Inhibition of immune response in pancreatic islet transplantation: the glycan approach.

VIEIRO, MERCEDES; BARRIONUEVO, PAULA; CEBALLOS, CANDELA; HYON, SH; ISTURIZ, MARTÍN; ARGIBAY , PABLO

Viena, Austria.

Congreso; XX International Congress of The Transplantation Society.; 2004

Transplantation Society.

Aims: Host immune response has precluded clinical islet transplantation from becoming a consistent therapy for type 1 diabetic patients. O-glycosylated proteins, present in immune-reactive cells, may have a main role both in primary, unspecific, and in cellular, specific rejection. The purpose of this study was to investigate if the cleavage of these molecules could have an immunomodulatory effect on islet allotransplantation. Methods: We investigated the putative immunomodulatory effect of O-sialoglycoprotein endopeptidase (O-SEP) by studying three endpoints: a) proliferation in allogeneic mixed islet/lymphocyte reactions (MILR), b) expression of IA-d on monocytes in an in-vivo mouse model of allogeneic islet transplantation, and c) posttransplant graft function in an in-vivo model of allotransplantation. a) Allogeneic 5-day MILRs were established between islets from C57BL/6 and lymphocytes from C3H mice (50 islets/2x105 lymphocytes). Irradiated islets were used as stimulator cells with (treated group) or without (control group) previous incubation with O-SEP (0.12 mg/mL, at 37°C in 5% CO2 for 30 min). Tritiated thymidine was added 18 h before harvesting. Lymphocyte reaction was assessed by calculation of the allogeneic proliferative response (APR, cpm) and the proliferative inhibition rate of treated vs. control islets. Results are representative of five replicate experiments. b) Treated and untreated islets from C57BL/6 were transplanted into subcutaneous glass capsules of Balb-c mice and after three weeks of transplant animals were sacrificed. Expression of IA-d on monocytes was analyzed by flow cytometry as a percentage of the expression of IA-d in non transplanted mice. c) Two hundred pancreatic islets from C57BL/6 were transplanted under the kidney capsule of C3H mice previously made diabetic by injection of streptozotocin (270 mg/kg).  Four groups were tested: 1) islets treated with O-SEP plus cyclosporin (CsA) (n=6), 2) islets treated with CsA (n=6), 3) islets treated with O-SEP (n=6), and 4) non treated islets (n=8). Glycemia was monitored daily after transplantation and diabetes was considered as reversed upon three consecutive glycemias <250 mg/dL. Results: a) APR was maximal when allogeneic lymphocytes were mixed with untreated, control islets while APR significantly decreased with endopeptidase treated islets (individual data for control vs. treated islets, 7 294 vs. 1 525; 864 vs. 477; 689 vs. 172; 455 vs. 232; 1 987 vs. 728; respectively) (p<0.05, Wilcoxon test). Mean proliferative inhibition rate was 62%. b) Percentage of the expression of IA-d on monocytes, a molecule involved in inflammatory immune response, was also maximal in untreated islets (individual data for control vs. treated islets, 153, 99, 132, 145 vs. 80, 54, 94, 70, respectively) (p<0.01; paired t test). c) Reversion of hyperglycemia were significantly different (p<0.002, Wilcoxon Mann-Whitney U-test) between group 1) versus groups 2, 3 and 4. Duration of reversion varied from 3 to 7 days. Conclusions: These results suggest that treatment of islets with O-sialoglycoprotein endopeptidase may modulate the allogeneic immune reaction at different levels, both: in vitro and in vivo. Pretransplant islet treatment with O-sialoglycoprotein endopeptidase associated with CsA could have a protective effect over graft function during the early posttransplant period.