Activation of adrenergic receptor in H9c2 cardiac myoblasts co-stimulates Nox2 and the derived ROS mediate the downstream responses
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In recent years, NADPH oxidases (Noxes) have emerged as an important player in cardiovascular pathophysiology. Despite the growing evidences on the role of specific Nox isoforms, mechanisms of their activation, targets of reactive oxygen species (ROS) generated, and their downstream effects are poorly understood as yet. In this study, we treated H9c2 cardiac myoblasts with norepinephrine (NE, 2 µM), inducing ROS generation that was inhibited by Nox2-specific peptide inhibitor gp91ds-tat. Organelle-specific hydrogen peroxide-sensitive probe HyPer showed that the site of ROS generation is primarily in the cytosol, to some extent in the endoplasmic reticulum (ER) but not the mitochondria. Modulation of mRNAs of marker genes of cardiac hypertrophy i.e. induction in ANP and β-MHC, and reduction in α-MHC by NE treatment was prevented by specific inhibition of Nox2 by gp91ds-tat. Induction of ANP and β-MHC at the protein level were also attenuated by the inhibition of Nox2. Induction of c-Jun and FosB, the two members of the transcription factor family AP-1, were also blocked by the inhibition of Nox2 by gp91ds-tat. Induction of promoter-reporter constructs harboring multiple AP-1 elements and the upstream of FosB and ANP genes by NE were also blocked by the inhibition of Nox2 by gp91ds-tat and a dominant negative mutant of p22phox, a constituent of Nox2 that prevents its activation. This study for the first time establishes the significant role of Nox2 in mediating the NE-induced pathological adrenergic signaling in cardiac myoblasts.
KeywordsRedox signaling Reactive oxygen species NADPH oxidase Cardiac hypertrophy Norepinephrine gp91ds-tat
This work was supported by The Department of Biotechnology, Government of India, under Grant (BT/PR4268/BRB/10/1016/2011), awarded to SKG. NS is a recipient of a JRF/SRF from the Indian Council of Medical Research, Government of India.
Compiance with ethical standards
This article does not contain any studies performed with animals.
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