Microglial TIA1-mediated stress granules promote neuroinflammation and aggravate neuron loss in mice after ischemic stroke by inhibiting IGF2 signaling.

Rationale: Microglia cells as niche homeostasis monitor with rapid responses to acute ischemic stroke (IS). T-cell intracellular antigen 1 (TIA1), a core component of stress granules (SGs), is involved in cellular stress responses such as hypoxia, but its roles and mechanisms in regulating microglial responses during IS remain unclear. Methods: To evaluate the function of microglial TIA1 in IS, we established a mouse model of IS by using photothrombotic method. Furthermore, conditional knockout (CKO) of Tia1 in microglia mice (Tia1 (Cx3cr1)-CKO mice) was generated and then Tia1 (Cx3cr1)-CKO IS mice and their littermate controls (Tia1 (f/f) IS mice) were used as experimental subjects. The behavioral tests, immunostaining, Laser speckle contrast imaging (LSCI), TTC staining, Nissl staining, quantitative real-time PCR (qPCR) and Western blotting were used to assess the effects of microglial Tia1 deletion in IS progression. In vitro, we utilized the microglia cell line (HMC3 cells) and primary cultured microglia to establish an OGD model, and generated stable TIA1-knockdown or TIA1-overexpressing HMC3 cell lines, and employed a co-culture system of HMC3 and N2a cells to further explore the roles of microglial Tia1 signaling in IS. Through RNA sequencing (RNA-seq) of control HMC3 cells and Tia1-knockdown HMC3 cells, we investigated in depth the role and molecular mechanism of TIA1-mediated insulin-like growth factor 2 (IGF2) signaling pathway in microglia during IS progression. Results: Microglial TIA1 was significantly upregulated in mice during the acute phase of IS. Microglial Tia1 knockout suppressed microglial pro-inflammatory responses, enhanced anti-inflammatory responses, promoted phagocytic clearance of infarct debris, alleviated neuronal death, and improved motor deficits in post-IS mice. In vitro, TIA1 promoted pro-inflammatory responses to exacerbate neuronal cell death and inhibited phagocytic ability of microglia cells after OGD. Mechanistically, Tia1 deletion in microglia impaired SG formation, reduced sequestration of Igf2 mRNA into SGs, upregulated IGF2 expression, and IGF2 signaling enhanced anti-inflammatory responses and phagocytic capacity while suppressing pro-inflammatory activation in microglia. Conclusions: These findings identify a previously unrecognized function of microglial TIA1 in modulating microglia homeostasis and sustaining pro-inflammatory responses via SGs-mediated Igf2 mRNA sequestration after IS, providing a novel therapeutic target for IS treatment.