Abstract 312: Selective Inhibition of Myocardial NADPH Oxidase Significantly Decreases Heart Size in Transgenic (Tg) Mice with Cardiac-Specific Overexpression of a Dominant-Negative (DN) Mutant of p67 phox

Abstract

The role of NADPH oxidase (NOX) was recently suggested in cardiac diseases including hypertrophy. Overexpression of a DN mutant of p67 phox , a cytosolic subunit of NOX, DN-p67 with a V204A point mutation completely inhibited NOX enzymatic function of superoxide generation. We found that overexpression of DN-p67 in cultured adult rat cardiac myocytes attenuated α 1 -adrenoceptor-induced hypertrophy , we thus hypothesized that NOX/p67 phox plays a critical role in cardiac hypertrophy and remodeling in vivo . Accordingly, we recently generated a Tg mouse model in C57BL/6 strain with cardiac-specific overexpression of this DN-p67 mutant along with an IRES-initiated EGFP tag protein using the cardiac-specific α-MHC promoter. Hearts from 1.5-, 3- and 6-month (M) old heterozygous Tg mice and their Non-Tg (NTg) littermates were harvested and compared. The heart weight/body weight (HW/BW) ratio in Tg group was significantly smaller (overall: 14%↓; 1.5-M: 10%↓; 3-M: 16%↓; 6-M: 12%↓) vs. the NTg group ( p < 0.01, n=23). This result was confirmed by 2D echocardiography (2D-echo) measuring the left ventricular myocardial area (LVMA) determined by subtracting endocardial area from epicardial area in short axis view at the end diastole. The LVMA (mm 2 ) in 3-M and 6-M mice was significantly ( p < 0.05, n=21) smaller in Tg (0.44±0.02) than the NTg (0.50±0.02) with equal average BW in both groups. LV internal dimensions and systolic function were preserved without difference in Tg vs. NTg. The LV wall thickness is smaller in Tg vs. NTg by both 2D-echo and hematoxylin/eosin staining of LV cross-sections, but no change in fibrosis was detected in both groups by trichrome staining (n=4). Interestingly, the expression of NOX catalytic subunit gp91 phox in Tg hearts was significantly decreased (~3-fold↓, n=5) vs. NTg by Western blot, whereas the cardiac expression of other NOX subunits p47 phox and p22 phox remained the same in both groups. In summary, we have generated a viable Tg mouse model with cardiac-specific overexpression of a DN p67 phox , which shows significant reduction in both myocardial mass/size and gp91 phox expression with normal cardiac function and development. Our results first show the novel and critical role of NOX/p67 phox in normal cardiac growth and heart development. This research has received full or partial funding support from the American Heart Association, AHA National Center.