Pancreatic β-cells expressing GLP-1 are resistant to the toxic effects of immunosuppressive drugs

Abstract

Glucose intolerance is often observed after pancreatic islet cell transplantation. The administration of immunosuppressive agents (ISD), necessary to avoid tissue rejection, is in part responsible for hyperglycemia. To investigate whether mouse insulinoma (MIN6) cells transfected with the glucagon like peptide-1 (GLP-1) fragment of the proglucagon gene (RIP/GLP-1 MIN6 cells) are resistant to the toxicity derived from the administration of ISD. RIP/GLP-1 MIN6 cells, as well as parental MIN6 cells, were exposed to a cocktail of ISD. The secretion of insulin and the expression of apoptosis-related proteins were investigated by RIA and western blot analysis. Cell apoptosis was quantified by FACS analysis. Finally, to study whether the antiapoptotic action of GLP-1 was a function of its effect on insulin secretion, or rather it was a direct effect of GLP-1, cells were cultured with or without diazoxide or exendin-9. GLP-1 improved the functional activity and the viability of cells exposed to ISD. The insulin secretion of RIP/GLP-1 MIN6 cells after exposure to ISD was preserved. The expression of GLP-1 by -cells reduced the number of apoptotic cells and increased the expression of the antiapoptotic protein Bcl-2. GLP-1 also decreased the abundance of the proapoptotic markers PARP-p85 and Smac/Diablo. Treatment of cells with the diazoxide did not abolish the protective advantage that cells transfected with GLP-1 had; conversely the exposure of cells to exendin-9 was associated with a restored susceptibility to apoptosis. This report demonstrates that GLP-1 is capable of preserving -cell function and protecting cells from apoptotic cell death.

Hes3 Is Expressed in the Adult Pancreatic Islet and Regulates Gene Expression, Cell Growth, and Insulin Release

“…GLP-1 analogs activate the Akt pathway which, in NSCs, leads to phosphorylation of STAT3-serine, Hes3 transcription, and improved cell survival (10). In MIN6 cells, where Exendin-4 protects against pro-apoptotic insults (49), high concentrations of Exendin-4 significantly increased nuclear Hes3 expression in CC and DC. Although in this context it is not clear if changes in Hes3 expression/localization are responsible for the protective effects, it will be of interest to address this issue, especially with primary cell systems.…”

Mechanisms of action of glucagon-like peptide 1 in the pancreas

“…In biopsies taken from pancreata in 20 simultaneous kidney-pancreas transplant recipients on immunosuppressive regimens including FK506 and cyclosporin A, islet cell toxicity was observed in the form of cytoplasmic swelling, vacuolization and loss of secretory vesicles (Drachenberg et al, 1999). In the MIN6 cells D'Amico and colleagues (D'Amico et al, 2005) did observe increased necrosis and apoptosis, caspase 3 levels and levels of the proapoptotic markers PARP and Smac/Diablo in response to the cocktail of immunosuppressive drugs that they administered. The cells expressing GLP-1 exhibited high levels of Bcl-2 and were resistant to the effects of the immunosuppressive drugs.…”

“…Perfetti and co-workers expressed the GLP-1 fragment of the proglucagon gene under control of the rat insulin II promoter in MIN6 cells (D'Amico et al, 2005) and showed that this conferred protection against a cocktail of immunosuppressive reagents (sirolimus 25 ng/ml, mycophenolate 17.5 μg/ml and FK506 75 ng/ml) a commonly used regimen in organ transplantation. As discussed above in section 5.2, FK506 inhibits insulin synthesis but it has also been shown to cause reversible toxic effects to pancreatic islets.…”

Current status of pancreatic islet transplantation

“…Additionally, anti-apoptotic peptides, such as caspaseinhibitory peptides [IAP (inhibitor of apoptosis protein) family] [65] and hormones that promote islet growth and function in vivo [EGF (epidermal growth factor), gastrin, GLP-1 (glucagon-like peptide-1) and exendin-4] [66][67][68] are being investigated for their possible role to promote islet engraftment.…”