Effects of prolonged exposure to pancreatic glucagon on the function, antigenicity and survival of isolated human islets

Background Certain clinical conditions are associated with inappropriately high levels of circulating glucagon. To date, little information is availab le about the direct effects of prolonged exposure of human islet cells to p ancreatic glucagon. In the present study we evaluated the function, antigen icity and survival of human islets exposed for 24h to human pancreatic gluc agon. Methods We prepared human islets of Langerhans by collagenase digestion and density-gradient purification, incubated them for 24h with 44 or 430 pmol/ l pancreatic glucagon at physiological (5.5 mmol/l) glucose level, and eval uated their insulin release function, which was then compared with that obt ained from islets kept at high (11.1 mmol/l) glucose concentration. In addi tion, aliquots of the islets were evaluated to assess their chemotactic pro perties towards human monocyte-macrophage cells, and their potency to induc e cytokine release from human lymphocytes. Finally, survival of the islet c ells cultured under varying conditions was evaluated, and an assessment was performed of mRNA expression of Bcl-2 and Bax proteins. Results The insulin secretion results demonstrated that, compared to the co ntrol islets, the islets previously exposed to either 44 or 430 pmol/l gluc agon exhibited changes in insulin release in response to glucose, consistin g of augmented secretion at low glucose challenge, and no further significa nt increase at high glucose stimulation, similar to the effects observed wi th islets pre-cultured with high glucose. These effects were reversible, as documented by the recovery of normal islet sensitivity to glucose after an additional 24-h culture in medium lacking glucagon. Compared to control is lets, the culture medium from islets pre-cultured with high glucagon or hig h glucose showed an increased chemotactic potency towards human monocyte-ma crophage cells. In addition, human lymphocytes released a greater amount of tumour necrosis factor alpha when co-cultured with the islets pre-exposed to high glucagon or high glucose, whereas no significant difference was obs erved (in comparison with control islets) as regards the release of gamma-i nterferon, interleukin-2 and interleukin-10. The TUNEL technique and RT-PCR showed, respectively, no major difference in cell survival and expression of mRNA encoding for Bcl-2 and Bar protein between control islets and islet s kept for 24 h in the presence of high glucagon or high glucose. Conclusions Our results show that in vitro exposure of human islets to panc reatic glucagon for 24 h causes changes in the function and antigenicity of isolated human islets that are similar to the changes observed after pre-c ulture with increased glucose levels. Under our experimental conditions, th ese changes were not accompanied by any evidence of cytotoxicity. Copyright (C) 2000 John Wiley & Sons, Ltd.