LncRNA MAARS, macrophage apoptosis, and atherosclerosis

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Atherosclerosis, a chronic arterial disease, involves multiple cellular processes including the accumulation of intimal macrophages. Macrophage apoptosis is increased with progression of atherosclerosis, leading to increased cell death and accumulation of cellular debris. This in turn may abrogate macrophage efferocytosis, an important event for clearance of apoptotic or necrotic cells. Therefore, improving the efficiency of macrophages in the clearance of intra-lesional cellular debris may provide a novel therapeutic approach to limit atherosclerotic progression. Long non-coding RNAs (lncRNAs) have garnered widespread attention as emerging regulators of diverse biological processes relevant to atherosclerosis. However, the identity and roles of specific lncRNAs within atherosclerotic lesions are not well defined. Using RNA-Seq profiling to identify lncRNAs derived specifically from the aortic intima of LDLR-/- mice during lesion progression and regression phases, we identify the lncRNA MAARS (Macrophage-Associated Atherosclerosis lncRNA Sequence). MAARS was the highest expressed lncRNA with a 300-fold increase after lesion progression and decreased by 70% with regression. MAARS is a polyadenylated, macrophage- and nuclear-specific, lncRNA. Kinetic studies showed that MAARS expression is markedly induced in macrophages differentiated from bone marrow, PBMCs, or splenocytes. Our preliminary data demonstrate that systemic delivery of inhibitors to MAARS strongly reduced lesion size, independent of effects on circulating lipid profile, but rather by decreased macrophage apoptosis and increased efferocytosis in the vessel wall. Deficiency of MAARS reduced macrophage apoptosis induced by different stimuli and increased macrophage efferocytosis in vitro. Mechanistically, lncRNA pulldown assays in combination with LC-MS/MS analysis showed that MAARS interacts with HuR, an RNA-binding protein and important regulator of apoptosis. Preliminary studies show that HuR silencing increases macrophage apoptosis and that the MAARS-mediated effects on macrophage apoptosis may be HuR dependent. In addition, MAARS knockdown altered HuR nuclear-cytoplasmic trafficking, and regulated important apoptotic genes. These observations provide the foundation for the central hypothesis that MAARS deficiency, via regulatory effects on HuR and specific macrophage apoptotic signaling pathways, reduces macrophage apoptosis, improves cellular efferocytosis, and suppresses atherosclerosis. To address this further, in Aim1 we examine the role of MAARS in regulating HuR-mediated macrophage apoptosis and efferocytosis; in Aim2, we assess how alterations of MAARS expression affects short- and long-term atherosclerosis in vivo; and in Aim3, we examine the role of the MAARS-HuR signaling axis in human cells and atherosclerotic lesions. Our studies will address a major gap in our understanding of lncRNAs in atherosclerosis and inform how MAARS-mediated control of macrophage apoptosis and efferocytosis may provide new targets for therapy.