Abstract 448: Stress Granule Formation as a Molecular Indicator of Atherosclerotic and Vascular Disease Progression

Abstract

The molecular mechanisms by which atherogenic stress contributes to the pathophysiology of disease remain to be fully characterized. RNA-binding proteins (RBPs) regulate vascular cell (VSMCs and ECs) response to inflammatory stimuli by fine-tuning mRNA stability, and protein abundance. RBPs such as HuR and FXR1 regulate mRNA transcript abundance by binding AU-rich elements (AREs) in the 3’UTR permitting the transcript stabilization or destabilization, respectively. Most inflammatory transcripts including TNFα contain conserved AREs in their 3’UTR, and inflammatory stimuli activate RBPs to alter the landscape of inflammatory mediators. This post-transcriptional regulation of inflammatory transcripts by RBPs islocalized to structures called stress granules (SGs), which are mRNA-protein assemblies formed from non-translating mRNAs, ribosomal subunits, among other components. The manner by which these complexes form between mRNA and RBPs (mRNPs) under inflammatory or pathological conditions remains unknown, particularly with respect to vascular inflammatory disease. We hypothesize that the presence and quantity of stress granules are a critical cellular response to atherogenic stimuli. We identified SGs in cultured VSMCs and ECs treated with clotrimazole, a known inducer of SGs by co-staining for PABP, a SG marker, along with RBPs HuR and FXR1, both known to co-localize to SGs. VSMC treated with clotrimazole had increased eIF2α phosphorylation. LDLR-/- mice fed high fat diet for 14 weeks also demonstrated significantly increased PABP expression in the smooth muscle and endothelial cells of the aortic arch as compared to WT control. Atherogenic and hypoxic stimuli induced the formation of SGs in cultured primary human VSMCs and ECs. Interestingly, we found VSMCs pre-treated with anti-inflammatory cytokine IL-19 followed by clotrimazole significantly reduced the formation of SGs per cell in a time dependent manner (n=50, p<0.05) as well as reduction in PABP expression, suggesting anti-inflammatory treatment effects SG formation. Taken together, these results have led to the hypothesis that SG formation in atherosclerosis is driven by inflammatory stimuli, and that anti-inflammatory treatment may lessen atherosclerosis progression and plaque formation by reduction of SGs.