16 Myocardial Expression of Mutant P53 Protects Against Doxorubicin-Induced Cardiomyocyte Apoptosis and Preserves Heart Function.

Abstract

Doxorubicin, a topoisomerase II inhibitor that is commonly used to treat childhood cancer, induces p53-mediated cardiomyocyte apoptosis which can ultimately lead to heart failure. We hypothesized that cardiac-restricted expression of a dominant interfering p53 mutant (CB7) would protect cardiomyocytes from doxorubicin-induced apoptosis in vivo. Age-matched CB7 transgenic mice and control non-transgenic (NTG) mice were treated with a total of 20mg/kg of doxorubicin (2 intraperitoneal injections of 10mg/kg at 3-day interval). The prevalence of cardiomyocyte apoptosis was assessed via anti-activated caspase-3 immune reactivity and also via terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining. Cleavage of poly (ADP-ribose) polymerase (PARP, a substrate of caspase-3) and the level of the anti-apoptotic protein Bcl-xL were also monitored. Cardiac function was assessed using the two dimensional echocardiography. Our results show that the prevalence of cardiomyocyte apoptosis was reduced in CB7 mice as compared to NTG mice following doxorubicin injection (activated caspase 3 immune reactivity: 0.85± 0.14 positive cardiomyocytes/mm² vs. 1.98±0.24 positive cardiomyocytes/mm², p<0.05, n=6; TUNEL staining: 0.045±0.006% of total cardiomyocyte nuclei vs. 0.146±0.009%, p<0.01, n=5). In agreement with this, caspase-dependent PARP cleavage was not detected in the doxorubicin-treated CB7 hearts but was detected in the doxorubicin-treated NTG. Bcl-xL expression was induced to a greater extent in CB7 mice as compared to NTG mice. Finally, fractional shortening was better preserved in the CB7 mice as compared to the NTG mice (7.79±1.35% reduction post-injection vs. 28.87±5.48%, p<0.05, n=6). Interestingly, the extent of functional preservation observed in DOX treated CB7 mice appeared to exceed that which would be expected based on the reduced levels of cardiomyocyte apoptosis. In conclusion, cardiac expression of CB7 protects against doxorubicin-induced cardiomyocyte apoptosis and preserves heart function. The anti-apoptotic effects of CB7 might result from the increased level of Bcl-xL. The disproportionate preservation of cardiac function suggests that antagonization of p53 may engender additional cardioprotective activity.

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