17β-estradiol and testosterone protect mitochondria against oxidative stress in skeletal muscle cells

Abstract

We have previously shown that testosterone (T) and 17β-estradiol (E2) protect C2C12 muscle cells against apoptosis induced by hydrogen peroxide (H2O2). Since we also showed the presence of estrogen and androgen receptors in mitochondria, this work was focused on the effects of both steroids on this organelle, which result in cellular survival. Specifically, we evaluated the actions of T and E2 on the mitochondrial membrane potential with JC-1 dye and on the mitochondrial permeability transition pore (MPTP) by the calcein-acetoxymethylester/cobalt method, using fluorescence microscopy and flow cytometry. We demonstrated that T and E2 prevented MPTP opening and the loss of mitochondrial membrane potential induced by H2O2. In addition, it was observed that H2O2 increased voltage-dependent anion channel (VDAC) protein expression levels and induced translocation of Bax to mitochondria. However, in the presence of the steroid Bax translocation was abrogated suggesting that members of the Bcl-2 family may be regulated by E2 and T. The observed effects triggered by E2 and T were reflected on mitochondrial morphology. Microscopic analysis of C2C12 cells and primary cultures of mouse skeletal muscle, with Janus Green and Mitotracker staining revealed a protective effect of the steroids against oxidative stress damage which included mitochondrial redistribution and pyknosis of the organelle.