Abstract 13914: Low-Density Lipoprotein Delivery of Microbial Small RNAs Drives Atherosclerosis Through Macrophage Toll-Like Receptor 8

Low-density lipoproteins (LDL) are a causal factor for atherosclerosis, a key pathological process in cardiovascular disease. Within atherosclerotic lesions, macrophages present a spectrum of phenotypes that mediate both disease pathogenesis and resolution. Inflammatory macrophage phenotypes are pro-atherogenic, but the natural factors that promote polarization remain to be identified. Here, we demonstrate that microbial small RNAs (msRNA) are enriched on LDL and drive pro-inflammatory macrophage polarization and cytokine secretion via activation of toll-like receptor 8 (TLR8). Removal of msRNA cargo during LDL reconstitution yields particles that are taken up by macrophages but fail to stimulate inflammatory activation. Using loss-of-function and gain-of-function studies, we demonstrate that TLR8 is both necessary and sufficient for LDL-induced activation of NF-kappa-B, a central regulator of inflammatory macrophage polarization. TLR8 recognition of msRNA was found to be antagonized by non-targeting locked-nucleic acids (nt-LNA). Pre-treatment of macrophages with a nt-LNA was found to prevent LDL-induced macrophage polarization in vitro , and re-organize lesion macrophage phenotypes in vivo , as determined by single-cell RNA sequencing. Critically, we demonstrate that nt-LNA treatment reduced plaque burden in 4-week and 10-week atherosclerosis progression studies in Apoe -/- mice fed a western diet. Moreover, in a regression model using male and female Ldlr -/- mice with advanced lesions, nt-LNA treatment in combination with dietary intervention promoted lesion regression relative to mice receiving only dietary intervention. These results identify LDL-msRNA as stimuli for atherosclerosis-associated inflammation and support alternative functions of LDL beyond cholesterol transport.