Mechanism of Hypoxia-Induced HMGB1 Regulating NLRP3 Inflammasome/Caspase-1 Pathway-Mediated Pyroptosis in Myocardial Ischemia Reperfusion Injury Through the Nrf2/HO-1 Pathway

Objective: Myocardial ischemia-reperfusion injury (MIRI) represents an inevitable risk event for acute myocardial infarction. We explored the mechanism of hypoxia-induced high-mobility group box 1 (HMGB1) promoting MIRI by modulating the NLRP3 inflammasome/Caspase-1 pathway-mediated Pyroptosis via the Nrf2/HO-1 pathway.

Methods: In vitro cultured mouse cardiomyocytes were exposed to hypoxia/reoxygenation (H/R) to establish an MIRI cell model, then treated with short hairpin-HMGB1, a NLRP3 Agonist (Nigericin), and a Nrf2 inhibitor (ML385). Cell viability and injury were assessed via MTT and LDH assays. HMGB1 (nuclear/cytoplasm), Nrf2 (nuclear/cytoplasm), HO-1, NLRP3, ASC, cleaved Caspase-1, and GSDMD-N protein levels, and IL-1β and IL-18 levels in cell supernatants were determined by western blot and ELISA. HMGB1 and Nrf2 distribution were analyzed by immunofluorescence, with their interaction verified by co-immunoprecipitation. An MIRI mouse model was developed and treated with HMGB1 Box A for in vivo verification.

Results: H/R induction declined the nuclear HMGB1 protein level and cell viability, and intensified the cytoplasmic HMGB1 protein level, cell damage, and pyroptosis-related protein and inflammatory cytokine levels, which were averted by HMGB1 knockdown. NLRP3 activation partially reversed HMGB1 knockdown's effect on improving cardiomyocyte Pyroptosis. Hypoxia-induced HMGB1 inhibited Nrf2/HO-1 activation by interacting with Nrf2. Nrf2/HO-1 suppression partly counteracted HMGB1 knockdown's suppressive effects on NLRP3 inflammasome activation and Pyroptosis. HMGB1 suppressed the Nrf2/HO-1 axis to enhance NLRP3 inflammasome/Caspase-1 pathway-mediated Pyroptosis, thereby exacerbating MIRI in vivo.

Conclusion: Hypoxia induces HMGB1's nucleus-to-cytoplasm translocation, which binds to Nrf2 to repress Nrf2 nuclear translocation to suppress Nrf2/HO-1 activation to promote NLRP3 inflammasome/Caspase-1-mediated Pyroptosis, thereby exacerbating MIRI.

Caspase‐1; NLRP3 inflammasome; Pyroptosis; heme oxygenase‐1; high mobility group box 1 protein; myocardial ischemia–reperfusion injury; nuclear factor erythroid 2‐related factor 2; nuclear translocation.