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题目:: An efficient experimental strategy for mouse embryonic stem cell differentiation and separation of a cytokeratin-19-positive population of insulin-producing cells.
作者:: Naujok(O),Francini(F),Jörns(A),Lenzen(S)
状态:: 发布时间2008-08-14 , 更新时间 2011-11-17
期刊:: Cell Prolif
摘要:: Embryonic stem cells are a potential source for insulin-producing cells, but existing differentiation protocols are of limited efficiency. Here, the aim has been to develop a new one, which drives development of embryonic stem cells towards insulin-producing cells rather than to neuronal cell types, and to combine this with a strategy for their separation from insulin-negative cells.,The cytokeratin-19 (CK19) promoter was used to control the expression of enhanced yellow fluorescence protein in mouse embryonic stem cells during their differentiation towards insulin-producing cells, using a new optimized four-stage protocol. Two cell populations, CK19(+) and CK19(-) cells, were successfully fluorescence sorted and analysed.,The new method reduced neuronal progeny and suppressed differentiation into glucagon- and somatostatin-producing cells. Concomitantly, beta-cell like characteristics of insulin-producing cells were strengthened, as documented by high gene expression of the Glut2 glucose transporter and the transcription factor Pdx1. This novel protocol was combined with a cell-sorting technique. Through the combined procedure, a fraction of glucose-responsive insulin-secreting CK19(+) cells was obtained with 40-fold higher insulin gene expression and 50-fold higher insulin content than CK19(-) cells. CK19(+) cells were immunoreactive for C-peptide and had ultrastructural characteristics of an insulin-secretory cell.,Differentiated CK19(+) cells reflect an endocrine precursor cell type of ductal origin, potentially suitable for insulin replacement therapy in diabetes.
语言:: eng
DOI:: 10.1111/j.1365-2184.2008.00541.x

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