Stage-dependent effect of pancreatic transplantation on diabetic ocular complications in the Spontaneously Diabetic Torii rat.

BACKGROUND: 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. METHODS: A spontaneous type 2 diabetic model of Spontaneously Diabetic Torii (SDT; RT1) rats was used in the present study, and syngeneic PTx was performed. RESULTS: 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. CONCLUSIONS: 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.

Development of donor-specific immunoregulatory T-cells after local CTLA4Ig gene transfer to pancreatic allograft.

BACKGROUND: 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. METHODS: 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. RESULTS: 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). CONCLUSION: 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.

Reversal of diabetes in mice by xenotransplantation of a bioartificial pancreas in a prevascularized subcutaneous site.

BACKGROUND: The subcutaneous site has been regarded as a potential site for a bioartificial pancreas. Transplantation of islets, encapsulated by the development of diverse biocompatible materials and structural designs, can reverse hyperglycemia in diabetic recipients. METHODS: Approximately 750 Sprague-Dawley rat islets macroencapsulated in an agarose/poly (styrene sulfonic acid) mixed gel were implanted into a prevascularized subcutaneous site. The site was constructed by subcutaneous injection of basic fibroblast growth factor (bFGF)-impregnated gelatin microspheres in streptozotocin-induced C57BL/6 diabetic mice. Diabetic mice treated with bFGF-free gelatin microspheres and diabetic mice without any treatment undergoing the same subcutaneous transplantation were used as controls. After transplantation, non-fasting blood glucose, body weight, intraperitoneal glucose tolerance test, and histologic evaluations were processed. RESULTS: All the recipients undergoing the subcutaneous xenograft returned to normoglycemia within 1 week after transplantation. Eight of 10 recipients in the bFGF+ group maintained normoglycemia for a period of 38-101 days and gradually gained increase of body weight. Two of 10 recipients became hyperglycemic again when the grafts were respectively retrieved at days 31 and 63. Intraperitoneal glucose tolerance tests at month 1 and 2 revealed significant ameliorated glucose tolerance but a tendency to reduced glucose tolerance when compared respectively with those of the streptozotocin-induced diabetic mice and normal mice. Histologic examination revealed that islets within the retrieved grafts at days 31 and 63 were viable and intact; no fibrotic overgrowth was present around the surface of grafts. CONCLUSIONS: A successfully prevascularized subcutaneous site could be constructed by a tissue bioengineering approach. Xenotransplantation of the agarose/poly (styrene sulfonic acid) mixed gel-based bioartificial pancreas in the prevascularized subcutaneous site could reverse diabetes in mice.

Predicting operational tolerance in pediatric living-donor liver transplantation by absence of HLA antibodies.

BACKGROUND: The role of anti-human leukocyte antigen (HLA) antibodies in operational tolerance (OT) after pediatric living-donor liver transplantation (LDLT) remains inconclusive. We investigated whether the presence of HLA antibodies impeded the development of OT. METHODS: We retrospectively examined the prevalence of anti-HLA antibodies in pediatric LDLT recipients before transplantation and at 3 weeks after transplantation and analyzed the significance of those antibodies in relation to later OT. Forty pediatric LDLTs were performed between April 1996 and December 2000 and followed up through July 2011, with sera available for measurement of HLA antibodies. Seventeen patients achieved OT (mean follow-up, 4571.9±544.7 days) and 23 patients did not achieve OT (mean follow-up, 4532.0±425.4 days). Protocol liver biopsy was done for 14 OT patients and 16 non-OT patients. Their sera were tested for anti-HLA class I and II antibodies using the LABScreen single antigen beads test, in which a 1000 mean fluorescence value was considered positive. RESULTS: The prevalence of antibodies after transplantation in non-OT patients was higher than in OT patients (95.2% vs. 73.3%; P<0.001). The highest mean fluorescence intensity of antibodies was significantly higher in non-OT patients than in OT patients. The prevalence of HLA-B, HLA-C, HLA-DQ, and HLA-DR antibodies was significantly higher in non-OT patients than in OT patients. The highest mean fluorescence intensity of HLA-A, HLA-B, and HLA-DQ observed in non-OT patients was significantly higher than those in OT patients. CONCLUSIONS: In our study, posttransplantation HLA antibodies were associated with the future absence of OT. A prospective study with more patients is necessary to confirm the predictive value of HLA antibodies for OT.

Pancreas transplantation using type I and type II spontaneously diabetic rats--our experimental experience.

Pancreas transplantation (PTx) is the only therapy that can cure type 1 diabetes mellitus. With the recent advance of surgical procedures and immunosuppression, the outcome of PTx has become better than it used to be before, but some problems still remain. It is rather difficult to induce tolerance and to reverse rejection once it occurred because pancreas graft itself has a strong immunogenicity. Another important issue is regarding the recurrence of autoimmune disease in the pancreatic graft, therefore, some animal models are necessary to delineate and regulate those immune responses specific for PTx. Recently, PTx is also clinically applicable for type 2 diabetic patients with end-stage renal disease. It has been shown that insulin resistance was improved by PTx in type 2 diabetic recipients. In the current study, we have introduced some useful type 1 and type 2 diabetic models mainly based on our experimental experiences.

Development of islet-like cell clusters after pancreas transplantation in the spontaneously diabetic Torri rat.

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.

Consecutive organ transplantation in inbred rats: long-term results--Are organs recyclable?

Using 9-12-month-old Lewis rats, our laboratory performed consecutive organ transplantations to assess the technical feasibility and observe the histopathologies of prolonged transplants and implants over a rat's life span. This study includes a total of 688 cases of consecutive syngeneic organ transplantation, including pancreaticoduodenum (PD), en-bloc liver-pancreaticoduodenum-spleen-stomach (LPdSpSt), and spleen bearing ovary or testicle. A 52-month-old consecutively transplanted pancreas showed normal acini and islet cells. Consecutively transplanted LPdSpSt cases at 25-28 months preserved normal histology. After 26.5 months of ovarian follicle implantation into the spleen, the pathology of the organ exhibited typical dysgerminoma. However, 29.5-month-old infantile testicle implants in the spleen showed seminoma. Our studies suggest that transplanted organs can be reusable and retransplanted, and we hope this could be one of the clues to solve donor-organ shortages in the future.

Immunological characteristics of pancreas transplantation: review and our experimental experience.

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.