YAP1 silencing attenuated lung injury/fibrosis but worsened diaphragmatic function by regulating oxidative stress and inflammation response in mice

Oxidative stress is a crucial mechanism in the pathophysiology of lung injury/fibrosis and diaphragmatic dysfunction. Yes-associated protein 1 (YAP1) is a key oxidative stress response regulator. However, how lung injury/fibrosis and the subsequent YAP1 silencing treatment affect diaphragmatic function remains largely uncharacterized. In this study, mice models of acute lipopolysaccharide (LPS) and paraquat exposure were used to establish acute lung injury and chronic pulmonary fibrosis. AT2 and C2C12 cells were co-cultured under LPS and paraquat challenge. YAP1 was interfered with shRNA given in vivo and verteporfin administration in vitro. Pulmonary histology, contractile properties, and cross-sectional areas (CSAs) of the diaphragm and gastrocnemius were evaluated. Histological and biochemical analyses were performed for targeted biomarker determination. We found that LPS and paraquat caused significant lung injury/fibrosis and significantly reduced the diaphragmatic-specific force and CSAs compared with the control. YAP1 silencing alleviated inflammatory cell infiltration or collagen deposition in the lungs yet worsened the already impaired diaphragmatic function by increasing inflammatory cytokines (IL-6 and TNF-α), mitochondrial reactive oxidative species (ROS) emission, protein degradation (Murf-1, atrogin-1, and calpain), and decreasing antioxidant capabilities (superoxide dismutase 2 and glutathione peroxidase). No significant improvements were observed in diaphragmatic function by transient YAP1 knockdown in the gastrocnemius. In vitro, LPS- or paraquat-caused cytotoxicity in AT2 cells was mostly alleviated by verteporfin in a concentration that was 20-fold higher than that in C2C12 cells (20 and 1 μg/mL, respectively). Finally, 0.5 μg/mL of verteporfin significantly ameliorated hydrogen peroxide-induced proteolytic activity and antioxidant enzyme suppression in C2C12 cells, whereas 2 μg/mL of verteporfin deteriorated the same. Collectively, lung injury/fibrosis adversely affects the diaphragm. YAP1 inhibition alleviates lung injury/fibrosis but worsens diaphragmatic function potentially by enhancing inflammatory cytokines and ROS-mediated protein degradation. This disparity might be attributed to differences in susceptibility to YAP1 inhibition between muscles and the lungs.