TRPV4/IP3R-1-mediated endothelial pyroptosis drives pulmonary microvascular endothelial permeability in endotoxin-induced acute lung injury.

BACKGROUND

Sepsis-induced acute lung injury (ALI) is characterized by edema resulting from increased vascular permeability. Transient receptor potential vanilloid 4 (TRPV4) interacts with inositol 1,4,5-trisphosphate receptor type 1 (IP3R-1) through calmodulin-binding domains and regulates vascular permeability. However, the specific mechanisms underlying the roles of TRPV4 and IP3R-1 in endothelial pyroptosis and vascular permeability remain unclear.

METHODS

EMPs were measured in septic patients and controls, and co-cultured with human pulmonary microvascular endothelial cells (HPMECs). LPS-induced ALI was assessed in wild-type or Gsdmd-/- mice with pharmacological modulation of TRPV4, as well as in Trpv4-/- mice. Pyroptosis was detected in IP3R-1-silenced HPMECs treated with LPS and TRPV4 agonist. IP3R-1/GSDMD interaction was confirmed by structural prediction, Co-IP, and immunofluorescence. Intracellular Ca2+ was detected by flow cytometry, and tissue Ca2+ was visualized by in vivo imaging.

RESULTS

Elevated EMPs in septic patients enhanced endothelial permeability, upregulated TRPV4 and GSDMD-NT generation in HPMECs. TRPV4 promotes HPMECs permeability and pyroptosis in an IP3R-1-dependent manner. TRPV4 inhibition attenuated lung injury and ameliorated pulmonary dysfunction, while Trpv4 knockdown improved survival. Knockdown of IP3R-1 reduced intracellular Ca2+ and mtDNA levels in HPMECs. In vivo imaging revealed attenuated Ca2+ accumulation in Trpv4-/- mice. The interaction between TRPV4/IP3R-1 and GSDMD was Ca2+-dependent.

CONCLUSIONS

TRPV4 disrupts HPMEC integrity via IP3R-1 and triggers GSDMD-mediated pyroptosis through a Ca2+-dependent mechanism, exacerbating LPS-induced ALI.