Abstract / Summary
Abstract Microglia play dual roles in brain injury and repair after ischemic stroke. Polarizing microglia toward a pro-reparative, anti-inflammatory phenotype represents a potential strategy to mitigate ischemic damage, but the relationships among hypercapnia, microglial phenotypic changes, and post-ischemic pro-angiogenic signaling remain incompletely defined. In this study, we used a mouse transient middle cerebral artery occlusion (tMCAO) model and administered 8% CO₂ both before ischemia induction and after reperfusion to assess its effects on early neurological injury. We performed behavioral testing, Western blotting, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, chick chorioallantoic membrane (CAM) assay, and arterial blood gas analysis. Our results showed that hypercapnia reduced infarct volume and improved neurological outcomes within 72 h after reperfusion. It increased the expression of tight junction proteins (zonula occludens-1 [ZO-1], occludin) and angiogenic factors (brain-derived neurotrophic factor [BDNF], vascular endothelial growth factor A [VEGFA]), suppressed pro-inflammatory cytokines while elevating anti-inflammatory cytokines, and upregulated the pro-reparative microglial markers arginase-1 (Arg1) and triggering receptor expressed on myeloid cells 2 (TREM2) as well as mitochondrial aspartyl-tRNA synthetase (DARS2). Pearson correlation analysis confirmed a positive association between Arg1/TREM2 and VEGFA expression. These findings indicate that combined preconditioning and post-ischemic hypercapnia attenuates acute ischemic brain injury, which is associated with an anti-inflammatory phenotypic shift and enhanced pro-angiogenic signaling. The present data demonstrate association rather than causality; the causal roles of DARS2 and microglia-mediated angiogenesis require further investigation using cell-specific and functional approaches.