Abstract Body: Introduction: Atherosclerotic cardiovascular disease remains the leading cause of global mortality. Although atherosclerosis develops gradually over decades, acute viral infections can abruptly heighten vascular inflammation and trigger myocardial infarction or stroke. The IL-1β pathway is a central contributor to cardiovascular risk, yet the upstream drivers that connect viral infection to inflammasome activation are not well defined. Viral infections robustly induce transcription of retrotransposons, particularly short interspersed nuclear elements (SINEs) such as Alu RNAs, yet their contribution to vascular inflammation has not been explored.

Hypothesis: Infection-induced Alu RNAs are critical drivers of vascular inflammation, and their targeted inhibition represents a promising therapeutic strategy to prevent infection-associated cardiovascular events.

Methods: Primary human coronary artery endothelial cells (HCAECs) were transfected with either control siRNA or an Alu siRNA designed to bind consensus Alu repeat regions. After a 24-hour transfection period, cells were infected with HSV-1 (MOI 5). Twenty hours post-infection, inflammatory gene expression (IL1B, IL6, and CASP1) was quantified by RT-qPCR (n=3) and analyzed using unpaired two-tailed t-tests. Protein-level assessments included pro–IL-1β by western blot and active IL-1β by ELISA (n=3). To verify Alu RNA target engagement, AGO2 cross-linking immunoprecipitation (CLIP) followed by Alu-specific RT-PCR.

Results: HSV-1 infection of HCAECs induced a dose-dependent increase in Alu RNA expression (MOI 1, 5, and 10 vs. MOI 0) and upregulated IL1B, IL6, and CASP1. Compared with control siRNA, Alu siRNA significantly reduced HSV-1–induced expression of IL1B, IL6, and CASP1 (p<0.05) in primary HCAECs. Protein analyses showed parallel decreases in HSV-1–induced pro–IL-1β by western blot and active IL-1β by ELISA. AGO2 CLIP combined with Alu-specific RT-PCR demonstrated enrichment of Alu RNA in cells treated with Alu-siRNA and infected with HSV-1 compared to cells treated with control siRNA and infected with HSV-1, confirming direct targeting of endogenous Alu sequences.

Conclusion: These findings identify Alu RNA as a previously unrecognized, modifiable driver of virus-induced inflammatory signaling in HCAECs. Targeted silencing of Alu transcripts suppresses inflammasome-related cytokine activation, highlighting Alu-directed therapy as a novel strategy to mitigate infection-associated cardiovascular risk.