MHY1485

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are severe clinical conditions with limited treatment options. Toosendanin (TSN), a triterpenoid compound with anti-inflammatory effects, has unclear efficacy in ALI.

This study aimed to evaluate TSN's protective effects on ALI and the related mechanisms.

Lipopolysaccharide (LPS)-induced ALI models were developed in vivo and in vitro. Endothelial permeability was measured using Evans Blue dye; lipid reactive oxygen species (ROS) and apoptosis were assessed using flow cytometry. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were determined, and cell viability was measured. mRNA and protein expression were quantified using qRT-PCR and Western blotting. Network pharmacology and surface plasmon resonance were used to identify and validate TSN's targets.

TSN reduced endothelial permeability and LPS-induced ALI. It lowered ROS levels, lipid peroxidation, endoplasmic reticulum (ER) stress, and apoptosis, both in vitro and in vivo. Network pharmacology identified mTOR as a key target of TSN, and surface plasmon resonance analysis confirmed TSN's direct binding to mTOR, underscoring mTOR's role in TSN's protective effects against ALI. Western blotting showed that TSN inhibits mTOR and its phosphorylation. In vitro, the mTOR activator MHY1485 reversed TSN's protective effects, increasing ER stress, apoptosis, and endothelial permeability. In vivo, TSN and rapamycin synergistically protected against ALI.

This study is the first to demonstrate that TSN protects against ALI by targeting the mTOR pathway, regulating ER stress and apoptosis and mitigating endothelial damage. These findings suggest a novel approach for ALI treatment and underscore TSN's potential clinical value.

The processes of autophagy, including autophagosome formation, fusion of autophagosomes with lysosomes, and degradation of autophagosomes by lysosomes, are regulated by various mechanisms. We recently found that treatment with resveratrol, an activator of the NAD⁺-dependent protein deacetylase Sirtuin-1 (SIRT1), in a mouse model prevented autophagosome accumulation in the heart with high mTORC1 activity. In this study, we investigated whether SIRT1 mediates the effects of resveratrol on autophagosome elimination using a cardiomyocyte model. In H9c2 cardiomyocytes, treatment with the mTORC1 activator MHY1485 induced autophagosome accumulation accompanied by increases in fragmented mitochondria within the autophagosomes and levels of intracellular reactive oxygen species (ROS), indicative of impaired autophagy-mediated elimination of mitochondria and resultant oxidative stress. MHY1485 suppressed the fusion of autophagosomes with lysosomes. Co-treatment with resveratrol attenuated the MHY1485-induced increases in autophagosomes, mitochondria within autophagosomes, and levels of ROS. Knockdown of Sirt1 reversed the reductions in autophagosomes and ROS levels induced by resveratrol under the condition of MHY1485 treatment. Neither resveratrol treatment nor Sirt1 knockdown modulated the phosphorylation levels of UVRAG, a target of mTORC1 for suppression of autophagosome-lysosome fusion. Our findings suggest that SIRT1 mediates the resveratrol-induced promotion of autophagosome elimination in cells with high mTORC1 activity.