Mn₃O₄ nanozyme-based anti-inflammatory therapy modulates microglial phenotype by downregulating TLR4/NOX2 expression and further alleviates Alzheimer's disease pathology

<b>Rationale:</b> Evidence shows that neuroinflammation mediated by microglial activation plays an important role in Alzheimer's disease (AD) pathogenesis. However, the relationship between microglial phenotype and fibrillar β-amyloid (fAβ) pathology in anti-inflammatory treatment of AD remains unclear. <b>Methods:</b> We designed a water-soluble Mn₃O₄ nanozymes and demonstrated its ability to reverse lipopolysaccharide (LPS)-induced microglial transition from M1 to M2 phenotype by clearing reactive oxygen species (ROS). <b>Results:</b> In 5×FAD transgenic mice, intranasal (IN) instillation of Mn₃O₄ nanozymes initially promoted M2 microglial polarization and significantly reduced neuroinflammation after 4 weeks of treatment. After 8 weeks of continuous treatment, they further alleviate fAβ pathology and improved learning and memory deficits in 5×FAD mice. The excellent anti-inflammatory effect of Mn₃O₄ nanozymes is achieved by inhibiting the Toll-like receptor 4 (TLR4)/nicotinamide adenine dinucleotide phosphate (NAPDH) oxidase isoform 2 (NOX2) pathway to clear ROS. <b>Conclusions:</b> This study reveals the molecular mechanism of Mn₃O₄ nanozymes modulating microglia phenotype to attenuate neuroinflammation primarily through inhibition of the TLR4/NOX2 pathway and highlights the temporal sequence of anti-inflammatory treatment in regulating microglial phenotype and improving fAβ pathology, providing new insights for the anti-inflammatory treatment of AD and other neurological diseases.