Expression of progenitor cell markers during expansion of sorted human pancreatic beta cells.

Functional pancreatic beta cell mass is dynamic and although fully differentiated, beta cells are capable of reentering the cell cycle upon appropriate stimuli. Stimulating regeneration-competent cells in situ is clearly the most desirable way to restore damaged tissue. Regeneration by dedifferentiation and transdifferentiation is a potential source of cells exhibiting a more developmentally immature phenotype and a wide differentiation potential. In this context and to gain a better understanding of the transformation induced in human beta cells during forced in vitro expansion, we focused on identifying differences in gene expression along with phenotypical transformation between proliferating and quiescent human beta cells. FACS-purified beta cells from three different human pancreata were cultured during 3-4 months (8-10 subcultures) on HTB-9 cell matrix with hepatocyte growth factor. Gene expression profiling was performed on cells from each subculture on "in-house" pancreas-specific microarrays consisting of 218 genes and concomitant morphological transformations were studied by immunocytochemistry. Immunocytochemical studies indicated a shift from epithelial to neuroepithelial cell phenotype, including progenitor cell features such as protein gene product 9.5 (PGP 9.5), Reg, vimentin, and neurogenin 3 protein expression. The expression of 49 genes was downregulated, including several markers of endocrine differentiation while 76 were induced by cell expansion including several markers of progenitor cells. Their pattern also argues for the transdifferentiation of beta cells into progenitor cells, demonstrating neuroepithelial features and overexpressing both PBX1, a homeodomain protein that can bind as a heterodimer with PDX1 and could switch the nature of its transcriptional activity, and neurogenin 3, a key factor for the generation of endocrine islet cells. Our study of the machinery that regulates human beta cell expansion and dedifferentiation may help elucidate some of the critical genes that control the formation of adult pancreatic progenitor cells and hence design targets to modify their expression in view of the production of insulin-secreting cells.