Abstract Body (Do not enter title and authors here): Background: Thoracic aortic aneurysm (TAA) is an asymptomatic, life-threatening disease with high mortality greater than 80% after rupture. The assembly of cytoskeletal structural proteins, e.g. Filamin A (FLNA) with extracellular matrix (ECM), which helps in maintaining aortic structural integrity and function, is highly disrupted in TAA. Besides surgical interventions, no medical therapies are available to blunt TAA progression and rupture. miR-146a, a short non-coding microRNA, is well known to regulate inflammatory and auto-immune processes under cardiovascular diseases. Increased miR-146a has been observed in plasma and dissected aortic tissue of TAA patients.
Hypothesis: miR-146a promotes actin cytoskeleton disassembly in the aorta by repressing FLNA, which subsequently decreases ECM production and accelerates TAA rupture.
Aim: To examine the effect of miR-146a deficiency on TAA rupture in mice induced by Lysyl oxidase inhibitor, β-aminopropionitrile (BAPN).
Methods: Three-week-old male and female C57BL/6J miR146a wild type (WT) or deficient (KO) [n=12-18/per group] mice were administered with either vehicle or BAPN (0.5% wt/vol) in drinking water for 28 days. TAA was examined by in vivo ultrasound aortic lumen measurements and ex vivo aortic external width measurements. Mortality by TAA rupture was calculated.
Results: BAPN administration promoted TAA development equivalently in both WT and KO male (WT=67%,12/18; KO=62%,8/13) and female (WT=75%,9/12; KO=50%,8/16) mice compared to vehicle control. miR-146a deficiency significantly protected mice from BAPN-induced TAA rupture (Male-WT=33%,6/18; KO=0%,0/13; Female–WT=33%,4/12; KO=0%,0/16; P<0.05) and improved survival rate (P<0.05). Histological and immunofluorescent analyses showed that BAPN-induced TAA is associated with increased medial elastin breaks, less ECM-collagen, and cytoskeletal disassembly as evidenced by decreased filamentous F-actin in the aortic media only in WT mice, not in miR-146a KO mice. In silico target prediction identified miR-146a binding sites in the cytoskeletal structural protein FLNA 3’UTR. Western blot and immunohistochemical analyses revealed a strong reduction of aortic FLNA protein especially in the SMC-rich aortic medial layer, whereas miR-146a deficiency prevented BAPN-induced loss of aortic FLNA protein.
Conclusion: These findings suggest that miR-146a plays a critical role in mediating TAA rupture by influencing aortic cytoskeletal-ECM structural assembly and integrity.