Adeno-associated virus vector mediated gene transfer to pancreatic beta cells

Insulin-dependent diabetes mellitus (IDDM) or type 1 diabetes is an autoimm une disease that results in destruction of the insulin-producing pancreatic islet beta cells. Several factors induce the invasion of immune cells into islets and trigger inflammation. Gene therapy approaches targeting the isl et cells could be an effective treatment to prevent the onset or reverse ty pe I diabetes. Allogeneic islet transplantation provides short-term treatme nt. However, genetically modified islets, which resist the host immune resp onse, could provide long-term solutions. Adeno-associated virus (AAV) is em erging as a prominent vector system for delivering therapeutic genes for hu man gene therapy. AAV vector can transduce nondividing cells and provide lo ng-term gene expression by integrating into host chromosome. Therefore, it is an appropriate vector system for islet cell gene therapy. To test the ef ficacy of AAV vector to transduce pancreatic endocrine cells, we constructe d AAV vectors using plasmid pSub201. Wild-type AAV DNA analogue from plasmi d psub201 was subcloned into a cloning plasmid pSP72 and AAV vectors were c onstructed by inserting the transgenes with heterologous promoter in place of AAV open reading frames (rep and cap). In this report we demonstrate the transduction of pancreatic islet cells with AAV vectors encoding bacterial beta-galactosidase enzyme or enhanced green fluorescent protein (EGFP) as reporter gene. Dispersed porcine and rat islet cells can be transduced by A AV vector, with an efficiency of 47% and 38%, respectively In particular po rcine islet insulin producing beta cells were transduced with an efficiency of 39%. Intact rat islet cells were transduced with an efficiency of 26% a s estimated by FAGS analysis following transduction with an AAV vector enco ding EGFP. Transduction of intact mt islets with an AAV vector did not alte r glucose-induced insulin secretion. AAV vector transduction was higher in transformed islet cell lines INS-1 and RIN m5F with an efficiency of 65% an d 57%, respectively. These new results suggest that AAV vectors will provid e an improved method of gene delivery to pancreatic islets and isolated pan creatic beta cells.