Aim: One of the hallmarks of type 2 diabetes (T2D) is impaired beta cell function, which develops in part as a result of widespread cellular de-differentiation. The current state-of-the-art defines beta cells as a heterogeneous islet population, with the existence of individual subpopulations including specialised pacemakers. We sought here to understand how overexpression-induced beta cell maturity affects such heterogeneity, before examining the influence of this on islet function and insulin secretion.
Materials and methods: An adenoviral polycistronic construct for Ngn3, MafA, Pdx1 and mCherry (Ad3-NPM) was used for inducing overexpression in adult mouse islets. PATagRFP- or non-transduced (CT) islets served as controls. Gene expression was quantified by qRT-PCR. Pdx1 and insulin content was analysed by immunohistochemistry using a Zeiss LSM780 confocal microscope. Nipkow spinning disk microscopy was used for quantifying Ca++, ATP and cAMP dynamics in live islets. Glucose-stimulated (GSIS) and incretin-stimulated insulin secretion (ISIS) were assessed by HTRF assay.
Results: Ad3-NPM treatment increased Pdx1 and MafA expression in mouse islets, while Ngn3 levels remained unchanged. Immunohistochemistry showed that Pdx1 overexpression preferentially occurred in immature beta cells, inducing cellular homogeneity. Ca++ levels showed a marked decrease (ΔF= 0.81 in CT vs 0.44 AU in Ad3-NPM-islets; P<0.01), accompanied by reduced beta cell-beta cell coordination and a reduced number of beta cell pacemakers, i.e. hubs (12.6 vs 5.6% hubs, CT vs Ad3-NPM; P<0.05). The ATP/ADP ratio was slightly higher in Ad3-NPM-transduced islets, although cAMP responses to glucose were sharply reduced. Basal insulin release was increased following overexpression, with impairments in both GSIS and ISIS (7.5-fold vs 5-fold after glucose stimulation and 98-fold vs 50-fold after exendin-4 stimulation; CT vs Ad3-NPM-islets).
Summary: Induced beta cell maturity leads to islet failure and lowered insulin secretory capacity. This work underlines the importance of maintaining subtle differences in beta cell maturity for normal islet function.