Abstract 616: Cardiomyocyte-Specific Over-Expression of Plasma Membrane Ca 2+ ATPase 4-C-I Improves Cardiac Performance and Reduces Compensatory Hypertrophy after Infarction
Background: The role of plasmalemmal Ca2+ ATPase (PMCA) pumps in cardiomyocytes (CM) was explored by over-expressing the caveolin- and nNOS-associating isoform PMCA4-C-I (P4b) with an αMHC promoter-driven Doxycycline (Dox)-responsivetranscriptional activator (tTA) in transgenic mice.
Methods & Results: All genotypes appeared with expected frequencies, and none altered mouse survival. RT-PCR, Western blot and immunochemistry confirmed Dox-regulated, CM-restricted, caveolin-localized expression of P4b in binary transgenic (BT: tTA+/P4b+) but not non-binary transgenic (NBT) littermates. Westerns and coupled enzyme assays revealed 12-fold higher PMCA protein and 6-fold higher PMCA activity in BT vs. NBT mice (p<0.05). Real-time PCR confirmed adaptive changes in levels of other Ca2+-regulatory genes. By contrast, total and neuronal NOS activities, as determined by L-[3H] arginine to L-[3H] citrulline conversion (n=6), were reduced in BT vs. NBT littermates (2.3±0.13 vs. 4.3±0.14; 0.32±0.19 vs. 1.3±0.25 pmol/min/mg, p<0.05). Trans-carotid Millar catheterization of adult mice revealed elevated systolic BP in BT vs. NBT and Dox-treated BT mice (p<0.001) before MI, but no significant difference 2 weeks after MI. Although HR did not vary, BT mice showed improved LV performance at baseline (p<0.05) and a similar trend post-MI. BT mice also showed cardiac hypertrophy with higher heart-to-body weight ratios (5.3±0.3 vs. 4.8±0.1; mg/g; p<0.05). Interestingly, this phenotype was inverted post-MI (8±1 vs. 9±1 mg/g; p<0.05), suggesting beneficial effects of P4b expression on cardiac remodeling. M-mode echo showed smaller LV diameters and higher %FS in basal and post MI conditions (<0.05) in BT mice. Histomorphometry revealed smaller infarcts in BT mice (25% vs. 36%; p<0.05). Freshly isolated CM confirmed hypertrophy of BT vs. NBT cells at baseline, but electrical field stimulation of Indo-loaded CM revealed no significant differences in Ca2+-transients or diastolic Ca2+ concentrations.
Conclusion: Cardiac P4b levels regulate nNOS activity, cardiac mass and contractility, such that P4b over-expression heightens cardiac performance and limits infarct size and cardiac hypertrophy post-MI.