Redox regulation of cardiomyocyte cell cycling via an ERK1/2 and c-Myc-dependent activation of cyclin D2 transcription

• Published in: Journal of molecular and cellular cardiology Vol. 79; pp. 54 - 68
• Main Authors: Murray, Thomas V.A, Smyrnias, Ioannis, Schnelle, Moritz, Mistry, Rajesh K, Zhang, Min, Beretta, Matteo, Martin, Daniel, Anilkumar, Narayana, de Silva, Shana M, Shah, Ajay M, Brewer, Alison C
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.02.2015; Academic Press
• Subjects: Animals; Animals, Newborn; Base Sequence; Binding Sites - genetics; c-Myc; Cardiomegaly - genetics; Cardiomegaly - pathology; Cardiomyocyte proliferation; Cardiovascular; Cell Cycle - genetics; Cell Proliferation; Conserved Sequence - genetics; Cyclin D2; Cyclin D2 - genetics; Cyclin D2 - metabolism; ERK1/2; Extracellular Signal-Regulated MAP Kinases - metabolism; Mice, Transgenic; Molecular Sequence Data; Myocardium - metabolism; Myocardium - pathology; Myocytes, Cardiac - cytology; Myocytes, Cardiac - enzymology; NADPH Oxidase 4; NADPH Oxidases - metabolism; Nox4; Original; Oxidation-Reduction; Phosphorylation; Promoter Regions, Genetic - genetics; Proto-Oncogene Proteins c-myc - metabolism; Rats; Reactive Oxygen Species - metabolism; Redox signalling; Transcription, Genetic; Up-Regulation - genetics; Cardiomyocyte proliferation; adenoviral Nox4; polyethylene glycol; c-Myc; neonatal rat cardiomyocyte; Cyclin D2; AdNox4; ERK1/2; PEG; Tg; bromodeoxyuridine; adenoviral β-galactosidase; AdβGal; Nox4; wild type; transgenic; Redox signalling; Wt; BrdU; NRC
• ISSN: 0022-2828; 1095-8584
• DOI: 10.1016/j.yjmcc.2014.10.017

Abstract Adult mammalian cardiomyocytes have a very limited capacity to proliferate, and consequently the loss of cells after cardiac stress promotes heart failure. Recent evidence suggests that administration of hydrogen peroxide (H2 O2 ), can regulate redox-dependent signalling pathway(s) to promote cardiomyocyte proliferation in vitro , but the potential relevance of such a pathway in vivo has not been tested. We have generated a transgenic (Tg) mouse model in which the H2 O2 -generating enzyme, NADPH oxidase 4 (Nox4), is overexpressed within the postnatal cardiomyocytes, and observed that the hearts of 1–3 week old Tg mice pups are larger in comparison to wild type (Wt) littermate controls. We demonstrate that the cardiomyocytes of Tg mouse pups have increased cell cycling capacity in vivo as determined by incorporation of 5-bromo-2′-deoxyuridine. Further, microarray analyses of the transcriptome of these Tg mouse hearts suggested that the expression of cyclin D2 is significantly increased. We investigated the molecular mechanisms which underlie this more proliferative phenotype in isolated neonatal rat cardiomyocytes (NRCs) in vitro , and demonstrate that Nox4 overexpression mediates an H2 O2 -dependent activation of the ERK1/2 signalling pathway, which in turn phosphorylates and activates the transcription factor c-myc. This results in a significant increase in cyclin D2 expression, which we show to be mediated, at least in part, by cis -acting c-myc binding sites within the proximal cyclin D2 promoter. Overexpression of Nox4 in NRCs results in an increase in their proliferative capacity that is ablated by the silencing of cyclin D2. We further demonstrate activation of the ERK1/2 signalling pathway, increased phosphorylation of c-myc and significantly increased expression of cyclin D2 protein in the Nox4 Tg hearts. We suggest that this pathway acts to maintain the proliferative capacity of cardiomyocytes in Nox4 Tg pups in vivo and so delays their exit from the cell cycle after birth.