Angiotensin II and hypoxia induce autophagy in cardiomyocytes via activating specific protein kinase C subtypes.

Abstract

The purpose of this study was to explore the role of protein kinase C (PKC) isozymes and reactive oxygen species (ROS) in hypoxia and angiotensin (Ang) II-induced autophagy. Primary cardiomyocytes were isolated from Sprague-Dawley (SD) neonatal rats and cultured in hypoxia and/or Ang II conditions. Dihydroethidium fluorescence staining was used to detect the content of ROS. Cardiomyocyte autophagy was determined using Monodansylcadaverine fluorescence staining and Western blot. We also inhibited ROS production to explore the relationship between ROS and autophagy. ELISA was used to detect the contents of PKC δ and PKC ε. After inhibition of PKC δ activation and PKC ε expression by lentiviral siRNA, ROS content and autophagy of cultured cardiomyocytes were detected. Hypoxia and Ang II stimulation increased autophagy in cardiomyocytes, accompanied by increased intracellular ROS production. Inhibiting ROS following hypoxia or Ang II stimulation significantly suppressed autophagy in comparison with hypoxia or Ang II stimulation group. Inhibiting PKC δ significantly reduced ROS production and autophagy activity following hypoxia or accompanied with Ang II stimulation except Ang II stimulation alone. Knockdown of PKC ε notably decreased ROS production and autophagy in response to Ang II alone and in combination with hypoxia rather than hypoxia alone. Both hypoxia and Ang II stimulation can induce autophagy in cardiomyocytes through increasing intracellular ROS. However, hypoxia and Ang II stimulation induced myocardial autophagy via PKC δ and PKC ε, respectively.