Sulforaphane Inhibits LPS-induced Macrophage PANoptosis via TLR4/NFκB Pathway: A Potential Therapeutic Strategy for Acute Lung Injury.

Abstract

Sepsis-induced acute lung injury (ALI) has a high mortality rate, and cytokine storm is its feature. PANoptosis is a new type of cell death including apoptosis, pyroptosis and necroptosis. The aim of this study is to detect the PANoptosis level of lung macrophages, and to elucidate the new mechanism of sulforaphane (SFN) in sepsis-induced ALI. In septic animal model, the fluorescent staining of Caspase-8, GSDMD and p-MLKL and ASC/Caspase-8/RIPK3 PANoptosome in lung macrophages was performed. Lipopolysaccharide (LPS) was used to induce macrophages to construct cell model of sepsis. The proportion of dead cells was detected by PI staining, and the expression of Bax, GSDMD-N, NLRP3 and p-MLKL was detected by western blotting. Search for the target genes of SFN and sepsis by network pharmacology. Molecular docking analysis confirmed the binding between SFN and TLR4. The protein levels of TLR4, P65 and p-P65 were detected by western blotting. The transcriptional levels of inflammatory factors were detected by qPCR. The expression of Caspase-8, GSDMD, p-MLKL and PANoptosome in septic lung macrophages was significantly increased, suggesting PANoptosis was up-regulated. LPS induced macrophages death and increased protein levels of Bax, GSDMD-N, NLRP3 and p-MLKL, which were reversed by pretreatment with SFN. Network pharmacology and molecular docking demonstrated that SFN could bind to TLR4 and inhibit NFκB pathway. The mRNA levels of pro-inflammatory factors IL6, CXCL16, iNOS and IL18 were down-regulated by SFN. SFN might alleviate LPS-induced macrophage PANoptosis through TLR4/NFκB pathway, thereby inhibiting macrophage inflammation and becoming a potential therapeutic drug for sepsis-induced ALI.