Mechanisms of Rapamycin Toxicity in Pancreatic Beta Cells
thesisposted on 2011-06-16, 08:41 authored by Adam Barlow
Islet transplantation offers a potential cure for type I diabetes mellitus. The publication of the landmark Edmonton study in 2000, which reported insulin independence in seven consecutive patients, prompted increased interest in this therapy. However, enthusiasm was tempered when the 5 year follow up results of this study were published, with only 10% of recipients maintaining insulin independence. The cause for this graft loss is multifactorial, but there is in vivo and in vitro evidence that suggests immunosuppressive drug toxicity plays an important role. The aim of this thesis was to establish the effects of rapamycin, one of the primary immunosuppressants used in islet transplantation, on murine β cells and islets and elucidate the mechanisms of any toxicity seen. The intracellular target for rapamycin is mTOR which exists in two complexes, mTORC1 and mTORC2. mTORC1 primarily regulates cell size and proliferation; whereas mTORC2 plays a key role in regulating cell survival via protein kinase B (PKB). This thesis has demonstrated that rapamycin treatment results in significant reductions in glucose stimulated insulin secretion in the MIN6 mouse insulinoma cell line and isolated rat islets, as well as increased apoptosis in these cell types. Furthermore, it has shown that prolonged rapamycin results in inhibition of mTORC2 assembly, with resultant inhibition of PKB phosphorylation and activity. Overcoming rapamycin induced mTORC2 inhibition with an adenovirus encoding constitutively active PKB ameliorates the detrimental effects of rapamycin on both MIN6 cells and rat islets. This suggests that rapamycin toxicity is mediated predominantly via mTORC2 rather than mTORC1 inhibition. This work brings into question the use of rapamycin as an immunosuppressant in islet transplantation and also highlights the key role of PKB in β cell survival. Therapies resulting in PKB activation may have the potential to improve outcomes of islet transplantation.