Establishment and identification of pancreatic stem cell strain derived from islets of Kunming mice under feeder layer conditions / 中国组织工程研究

ABSTRACT

BACKGROUND:Until now, little has been reported on establishment of pancreatic stem cell strains and lines,and the purification of pancreatic stem cells is difficult since the cell line establish rate is low.OBJECTIVE:To explore a more rational and effective technique of in vitro separation and continuous passage of pancreatic stem cells, with the hope to establish cell strains and even cell lines and to lay the foundation for the follow-up study of pancreatic stem cells in the treatment of diabetes.METHODS:Firstly, Percoll discontinuous density gradient centrifugation method was applied to separate the mouse pancreatic endocrine portion from the exocrine portion, then to obtain cell strains with highly proliferative ability and low differentiation from pancreatic endocrine portion-the islet. We used mouse embryonic fibroblasts treated with mitomycin C as a feeder layer, for in vitro continuous culture of islet-derived pancreatic stem cells under feeder layer conditions until they were transferred to the 30th passage to establish cell lines. Then pancreatic stem cell line derived from pancreatic islet was detected and identified by a series of tests including growth characteristic test, morphological observation, related molecular marker identification and differentiation characteristic identification.RESULTS AND CONCLUSION:In the continuous process of passage, pancreatic stem cells showed active proliferative ability, and maintained the typical morphological characteristics of stem cells and expression of pancreatic stem cell marker-Nestin. After induction, pancreatic stem cells showed insulin gene expression,reflecting their differentiation potential. Therefore, under the condition of feeder layer, the pancreatic stem cell line derived from Kunming mice was successfully established and the related identification was completed,which lays the foundation for the following research.