Cell Transplantation 22(9) Abstracts

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Cell Transplantation, Vol. 22, pp. 1507-1517, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X661292
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Lentiviral-Encoded shRNA Silencing of Proteoglycan Decorin Enhances Tendon Repair and Regeneration Within a Rat Model

Ping Lu,*†‡1 Guo-Rong Zhang,*†§1 You-Zhi Cai,*†‡ Boon Chin Heng,¶ Hao Ren,*†‡ Lin-Lin Wang,*†‡ Junfeng Ji,‡# Xiao-Hui Zou,‡# and Hong-Wei Ouyang*†‡**

*Center for Stem Cells and Tissue Engineering, School of Medicine, Zhejiang University, Hangzhou, China
†Department of Sports Medicine, Zhejiang University, Hangzhou, China
‡Zhejiang Provincial Key Laboratory of Tissue Engineering and Regenerative Medicine, Hangzhou, China
§Department of Basic Medicine, Changchun Medical College, Changchun, China
¶Department of Biosystems Science and Engineering, ETH-Zurich, Switzerland
#Central Laboratory, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China
**Department of Orthopaedic Surgery, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China

Injured tendons often heal with scar tissue formation, resulting in uniformly smaller collagen fibrils and poor mechanical properties. The small leucine-rich proteoglycan decorin is well known to regulate fusion of collagen fibrils. Rat patellar tendon cells were transfected with lentiviral-encoded shRNA that specifically targets decorin. Silencing of decorin expression resulted in decreased cell growth. Three types of scaffold-free engineered tendons with different mix ratios of anti-decorin shRNA-treated cells to untreated cells at 1:0 (DCN), 1:1 (MIX), and 0:1 (CON) were utilized for repair of injured patellar tendons. Four weeks after implantation in situ, the MIX group manifested the best results (best coordination of histology, more mature collagen deposition, and larger collagen fibril diameter). Although the DCN group exhibited the largest collagen fibril diameter, this was associated with abnormal shape. Hence, regulation of decorin expression to an appropriate level is crucial for tendon repair with gene therapy.

Key words: Decorin; Short hairpin RNA (shRNA); Lentivirus; Patellar tendon; Engineered tendon

Received December 5, 2011; final acceptance August 20, 2012. Online prepub date: January 2, 2013.
1These authors provided equal contribution to this work.
Address correspondence to Xiao-Hui Zou, Center for Stem Cell and Tissue Engineering, School of Medicine, Zhejiang University, Yu Hang Tang Road #866, Hangzhou, China 310058. Tel/Fax: 0086-0571-88208262; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1519-1528, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X647261
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Cartilage Tissue Formation From Dedifferentiated Chondrocytes by Codelivery of BMP-2 and SOX-9 Genes Encoding Bicistronic Vector

Byung-Hyun Cha,* Jae-Hwan Kim,* Sun-Woong Kang,* Hyun-Jin Do,* Ju-Woong Jang,† Yon Rak Choi,† Hansoo Park,‡ Byung-Soo Kim,§ and Soo-Hong Lee*

*Department of Biomedical Science, CHA University, Seoul, Republic of Korea
†Korea Bone Bank, Seoul, Republic of Korea
‡Department of Integrative Engineering, Chung-Ang University, Seoul, Republic of Korea
§School of Chemical and Biological Engineering, Seoul National University, Seoul, Republic of Korea

Articular cartilage, when damaged by degenerative disease or trauma, has limited ability for self-repair. Recently, many trials have demonstrated that gene therapy combined with tissue engineering techniques would be a promising approach for cartilage regeneration. Bone morphogenetic protein 2 (BMP-2) is an important signal for upregulation of osteogenesis and chondrogenesis of stem cells. Sex-determining region Y box gene 9 (SOX-9) has also been reported as one of the key transcription factors for chondrogenesis. We hypothesized that codelivery of BMP-2 and SOX-9 genes would result in improved efficiency of recovery of normal chondrogenic properties in dedifferentiated chondrocytes. To this aim, we constructed a bicistronic vector encoding the BMP-2 and SOX-9 genes linked to the “self-cleaving” 2A peptide sequence. After gene delivery to dedifferentiated chondrocytes using a microporator transfection system, we confirmed over 65% delivery efficiency of the BMP-2 and SOX-9 genes. According to RT-PCR analysis and Alcian blue staining, simultaneous delivery of BMP-2/SOX-9 resulted in significantly increased expression of chondrogenesis-related markers (type II collagen and aggrecan) and GAG matrix formation compared with individual delivery of the BMP-2 or SOX-9 gene. Six weeks after in vivo transplantation, BMP-2/SOX-9 genes also showed a significant increase in cartilage formation compared with the BMP-2 or SOX-9 gene. These results demonstrate that codelivery of two chondrogenic lineage-determining genes can enhance normal chondrogenic properties of dedifferentiated chondrocytes followed by improved cartilage formation.

Keywords: Bone morphogenetic protein 2 (BMP-2); Sex-determining region Y box gene 9 (SOX-9); Codelivery; Dedifferentiation; Cartilage tissue formation

Received May 16, 2011; final acceptance September 20, 2011. Online prepub date: July 5, 2012.
Address correspondence to Dr. Soo-Hong Lee, Department of Biomedical Science, CHA University, 502 Acecord Building 3rd Floor, Yatop-dong, Bundang-gu, Gyunggi-do 463-840, Korea. Tel: +82-31-8017-9415; Fax: +82-31-8017-9892; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1529-1539, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657693
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Glial Cell Line-Derived Neurotrophic Factor Partially Ameliorates Motor Symptoms Without Slowing Neurodegeneration in Mice With Respiratory Chain-Deficient Dopamine Neurons

Fredrik H. Sterky,*†1 Karin Pernold,†1 Brandon K. Harvey,‡ Eva Lindqvist,† Barry J. Hoffer,§ and Lars Olson†

*Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden
†Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden
‡Intramural Research Program, National Institute on Drug Abuse, NIH, Baltimore, MD, USA
§Department of Neurosurgery, Case Western Reserve University School of Medicine, Cleveland, OH, USA

Degeneration of midbrain dopamine neurons causes the striatal dopamine deficiency responsible for the hallmark motor symptoms of Parkinson’s disease (PD). Intraparenchymal delivery of neurotrophic factors, such as glial cell line-derived neurotrophic factor (GDNF), is a possible future therapeutic approach. In animal PD models, GDNF can both ameliorate neurodegeneration and promote recovery of the dopamine system following a toxic insult. However, clinical studies have generated mixed results, and GDNF has not been efficacious in genetic animal models based on α-synuclein overexpression. We have tested the response to GDNF in a genetic mouse PD model with progressive degeneration of dopamine neurons caused by mitochondrial impairment. We find that GDNF, delivered to the striatum by either an adeno-associated virus or via miniosmotic pumps, partially alleviates the progressive motor symptoms without modifying the rate of neurodegeneration. These behavioral changes are accompanied by increased levels of dopamine in the midbrain, but not in striatum. At high levels, GDNF may instead reduce striatal dopamine levels. These results demonstrate the therapeutic potential of GDNF in a progressively impaired dopamine system.

Key words: Glial cell line-derived neurotrophic factor (GDNF); Mitochondria; Parkinson’s disease (PD); Trophic support; Gene therapy

Received April 4, 2012; final acceptance August 2, 2012. Online prepub date: October 8, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Lars Olson, Department of Neuroscience, Karolinska Institutet, Retzius väg 8, SE-17177 Stockholm, Sweden. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1541-1552, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657936
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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In Vivo and In Vitro Characterization of the Angiogenic Effect of CTX0E03 Human Neural Stem Cells

Caroline Hicks,1 Lara Stevanato,1 Robert P. Stroemer, Ellen Tang, Sheila Richardson, and John D. Sinden

ReNeuron Limited, Surrey Research Park, Guildford, Surrey, UK

CTX0E03 is a human neural stem cell line previously reported to reduce sensory motor deficits in a middle cerebral artery occlusion (MCAo) model of stroke. The objective of this study was to investigate if CTX0E03 treatment promotes angiogenesis. As stroke leads to damage of the vasculature in the brain, angiogenesis may contribute to the functional recovery. To test this hypothesis, the angiogenic activity of CTX0E03 was assessed both in vitro and in vivo. In vitro, CTX0E03 expression of trophic and proangiogenic factors was determined by real-time RT-PCR, Western blot, and ELISA, and its angiogenic activity was investigated in well-established angiogenesis assays. In vivo, angiogenesis was investigated in naive mice and MCAo rat brain and was evaluated by immunohistochemistry (IHC) using Von Willebrand factor (VWF), a marker of blood vessel formation, and BrdU/CD31 double labeling in naive mice only. In vitro results showed that CTX0E03-conditioned medium and coculture significantly increased total tubule formation compared with controls (p = 0.002 and p = 0.0008, respectively). Furthermore, CTX0E03 cells were found to be in direct association with the tubules by ICC. In vivo CTX0E03-treated brains demonstrated a significant increase in areas occupied by VWF-positive microvessels compared with vehicle-treated naive mice (two-way ANOVA, Interaction p < 0.05, Treatment p < 0.0001, Time p < 0.0) and MCAo rat (p = 0.001 unpaired t test, Welch’s correction). CTX0E03-treated naive mouse brains showed an increase in BrdU/CD31 colabeling. In conclusion, in vitro CTX0E03 cells express proangiogenic factors and may promote angiogenesis by both release of paracrine factors and direct physical interaction. Furthermore, in vivo CTX0E03-treated rodent brains exhibited a significant increase in microvessels at the site of implantation compared with vehicle-injected groups. Taken together these data suggest that CTX0E03 cell therapy may provide significant benefit to stroke patients through upregulation of angiogenesis in the ischemic brain.

Key words: Neural stem cells; Trophic factors; Angiogenesis; Cell transplantation; Stroke

Received November 18, 2011; final acceptance October 8, 2012. Online prepub date: October 12, 2012.
1These authors share first authorship.
Address correspondence to John D. Sinden, Ph.D., CSO ReNeuron, 10 Nugent Road, Surrey Research Park, Guildford, Surrey, GU2 7AF, UK. Tel: +44-1483-269119; Fax: +44-1483-534864; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1553-1568, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X662390
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Environmental Enrichment Synergistically Improves Functional Recovery by Transplanted Adipose Stem Cells in Chronic Hypoxic-Ischemic Brain Injury

Jung Hwa Seo,*†1 Hyongbum Kim,‡1 Eun Sook Park,* Jong Eun Lee,§¶ Dong Wook Kim,¶# Hyun Ok Kim,**†† Sang Hee Im,*‡‡ Ji Hea Yu,*¶ Ji Yeon Kim,*,** Min-Young Lee,* Chul Hoon Kim,¶§§ and Sung-Rae Cho*†¶††¶¶

*Department and Research Institute of Rehabilitation Medicine, Yonsei University College of Medicine, Seoul, Korea
†Graduate Program of Nano Science and Technology, Yonsei University, Seoul, Korea
‡Graduate School of Biomedical Science and Engineering/College of Medicine, Hanyang University, Seoul, Korea
§Department of Anatomy, Yonsei University College of Medicine, Seoul, Korea
¶Brain Korea 21 Project for Medical Science, Yonsei University, Seoul, Korea
#Department of Physiology, Yonsei University College of Medicine, Seoul, Korea
**Department of Laboratory Medicine, Yonsei University College of Medicine, Seoul, Korea
††Yonsei Stem Cell Research Center, Avison Biomedical Research Center, Seoul, Korea
‡‡Department of Rehabilitation Medicine, Kwandong University College of Medicine, Seoul, Korea
§§Department of Pharmacology, Yonsei University College of Medicine, Seoul, Korea
¶¶Rehabilitation Institute of Neuromuscular Disease, Yonsei University College of Medicine, Seoul, Korea

We investigated the effects of environmental enrichment (EE) on the function of transplanted adipose stem cells (ASCs) and the combined effect of EE and ASC transplantation on neurobehavioral function in an animal model of chronic hypoxic-ischemic (HI) brain injury. HI brain damage was induced in 7-day-old mice by unilateral carotid artery ligation and exposure to hypoxia (8% O2 for 90 min). At 6 weeks of age, the mice were randomly injected with either ASCs or PBS into the striatum and were randomly assigned to either EE or standard cages (SC), comprising ASC-EE (n = 18), ASC-SC (n = 19), PBS-EE (n = 12), PBS-SC (n = 17), and untreated controls (n = 23). Rotarod, forelimb-use asymmetry, and grip strength tests were performed to evaluate neurobehavioral function. The fate of transplanted cells and the levels of endogenous neurogenesis, astrocyte activation, and paracrine factors were also measured. As a result, EE and ASC transplantation synergistically improved rotarod latency, forelimb-use asymmetry, and grip strength compared to those of the other groups. The number of engrafted ASCs and βIII-tubulin+ neurons derived from the transplanted ASCs was significantly higher in mice in EE than those in SC. EE and ASC transplantation also synergistically increased BrdU+βIII-tubulin+ neurons, GFAP+ astrocytic density, and fibroblast growth factor 2 (FGF2) level but not the level of CS-56+ glial scarring in the striatum. In conclusion, EE and ASC transplantation synergistically improved neurobehavioral functions. The underlying mechanisms of this synergism included enhanced repair processes such as higher engraftment of the transplanted ASCs, increased endogenous neurogenesis and astrocytic activation coupled with upregulation of FGF2.

Key words: Environmental enrichment (EE); Adipose stem cells (ASCs); Cell transplantation; Neurogenesis; Astrocyte; Fibroblast growth factor 2 (FGF2)

Received September 4, 2011; final acceptance January 20, 2012. Online prepub date: February 4, 2013.
1These authors provided equal contribution to this work.
Address correspondence to Sung-Rae Cho, M.D., Ph.D., Yonsei University College of Medicine, Department and Research Institute of Rehabilitation Medicine, 134 Shinchon-dong, Seodaemun-gu, Seoul, South Korea 120-752. Tel: +82-2-2228-3715; Fax: +82-2-363-2795; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1569-1576, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658674
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Intravenous Administration of Human Umbilical Tissue-Derived Cells Improves Neurological Function in Aged Rats After Embolic Stroke

Li Zhang,* Li Yi,* Michael Chopp,*† Brian C. Kramer,‡ Michael Romanko,‡ Anna Gosiewska,‡ and Klaudyne Hong‡

*Department of Neurology, Henry Ford Hospital, Detroit, MI, USA
†Department of Physics, Oakland University, Rochester, MI, USA
‡Advanced Technologies and Regenerative Medicine (ATRM), LLC, a Johnson & Johnson Company, Somerville, NJ, USA

Intravenous administration of human umbilical tissue-derived cells (hUTC) improves neurological function in young adult rats after stroke. However, stroke is a major cause of death and disability in the aged population, with the majority of stroke patients 65 years and older. The present study investigated the effect of hUTC on aged rats after embolic stroke. Rats at the age of 18–20 months were subjected to embolic middle cerebral artery (MCA) occlusion. Two groups of eight animals each were compared. The investigational group was injected intravenously with 1 × 107 cells/kg in serum-free culture medium (vehicle) 24 h after stroke onset, and the control group was treated with vehicle only at the same time poststroke. Intravenous administration of hUTC significantly improved neurological functional recovery without reducing infarct volume compared to vehicle-treated aged rats. Additionally, hUTC treatment significantly enhanced synaptogenesis and vessel density in the ischemic boundary zone (IBZ). Moreover, hUTC treatment resulted in a trend toward increased progenitor cell proliferation in the subventricular zone (SVZ) compared to vehicle-treated aged rats. Intravenous administration of hUTC improved functional recovery in aged rats after stroke. The enhancement of synaptogenesis and vessel density may contribute to the beneficial effects of hUTC in the treatment of stroke in the aged animal.

Key words: Cerebral infarct; Human umbilical cord; Synaptogenesis; Stroke recovery; Aged rat model

Received October 11, 2011; final acceptance August 24, 2012. Online prepub date: October 31, 2012.
Address correspondence to Michael Chopp, Ph.D., Henry Ford Hospital, 2799 W. Grand Boulevard, Detroit, MI 48202, USA. Tel: +1 313 916 2227; Fax: +1 313 916 1318; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1577-1590, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X659907
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Transplantation of Human Umbilical Cord Blood or Amniotic Epithelial Stem Cells Alleviates Mechanical Allodynia After Spinal Cord Injury in Rats

Dae-Hyun Roh,*1 Min-Soo Seo,†1,2 Hoon-Seong Choi,‡ Sang-Bum Park,† Ho-Jae Han,‡ Alvin J. Beitz,§ Kyung-Sun Kang,† and Jang-Hern Lee‡

*Department of Maxillofacial Tissue Regeneration, School of Dentistry, Kyung Hee University, Seoul, South Korea
†Adult Stem Cell Research Center, College of Veterinary Medicine, Seoul National University, Seoul, South Korea
‡Department of Veterinary Physiology, College of Veterinary Medicine and Research, Institute for Veterinary Science, Seoul National University, Seoul, South Korea
§Department of Veterinary and Biomedical Sciences, College of Veterinary Medicine, University of Minnesota, St. Paul, MN, USA

Stem cell therapy is a potential treatment for spinal cord injury (SCI), and a variety of different stem cell types have been grafted into humans suffering from spinal cord trauma or into animal models of spinal injury. Although several studies have reported functional motor improvement after transplantation of stem cells into injured spinal cord, the benefit of these cells for treating SCI-induced neuropathic pain is not clear. In this study, we investigated the therapeutic effect of transplanting human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) or amniotic epithelial stem cells (hAESCs) on SCI-induced mechanical allodynia (MA) and thermal hyperalgesia (TH) in T13 spinal cord hemisected rats. Two weeks after SCI, hUCB-MSCs or hAESCs were transplanted around the spinal cord lesion site, and behavioral tests were performed to evaluate changes in SCI-induced MA and TH. Immunohistochemical and Western blot analyses were also performed to evaluate possible therapeutic effects on SCI-induced inflammation and the nociceptive-related phosphorylation of the NMDA NR1 receptor subunit. While transplantation of hUCB-MSCs showed a tendency to reduce MA, transplantation of hAESCs significantly reduced MA. Neither hUCB-MSC nor hAESC transplantation had any effect on SCI-induced TH. Transplantation of hAESCs also significantly reduced the SCI-induced increase in NMDA receptor NR1 subunit phosphorylation (pNR1) expression in the spinal cord. Both hUCB-MSCs and hAESCs reduced the SCI-induced increase in spinal cord expression of the microglial marker, F4/80, but not the increased expression of GFAP or iNOS. Taken together, these findings demonstrate that the transplantation of hAESCs into the injured spinal cord can suppress mechanical allodynia, and this effect seems to be closely associated with the modulation of spinal cord microglia activity and NR1 phosphorylation.

Keywords: Spinal cord injury (SCI); Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs); Human amniotic epithelial stem cells (hAESCs); Mechanical allodynia (MA); NMDA receptor

Received May 23, 2012; final acceptance November 26, 2012. Online prepub date: January 2, 2013.
1These authors provided equal contribution to this work.
2Present address: Laboratory Animal Center, Daegu-Gyeongbuk Medical Innovation Foundation, Deagu, Korea.
Address correspondence to Kyung-Sun Kang, D.V.M., Ph.D., Adult Stem Cell Research Center, College of Veterinary Medicine, Seoul National University, Seoul 151-742, South Korea. Tel.: +82-2-880-1241; Fax: +82-2-876-7610; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Jang-Hern Lee, D.V.M., Ph.D., Department of Veterinary Physiology, College of Veterinary Medicine, Seoul National University, Seoul 151-742, South Korea. Tel: +82-2-880-1272; Fax: +82-2-885-2732; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1591-1612, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X663532
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Transplantation of Autologous Olfactory Ensheathing Cells in Complete Human Spinal Cord Injury

Pawel Tabakow,* Wlodzimierz Jarmundowicz,* Bogdan Czapiga,* Wojciech Fortuna,† Ryszard Miedzybrodzki,† Marcin Czyz,* Juliusz Huber,‡ Dariusz Szarek,* Stefan Okurowski,§ Pawel Szewczyk,¶ Andrzej Gorski,†# and Geoffrey Raisman**

*Department of Neurosurgery, Wroclaw Medical University, Wroclaw, Poland
†Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland
‡Department of Pathophysiology of Locomotor Organs, Karol Marcinkowski Medical University, Poznan, Poland
§Neurorehabilitation Center for Treatment of Spinal Cord Injuries AKSON, Wroclaw, Poland
¶Department of General Radiology, Interventional Radiology and Neuroradiology, Wroclaw Medical University, Wroclaw, Poland
#Transplantation Institute, Warsaw Medical University, Warsaw, Poland
**Spinal Repair Unit, UCL Institute of Neurology, Queen Square, London, UK

Numerous studies in animals have shown the unique property of olfactory ensheathing cells to stimulate regeneration of lesioned axons in the spinal cord. In a Phase I clinical trial, we assessed the safety and feasibility of transplantation of autologous mucosal olfactory ensheathing cells and olfactory nerve fibroblasts in patients with complete spinal cord injury. Six patients with chronic thoracic paraplegia (American Spinal Injury Association class A-ASIA A) were enrolled for the study. Three patients were operated, and three served as a control group. The trial protocol consisted of pre- and postoperative neurorehabilitation, olfactory mucosal biopsy, culture of olfactory ensheathing cells, and intraspinal cell grafting. Patient’s clinical state was evaluated by clinical, neurophysiological, and radiological tests. There were no adverse findings related to olfactory mucosa biopsy or transplantation of olfactory ensheathing cells at 1 year after surgery. There was no evidence of neurological deterioration, neuropathic pain, neuroinfection, or tumorigenesis. In one cell-grafted patient, an asymptomatic syringomyelia was observed. Neurological improvement was observed only in transplant recipients. The first two operated patients improved from ASIA A to ASIA C and ASIA B. Diffusion tensor imaging showed restitution of continuity of some white matter tracts throughout the focus of spinal cord injury in these patients. The third operated patient, although remaining ASIA A, showed improved motor and sensory function of the first spinal cords segments below the level of injury. Neurophysiological examinations showed improvement in spinal cord transmission and activity of lower extremity muscles in surgically treated patients but not in patients receiving only neurorehabilitation. Observations at 1 year indicate that the obtaining, culture, and intraspinal transplantation of autologous olfactory ensheathing cells were safe and feasible. The significance of the neurological improvement in the transplant recipients and the extent to which the cell transplants contributed to it will require larger numbers of patients.

Key words: Human; Olfactory ensheathing cells (OECs); Spinal cord injury (SCI); Transplantation

Received February 4, 2012; final acceptance August 22, 2012. Online prepub date: April 2, 2013.
Address correspondence to Pawel Tabakow, M.D., Ph.D., Department of Neurosurgery, Wroclaw Medical University, Borowska str. 213, 50-556 Wroclaw, Poland. Tel: +48 606 137 846; Fax: +48 71 734 34 09; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1613-1625, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X658791
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Transplantation of Bone Marrow Stromal Cell-Derived Neural Precursor Cells Ameliorates Deficits in a Rat Model of Complete Spinal Cord Transection

Misaki Aizawa-Kohama,*† Toshiki Endo,† Masaaki Kitada,* Shohei Wakao,* Akira Sumiyoshi,‡ Dai Matsuse,* Yasumasa Kuroda,§ Takahiro Morita,*† Jorge J. Riera,‡ Ryuta Kawashima,‡ Teiji Tominaga,† and Mari Dezawa*§

*Department of Stem Cell Biology and Histology, Tohoku University Graduate School of Medicine, Sendai, Japan
†Department of Neurosurgery, Tohoku University Graduate School of Medicine, Sendai, Japan
‡Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan
§Department of Anatomy and Anthropology, Tohoku University Graduate School of Medicine, Sendai, Japan

After severe spinal cord injury, spontaneous functional recovery is limited. Numerous studies have demonstrated cell transplantation as a reliable therapeutic approach. However, it remains unknown whether grafted neuronal cells could replace lost neurons and reconstruct neuronal networks in the injured spinal cord. To address this issue, we transplanted bone marrow stromal cell-derived neural progenitor cells (BM-NPCs) in a rat model of complete spinal cord transection 9 days after the injury. BM-NPCs were induced from bone marrow stromal cells (BMSCs) by gene transfer of the Notch-1 intracellular domain followed by culturing in the neurosphere method. As reported previously, BM-NPCs differentiated into neuronal cells in a highly selective manner in vitro. We assessed hind limb movements of the animals weekly for 7 weeks to monitor functional recovery after local injection of BM-NPCs to the transected site. To test the sensory recovery, we performed functional magnetic resonance imaging (fMRI) using electrical stimulation of the hind limbs. In the injured spinal cord, transplanted BM-NPCs were confirmed to express neuronal markers 7 weeks following the transplantation. Grafted cells successfully extended neurites beyond the transected portion of the spinal cord. Adjacent localization of synaptophysin and PSD-95 in the transplanted cells suggested synaptic formations. These results indicated survival and successful differentiation of BM-NPCs in the severely injured spinal cord. Importantly, rats that received BM-NPCs demonstrated significant motor recovery when compared to the vehicle injection group. Volumes of the fMRI signals in somatosensory cortex were larger in the BM-NPC-grafted animals. However, neuronal activity was diverse and not confined to the original hind limb territory in the somatosensory cortex. Therefore, reconstruction of neuronal networks was not clearly confirmed. Our results indicated BM-NPCs as an effective method to deliver neuronal lineage cells in a severely injured spinal cord. However, reestablishment of neuronal networks in completed transected spinal cord was still a challenging task.

Key words: Cell transplantation; Functional magnetic resonance imaging (fMRI); Bone marrow stromal cells (BMSCs); Neural progenitor cells (NPCs); Spinal cord injury (SCI)

Received October 7, 2011; final acceptance August 15, 2012. Online prepub date: October 31, 2012.
Address correspondence to Toshiki Endo, Department of Neurosurgery, Tohoku University Graduate School of Medicine, 1-1 Seiryo, Aoba, 980-8574, Sendai, Japan. Tel: +81-22-717-7230; Fax: +81-22-717-7233; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Mari Dezawa, Department of Stem Cell Biology and Histology and Department of Anatomy and Anthropology, Tohoku Univeristy Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, 980-8575, Sendai, Japan. Tel: +81-22-717-8025; Fax: +81-22-717-8030; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1627-1635, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657729
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
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Analyzing Migratory Properties of Human CD133+ Stem Cells In Vivo After Intraoperative Sternal Bone Marrow Isolation

Peter Donndorf,*1 Dritan Useini,*1 Cornelia A. Lux,* Brigitte Vollmar,† Evgenya Delyagina,* Michael Laupheimer,* Alexander Kaminski,* and Gustav Steinhoff*

*Reference and Translation Centre for Cardiac Stem Cell Therapy, Department for Cardiac Surgery, University of Rostock, Rostock, Germany
†Institute of Experimental Surgery, University of Rostock, Rostock, Germany

Human bone marrow stem cell populations have been applied for cardiac regeneration purposes within different clinical settings in the recent past. The migratory capacity of applied stem cell populations towards injured tissue, after undergoing specific peri-interventional harvesting and isolation procedures, represents a key factor limiting therapeutic efficacy. We therefore aimed at analyzing the migratory capacity of human cluster of differentiation (CD) 133+ bone marrow stem cells in vivo after intraoperative harvesting from the sternal bone marrow. Human CD133+ bone marrow stem cells were isolated from the sternal bone marrow of patients undergoing cardiac surgery at our institution. Migratory capacity towards stromal cell-derived factor-1α (SDF-1α) gradients was tested in vitro and in vivo by intravital fluoresecence microscopy, utilizing the cremaster muscle model in severe combined immunodeficient (SCID) mice and analyzing CD133+ cell interaction with the local endothelium. Furthermore, the role of a local inflammatory stimulus for CD133+ cell interaction with the endothelium was studied. In order to describe endothelial response upon chemokine stimulation laser scanning microscopy of histological cremaster muscle samples was performed. SDF-1α alone was capable to induce relevant early CD133+ cell interaction with the endothelium, indicated by the percentage of rolling CD133+ cells (45.9 ± 1.8% in “SDF-1” vs. 17.7 ± 2.7% in “control,” p < 0.001) and the significantly reduced rolling velocity after SDF-1α treatment. Furthermore, SDF-1α induced firm endothelial adhesion of CD133+ cells in vivo. Firm endothelial adhesion, however, was significantly enhanced by additional inflammatory stimulation with tumor necrosis factor-α (TNF-α) (27.9 ± 4.3 cells/mm2 in “SDF-1 + TNF” vs. 2.2 ± 1.1 cells/mm2 in “control,” p < 0.001). CD133+ bone marrow stem cells exhibit sufficient in vivo homing towards SDF-1α gradients in an inflammatory microenvironment after undergoing standardized intraoperative harvesting and isolation from the sternal bone marrow.

Key words: Bone marrow stem cell (BMSC); Isolation; Migration; Stromal cell-derived factor-1α (SDF-1α); Cardiac surgery

Received January 25, 2012; final acceptance August 22, 2012. Online prepub date: October 8, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Peter Donndorf, M.D., Department of Cardiac Surgery University of Rostock, Schillingallee 35, 18057 Rostock, Germany. Tel/Fax: +49-381494-6101/-6102; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1637-1650, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X661427
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Umbilical Cord Blood Mononuclear Cell-Conditioned Media Inhibits Hypoxic-Induced Apoptosis in Human Coronary Artery Endothelial Cells and Cardiac Myocytes by Activation of the Survival Protein Akt

Hua Jin,* Paul R. Sanberg,† and Robert J. Henning*

*Center for Cardiovascular Research and James A. Haley VA Medical Center, Tampa, FL, USA
†Center of Excellence for Aging and Brain Repair, University of South Florida Morsani College of Medicine, Tampa, FL, USA

We have previously demonstrated in acute myocardial infarctions that human umbilical cord blood mononuclear cells (HUCBCs), which contain hematopoietic, endothelial, and mesenchymal stem cells, reduce acute myocardial infarction size by ³50% and preserve LV contractility. We hypothesize that the beneficial effects of HUCBCs are due to secretion of biologically active factors that activate in cardiac endothelial cells and myocytes the cell survival protein Akt. We determined by protein microarrays the growth factors and anti-inflammatory cytokines secreted by HUCBCs into culture media during 12 h of hypoxia (1% O2). We then determined by Western blots the effects of cell-free media from hypoxic-conditioned HUCBCs (HUCM) on activation of the cell survival protein Akt in human coronary artery endothelial cells and cardiac myocytes in culture during 24 h of 1% O2. We also determined in separate experiments endothelial cell and myocyte apoptosis by caspase-3 and Annexin V. In the present experiments, HUCBCs secreted multiple growth factors, anti-inflammatory cytokines, and inhibitors of metalloproteinase during normoxia and hypoxia. Human cord blood cells increased the concentration in culture media of angiopoietin, hepatocyte growth factor, interleukin-4, insulin-like growth factor, placental growth factor, vascular endothelial cell growth factor, angiogenin, and stem cell factor by 100 to >10,000% during 12 h of 1% O2 (p < 0.001). HUCM, which contained these biological factors, significantly increased Akt phosphorylation/activation in coronary artery endothelial cells and cardiac myocytes subjected to 24 h of 1% O2 by more than 60% (p < 0.05) and increased the antiapoptotic protein Bcl-2 expression by 34–50% in comparison with endothelial cells and myocytes treated without HUCM in 1% O2 (p < 0.05). HUCM also significantly decreased caspase-3 activity and decreased hypoxic endothelial cell and cardiac myocyte apoptosis by more than 40% in comparison with cells cultured without HUCM (p < 0.05). Inhibition of Akt activation in endothelial cells and myocytes by the sensitive and specific antagonist API-1 during 24 h of hypoxia nearly completely prevented the beneficial effects of HUCM on inhibiting caspase-3 activity and apoptosis. We conclude that HUCBCs secrete biologically active factors during hypoxia that activate survival proteins in endothelial cells and myocytes that significantly limit apoptosis.

Key words: Umbilical cord cells; Stem cells; Paracrine actions; Growth factors; Anti-inflammatory cytokines; Coronary artery endothelial cells; Cardiac myocytes; Apoptosis; Caspase; Akt (protein kinase B)

Received September 5, 2012; final acceptance December 23, 2012. Online prepub date: January 16, 2013.
Address correspondence to Robert J. Henning, M.D., Center for Cardiovascular Research and James A. Haley VA Medical Center, 13000 Bruce B. Downs Blvd., Tampa, FL 33612, USA. Tel: +813-978-5873; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1651-1666, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657675
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Human Umbilical Cord Perivascular Cells Exhibit Enhanced Cardiomyocyte Reprogramming and Cardiac Function After Experimental Acute Myocardial Infarction

Gustavo Yannarelli,* Victor Dayan,* Natalia Pacienza,† Chyan-Jang Lee,† Jeffrey Medin,† and Armand Keating*‡

*Cell Therapy Program, Prince Margaret Hospital, University Health Network, Toronto, ON, Canada
†University Health Network, Toronto, ON, Canada
‡Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, ON, Canada

We were interested in evaluating the ability of the mesenchymal stromal cell (MSC) population, human umbilical cord perivascular cells (HUCPVCs), to undergo cardiomyocyte reprogramming in an established coculture system with rat embryonic cardiomyocytes. Results were compared with human bone marrow-derived (BM) MSCs. The transcription factors GATA4 and Mef 2c were expressed in HUCPVCs but not BM-MSCs at baseline and, at 7 days, increased 7.6- and 3.5-fold, respectively, compared with BM-MSCs. Although cardiac-specific gene expression increased in both cell types in coculture, upregulation was more significant in HUCPVCs, consistent with Mef 2c-GATA4 synergism. Using a lentivector with eGFP transcribed from the α-myosin heavy chain (α-MHC) promoter, we found that cardiac gene expression was greater in HUCPVCs than BM-MSCs after 14 days coculture (52 ± 17% vs. 29 ± 6%, respectively). A higher frequency of HUCPVCs expressed α-MHC protein compared with BM-MSCs (11.6 ± 0.9% vs. 5.3 ± 0.3%); however, both cell types retained MSC-associated determinants. We also assessed the ability of the MSC types to mediate cardiac regeneration in a NOD/SCID γ mouse model of acute myocardial infarction (AMI). Fourteen days after AMI, cardiac function was significantly better in cell-treated mice compared with control animals and HUCPVCs exhibited greater improvement. Although human cells persisted in the infarct area, the frequency of α-MHC expression was low. Our results indicate that HUCPVCs exhibit a greater degree of cardiomyocyte reprogramming but that differentiation for both cell types is partial. We conclude that HUCPVCs may be preferable to BM-MSCs in the cell therapy of AMI.

Key words: Human umbilical cord perivascular cells (HUCPVCs); Mesenchymal stromal cells (MSCs); Cardiomyocyte differentiation; Cell therapy

Received January 25, 2012; final acceptance August 16, 2012. Online prepub date: October 4, 2012.
Address correspondence to Gustavo Yannarelli, Ph.D., Princess Margaret Hospital, University Health Network, 610 University Ave. Suite 4-605, Toronto, Canada, M5G2M9. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1667-1681, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657765
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Amelioration of Paraquat-Induced Pulmonary Injury by Mesenchymal Stem Cells

Hsin-Lin Tsai,*† Jei-Wen Chang,*‡ Hui-Wen Yang,§ Chang-Wei Chen,§ Chen-Chang Yang,*¶# An-Hang Yang,*,** Chin-Su Liu,*† Tai-Wai Chin,*† Chou-Fu Wei,†† and Oscar K. Lee§,‡‡

*Institute of Clinical Medicine, School of Medicine, National Yang-Ming University, Taipei, Taiwan
†Division of Pediatric Surgery, Department of Surgery, Taipei Veterans General Hospital, Taipei, Taiwan
‡Department of Pediatrics, Taipei Veterans General Hospital, Taipei, Taiwan
§Department of Medical Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan
¶Department of Environmental and Occupational Medicine, School of Medicine, National Yang-Ming University, Taipei, Taiwan
#Division of Clinical Toxicology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan
**Division of Ultrastructural and Molecular Pathology, Department of Pathology, Taipei Veterans General Hospital, Taipei, Taiwan
††Department of Surgery, Taipei Medical University Hospital, Taipei, Taiwan
‡‡Stem Cell Research Center, National Yang-Ming University, Taipei, Taiwan

Acute paraquat (PQ) poisoning induces redox cycle and leads to fatal injury of lung. Clinical management is supportive in nature due to lack of effective antidote, and the mortality is very high. Mesenchymal stem cells (MSCs) process the properties of immunomodulation, anti-inflammatory, and antifibrotic effects and oxidative stress resistance. MSC transplantation may theoretically serve as an antidote in PQ intoxication. In this study, we examined the potential therapeutic effects of MSCs in PQ-induced lung injury. The degree of PQ toxicity in the rat type II pneumocyte cell line, L2, and MSCs was evaluated by examining cell viability, ultrastructural changes, and gene expression. L2 cells treated with 0.5 mM PQ were cocultured in the absence or presence of MSCs. For the in vivo study, adult male SD rats were administered an intraperitoneal injection of PQ (24 mg/kg body weight) and were divided into three groups: group I, control; group II, cyclophosphamide and methylprednisolone; group III, MSC transplantation 6 h after PQ exposure. MSCs were relatively resistant to PQ toxicity. Coculture with MSCs significantly inhibited PQ accumulation in L2 cells and upregulated the expression of antioxidative heme oxygenase 1 and metallothionein 1a genes, reversed epithelial-to-mesenchymal transition, and increased the viability of PQ-exposed L2 cells. Treatment with MSCs resulted in a significant reduction in severity of liver and renal function deterioration, alleviated lung injury, and prolonged the life span of rats. Altogether, our results suggest that MSCs possess antidote-like effect through multifactorial protection mechanism. The results of this preclinical study demonstrate that transplantation of MSCs may be a promising therapy and should be further validated clinically.

Key words: Mesenchymal stem cells (MSCs); Paraquat (PQ); Oxidative stress; Pulmonary injury

Received March 14, 2012; final acceptance August 26, 2012. Online prepub date: October 8, 2012.
Address correspondence to Oscar K. Lee, Department of Medical Research and Education, Taipei Veterans General Hospital, No. 201, Shi-Pai Rd., Sec. 2, Peitou, Taipei 11217, Taiwan. Tel: +886-2-2875-7557; Fax: +886-2-2875-7657; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1683-1694, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657756
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Amniotic Fluid Stem Cells Rescue Both In Vitro and In Vivo Growth, Innervation, and Motility in Nitrofen-Exposed Hypoplastic Rat Lungs Through Paracrine Effects

F. Pederiva,* M. Ghionzoli,† A. Pierro,† P. De Coppi,†1 and J. A. Tovar*1

*Department of Pediatric Surgery and Research Laboratory, Hospital Universitario La Paz, Madrid, Spain
†Surgery Unit, UCL Institute of Child Health and Great Ormond Street Hospital for Children, London, UK

Lung hypoplasia can be prevented in vitro by retinoic acid (RA). Recent evidence suggests that amniotic fluid stem (AFS) cells may integrate injured lungs and influence their recovery. We tested the hypothesis that AFS cells might improve lung growth and motility by paracrine mechanisms. Pregnant rats received either nitrofen or vehicle on E9.5. In vitro E13 embryonic lungs were cultured in the presence of culture medium alone or with RA, basophils, or AFS cells. In vivo green fluorescent protein-expressing (GFP+) rat AFS cells were transplanted in nitrofen-exposed rats on E10.5. E13 lung explants were cultured before analysis. The surface, the number of terminal buds, and the frequency of bronchial contractions were assessed. Protein gene product 9.5 (PGP 9.5) and α-actin protein levels were measured. The lung explants transplanted with AFS cells were stained for α-actin, PGP 9.5, and TTF-1. The levels of FGF-10, VEGFα, and TGF-β1 secreted by the AFS cells in the culture medium were measured. Comparison between groups was made by ANOVA. In vitro, the surface, the number of terminal buds, and the bronchial peristalsis were increased in nitrofen + AFS cell explants in comparison with nitrofen-exposed lungs. While nitrofen + RA lungs were similar to nitrofen + AFS ones, basophils did not normalize these measurements. PGP 9.5 protein was decreased in nitrofen lungs, but after adding AFS cells, the value was similar to controls. No differences were found in the expression of α-actin. In vivo, the surface, number of terminal buds, and peristalsis were similar to control after injection of AFS cells in nitrofen-exposed rats. Colocalization with TTF-1-positive cells was found. The levels of FGF-10 and VEGFα were increased in nitrofen + AFS cell explants, while the levels of TGF-β1 were similar to controls. Lung growth, bronchial motility, and innervation were decreased in nitrofen explants and rescued by AFS cells both in vitro and in vivo, similarly to that observed before with RA. The AFS cell beneficial effect was probably related to paracrine action of growth factor secretion.

Key words: Diaphragmatic hernia; Amniotic fluid stem (AFS) cells; Intrinsic innervation; Nitrofen; Peristalsis; Lung hypoplasia; Retinoic acid (RA)

Received November 16, 2011; final acceptance August 27, 2012. Online prepub date: October 8, 2012.
1These authors provided equal contribution to this work.
Address correspondence to Dr. Federica Pederiva, Department of Pediatric Surgery, Hospital Universitario “La Paz,” Paseo de la Castellana, 261, 28046 Madrid, Spain. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1695-1708, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657738
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Adoptive Transfer of Allogeneic Liver Sinusoidal Endothelial Cells Specifically Inhibits T-Cell Responses to Cognate Stimuli

Masataka Banshodani, Takashi Onoe, Masayuki Shishida, Hiroyuki Tahara, Shinji Hashimoto, Yuka Igarashi, Yuka Tanaka, and Hideki Ohdan

Department of Surgery, Division of Frontier Medical Science, Programs for Biomedical Research, Graduate School of Biomedical Sciences, Hiroshima University, Minami-ku, Hiroshima, Japan

Although it is well known that liver allografts are often accepted by recipients, leading to donor-specific tolerance of further organ transplants, the underlying mechanisms remain unclear. We had previously used an in vitro model and showed that mouse liver sinusoidal endothelial cells (LSECs) selectively suppress allospecific T-cells across major histocompatibility complex (MHC) barriers. In the present study, we established an in vivo model for evaluating the immunomodulatory effects of allogeneic LSECs on corresponding T-cells. Allogeneic BALB/cA LSECs were injected intraportally into recombination activating gene 2 γ-chain double-knockout (RAG2/gc-KO, H-2b) mice lacking T, B, and natural killer (NK) cells. In order to facilitate LSEC engraftment, the RAG2/gc-KO mice were injected intraperitoneally with monocrotaline 2 days before the adoptive transfer of LSECs; this impaired the host LSECs, conferring a proliferative advantage to the transplanted LSECs. After orthotopic allogeneic LSEC engraftment, the RAG2/gc-KO mice were immune reconstituted intravenously with C57BL/6 splenocytes. After immune reconstitution, mixed lymphocyte reaction (MLR) assay using splenocytes from the recipients revealed that specific inhibition of host CD4+ and CD8+ T-cell proliferation was greater in response to allostimulation with irradiated BALB/cA splenocytes rather than to stimulation with irradiated third party SJL/jorllco splenocytes. This inhibitory effect was attenuated by administering anti-programmed death ligand 1 (PD-L1) monoclonal antibody during immune reconstitution in the above-mentioned mice, but not in RAG2/gc-KO mice engrafted with Fas ligand (FasL)-deficient BALB/cA LSECs. Furthermore, engraftment of allogeneic BALB/cA LSECs significantly prolonged the survival of subsequently grafted cognate allogeneic BALB/cA hearts in RAG2/gc-KO mice immune reconstituted with bone marrow transplantation from C57BL/6 mice. In conclusion, murine LSECs have been proven capable of suppressing T-cells with cognate specificity for LSECs in an in vivo model. The programmed death 1/PD-L1 pathway is likely involved in these suppressive effects.

Key words: Endothelial cells; Tolerance; Transplantation; Alloreactive T-cells; Programmed death ligand 1 (PD-L1)

Received June 28, 2011; final acceptance August 25, 2012. Online prepub date: October 8, 2012.
Address correspondence to Hideki Ohdan, M.D., Ph.D., Department of Surgery, Division of Frontier Medical Science, Programs for Biomedical Research, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8551, Japan. Tel: +81-82-257-5220; Fax: +81-82-257-5224; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1709-1722, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657684
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Roles of Islet Toll-Like Receptors in Pig to Mouse Islet Xenotransplantation

Han Ro,*† Eun Won Lee,† Joo Ho Hong,† Kyu Hyun Han,† Hye-Jung Yeom,† Hwa Jung Kim,† Myung-Gyu Kim,* Hye Seung Jung,‡ Kook-Hwan Oh,§ Kyong Soo Park,‡ Curie Ahn,*†§ and Jaeseok Yang*†

*Transplantation Center, Seoul National University Hospital, Seoul, Republic of Korea
†Transplantation Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea
‡Division of Endocrinology and Metabolism, Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea
§Division of Nephrology, Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea

Although innate immunity plays important roles in xenograft rejection, there have been few studies on the role of toll-like receptors (TLRs) in xenotransplantation. Furthermore, most studies focused on the recipient’s TLRs. Therefore, we investigated whether TLRs in porcine islets can contribute to islet xenograft rejection. Adult porcine islets were isolated and stimulated by polyinosinic/polycytidylic acid (poly I:C) or lipopolysaccharide (LPS). Both poly I:C and LPS stimulation in porcine islets induced expression of chemokines (RANTES, MCP-1, IP-10, and IL-8), cytokines (IL-6 and type I interferons), and adhesion molecules (VCAM-1 and ICAM-1). Porcine islet supernatants stimulated by TLR agonists induced chemotaxis of human leukocytes. They also induced procoagulant activation (tissue factor and fgl-2). However, TLR stimulation did not influence insulin secretion. When porcine MyD88 was knocked down using shRNA lentivirus, TLR-mediated induction of proinflammatory mediators and procoagulants was attenuated. When LPS was injected to MyD88 or TLR4 knockout mice after porcine islet transplantation, LPS stimulation on donor islets interfered with islet xenograft tolerance induction by anti-CD154 antibodies. Inflammatory cell infiltration and expression of proinflammatory chemokines and cytokines in islet xenografts also increased. In conclusion, TLR activation in porcine islets induced both a proinflammatory and procoagulant response and thereby contributed to xenograft rejection.

Key words: Islet transplantation; Rejection; Toll-like receptors (TLRs); Xenotransplantation

Received December 10, 2011; final acceptance August 25, 2012. Online prepub date: October 8, 2012.
Address correspondence to Jaeseok Yang, Transplantation Center, Seoul National University Hospital, 28 Yeongeon-dong, Jongno-gu, Seoul, 110-744, Republic of Korea. Tel: +82-2-2072-4128; Fax: +82-2-2072-4129; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Curie Ahn, Transplantation Center, Seoul National University Hospital, 28 Yeongeon-dong, Jongno-gu, Seoul, 110-744, Republic of Korea. Tel: +82-2-2072-2222; Fax: +82-2-763-6317; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 22, pp. 1723-1733, 2013
0963-6897/13 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X657873
E-ISSN 1555-3892
Copyright © 2013 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

A Physiological Pattern of Oxygenation Using Perfluorocarbon-Based Culture Devices Maximizes Pancreatic Islet Viability and Enhances β-Cell Function

Chris A. Fraker,* Sirlene Cechin,* Silvia Álvarez-Cubela,* Felipe Echeverri,† Andrés Bernal,† Ramón Poo,† Camillo Ricordi,*‡ Luca Inverardi,*§ and Juan Domínguez-Bendala*‡1

*Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
†Biorep Technologies, Miami, FL, USA
‡Department of Surgery, University of Miami Miller School of Medicine, Miami, FL, USA
§Department of Medicine, University of Miami School of Medicine, Miami, FL, USA

Conventional culture vessels are not designed for physiological oxygen (O2) delivery. Both hyperoxia and hypoxia—commonly observed when culturing cells in regular plasticware—have been linked to reduced cellular function and death. Pancreatic islets, used for the clinical treatment of diabetes, are especially sensitive to sub- and supraphysiological O2 concentrations. A result of current culture standards is that a high percentage of islet preparations are never transplanted because of cell death and loss of function in the 24–48 h postisolation. Here, we describe a new culture system designed to provide quasiphysiological oxygenation to islets in culture. The use of dishes where islets rest atop a perfluorocarbon (PFC)-based membrane, coupled with a careful adjustment of environmental O2 concentration to target the islet physiological pO2 range, resulted in dramatic gains in viability and function. These observations underline the importance of approximating culture conditions as closely as possible to those of the native microenvironment, and fill a widely acknowledged gap in our ability to preserve islet functionality in vitro. As stem cell-derived insulin-producing cells are likely to suffer from the same limitations as those observed in real islets, our findings are especially timely in the context of current efforts to define renewable sources for transplantation.

Key words: Oxygenation; Cell culture; Cell viability; Islets; Perfluorocarbon (PFC)

Received July 10, 2012; final acceptance August 26, 2012. Online prepub date: October 12, 2012.
Address correspondence to Juan Domínguez-Bendala, Diabetes Research Institute, University of Miami Miller School of Medicine, 1450 NW 10th Avenue, Miami, FL, 33136, USA. Tel: +1-305-243-4092; Fax: +1-305-243-4404; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it