In vitro generation of insulin-producing beta cells from adult exocrine pancreatic cells

Abstract

Aims/hypothesis: Transplantation of insulin-producing beta cells from donors can cure diabetes, but they are available in insufficient quantities. In this study, we investigated the possibility of generating insulin-producing cells from adult rat exocrine cells cultured in the presence of growth factors.

Methods: Rat exocrine pancreatic cells were isolated and treated in vitro with epidermal growth factor (EGF) and leukaemia inhibitory factor (LIF). Analysis was performed by immunocytochemistry, DNA measurement and radioimmunoassay. Cells were transplanted to alloxan-treated (70 mg/kg) nude mice and glycaemia was monitored for 21 days. Nephrectomy was performed on day 15.

Results: In a 3-day culture period, addition of LIF plus EGF to the medium resulted in an 11-fold increase of the beta cell mass. This could not be attributed to the very low mitotic activity of contaminating beta cells. Furthermore, when contaminating beta cells were initially destroyed with alloxan, this effect was even more pronounced. The newly formed cells secreted insulin in response to glucose and were immunoreactive for C-peptide-I, Pdx-1 and GLUT-2, which are characteristics of mature beta cells. Electron microscopy showed that they also contained insulin-immunoreactive secretory granules. Some insulin-positive cells were immunoreactive for amylase and cytokeratin-20, or were binucleated, which are characteristics of exocrine cells. The cells were able to restore normoglycaemia when transplanted to alloxan-diabetic mice, and hyperglycaemia recurred upon removal of the graft.

Conclusions/interpretation: Our study shows that functional beta cells can be generated from exocrine tissue by transdifferentiation and thereby may offer a new perspective for beta cell therapy.