Nitroxoline attenuates sepsis-associated pancreatic injury through inhibiting NLRP3-inflammasome mediated pyroptosis

Pancreatic injury constitutes a prevalent and clinically critical complication in the setting of sepsis, typified by both an increased occurrence and a substantial mortality risk. This study investigated the mechanism by which nitroxoline (NTX), a novel NLRP3 inflammasome inhibitor, alleviates pancreatic injury in septic mice through modulating classical pyroptosis pathways.

Sepsis-associated pancreatic injury (SPI) was induced in experimental animals by performing cecal ligation and puncture procedures. For in vitro mechanistic studies, primary pancreatic acinar cells and macrophages were harvested from pancreatic tissues and maintained in culture. Histopathological alterations indicative of pancreatic injury were evaluated through HE staining. Pyroptosis in pancreatic cells and macrophages was identified using Hoechst33342/PI dual staining. IHC analysis was employed to detect Caspase-1 expression in pancreatic tissue sections, whereas IF staining was utilized to measure Caspase-1 levels in isolated pancreatic acinar cells. ELISA was conducted to quantify the serum levels of inflammatory cytokines including TNF-α, IL-6, IL-1β and IL-18. Western blotting was performed to examine the protein abundance of NLRP3, Caspase-1 p20 and GSDMD-N.

NTX treatment relieved pancreatic tissue lesions in SPI-model mice, accompanied by a significant decrease in injury scores and the extent of necrotic regions. The protective effect conferred by high-dose NTX was superior to that achieved in the group receiving VX-765. Within the SPI group, NTX prominently downregulated the expression of NLRP3, Caspase-1 p20 and GSDMD-N, and high-dose NTX displayed a more favorable inhibitory action in comparison to VX-765. Exposure to gradient concentrations of NTX substantially ameliorated LPS-induced pyroptotic events and decreased the ratio of pyroptotic pancreatic acinar cells. Following LPS challenge, the levels of pyroptosis activation-related proteins NLRP3, Caspase-1 p20 and GSDMD-N were markedly increased in pancreatic acinar cells, and these increases were abrogated upon NTX administration. Additionally, NTX significantly diminished pyroptosis in macrophages and the expression of pyroptosis-associated proteins in the SPI model.

The protective efficacy of NTX in the context of SPI was achieved via the regulation of pathological pyroptotic activation in both pancreatic acinar cells and macrophages, with the NLRP3-Caspase-1-GSDMD pyroptotic pathway serving as the key underlying molecular mechanism.