TREM1/DAP12-mediated neuron-microglia crosstalk in ischemic stroke: Huangxiong formula reduces Neuroinflammation and neuronal Pyroptosis.

BACKGROUND: The inflammatory cascade drives the progression of ischemic stroke (IS), with neuron-microglia interactions playing a critical role in maintaining brain homeostasis. Huangxiong Formula (HXF) demonstrates clinical efficacy in IS; however, its precise neuroprotective mechanism remains unclear. This study investigates whether HXF mitigates neuroinflammation by modulating microglial polarization and neuronal pyroptosis. METHODS: In vivo, HXF was administered to mice in a middle cerebral artery occlusion/reperfusion (MCAO/R) model. In vitro, a TREM1-overexpressing BV2 microglial cell line was established, and HT22 neurons were treated with conditioned medium derived from LPS-stimulated microglia. HXF-containing cerebrospinal fluid (HXF-CSF) was used for microglial treatment. RESULTS: HXF significantly improved neurological scores, enhanced cerebral perfusion, and reduced infarct volume in MCAO/R mice. Mechanistically, HXF shifted microglial polarization from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype and inhibited neuronal pyroptosis, as evidenced by reduced levels of IL-18, IL-1β, and cleaved caspase-1, along with suppression of the TREM1/DAP12 pathway. In vitro, HXF-CSF directly suppressed microglial M1 polarization and cytokine release. Conditioned medium derived from HXF-CSF-treated microglia markedly reduced pyroptosis in HT22 neurons. TREM1 overexpression in microglia abolished these protective effects. CONCLUSION: HXF alleviates neuroinflammation and neuronal pyroptosis by inhibiting microglial M1 polarization through the TREM1/DAP12 pathway. Microglial TREM1 represents a key therapeutic target, supporting the clinical application of HXF for ischemic stroke.