Abstract 624: MiR-33 Antagonism Prevents the Progression of Atherosclerosis by Promoting an M2 Macrophage Phenotype

Abstract

MicroRNA-33 (miR-33) plays a central role in regulating HDL biogenesis and reverse cholesterol transport (RCT) via post-transcriptional repression of cholesterol efflux genes (ABCA1, ABCG1, NPC1). Delivery of miR-33 inhibitors to mice and non-human primates fed a chow diet raises plasma HDL cholesterol (HDL-C) and increases RCT, suggesting miR-33 as a promising therapeutic target for the treatment of atherosclerosis. However, recent studies in mice have shown that the HDL-raising effects of anti-miR33 are blunted during Western diet feeding, and its efficacy in reducing atherosclerosis progression under these conditions remains controversial. To further test the effects of miR-33 inhibition during atherogenesis, we treated Ldlr −/− mice with control or anti-miR33 oligonucleotides for 8 weeks coincident with Western diet feeding. Morphometric analyses revealed that anti-miR33 reduced atherosclerosis progression by 40% in both the aortic root and the aorta en face without altering plasma total or HDL cholesterol. Because anti-miR33 accumulates in plaque macrophages where it may directly alter gene expression, we performed Nanostring profiling of plaque macrophages isolated by laser capture microdissection. We found a marked increase in markers of alternatively activated M2 macrophages (Arg1, Fizz1, Cd206, Ym1) in anti-miR33-treated mice compared to controls. Moreover, in vitro transfection of peritoneal macrophages with anti-miR33 reduced the expression of M1 markers (IL-1, TNFα) and increased the expression of M2 markers (Arg1, IL-10, Fizz1), suggesting that miR-33 directly controls the inflammatory polarization of macrophages. This regulation of macrophage polarization by miR-33 was found to be independent of its effects on ABCA1, but dependent on the targeting of AMP Kinase, a key integrator of cellular energy homeostasis. Notably, inhibitors of AMPK blocked the anti-inflammatory effects of miR-33 inhibition in vitro and in vivo . Collectively, these results identify a novel role for miR-33 in the regulation of macrophage inflammation and show that antagonism of miR-33 prevents atherogenesis, in part, by reducing plaque inflammation through the polarization of macrophages from a pro-inflammatory M1 to a reparative M2 state.