Cigarette smoke exposure impairs β-cell function through activation of oxidative stress and ceramide accumulation

Objectives Epidemiological studies indicate that first- and second-hand cigarette smoke (CS) exposure are important risk factors for the development of type 2 diabetes (T2D). Additionally, elevated diabetes risk has been reported to occur within a short period of time after smoking cessation, and health risks associated with smoking are increased when combined with obesity. At present, the mechanisms underlying these associations remain incompletely understood. The objective of this study was to test the impact of CS exposure on pancreatic β-cell function using rodent and in vitro models. Methods Beginning at 8 weeks of age, C57BL/6 J mice were concurrently fed a high-fat diet (HFD) and exposed to CS for 11 weeks, followed by an additional 11 weeks of smoking cessation with continued HFD. Glucose tolerance testing was performed during CS exposure and during the cessation period. Cultured INS-1 β-cells and primary islets were exposed ex vivo to CS extract (CSE), and β-cell function and viability were tested. Since CS increases ceramide accumulation in the lung and these bioactive sphingolipids have been implicated in pancreatic β-cell dysfunction in diabetes, islet and β-cell sphingolipid levels were measured in islets from CS-exposed mice and in CSE-treated islets and INS-1 cells using liquid chromatography-tandem mass spectrometry. Results Compared to HFD-fed, ambient air-exposed mice, HFD-fed and CS-exposed mice had reduced weight gain and better glucose tolerance during the active smoking period. Following smoking cessation, CS-mice exhibited rapid weight gain and had accelerated worsening of their glucose tolerance. CS-exposed mice had higher serum proinsulin/insulin ratios, indicative of β-cell dysfunction, significantly lower β-cell mass (p = 0.017), reduced β-cell proliferation (p = 0.006), and increased islet ceramide content compared to non-smoking control mice. Ex vivo exposure of isolated islets to CSE was sufficient to increase islet ceramide levels, which was correlated with reduced insulin gene expression and glucose-stimulated insulin secretion, and increased β-cell oxidative and endoplasmic reticulum (ER) stress. Treatment with the antioxidant N-acetylcysteine markedly attenuated the effects of CSE on ceramide levels, restored β-cell function and survival, and increased cyclin D2 expression, while also reducing activation of β-cell ER and oxidative stress. Conclusions Our results indicate that CS exposure leads to impaired insulin production, processing, secretion and reduced β-cell viability and proliferation. These effects were linked to increased β-cell oxidative and ER stress and ceramide accumulation. Mice fed HFD continued to experience detrimental effects of CS exposure even during smoking cessation. Elucidation of the mechanisms by which CS exposure impairs β-cell function in synergy with obesity will help design therapeutic and preventive interventions for both active and former smokers

A Role for MicroRNA-146a-5p Mediated Regulation of Stromal Interaction Molecule 1 and Store Operated Calcium Entry in the Pancreatic Beta-Cell in Response to Cytokine Mediated Stress

Indiana University-Purdue University Indianapolis (IUPUI)Store-operated Ca2+ entry (SOCE) is involved in the maintenance of endoplasmic reticulum (ER) Ca2+ levels. The SOCE involves Stromal Interaction Molecule 1 (STIM1), distributed throughout the ER, and Orai1 channels, dispersed on the plasma membrane. SOCE is activated by the depletion of ER Ca2+ causing STIM1 to induce ER expansion and recruits Orai1 channels thus replenishing ER Ca2+. We reported downregulation of STIM1 in human islets from donors with type 2 diabetes (T2D) and in INS-1 β-cells treated with cytokines, and loss of STIM1 expression impairs β-cell SOCE, ER stress, and reduced insulin secretion. However, the regulatory mechanisms of STIM1 downregulation are unknown. To test this, actinomycin D and cycloheximide chase assay was performed to define whether IL-1β treatment impacted STIM1 mRNA or protein half-life. IL-1β had no impact on mRNA or protein decay. MicroRNAs (miRNAs), a class of small non-coding RNAs can regulate gene expression post-transcriptionally by binding to complementary regions in the 3’ untranslated region (UTR) of target mRNAs, affecting mRNA stability and translatability. The objective of this study was to establish miRNA regulation of STIM1 expression and altered SOCE. To identify potential miRNA candidates, RNA sequencing was done in human islets, treated with IL-1β and IFN-γ for 24 hrs. A total of 20 miRNAs were differentially expressed using a FC value of ≥ 1.5 and a p value of < 0.05. Of these, two miRNAs (miR-146a-5p and miR-4640-5p) were predicted by TargetScan to bind the 3’UTR of STIM1.To validate these findings, INS-1 β-cells, and human islets were treated with or without IL-1β. Only miR-146a-5p was upregulated in both systems. Consistent with inverse correlation, INS-1 β-cells transfected with miR-146a-5p mimic showed reduced STIM1 expression. To test whether miR-146a-5p inhibition preserves STIM1 expression, INS1 cells were treated with miR-146a-5p inhibitor along with IL-1β and inhibition of miR-146a-5p led to partial preservation of STIM1 expression. Future studies will test the effect of miR-146a-5p mimics and inhibitors on SOCE. The results indicate that the stress induced by IL-1β leads to induction of miR-146a-5p, which may then target STIM1 mRNA. Such studies could enable broader implementation of miRNA in βcell dysfunction

The Contribution of Transcriptional Coregulators in the Maintenance of β-cell Function and Identity

Islet β-cell dysfunction that leads to impaired insulin secretion is a principal source of pathology of diabetes. In type 2 diabetes, this breakdown in β-cell health is associated with compromised islet-enriched transcription factor (TF) activity that disrupts gene expression programs essential for cell function and identity. TF activity is modulated by recruited coregulators that govern activation and/or repression of target gene expression, thereby providing a supporting layer of control. To date, more than 350 coregulators have been discovered that coordinate nucleosome rearrangements, modify histones, and physically bridge general transcriptional machinery to recruited TFs; however, relatively few have been attributed to β-cell function. Here, we will describe recent findings on those coregulators with direct roles in maintaining islet β-cell health and identity and discuss how disruption of coregulator activity is associated with diabetes pathogenesis