Cell Transplantation 19(12) Abstracts

Return to Cell Transplantation main page>

Cell Transplantation, Vol. 19, pp. 1511–1522, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X514279
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Review

Bone Marrow and Pancreatic Islets: An Old Story With New Perspectives

Fabio Ciceri* and Lorenzo Piemonti†

*Haematology and BMT Unit, San Raffaele Scientific Institute, Milan, Italy
†San Raffaele Diabetes Research Institute (HSR-DRI), San Raffaele Scientific Institute, Milan, Italy

In the past years, in the field of β-cell replacement for diabetes therapy, the easy availability of bone marrow (BM) and the widely consolidated clinical experience in the field of hematology have contributed to the development of strategy to achieve donor-specific transplantation tolerance. Recently, the potential role of BM in diabetes therapy has been reassessed from a different point of view. Diverse groups investigated the contribution of BM cells to β-cell replacement as direct differentiation into insulin-producing cells. More importantly, while direct differentiation is highly unlikely, a wide array of experimental evidences indicates that cells of BM origin are capable of facilitating the survival or the endogenous regeneration of β-cells through an as yet well-defined regeneration process. These new experimental in vitro and in vivo data will expand in the near future the clinical trials involving BM or BM-derived cells to cure both type 1 and type 2 diabetes in humans. In this review we recapitulate the history of use of BM in diabetes therapy and we provide clinically relevant actual information about the participation of BM and BM-derived stem cells in islet cell regeneration processes. Furthermore, new aspects such as employing BM as “feeder tissue” for pancreatic islets and new clinical use of BM in diabetes therapy are discussed.

Key words: Bone marrow; Bone marrow-derived cells; Pancreatic islets; Diabetes

Address correspondence to Lorenzo Piemonti, Diabetes Research Institute, San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy. Tel: 39 02 26432706; Fax: 39 02 26432871; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1523–1535, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X515872
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Review

Pancreatic Islet Culture and Preservation Strategies: Advances, Challenges, and Future Outlook

Jamal Daoud,* Lawrence Rosenberg,† and Maryam Tabrizian*

*Department of Biomedical Engineering, Faculty of Medicine, McGill University, Montreal, QC, Canada
†Department of Surgery, Faculty of Medicine, McGill University, Montreal, QC, Canada

Postisolation islet survival is a critical step for achieving successful and efficient islet transplantation. This involves the optimization of islet culture in order to prolong survival and functionality in vitro. Many studies have focused on different strategies to culture pancreatic islets in vitro through manipulation of culture media, surface modified substrates, and the use of various techniques such as encapsulation, embedding, scaffold, and bioreactor culture strategies. This review aims to present and discuss the different methodologies employed to optimize pancreatic islet culture in vitro as well as address their respective advantages and drawbacks.

Key words: Islet culture; Surface modification; Encapsulation; Scaffolds; Bioreactors

Address correspondence to to Maryam Tabrizian, Duff Medical Building, 3775 University Street Room 313, Montreal, QC H3A 2B4, Canada. Tel: (514) 398-8129; Fax: (514) 398-7461; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1537–1546, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X51660
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Antiproinflammatory Effects of Iodixanol (OptiPrep)-Based Density Gradient Purification on Human Islet Preparations

A. Mita,*†† C. Ricordi,*†‡ S. Messinger,*¶ A. Miki,* R. Misawa,*†† S. Barker,* R. D. Molano,* R. Haertter,* A. Khan,* S. Miyagawa,¶ A. Pileggi,* L. Inverardi,*§ R. Alejandro,*# B. J. Hering,** and H. Ichii*†‡

*Cell Transplant Center, Diabetes Research Institute, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
†DeWitt Family Department of Surgery, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
‡Miami Transplant Institute, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
§Department of Epidemiology and Public Health, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
¶Department of Microbiology and Immunology, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
#Department of Medicine, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA
**Department of Surgery, Diabetes Institute for Immunology and Transplantation, University of Minnesota Medical School, Minneapolis, MN, USA
††Division of Transplantation, Department of Surgery, Shinshu University School of Medicine, Matsumoto, Nagano, Japan

Islet isolation and purification using a continuous density gradient may reduce the volume of tissue necessary for implantation into patients, therefore minimizing the risks associated with intraportal infusion in islet transplantation. On the other hand, the purification procedure might result in a decreased number of islets recovered due to various stresses such as exposure to cytokine/chemokine. While a Ficoll-based density gradient has been widely used in purification for clinical trials, purification with iodixanol (OptiPrep) has been recently reported in islet transplant series with successful clinical outcomes. The aim of the current study was to compare the effects of the purification method using OptiPrep-based and Ficoll-based density gradients. Human islet isolations were performed using a modified automated method. After the digestion phase, prepurification digests were divided into two groups and purified using a semiautomated cell processor with either a continuous Ficoll- or OptiPrep-based density gradient. The quantity, purity, viability, and cellular composition of islet preparations from each group were assessed. Cytokine/chemokine and tissue factor production from islet preparations after 48-h culture were also measured. Although islet purity, postpurification IEQ, islet recovery rate, FDA/PI, and fractional β-cell viability were comparable, β-cell mass after 48-h culture significantly improved in the OptiPrep group when compared to the Ficoll group. The production of cytokine/chemokine including IL-1 β, TNF-α, IFN-γ, IL-6, IL-8, MIP-1 β, MCP-1, and RANTES but not tissue factor from the OptiPrep group was significantly lower during 48-h culture after isolation. Each preparation contained the similar number of ductal cells and macrophages. Endotoxin level in both gradient medium was also comparable. The purification method using OptiPrep gradient media significantly reduced cytokine/chemokine production but not tissue factor from human islet preparations and improved â-cell survival during pretransplant culture. Our results suggest that the purification method using OptiPrep gradient media may be of assistance in increasing successful islet transplantation.

Key words: Islet; Transplantation; OptiPrep; Ficoll; Cytokine; Purification

Address correspondence to Hirohito Ichii, M.D., Ph.D., at his current address: Division of Transplant, Department of Surgery, University of California Irvine, 333 City Boulevard West Suite 1205, Irvine, CA 92697, USA. Tel: 714-456-8429; Fax: 714-456-8796; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1547–1561, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X513973
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Choice of Immunosuppression Influences Cytomegalovirus DNAemia in Cynomolgus Monkey (
Macaca fascicularis) Islet Allograft Recipients

Dongmei Han,* Dora M. Berman,*† Melissa Willman,* Peter Buchwald,*‡ Daniel Rothen,§ Norman M. Kenyon,* and Norma S. Kenyon*†¶#

*Diabetes Research Institute, University of Miami School of Medicine, Miami, FL, USA
†Department of Surgery, University of Miami School of Medicine, Miami, FL, USA
‡Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, Miami, FL, USA
§Division of Veterinary Resources, University of Miami School of Medicine, Miami, FL, USA
¶Department of Medicine, University of Miami School of Medicine, Miami, FL, USA
#Department of Microbiology and Immunology, University of Miami School of Medicine, Miami, FL, USA

This retrospective study reviews the results of our experience with the occurrence of CMV DNAemia in islet cell transplanted cynomolgus monkeys subjected to different immunosuppressive protocols, including induction treatment with thymoglobulin (TMG), with a combination of thymoglobulin and fludarabine (FLUD), with cyclophosphamide, or with daclizumab. CMV DNA in the peripheral blood (CMV DNAemia) of 47 monkeys was quantified by real-time PCR on a weekly to biweekly basis. As compared to other immunosuppressive regimens, and in association with greater decreases in WBC, lymphocyte, CD3+CD4+, and CD3+CD8+ lymphocyte counts, frequent CMV DNAemia occurred earlier (within the first month posttransplant), and was of greater severity and duration in recipients of TMG ± FLUD. Treatment of recipients with alternative induction agents that resulted in less dramatic reductions in WBC and lymphocyte counts, however, resulted in occurrence of CMV DNAemia after postoperative day 60. The frequency, average intensity, duration, and area under the curve (AUC) for CMV DNAemia in animals receiving TMG ± FLUD were 75–100%, 4.02 ± 1.75 copies/ng DNA, 23.0 ± 5.3 days, and 367.0 ± 121.1 days × copies/ng DNA, respectively; corresponding values in animals receiving other treatments (0–44%, 0.19 ± 0.10 copies/ng DNA, 0.5 ± 0.3 days, and 75.4 ± 40.2 days × copies/ng DNA, respectively) were significantly different. The value of WBC, T and B cells at the nadir of cell depletion greatly affects the occurrence of CMV DNAemia. No animals developed CMV DNAemia within the next 3 weeks when the lowest value of WBC, lymphocyte, CD3+, CD3+CD4+, CD3+CD8+, or CD20+ cells was above 4500, 1800, 300, 200, 150, or 300 cells/μl, respectively. Oral valganciclovir prophylaxis did not completely prevent the appearance of CMV DNAemia.

Key words: Cytomegalovirus; Nonhuman primates; Transplantation; Immunosuppression; Infection

Address correspondence to Dongmei Han, Ph.D., Diabetes Research Institute, University of Miami School of Medicine, 1450 NW 10th Avenue (R-134), Miami, FL 33136, USA. Tel: (305) 243-2275; Fax: (305) 243-1042; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1563–1572, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X515881
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Lack of Evidence for Recipient Precursor Cells Replenishing β-Cells in Transplanted Islets

Yoshiyuki Hamamoto,*†‡ Tomoyuki Akashi,*† Akari Inada,*† Susan Bonner-Weir,*† and Gordon C. Weir*†

*Section on Islet Transplantation and Cell Biology, Research Division, Joslin Diabetes Center, Boston, MA, USA
†Department of Medicine, Harvard Medical School, Boston, MA, USA
‡Center for Diabetes and Endocrinology, Tazuke Kofukai Foundation Medical Institute Kitano Hospital, Osaka, Japan

Bone marrow and tissue precursor cells have been postulated to replenish grafts of transplanted islets. Several investigators have reported that bone marrow cells can promote the regeneration of injured islets. In this study, we investigated the potential of recipient-derived precursor cells to form new pancreatic endocrine cells in islet grafts transplanted under the kidney capsule. Mouse insulin promoter (MIP)-green fluorescence protein (GFP) mice, which express GFP only in β-cells, or β-actin GFP mice, which express GFP ubiquitously, were used to determine if the recipient-derived cells differentiate into β-cells or other types of endocrine cells. We transplanted MIP-GFP islets into wild-type mice, wild-type islets into MIP-GFP mice, β-actin GFP islets into wild-type mice, and wild-type islets into β-actin GFP mice. β-Actin GFP bone marrow cells were then injected into wild-type mice to evaluate the potential role of bone marrow stem cells to provide new islet cells to the graft. No β-cells with green fluorescence were seen in the graft when wild-type islets were transplanted into MIP-GFP mice. When wild-type islets were transplanted into β-actin GFP mice, no β-cells with GFP staining could be identified in the grafts. Similarly, no endocrine cells with GFP staining could be identified in the grafts after injection of β-actin GFP bone marrow cells into wild-type islet-transplanted wild-type mice. This study provides further support for the concept that recipient precursor cells do not produce new β-cells in grafts of transplanted islets.

Key words: Islet precursor cell; Islet transplantation; Regeneration; Bone marrow; Stem cell

Address correspondence to Gordon C. Weir, M.D., Section on Islet Transplantation and Cell Biology, Joslin Diabetes Center, One Joslin Place, Boston, MA 02215, USA. Tel: (617) 732-2581; Fax: (617) 732-2650; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1573–1585, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X515863
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Assessment of Human Islet Labeling With Clinical Grade Iron Nanoparticles Prior to Transplantation for Graft Monitoring by MRI

Frederic Ris,*†1 Matthieu Lepetit-Coiffe,‡1 Paolo Meda,§ Lindsey A. Crowe,‡ Christian Toso,*† Mathieu Armanet,* Nadja Niclauss,* Ge´raldine Parnaud,* Laurianne Giovannoni,* Domenico Bosco,* Philippe Morel,† Jean-Paul Vallee,‡ and Thierry Berney*†

*Cell Isolation and Transplantation Center, Geneva University Hospitals and University of Geneva, School of Medicine, Geneva, Switzerland
†Department of Surgery, Geneva University Hospitals and University of Geneva, School of Medicine, Geneva, Switzerland
‡Department of Radiology, Geneva University Hospitals and University of Geneva, School of Medicine, Geneva, Switzerland
§Department of Cellular Physiology and Metabolism, Geneva University Hospitals and University of Geneva, School of Medicine, Geneva, Switzerland

Ex vivo labeling of islets with superparamagnetic iron oxide (SPIO) nanoparticles allows posttransplant MRI imaging of the graft. In the present study, we compare two clinical grade SPIOs (ferucarbotran and ferumoxide) in terms of toxicity, islet cellular uptake, and MRI imaging. Human islets (80–90% purity) were incubated for 24 h with various concentrations of SPIOs (14–280 μg/ml of iron). Static incubations were performed, comparing insulin response to basal (2.8 mM) or high glucose stimulation (16.7 mM), with or without cAMP stimulation. Insulin and Perl’s (assessment of iron content) staining were performed. Electronic microscopy analysis was performed. Labeled islets were used for in vitro or in vivo imaging in MRI 1.5T. Liver section after organ removal was performed in the same plane as MRI imaging to get a correlation between histology and radiology. Postlabeling islet viability (80 ± 10%) and function (in vitro static incubation and in vivo engraftment of human islets in nude mice) were similar in both groups. Iron uptake assessed by electron microscopy showed iron inclusions within the islets with ferucarbotran, but not with ferumoxide. MRI imaging (1.5T) of phantoms and of human islets transplanted in rats, demonstrated a strong signal with ferucarbotran, but only a weak signal with ferumoxide. Signal persisted for >8 weeks in the absence of rejection. An excellent correlation was observed between radiologic images and histology. The hepatic clearance of intraportally injected ferucarbotran was faster than that of ferumoxide, generating less background. A rapid signal decrease was observed in rejecting xenogeneic islets. According to the present data, ferucarbotran is the most appropriate of available clinical grade SPIOs for human islet imaging.

Key words: Islet imaging; Islet transplantation; Iron nanoparticles; Magnetic resonance; Imaging

1These authors provided equal contribution to this study.
Address correspondence to Frederic Ris, M.D., Cell Isolation and Transplantation Center, Department of Surgery, Geneva University Hospitals, 4, rue Gabrielle-Perret-Gentil, 1211 Geneva 14, Switzerland. Tel: +41 22 372 33 03; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1587–1597, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X514323
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Intraoperative Intracerebral MRI-Guided Navigation for Accurate Targeting in Nonhuman Primates

Marina E. Emborg,*† Valerie Joers,* Ronald Fisher,‡ Kevin Brunner,* Victoria Carter,* Chris Ross,¶ Raghu Raghavan,# Martin Brady,# James Raschke,* Ken Kubota,** and Andrew Alexander*†‡§

*Preclinical Parkinson’s Research Program, Wisconsin National Primate Research Center, University of Wisconsin, Madison, WI, USA
†Department of Medical Physics, University of Wisconsin, Madison, WI, USA
‡Waisman Center, University of Wisconsin, Madison, WI, USA
§Department of Psychiatry, University of Wisconsin, Madison, WI, USA
¶Engineering Resources Group Inc., Hialeah, FL, USA
#Therataxis, Baltimore, MD, USA
*Kinetics Foundation, Los Altos, CA, USA

During in vivo intracerebral infusions, the ability to perform accurate targeting towards a 3D-specific point allows control of the anatomical variable and identification of the effects of variations in other factors. Intraoperative MRI navigation systems are currently being used in the clinic, yet their use in nonhuman primates and MRI monitoring of intracerebral infusions has not been reported. In this study rhesus monkeys were placed in a MRI-compatible stereotaxic frame. T1 MRIs in the three planes were obtained in a 3.0T GE scanner to identify the target and plan the trajectory to ventral postcommisural putamen. A craniotomy was performed under sterile surgical conditions at the trajectory entry point. A modified MRI-compatible trajectory guide base (Medtronic Inc.) was secured above the cranial opening and the alignment stem applied. Scans were taken to define the position of the alignment stem. When the projection of the catheter in the three planes matched the desired trajectory to the target, the base was locked in position. A catheter replaced the alignment stem and was slowly introduced to the final target structure. Additional scans were performed to confirm trajectory and during the infusion of a solution of gadoteridol (ProHance, Bracco Diagnostics; 2 mM/L) and bromophenol blue (0.16 mg/ml) in saline. Monitoring of the pressure in the infusion lines was performed using pressure monitoring and infusion pump controller system (Engineering Resources Group Inc.) in combination with a MRI-compatible infusion pump (Harvard). MRI during infusion confirmed successful targeting and matched postmortem visualization of bromophenol blue. Assessment of the accuracy of the targeting revealed an overall 3D mean ± SD distance error of 1.2 ± 0.6 mm and angular distance error of 0.9 ± 0.5 mm. Our results in nonhuman primates confirm the accuracy of intraoperative MRI intracerebral navigation combined with an adaptable, pivot point-based targeting system and validates its use for preclinical intracerebral procedures.

Key words: Magnetic resonance imaging (MRI); Stereotaxic surgery; Intracerebral infusions; Convection-enhanced delivery; Intracerebral targeting; Putamen

Address correspondence to Marina E. Emborg, M.D., Ph.D., Preclinical Parkinson’s Research Program, Wisconsin National Primate Research Center, University of Wisconsin-Madison, 1223 Capitol Court, Madison, WI 53715, USA. Tel: 608-262-9714; Fax: 608-263-3524; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1599–1607, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X513982
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Impact of Escaped Bone Marrow Mesenchymal Stromal Cells on Extracardiac Organs After Intramyocardial Implantation in a Rat Myocardial Infarction Model

Wei Wang,*†1 Peifeng Jin,†‡1 Lei Wang,†§ Zhikai Yang,*† Shengshou Hu,† Bingren Gao,* and Hao Zhang†

*Department of Thoracic and Cardiovascular Surgery, Second Hospital Of Lanzhou University, Lanzhou, China
†Department of Surgery and Research Center for Cardiac Regenerative Medicine, Cardiovascular Institute and Fu Wai Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
‡Department of Thoracic and Cardiovascular Surgery, First Affiliated Hospital of Wenzhou Medical College, Wenzhou, China
§Department of Thoracic and Cardiovascular Surgery, Affiliated Hospital of Weifang Medical College, Weifang, China

Cell escape occurs after intramyocardial injection for treatment of myocardial infarction (MI) and then the migrated cells might be entrapped by extracardiac organs. We investigated the fate of migrated bone marrow-derived mesenchymal stromal cells (MSCs) and their impact on lung, liver, and spleen. MI model was created by coronary artery ligation in female Lewis rats. Three weeks after the ligation, bromodeoxyuridine (BrdU)-labeled male MSCs were directly injected into the infarcted area in the cell transplantation group (n = 22). The same volume of phosphate-buffered solution (PBS) was injected in the control group (n = 21). In the sham group (n = 10) intramyocardial injection of the same volume of PBS was performed in healthy rats. Four weeks later, echocardiography was performed and the cell retention was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Immunohistochemistry study was performed to identify the migrated cells. Heart function was improved after the cell injection. qRT-PCR results showed the percentage of retained cells in heart, spleen, liver, and lung ranked 3.63 ± 0.48%, 0.77 ± 0.13%, 0.68 ± 0.10%, 0.62 ± 0.11%, respectively, after cell transplantation. The implanted MSCs that escaped to liver, spleen, and lung did not differentiate into fibroblast, myofibroblast, or alveolar epithelial cells. However, the migrated MSCs in liver expressed functional hepatocyte marker. In conclusion, cell migration after intramyocardial injection did not result in deterioration of lung, liver, and spleen function. Our study might pave the way for new safety investigation of emerging cell resources and their impact on target and untargeted organs.

Key words: Bone marrow mesenchymal stromal cells; Myocardial infarction; Lung; Liver; Spleen

1
These authors provided equal contribution to this work.
Address correspondence to Hao Zhang, Department of Surgery and Research Center for Cardiac Regenerative Medicine, Cardiovascular Institute and Fu Wai Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100037, China. Tel: +86 10 88396050; Fax: +86 10-88396050; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Bingren Gao, Department of Thoracic and Cardiovascular Surgery, Second Hospital of Lanzhou University, Lanzhou, 730030, China. Tel: +86 931 8942306; Fax: +86 931 8458109; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1609–1622, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X516583
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Characterization of Tolerance Induction Through Prenatal Marrow Transplantation: The Requirement for a Threshold Level of Chimerism to Establish Rather Than Maintain Postnatal Skin Tolerance

Jeng-Chang Chen,*† Ming-Ling Kuo,‡ Liang-Shiou Ou,§ Pei-Yeh Chang,* Marcus O. Muench,¶ Chia-Rui Shen,# Hsueh-Ling Chang,** Hsiu-Yueh Yu,** and Ren-Huei Fu§

*Department of Surgery, Chang Gung Memorial Hospital, Chang Gung University, Taoyuan, Taiwan
†Graduate Institute of Clinical Medical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan
‡Department of Microbiology and Immunology, Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan
§Department of Pediatrics, Chang Gung Memorial Hospital, Taoyuan, Taiwan
¶Blood Systems Research Institute and Department of Laboratory Medicine, University of California, San Francisco, CA, USA
#Graduate Institute of Medical Biotechnology and Laboratory Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan
**Pediatric Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan

Hematopoietic chimerism resulting from prenatal marrow transplantation does not consistently result in allotolerance for unidentified causes. In a C57BL/6-into-FVB/N murine model, we transplanted T-cell-depleted adult marrow on gestational day 14 to elucidate the immunological significance of chimerism towards postnatal tolerance. Postnatally, chimerism was examined by flow cytometry, and tolerance by skin transplantation and mixed lymphocyte reaction. Regulatory T cells were quantified by FoxP3 expression. Peripheral chimerism linearly related to thymic chimerism, and predicted the degree of graft acceptance with levels >3% at skin placement, yielding consistent skin tolerance. Low- and high-level chimeras had lower intrathymic CD3high expression than microchimeras or untransplanted mice. Regardless of the skin tolerance status in mixed chimeras, donor-specific alloreactivity by lymphocytes was suppressed but could be partially restored by exogenous interleukin-2. Recipients that lost peripheral chimerism did not accept donor skin unless prior donor skin had engrafted at sufficient chimerism levels, suggesting that complete tolerance can develop as a consequence of chimerism-related immunosuppression of host lymphocytes and the tolerogenic effects of donor skin. Thus, hematopoietic chimerism exerted immunomodulatory effects on the induction phase of allograft tolerance. Once established, skin tolerance did not fade away along with spontaneous regression of peripheral and tissue chimerism, as well as removal of engrafted donor skin. Neither did it break following in vivo depletion of increased regulatory T cells, and subcutaneous interleukin-2 injection beneath the engrafted donor skin. Those observations indicate that the maintenance of skin tolerance is multifaceted, neither solely dependent upon hematopoietic chimerism and engrafted donor skin nor on the effects of regulatory T cells or clonal anergy. We conclude that hematopoietic chimerism generated by in utero hematopoietic stem cell transplantation is critical to establish rather than maintain postnatal skin tolerance. Therefore, the diminution of hematopoietic chimerism below a threshold level does not nullify an existing tolerance state, but lessens the chance of enabling complete tolerance.

Key words: Chimerism; Hematopoietic stem cells; In utero; Marrow transplantation; Transplantation tolerance

Address correspondence to Jeng-Chang Chen, M.D., Department of Surgery, Chang Gung Memorial Hospital, 5, Fu-Shin Street, Kweishan, 333, Taoyuan, Taiwan. Tel: +886-3-3281200, ext. 8227; Fax: +886-3-3287261; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Ming-Ling Kuo, Ph.D., Department of Microbiology and Immunology, Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, 259, Wen-Hwa 1st Road, Kweishan, 333, Taoyuan, Taiwan. Tel: +886-3-2118800, ext. 3319; Fax: +886-3-2118293; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1623–1633, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X514297
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Evaluation of Alginate Microspheres for Mesenchymal Stem Cell Engraftment on Solid Organ

E. Trouche,* S. Girod Fullana,†‡ C. Mias,* C. Ceccaldi,†‡ F. Tortosa,*† M. H. Seguelas,* D. Calise,* A. Parini,*† D. Cussac,*†1 and B. Sallerin*†1

*INSERM U858, Toulouse, France
†Université Toulouse, UPS, Faculté des Sciences Pharmaceutiques, Toulouse, France
‡CIRIMAT, UPS-INPT-CNRS 5085, Toulouse, France

Mesenchymal stem cells (MSCs) may be used as a cell source for cell therapy of solid organs due to their differentiation potential and paracrine effect. Nevertheless, optimization of MSC-based therapy needs to develop alternative strategies to improve cell administration and efficiency. One option is the use of alginate microencapsulation, which presents an excellent biocompatibility and an in vivo stability. As MSCs are hypoimmunogenic, it was conceivable to produce microparticles with [alginate-poly-L-lysine-alginate (APA) microcapsules] or without (alginate microspheres) a surrounding protective membrane. Therefore, the aim of this study was to determine the most suitable microparticles to encapsulate MSCs for engraftment on solid organ. First, we compared the two types of microparticles with 4 × 106 MSCs/ml of alginate. Results showed that each microparticle has distinct morphology and mechanical resistance but both remained stable over time. However, as MSCs exhibited a better viability in microspheres than in microcapsules, the study was pursued with microspheres. We demonstrated that viable MSCs were still able to produce the paracrine factor bFGF and did not present any chondrogenic or osteogenic differentiation, processes sometimes reported with the use of polymers. We then proved that microspheres could be implanted under the renal capsule without degradation with time or inducing impairment of renal function. In conclusion, these microspheres behave as an implantable scaffold whose biological and functional properties could be adapted to fit with clinical applications.

Key words: Mesenchymal stem cells; Alginate; Microspheres

1
These authors provided equal contribution to this work.
Address correspondence to Brigitte Sallerin, I2MR, INSERM U858, CHU Rangueil, BP 84225, 31432 Toulouse Cedex 4, France. Tel: 33-5-65-25-68-43; Fax: 33-5-65-25-98-16; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1635–1644, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X516637
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Equivalence for Diabetic Wound Healing

Ji Yeon Kim,*† Sun-Hwa Song,* Koung Li Kim,*§ Jeong-Jae Ko,* Ji-Eun Im,* Se Won Yie,† Young Keun Ahn,‡ Duk-Kyung Kim,§ and Wonhee Suh*

*Department of Biomedical Science, College of Life Science, CHA University, Gyeonggi-do, Korea
†Department of Molecular Bioscience, Division of Bioscience and Biotechnology, Kangwon National University, Chuncheon, Korea
‡Department of Cardiology, Chonnam National University Medical Center, Gwangju, Korea
§Department of Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul Korea

Transplantation of human cord blood-derived endothelial progenitor cells (EPCs) is reported to contribute to neovascularization in various ischemic diseases. However, the possible beneficial role and underlying mechanisms in diabetes-impaired wound healing have been less well characterized. In this study, EPC transplantation stimulated keratinocyte and fibroblast proliferation substantially as early as 3 days after injury, leading to significantly accelerated wound closure in streptozotocin-induced diabetic nude mice, compared to PBS control. RT-PCR analysis showed that EPCs secreted various wound healing-related growth factors. Among them, keratinocyte growth factor and platelet-derived growth factor were highly expressed in the EPCs and were present at substantial levels in the EPC-injected dermal tissue. Using EPC-conditioned medium (CM), we found that paracrine factors from EPCs directly exerted mitogenic and chemotactic effects on keratinocytes and fibroblasts. Moreover, injection of EPC-CM alone into the same diabetic wound mice promoted wound healing and increased neovascularization to a similar extent as achieved with EPC transplantation. These results indicate that the beneficial effect of EPC transplantation on diabetic wounds was mainly achieved by their direct paracrine action on keratinocytes, fibroblasts, and endothelial cells, rather than through their physical engraftment into host tissues (vasculogenesis). In addition, EPC-CM was shown to be therapeutically equivalent to EPCs, at least for the treatment of diabetic dermal wounds, suggesting that conditioned medium may serve as a novel therapeutic option that is free from allograft-associated immune rejection concern.

Key words: Endothelial progenitor cells (EPCs); Diabetes mellitus; Paracrine signaling; Wound healing; Animal model

Address correspondence to Wonhee Suh, Ph.D., Department of Biomedical Science, College of Life Science, CHA University, CHA Stem Cell Institute, 606-16 Yeoksam1-dong, Kangnam-gu, Seoul, 135-907, Korea. Tel: 82-2-3468-3668; Fax: 82-2-538-4102; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1645–1657, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X516628
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Delivery of a Therapeutic Protein by Immune-Privileged Sertoli Cells

Katelyn Halley,1 Emily L. Dyson,1 Gurvinder Kaur, Payal Mital, Peter M. Uong, Brinda Dass, Sherry N. Crowell, and Jannette M. Dufour

Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX, USA

Immune-privileged Sertoli cells survive long term after allogeneic or xenogeneic transplantation without the use of immunosuppressive drugs, suggesting they could be used as a vehicle to deliver therapeutic proteins. As a model to test this, we engineered Sertoli cells to transiently produce basal levels of insulin and then examined their ability to lower blood glucose levels after transplantation into diabetic SCID mice. Mouse and porcine Sertoli cells transduced with a recombinant adenoviral vector containing furin-modified human proinsulin cDNA expressed insulin mRNA and secreted insulin protein. Transplantation of 5–20 million insulin-expressing porcine Sertoli cells into diabetic SCID mice significantly decreased blood glucose levels in a dose-dependent manner, with 20 million Sertoli cells decreasing blood glucose levels to 9.8 ± 2.7 mM. Similar results were obtained when 20 million insulin-positive, BALB/c mouse Sertoli cells were transplanted; blood glucose levels dropped to 6.3 ± 2.4 mM and remained significantly lower for 5 days. To our knowledge, this is the first study to demonstrate Sertoli cells can be engineered to produce and secrete a clinically relevant factor that has a therapeutic effect, thus supporting the concept of using immune-privileged Sertoli cells as a potential vehicle for gene therapy.

Key words: Sertoli cell; Gene therapy; Transplantation; Insulin; Diabetes

1
These authors provided equal contribution to this work.
Address correspondence to Dr. Jannette M. Dufour, Cell Biology and Biochemistry, 3601 4th St. STOP 6540, Texas Tech University Health Sciences Center, Lubbock, TX, 79430, USA. Tel: (806) 743-2616; Fax: (806) 743-2990; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation
, Vol. 19, pp. 1659–1670, 2010
0963-6897/10 $90.00 + .00
DOI: 10.3727/096368910X516619
E-ISSN 1555-3892
Copyright © 2010 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Intravital Two-Photon Microscopy Assessment of Renal Protection Efficacy of siRNA for p53 in Experimental Rat Kidney Transplantation Models

Ryoichi Imamura,* Yoshitaka Isaka,† Ruben M. Sandoval,‡ Asaf Ori,§ Swetlana Adamsky,§ Elena Feinstein,§ Bruce A. Molitoris,‡ and Shiro Takahara¶

*Department of Urology, Osaka University Graduate School of Medicine, Osaka, Japan
†Department of Nephrology, Osaka University Graduate School of Medicine, Osaka, Japan
‡Department of Medicine, Division of Nephrology, Indiana Center for Biological Microscopy, Indiana University, Bloomington, IN, USA
§Quark Pharmaceuticals Inc., Fremont, CA, USA
¶Department of Advanced Technology for Transplantation, Osaka University Graduate School of Medicine, Osaka, Japan

Renal ischemia-reperfusion (I/R) injury, which is unavoidable in renal transplantation, frequently influences both short- and long-term allograft survival. Despite decades of laboratory and clinical investigations, and the advent of renal replacement therapy, the overall mortality rate due to acute tubular injury has changed little. I/R-induced DNA damage results in p53 activation in proximal tubule cells (PTC), leading to their apoptosis. Therefore, we examined the therapeutic effect of temporary p53 inhibition in two rat renal transplantation models on structural and functional aspects of injury using intravital two-photon microscopy. Nephrectomized Sprague-Dawley rats received syngeneic left kidney transplantation either after 40 min of intentional warm ischemia or after combined 5-h cold and 30-min warm ischemia of the graft. Intravenously administrated siRNA for p53 (siP53) has previously been shown to be filtered and reabsorbed by proximal tubular epithelial cells following the warm ischemia/reperfusion injury in a renal clamp model. Here, we showed that it was also taken up by PTC following 5 h of cold ischemia. Compared to saline-treated recipients, treatment with siP53 resulted in conservation of renal function and significantly suppressed the I/R-induced increase in serum creatinine in both kidney transplantation models. Intravital two-photon microscopy revealed that siP53 significantly ameliorated structural and functional damage to the kidney assessed by quantification of tubular cast formation and the number of apoptotic and necrotic tubular cells and by evaluation of blood flow rate. In conclusion, systemic administration of siRNA for p53 is a promising new approach to protect kidneys from I/R injury in renal transplantation.

Key words: Apoptosis; p53; Ischemia-reperfusion injury; Kidney; Small interfering RNA (siRNA)

Address correspondence to Yoshitaka Isaka, M.D., Ph.D., Department of Nephrology, Osaka University Graduate School of Medicine, Suita, Osaka, 565-0871 Japan. Tel: +81-6-6879- 3857; Fax: +81-6-6879-3857; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it