关键词: 胰腺, 胰

Abstract: Objective To establish the method of isolation,culture, and differentiation of Nestin positive cells of non-islet cells in adult rats in vitro. Methods The whole pancreas of three adult rats were digested in vitro with the intraductal collagenase distention method, then both islets and non-islets were collected and cultured with 10% fetal bovine serum (pH 7.6) and serum-free RPMI 1640 (pH 7.4).The formation process of new islet-like cell masses were examined by light microscopy,and the glucose-stimulated insulin release　(SI) from 40 just isolated islets and 40 new produced islet-like cell masses were tested.The expression of Nestin was tested by immunocytochemistry. Results Functioning islets in good morphology could be obtained from pancreas of adult rats by the intraductal collagenase distention method；there were adherent cells after 36 h culture,but no islets; the Nestin positive cells proliferated fast and formed new islet-like cell masses after 18-22 d. Nestin expression could be confirmed among adherent cells，but no insulin and glucagon, and Nestin positive cells can be seen in passage cells of new produced islet-like cell masses. Insulin secretion levels in 40 new isolated islets were (63.6±4.0)，(202.2±14.8)mIU•L^-1 in 3.3 and 16.7 mmol•L^-1 glucose, respectively;and (3.5±0.2)，(3.3±0.2)mIU•L^-1,respectively, in 40 new produced islet-like cell masses (P<0.001). Conclusion Functioning islets can be obtained from the pancreas of adult rats. Nestin positive cells of non-islet cells can be obtained from pancreas of adult rats in pH 7.6 condition through culture in vitro,and can form islet-like cell masses;Nestin positive cells of non-islet cells in pancreas of adult rats possess the characters of islet stem cells.

Key words: pancreas, islets of langerhans, cells, cultured