Relaxin protects cardiac muscle cells from hypoxia/reoxygenation injury: involvement of the Notch‐1 pathway

Abstract

In animal models, the cardiotropic hormone relaxin has been shown to protect the heart against ischemia and reperfusion-induced damage, acting by multiple mechanisms that primarily involve the coronary vessels. This in vitro study evaluates whether relaxin also has a direct protective action on cardiac muscle cells. H9c2 rat cardiomyoblasts and primary mouse cardiomyocytes were subjected to hypoxia and reoxygenation. In some experiments, relaxin was added preventatively before hypoxia; in others, at reoxygenation. To elucidate its mechanisms of action, we focused on Notch-1, which is involved in heart pre-and postconditioning to ischemia. Inactivated RLX was used as negative control. Relaxin (17 nmol/L, EC 50 4.7 nmol/L), added 24 h before hypoxia or at reoxygenation, protected against cardiomyocyte injury. In fact, relaxin significantly increased cell viability (assayed by trypan blue and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide), decreased apoptosis (assayed by TUNEL and bax/bcl-2 ratio), and reduced nitroxidative damage (assayed by nitrotyrosine expression and 8-hydroxy-deoxyguanosine levels). These effects were partly attributable to the ability of relaxin to upregulate Notch-1 signaling; indeed, blockade of Notch-1 activation with the specific inhibitor DAPT reduced relaxin-induced cardioprotection during hypoxia and reoxygenation. This study adds new mechanistic insights on the cardioprotective role of relaxin on ischemic and oxidative damage.-Boccalini, G., Sassoli, C., Formigli, L., Bani, D., Nistri, S. Relaxin protects cardiac muscle cells from hypoxia/reoxygenation injury: involvement of the Notch-1 pathway. FASEB J. 29, 239-249 (2015). www.fasebj.orgCARDIOVASCULAR DISEASE is the leading cause of death in modern society worldwide (1). In particular, cardiac dysfunction caused by nonlethal myocardial infarction remains one of the most challenging clinical problems. The pathogenic occurrences in the infarcted heart are dependent on ischemia/reperfusion (I/R). The mechanisms of I/R injury are complex and involve multiple events, basically nitroxidative stress (2, 3) and inflammation (4), that play a key role in the progression of cardiac damage and in the development of myocardial fibrosis and heart failure (5). Hence, there is a major interest in the identification of new therapeutic agents that can prevent or reduce I/R-induced myocardial injury. In this context, the hormone relaxin (RLX) emerges as a feasible candidate. Best known for its effects on reproduction (6), RLX has been validated as a bona fide cardiovascular hormone (7-9). Studies in rats have shown that immunoreactive RLX is produced by cultured atrial cardiomyocytes (10) and that RLX receptors are expressed by the atrial and ventricular myocardium (11) and ventricular cardiomyocytes (12). RLX is capable of enhancing cardiac blood perfusion by dilating the preexisting coronary vessels (13) and promoting neo-angiogenesis (14, 15). The increased blood supply to the heart induced by RLX, accompanied by its clear-cu...

MicroRNA-449a Inhibition Protects H9C2 Cells Against Hypoxia/Reoxygenation-Induced Injury by Targeting the Notch-1 Signaling Pathway

“…In H9C2 cells, the activation of the Notch 1 gene was demonstrated to effectively promote cell proliferation and inhibit apoptosis in ischemic post-conditioning [39]. Consistently, Notch-1 can protect cardiomyocytes against hypoxia or I/R injury [38]. All these findings support that Notch activation is a favorable signal in myocardial I/R injury.…”

Notch Signaling in Ischemic Damage and Fibrosis: Evidence and Clues from the Heart

“…Targeting Notch signaling, for example by soluble Notch ligands or Notch pathway activating molecules delivered intramyocardially or embedded into suitable biomaterials (Gude et al, 2008; Limana et al, 2013; Boccalini et al, 2015; Boopathy et al, 2015), may be a worthwhile therapeutic approach to cardiovascular disease.…”

The impact of neuronal Notch-1/JNK pathway on intracerebral hemorrhage-induced neuronal injury of rat model

“…In recent years, the disrupted Notch-1 expression is associated with some neurodegenerative or cardiovascular diseases [18, 19]. During early neural development, Notch is highly expressed in the neural precursor cells and plays a critical role in regulating neural proliferation and differentiation [6].…”