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ABSTRACT: Nerve growth factor (NGF) has been reported to play an important regulatory role in pancreatic beta-cell function. However, the usefulness of NGF in a transplantation setting is unknown.
A marginal number of islet cells (260 islet equivalents/recipient) cultured for 24 hr with NGF (500 ng/ml) was syngeneically transplanted under the kidney capsule of streptozotocin-induced diabetic Balb/c mice. Fluorescence microscopy was used to evaluate islet viability. Islet function was evaluated in vitro and in vivo by static assay and glucose tolerance test, respectively.
In vitro, improved viability and survival were found in murine islets cultured in serum-free medium for 96 hr with 500 ng/ml NGF (P<0.05). NGF-treated islets had more insulin secretion than islets cultured without NGF in response to 2.8 mmol/L glucose (P<0.05), and 20 mmol/L glucose conditions. In vivo, 67% of recipients with a submarginal number of islets cultured in NGF attained normoglycemia for more than 120 days, whereas transplanted islets without NGF treatment survived a maximum of 13 days in control mice. At posttransplant day 4, recipients of NGF-cultured islets showed significant improvement of glucose tolerance. On immunohistochemistry, the kidney capsules containing NGF-cultured islets displayed higher insulin content, and more dilated neoplastic microvessels than control renal capsules. The number of apoptotic cells using TUNEL staining decreased by nearly 50% in NGF-cultured islet grafts in comparison to control islet grafts.
The above data suggest potential advantages of NGF for islet survival following transplantation. This neurotrophic approach may prove beneficial in human islet transplantation.
Transplantation 03/2006; 81(4):519-24. · 4.00 Impact Factor
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ABSTRACT: The mechanism involved in the spontaneous acceptance of liver allografts in some rat strain combinations remains unclear. Immunoregulatory NKR-P1TCRalphabetaT (NKT) cells primarily produce IL-4 and IFN-gamma, and enhance the polarization of immune responses to Th2 and Th1, respectively. The aim of this study was to clarify the role of graft-derived NKT cells in inducing the spontaneous acceptance of rat orthotopic liver transplantation (OLTx)
The experimental groups were divided as follows: Group 1, BN to LEW "low responder (acceptor)" combination; Group 2, DA to LEW "high responder (rejector)" combination; naïve BN (Group 3) or LEW recipients (Group 4) with liver allografts from irradiated BN donors. The recipients had liver allografts from irradiated donors reconstituted from the following cell populations 24 hr before harvesting, spleen cells (SPCs, Group 5), IgSPCs (Group 6), IgNKR-P1SPCs (Group 7), and IgTCRabSPCs (Group 8)
In Group 1, the percent of graft-derived NKT cells harvested on day 7 posttransplant were significantly higher than in Group 2. In the case of BN liver allografts that had been irradiated and reconstituted with cell populations including NKT cells (Groups 5 and 6), the mean graft survival (MST) was extended to 39.2+/-5.7 and 38.8+/-8.0 days, respectively. In contrast, when NKT cells were excluded (Groups 7 and 8), the grafts were acutely rejected within MST of 17.8+/-4.0 and 18.8+/-7.7 days, respectively. The concentrations of IL-10 and TGF-beta, but not IL-4 in IgGICs culture supernatants were predominant in the acceptor, whereas those with IFN-gamma predominated in the rejector.
Graft-derived NKT cells might be responsible for spontaneous acceptance in the rat OLTx.
Transplantation 01/2006; 80(12):1749-55. · 4.00 Impact Factor
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ABSTRACT: Background. The mechanism involved in the spontaneous acceptance of liver allografts in some rat strain combinations remains unclear. Immunoregulatory NKR-P1+TCRαβ+T (NKT) cells primarily produce IL-4 and IFN-γ, and enhance the polarization of immune responses to Th2 and Th1, respectively. The aim of this study was to clarify the role of graft-derived NKT cells in inducing the spontaneous acceptance of rat orthotopic liver transplantation (OLTx)
Methods. The experimental groups were divided as follows: Group 1, BN to LEW low responder (acceptor) combination; Group 2, DA to LEW high responder (rejector) combination; naïve BN (Group 3) or LEW recipients (Group 4) with liver allografts from irradiated BN donors. The recipients had liver allografts from irradiated donors reconstituted from the following cell populations 24 hr before harvesting, spleen cells (SPCs, Group 5), Ig-SPCs (Group 6), Ig-NKR-P1-SPCs (Group 7), and Ig-TCRab-SPCs (Group 8)
Results. In Group 1, the percent of graft-derived NKT cells harvested on day 7 posttransplant were significantly higher than in Group 2. In the case of BN liver allografts that had been irradiated and reconstituted with cell populations including NKT cells (Groups 5 and 6), the mean graft survival (MST) was extended to 39.2±5.7 and 38.8±8.0 days, respectively. In contrast, when NKT cells were excluded (Groups 7 and 8), the grafts were acutely rejected within MST of 17.8±4.0 and 18.8±7.7 days, respectively. The concentrations of IL-10 and TGF-β, but not IL-4 in Ig-GICs culture supernatants were predominant in the acceptor, whereas those with IFN-γ predominated in the rejector.
Conclusions. Graft-derived NKT cells might be responsible for spontaneous acceptance in the rat OLTx.
Transplantation 12/2005; 80(12):1749-1755. · 4.00 Impact Factor
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ABSTRACT: Pancreas transplantation (PTx) has evolved as a clinical therapy to achieve sustained euglycemia. However, it remains unclear if naive diseased islets of the pancreas benefit from the avoidance of glucose toxicity by PTx. In the present study, using an animal model of type 2 diabetes, the Spontaneously Diabetic Torii (SDT; RT1a) rat, we syngeneically transplanted nondiabetic 10-week-old pancreaticduodenal grafts into diabetic 25-week-old recipients. In the control SDT rats that received no treatment, hyperglycemia developed with a mean onset time of 25 +/- 3.9 weeks of age. Few normal islet cells were found from 25 weeks and none at 40 weeks. However, in the PTx rats, the onset age (graft age) of diabetes was significantly prolonged (47 +/- 18.2 weeks). Moreover, we found that the beta-cell mass was significantly increased in the naive pancreases of 40-week-old PTx recipients (PTx40-naive). Interestingly, islet-like cell clusters of varying size were found close to ductal structures of PTx40-naive pancreases, suggesting that these cells are derived from ductal cells. Furthermore, pancreatic and duodenal homeobox factor-1 (PDX-1) was more clearly expressed in the nuclei of PTx40-naive pancreatic islet-like cell clusters. Our results demonstrate the development of duct-derived beta cells in the pancreas of type 2 diabetic recipients after PTx.
American Journal of Transplantation 11/2005; 5(10):2360-7. · 6.39 Impact Factor
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ABSTRACT: CTLA4Ig gene transfer directly to graft tissue might have the potential to avoid the need for systemic immunosuppression. In our previous studies of bio-breeding (BB) rats, local adenovirus-mediated CTLA4Ig gene transfer protected the pancreas from autoimmune and alloimmune responses. This study investigated the potency of local CD28/B7 costimulatory blockade for induction of donor-specific tolerance and further examined the existing mechanisms.
Brown Norway (BN; RT1)-pancreaticoduodenal grafts transfected with Ad.CTLA4Ig via intraarterial ex vivo perfusion were transplanted into streptozotocin-induced diabetic Lewis (LEW; RT1) rats.
Ad.CTLA4Ig transduced grafts combined with a short course of FK506 resulted in indefinitely prolonged survival (>156 days vs. 19.5 days with FK506 alone). CTLA4Ig was predominantly expressed in grafts on day 4. The expression was gradually diminished and was only slightly detectable at day >100. The proliferative responses against BN antigen were remarkably enhanced among recipients with rejected grafts, but the T-cells from tolerant recipients (>100 days) showed poor cytotoxic responses. On adoptive transfer assay, the splenic T-cells of tolerant recipients were able to suppress the rejection of BN, but not third-party Wistar Furth (WF; RT1) hearts in irradiated (480 cGy) LEW recipients. The percentage of CD4CD25 splenic T-cells was significantly increased in tolerant recipients (13.53 +/- 4.06% vs. 6.06 +/- 0.56% in naive rats).
CTLA4Ig gene transfer to the pancreaticoduodenal allograft combined with a short course of FK506 induces donor-specific tolerance. The mechanism of maintaining tolerance could be explained by development of splenic T suppressor cells.
Transplantation 07/2004; 78(1):59-64. · 4.00 Impact Factor
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ABSTRACT: In terms of the temporal relationship between pancreas transplantation (PTx) and reversal of diabetic ocular complications, it has been difficult but important to determine a "point of no return." Thus, it is of great clinical interest to evaluate the efficacy of PTx on diabetic ocular complications.
A spontaneous type 2 diabetic model of Spontaneously Diabetic Torii (SDT; RT1) rats was used in the present study, and syngeneic PTx was performed.
In the control SDT rats that received no treatment, hyperglycemia (>250 mg/dL) was developed from 25.2+/-3.9 weeks of age. Lens opacity was observed in all rats at 15 weeks after the onset of diabetes. Fluorescein angiography and immunohistochemistry detected the nonperfusion area and neovascularization in the retina at 5 weeks of diabetes. Daily insulin treatment could not prevent or reverse the ocular changes in our experiment. Fluorescein filling defect of the retinal vessels was observed at 10 weeks of diabetes. However, in the PTx rats, normoglycemia was achieved at all experimental time points. Diabetic cataract and retinopathy could have been prevented and improved if PTx had been performed at 5 weeks, but not at 10 weeks after the onset of diabetes. With PTx treatment, an inhibition of angiogenesis in the retina at 5 weeks after the onset of diabetes was demonstrated by immunohistochemistry.
Our results indicate that the potential use of the SDT rat for diabetes study and the positive effect of PTx performed before the "point of no return" could prevent and cure diabetic ocular complications.
Transplantation 03/2004; 77(5):658-63. · 4.00 Impact Factor
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ABSTRACT: Pancreas and islet transplantation is the only treatment that can cure type 1 diabetes mellitus (DM). With the recent advances of operative procedure and immunosuppression, pancreas graft survivals have become better than before, but some problems still remain. It is extremely difficult to establish tolerance and to reverse rejection once it has been initiated because the pancreas graft itself has a strong immunogenicity. It is also known that pancreatic graft failure is sometimes due to autoimmune recurrence. In the clinical situation, however, these immunologic events actually coexist within the pancreatic graft. Thus, it is rather difficult to analyze each of them independently, but possible to delineate each of these immunologic mechanisms with using animal models of type 1 DM such as BB (BioBreeding) rats or NOD (nonobese diabetic) mice. In the current study, we reviewed the immunological characteristics in pancreas transplantation (PTx) based on our experimental experiences together with that of others and investigated the possibility of tolerance induction in PTx.
Pancreas 08/2003; 27(1):31-7. · 2.39 Impact Factor
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ABSTRACT: Pancreas and islet transplantation is the only treatment that can cure type 1 diabetes mellitus (DM). With the recent advances of operative procedure and immunosuppression, pancreas graft survivals have become better than before, but some problems still remain. It is extremely difficult to establish tolerance and to reverse rejection once it has been initiated because the pancreas graft itself has a strong immunogenicity. It is also known that pancreatic graft failure is sometimes due to autoimmune recurrence. In the clinical situation, however, these immunologic events actually coexist within the pancreatic graft. Thus, it is rather difficult to analyze each of them independently, but possible to delineate each of these immunologic mechanisms with using animal models of type 1 DM such as BB (BioBreeding) rats or NOD (nonobese diabetic) mice. In the current study, we reviewed the immunological characteristics in pancreas transplantation (PTx) based on our experimental experiences together with that of others and investigated the possibility of tolerance induction in PTx.
Pancreas 06/2003; 27(1):31-37. · 2.39 Impact Factor
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ABSTRACT: Adenovirus-mediated CTLA4Ig gene transfer has been reported to enhance graft survival in several rodent transplantation models. In this study, we investigated the efficacy of ex vivo and systemic transfer of the CTLA4Ig gene by adenoviral vectors in pancreatic islet allo-transplantation. Islet grafts from BN rats were transplanted to chemically induced diabetic LEW rats. First, ex vivo CTLA4Ig gene transfer into isolated islets was performed prior to transplantation. Survival of transduced grafts under the kidney capsule was slightly prolonged (8.6+/-1.3 days) compared with survival of untransduced grafts (6.7+/-1.2 days); when combined with a short course of FK506, graft survival was further extended (32.6+/-10.7 days vs. 13.7+/-1.0 days with FK506 alone). Secondly, systemic gene transfer was accomplished by intravenous administration immediately after the transplantation procedure. In these animals, islet grafts under the kidney capsule survived longer (15.2+/-3.3 days) than in controls (6.7+/-1.2 days), and when FK506 was administered perioperatively, all the islet grafts survived for more than 100 days. In systemically transduced recipients, the survival of islet grafts transplanted into the liver was not significantly different from that of the grafts placed under the kidney capsule. In order to examine organ-specific immunogenicity, heterotopic BN cardiac grafts were transplanted to LEW rats intra-abdominally, with the virus transferred systemically as in the islet model. In contrast to the islet grafts, all the cardiac grafts were accepted for longer than 100 days, even without FK506 therapy. Finally, the LEW recipients with long-surviving islet or cardiac grafts were re-transplanted with islet grafts from the same donor strain (BN) on day 100. The second islet grafts survived longer than 100 days in half of the cardiac recipients, but consistently failed in the islet recipients. We conclude that in this transplant model, CTLA4Ig gene transfer and FK506 treatment synergistically improved islet graft survival, systemic transfer of the gene was more effective than ex vivo transfer to the islets, and donor-specific tolerance could not be achieved for islet transplantation but was achieved for cardiac transplantation.
Transplant Immunology 11(1):91-100. · 1.46 Impact Factor
