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G S Korbutt

Publications and source records attributed to G S Korbutt.

At least 19 recordsLinked to original sources
PubMed2001

Clinical outcomes and insulin secretion after islet transplantation with the Edmonton protocol.

Islet transplantation offers the prospect of good glycemic control without major surgical risks. After our initial report of successful islet transplantation, we now provide further data on 12 type 1 diabetic patients with brittle diabetes or problems with hypoglycemia previous to 1 November 2000. Details of metabolic control, acute complications associated with islet transplantation, and long-term complications related to immunosuppression therapy and diabetes were noted. Insulin secretion, both acute and over 30 min, was determined after intravenous glucose tolerance tests (IVGTTs). The median follow-up was 10.2 months (CI 6.5-17.4), and the longest was 20 months. Glucose control was stable, with pretransplant fasting and meal tolerance-stimulated glucose levels of 12.5+/-1.9 and 20.0+/-2.7 mmol/l, respectively, but decreased significantly, with posttransplant levels of 6.3+/-0.3 and 7.5+/-0.6 mmol/l, respectively (P < 0.006). All patients have sustained insulin production, as evidenced by the most current baseline C-peptide levels 0.66+/-0.06 nmol/l, increasing to 1.29+/-0.25 nmol/l 90 min after the meal-tolerance test. The mean HbA1c level decreased from 8.3+/-0.5% to the current level of 5.8+/-0.1% (P < 0.001). Presently, four patients have normal glucose tolerance, five have impaired glucose tolerance, and three have post-islet transplant diabetes (two of whom need oral hypoglycemic agents and low-dose insulin (<10 U/day). Three patients had a temporary increase in their liver-function tests. One patient had a thrombosis of a peripheral branch of the right portal vein, and two of the early patients had bleeding from the hepatic needle puncture site; but these technical problems were resolved. Two patients had transient vitreous hemorrhages. The two patients with elevated creatinine levels pretransplant had a significant increase in serum creatinine in the long term, although the mean serum creatinine of the group was unchanged. The cholesterol increased in five patients, and lipid-lowering therapy was required for three patients. No patient has developed cytomegalovirus infection or disease, posttransplant lymphoproliferative disorder, malignancies, or serious infection to date. None of the patients have been sensitized to donor antigen. In 11 of the 12 patients, insulin independence was achieved after 9,000 islet equivalents (IEs) per kilogram were transplanted. The acute insulin response and the insulin area under the curve (AUC) after IVGTT were consistently maintained over time. The insulin AUC from the IVGTT correlated to the number of islets transplanted, but more closely correlated when the cold ischemia time was taken into consideration (r = 0.83, P < 0.001). Islet transplantation has successfully corrected labile type 1 diabetes and problems with hypoglycemia, and our results show persistent insulin secretion. After a minimum of 9,000 IEs per kilogram are provided, insulin independence is usually attained. An elevation of creatinine appears to be a contraindication to this immunosuppressive regimen. For the subjects who had labile type 1 diabetes that was difficult to control, the risk-to-benefit ratio is in favor of islet transplantation.

Adult↗
PubMed2000

Glucose-dependent insulin release from genetically engineered K cells.

Genetic engineering of non-beta cells to release insulin upon feeding could be a therapeutic modality for patients with diabetes. A tumor-derived K-cell line was induced to produce human insulin by providing the cells with the human insulin gene linked to the 5'-regulatory region of the gene encoding glucose-dependent insulinotropic polypeptide (GIP). Mice expressing this transgene produced human insulin specifically in gut K cells. This insulin protected the mice from developing diabetes and maintained glucose tolerance after destruction of the native insulin-producing beta cells.

Animals↗
PubMed2000

Single injection of insulin delays the recurrence of diabetes in syngeneic islet-transplanted diabetic NOD mice.

BACKGROUND: Insulin has been implicated in the pathogenesis of type 1 diabetes and oral administration of insulin has been shown to delay the onset of diabetes in NOD mice. In this study we determined whether a single footpad injection of insulin will protect syngeneic islet grafts from autoimmune destruction when placed under the kidney capsule of diabetic NOD mice. METHODS: Five hundred islets were transplanted under the kidney capsule of diabetic female NOD mice in conjunction with a single footpad injection of either pork insulin in saline or mixed with incomplete Freund's adjuvant (IFA). Control groups received either IFA or saline alone. RESULTS: Seven of 11 animals (63.6%) given insulin in IFA exhibit long-term graft survival (>75 days; mean +/- SEM >85.4+/-16.1) whereas only 3 of 12 animals (25.0%) in the IFA group had graft survival longer than 75 days (mean +/- SEM >41.9+/-12.8 days). In contrast, none of the animals that received insulin in saline (17.3+/-2.5 days) and saline only (16.1+2.0 days) exhibit prolonged graft survival. CONCLUSION: These results suggest that a single footpad injection of insulin can protect the islet graft from immune attack in NOD mice.

Animals↗
PubMed2000

Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen.

BACKGROUND: Registry data on patients with type 1 diabetes mellitus who undergo pancreatic islet transplantation indicate that only 8 percent are free of the need for insulin therapy at one year. METHODS: Seven consecutive patients with type 1 diabetes and a history of severe hypoglycemia and metabolic instability underwent islet transplantation in conjunction with a glucocorticoid-free immunosuppressive regimen consisting of sirolimus, tacrolimus, and daclizumab. Islets were isolated by ductal perfusion with cold, purified collagenase, digested and purified in xenoprotein-free medium, and transplanted immediately by means of a percutaneous transhepatic portal embolization. RESULTS: All seven patients quickly attained sustained insulin independence after transplantation of a mean (+/-SD) islet mass of 11,547+/-1604 islet equivalents per kilogram of body weight (median follow-up, 11.9 months; range, 4.4 to 14.9). All recipients required islets from two donor pancreases, and one required a third transplant from two donors to achieve sustained insulin independence. The mean glycosylated hemoglobin values were normal after transplantation in all recipients. The mean amplitude of glycemic excursions (a measure of fluctuations in blood glucose concentrations) was significantly decreased after the attainment of insulin independence (from 198+/-32 mg per deciliter [11.1+/-1.8 mmol per liter] before transplantation to 119+/-37 mg per deciliter [6.7+/-2.1 mmol per liter] after the first transplantation and 51+/-30 mg per deciliter [2.8+/-1.7 mmol per liter] after the attainment of insulin independence; P<0.001). There were no further episodes of hypoglycemic coma. Complications were minor, and there were no significant increases in lipid concentrations during follow-up. CONCLUSIONS: Our observations in patients with type 1 diabetes indicate that islet transplantation can result in insulin independence with excellent metabolic control when glucocorticoid-free immunosuppression is combined with the infusion of an adequate islet mass.

Adult↗
PubMed2000

Microencapsulation of neonatal porcine islets: protection from human antibody/complement-mediated cytolysis in vitro and long-term reversal of diabetes in nude mice.

BACKGROUND: Recently, we have developed a simple and reliable method to efficiently isolate large numbers of neonatal porcine islets (NPI). We and others have shown that NPI are susceptible to cytolysis by the activation of human complement in vitro. Microencapsulation of islets may be one strategy to protect NPI from this form of rejection. We examined whether microencapsulation can prevent lysis of NPI induced by human antibody and complement in vitro and also assessed their ability to reverse hyperglycemia in diabetic nude mice. METHODS: NPI were microencapsulated with purified alginate, cultured for 2 days, then tested for sensitivity to fresh human serum using an established in vitro cytotoxicity assay or transplanted into alloxan-induced diabetic nude mice. RESULTS: Incubation of nonencapsulated NPI for 24 hr in the presence of fresh human serum resulted in a 53% loss of cellular insulin content, a 51% reduction in recoverable DNA content, and a marked reduction of insulin secretory responsiveness when compared with controls cultured in heat-inactivated human serum. In contrast, exposure of encapsulated islets to fresh human serum had no cytotoxic effect on the islets. Transplantation of 2000 encapsulated NPI i.p. into diabetic nude mice (n=16) corrected hyperglycemia in all mice within 8 weeks. Similar results were obtained when 2000 nonencapsulated NPI were implanted under the kidney capsule (n=10); however recipients of nonencapsulated NPI placed i.p. failed to obtain euglycemia and survived for only 3 weeks posttransplantation. CONCLUSION: Microencapsulation protects NPI from the cytotoxic effects of human antibody and complement and allows for long-term reversal of diabetes in nude mice.

Animals↗
PubMed2000

Testicular Sertoli cells exert both protective and destructive effects on syngeneic islet grafts in non-obese diabetic mice.

AIMS/HYPOTHESIS: Testicular Sertoli cells protect allogeneic islet grafts from rejection after transplantation into animals with chemically induced diabetes. The aims of this study were to determine whether Sertoli cells can protect syngeneic islets from autoimmune destruction after transplantation into non-obese diabetic (NOD) mice and, if so, whether protection is due to Sertoli cell expression of Fas ligand (FasL), believed to be the mechanism that protects against allograft rejection. METHODS: We compared the survival of syngeneic islets transplanted under the renal capsule of nonobese diabetic mice, alone and together with purified Sertoli cells prepared from testes of newborn nonobese diabetic mice. Additionally, we examined the composition of the islet and Sertoli cell co-transplants by immunohistochemistry to determine whether islet graft survival correlated with Sertoli cell expression of Fas ligand. RESULTS: Sertoli cell doses of 1, 2 and 4 x 10(6) cells produced a dose-dependent prolongation of median islet graft survival from 11 days (islets alone) to 32 days (islets + 4 x 10(6) Sertoli cells); addition of 8 x 10(6) Sertoli cells to the islet grafts decreased, however, median survival to 8 days. Immunohistochemical analysis of the islet and Sertoli cell co-transplants showed a correlation between Fas ligand expression by Sertoli cells and graft infiltration by neutrophilic leucocytes, leading to islet beta-cell destruction and diabetes recurrence. CONCLUSION/INTERPRETATION: Sertoli cells exert opposing effects on survival of syngeneic islet grafts in nonobese diabetic mice: Fas ligand-dependent neutrophil infiltration and graft destruction, and Fas ligand-independent protection of the graft from autoimmune destruction.

Animals↗
PubMed2000

Testicular sertoli cells protect islet beta-cells from autoimmune destruction in NOD mice by a transforming growth factor-beta1-dependent mechanism.

Testicular Sertoli cells protect pancreatic islet grafts from allo- and autoimmune destruction; however, the mechanism(s) of protection is unclear. The aim of this study was to determine whether Fas ligand (FasL) and/or transforming growth factor (TGF)-beta, immunoregulatory proteins produced by Sertoli cells, might mediate the protective effects of these cells against autoimmune destruction of islet beta-cells. Sertoli cells were purified from testes of NOD mice and implanted under the right renal capsule of diabetic NOD mice, whereas NOD islets were implanted under the left renal capsule. Of the mice that received islet and Sertoli cells grafts, 64% (9 of 14) remained normoglycemic at 60 days posttransplantation compared with 0% (0 of 6) of the mice that received islet grafts alone. Immunohistochemical examination of Sertoli cell grafts in normoglycemic mice revealed that TGF-beta1 expression by Sertoli cells remained high, whereas FasL expression by Sertoli cells decreased progressively posttransplantation. Also, plasma levels of TGF-beta1 were significantly elevated in mice that received Sertoli cells and islet grafts, and anti-TGF-beta1 antibody administration completely abrogated the protective effect of Sertoli cells on islet graft survival, whereas anti-FasL antibody did not. Islet graft destruction in anti-TGF-beta1-treated mice was associated with increases in interferon (IFN)-gamma-producing cells and decreases in interleukin (IL)-4-producing cells in the islet grafts. We conclude that 1) Sertoli cell production of TGF-beta1, not FasL, protects islet beta-cells from autoimmune destruction and 2) TGF-beta1 diverts islet-infiltrating cells from a beta-cell-destructive (IFN-gamma+) phenotype to a nondestructive (IL-4+) phenotype.

Animals↗
PubMed1999

Potential application of neonatal porcine islets as treatment for type 1 diabetes: a review.

Islet transplantation has been shown to be a viable option for treating patients with type 1 diabetes. However, widespread clinical application of this treatment will necessitate an alternative source of insulin-producing tissue. Porcine pancreata may be a potential source of islets since pigs are inexpensive, readily available, and exhibit morphological and physiological characteristics comparable to humans. Recently, we developed a simple, standardized procedure for isolating large numbers of neonatal porcine islets with a reproducible and defined cellular composition. Following nine days of in vitro culture, tissue from one neonatal pig pancreas yielded approximately 50,000 islet cell aggregates, consisting of primarily epithelial cells (57%) and pancreatic endocrine cells (35%). In addition, neonatal porcine islets were responsive to glucose challenge in vitro and were capable of correcting hyperglycemia in alloxan-induced diabetic nude mice. Although neonatal porcine islets constitute an attractive alternative source of insulin-producing tissue for clinical transplantation, many aspects such as the immunological responses to these tissue and the latent period (2 to 8 weeks) between transplantation of these islets and the reversal of hyperglycemia need further investigation. This article discusses these issues and presents possible solutions to problems that may hinder the potential application of neonatal porcine islets for transplantation into patients with type 1 diabetes.

Animals↗
PubMed1999

Transfection of human pancreatic islets with an anti-apoptotic gene (bcl-2) protects beta-cells from cytokine-induced destruction.

Apoptosis has been identified as a mechanism of pancreatic islet beta-cell death in autoimmune diabetes. Proinflammatory cytokines are candidate mediators of beta-cell death in autoimmune diabetes, and these cytokines can induce beta-cell death by apoptosis. In the present study, we examined whether transfection of human islet beta-cells with an anti-apoptotic gene, bcl-2, can prevent cytokine-induced beta-cell destruction. Human islet beta-cells were transfected by a replication-defective herpes simplex virus (HSV) amplicon vector that expressed the bcl-2 gene (HSVbcl-2) and, as a control, the same HSV vector that expressed a beta-galactosidase reporter gene (HSVlac). Two-color immunohistochemical staining revealed that 95+/-3% of beta-cells transfected with HSVbcl-2 expressed Bcl-2 protein compared with 14+/-3% of beta-cells transfected with HSVlac and 19+/-4% of nontransfected beta-cells. The bcl-2-transfected beta-cells were fully protected from impaired insulin secretion and destruction resulting from incubation for 5 days with the cytokine combination of interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma. In addition, the bcl-2-transfected islet cells were significantly protected from cytokine-induced lipid peroxidation and DNA fragmentation. These results demonstrate that cytokine-induced beta-cell dysfunction and death involve mechanisms subject to regulation by an anti-apoptotic protein, Bcl-2. Therefore, bcl-2 gene therapy has the potential to protect human beta-cells in pancreatic islets, or islet grafts, from immune-mediated damage in type 1 diabetes.

Apoptosis↗
PubMed1999

Neonatal porcine islet cells induce human CD4+, but not CD8+, lymphocyte proliferation and resist cell-mediated cytolytic injury in vitro.

Xenotransplantation of porcine tissue to human recipients promises to alleviate the organ shortage. Human antibody-mediated and cell-mediated immune responses against porcine grafts, however, represent barriers to successful xenotransplantation. We compared neonatal porcine islet cells (NPICs) and neonatal porcine splenocytes for the ability to stimulate proliferation of human peripheral blood lymphocytes (PBLs), and for their susceptibility to human natural killer (NK) and cytotoxic T-lymphocyte (CTL)-mediated lysis. Human peripheral blood CD4+ lymphocytes showed strong proliferation in response to NPICs, likely because of occasional swine leukocyte antigen (SLA) class II+ cells in the NPIC preparations. In contrast, human peripheral blood CD8+ lymphocytes did not proliferate in response to NPICs, although they showed clear responses to both porcine splenocytes and endothelial cells. Both human CTL-raised-against-porcine splenocytes and endogenous NK cells lysed porcine splenocytes, but the same cells showed little or no lytic activity against NPICs. Lysis of porcine splenocyte targets was completely abrogated by pretreatment of the human NK or CTL populations with concana-mycin A, suggesting a perforin-dependent effector mechanism. Pretreatment of the NPIC targets with proinflammatory porcine cytokines to upregulate SLA class I expression failed to enhance human CTL-mediated lysis. However, lysis of NPICs by human CTLs could be elicited when a lectin was added to form stable effector:target cell conjugates. It appears that NPICs do not express sufficiently high levels of co-stimulatory and/or adhesion molecules to either activate human CD8+ T-cells or to be effective targets for activated human CTLs. These data suggest that NPICs may not be destroyed by NK- or CTL-mediated lytic mechanisms after transplantation into humans.

Animals↗
PubMed1998

Improved survival of biolistically transfected mouse islet allografts expressing CTLA4-Ig or soluble Fas ligand.

BACKGROUND: Pancreatic islet transplantation is limited because of immune rejection of the transplanted tissue. Long-term survival of allogeneic pancreatic islet grafts in the absence of systemic immunosuppressive agents should be possible by transfecting the islets directly with DNA encoding immunoregulatory molecules. Localized production of these molecules should affect only the immune cells that come into the vicinity of the foreign tissue. We investigated whether local expression of human CTLA4-Ig or soluble human Fas ligand from biolistically transfected mouse islets would have a protective effect on allograft survival. METHODS: Isolated CBA (H2k) islets were biolistically transfected using the gene gun. The experimental groups were naked gold particles (n=6), empty vector DNA (n=5), DNA encoding human CTLA4-Ig (n=8), or soluble human Fas ligand (n=5). Secretion of the transfected gene product was confirmed by screening islet culture supernatants for protein production using a sandwich ELISA. The blasted islets were transplanted under the kidney capsule of alloxan-diabetic BALB/c (H2d) recipients. RESULTS: Control grafts survived for 23 days, on average. CTLA4-Ig-transfected islets showed a bimodal distribution: 50% of cases survived > or = 46 days and 50% were similar to the controls. In the soluble human Fas ligand group, 80% of grafts survived > or = 50 days. There was no correlation between graft survival times and pretransplant levels of protein production. CONCLUSION: Our results indicate that local production of human CTLA4-Ig or soluble human Fas ligand by biolistically transfected islets can promote allograft survival. This approach should be valuable as a potential immunoprotective therapeutic strategy in tissue transplantation.

Abatacept↗