MicroRNAs are small non-coding RNAs with ~22 nucleotides in size involved in the modulation of gene expression. Previously, we observed that miR-146a-5p is upregulated during inflammatory stress in pancreatic islets and pancreatic tissue sections from T1D donors. Although others have linked overexpression of miR-146a-5p to β cell death and impaired insulin secretion, the molecular mechanisms mediating these effects remain unclear. To further understand miR-146a-5p role in β cell health and disease, we have generated stable cell lines that are either overexpressed (OE) or inhibited (ZIP) for miR-146a-5p and assessed for β cell function under inflammatory (IL-1β, IFN-γ, and TNF-α) conditions to model T1D in vitro. Treatment of OE cells with or without pro-inflammatory cytokines increased expression of cleaved caspase-3 and Annexin-V-positive cell death and reduced insulin secretion in both untreated and cytokine treatment. However, ZIP cells are protected from inflammatory stress showing decreased apoptosis and improved insulin secretion. Furthermore, ZIP cells were found to have increased mitochondrial DNA copy numbers, mitochondrial membrane potential, and basal and maximal respiration rate. Notably, cytokines-treated OE cells showed reduced basal respiration rate and ATP production compared to cytokines-treated ZIP cells. In addition, mitochondria isolated from MIN6 cells showed enriched miR-146a-5p under cytokine treatment, suggesting inflammation-mediated enrichment of miR-146a-5p. Finally, RNA sequencing analysis of OE and ZIP cells identified the RNA transcripts regulated by miR-146a-5p, including MAPT, MYRIP, NRG1, and GPX1; pathway enrichment analysis showed pathways related to insulin secretion, apoptosis, and mitochondrial function. Taken together, findings from our study show for the first time that miR-146a-5p upregulation promotes β cell dysfunction and death by suppressing mitochondrial function during T1D pathogenesis.
P.Krishnan: None. R.C.S.Branco: None. S.A.Weaver: Employee; Eli Lilly and Company. C.Lee: None. F.Syed: None. C.Evans-molina: Advisory Panel; Provention Bio, Inc., DiogenX, Avotres Inc., Neurodon, MaiCell Therapeutics, Other Relationship; Isla Technology, Bristol-Myers Squibb Company, Nimbus Therapeutics, Research Support; Lilly, Astellas Pharma Inc.
JDRF (2-SRA-2019-834-S-B, 2-SRA-2018-493-A-B); National Institutes of Health (DK127308, U01DK127786, UC4DK104166)