Esketamine attenuated sepsis-induced organ injury in mice via the TGF- -Smad3 signaling pathway.

Sepsis remains a critical global healthcare challenge, with a high mortality rate of 30.8% among critical illnesses, second only to cancer. Currently, there is a lack of effective clinical treatments. The primary cause of early-stage mortality is organ damage and failure induced by an excessive inflammatory response. Macrophages, particularly their polarization states, are deeply involved in driving this pathological inflammatory cascade. This study aimed to investigate the effect of Esketamine (EK) on macrophage polarization and its subsequent role in ameliorating organ injury in a mouse model of sepsis induced by Escherichia coli. EK was investigated given its documented anti-inflammatory properties. In vitro, bone marrow-derived macrophages (BMDMs) were stimulated with lipopolysaccharide (LPS) in the presence or absence of EK. In vivo, a septic mouse model was established, with a cohort receiving EK treatment. The expression of macrophage polarization markers was assessed at both transcriptional and protein levels using quantitative polymerase chain reaction (qPCR) and western blotting, respectively. Concurrently, the impact of EK on injury to major organs in septic mice was evaluated. In vitro, EK (50-200 ng/mL) pretreatment of RAW264.7 and BMDM cells significantly suppressed LPS-induced M1 marker expression (TNF-α, IL-1 , IL-6, iNOS) and enhanced M2 marker expression (Arg-1, CD206, IL-10, Fizz1) at both mRNA and protein levels (P < 0.01). In vivo, C57BL/6 mice (n = 8/group) received EK (10 mg/kg, i.p.) 6 h prior to E. coli challenge. EK pretreatment markedly attenuated lung histopathological injury, reduced serum ALT/AST/lactate levels, and decreased TNF-α and IL-6 concentrations in serum and BALF (P < 0.01). Mechanistically, EK induced Smad3 phosphorylation, and blockade with SIS3 (2 mg/kg, i.p.) abolished both M2 polarization in vitro and organ protection in vivo. Our findings suggested that EK reprograms macrophages by shifting their polarization from the pro-inflammatory M1 phenotype toward the anti-inflammatory M2 phenotype. Therefore, EK may represent a potential therapeutic agent for sepsis and related inflammatory conditions not only due to its direct anti-inflammatory property but also because it can remodel the immune landscape by promoting the M2 macrophage phenotype.