Effects of Mechanical Ventilation and Autophagy on Diaphragm Oxidative Stress and Proteolysis

Abstract

Mechanical ventilation (MV) is a life-saving measure for patients in respiratory failure. However, prolonged MV results in diaphragm weakness due to muscle fiber atrophy and contractile dysfunction. Therefore, identifying the signaling pathways responsible for ventilator-induced diaphragm dysfunction (VIDD) is important. While previous reports demonstrate an increase in the autophagy/lysosomal system during MV, the contribution of autophagy to VIDD is unknown. Thus, these experiments were designed to determine the effects of accelerated autophagy on the diaphragm during MV. Cause and effect was determined by inhibiting MV-induced autophagy via adeno-associated virus overexpression of mutated autophagy-related protein 5 (ATG5) in the diaphragm of rats. Our results reveal that inhibiting autophagy prevented the MV-induced reduction in both diaphragm muscle fiber size and force generation. In addition, overexpression of mutated ATG5 prior to MV caused a significant reduction in MV-induced diaphragm lipid peroxidation and mitochondrial ROS production. Also, transduction of mutated ATG5 resulted in increased diaphragm cytosolic catalase content compared to mechanically ventilated control animals. Finally, our results show a regulatory cross-talk may exist between autophagy, calpain and caspase-3. Specifically, inhibition of autophagy resulted in a reduction of the MV-induced increase in proteolytic activity of calpain and caspase-3. Therefore, our data indicate that MV-induced autophagy is detrimental to the diaphragm and inhibition of autophagic signaling protects against VIDD by reducing diaphragmatic oxidative stress and proteolysis.