Abstract WP284: Ifi27l2a acts as a cytokine that promotes a pro-inflammatory phenotype in microglia and enhances neuroinflammation

Interferon alpha-inducible protein 27 like 2A (Ifi27l2a) was originally identified as an interferon-stimulated gene (ISG) involved in host defense mechanisms during viral invasion. We recently showed that intracellular Ifi27l2a also plays a critical role in regulating microglial activity and promoting the spread of neuroinflammation. However, the functions of extracellular Ifi27l2a in inflammatory conditions are presently undefined. We now hypothesize that released Ifi27l2a protein acts as a novel cytokine and contributes to the intercellular spread of neuroinflammation. Methods: Circulating Ifi27l2a protein was measured in plasma of aged mice following experimentally-induced endotoxemia (systemic administration of lipopolysaccharide; LPS). Sim-A9 cells and primary microglia were treated with recombinant Ifi27l2a (rIfi27l2a) to assess its cytokine-like effects on mitochondrial function, microglial phenotype, and metabolic reprogramming. RNA-seq and targeted metabolomic analysis were performed to characterize transcriptional and metabolic changes following rIfi27l2a exposure. Results: LPS administration significantly increased plasma Ifi27l2a levels in aged mice (3.2-fold induction; n=4–5, p<0.05). Elevated Ifi27l2a levels were also detected in the culture media of stimulated primary microglia (n=8, p<0.01). In human plasma samples from male stroke patients (n=11), IFI27L2 levels gradually increased over a 7-day period following stroke onset, compared to the initial 24-hour time point (p<0.01). Treatment of microglia with rIfi27l2a led to significant activation of the inflammasome, as indicated by increased Caspase-1 activity (n=5, p<0.05). rIfi27l2a also enhanced ATP production and induced a metabolic shift toward glycolysis, as measured by Seahorse cell analysis (n=10-21, p<0.05). Expression of inflammatory marker genes ( Il1b , Tnfa ) were significantly upregulated in rIfi27l2a-treated microglia. RNA-seq (n=5) revealed robust induction of proinflammatory genes, including members of the Mmp family. Furthermore, levels of itaconate, a metabolite produced by Acod1, were significantly elevated in rIfi27l2a-treated microglia (n=5, p<0.001). Conclusions: Extracellular Ifi27l2a acts as a novel cytokine that promotes microglial polarization toward a proinflammatory phenotype. Targeting Ifi27l2a-mediated signaling may offer a promising therapeutic approach to attenuate neuroinflammation and enhance functional recovery following stroke.