β-Hydroxybutyrate induces microglial M2 polarization by inhibiting the NF-κB/NLRP3 pathway to ameliorate cerebral ischemia–reperfusion injury

Abstract

Objective To investigate the neuroprotective effects of β-hydroxybutyrate (BHB) on cerebral ischemia–reperfusion injury (CIRI) and its underlying molecular mechanisms; to clarify the association between ketone bodies and the severity of CIRI under obese conditions. Methods CIRI rat model was established using modified middle cerebral artery occlusion/reperfusion (MCAO/R), while obesity was induced using high fat diet. BHB was administered via subcutaneous osmotic pumps. Lipopolysaccharide (LPS) was intraperitoneally injected to forcibly activate the nuclear factor-κB (NF-κB)/NOD-like receptor protein 3 (NLRP3) pathway. Neurological deficits were evaluated using the Zea–Longa 5-point scoring system. Serum BHB levels were measured by colorimetric assay. Immunofluorescence was performed to detect CD86 (M1 phenotype) and CD206 (M2 phenotype) levels. Inflammatory cytokines in brain tissues were measured. Levels of phosphorylated p65 (p-p65), cleaved caspase-1, and ionized calcium-binding adapter molecule 1 (Iba1) were analyzed by Western blotting. Results CIRI rats with obesity exhibited increased serum BHB levels, reduced neurological deficit scores, and markedly decreased infarct volumes. Moreover, levels of p-p65, cleaved caspase-1, Iba1, and pro-inflammatory cytokines, were decreased, while Iba1+CD206+ co-localization was increased. BHB treatment for CIRI rats with normal weight reduced neurological deficit scores, increased serum BHB, and decreased infarct volume. Following further LPS administration, neurological deficit scores and infarct volume were enhanced, with no difference in serum BHB levels. Conclusion Elevated endogenous BHB under obese conditions exerts neuroprotective effects against CIRI, and exogenous BHB can mimic this effect. Mechanistically, BHB inhibits activation of the NF-κB/NLRP3 pathway and promotes microglial polarization toward the anti-inflammatory M2 phenotype, thereby alleviating CIRI.