title: Diabetes-induced increased oxidative stress in cardiomyocytes is sustained by a positive feedback loop involving Rho kinase and PKCβ2.
abstract: We previously reported that acute inhibition of the RhoA/Rho kinase (ROCK) pathway normalized contractile function of diabetic rat hearts, but the underlying mechanism is unclear. Protein kinase C (PKC) β(2) has been proposed to play a major role in diabetic cardiomyopathy at least in part by increasing oxidative stress. Further evidence suggests that PKC positively regulates RhoA expression through induction of inducible nitric oxide synthase (iNOS) in diabetes. However, in preliminary studies, we found that inhibition of ROCK itself reduced RhoA expression in diabetic hearts. We hypothesized that there is an interaction between RhoA/ROCK and PKCβ(2) in the form of a positive feedback loop that sustains their activation and the production of reactive oxygen species (ROS). This was investigated in cardiomyocytes isolated from diabetic and control rat hearts, incubated with or without cytochalasin D or inhibitors of ROCK, RhoA, PKCβ(2), or iNOS. Inhibition of RhoA and ROCK markedly attenuated the diabetes-induced increases in PKCβ(2) activity and iNOS and RhoA expression in diabetic cardiomyocytes, while having no effect in control cells. Inhibition of PKCβ(2) and iNOS also normalized RhoA expression and ROCK overactivation, whereas iNOS inhibition reversed the increase in PKCβ(2) activity. Each of these treatments also normalized the diabetes-induced increase in production of ROS. Actin cytoskeleton disruption attenuated the increased expression and/or activity of all of these targets in diabetic cardiomyocytes. These data suggest that, in the diabetic heart, the RhoA/ROCK pathway contributes to contractile dysfunction at least in part by sustaining PKCβ(2) activation and ROS production via a positive feedback loop that requires an intact cytoskeleton.
authors.affiliation: Molecular and Cellular Pharmacology Research Group, Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3.
authors.forename: Hesham
authors.initials: H
authors.lastname: Soliman
authors.forename: Anthony
authors.initials: A
authors.lastname: Gador
authors.forename: Yi-Hsuan
authors.initials: YH
authors.lastname: Lu
authors.forename: Guorong
authors.initials: G
authors.lastname: Lin
authors.forename: Girish
authors.initials: G
authors.lastname: Bankar
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: MacLeod
mesh_terms: D008841:Actin Cytoskeleton / Q000201:enzymology; D000818:Animals; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism / Q000473:pathology; D058065:Diabetic Cardiomyopathies / Q000378:metabolism* / Q000473:pathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D025461:Feedback, Physiological / Q000502:physiology*; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology; D008297:Male; D009200:Myocardial Contraction / Q000502:physiology; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology; D010766:Phosphorylation / Q000502:physiology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D064546:Protein Kinase C beta; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D020742:rhoA GTP-Binding Protein; D005947:Glucose