Fujimoto, Hirokazu

Increasing age is the greatest risk factor for development and progression of calcific aortic valve disease (CAVD), and mitochondrial dysfunction has been implicated in the pathogenesis of cardiovascular calcification. Previous findings by our group suggested a significant reduction in expression of multiple sirtuin (SIRT) isoforms in normocholesterolemic aged mouse aortic valves, however, it is unclear if losses in SIRT3—a major mitochondrial SIRT isoform—modulates progression of aortic valve calcification. Therefore, we hypothesized that loss of SIRT3 in a hypercholesterolemic mouse model of CAVD will result in augmented calcium burden in aortic valve, increased osteogenic signaling, and impaired aortic valve function. To test this we used mice that were Ldlr -/- /ApoB 100/100 mice that were either wild-type (LA-SIRT3 +/+ ) or null for SIRT3 (LA-SIRT3 -/- ) and fed a western diet (TD88137) for 12 months. Alizarin red was used to quantitate calcium burden, qRT-PCR was used to measure changes in mRNA levels, and high-resolution echocardiography was used to assess aortic valve function (cusp separation distance). In line with our hypothesis, we observed a substantial increase in calcium burden in LA-SIRT3 -/- mice compared to their LA-SIRT3 +/+ littermates(11.7±4.0, 4.8±1.8; respectively). Interestingly, expression of Runx2 and osterix—classic markers of osteogenic differentiation—were decreased in LA-SIRT3 -/- mice compared to their LA-SIRT3 +/+ littermates. Expression of Msx2, were markedly increased in aortic valve tissue from LA-SIRT3-deficient mice. Despite these histological and molecular changes, SIRT3 deletion did not alter cusp separation distance (LA-SIRT3 +/+ = 0.81±0.04; LA-SIRT3 -/- = 0.81±0.02). Collectively, this data suggest losses in SIRT3 can contribute to accelerated valve calcification through unclear molecular mechanisms, but these changes are not sufficient to drive reductions in cusp separation distance. Additional experiments delineating the histopathological and molecular sequelae of SIRT3 deletion will be critical to understanding its role in the pathogenesis of CAVD.