Cell Transplantation 22(11) Abstracts

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Cell Transplantation, Vol. 22, pp. 1959-1970, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X661382
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Functionalizable Silica-Based Micron-Sized Iron Oxide Particles for Cellular Magnetic Resonance Imaging


Nathanael Raschzok,* Carolin M. Langer,* Christian Schmidt,† Karl H. Lerche,† Nils Billecke,* Kerstin Nehls,* Natalie B. Schlüter,* Annekatrin Leder,* Susanne Rohn,* Martina T. Mogl,* Lutz Lüdemann,‡ Lars Stelter,§ Ulf K. Teichgräber,§¶ Peter Neuhaus,* and Igor M. Sauer*

*General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Universitätsmedizin Berlin, Berlin, Germany
†Microparticles GmbH, Berlin, Germany
‡Klinik für Strahlentherapie, Universitätsklinikum Essen, Essen, Germany
§Department of Radiology, Charité-Universitätsmedizin Berlin, Berlin, Germany
¶Department of Radiology, Universitätsklinikum Jena, Jena, Germany

Cellular therapies require methods for noninvasive visualization of transplanted cells. Micron-sized iron oxide particles (MPIOs) generate a strong contrast in magnetic resonance imaging (MRI) and are therefore ideally suited as an intracellular contrast agent to image cells under clinical conditions. However, MPIOs were previously not applicable for clinical use. Here, we present the development and evaluation of silica-based micron-sized iron oxide particles (sMPIOs) with a functionalizable particle surface. Particles with magnetite content of >40% were composed using the sol-gel process. The particle surfaces were covered with COOH groups. Fluorescein, poly-L-lysine (PLL), and streptavidin (SA) were covalently attached. Monodisperse sMPIOs had an average size of 1.18 μm and an iron content of about 1.0 pg Fe/particle. Particle uptake, toxicity, and imaging studies were performed using HuH7 cells and human and rat hepatocytes. sMPIOs enabled rapid cellular labeling within 4 h of incubation; PLL-modified particles had the highest uptake. In T2*-weighted 3.0 T MRI, the detection threshold in agarose was 1,000 labeled cells, whereas in T1-weighted LAVA sequences, at least 10,000 cells were necessary to induce sufficient contrast. Labeling was stable and had no adverse effects on labeled cells. Silica is a biocompatible material that has been approved for clinical use. sMPIOs could therefore be suitable for future clinical applications in cellular MRI, especially in settings that require strong cellular contrast. Moreover, the particle surface provides the opportunity to create multifunctional particles for targeted delivery and diagnostics.

Key words: Silica; Magnetic resonance imaging (MRI); Theranostics; Cell transplantation; Hepatocyte transplantation

Received January 7, 2012; final acceptance September 25, 2012. Online prepub date: January 2, 2013.
Address correspondence to Dr. Nathanael Raschzok, M.D., General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Campus Virchow Klinikum, Charité - Universitätsmedizin Berlin, Augustenburger Platz 1, 13353 Berlin, Germany. Tel: +49-30-450652356; Fax: +49-30-450559987; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1971-1980, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657747
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Noninvasive In Vivo Tracking of Mesenchymal Stem Cells and Evaluation of Cell Therapeutic Effects in a Murine Model Using a Clinical 3.0 T MRI

Florian Drey,*1 Yeong-Hoon Choi,*†1 Klaus Neef,*† Birgit Ewert,* Arne Tenbrock,* Philipp Treskes,*† Henning Bovenschulte,‡ Oliver J. Liakopoulos,* Meike Brenkmann,* Christof Stamm,§ Thorsten Wittwer,*† and Thorsten Wahlers*†

*Department of Cardiothoracic Surgery, University of Cologne, Cologne, Germany
†Center of Molecular Medicine Cologne, University of Cologne, Cologne, Germany
‡Department of Radiology, University of Cologne, Cologne, Germany
§Department of Cardiothoracic and Vascular Surgery, German Heart Institute Berlin, Berlin, Germany

Cardiac cell therapy with mesenchymal stem cells (MSCs) represents a promising treatment approach for endstage heart failure. However, little is known about the underlying mechanisms and the fate of the transplanted cells. The objective of the presented work is to determine the feasibility of magnetic resonance imaging (MRI) and in vivo monitoring after transplantation into infarcted mouse hearts using a clinical 3.0 T MRI device. The labeling procedure of bone marrow-derived MSCs with micron-sized paramagnetic iron oxide particles (MPIOs) did not affect the viability of the cells and their cell type-defining properties when compared to unlabeled cells. Using a clinical 3.0 T MRI scanner equipped with a dedicated small animal solenoid coil, 105 labeled MSCs could be detected and localized in the mouse hearts for up to 4 weeks after intramyocardial transplantation. Weekly ECG-gated scans using T1-weighted sequences were performed, and left ventricular function was assessed. Histological analysis of hearts confirmed the survival of labeled MSCs in the target area up to 4 weeks after transplantation. In conclusion, in vivo tracking of labeled MSCs using a clinical 3.0 T MRI scanner is feasible. In combination with assessment of heart function, this technology allows the monitoring of the therapeutic efficacy of regenerative therapies in a small animal model.

Key words: Magnetic resonance imaging (MRI); Mesenchymal stem cells (MSCs); Cell tracking; Cardiac function; Myocardial infarction

Received November 11, 2011; final acceptance June 30, 2012. Online prepub date: October 8, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Yeong-Hoon Choi, M.D., Department of Cardiothoracic Surgery, Heart Center of the University, Center of Molecular Medicine Cologne, University of Cologne, Kerpener Str. 62, 50924 Cologne, Germany. Tel: +49 (221) 478-32452; Fax: +49 (221) 478-32509; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1981-2000, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657990
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

GMP-Compliant Isolation and Expansion of Bone Marrow-Derived MSCs in the Closed, Automated Device Quantum Cell Expansion System

Markus T. Rojewski,*†1 Natalie Fekete,*†1 Stefano Baila,‡ Kim Nguyen,§ Daniel Fürst,† Delbert Antwiler,§ Julia Dausend,* Ludwika Kreja,¶ Anita Ignatius,¶ Luc Sensebé,# and Hubert Schrezenmeier*†

*Institut für Transfusionsmedizin, Universität Ulm, Ulm, Germany
†Institut für klinische Transfusionsmedizin und Immungenetik Ulm, DRK-Blutspendedienst Baden-Württemberg – Hessen gemeinnützige GmbH, Ulm, Germany
‡Terumo BCT, Zaventem, Belgium
§Terumo BCT, Lakewood, CO, USA
¶Institute of Orthopaedic Research and Biomechanics, Center of Musculoskeletal Research, University of Ulm, Ulm, Germany
#EFS Pyrénées Méditerranée, Toulouse, France

The estimated frequency of MSCs in BM is about 0.001–0.01% of total nucleated cells. Most commonly, one applied therapeutic cell dose is about 1–5 million MSCs/kg body weight, necessitating a reliable, fast, and safe expansion system. The limited availability of MSCs demands for an extensive ex vivo amplification step to accumulate sufficient cell numbers. Human platelet lysate (PL) has proven to be a safe and feasible alternative to animal-derived serum as supplement for MSC cultivation. We have investigated the functionally closed automated cell culture hollow fiber bioreactor Quantum cell expansion system as an alternative novel tool to conventional tissue flasks for efficient clinical-scale MSC isolation and expansion from bone marrow using PL. Cells expanded in the Quantum system fulfilled MSC criteria as shown by flow cytometry and adipogenic, chondrogenic, and osteogenic differentiation capacity. Cell surface expression of a variety of chemokine receptors, adhesion molecules, and additional MSC markers was monitored for several passages by flow cytometry. The levels of critical media components like glucose and lactate were analyzed. PDGF-AA, PDGF-AB/BB, bFGF, TGF-β1, sICAM-1, sVCAM-1, RANTES, GRO, VEGF, sCD40L, and IL-6 were assessed using a LUMINEX platform. Originally optimized for the use of fetal calf serum (FCS) as supplement and fibronectin as coating reagent, we succeeded to obtain an average of more than 100 × 106 of MSCs from as little as 18.8–28.6 ml of BM aspirate using PL. We obtained similar yields of MSCs/μl BM in the FCS-containing and the xenogen-free expansion system. The Quantum system reliably produces a cellular therapeutic dose in a functionally closed system that requires minimal manipulation. Both isolation and expansion are possible using FCS or PL as supplement. Coating of the hollow fibers of the bioreactor is mandatory when loading MSCs. Fibronectin, PL, and human plasma may serve as coating reagents.

Key words: Mesenchymal stem cells (MSCs); Good manufacturing practice (GMP); Bioreactor; Large scale

Received April 26, 2012; final acceptance September 20, 2012. Online prepub date: October 25, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Markus T. Rojewski, Institut für Transfusionsmedizin, Universität Ulm, Helmholtzstraße 10, D-89081 Ulm, Germany. Tel: +49-7 31-1 50 68 33; Fax: +49-7 31-15 05 00; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2001-2015, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657909
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Restoration of Intracortical and Thalamocortical Circuits After Transplantation of Bone Marrow Mesenchymal Stem Cells Into the Ischemic Brain of Mice

Mingke Song, Osama Mohamad, Xiaohuan Gu, Ling Wei, and Shan Ping Yu

Department of Anesthesiology, Emory University School of Medicine, Atlanta, GA, USA

Transplantation of bone marrow mesenchymal stem cells (BMSCs) provides a promising regenerative medicine for stroke. Whether BMSC therapy could repair ischemia-damaged neuronal circuits and recover electrophysiological activity has largely been unknown. To address this issue, BMSCs were implanted into the ischemic barrel cortex of adult mice 1 and 7 days after focal barrel cortex stroke. Two days after the first transplantation (3 days after stroke), the infarct volume determined by TTC staining was significantly smaller in BMSC-treated compared to vehicle-treated stroke mice. The behavioral corner test showed better long-term recovery of sensorimotor function in BMSC-treated mice. Six weeks poststroke, thalamocortical slices were prepared and neuronal circuit activity in the peri-infarct region of the barrel cortex was determined by extracellular recordings of evoked field potentials. In BMSC-transplanted brain slices, the ischemia-disrupted intracortical activity from layer 4 to layer 2/3 was noticeably recovered, and the thalamocortical circuit connection was also partially restored. In contrast, much less and slower recovery was seen in control animals of barrel cortex stroke. Immunohistochemical staining disclosed that the density of neurons, axons, and blood vessels in the peri-infarct region was significantly higher in BMSC-treated mice, accompanied with enhanced local blood flow recovery. Western blotting showed that BMSC treatment increased the expression of stromal cell-derived factor-1 (SDF-1), vascular endothelial growth factor (VEGF), and brain-derived neurotrophic factor (BDNF) in the peri-infarct region. Moreover, the expression of the axonal growth associated protein-43 (GAP-43) was markedly increased, whereas the axonal growth inhibiting proteins ROCK II and NG2 were suppressed in the BMSC-treated brains. BMSC transplantation also promoted directional migration and survival of doublecortin (DCX)-positive neuroblasts in the peri-infarct region. The present investigation thus provides novel evidence that BMSC transplantation has the potential to repair the ischemia-damaged neural networks and restore lost neuronal connections. The recovered circuit activity likely contributes to the improved sensorimotor function after focal ischemic stroke and BMSC transplantation.

Key words: Bone marrow mesenchymal stem cells (BMSCs); Barrel cortex ischemic stroke; Neuronal circuit; Thalamocortical slices; Evoked field potential; Doublecortin (DCX)

Received October 20, 2011; final acceptance July 26, 2012. Online prepub date: October 12, 2012.
Address correspondence to Shan Ping Yu, 101 Woodruff Circle, Woodruff Memorial Research Building, Suite 620, Department of Anesthesiology, Emory University School of Medicine, Atlanta, GA 30322, USA. Tel: +1-404-712-8678; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2017-2028, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657305
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Adult Periodontal Ligament-Derived Cells Integrate and Differentiate After Implantation Into the Adult Mammalian Brain

Carlos Bueno,* Carmina Ramirez,* Francisco J. Rodríguez-Lozano,† Rafael Tabarés-Seisdedos,‡ Mónica Rodenas,* Jose M. Moraleda,† Jonathan R. Jones,* and Salvador Martinez*

*Instituto de Neurociencias de Alicante (UMH-CSIC), Sant Juan, Alicante, Spain
†Cell Therapy Unit, Hospital Universitary Virgen de la Arrixaca, Faculty of Medicine, University of Murcia, Murcia, Spain
‡Department of Medicine, University of Valencia, Centro de Investigación Biomédica en Red en el area de Salud Mental (CIBERSAM), Valencia, Spain

Previous studies suggest that neural crest (NC)-derived stem cells may reside in NC derivatives including the human periodontal ligament (hPDL). The isolation and manipulation of autologous NC-derived cells could be an accessible source of adult neural stem cells for their use in cell replacement and gene transfer to the diseased central nervous system. In this study, we examined the expression of NC markers and neural differentiation potential of hPDL-derived cells both in vitro and in vivo. In vitro we found that hPDL-derived cells expressed stem cell markers (Oct3/4, Nestin, Sox2, and Musashi-1) and a subset of NC cell markers (Slug, p75NTR, Twist, and Sox9). hPDL-derived cells differentiated into neural-like cells based on cellular morphology and neural marker expression (TUJ1, MAP2, MAP1b, GAD65/67, GABA, NeuN, ChAT, GAT1, synaptophysin, GFAP, NG2, and O4). In vivo, hPDL-derived cells survive, migrate, and give rise to DCX+, NF-M+, GABA+, GFAP+, and NG2+ cells after grafting the adult mouse brain. Some of the grafted hPDL-derived cells were located in stem cell niches such as the ventricular epithelium and the subventricular zone of the anterolateral ventricle wall as well as in the subgranular zone of the hippocampal dentate gyrus. Thus, the hPDL contains stem cells that originate from the NC and can differentiate into neural cells. The engraftment and differentiation properties of hPDL-derived cells in the adult brain indicate that they are a potential stem cell source to be used in neuroregenerative and/or neurotrophic medicine.

Key words: Tissue-specific stem cells; Periodontal ligament (PDL); Neural crest (NC); Neural stem cells; Stem cell transplantation; Cell therapy

Received December 12, 2011; final acceptance June 25, 2012. Online prepub date: October 3, 2012.
Address correspondence to Salvador Martinez, M.D., Ph.D., Instituto de Neurociencias, UMH-CSIC, Campus de San Juan, E-03550-Alicante, Spain. Tel: +34-96-591-9556; Fax: +34-96-591-9555; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2029-2039, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658953
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Sciatic Nerve Regeneration by Cocultured Schwann Cells and Stem Cells on Microporous Nerve Conduits

Lien-Guo Dai,*† Guo-Shiang Huang,‡ and Shan-hui Hsu‡§¶

*Department of Chemical Engineering, National Chung Hsing University, Taichung, Taiwan
†Department of Orthopedics, Kuang Tien General Hospital, Taichung, Taiwan
‡Institute of Polymer Science and Engineering, National Taiwan University, Taipei, Taiwan
§Research Center for Developmental Biology and Regenerative Medicine, National Taiwan University, Taipei, Taiwan
¶Institute of Biomedical Engineering, National Chung Hsing University, Taichung, Taiwan

Cell transplantation is a useful therapy for treating peripheral nerve injuries. The clinical use of Schwann cells (SCs), however, is limited because of their limited availability. An emerging solution to promote nerve regeneration is to apply injured nerves with stem cells derived from various tissues. In this study, different types of allogeneic cells including SCs, adipose-derived adult stem cells (ASCs), dental pulp stem cells (DPSCs), and the combination of SCs with ASCs or DPSCs were seeded on nerve conduits to test their efficacy in repairing a 15-mm-long critical gap defect of rat sciatic nerve. The regeneration capacity and functional recovery were evaluated by the histological staining, electrophysiology, walking track, and functional gait analysis after 8 weeks of implantation. An in vitro study was also performed to verify if the combination of cells led to synergistic neurotrophic effects (NGF, BDNF, and GDNF). Experimental rats receiving conduits seeded with a combination of SCs and ASCs had the greatest functional recovery, as evaluated by the walking track, functional gait, nerve conduction velocity (NCV), and histological analysis. Conduits seeded with cells were always superior to the blank conduits without cells. Regarding NCV and the number of blood vessels, conduits seeded with SCs and DPSCs exhibited better values than those seeded with DPSCs only. Results from the in vitro study confirmed the synergistic NGF production from the coculture of SCs and ASCs. It was concluded that coculture of SCs with ASCs or DPSCs in a conduit promoted peripheral nerve regeneration over a critical gap defect.

Key words: Schwann cells (SCs); Adipose-derived adult stem cells (ASCs); Dental pulp stem cells (DPSCs); Peripheral nerve injury; Nerve regeneration; Conduits

Received April 3, 2012; final acceptance September 7, 2012. Online prepub date: November 27, 2012.
Address correspondence to Shan-hui Hsu, Institute of Polymer Science and Engineering, National Taiwan University, No. 1, Sec. 4 Roosevelt Road, Taipei 10617, Taiwan, R.O.C. Tel: +886-2-33665313; Fax: +886-2-33665237; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2041-2051, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X663533
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Application of Ex Vivo Expanded Umbilical Cord-Derived Mesenchymal Stem Cells: Enhance Hematopoiesis After Cord Blood Transplantation

Kang-Hsi Wu,*† Chris Tsai,‡ Han-Ping Wu,§ Martin Sieber,‡ Ching-Tien Peng,†¶ and Yu-Hua Chao#**††

*School of Chinese Medicine, China Medical University, Taichung, Taiwan
†Department of Pediatrics, China Medical University Hospital, Taichung, Taiwan
‡Bionet Corp, Taipei, Taiwan
§Department of Pediatrics, Buddhist Tzu-Chi General Hospital, Taichung Branch, Taichung, Taiwan
¶Department of Biotechnology and Bioinformatics, Asia University, Taichung, Taiwan
#Department of Pediatrics, Chung Shan Medical University Hospital, Taichung, Taiwan
**Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan
††School of Medicine, Chung Shan Medical University, Taichung, Taiwan

Delayed hematopoietic reconstitution after cord blood (CB) transplantation (CBT) needs to be overcome. Bone marrow-derived mesenchymal stem cells (BMMSCs) have been found to enhance engraftment after hematopoietic stem cell transplantation. However, getting BMMSCs involves an invasive procedure. In this study, umbilical cord-derived mesenchymal stem cells (UCMSCs) were isolated from Wharton’s jelly and cryopreserved in the UCMSCs bank. Compared with BMMSCs, we found that UCMSCs had superior proliferative potential. We found that NOD/SCID mice cotransplanted with CB and UCMSCs demonstrated significant human CD45+ cell engraftment compared with those transplanted with CB alone. Then, 20 patients with high-risk leukemia were prospectively randomized to either receive cotransplantation of CB and ex vivo expanded banked UCMSCs or to receive CBT alone. No serious adverse events were observed in the patients receiving UCMSC infusion. The time to undergo neutrophil engraftment and platelet engraftment was significantly shorter in the eight patients receiving cotransplantation than that in the 12 patients receiving CBT alone (p = 0.003 and p = 0.004, respectively). Thus, application of ex vivo expanded banked UCMSCs in humans appears to be feasible and safe. UCMSCs can enhance engraftment after CBT, but further studies are warranted.

Key words: Mesenchymal stem cells (MSCs); Umbilical cord; Cord blood transplantation (CBT)

Received November 1, 2011; final acceptance September 23, 2012. Online prepub date: February 26, 2013.
Address correspondence to Yu-Hua Chao, M.D., Department of Pediatrics, Chung Shan Medical University Hospital, No. 110, Sec. 1, Chien-Kuo N. Road, Taichung 402, Taiwan. Tel: +886-4-24739595 ext. 21728; Fax: +886-4-24710934; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2053-2061, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658827
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Increasing Follicular and Stromal Cell Proliferation in Cryopreserved Human Ovarian Tissue After Long-Term Precooling Prior to Freezing: In Vitro Versus Chorioallantoic Membrane (CAM) Xenotransplantation

Vladimir Isachenko, Evgenia Isachenko, Peter Mallmann, and Gohar Rahimi

Research Group for Reproductive Medicine and IVF-Laboratory, CAM-Xenotransplantation Group, Department of Obstetrics and Gynecology, Cologne University, Cologne, Germany

A positive effect on the future development of cells, which have been cooled to low suprazero temperatures and then thawed, has been observed before and is not new. The aim of this study was to test the effectiveness of postthawing culture of human ovarian tissue, which was either frozen just after operative removal or cooled after removal to 5°C for 24 h before cryopreservation. Ovarian fragments from six patients were divided into small pieces in the form of cortex with medulla and randomly divided into the following four groups: Group 1 consisted of pieces that just after removal had been cultured in vitro for 8 days in a big volume of medium with mechanical agitation; Group 2 included pieces cooled after operation to 5°C for 24 h and then cultured in vitro for 8 days; Group 3 was comprised of pieces frozen–thawed just after operation and then cultured for 5 days in the chorioallantoic membrane (CAM) culture system; and the pieces in Group 4 were cooled after operation to 5°C for 24 h, frozen–thawed, and then cultured in the CAM system for 5 days. The effectiveness of the tissue culture was evaluated by the development of follicles and by the intensiveness of proliferation in the tissue (by expression of cytokeratin and Ki-67). For Groups 1, 2, 3, and 4, the mean densities of follicles per 1 mm3 was 12.9, 12.2, 12.4, and 16.1, respectively (p1–2 > 0.1; p3–4 < 0.05). For these groups, 87%, 95%, 71%, and 84% of the preantral follicles were morphologically normal (p1–2,3–4 < 0.05). The immunohistochemical analysis showed increased proliferation after cooling of fresh and cryopreserved tissue. Long-term (24 h) cooling of ovarian tissue to 5°C before cryopreservation increases the viability of the cells in the tissue after thawing. Additionally, the efficacy of the CAM system for the culture of thawed human ovarian tissue was demonstrated.

Key words: Human ovarian tissue; Freezing; Proliferation; Cytokeratin; Ki-67

Received May 22, 2012; final acceptance September 28, 2012. Online prepub date: November 1, 2012.
Address correspondence to Vladimir Isachenko, University Maternal Hospital, Department of Obstetrics and Gynecology, Cologne University, Kerpener Str. 34, 50931 Cologne, Germany. Tel: +49-221-4783975; Fax: +49-221-47886201; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2063-2077, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657855
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

A New Nonenzymatic Method and Device to Obtain a Fat Tissue Derivative Highly Enriched in Pericyte-Like Elements by Mild Mechanical Forces From Human Lipoaspirates

Francesca Bianchi,*† Margherita Maioli,*‡ Erika Leonardi,§ Elena Olivi,*† Gianandrea Pasquinelli,¶ Sabrina Valente,¶ Armando J. Mendez,§ Camillo Ricordi,§ Mirco Raffaini,# Carlo Tremolada,# and Carlo Ventura*†

*Laboratory of Molecular Biology and Stem Cell Engineering-National Institute of Biostructures and Biosystems, Bologna, Italy
†Cardiovascular Department, S. Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy
‡Department of Biomedical Sciences, University of Sassari, Sassari, Italy
§Diabetes Research Institute, Miller School of Medicine, University of Miami, Miami, FL, USA
¶Surgical Pathology Unit, Department of Hematology, Oncology and Clinical Pathology, University of Bologna, Bologna, Italy
#Istituto Image, Diabetes Research Institute [DRI] Federation, Milan, Italy

Adipose tissue contains multipotent elements with phenotypic and gene expression profiles similar to human mesenchymal stem cells (hMSCs) and pericytes. The chance of clinical translation of the multilineage potential of these cells is delayed by the poor/negligible cell survival within cryopreserved lipoaspirates, the difficulty of ex vivo expansion, and the complexity of current Good Manufacturing Practice (cGMP) requirements for expanded cells. Hence, availability of a minimally manipulated, autologous, hMSC/pericyte-enriched fat product would have remarkable biomedical and clinical relevance. Here, we present an innovative system, named Lipogems, providing a nonexpanded, ready-to-use fat product. The system uses mild mechanical forces in a completely closed system, avoiding enzymes, additives, and other manipulations. Differently from unprocessed lipoaspirate, the nonexpanded Lipogems product encompasses a remarkably preserved vascular stroma with slit-like capillaries wedged between adipocytes and stromal stalks containing vascular channels with evident lumina. Immunohistochemistry revealed that Lipogems stromal vascular tissue included abundant cells with pericyte/hMSC identity. Flow cytometry analysis of nonexpanded, collagenase-treated Lipogems product showed that it was comprised with a significantly higher percentage of mature pericytes and hMSCs, and lower amount of hematopoietic elements, than enzymatically digested lipoaspirates. Differently from the lipoaspirate, the distinctive traits of freshly isolated Lipogems product were not altered by cryopreservation. Noteworthy, the features of fresh product were retained in the Lipogems product obtained from human cadavers, paving the way to an off-the-shelf strategy for reconstructive procedures and regenerative medicine. When placed in tissue culture medium, the Lipogems product yielded a highly homogeneous adipose tissue-derived hMSC population, exhibiting features of hMSCs isolated from other sources, including the classical commitment to osteogenic, chondrogenic, and adipogenic lineages. Moreover, the transcription of vasculogenic genes in Lipogems-derived adipose tissue hMSCs was enhanced at a significantly greater extent by a mixture of natural provasculogenic molecules, when compared to hMSCs isolated from enzymatically digested lipoaspirates.

Key words: Adipose tissue; Lipoaspirates; Stromal vascular architecture; Stem cells; Nonenzymatic isolation

Received January 20, 2012; final acceptance September 22, 2012. Online prepub date: October 8, 2012.
Address correspondence to Prof. Carlo Ventura, Laboratory of Molecular Biology and Stem Cell Engineering-National Institute of Biostructures and Biosystems, Strada Maggiore 42, 40125 Bologna, Italy. Tel/Fax: +39-051-340339; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2079-2090, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657927
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Embryonic Stem Cell-Derived Endothelial Cells as Cellular Delivery Vehicles for Treatment of Metastatic Breast Cancer

Weijun Su,*1 Lina Wang,*†1 Manqian Zhou,*‡ Ze Liu,* Shijun Hu,§ Lingling Tong,* Yanhua Liu,* Yan Fan,* Deling Kong,¶ Yizhou Zheng,† Zhongchao Han,† Joseph C. Wu,§ Rong Xiang,* and Zongjin Li*¶

*Nankai University School of Medicine, Tianjin, China
†State Key Lab of Experimental Hematology, Chinese Academy of Medical Sciences, Tianjin, China
‡Department of Oncology, Tianjin People’s Hospital, Tianjin, China
§Department of Medicine, Division of Cardiology, Stanford University School of Medicine, Stanford, CA, USA
¶The Key Laboratory of Bioactive Materials, Ministry of Education, Nankai University, The College of Life Science, Tianjin, China

Endothelial progenitor cells (EPCs) have shown tropism towards primary tumors or metastases and are thus potential vehicles for targeting tumor therapy. However, the source of adult EPCs is limited, which highlights the need for a consistent and renewable source of endothelial cells for clinical applications. Here, we investigated the potential of human embryonic stem cell-derived endothelial cells (hESC-ECs) as cellular delivery vehicles for therapy of metastatic breast cancer. In order to provide an initial assessment of the therapeutic potency of hESC-ECs, we treated human breast cancer MDA-MB-231 cells with hESC-EC conditioned medium (EC-CM) in vitro. The results showed that hESC-ECs could suppress the Wnt/β-catenin signaling pathway and thereby inhibit the proliferation and migration of MDA-MB-231 cells. To track and evaluate the possibility of hESC-EC-employed therapy, we employed the bioluminescence imaging (BLI) technology. To study the therapeutic potential of hESC-ECs, we established lung metastasis models by intravenous injection of MDA-MB-231 cells labeled with firefly luciferase (Fluc) and green fluorescent protein (GFP) to NOD/SCID mice. In mice with lung metastases, we injected hESC-ECs armed with herpes simplex virus truncated thymidine kinase (HSV-ttk) intravenously on days 11, 16, 21, and 26 after MDA-MB-231 cell injection. The NOD/SCID mice were subsequently treated with ganciclovir (GCV), and the growth status of tumor was monitored by Fluc imaging. We found that MDA-MB-231 tumors were significantly inhibited by intravenously injected hESC-ECs. The tumor-suppressive effects of the hESC-ECs, by inhibiting Wnt/b-catenin signaling pathway and inducing tumor cell death through bystander effect in human metastatic breast cancer model, provide previously unexplored therapeutic modalities for cancer treatment.

Key words: Human embryonic stem cells (hESCs); Endothelial cells (ECs); Molecular imaging; Cellular vehicle; Cancer therapy

Received February 23, 2012; final acceptance October 4, 2012. Online prepub date: October 12, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Zongjin Li, M.D., Ph.D., Nankai University School of Medicine, 94 Weijin Road, Tianjin, 300071, China. Tel: +86-22-23509475; Fax: +86-22-23509505; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Rong Xiang, M.D., Ph.D., Nankai University School of Medicine, 94 Weijin Road, Tianjin, 300071, China. Tel: +86-22-23509505; Fax: +86-22-23509505; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2091-2103, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658700
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Decompression of Inflammatory Edema Along With Endothelial Cell Therapy Expedites Regeneration After Renal Ischemia-Reperfusion Injury

Tanja Herrler,* Hao Wang,† Anne Tischer,* Nina Schupp,‡ Sebastian Lehner,† Andreas Meyer,* Jens Wallmichrath,* Antje Habicht,‡ Bechara Mfarrej,‡ Hans-Joachim Anders,§ Peter Bartenstein,† Karl-Walter Jauch,* Marcus Hacker,† and Markus Guba*

*Department of Surgery, Campus Großhadern, University of Munich, Munich, Germany
†Department of Nuclear Medicine, University of Munich, Munich, Germany
‡Transplantation Center, University of Munich, Munich, Germany
§Nephrologisches Zentrum, Medizinische Klinik und Poliklinik IV, Klinikum der Universität München, Universität München, Munich, Germany

Increased pressure due to postischemic edema aggravates renal ischemia-reperfusion injury (IRI). Prophylactic surgical decompression using microcapsulotomy improves kidney dysfunction after IRI. Supportive cell therapy in combination with microcapsulotomy might act synergistically protecting kidney function against IRI. The effects of therapeutic endothelial cell application alone and in combination with microcapsulotomy were investigated in a xenogenic murine model of 45-min warm renal ischemia. Renal function and perfusion were determined before as well as 2 and 18 days postischemia by 99mTc-MAG3 imaging and laser Doppler. Histological analysis included H&E stains and immunohistology for endothelial marker MECA-32, cell proliferation marker Ki-67, and macrophage marker F4/80. Histomorphological changes were quantified using a tubular injury score. Ischemia of 45 min led to severe tissue damage and a significant decrease in renal function and perfusion. Microcapsulotomy and cell therapy alone had no significant effect on renal function, while only surgical decompression significantly increased blood flow in ischemic kidneys. However, the combination of both microcapsulotomy and cell therapy significantly improved kidney function and perfusion. Combination therapy significantly reduced morphological injury of ischemic kidneys as determined by a tubular injury score and MECA-32 staining. Macrophage infiltration evidenced by F4/80 staining was significantly reduced. The Ki-67 proliferation index was increased, suggesting a regenerative environment. While microcapsulotomy and cell therapy alone have limited effect on renal recovery after IRI, combination therapy showed synergistic improvement of renal function, perfusion, and structural damage. Microcapsulotomy may create a permissive environment for cell therapy to work.

Key words: Cell therapy decompression; Endothelial cells; Ischemia-reperfusion injury (IRI); Kidney; Renal function

Received November 9, 2011; final acceptance September 30, 2012. Online prepub date: October 31, 2012.
Address correspondence to Tanja Herrler, M.D., Department of Surgery, Campus Großhadern, University of Munich, Munich, Germany. Tel: +49-89-7095-0; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2105-2118, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X661300
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Design of Biomimetic Vascular Grafts With Magnetic Endothelial Patterning

Delphine Fayol,* Catherine Le Visage,† Julia Ino,† Florence Gazeau,* Didier Letourneur,† and Claire Wilhelm*

*Laboratoire Matière et Systèmes Complexes (MSC), UMR 7057 CNRS and Université Paris Diderot, Paris, France
†Inserm, U698, Bio-ingénierie Cardiovasculaire, Université Paris Diderot, CHU X. Bichat, Paris, France

The development of small diameter vascular grafts with a controlled pluricellular organization is still needed for effective vascular tissue engineering. Here, we describe a technological approach combining a tubular scaffold and magnetically labeled cells to create a pluricellular and organized vascular graft, the endothelialization of which could be monitored by MRI prior to transplantation. A novel type of scaffold was developed with a tubular geometry and a porous bulk structure enabling the seeding of cells in the scaffold pores. A homogeneous distribution of human mesenchymal stem cells in the macroporous structure was obtained by seeding the freeze-dried scaffold with the cell suspension. The efficient covering of the luminal surface of the tube was then made possible thanks to the implementation of a magnetic-based patterning technique. Human endothelial cells or endothelial progenitors were magnetically labeled with iron oxide nanoparticles and successfully attracted to the 2-mm lumen where they attached and formed a continuous endothelium. The combination of imaging modalities [fluorescence imaging, histology, and 3D magnetic resonance imaging (MRI)] evidenced the integrity of the vascular construct. In particular, the observation of different cell organizations in a vascular scaffold within the range of resolution of single cells by 4.7 T MRI is reported.

Key words: Biomimetic material; Scaffold; Endothelial cells; Mesenchymal stem cells (MSCs); Endothelialization; Iron oxide nanoparticles; Magnetic patterning; Magnetic resonance imaging (MRI)

Received November 9, 2011; final acceptance September 22, 2012. Online prepub date: January 2, 2013.
Address correspondence to Claire Wilheim, Laboratoire Matière et Systèmes Complexes (MSC), UMR 7057 CNRS and Université Paris Diderot, Paris, France. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2119-2133, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657701
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Revascularization of Pancreatic Islet Allografts Is Enhanced by α-1-Antitrypsin Under Anti-Inflammatory Conditions

Keren Bellacen, Noa Kalay, Eyal Ozeri, Galit Shahaf, and Eli C. Lewis

Faculty of Health Sciences, Department of Clinical Biochemistry and Pharmacology, Ben-Gurion University of the Negev, Beer-Sheva, Israel

Pancreatic islets are a highly vascularized entity, and their transplantation into diabetic individuals requires optimal revascularization. In addition, β-cells in islets are extremely sensitive to inflammation. α-1-Antitrypsin (AAT), a circulating serine-protease inhibitor that is available for clinical use as an affinity-purified human product, has been shown to protect islets from graft failure in mouse transplantation models and to achieve readily vascularized islet grafts. AAT is known to induce vascular endothelial growth factor (VEGF) expression and release, as well as protect from proteolytic cleavage of VEGF by elastase, promote viability of endothelial cells, and enhance migration of myocytes. Our aim was to examine whether AAT enhances vasculogenesis toward islet grafts. We employed Matrigel-islet plugs as means to introduce islets in an explantable isolated compartment and examined vessel formation, vessel maturation, and inflammatory profile of explants 9 days after implantation. Also, we examined primary epithelial cell grafts that were prepared from lungs of mice that are transgenic for human AAT. In addition, aortic ring sprouting assay was performed, and HUVEC tube formation assays were studied in the presence of AAT. Our findings indicate that islet grafts exhibit mature vessels in the presence of AAT, as demonstrated by morphology, as well as expression of endothelial CD31, smooth muscle actin (SMA), and von Willebrand factor (vWF). Epithelial cells that express human AAT achieved a similar positive outcome. Aortic ring sprouting was enhanced in AAT-treated cultures and also in cultures that contained primary epithelial cells from human AAT transgenic animals in the absence of added AAT. According to the tube formation assay, HUVECs exhibited superior responses in the presence of AAT. We conclude that vasculogenesis toward islet grafts is enhanced in the presence of AAT. Together with the remarkable safety profile of AAT, the study supports its use in the relevant clinical setups.

Key words: Diabetes; Angiogenesis; Inflammation; Transgenic; Transplantation

Received January 12, 2011; final acceptance September 14, 2012. Online prepub date: October 8, 2012.
Address correspondence to Eli C. Lewis, Faculty of Health Sciences, Department of Clinical Biochemistry and Pharmacology, Ben-Gurion University of the Negev, POB 151, Beer-Sheva, Israel. Tel/Fax: +972-8-647-9981; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2135-2145, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657891
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Overexpression of Galectin-9 in Islets Prolongs Grafts Survival via Downregulation of Th1 Responses

Feng-Cheng Chou,* Chih-Chi Kuo,† Yen-Ling Wang,‡ Ming-Hong Lin,† B. Linju Yen,§ Deh-Ming Chang,¶ and Huey-Kang Sytwu*

*Department and Graduate Institute of Microbiology and Immunology, National Defense Medical Center, Taipei, Taiwan
†Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei, Taiwan
‡Center for Vascularized Composite Allotransplantation, Chang Gung Memorial Hospital, Linkou, Taiwan
§Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan
¶Department of Internal Medicine, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan

The differential activation of T helper (Th) cells and production of cytokines contribute to graft rejection or tolerance. In general, the Th1-type cytokines and cytotoxic T-cells are detected consistently in a host who is undergoing rejection, whereas Th2 responses are linked to a tolerance condition. Galectin-9 modulates Th1 cell immunity by binding to the T-cell immunoglobulin mucin-3 (Tim-3) molecule expressed on the Th1 cells. We investigate whether overexpression of galectin-9 in islets prolongs grafts survival in diabetic recipients. Islets were transduced with lentiviruses carrying galectin-9 and were then transplanted to streptozotocin-induced diabetic NOD/SCID recipients. The normoglycemic recipients then received splenocytes from diabetic NOD mice. Blood glucose concentration was monitored daily after adoptive transfer. The histology of the islet grafts and flow cytometric analyses were assessed at the end of the study. Overexpression of galectin-9 in islets prolonged graft survival in NOD/SCID mice after challenge with diabetogenic splenocytes (mean graft survival, 38.5 vs. 26.0 days, n = 10, respectively; p = 0.0096). The galectin-9-overexpressed grafts showed decreased infiltration of IFN-γ-producing CD4+ and CD8+ T-cells, but not of IL-17-producing CD4+ T-cells. Strikingly, this islet-specific genetic manipulation did not affect the systemic lymphocyte composition, indicating that galectin-9 may regulate T-cell-mediated inflammation in situ. We demonstrate that galectin-9 protects grafts from Th1 and Tc1 cell-mediated rejections, suggesting that galectin-9 has preventive and/or therapeutic benefit in transplant therapy for autoimmune diabetes and may be applied further to the transplantation of other organs or tissues.

Key words: Islet transplantation; Galectin-9; T-cell immunoglobulin mucin-3 (Tim-3); T helper 1 (Th1) cells; Autoimmune diabetes

Received November 16, 2011; final acceptance September 29, 2012. Online prepub date: October 12, 2012.
Address correspondence to Huey-Kang Sytwu, M.D., Ph.D., Department and Graduate Institute of Microbiology and Immunology, National Defense Medical Center, 161, Section 6, MinChuan East Road, Neihu, Taipei 114 Taiwan. Tel: +886-2-87923100, ext. 18540; Fax: +886-2-87921774; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2147-2159, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658728
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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A Preexistent Hypoxic Gene Signature Predicts Impaired Islet Graft Function and Glucose Homeostasis

James Cantley,*†1 Stacey N. Walters,†‡1 Min-Ho Jung,§ Anita Weinberg,†‡ Mark J. Cowley,†¶ Tess P. Whitworth,*† Warren Kaplan,†# Wayne J. Hawthorne,** Philip J. O’Connell,** Gordon Weir,§ and Shane T. Grey†‡

*Diabetes and Obesity Research Program, Garvan Institute, Darlinghurst, New South Wales, Australia
†St. Vincent’s Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
‡Immunology Program, Garvan Institute, Darlinghurst, New South Wales, Australia
§Islet Cell and Regenerative Biology, Joslin Diabetes Center, Boston, MA, USA
¶Cancer Program, Garvan Institute, Darlinghurst, New South Wales, Australia
#Peter Wills Bioinformatics Centre, Garvan Institute, Darlinghurst, New South Wales, Australia
**The Centre for Transplant and Renal Research, Westmead Hospital, Westmead, New South Wales, Australia

We examined whether hypoxic exposure prior to the event of transplantation would have a positive or negative effect upon later islet graft function. Mouse islets exposed to hypoxic culture were transplanted into syngeneic recipients. Islet graft function, β-cell physiology, as well as molecular changes were examined. Expression of hypoxia-response genes in human islets pre- and posttransplant was examined by microarray. Hypoxia-preexposed murine islet grafts provided poor glycemic control in their syngeneic recipients, marked by persistent hyperglycemia and pronounced glucose intolerance with failed first- and second-phase glucose-stimulated insulin secretion in vivo. Mechanistically, hypoxic preexposure stabilized HIF-1α with a concomitant increase in hypoxic-response genes including LDHA, and a molecular gene set, which would favor glycolysis and lactate production and impair glucose sensing. Indeed, static incubation studies showed that hypoxia-exposed islets exhibited dysregulated glucose responsiveness with elevated basal insulin secretion. Isolated human islets, prior to transplantation, express a characteristic hypoxia-response gene expression signature, including high levels of LDHA, which is maintained posttransplant. Hypoxic preexposure of an islet graft drives a HIF-dependent switch to glycolysis with subsequent poor glycemic control and loss of GSIS. Early intervention to reverse or prevent these hypoxia-induced metabolic gene changes may improve clinical islet transplantation.

Key words: Islet transplantation; Hypoxia; Glycolysis; Hypoxia-inducible factor-1α (HIF-1α)

Received March 5, 2012; final acceptance September 14, 2012. Online prepub date: October 31, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Shane T. Grey, Gene Therapy and Autoimmunity Group, Immunology Program, Garvan Institute of Medical Research, 384 Victoria Street, Darlinghurst, New South Wales 2010, Australia. Tel: +011-61-2-9295 8104; Fax: +011-61-2-9295 8404; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2161-2173, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657864
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Improvement of Pig Islet Function by In Vivo Pancreatic Tissue Remodeling: A “Human-Like” Pig Islet Structure With Streptozotocin Treatment

Sophie Vériter,* Najima Aouassar,† Gwen Beaurin,* Rose-Marie Goebbels,† Pierre Gianello,* and Denis Dufrane†

*Pôle de Chirurgie Expérimentale et Transplantation, Laboratory of Experimental Surgery, Université Catholique de Louvain, Secteur des Sciences de la Santé, Brussels, Belgium
†Endocrine Cell Therapy Unit, Center of Tissular and Cellular Therapy, St. Luc Hospital, Brussels, Belgium

Pig islets demonstrate significantly lower insulin secretion after glucose stimulation than human islets (stimulation index of ~12 vs. 2 for glucose 1 and 15 mM, respectively) due to a major difference in β- and α-cell composition in islets (60% and 25% in humans and 90% and 8% in pigs, respectively). This leads to a lower rise in 3′,5′-cyclic adenosine monophosphate (cAMP) in pig β-cells. Since glucagon is the major hormonal effector of cAMP in b-cells, we modified pig islet structure in vivo to increase the proportion of α-cells per islet and to improve insulin secretion. Selected doses (0, 30, 50, 75, and 100 mg/kg) of streptozotocin (STZ) were intravenously injected in 32 young pigs to assess pancreatic (insulin and glucagon) hormone levels, islet remodeling (histomorphometry for α- and β-cell proportions), and insulin and glucagon secretion in isolated islets. Endocrine structure and hormonal content of pig islets were compared with those of human islets. The dose of STZ was significantly correlated with reductions in pancreatic insulin content (p < 0.05, r2 = 0.77) and the proportion of β-cells (p < 0.05, r2 = 0.88). A maximum of 50 mg/kg STZ was required for optimal structure remodeling, with an increased proportion of α-cells per islet (26% vs. 48% α-cells per islet for STZ <50 mg/kg vs. >75 mg/kg; p < 0.05) without β-cell dysfunction. Three months after STZ treatment (30/50 mg/kg STZ), pig islets were isolated and compared with isolated control islets (0 mg/kg STZ). Isolated islets from STZ-treated (30/50 mg/kg) pigs had a higher proportion of α-cells than those from control animals (32.0% vs. 9.6%, respectively, p < 0.05). After in vitro stimulation, isolated islets from STZ-treated pigs demonstrated significantly higher glucagon content (65.4 vs. 21.0 ng/ml, p < 0.05) and insulin release (144 μU/ml) than nontreated islets (59 μU/ml, p < 0.05), respectively. Low-dose STZ (<50 mg/kg) can modify the structure of pig islets in vivo and improve insulin secretion after isolation.

Key words: Pig islet xenotransplantation; Islet structure; Pancreatic tissue remodeling; α-Cell; β-Cell stimulation

Received February 13, 2012; final acceptance August 21, 2012. Online prepub date: October 8, 2012.
Address correspondence to Denis Dufrane, M.D., Ph.D., Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain/CHEX, Avenue Hippocrate 55/70, B-1200 Brussels, Belgium. Tel: +32 (0)2 764 55 87; Fax: +32 (0)2 764 95 20; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 2175-2186, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657783
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Pancreatic Islet Preparations Release HMGB1: (Ir)Relevance for Graft Engraftment

Rita Nano,* Leda Racanicchi,* Raffaella Melzi,* Alessia Mercalli,* Paola Maffi,† Valeria Sordi,* Zhidong Ling,‡ Marina Scavini,* Olle Korsgren,§ Barbara Celona,¶ Antonio Secchi,†# and Lorenzo Piemonti*

*San Raffaele Diabetes Research Institute (HSR-DRI), San Raffaele Scientific Institute, Milan, Italy
†Transplant Unit, Department of Medicine, San Raffaele Scientific Institute, Milan, Italy
‡Diabetes Research Center, Brussels Free University-VUB, Brussels, Belgium
§Department of Immunology, Genetics and Pathology, Division of Immunology, Uppsala University, Uppsala, Sweden
¶Division of Genetics and Cell Biology, San Raffaele Scientific Institute, Milan, Italy
#University Vita-Salute San Raffaele, Milan, Italy

High levels of donor-derived high-mobility group box 1 (HMGB1) protein have been associated with poor islet graft outcome in mouse models. The aim of our work was to determine whether HMGB1 released by human islets had independent proinflammatory effects that influence engraftment in humans. Human islet preparations contained and released HMGB1 in different amounts, as determined by Western blot and ELISA (median 17 pg/ml/IEQ/24 h; min–max 0–211, n = 74). HMGB1 release directly correlated with brain death, donor hyperamilasemia, and factors related to the pancreas digestion procedure (collagenase and digestion time). HMGB1 release was significantly positively associated with the release of other cytokines/chemokines, particularly with the highly released “proinflammatory” CXCL8/IL-8, CXCL1/GRO-α, and the IFN-γ-inducible chemokines CXCL10/IP-10 and CXCL9/MIG. HMGB1 release was not modulated by Toll-like receptor 2, 3, 4, 5, and 9 agonists or by exposure to IL-1β. When evaluated after islet transplantation, pretransplant HMGB1 release was weakly associated with the activation of the coagulation cascade (evaluated as serum cross-linked fibrin products), but not with the immediate posttransplant inflammatory response. Concordantly, HMGB1 did not affect short-term human islet function. Our data show that human islet HMGB1 release is a sign of “damaged” islets, although without any independent direct role in graft failure.

Key words: Islet transplantation; Engraftment; High-mobility group box 1 (HMGB1); Human

Received March 19, 2012; final acceptance September 11, 2012. Online prepub date: October 8, 2012.
Address correspondence to Rita Nano or 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