Abstract 4361211: The RNA-binding protein HuR controls atherosclerotic disease progression of carotid and femoral arteries

Introduction: Peripheral artery disease (PAD) is an escalating health burden with rising incidence in aging populations driven by its strong association with cardiovascular morbidity and mortality. The RNA-binding protein HuR may represent a therapeutic target by regulating thrombosis and vascular inflammation in PAD. Methods: HuR expression was assessed by single-cell RNA sequencing and in peripheral blood mononuclear cells from 977 deeply phenotyped individuals with vascular disease. Carotid and femoral artery ultrasound was used to assess intima-media thickness, maximum wall thickness, and plaque count. Participants were prospectively followed for major adverse cardiovascular events (MACE). Mechanistically, the role of endothelial HuR was investigated through silencing and overexpression, RNA stability assays, and pharmacological inhibition. The HuR-mRNA interactome was defined by iCLIP-seq, RIP-seq, and mRNA-seq. Results: Single-cell analysis revealed elevated HuR expression in vascular endothelium and infiltrated immune cells from Ldlr -/- mice on high-fat diet vs. chow and human carotid plaques. In humans, elevated HuR expression was independently associated with coronary artery disease (OR=2.67, highest vs. lower tertiles), increased C-reactive protein, and greater mean carotid IMT, after adjustment for traditional risk factors (P<0.05). In healthy individuals, baseline HuR levels predicted accelerated subclinical atherosclerosis progression, reflected by increased carotid and femoral plaque number (P<0.001) and higher maxWT increase (P for interaction=0.045). High HuR levels also correlated with greater MACE incidence over a median 48-month follow-up (log-rank P=0.009). Mechanistically, HuR iCLIP-seq, RIP-seq, and transcriptomics in endothelial cells under basal and TNF-α-stimulated conditions revealed multiple HuR binding sites in AUUUA- and UUUU(G/U)-rich 3′UTRs of pro-inflammatory and pro-thrombotic transcripts regulating adhesion, TGFBR signaling, migration, and differentiation. Pharmacological inhibition or siRNA-mediated HuR silencing suppressed endothelial inflammatory gene expression, whereas HuR overexpression alone induced a pro-inflammatory phenotype via stabilization of its mRNA targetome. Expression of HuR target mRNAs strongly correlated with HuR levels in vascular diseases. Conclusion: HuR inhibition poses as a promising therapeutic target in peripheral artery disease by controling the RNA stability of several pro-atherosclerotic genes.