MicroRNAs and histone deacetylase inhibition-mediated protection against inflammatory β-cell damage.

Anna Lindeløv Vestergaard,Christine Bruun,Jonathan Lucien Stahl,Peter De Hemmer Horskjær,Frida Greek Kofod,Farzaneh Taheri Sotudeh,Romano Regazzi,Claus Heiner Bang-Berthelsen,Flemming Pociot,Lukas Adrian Berchtold,Lisa Christen,Klaus Stensgaard Frederiksen,Thomas Mandrup-Poulsen,Joachim Størling,Tina Fløyel,Simranjeet Kaur,Bernard Mari

doi:10.1371/journal.pone.0203713

Abstract

Inflammatory β-cell failure contributes to type 1 and type 2 diabetes pathogenesis. Pro-inflammatory cytokines cause β-cell dysfunction and apoptosis, and lysine deacetylase inhibitors (KDACi) prevent β-cell failure in vitro and in vivo, in part by reducing NF-κB transcriptional activity. We investigated the hypothesis that the protective effect of KDACi involves transcriptional regulation of microRNAs (miRs), potential new targets in diabetes treatment. Insulin-producing INS1 cells were cultured with or without the broad-spectrum KDACi Givinostat, prior to exposure to the pro-inflammatory cytokines IL-1β and IFN-γ for 6 h or 24 h, and miR expression was profiled with miR array. Thirteen miRs (miR-7a-2-3p, miR-29c-3p, miR-96-5p, miR-101a-3p, miR-140-5p, miR-146a-5p, miR-146b-5p, miR-340-5p, miR-384-5p, miR-455-5p, miR-466b-2-3p, miR-652-5p, and miR-3584-5p) were regulated by both cytokines and Givinostat, and nine were examined by qRT-PCR. miR-146a-5p was strongly regulated by cytokines and KDACi and was analyzed further. miR-146a-5p expression was induced by cytokines in rat and human islets. Cytokine-induced miR-146a-5p expression was specific for INS1 and β-TC3 cells, whereas α-TC1 cells exhibited a higher basal expression. Transfection of INS1 cells with miR-146a-5p reduced cytokine signaling, including the activity of NF-κB and iNOS promoters, as well as NO production and protein levels of iNOS and its own direct targets TNF receptor associated factor 6 (TRAF6) and interleukin-1 receptor-associated kinase 1 (IRAK1). miR-146a-5p was elevated in the pancreas of diabetes-prone BB-DP rats at diabetes onset, suggesting that miR-146a-5p could play a role in type 1 diabetes development. The miR array of cytokine-exposed INS1 cells rescued by KDACi revealed several other miRs potentially involved in cytokine-induced β-cell apoptosis, demonstrating the strength of this approach.

Highlights

Reduction of functional β-cell mass is a feature of both type 1 and type 2 diabetes (T1D and T2D, respectively), and inflammatory mechanisms including pro-inflammatory cytokines have been implicated as mediators of β-cell apoptosis in both disorders [1,2,3]
Given the higher efficacy of Givinostat in lowering Nitric oxide (NO) production, this KDACi was chosen for the miR array study
INS1 cells were exposed to Givinostat or no treatment for 1 h and to media with or without 150 pg/ml IL1β and 0.1 ng/ml IFN-γ for 6 h or 24 h

Summary

Introduction

Reduction of functional β-cell mass is a feature of both type 1 and type 2 diabetes (T1D and T2D, respectively), and inflammatory mechanisms including pro-inflammatory cytokines have been implicated as mediators of β-cell apoptosis in both disorders [1,2,3]. Cytokine-induced β-cell apoptosis requires active gene expression and protein translation [11]. We recently discovered that oral inhibitors of lysine deacetylases (KDACs), proven to be effective and safe in other inflammatory disorders such as systemic onset juvenile idiopathic arthritis [12] and graft-versus-host disease [13], prevent cytokine-induced β-cell apoptosis [14,15,16,17,18,19]. An additional mechanism could be hyperacetylation of histones upregulating expression of anti-apoptotic microRNAs (miRs). These in turn could act by e.g. repressing the translation of proteins that promote βcell death via activation of the intrinsic (mitochondrial) death pathway

Methods

Results

Conclusion