Cell Transplantation 23(7) Abstracts

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Cell Transplantation, Vol. 23, pp. 791-803, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X666430
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Optimization of Culture Conditions for Stem Cells Derived From Human Anterior Cruciate Ligament and Bone Marrow

Ming-Te Cheng,*† Chien-Lin Liu,*‡ Tain-Hsiung Chen,*‡ and Oscar K. Lee‡§¶

*Department of Surgery, School of Medicine, National Yang-Ming University, Taipei, Taiwan
†Taoyuan General Hospital, Taoyuan County, Taiwan
‡Department of Orthopedics and Traumatology, Taipei Veterans General Hospital, Taipei, Taiwan
§Department of Medical Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan
¶Stem Cell Research Center, National Yang-Ming University, Taipei, Taiwan

Tissue engineering with stem cells is a fascinating approach for treating anterior cruciate ligament (ACL) injuries. In our previous study, stem cells isolated from the human anterior cruciate ligament were shown to possess extensive proliferation and differentiation capabilities when treated with specific growth factors. However, optimal culture conditions and the usefulness of fetal bovine serum (FBS) as a growth factor in in vitro culture systems are yet to be determined. In this study, we compared the effects of different culture media containing combinations of various concentrations of FBS and the growth factors basic fibroblastic growth factor (bFGF) and transforming growth factor-β1 (TGF-β1) on the proliferation and differentiation of ligament-derived stem cells (LSCs) and bone marrow mesenchymal stem cells (BMSCs). We found that α-MEM plus 10% FBS and bFGF was able to maintain both LSCs and BMSCs in a relatively undifferentiated state but with lower major extracellular matrix (ECM) component gene expression and protein production, which is beneficial for stem cell expansion. However, the differentiation and proliferation potentials of LSCs and BMSCs were increased when cultured in MesenPRO, a commercially available stem cell medium containing 2% FBS. MesenPRO in conjunction with TGF-β1 had the greatest ability to induce the differentiation of BMSCs and LSCs to ligament fibroblasts, which was evidenced by the highest ligamentous ECM gene expression and protein production. These results indicate that culture media and growth factors play a very important role in the success of tissue engineering. With α-MEM plus 10% FBS and bFGF, rapid proliferation of stem cells can be achieved. In this study, MesenPRO was able to promote differentiation of both LSCs and BMSCs to ligament fibroblasts. Differentiation was further increased by TGF-β1. With increasing understanding of the effects of different culture media and growth factors, manipulation of stem cells in the desired direction for ligament tissue engineering can be achieved.

Key words: Tissue engineering; Anterior cruciate ligament (ACL); Culture medium; Fetal bovine serum (FBS); Growth factors

Received October 3, 2010; final acceptance December 31, 2012. Online prepub date: April 12, 2013.
Address correspondence to Oscar K. Lee, M.D., Ph.D., Stem Cell Research Center, National Yang-Ming University, No. 201, Sec 2, Shih-Pai Road, Taipei 11217, Taiwan. Tel: +886-2-2875-7391; 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. 23, pp. 805-817, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368913X664856
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Cultured Hepatocytes Adopt Progenitor Characteristics and Display Bipotent Capacity to Repopulate the Liver

Petra Krause,* Kirsten Unthan-Fechner,† Irmelin Probst,† and Sarah Koenig*

*Department of General and Visceral Surgery, University Medical Centre Goettingen, Goettingen, Germany
†Department of Biochemistry I, Centre of Biochemistry and Molecular Cell Biology, University Medical Centre Goettingen, Goettingen, Germany

Clinical studies have proved the therapeutic potential of hepatocyte transplantation as a promising alternative to whole organ liver transplantation in the treatment of hereditary or end-stage liver disease. However, donor shortage seriously restricts cell availability, and the lack of appropriate cell culture protocols for the storage and maintenance of donor cells constitutes a significant obstacle. The aim of this study was to stimulate mature hepatocytes in culture to multiply in vitro and track their fate on transplantation. Rat hepatocytes isolated nonenzymatically were cultured serum free for up to 10 days. They were stimulated into proliferation in the presence of growth factors and conditioned media from nonparenchymal and hepatocyte culture supernatants, as well as 10 mM lithium chloride (LiCl). Cell proliferation was assessed by determining DNA content. Additionally, the extent of cell differentiation was estimated using immunofluorescence staining of hepatic, biliary, progenitor, and mesenchymal markers and gene expression analyses. Transplantation studies were performed on the Fischer CD26-mutant rat following pretreatment with retrorsine and partial hepatectomy. Proliferating hepatocytes increasingly adopted precursor characteristics, expressing progenitor (OV6, CD133), hepatic lineage (CK18), biliary (CD49f, CK7, CK19), and mesenchymal (vimentin) markers. The supplement of LiCl further enhanced the proliferative capacity by 30%. Transplantation studies revealed extensive repopulation by large donor hepatocyte clusters. Furthermore, bile duct-like structures deriving from donor cells proved to be immunoreactive to ductular markers and formed in close proximity to endogenous bile ducts. Mature hepatocytes reveal their potential to “switch” between phenotypes, adopting progenitor characteristics during proliferation in vitro. Following transplantation, these “retrodifferentiated” cells further expanded in vivo, thereby generating bipotentially differentiated progenies (hepatocytes and bile duct-like structures). This apparent plasticity of mature hepatocytes may open new approaches for cell-based strategies to treat liver disease.

Key words: Liver cell transplantation; Differentiation; Retrodifferentiation; Hepatocyte culture; Progenitor cells; Oval cells

Received October 30, 2012; final acceptance February 22, 2013. Online prepub date: March 5, 2013
Address correspondence to Sarah Koenig, Department of General and Visceral Surgery, University Medical Centre Goettingen, Robert-Koch-Str. 40, 37075 Goettingen, Germany. Tel: +49-551 39-8977; Fax: +49-551 39-6109; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 819-830, 2014
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DOI: http://dx.doi.org/10.3727/096368913X664847
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Cold Storage of Rat Hepatocyte Spheroids

Hongling Liu,*† Yue Yu,*‡ Jaime Glorioso,* Shennen Mao,* Brian Rodysil,* Bruce P. Amiot,§ Piero Rinaldo,¶ and Scott L. Nyberg*

*Division of Experimental Surgery, Mayo Clinic College of Medicine, Rochester, MN, USA
†Liver Failure Diagnosis and Treatment Center, 302 Military Hospital, Beijing, P.R. China
‡Liver Transplantation Center, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province, P.R. China
§Brami Biomedical, Inc., Minneapolis, MN, USA
¶Department of Laboratory Medicine and Pathology, Mayo Clinic College of Medicine, Rochester, MN, USA

Cell-based therapies for liver disease rely on a high-quality supply of hepatocytes and a means for storage during transportation from site of isolation to site of usage. Unfortunately, frozen cryopreservation is associated with unacceptable loss of hepatocyte viability after thawing. The purpose of this study was to optimize conditions for cold storage of rat hepatocyte spheroids without freezing. Rat hepatocytes were isolated by a two-step perfusion method; hepatocyte spheroids were formed during 48 h of rocked culture in serum-free medium (SFM). Spheroids were then maintained in rocked culture at 37°C (control condition) or cold stored at 4°C for 24 or 48 h in six different cold storage solutions: SFM alone; SFM + 1 mM deferoxamine (Def); SFM + 1 mM cyclosporin A (CsA); SFM + 1 mM Def + 1 mM CsA, University of Wisconsin (UW) solution alone, UW + 1 mM Def. Performance metrics after cold storage included viability, gene expression, albumin production, and functional activity of cytochrome P450 enzymes and urea cycle proteins. We observed that cold-induced injury was reduced significantly by the addition of the iron chelator (Def) to both SFM and UW solution. Performance metrics (ammonia detoxification, albumin production) of rat hepatocyte spheroids stored in SFM + Def for 24 h were significantly increased from SFM alone and approached those in control conditions, while performance metrics after cold storage in SFM alone or cold storage for 48 h were both significantly reduced. A serum-free medium supplemented with Def allowed hepatocyte spheroids to tolerate 24 h of cold storage with less than 10% loss in viability and functionality. Further research is warranted to optimize a solution for extended cold storage of hepatocyte spheroids.

Key words: Bioartificial liver; Spheroid; Hepatocyte; Cold storage

Received August 10, 2012; final acceptance February 22, 2013. Online prepub date: March 5, 2013.
Address correspondence to Scott L. Nyberg, M.D., Ph.D., Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA. Tel: +1-507-266-6772; Fax: +1-507-266-2810; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 831-843, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X661391
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Isolation and Characterization of Antler-Derived Multipotent Stem Cells

Min-Soo Seo,1,2 Sang-Bum Park,1 Soon-Won Choi, Jae-Jun Kim, Hyung-Sik Kim, and Kyung-Sun Kang

Adult Stem Cell Research Center, College of Veterinary Medicine, Seoul National University, Seoul, Republic of Korea

Recent studies have reported that stem cells can be isolated from various tissues such as bone marrow, fatty tissue, umbilical cord blood, Wharton’s jelly, and placenta. These types of stem cell studies have also arisen in veterinary medicine. Deer antlers show a seasonal regrowth of tissue, an unusual feature in mammals. Antler tissue therefore might offer a source of stem cells. To explore the possibility of stem cell populations within deer antlers, we isolated and successfully cultured antler-derived multipotent stem cells (MSCs). Antler MSCs were maintained in a growth medium, and the proliferation potential was measured via an assay called the cumulative population doubling level. Immunophenotyping and immunostaining revealed the intrinsic characteristic stem cell markers of antler MSCs. To confirm the ability to differentiate, we conducted osteogenic, adipogenic, and chondrogenic induction under the respective differentiation conditions. We discovered that antler MSCs have the ability to differentiate into multiple lineages. In conclusion, our results show that deer antler tissue may contain MSCs and therefore may be a potential source for veterinary regenerative therapeutics.

Key words: Deer horn; Antler; Stem cells; Characterization

1These authors provided equal contribution to this work.
2Present address: Laboratory Animal Center, Daegu-Gyeongbuk Medical Innovation Foundation, Deagu, Korea.
Received May 8, 2012; final acceptance December 17, 2012. Online prepub date: January 2, 2013. Address correspondence to Kyung-Sun Kang, College of Veterinery Medicine, Seoul National University, Seoul 151-742, Republic of 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


Cell Transplantation, Vol. 23, pp. 845-854, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368913X664559
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Factors Affecting Successful Isolation of Human Corneal Endothelial Cells for Clinical Use

Jin San Choi,*†‡ Eun Young Kim,*§ Min Jeong Kim,*§ Faraaz A Khan,* Matthew Giegengack,*¶ Ralph D’Agostino Jr.,# Tracy Criswell,* Gilson Khang,†§ and Shay Soker*

*Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, USA
†Department of Advanced Organic Materials Engineering, Chonbuk National University, Backjedaero, Dukjin, Jeonju, Republic of Korea
‡Ocular Systems, Inc., Winston-Salem, NC, USA
§Department of BIN Fusion Technology and Polymer Fusion Research Center, Chonbuk National University, Backjedaero, Dukjin, Jeonju, Republic of Korea
¶Department of Ophthalmology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, USA
#Public Health Sciences—Department of Biostatistics, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, USA

Corneal transplantation is a common transplant procedure used to improve visual acuity by replacing the opaque or distorted host tissue with clear healthy donor tissue. However, its clinical utility is limited due to a lack of donor supply of high-quality corneas. Bioengineered neocorneas, created using an expandable population of human donor-derived corneal endothelial cells (HCECs), could address this shortage. Thus, the objective of this study was to evaluate HCEC sourcing with various isolation methods, including enzymatic digestion, culture medium components, and adhesive proteins. HCECs were obtained from corneas obtained from various aged donors after endothelial keratoplasty. Under a dissection microscope, the Descemet’s membrane, including the attached corneal endothelium, was stripped from the stroma, and the cells were isolated and expanded by explant culture or by enzymatic digestion with enzymes such as collagenase II, dispase, or trypsin. In order to improve the initial cell attachment, tissue culture plates were coated with collagen IV, fibronectin, or fibronectin–collagen combination coating mix (FNC) before cell plating. We were able to successfully obtain HCECs from 32% (86/269) of donor corneas. Donor age and isolation method influenced the characteristics of the resulting in vitro HCEC culture. Under all conditions tested, FNC-coated plates showed higher quality cultures than the other coatings tested. These results suggest that donor age and HCEC isolation methodology are the two factors that most directly affect the quality of the resulting HCEC culture in vitro. These factors should guide the methodological development for the clinical expansion of HCECs for the generation of bioengineered neocorneas.

Key words: Cornea; Endothelial cells; Isolation; Transplantation; Tissue engineering

Received August 22, 2012; final acceptance February 6, 2013. Online prepub date: February 26, 2013.
Address correspondence to Shay Soker, Ph.D., Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157, USA. Tel: +1-336-713-7295; Fax: +1-336-713-7290; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 855-872, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368913X667024
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Survival and Integration of Developing and Progenitor-Derived Retinal Ganglion Cells Following Transplantation

Jonathan Hertz,*† Bo Qu,*‡ Ying Hu,* Roshni D. Patel,* Daniel A. Valenzuela,* and Jeffrey L. Goldberg*†§

*Bascom Palmer Eye Institute, Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, FL, USA
†Neuroscience Graduate Program, University of Miami Miller School of Medicine, Miami, FL, USA
‡Department of Ophthalmology at the 4th Affiliated Hospital, China Medical University, Shen Yang, Liao Ning Province, China
§Shiley Eye Center, UC San Diego, La Jolla, CA, USA

There is considerable interest in transplanting stem cells or progenitors into the injured nervous system and enhancing their differentiation into mature, integrated, functional neurons. Little is known, however, about what intrinsic or extrinsic signals control the integration of differentiated neurons, either during development or in the adult. Here we ask whether purified, postmitotic, differentiated retinal ganglion cells (RGCs) directly isolated from rat retina or derived from in vitro-differentiated retinal progenitor cells can survive, migrate, extend neurites, and form morphologic synapses in a host retina, in vivo and ex vivo. We found that acutely purified primary and in vitro-differentiated RGCs survive transplantation and migrate into deeper retinal layers, including into their normal environment, the ganglion cell layer (GCL). Transplanted RGCs from a wide range of developmental ages, but not from adults, were capable of extending lengthy neurites in the normal and injured adult rat retina ex vivo and to a lesser degree after transplantation in vivo. We have also demonstrated that RGCs may be differentiated and purified from retinal precursor cultures and that they share many of the same cell biological properties as primary RGCs. We have established that progenitor-derived RGCs have similar capacity for integration as developing primary RGCs but appear to form a lower number of presynaptic punctae. This work provides insight for further understanding of the integration of developing RGCs into their normal environment and following injury.

Key words: Retinal ganglion cell (RGC); Neural transplantation; Integration; Retina

Received June 29, 2011; final acceptance April 4, 2013. Online prepub date: April 29, 2013.
Address correspondence to Jeffrey L. Goldberg, Shiley Eye Center, 9415 Campus Point Dr. #0946, La Jolla, CA 92093, USA. Tel: +1-858-534-9794; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 873-887, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368913X664586
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Characteristics of Mouse Adipose Tissue-Derived Stem Cells and Therapeutic Comparisons Between Syngeneic and Allogeneic Adipose Tissue-Derived Stem Cell Transplantation in Experimental Autoimmune Thyroiditis

Eun Wha Choi,*†1 Il Seob Shin,‡1 So Young Park,* Eun Ji Yoon,‡ Sung Keun Kang,‡ Jeong Chan Ra,‡ and Sung Hwa Hong*§

*Laboratory Animal Research Center, Samsung Biomedical Research Institute, Gangnam-gu, Seoul, Republic of Korea
†School of Medicine, Sungkyunkwan University, Gangnam-gu, Seoul, Republic of Korea
‡Stem Cell Research Center, K-STEMCELL, Gasan-dong, Geumcheon-gu, Seoul, Republic of Korea
§Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Gangnam-gu, Seoul, Republic of Korea

Previously, we found that the intravenous administration of human adipose tissue-derived mesenchymal stem cells was a promising therapeutic option for autoimmune thyroiditis even when the cells were transplanted into a xenogeneic model without an immunosuppressant. Therefore, we explored the comparison between the therapeutic effects of syngeneic and allogeneic adipose tissue-derived stem cells on an experimental autoimmune thyroiditis mouse model. Experimental autoimmune thyroiditis was induced in C57BL/6 mice by immunization with porcine thyroglobulin. Adipose tissue-derived stem cells derived from C57BL/6 mice (syngeneic) or BALB/c mice (allogeneic) or saline as a vehicle control were administered intravenously four times weekly. Blood and tissue samples were collected 1 week after the last transplantation. Adipose tissue-derived stem cells from mice were able to differentiate into multiple lineages in vitro; however, mouse adipose tissuederived stem cells did not have immunophenotypes identical to those from humans. Syngeneic and allogeneic administrations of adipose tissue-derived stem cells reduced thyroglobulin autoantibodies and the inflammatory immune response, protected against lymphocyte infiltration into the thyroid, and restored the Th1/Th2 balance without any adverse effects. However, different humoral immune responses were observed for infused cells from different stem cell sources. The strongest humoral immune response was induced by xenogeneic transplantation, followed by allogeneic and syngeneic administration, in that order. The stem cells were mostly found in the spleen, not the thyroid. This migration might be because the stem cells primarily function in systemic immune modulation, due to being given prior to disease induction. In this study, we confirmed that there were equal effects of adipose tissue-derived stem cells in treating autoimmune thyroiditis between syngeneic and allogeneic transplantations.

Key words: Adipose tissue-derived stem cells; Autoimmune thyroiditis; Autoimmune disease; Anti-thyroglobulin antibody; Immunophenotype; Mouse

Received September 22, 2011; final acceptance February 19, 2013. Online prepub date: March 5, 2013.
1These authors provided equal contribution to this work.
Address correspondence to Eun Wha Choi, Laboratory Animal Research Center, Samsung Biomedical Research Institute, 50 Irwon-dong, Gangnam-gu, Seoul 135-710, Republic of Korea. Tel: +82-2-3410-3700; Fax: +82-2-3410-3628; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 889-899, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368913X665594
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Transplantation of Allogenic Fetal Membrane-Derived Mesenchymal Stem Cells Protects Against Ischemia/Reperfusion-Induced Acute Kidney Injury

Hidetoshi Tsuda,*† Kenichi Yamahara,† Kentaro Otani,† Masayoshi Okumi,‡ Koji Yazawa,‡ Jun-ya Kaimori,*§ Akihiko Taguchi,†¶ Kenji Kangawa,# Tomoaki Ikeda,†** Shiro Takahara,* and Yoshitaka Isaka§

*Department of Advanced Technology for Transplantation, Osaka University Graduate School of Medicine, Osaka, Japan
†Department of Regenerative Medicine and Tissue Engineering, National Cerebral and Cardiovascular Center Research Institute, Osaka, Japan
‡Department of Urology, Osaka University Graduate School of Medicine, Suita, Japan
§Department of Geriatric and Nephrology, Osaka University Graduate School of Medicine, Suita, Japan
¶Department of Regenerative Medicine Research, Institute of Biomedical Research and Innovation, Kobe, Japan
#Department of Biochemistry, National Cerebral and Cardiovascular Center, Osaka, Japan
**Department of Obstetrics and Gynecology, Mie University Graduate School of Medicine, Tsu, Japan

Mesenchymal stem cells (MSCs) are an attractive therapeutic cell source for treating renal diseases. MSC administration has been shown to improve renal function, although the underlying mechanisms are not completely understood. We recently showed that allogenic fetal membrane-derived MSCs (FM-MSCs), which are available noninvasively in large amounts, had a renoprotective effect in an experimental glomerulonephritis model. Here we investigated whether allogenic FM-MSC administration could protect kidneys from ischemia/ reperfusion (I/R) injury. Lewis rats were subjected to right nephrectomy and left renal I/R injury by clamping the left renal artery as an acute kidney injury (AKI) model. After declamping, FM-MSCs (5 × 105 cells) obtained from major histocompatibility complex (MHC)-mismatched ACI rats were intravenously administered. I/R-injured rats exhibited increased serum creatinine and BUN, whereas FM-MSC administration significantly ameliorated renal function. Histological analysis revealed that FM-MSC administration significantly suppressed tubular apoptosis and infiltration of macrophages and T-cells. Administration of FM-MSCs mainly homed into the lung, but increased serum IL-10 levels. Of interest is that renoprotective effects of FM-MSCs were abolished by using anti-IL-10 neutralization antibody, suggesting that IL-10 would be one of the candidate factors to protect rat kidney from I/R injury in this model. We concluded that allogenic FM-MSC transplantation is a potent therapeutic strategy for the treatment of AKI.

Key words: Fetal membrane (FM); Cell therapy; Acute kidney injury (AKI)

Received August 9, 2012; final acceptance March 8, 2013. Online prepub date: April 2, 2013.
Address correspondence to Yoshitaka Isaka, M.D., Ph.D., Department of Geriatric and Nephrology, Osaka University Graduate School of Medicine, Suita, Japan. E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 901-912, 2014
0963-6897/14 $90.00 + .00
DOI: http://dx.doi.org/10.3727/096368912X667015
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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Molecular Imaging of Expression of Vascular Endothelial Growth Factor A (VEGF A) in Femoral Bone Grafts Transplanted Into Living Mice

Olga Strachna,*1 Daniel Torrecilla,*1 Marie K. Reumann,† Inna Serganova,‡ Jihye Kim,* Simone Gieschler,* Adele L. Boskey,§ Ronald G. Blasberg,‡ and Philipp Mayer-Kuckuk¶

*Research Division, Hospital for Special Surgery, New York, NY, USA
†BG Trauma Center, Eberhard-Karls-University, Tuebingen, Germany
‡Center for Molecular Imaging in Cancer, Memorial Sloan-Kettering Cancer Center, New York, NY, USA
§Mineralized Tissue Laboratory, Hospital for Special Surgery, New York, NY, USA
¶Bone Cell Biology and Imaging Laboratory, Department of Orthopedics, MRI, Technical University Munich, Munich, Germany

The biology of cells transplanted with bone grafts is incompletely understood. Focusing on the early angiogenic response postgrafting, we report a mouse femur graft model in which grafts were derived from mice transgenic for a firefly luciferase (FLuc) bioluminescence reporter gene driven by a promoter for the angiogenic signaling molecule vascular endothelial growth factor (VEGF). Upon transplantation into wild-type (wt) mice, in vivo bioluminescence imaging (BLI) permitted longitudinal visualization and measurements of VEGF promoter activity in the transplanted graft cells and demonstrated a lag period of 7 days posttransplantation prior to robust induction of the promoter. To determine cellular mediators of VEGF induction in graft bone, primary graft-derived osteoblastic cells (GDOsts) were characterized. In vitro BLI on GDOsts showed hypoxia-induced VEGF expression and that this induction depended on PI3K signaling and, to a lesser degree, on the MEK pathway. This transcriptional regulation correlated with VEGF protein production and was validated in GDOsts seeded on demineralized bone matrix (DBM), a bone graft substitute material. Together, combined imaging of VEGF expression in living animals and in live cells provided clues about the regulation of VEGF in cells post-bone grafting. These data are particularly significant toward the development of future smart bone graft substitutes.

Key words: Bone grafting; Vascular endothelial growth factor (VEGF); Molecular imaging; Bioluminescence imaging; Osteoblasts; Signaling pathways

Received January 6, 2012; final acceptance December 31, 2012. Online prepub date: April 12, 2013.
1These authors provided equal contribution to this work.
Address correspondence to Dr. Philipp Mayer-Kuckuk, Bone Cell Biology and Imaging Laboratory, Department of Orthopedics, MRI, Technical University Munich, Munich, Germany. Fax: +49-(89)-4140-7881; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Cell Transplantation, Vol. 23, pp. 913-919, 2014
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DOI: http://dx.doi.org/10.3727/096368913X665567
E-ISSN 1555-3892
Copyright © 2014 Cognizant Comm. Corp.
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A Preconditioning Regimen With a PKCε Activator Improves Islet Graft Function in a Mouse Transplant Model

Diana Hamilton,* Caitlin Rugg,* Nicolynn Davis,* Manana Kvezereli,* Bashir Akhavan Tafti,† Stephan Busque,† and Magali Fontaine*‡

*Department of Pathology, Stanford University, Stanford, CA, USA
†Department of Surgery, Stanford University, Stanford, CA, USA
‡Department of Pathology, University of Maryland School of Medicine, Baltimore, MD, USA

Transplantation of islets isolated from deceased donor pancreata is an attractive method of b-cell replacement therapy for patients with type 1 diabetes (T1D). However, the loss of islet cell viability and function during the peritransplant period is a limiting factor to long-term islet engraftment. Activation of the isoenzyme PKCε may improve islet survival and function. The current study assesses the effects of PKCε activation on islet graft function in a syngeneic streptozotocin-induced diabetic mouse model. Islets were isolated from wild-type BALB/c mice preconditioned with either a PKCε activator (ψεRACK) or a TAT carrier control peptide. Islets were further treated with the same agents during isolation, purification, and incubation prior to transplantation. Two hundred seventy-five islet equivalents were transplanted under the kidney capsule of streptozotocin-induced diabetic BALB/c mice. Islet function was assessed by measurement of blood glucose levels every 3 days for 42 days after transplant and through an intraperitoneal glucose tolerance test (IPGTT). The time for return to euglycemia in mice transplanted with islets treated with yeRACK was improved at 14 ± 6 days versus 21 ± 6 days with TAT-treated islets. The IPGTT showed a 50% reduction in the area under the curve associated with an improved insulin response in mice transplanted with yeRACK-treated islets compared to TAT-treated islets. A preconditioning regimen using PKCε agonist before pancreatic recovery and during islet isolation improves islet graft function and resistance to high glucose stress after transplantation.

Key words: Islet; Transplantation; Protein kinase C ε (PKCε); Diabetes

Received November 13, 2012; final acceptance March 6, 2013. Online prepub date: April 2, 2013.
Address correspondence to Magali J. Fontaine, Department of Pathology, University of Maryland School of Medicine, 22 S. Greene Street N2W50a, Baltimore, MD 21201, USA. Tel: 410 328 3834; Fax: 433 462 3248; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it or Stephan Busque, Department of Surgery, Multi-Organ Transplantation, Stanford University, 750 Welch Rd, Ste 319, MC 5731, Palo Alto, CA 94304, USA. Tel: +1-650-498-6189; Fax: +1-650-498-5690; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it