Background: Limb ischemia-reperfusion-induced acute lung injury (LIR-ALI) remains without effective therapies, with macrophage polarization playing a pivotal role in its pathology. This study explored the protective effects of esketamine in LIR-ALI, focusing on its modulation of macrophage polarization through the TLR4/NF-κB/NLRP3 pathway.

Methods: Sprague-Dawley rats subjected to LIR (bilateral femoral artery clamping for 2 hours followed by reperfusion) under mechanical ventilation were treated with esketamine (5 mg/kg) or saline. Lung injury, inflammatory cytokines (Enzyme-linked immunosorbent assay, ELISA), and macrophage polarization markers (iNOS, Arg-1) were evaluated. In vitro, THP-1 macrophages were pretreated with esketamine (50 μM), stimulated with LIR-ALI rat serum, and co-cultured with BEAS-2B epithelial cells. Macrophage polarization (NOS2, Arg-1, CD86, CD206), epithelial cell viability (CCK-8), inflammatory cytokines, oxidative stress markers (superoxide dismutase (SOD), malondialdehyde (MDA)), and TLR4/NF-κB/NLRP3 pathway proteins were assessed. LPS was used to assess pathway dependency.

Results: Esketamine significantly reduced LIR-induced lung pathological damage and inflammatory cell infiltration in rats. It lowered pro-inflammatory cytokines (IL-1β, IL-6, TNF-α) and elevated IL-10 in bronchoalveolar lavage fluid (BALF) (p < 0.01), while promoting M2 macrophage polarization (decreased iNOS, increased Arg-1) in lung tissue (p < 0.05). In vitro, esketamine redirected LIR-ALI serum-stimulated macrophages toward the M2 phenotype (decreased NOS2/CD86, increased Arg-1/CD206) (p < 0.001), enhanced BEAS-2B cell viability, and mitigated inflammation and oxidative stress (p < 0.01). These effects were linked to esketamine's inhibition of the TLR4/NF-κB/NLRP3 pathway in macrophages (p < 0.01). LPS reversed esketamine's protective effects, including its modulation of macrophage polarization and the signaling pathway (p < 0.05).

Conclusion: Esketamine mitigates LIR-ALI by promoting M2 macrophage polarization through the suppression of the TLR4/NF-κB/NLRP3 axis, suggesting its potential as a therapeutic strategy for LIR-ALI.