Mechanisms of Microglial NLRP3 Inflammasome Activation by Methylmalonic Acid in Methylmalonic Acidemia

Background Microglial activation is a critical pathogenic mechanism of neurological damage in methylmalonic acidemia (MMA). The NOD‐like receptor pyrin domain containing 3 (NLRP3) inflammasome is closely associated with neuroinflammatory processes. However, the mechanism of NLRP3 inflammasome activation in microglia by methylmalonic acid (MMAcid) remains unclear. Methods Mouse microglia BV2 cells and hippocampal neuronal HT22 cells were stimulated with different concentrations of MMAcid. Neuronal cells were treated with conditioned medium derived from differently treated microglia. Microscopy was used to observe morphological changes of cells. Cell counting kit‐8 (CCK‐8) assay was performed to assess cell viability (VB). In addition, the mRNA and protein levels were detected via qRT‐PCR, ELISA, and western blot. To further validate the mechanism, we used lentivirus infection and small interfering RNA (siRNA) transfection to overexpress (OE) and knockdown (KD) extracellular signal–regulated kinase (ERK) and analyzed the inflammatory pathway. Results Direct exposure of neuronal cells to MMAcid significantly reduced cell VB, accompanied by morphological features suggestive of pyroptosis. MMAcid activated microglia, which was accompanied by increased expression of inflammatory molecules, activation of the ERK/nuclear factor‐κB (NF‐κB)/NLRP3 signaling pathway, and elevated secretion of interleukin‐1 beta (IL‐1β) and tumor necrosis factor‐α (TNF‐α). Neuronal cell VB decreased when cocultured with the supernatant from MMAcid‐treated microglia. Overexpression of ERK further enhanced microglial activation and potentiated MMA‐induced neuronal pyroptosis‐related signaling. Conversely, knocking down ERK expression in microglia reduced their activation by MMAcid and decreased cytokine secretion, but it did not completely reverse MMA‐induced neuronal damage. Conclusion MMAcid is associated with microglial activation, accompanied by alterations in the ERK/NF‐κB/NLRP3 signaling pathway and increased release of inflammatory cytokines, which may contribute to neuronal pyroptosis–related changes.