Abstract
In-vitro expansion of β cells from adult human pancreatic islets could provide abundant cells for cell replacement therapy of diabetes. However, proliferation of β-cell-derived (BCD) cells is associated with dedifferentiation. Here we analyzed changes in microRNAs (miRNAs) during BCD cell dedifferentiation and identified miR-375 as one of the miRNAs greatly downregulated. We hypothesized that restoration of miR-375 expression in expanded BCD cells may contribute to their redifferentiation. Our findings demonstrate that overexpression of miR-375 alone leads to activation of β-cell gene expression, reduced cell proliferation, and a switch from N-cadherin to E-cadherin expression, which characterizes mesenchymal-epithelial transition. These effects, which are reproducible in cells derived from multiple human donors, are likely mediated by repression of PDPK1 transcripts and indirect downregulation of GSK3 activity. These findings support an important role of miR-375 in regulation of human β-cell phenotype, and suggest that miR-375 upregulation may facilitate the generation of functional insulin-producing cells following ex-vivo expansion of human islet cells.
Highlights
Beta-cell replacement by regeneration or transplantation is considered a promising therapy for diabetes
As with miR-375 overexpression, SB-216763 did not increase total β-catenin protein levels (Fig 6D and 6E), and resulted in growth arrest (Fig 6F). These findings suggest that GSK3 inhibition at least partially mediates the effect of miR-375 overexpression on BCD cell redifferentiation
Our findings demonstrate that restoration of normal levels of a single miRNA, miR-375, in BCD cells is sufficient for induction of β-cell gene expression, reduced cell proliferation, and a switch from NCAD to ECAD expression, which is characteristic of mesenchymal-epithelial transition
Summary
Beta-cell replacement by regeneration or transplantation is considered a promising therapy for diabetes. Expanded human β-cell-derived (BCD) cells, which constitute ~40% of cells in islet cell cultures [2], maintain open chromatin structure at β-cell genes [4], and can be redifferentiated in response to a combination of soluble factors termed Redifferentiation Cocktail (RC) [5]. These factors include activin A, exendin-4, nicotinamide, and high glucose concentrations, which have been shown to promote β-cell differentiation, in serum-free medium supplemented with B27 and insulin-transferrin-selenium. RC treatment leads to redifferentiation of only part of BCD cells. In search for improved redifferentiation approaches, we analyzed changes in PLOS ONE | DOI:10.1371/journal.pone.0122108 April 13, 2015
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