leupeptins and Cardiomegaly

The elevated expression of poly(ADP-ribose) polymerase-1 (PARP1) and increased PARP1 activity, namely, poly(ADP-ribosyl)ation (PARylation), have been observed in cardiac remodeling, leading to extreme energy consumption and myocardial damage. However, the mechanisms underlying the regulation of PARP1 require further study. WWP2, a HECT-type E3 ubiquitin ligase, is highly expressed in the heart, but its function there is largely unknown. Here, we clarified the role of WWP2 in the regulation of PARP1 and the impact of this regulatory process on cardiac remodeling. We determined that the knockout of WWP2 specifically in myocardium decreased the level of PARP1 ubiquitination and increased the effects of isoproterenol (ISO)-induced PARP1 and PARylation, in turn aggravating ISO-induced myocardial hypertrophy, heart failure, and myocardial fibrosis. Similar findings were obtained in a model of ISO-induced H9c2 cells with WWP2 knockdown, while the reexpression of WWP2 significantly increased PARP1 ubiquitination and decreased PAPR1 and PARylation levels. Mechanistically, coimmunoprecipitation results identified that WWP2 is a novel interacting protein of PARP1 and mainly interacts with its BRCT domain, thus mediating the degradation of PARP1 through the ubiquitin-proteasome system. In addition, lysine 418 (K418) and lysine 249 (K249) were shown to be of critical importance in regulating PARP1 ubiquitination and degradation by WWP2. These findings reveal a novel WWP2-PARP1 signal transduction pathway involved in controlling cardiac remodeling and may provide a basis for exploring new strategies for treating heart disorders related to cardiac remodeling.

Topics: Animals; Cardiomegaly; Fibrosis; Heart Failure; Humans; Isoproterenol; Leupeptins; Lysine; Male; Mice, Inbred C57BL; Mice, Transgenic; Models, Biological; Myocardium; Poly ADP Ribosylation; Poly(ADP-ribose) Polymerases; Proteasome Endopeptidase Complex; Protein Binding; Proteolysis; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination; Ventricular Remodeling

Proteasome inhibitors are involved in cell cycle control, growth and inflammatory signaling, and transcriptional regulation of mitotic cells. A recent study has suggested that specific proteasome inhibitor MG132 may suppress cardiomyocyte hypertrophy in vitro. However, the underlying molecular mechanisms are not clear. In this study, we investigated the effects of long-term MG132 treatment on cardiac hypertrophy and the related molecular mechanisms in vivo. MG132 (0.1 mg/kg/day) was intraperitoneally injected to rats with abdominal aortic banding (AAB) for 8 weeks. Results showed that treatment with MG132 significantly attenuated left ventricular (LV) myocyte area, LV weight/body weight, and lung weight/body weight ratios, decreased LV diastolic diameter and wall thickness, and increased fractional shortening in AAB rats. AAB induced the phosphorylation of ERK1/2, JNK1, and p38 in cardiac myocytes. The elevated phosphorylation levels of ERK1/2 and JNK1 in AAB rats were significantly reversed by MG132 treatment. In conclusion, our results suggested that long-term treatment with MG132 attenuates pressureoverload-induced cardiac hypertrophy and improves cardiac function in AAB rats through regulation of ERK1/2 and JNK1 signaling pathways.

Topics: Animals; Aorta; Cardiomegaly; Echocardiography; Heart Ventricles; Leupeptins; Male; MAP Kinase Signaling System; Phosphorylation; Pressure; Protease Inhibitors; Proteasome Inhibitors; Rats; Rats, Sprague-Dawley; Signal Transduction

Inhibitors of the proteasome interfere with transcriptional regulation of growth signaling pathways and block cell cycle progression of mitotic cells. As growth signaling pathways are highly conserved between mitotic and postmitotic cells, we hypothesized that proteasome inhibition might also be a valuable approach to interfere with hypertrophic growth of postmitotic cardiomyocytes. To test this hypothesis, we analyzed the effects of proteasome inhibition on hypertrophic growth of neonatal rat cardiomyocytes. Partial inhibition of the proteasome effectively suppressed cardiomyocyte hypertrophy as determined by reduced cell size, inhibition of hypertrophy-mediated induction of RNA and protein synthesis, reduced expression of several hypertrophic marker genes, and diminished transcriptional activation of the BNP promotor. Importantly, suppression of hypertrophic growth was independent of the hypertrophic agonist used. Expressional profiling and subsequent Western blot and kinase assays revealed that proteasome inhibition induced a cellular stress response with reduced expression of conserved growth signaling mediators and impaired G1/S phase transition of cardiomyocytes. In hypertensive Dahl-salt sensitive rats, inhibition of the proteasome with low doses of the FDA approved proteasome inhibitor Velcade significantly reduced hypertrophic heart growth. Our data provide important insight into the suppressive effects of proteasome inhibitors on hypertrophic growth of cardiomyocytes and establish low-dose proteasome inhibition as a new and broad-spectrum approach to interfere with cardiac hypertrophy.

Topics: Animals; Animals, Newborn; Biomarkers; Boronic Acids; Cardiomegaly; Cells, Cultured; Cysteine Proteinase Inhibitors; Dose-Response Relationship, Drug; Down-Regulation; Hypertrophy; Leupeptins; Male; Myocytes, Cardiac; Proteasome Inhibitors; Rats; Rats, Inbred Dahl; Signal Transduction; Ubiquitin