not yet proven to be an adequate source. In this study, islets were isolated from 20 juvenile porcine pancreata (~3 months; 25 kg Yorkshire pigs) immediately following procurement or after 24 h of hypothermic machine perfusion (HMP) preservation. The resulting islet preparations were characterized using a battery of tests during culture in silicone rubber membrane flasks. Islet biology assessment included oxygen consumption, insulin secretion, histopathology, and in vivo function. Islet yields were highest from HMP-preserved pancreata (2,242 ± 449 IEQ/g). All preparations comprised a high proportion (>90%) of small islets (<100 μm), and purity was on average 63 ± 6%. Morphologically, islets appeared as clusters on day 0, loosely disaggregated structures at day 1, and transitioned to aggregated structures comprising both exocrine and endocrine cells by day 6. Histopathology confirmed both insulin and glucagon staining in cultures and grafts excised after transplantation in mice. Nuclear staining (Ki-67) confirmed mitotic activity consistent with the observed plasticity of these structures. Metabolic integrity was demonstrated by oxygen consumption rates = 175 ± 16 nmol/min/mg DNA, and physiological function was intact by glucose stimulation after 6–8 days in culture. In vivo function was confirmed with blood glucose control achieved in nearly 50% (8/17) of transplants. Preparation and culture of juvenile porcine islets as a source for islet transplantation require specialized conditions. These immature islets undergo plasticity in culture and form fully functional multicellular structures. Further development of this method for culturing immature porcine islets is expected to generate small pancreatic tissue-derived organoids termed “pancreatites,” as a therapeutic product from juvenile pigs for xenotransplantation and diabetes research.

Key words: Islet transplantation; Porcine islets; Islet culture; Diabetes; Xenotransplantation

Received June 3, 2015; final acceptance August 3, 2016. Online prepub date: April 22, 2016.
1These authors provided equal contribution to this work.
Address correspondence to Bradley P. Weegman, Sylvatica Biotech Inc., 2231 Technical Parkway, Charleston, SC 29406, USA. Tel: +1-612-567-2723; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1777-1786, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368915X689721
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Determination of Islet Viability Using a Zinc-Specific Fluorescent Dye and a Semiautomated Assessment Method

Hirotake Komatsu, Keiko Omori, Mounika Parimi, Jeffrey Rawson, Fouad Kandeel, and Yoko Mullen

Division of Developmental and Translational Diabetes and Endocrinology Research, Department of Diabetes and Metabolic Researches, Beckman Research Institute of City of Hope, Duarte, CA, USA Islet transplantation is an effective therapy that allows the achievement of insulin independence in patients with type 1 diabetes (T1D). To ensure successful transplantation, islet viability and function are of great importance. Viability assessments most often use fluorescein diacetate (FDA)/propidium iodide (PI) staining. However, results using this method often do not correlate well with graft function. Because FDA nonspecifically penetrates all cells present in the islet preparation, including islets and contaminating acinar cells, its use often complicates viability assessments of the overall cell population. Furthermore, the manual method for determining viability percentages is highly subjective. Shortcomings of the conventional islet viability assay can be potentially improved by staining cells with Newport Green (NG). NG, is a zinc-specific fluorescent dye that specifically reacts with zinc-rich b cells. Two kinds of NG dyes, NG-DCF and NG-PDX, are currently available. We examined the zinc specificity of these NG dyes and compared NG staining with traditional FDA staining to explore the potential of NG dyes to improve islet viability assessment. Of the two NGs tested, NG-DCF showed the higher specificity toward a b-cell line as well as human islets. NG-DCF accurately identified the islet area, even in low-purity islets, while neither FDA nor NG-PDX did. Although NG-DCF staining required a longer incubation time, the addition of poloxamer F127 and incubation at 37°C allowed viability assessment to take place within 30 min. Unlike FDA/PI staining, NG-DCF/PI staining allowed for islet-specific assessment. We also introduced a semiautomated measurement to determine NG-DCF/PI staining results, which enabled us to obtain objective and reproducible results. NG-DCF/PI staining is easy and reliable, and this method permits highly objective islet-specific viability assessments.

Key words: Islet viability; Newport Green (NG); Zinc-specific fluorescent dye; Semiautomated method; Islet transplantation

Received May 15, 2015; final acceptance May 18, 2016. Online prepub date: October 22, 2015.
Address correspondence to Hirotake Komatsu, Department of Diabetes and Metabolic Research, Beckman Research Institute of City of Hope, 1500 E. Duarte Road, Duarte, CA 91010, USA. Tel: +1-626-359-8111, ext. 60971; Fax: +1-626-301-8136; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1787-1800, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691303
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Easy and Efficient Cell Tagging With Block Copolymer-Based Contrast Agents for Sensitive MRI Detection In Vivo

Barbara Argibay,* Jesse Trekker,†‡ Uwe Himmelreich,‡ Andrés Beiras,§ Antonio Topete,¶1 Pablo Taboada,¶ María Pérez-Mato,* Ramon Iglesias-Rey,* Tomas Sobrino,* José Rivas,# Francisco Campos,* and José Castillo*

*Clinical Neurosciences Research Laboratory, Clinical University Hospital, Health Research Institute of Santiago de Compostela (IDIS), Universidade de Santiago de Compostela, Santiago de Compostela, Spain
†IMEC, Department of Life Science Technology, Leuven, Belgium
‡Biomedical MRI, Department of Imaging and Pathology, KULeuven, Leuven, Belgium
§Department of Morphological Sciences, Universidade de Santiago de Compostela, Santiago de Compostela, Spain
¶Condensed Matter Physics Department, Universidad de Santiago de Compostela, Health Research Institute of Santiago de Compostela (IDIS), Santiago de Compostela, Spain
#Applied Physics Department, Campus Vida, Universidade de Santiago de Compostela, Health Research Institute of Santiago de Compostela (IDIS), Santiago de Compostela, Spain

Superparamagnetic iron oxide nanoparticles (MNPs) together with magnetic resonance imaging (MRI) are the preferred tools for monitoring the fate and biodistribution of administered cells in stem cell therapy studies. Commercial MNPs need transfection agents and long incubation times for sufficient cell labeling and further in vivo cell detection. In this work, we have synthesized MNPs coated with pluronic F127 and tetronic 908, and validated their applicability as contrast agents for MRI cell detection on two different cell types: rat mesenchymal stem cells (MSCs) and multipotent neural progenitor cell line from mice (C17.2). No transfection agent was needed for a complete MNP internalization, and the uptake was only dependent on MNP concentration in medium and limited on the incubation time. By combining in vivo MRI and ex vivo histology microscopy, we have demonstrated the MRI signal detected corresponded exclusively to labeled cells and not to free particles. Pluronic F127- and tetronic 908-coated MNPs represent promising contrast agents for stem cell tracking due to their ease of use in preparation, their efficiency for cell labeling, and their high sensitivity for in vivo cell detection.

Key words: Mesenchymal stem cells (MSCs); Magnetic resonance imaging (MRI); Cell tracking; Superparamagnetic nanoparticles; Intraparenchymal administration

Received December 1, 2015; final acceptance June 23, 2016. Online prepub date: April 18, 2016.
1Current address: Departamento de Fisiología, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Guadalajara 44340. México.
Address correspondence to Prof. José Castillo, Servicio de Neurologia, Hospital Clínico Universitario, Travesa da Choupana s/n, 15706 Santiago de Compostela, Spain. Tel: +34 981951098; Fax: +34 981915098; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it  or Dr. Francisco Campos, Servicio de Neurología, Hospital Clínico Universitario, Travesa da Choupana s/n, 15706 Santiago de Compostela, Spain. Tel: +34 981951098; Fax: +34 981915098; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1801-1817, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X690980
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Combined Use of Mesenchymal Stromal Cell Sheet Transplantation and Local Injection of SDF-1 for Bone Repair in a Rat Nonunion Model

Guangnan Chen,*†1 Tingting Fang,‡1 Yiying Qi,† Xiaofan Yin,* Tuoyu Di,† Gang Feng,† Zhong Lei,† Yuxiang Zhang,† and Zhongming Huang†§¶#

*Department of Orthopedic Surgery, Minhang Hospital, Fudan University, Shanghai, P.R. China
†Department of Orthopedic Surgery, The Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, P.R. China
‡Liver Cancer Institute, Zhongshan Hospital, Shanghai Medical School of Fudan University, Shanghai, P.R. China
§Department of Orthopaedic Surgery, Affiliated Jiangnan Hospital of Zhejiang Chinese Medical University, Hangzhou, P.R. China
¶Department of Orthopaedic Surgery, Xiaoshan Chinese Medical Hospital, Hangzhou, P.R. China
#Institute of Orthopaedics and Traumatology of Zhejiang Province, Hangzhou, P.R. China

Bone nonunion treatments pose a challenge in orthopedics. This study investigated the joint effects of using mesenchymal stem cell (MSC) sheets with local injection of stromal cell-derived factor-1 (SDF-1) on bone formation. In vitro, we found that migration of MSCs was mediated by SDF-1 in a dose-dependent manner. Moreover, stimulation with SDF-1 had no direct effect on the proliferation or osteogenic differentiation of MSCs. Furthermore, the results indicated elevated expression levels of bone morphogenetic protein 2, alkaline phosphatase, osteocalcin, and vascular endothelial growth factor in MSC sheets compared with MSCs cultured in medium. New bone formation in fractures was evaluated by X-ray, micro-computed tomography (micro-CT), hematoxylin and eosin (H&E) staining, Safranin-O staining, and immunohistochemistry in vivo. In the rat bone fracture model, the MSC sheets transplanted into the injured site along with injection of SDF-1 showed significantly more new bone formation within the gap. Moreover, at 8 weeks, complete bone union was obtained in this group. In contrast, the control group showed nonunion of the bone. Our study suggests a new strategy involving the use of MSC sheets with a local injection of SDF-1 for hard tissue reconstruction, such as the healing of nonunions and bone defects.

Key words: Mesenchymal stem cell (MSC) sheet; Stromal cell-derived factor (SDF); Fracture; Mesenchymal stem cells (MSCs)

Received November 17, 2015; final acceptance May 26, 2016. Online prepub date: February 15, 2016.
1These authors provided equal contribution to this work.
Address correspondence to Zhongming Huang, Department of Orthopaedic Surgery, Xiaoshan Chinese Medical Hospital, 152 Yu Cai Road, Hangzhou, Zhejiang 311200, P.R. China. Tel: +8613738193829; Fax: +86057183812172; E‑mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1819-1832, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691420
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Autogenous Mesenchymal Stem Cells From the Vertebral Body Enhance Intervertebral Disc Regeneration via Paracrine Interaction: An In Vitro Pilot Study

Eun-Kyung Shim,*†1 Jung-Seok Lee,*1 Dong-Eun Kim,‡ Seul Ki Kim,* Byung-Joo Jung,§ Eun-Young Choi,* and Chang-Sung Kim*†

*Department of Periodontology, Research Institute for Periodontal Regeneration, College of Dentistry, Yonsei University, Seoul, South Korea
‡Biomedical Research Institute, iBMT, Anyang, South Korea
§Department of Neurosurgery, Naeun Hospital, Anyang, South Korea

†Department of Applied Life Science, BK21 PLUS Project, Yonsei University College of Dentistry, Seoul, South Korea

Several in vivo studies have found that transplanting mesenchymal stem cells (MSCs) into degenerative intervertebral discs (IVDs) leads to regeneration of disc cells. Since the exact underlying mechanisms are not understood, we investigated the mechanisms of action of MSCs in regeneration of degenerative IVDs via paracrine actions. Human MSCs and degenerative disc cells from the same donor vertebrae were directly or indirectly cocultured. The multidifferentiation potential, cell proliferation, collagen synthesis, and mRNA expression levels were assessed. The proliferation rates of MSCs and degenerative disc cells were higher in the coculture system than in the monolayer cultures or in the conditioned medium of each cell type. During coculturing with nucleus pulposus (NP) cells, mRNA expression of the extracellular matrix (ECM) components aggrecan, versican (VCAN), SOX9, and type II and type VI collagen was significantly increased in MSCs, whereas mRNA expression for type V collagen was increased in MSCs cocultured with annulus fibrosus (AF) cells. In addition, the accumulation of total ECM collagen was greater in cocultured degenerative disc cells than in monocultured cells. During coculturing, MSCs downregulated the expression levels of various proinflammatory cytokine genes in degenerative NP [interleukin-1α (IL-1α), IL-1βIL-6, and tumor necrosis factor-α (TNF-α)] and AF cells (IL-1α and IL-6), which are involved in the degradation of ECM molecules. In association with the trophic effect of MSCs on degenerative disc cells, upregulation of growth factor mRNA expression was shown in MSCs cocultured with degenerative NP cells [epidermal growth factor (EGF), insulin-like growth factor-1 (IGF-1), osteogenic protein-1 (OP-1), growth and differentiation factor-7 (GDF-7), and transforming growth factor-β (TGF-β)] or degenerative AF cells (IGF-1OP-1, and GDF-7). In terms of MSC-based clinical approaches to IVD regeneration, implanting MSCs into a degenerative IVD may both stimulate MSC differentiation into an NP- or AF-like phenotype and stimulate the biological activation of degenerative disc cells for self-repair.

Key words: Annulus fibrosis (AF); Intervertebral disc degeneration; Mesenchymal stem cells (MSCs); Nucleus pulposus (NP); Vertebral body

Received March 17, 2015; final acceptance May 19, 2016. Online prepub date: April 5, 2016.
1These authors provided equal contribution to this work.
Address correspondence to Chang-Sung Kim, Department of Periodontology, Research Institute for Periodontal Regeneration, Department of Applied Life Science, BK21 PLUS Project, YonseiUniversity College of Dentistry 50-1, Yonsei-ro, Seodaemun-gu, Seoul 120-752, South Korea. Tel: +82-2-22283186; Fax: +82-2-3920398; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1833-1852, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691312
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
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Effects of the Post-Spinal Cord Injury Microenvironment on the Differentiation Capacity of Human Neural Stem Cells Derived From Induced Pluripotent Stem Cells

Clara López-Serrano,*1 Abel Torres-Espín,*1 Joaquim Hernández,* Ana B. Alvarez-Palomo,† Jordi Requena,† Xavier Gasull,‡§ Michael J. Edel,†¶#2 and Xavier Navarro*

*Group of Neuroplasticity and Regeneration, Institute of Neurosciences, Department of Cell Biology, Physiology and Immunology, Universitat Autònoma de Barcelona, and Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Bellaterra, Spain
†Control of Pluripotency Laboratory, Department of Physiological Sciences I, Faculty of Medicine, Universitat de Barcelona, Barcelona, Spain
‡Neurophysiology Lab, Department of Physiological Sciences I, Faculty of Medicine, Universitat de Barcelona, Barcelona, Spain
§Institut d’Investigacions Biomediques August Pi i Sunyer (IDIBAPS), Barcelona, Spain
¶University of Sydney Medical School, Westmead Children’s Hospital, Division of Pediatrics and Child Health, Westmead, Australia
#School of Anatomy, Physiology & Human Biology, and Centre for Cell Therapy and Regenerative Medicine (CCTRM), University of Western Australia, Nedlands, Australia

Spinal cord injury (SCI) causes loss of neural functions below the level of the lesion due to interruption of spinal pathways and secondary neurodegenerative processes. The transplant of neural stem cells (NSCs) is a promising approach for the repair of SCI. Reprogramming of adult somatic cells into induced pluripotent stem cells (iPSCs) is expected to provide an autologous source of iPSC-derived NSCs, avoiding the immune response as well as ethical issues. However, there is still limited information on the behavior and differentiation pattern of transplanted iPSC-derived NSCs within the damaged spinal cord. We transplanted iPSC-derived NSCs, obtained from adult human somatic cells, into rats at 0 or 7 days after SCI, and evaluated motor-evoked potentials and locomotion of the animals. We histologically analyzed engraftment, proliferation, and differentiation of the iPSC-derived NSCs and the spared tissue in the spinal cords at 7, 21, and 63 days posttransplant. Both transplanted groups showed a late decline in functional recovery compared to vehicle-injected groups. Histological analysis showed proliferation of transplanted cells within the tissue and that cells formed a mass. At the final time point, most grafted cells differentiated to neural and astroglial lineages, but not into oligodendrocytes, while some grafted cells remained undifferentiated and proliferative. The proinflammatory tissue microenviroment of the injured spinal cord induced proliferation of the grafted cells and, therefore, there are possible risks associated with iPSC-derived NSC transplantation. New approaches are needed to promote and guide cell differentiation, as well as reduce their tumorigenicity once the cells are transplanted at the lesion site.

Key words: Spinal cord injury (SCI); Cell therapy; Induced pluripotent stem cells (iPSCs); Neural stem cells (NSCs); Differentiation

Received December 1, 2015; final acceptance May 26, 2016. Online prepub date: April 5, 2016.
1These authors provided equal contribution to this work.
2For correspondence on iPSCs and iNSCs: E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it
Address correspondence to Xavier Navarro Acebes, Departamento de Biologia Celular, Fisiologia e Immunologia, Institut de Neurociències, CIBERNED, Av Can Domenech s/n, Ed. M, Campus UAB, Universitat Autònoma de Barcelona, E01893 Bellaterra, Spain. Tel: +34-935811966; Fax: +34-935812986: E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1853-1861, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691114
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
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Transient Microneedle Insertion Into Hippocampus Triggers Neurogenesis and Decreases Amyloid Burden in a Mouse Model of Alzheimer’s Disease

Shijie Song,*†‡ Xiaoyung Kong,*‡ Vasyl Sava,† Chuanhai Cao,§ Sandra Acosta,‡ Cesar Borlongan,‡ and Juan Sanchez-Ramos*†

*James Haley VA Medical Center, Tampa, FL, USA
†Department of Neurology, Morsani College of Medicine, University of South Florida, Tampa, FL, USA
‡Department of Neurosurgery, Morsani College of Medicine, University of South Florida, Tampa, FL, USA
§Byrd Alzheimer’s Institute, University of South Florida, Tampa, FL, USA

Targeted microlesions of the hippocampus have been reported to enhance neurogenesis in the subgranular zone (SGZ). The potential therapeutic impact of transient insertion of a microneedle was investigated in a mouse model of Alzheimer’s disease (AD). We tested the hypothesis that transient microinjury to the brain elicits cellular responses that mediate beneficial regenerative processes. Brief stereotaxic insertion and removal of a microneedle into the right hippocampus of 14-month-old APP/PS1 mouse brains resulted in (a) stimulation of hippocampal neurogenesis and (b) reduction of amyloid-β plaque number in the CA-1 region. This treatment also resulted in a trend toward improved performance in the radial arm water maze (RAWM). Further studies of fundamental cellular mechanisms of the brain’s response to microinjury will be useful for investigation of potential neuroprotective and deleterious effects of targeted microlesions and deep brain stimulation in AD.

Key words: Neurogenesis; Transgenic APP/PS1 mice; Microneedle stimulation; Alzheimer’s Disease (AD); Amyloid-β (Aβ)

Received November 10, 2015; final acceptance May 18, 2016. Online prepub date: February 18, 2016.
Address correspondence to Juan Sanchez-Ramos, Ph.D., M.D., Department of Neurology, Morsani College of Medicine, University of South Florida, 13220 Laurel Drive (FOB), Tampa, FL 33612, USA. Tel: (813) 974-5841; Fax: (813) 974-8032; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it  or Shijie Song, M.D., Department of Neurology, Morsani College of Medicine, University of South Florida, 13220 Laurel Drive (FOB), Tampa, FL 33612, USA. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1863-1877, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691150
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
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Human Neural Stem Cell Transplantation-Mediated Alteration of Microglial/Macrophage Phenotypes After Traumatic Brain Injury

Junling Gao,* Raymond J. Grill,† Tiffany J. Dunn,* Supinder Bedi,‡ Javier Allende Labastida,* Robert A. Hetz,‡ Hasen Xue,‡ Jason R. Thonhoff,* Douglas S. DeWitt,§ Donald S. Prough,§ Charles S. Cox Jr.,‡ and Ping Wu*¶

*Department of Neuroscience and Cell Biology, University of Texas Medical Branch at Galveston, Galveston, TX, USA
†Department of Integrative Biology and Pharmacology, University of Texas Health Science Center at Houston, Houston, TX, USA
‡Department of Pediatric Surgery, University of Texas Medical School at Houston, Houston, TX, USA
§Department of Anesthesiology, University of Texas Medical Branch at Galveston, TX, USA
¶Beijing Institute for Brain Disorders, Beijing, P.R. China

Neural stem cells (NSCs) promote recovery from brain trauma, but neuronal replacement is unlikely the sole underlying mechanism. We hypothesize that grafted NSCs enhance neural repair at least partially through modulating the host immune response after traumatic brain injury (TBI). C57BL/6 mice were intracerebrally injected with primed human NSCs (hNSCs) or vehicle 24 h after a severe controlled cortical impact injury. Six days after transplantation, brain tissues were collected for Western blot and immunohistochemical analyses. Observations included indicators of microglia/macrophage activation, M1 and M2 phenotypes, axonal injury detected by amyloid precursor protein (APP), lesion size, and the fate of grafted hNSCs. Animals receiving hNSCtransplantation did not show significant decreases of brain lesion volumes compared to transplantation procedures with vehicle alone, but did show significantly reduced injury-dependent accumulation of APP. Furthermore, intracerebral transplantation of hNSCs reduced microglial activation as shown by a diminished intensity of Iba1 immunostaining and a transition of microglia/macrophages toward the M2 anti-inflammatory phenotype. The latter was represented by an increase in the brain M2/M1 ratio and increases of M2 microglial proteins. These phenotypic switches were accompanied by the increased expression of anti-inflammatory interleukin-4 receptor α and decreased proinflammatory interferon-γ receptor β. Finally, grafted hNSCsmainly differentiated into neurons and were phagocytized by either M1 or M2 microglia/macrophages. Thus, intracerebral transplantation of primed hNSCs efficiently leads host microglia/macrophages toward an anti-inflammatory phenotype that presumably contributes to stem cell-mediated neuroprotective effects after severe TBI in mice.

Key words: Neural stem cells (NSCs); Microglia; Traumatic brain injury (TBI); Transplantation; Immunomodulation

Received October 23, 2015; final acceptance May 21, 2016. Online prepub date: March 14, 2016.
Address correspondence to Ping Wu, M.D., Ph.D., Department of Neuroscience and Cell Biology, University of Texas Medical Branch at Galveston, 301 University Boulevard, Research Building 17, Galveston, TX 77555-0620, USA. Tel: 409-772-9858; Fax: 409-747-2200; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it  or Charles S. Cox Jr., M.D., Department of Pediatric Surgery, University of Texas Medical School at Houston, Houston, TX 77030, USA. Tel: 713-500-7307; Fax: 715-500-7296; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1879-1892, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691178
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

Differentiation of Mouse Embryonic Stem Cells Toward Functional Pancreatic β-Cell Surrogates Through Epigenetic Regulation of Pdx1 by Nitric Oxide

Carmen Salguero-Aranda,*†‡§ Rafael Tapia-Limonchi,*¶1 Gladys Margot Cahuana,*¶ Ana Belen Hitos,*† Irene Diaz,*† Abdelkrim Hmadcha,*‡§ Mario Fraga,# Franz Martin,*†¶ BernatSoria,*†‡§ Juan Rigoberto Tejedo,*†‡¶ and Francisco Javier Bedoya*†‡¶

*Andalusian Center for Molecular Biology and Regenerative Medicine, Seville, Spain
†Center for Network Research in Diabetes and Metabolic Diseases (CIBERDEM) Instituto de Salud Carlos III, Madrid, Spain
‡Cell Therapy Network, Madrid (RED-TERCEL), Instituto de Salud Carlos III, Madrid, Spain
§Fundación Progreso y Salud, Seville, Spain
¶Pablo de Olavide University, Seville, Spain
#Department of Epigenetics, Oncologic Institute of Principado of Asturias, Oviedo, Spain

Pancreatic and duodenal homeobox 1 (Pdx1) is a transcription factor that regulates the embryonic development of the pancreas and the differentiation toward β cells. Previously, we have shown that exposure of mouse embryonic stem cells (mESCs) to high concentrations of diethylenetriamine nitric oxide adduct (DETA-NO) triggers differentiation events and promotes the expression of Pdx1. Here we report evidence that Pdx1 expression is associated with release of polycomb repressive complex 2 (PRC2) and P300 from its promoter region. These events are accompanied by epigenetic changes in bivalent markers of histones trimethylated histone H3 lysine 27 (H3K27me3) and H3K4me3, site-specific changes in DNA methylation, and no change in H3 acetylation. On the basis of these findings, we developed a protocol to differentiate mESCs toward insulin-producing cells consisting of sequential exposure to DETA-NO, valproic acid, and P300 inhibitor (C646) to enhance Pdx1 expression and a final maturation step of culture in suspension to form cell aggregates. This small moleculebased protocol succeeds in obtaining cells that express pancreatic β-cell markers such as PDX1, INS1, GCK, and GLUT2 and respond in vitro to high glucose and KCl.

Key words: Embryonic stem cells (ESCs); Nitric oxide (NO); Cell differentiation; Insulin-producing cells; Diabetes

Received March 17, 2015; final acceptance June 1, 2016. Online prepub date: March 14, 2016.
1Current address: Universidad de los Andes, Santiago de Chile, Chile.
Address correspondence to Francisco J. Bedoya, Av. Américo Vespucio s/n-Edif, CABIMER, Parque Cientifico y Tecnológico Cartuja 93, 41092 Seville, Spain. Tel: +34 954 467840; Fax: +34 954 461664; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 25, pp. 1893-1909, 2016
0963-6897/16 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368916X691439
E-ISSN 1555-3892
Copyright © 2016 Cognizant, LLC.
Printed in the USA. All rights reserved

PDGF Facilitates Direct Lineage Reprogramming of Hepatocytes to Functional β-Like Cells Induced by Pdx1 and Ngn3

Fang-Pei Chang,*† Candy Hsin-Hua Cho,† Chia-Rui Shen,‡ Chiao-Yun Chien,§ Ling-Wen Ting,† Hsuan-Shu Lee,§ and Chia-Ning Shen*†¶

*Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan
†Genomics Research Center, Academia Sinica, Taipei, Taiwan
‡Department of Medical Biotechnology and Laboratory Science, Chang Gung University, Tao-Yuan, Taiwan
§Institute of Biotechnology, National Taiwan University, Taipei, Taiwan
¶Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan

Islet transplantation has been proven to be an effective treatment for patients with type 1 diabetes, but a lack of islet donors limits the use of transplantation therapies. It has been previously demonstrated that hepatocytes can be converted into insulin-producing β-like cells by introducing pancreatic transcription factors, indicating that direct hepatocyte reprogramming holds potential as a treatment for diabetes. However, the efficiency at which functional β-cells can be derived from hepatocyte reprogramming remains low. Here we demonstrated that the combination of Pdx1 and Ngn3 can trigger reprogramming of mouse and human liver cells to insulin-producing cells that exhibit the characteristics of pancreatic β-cells. Treatment with PDGF-AA was found to facilitate Pdx1 and Ngn3-induced reprogramming of hepatocytes to β-like cells with the ability to secrete insulin in response to glucose stimulus. Importantly, this reprogramming strategy could be applied to adult mouse primary hepatocytes, and the transplantation of β-like cells derived from primary hepatocyte reprogramming could ameliorate hyperglycemia in diabetic mice. These findings support the possibility of developing transplantation therapies for type 1 diabetes through the use of β-like cells derived from autologous hepatocyte reprogramming.

Key words: Lineage reprogramming; β-Cells; Hepatocyte transdifferentiation; Platelet-derived growth factor (PDGF); Glucagon-like peptide-1 (GLP-1); Type 1 diabetes mellitus

Received March 22, 2016; final acceptance March 22, 2016. Online prepub date: April 5, 2016.
Address correspondence to Chia-Ning Shen, Ph.D., Genomics Research Center, Academia Sinica, 128 Academia Road, Section 2, Nankang, Taipei 115, Taiwan. Tel: +886-2-27899580, ext 301; Fax: +886-2-27899587; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it