Tetsu Nagai

Pancreatic Endocrine Dysfunction in Rats Not Expressing the Cholecystokinin-a Receptor

Summary Cholecystokinin (CCK) has been suggested to modulate insulin output. We have shown that Otsuka Long-Evans Tokushima Fatty (OLETF) rats show little or no expression of the CCK-A receptor gene in the pancreas. We examined whether the CCK-A and CCK-B receptor genes are expressed in the islets and the role of CCK-A receptor in insulin secretion. Gene expressions of CCK receptors were determined by the reverse-transcriptase polymerase chain reaction (RT-PCR) followed by Southern blot hybridization and Northern transfer analysis using LETO rats as controls. Pancreatic endocrine function was examined in perfusion (exogenous CCK stimulation) and meal ingestion (endogenous CCK stimulation) studies. CCK-A receptor mRNA was detected in the islets of LETO rats but not OLETF rats. Expression of the CCK-B receptor gene was detected in both strains by RT-PCR. Insulin secretion was impaired in OLETF rats, but the insulin contents of OLETF and LETO rats were not different. No abnormalities were detected histologically in either strain. These results suggest that the occurrence of pancreatic endocrine dysfunction in OLETF rats may be due to a defect in expression of the CCK-A receptor gene, not to insulin deficiency.

Expansion of intermediate T cell receptor cells expressing interleukin-2 receptor alpha- beta+, CD8alpha+ beta+, and lymphocyte function-associated antigen-1+ in the liver in association with intrahepatic islet xenograft rejection from rat to mouse: prevention of rejection with anti-interleukin-2 receptor beta monoclonal antibody treatment.

BACKGROUND The precise mechanisms involved in islet xenograft rejection remain unknown. The purpose of the present study was to determine cellular mechanisms responsible for islet xenograft rejection in the liver to facilitate finding a procedure for prevention of immune rejection. METHODS Hepatic mononuclear cells (MNC) as well as splenocytes, peripheral blood MNC, and thymocytes from streptozotocin-induced diabetic mice (BALB/c) rejecting the intrahepatic rat (Lewis) islet xenografts were isolated and examined by two-color FACS analysis. RESULTS The characteristic finding of the hepatic MNC from the mice rejecting islet xenografts compared with mice receiving isografts was a significant increase in the yield as well as in the percentage of the cells expressing CD3+ interleukin-2 receptor (IL-2R) alpha- beta+, CD3+ CD8alpha+ beta+, and T cell receptor (TCR) alphabeta+ lymphocyte function-associated antigen-1+. The expression of CD3 and TCR alphabeta of these T cells was found to be of intermediate intensity (TCR(int) cells). The expansion of these TCR(int) cells occurred predominantly in the liver. There was no significant difference in the cells expressing CD3+ IL-2R alpha+, CD3+ CD4+, CD3+ TCRgammadelta+, CD3- IL-2Rbeta+ (natural killer cells), and B220+ (B cells). In vivo administration of anti-IL-2Rbeta monoclonal antibody directed to the expanded cells produced a prevention of rejection. CONCLUSIONS These findings suggest that islet xenograft rejection in the liver from rat to mouse is an event for which the TCR(int) cells are responsible.

Amelioration of hyperglycemia in streptozotocin-induced diabetic rats receiving a marginal mass of islet grafts by troglitazone, an oral antidiabetic agent.

Troglitazone, a novel oral antidiabetic agent, was evaluated to determine whether it could have hypoglycemic effects in streptozotocin (STZ)-induced diabetic rats when a marginal mass of islets was transplanted and hyperglycemia persisted after transplantation. Lewis rats (RT1(1)) were used as both donors and recipients. Five hundred fresh islets were transplanted beneath the kidney capsule of STZ-induced diabetic recipients. Troglitazone was administered orally (0.34 mmol/kg/day) for 7 days before and for 60 days after islet transplantation. Neither troglitazone treatment without islet transplantation (n = 8) nor islet transplantation alone (n = 7) could produce normoglycemia (< 11 mmol/L) in diabetic recipients by 60 days after transplantation. In marked contrast, seven of 10 rats receiving islet grafts and treated with troglitazone became normoglycemia at 26.9 +/- 16.4 days (mean +/- SD; n = 7) after transplantation. Removal of the kidney bearing the grafts promptly resulted in the normoglycemic recipients (n = 4) becoming diabetic again. Light and electron microscopically, the intact islets with well-granulated beta cells could be observed in the transplant site of the normoglycemic recipients. These findings clearly demonstrate that the hyperglycemia in STZ-induced diabetic rats receiving an insufficient number of islet grafts to reverse diabetes was ameliorated by troglitazone treatment.

Improved human islet isolation by a tube method for collagenase infusion.

BACKGROUND Collagenase infusion into the pancreatic duct is an essential step in human islet isolation. We developed a new method for ductal canulation and collagenese infusion. METHODS A total of 53 pancreata were divided into two groups: group 1 (n=23), the new tube method, and group 2 (n=30), the standard angiocatheter method. In group 1, a polyethylene tube was inserted into the duct and pushed to the tail. The tail was first expanded, followed by expansion of the body and then the head, by pulling out the tube. RESULTS Total islet number and number/g pancreas (mean+/-SE) were significantly higher in group 1 (481,123+/-43,218 and 8,010+/-722) (mean+/-SE) than in group 2 (300,974+/-35,122 and 5,090+/-515, P<0.01). Total islet equivalent number and islet equivalent number per gram pancreas were also significantly higher in group 1 (319,176+/-39,354 and 5,455+/-652) than in group 2 (202,022+/-23,331 and 3,722+/-468, P<0.04). Islet purity and fragmentation showed no differences between the groups. CONCLUSIONS The tube method improved islet yields. We recommended this method for human islet isolation.