In vitro lymphocyte recognition of islet cells following in vivo priming with allogeneic murine pancreatic islets

Summary Non-islet-antigen specific treatments have been shown to alter the natural history of insulin dependent diabetes in both the non-obese diabetic (NOD) mouse and in recently diagnosed patients. However, concerns have been raised regarding the possibility that non-islet-antigen specific therapy may trade cell mediated autoimmunity for antibody dependent autoimmunity. Female NOD mice at approximately 70 days of age were treated with the non-islet-antigen specific agents complete Freund's adjuvant (CFA) and Bacillus Calmette-Guerin (BCG) and assayed for the development of antibody mediated autoimmunity at 300 days of age. Autoantibodies to red cells were not detected in any ofthe BCG {n = 19) or CFA [n = 15) treated animals, while 2 of 13 age-matched NOD animals had autoantibodies to red cells, shown by a positive direct Coomb's test. Anti-nuclear autoantibodies and complement deposition in the renal glomeruli were not significantly increased in the treated animals as compared to age-matched non-diabetic mice. The relative effectiveness of CFA and BCG treatment was examined in terms ofthe ability of these agents to preserve insulin containing islets. Complete Freund's adjuvant treatment was found to be more effective in preserving insulin containing islets when compared to BCG treatment. This study demonstrates that it is possible to inhibit the development of autoimmune diabetes without increasing the probability that treated animals will develop antibody dependent autoimmunity.

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Glucose-Stimulated Insulin Release by Individual Pancreatic Cells: Potentiation by Glyburide

Egan¹,

Asplin²,

Drumheller³

et al. 1991

Experimental Biology and Medicine

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To investigate the effect of glyburide on insulin secretion by individual beta cells from normal rats, we employed a reverse hemolytic plaque assay. Pancreata were harvested from female Wistar-Furth rats, the pancreatic islets isolated, and the latter dispersed into single cells. These cells were mixed with protein A-coated ox erythrocytes, the mixture was placed in a Cunningham chamber in the presence of insulin antiserum, and the cells were exposed to the various test substances. Having developed hemolytic plaques around the insulin-secreting cells with complement, the percentage of plaque-forming cells was determined and the plaque areas (reflecting the amount of insulin secreted) were quantitated. For the purpose of validation, we demonstrated that (i) plaque-forming (but not nonplaque-forming) cells could be identified as insulin secreting by an independent immunofluorescent technique, (ii), plaques did not form if insulin antiserum was deleted from the preparation, (iii) plaques failed to develop if insulin antiserum was preabsorbed with insulin, and (iv) incubation with non-protein A-coated RBC or omission of complement resulted in no plaque formation. In addition, both the percentage of plaque-forming cells and the mean plaque are increased upon exposure to glucose (0.75-20 mM) in a concentration-dependent manner at 5- and 60-min incubation times. Moreover, somatostatin suppressed the percentage of plaque-forming cells and diminished the mean plaque area of cells which continued to secrete insulin in response to glucose. Exposure of cells to 100 nM glyburide in the presence of 5 mM or 20 mM glucose had no effect on the percentage of plaque-forming cells present at 5 min or 60 min. Similarly, glyburide did not alter mean plaque area at 5 or 60 min when cells were co-incubated with 5 mM glucose. However, mean plaque area was markedly enhanced at 5 and 60 min in response to glyburide and 20 mM glucose. These results demonstrate that glyburide (i) does appear to enhance insulin secretion by an effect directly on the pancreatic beta cell; (ii) does not act by recruiting previously noninsulin-secreting cells into a secretory pool; (iii) does not potentiate the effect of glucose, at fed concentrations, on insulin secretion by individual cells; but (iv) does augment insulin secretion by beta cells stimulated with supraphysiologic concentrations of glucose.

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In vitro lymphocyte recognition of islet cells following in vivo priming with allogeneic murine pancreatic islets

Cited by 3 publications

References 16 publications

Chirurgie des Diabetes mellitus

Chirurgie des Diabetes mellitus

Regulation of autoimmune diabetes: Characteristics of non‐islet‐antigen specific therapies

Glucose-Stimulated Insulin Release by Individual Pancreatic Cells: Potentiation by Glyburide

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