Mdivi‐1 Attenuates Sepsis‐Associated Acute Lung Injury by Inhibiting M1 Alveolar Macrophage Polarization and Pyroptosis

<b>Background:</b> Dynamin-related protein 1 (DRP1)-dependent mitochondrial fission is a novel target for mitigating inflammatory diseases. This study aims to explore the effects of the DRP1 inhibitor Mdivi-1 on sepsis-induced acute lung injury (ALI). <b>Methods:</b> C57BL/6 mice were intraperitoneally injected with lipopolysaccharide (LPS) and then treated with or without Mdivi-1 2 h post-injection. RAW264.7 alveolar macrophages were stimulated with LPS and treated with or without NLRP3 inhibitors, Mito-TEMPO, or Mdivi-1. Hematoxylin and eosin (H&E) staining was used to observe pathological changes in lung tissues. The levels of inflammatory cytokines in lung tissue homogenates, serum, and cell culture medium were detected using enzyme-linked immunosorbent assays (ELISA). The mRNA expression of macrophage polarization markers, NLRP3 activation, and phosphorylation status of DRP1 were assessed. Flow cytometry was employed to evaluate the levels of macrophage apoptosis. Immunofluorescence was utilized to detect the levels of in vivo and in vitro macrophage polarization markers. Mitochondrial reactive oxygen species (Mito-ROS) were measured using a Mito-SOX assay kit. <b>Results:</b> Our results suggested that Mdivi-1 reduced lung tissue pathological injury, M1 alveolar macrophage polarization, NLRP3 activation, and DRP1 Ser616 phosphorylation. In vitro, LPS triggered abnormal accumulation of M1 polarization, NLRP3 activation, and excessive increase in Mito-ROS. NLRP3 inhibitors and Mito-TEMPO inhibited M1 alveolar macrophage polarization and pyroptosis-mediated tissue damage. Mito-TEMPO significantly inhibited NLRP3 activation. Furthermore, Mdivi-1 reduced ALI by inhibiting M1 polarization and pyroptosis. The mechanism of Mdivi-1 in reducing M1 alveolar macrophage polarization and pyroptosis may be related to the inhibition of DRP1-mediated mitochondrial fission, thus suppressing the Mito-ROS/NLRP3 pathway. Similar results were observed <i>in vitro</i> by knocking down DRP1. <b>Conclusion:</b> Inhibition of DRP1 by Mdivi-1 alleviates ALI by hindering Mito-ROS/NLRP3-mediated M1 alveolar macrophage polarization and pyroptosis, suggesting that DRP1-dependent mitochondrial fission is a potential therapeutic target for ALI.