title: Prohibitin confers cytoprotection against ISO-induced hypertrophy in H9c2 cells via attenuation of oxidative stress and modulation of Akt/Gsk-3β signaling.
abstract: Numerous hypertrophic stimuli, including β-adrenergic agonists such as isoproterenol (ISO), result in generation of reactive oxygen species (ROS) and alteration in the mitochondrial membrane potential (Δψ) leading to oxidative stress. This process is well associated with phosphorylation of thymoma viral proto-oncogene Akt (Ser473) and glycogen synthase kinase-3β (Gsk-3β) (Ser9), with resultant inactivation of Gsk-3β. In the present study, we found that the protective defensive role of prohibitin (PHB) against ISO-induced hypertrophic response in rat H9c2 cells is via attenuation of oxidative stress-dependent signaling pathways. The intracellular levels of mitochondrial membrane potential along with cellular ROS levels and mitochondrial superoxide generation were determined. In order to understand the regulation of Akt/Gsk-3β signaling pathway, we carried out immmunoblotting for key proteins of the pathway such as PTEN, PI3K, phosphorylated, and unphosphorylated forms of Akt, Gsk-3β, and immunofluorescence experiments of p-Gsk-3β. Enforced expression of PHB in ISO-treated H9c2 cells suppressed cellular ROS production with mitochondrial superoxide generation and enhanced the mitochondrial membrane potential resulting in suppression of oxidative stress which likely offered potent cellular protection, led to the availability of more healthy cells, and also, significant constitutive activation of Gsk-3β via inactivation of Akt was observed. Knockdown of PHB expression using PHB siRNA in control H9c2 cells reversed these effects. Overall, our results demonstrate that PHB confers cytoprotection against oxidative stress in ISO-induced hypertrophy and this process is associated with modulation of Akt/Gsk-3β signaling mechanisms as evident from our PHB overexpression and knockdown experiments.
authors.affiliation: Centre for Chemical Biology, CSIR-IICT (Indian Institute of Chemical Technology), Tarnaka, Hyderabad, Telangana, 500007, India.
authors.forename: Debabrata
authors.initials: D
authors.lastname: Chowdhury
authors.affiliation: Centre for Chemical Biology, CSIR-IICT (Indian Institute of Chemical Technology), Tarnaka, Hyderabad, Telangana, 500007, India.
authors.forename: Dinesh
authors.initials: D
authors.lastname: Kumar
authors.affiliation: Functional Genomics and Gene Silencing Group, CSIR-CCMB, Hyderabad, 500007, Telangana, India.
authors.forename: Utpal
authors.initials: U
authors.lastname: Bhadra
authors.affiliation: Centre for Chemical Biology, CSIR-IICT (Indian Institute of Chemical Technology), Tarnaka, Hyderabad, Telangana, 500007, India. anjana@iict.res.in.
authors.forename: Tangutur Anjana
authors.initials: TA
authors.lastname: Devi
authors.affiliation: Centre for Chemical Biology, CSIR-IICT (Indian Institute of Chemical Technology), Tarnaka, Hyderabad, Telangana, 500007, India. manika@iict.res.in.
authors.forename: Manika Pal
authors.initials: MP
authors.lastname: Bhadra
mesh_terms: D000818:Animals; D019610:Cytoprotection / Q000187:drug effects*; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism*; D006984:Hypertrophy / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control; D007545:Isoproterenol / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D000091402:Prohibitins; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D012097:Repressor Proteins / Q000096:biosynthesis*; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: C000717332:Phb protein, rat; D000091402:Prohibitins; D012097:Repressor Proteins; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D007545:Isoproterenol
keywords: Akt; Gsk-3β; Hypertrophy; Isoproterenol; Oxidative stress; Prohibitin