Inhibiting apoptosis and GSDME-mediated pyroptosis attenuates hepatic injury in septic mice.

ABSTRACT

BACKGROUND: Sepsis is characterized by severe inflammation and organ dysfunction resulting from a dysregulated organismal response to infection. Although pyroptosis has been presumably shown to be a major cause of multiple organ failure and septic death, whether gasdermin E (GSDME)-mediated pyroptosis occurs in septic liver injury and whether inhibiting apoptosis and GSDME-mediated pyroptosis can attenuate septic liver injury remain unclear. This study investigated the role of apoptosis and GSDME-mediated pyroptosis in septic liver injury. METHODS: Adult male C57BL/6 mice were randomly divided into four groups sham, cecal ligation puncture (CLP), CLP + Z-DEVD-FMK (a caspase-3 inhibitor, 5 mg/kg), and CLP + Ac-DMLD-CMK (a GSDME inhibitor, 5 mg/kg). Sepsis severity was assessed using the murine sepsis score (MSS). Hepatic tissue damage was observed by the hematoxylin-eosin staining method, the activities of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST), the levels of malondialdehyde (MDA), the concentrations of interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) were measured according to the related kits, and the changes in the hepatic tissue reactive oxygen species (ROS) levels were detected by immunofluorescence (IF). The protein expression levels of cleaved caspase-3, GSDME-N, IL-1ß, B-cell lymphoma-2 (Bcl-2), cytochrome C (Cyt-c), and acetaldehyde dehydrogenase 2 (ALDH2) were detected using western blotting. GSDME expression was detected by immunohistochemistry. RESULTS: Compared with the Sham group, CLP mice showed high sepsis scores and obvious liver damage. However, in the CLP + Z-DEVD-FMK and CLP + Ac-DMLD-CMK groups, the sepsis scores were reduced and liver injury was alleviated. Compared with the Sham group, the serum ALT and AST activities, MDA and ROS levels, and IL-1ß and TNF-α concentrations were increased in the CLP group, as well as the protein expression of cleaved caspase-3, GSDME-N, IL-1ß, Cyt-c, and GSDME positive cells (P < 0.05). However, the expression levels of Bcl-2 and ALDH2 protein were decreased (P < 0.05). Compared with the CLP group, the CLP + Z-DEVD-FMK and CLP + Ac-DMLD-CMK groups showed low sepsis scores, ALT and AST activities, MDA and ROS levels, decreased IL-1ß and TNF-α concentrations, and decreased expression of cleaved caspase-3, GSDME-N, IL-1ß protein expression, and GSDME positive cells (P < 0.05). The expression levels of Bcl-2 and ALDH2 protein were increased (P < 0.05). CONCLUSION: Apoptosis and GSDME-mediated pyroptosis are involved in the development of sepsis-induced hepatic injury. Inhibition of apoptosis and GSDME-mediated pyroptosis attenuates injury. ALDH2 plays a protective role by inhibiting apoptosis and pyroptosis.