Nrg - 1 To Protect The Mechanism Of Myocardial Cell Injury Caused By Adriamycin

Neuregulin-1attenuates doxorubicin-induced autophagy in neonatal rat cardiomyocytesBackground:Doxorubicin (DOX) is an effective antineoplastic drug and is frequently used in the treatment of hematologic and solid tumors including leukemia, breast cancer and sarcomas. However, its clinical benefit is limited by its cardiotoxicity. To date, treatments that to protect the heart from DOX-induced damage are limited. Therefore, the approaches to treating DOX-induced cardiotoxicity remains a critical issue in both cardiology and oncology. Recent studies suggest that increased autophagy accompanies overt presentation of cardiac damage and may contribute to DOX-induced cardiotoxicity. Autophagy is a cellular process of "self-digestion" through the lysosomal degradation pathway and is highly conserved across multiple phyla. Elucidated first in lower organisms such as yeast, it is a multiple-step, highly regulated mechanism for degrading proteins and organelles that have become damaged or need to be recycled for other reasons, such as utilization during calorie deprivation. Autophagy also has been implicated in various pathological processes and some data suggest that autophagy may serve as a cell death mechanism under certain circumstances. This autophagy-dependent cell death has been defined as "autophagic cell death" or "type II programmed cell death."Neuregulin (NRG)-1, a member of the neuregulin family, is expressed in many cell types and organs, including the heart. Neuregulin-1/erbB signaling is essential for embryonic cardiac development. Post-natal conditional erbB2-deficiency in cardiomyocytes results in severe cardiomyopathy and enhanced myocyte susceptibility to DOX-induced death. Recombinant human neuregulin-1(rhNRG-l,part of NRG-1) is a61-amino-acid peptide containing an EGF-like domain, which is the domain necessary for ErbB2/ErbB4activation. Previously, we reported that rhNRG-1could improve cardiac function in patients suffering from congestive heart failure (CHF), with significant increases in left ventricular (LV) ejection fraction (LVEF). Treatment also decreased end systolic and diastolic volume (ESV and EDV, respectively), demonstrating a beneficial effect on pathological remodeling. It has been reported that rhNRG-1can activate Erb2/4heterodimerization, thus improving cardiac function and survival in animal models of DOX-induced cardiomyopathy. However, the underlying molecular mechanism remains to be defined and the role of autophagy in the process is unclear.Based on these considerations, we tested the hypothesis that rhNRG-1can attenuate DOX-induced autophagy, using neonatal rat ventricular myocytes (NRVMs) as a model system. Here, we investigate the roles of rhNRG-1/erbB signaling and autophagy in DOX cardiotoxicity, using the NRVM model of DOX-induced cell autophagy. We demonstrate that rhNRG-1attenuated DOX-induced autophagy and this is accompanied by Akt activation and ROS reduction, resulting in the upregulation of Bcl-2protein levels. These signaling processes partially contribute to the ability of rhNRG-1to inhibit DOX-induced autophagy.Methods:Neonatal rat ventricular mocytes(NRVMs) were separated from two-day-old SD rats. NRVMs were subjected to various treatments in order to both induce autophagy and to determine the effects of rhNRG-1on the process. After treated with recombinant human neuregulin-1(rhNRG-1,1000ng/ml), DOX(1μM), the inhibitor of PI3K LY294002(10μM), NAC(50μM)or cells were transfected with Bcl-2siRNA. Activation of autophagy was determined by observing increases in the well-defined markers of autophagy (including protein levels of LC3and Beclin1, the formation of GFP-LC3puncta). The activation of Akt was detected by means of western blot analysis. Intracellular ROS was determined via fluorescent detection of dihydrodichlorofluorescein (DCF).Results:rhNRG-1inhibited DOX-induced autophagy, reduced reactive oxygen species production and increased protein expression of Bcl-2, effects that were recapitulated when the cells were treated with the antioxidant N-acetyl cysteine (NAC). These effects were blocked by the phosphoinositol-3-kinase (PI3-k) inhibitor LY294002, pointing to the involvement of the Akt pathway in mediating the process. Inhibition of Bcl-2expression with siRNA silencing also inhibited rhNRG-1’s ability to attenuate DOX-induced autophagy.Conclusion:RhNRG-1is a potent inhibitor of DOX-induced autophagy and multiple signaling pathways, including Akt and activation of ROS, play important roles in the anti-autophagy effect. rhNRG-1is a novel drug that may be effectively therapeutically in protecting further damage in DOX-induced damaged myocardium. Neuregulin-1protects against doxorubicin-induced apoptosis in cardiomyocytes through an Akt-dependent pathwayBackground:Doxorubicin (DOX) is an effective antineoplastic drug, and is frequently used in the treatment of hematologic and solid tumors, such as leukemia, breast cancer and sarcoma. However, the drug’s clinical benefit is limited by its cardiotoxicity. DOX-induced cardiomyopathy is characterized by irreversible left ventricular dysfunction and congestive heart failure with a poor prognosis. Nevertheless, to date, researchers and scientists have attempted a variety of approaches aimed at preventing the deleterious action of doxorubicin, but presently the ability of these treatments to protect the heart from damage remains limited. Therefore, the development of more therapies which may be used to prevent and/or treat the cardiotoxicity of doxorubicin remains a critical issue in both cardiology and oncology. Neuregulin (NRG)-1, a member of the neuregulin family, is expressed in many cell types and organs, including the heart. Neuregulin-1/erbB signaling is essential for embryonic cardiac development. Post-natal conditional erbB2-deficiency in cardiomyocytes may result in severe cardiomyopathy and enhanced myocyte susceptibility for DOX-induced death. There are at least31NRG-1isoforms derived from the NRG-1gene which are produced by utilizing different promoters and alternative splicing, and different groups use different ligands. Among these isoforms, recombinant human neuregulin-1(rhNRG-1, a component of NRG-1) is a61-amino-acid peptide containing an EGF-like domain, the domain which is necessary for ErbB2/ErbB4activation. The authors of this study previously reported that rhNRG-1is capable of improving cardiac function in patients suffering from congestive heart failure (CHF), with significant increases in left ventricular (LV) ejection fraction (LVEF). Treatment has also decreased end systolic and diastolic volume (ESV and EDV, respectively), demonstrating a beneficial effect on pathological remodeling. It has also been reported that rhNRG-1is capable of activating Erb2/4heterodimerization, thus improving cardiac function and survival in animal models of doxorubicin-induced cardiomyopathy. However, the underlying molecular mechanism has yet to be defined. Akt is known to regulate many survival pathways of the cardiac cells. Recent studies have provided evidence that the anti-apoptotic effects of rhNRG-1are at least partially mediated by the alteration of PI3K/Akt signaling pathway during H2O2-induced cardiomyocyte apoptosis, as well as ischemia/reperfusion injury in rat hearts. However, whether or not rhNRG-1is able to protect cardiomyocytes from DOX-induced apoptosis through the PI3K/Akt pathway has yet to be thoroughly investigated.In view of this, the authors of this paper postulate that the pretreatment of rhNRG-1possesses protective effects against DOX-induced injury in cardiomyocytes, and the activation of PI3K/Akt pathway occurs during the process.Methods:Neonatal rat ventricular mocytes(NRVMs) were separated from two-day-old SD rats. NRVMs were subjected to various treatments, in order to both induce apoptosis and determine the effects of rhNRG-1on the process. After treated with different concentration of recombinant human neuregulin-1(rhNRG-1,10ng/ml-1000ng/ml), DOX(1μM) or the inhibitor of PI3K LY294002(10μM). Activation of apoptosis was determined by observing increases in the protein levels of classic apoptosis markers (including cleaved caspase-3, cytochrome c, Bcl-2, BAX and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining). The activation of Akt was detected by means of western blot analysis.Results:The study results showed that doxorubicin increased the number of TUNEL positive cells, as well as the protein levels of cleaved caspase-3and cytochrome c, and reduced the ratio of Bcl-2/Bax. However, all of these effects were markedly antagonized by pretreament with rhNRG-1. It was then further demonstrated that the effects of rhNRG-1could be blocked by the phosphoinositole-3-kinase inhibitor LY294002, indicating the involvement of the Akt process in mediating the process.Conclusion:RhNRG-1is a potent inhibitor of doxorubicin-induced apoptosis, which acts through the PI3K-Akt pathway. RhNRG-1is a novel therapeutic drug which may be effective in preventing further damage from occurring in DOX-induced damaged myocardium.