OBJECTIVETo establish a new culture method to induce the differentiation of embryonic pancreatic cells into mature endocrine cells.

METHODSMouse embryos at day 12.5 were used and embryonic pancreata were isolated. The isolated embryonic pancreata were cultured on the filter for 7 days, which floated in the dish containing medium. During culture, the expression of pancreas duodenum homeobox-1 (PDX-1), a pancreatic stem cell marker, was examined at day 1. The expression of neurogenin 3 (Ngn3), a pancreatic progenitor cell marker, was examined at day 3. The expressions of endocrine and exocrine markers, insulin, glucagon, and carboxypeptidase (CPA) were examined at day 7 by immunohistochemistry. The kinetics of pancreatic marker expression during culture was assayed by real-time PCR.

RESULTSMany pancreatic stem cells still existed in embryonic pancreata cultured for 1 day; meanwhile, these pancreatic stem cells proliferated in high rate. A large amount of pancreatic progenitor cells were found in embryonic pancreata cultured for 3 days.Pancreatic stem/progenitor cells differentiated into mature endocrine and exocrine cells in embryonic pancreata after having been cultured for 7 days. Furthermore, the expression pattern of pancreatic marker is consistent with that in vivo.

CONCLUSIONWe successfully established a new culture method, with which embryonic pancreatic cells can efficiently differentiate into mature endocrine cell.