Uncovering a new player in ischemic stroke: a study of intra-arterial interferon-gamma-producing monocytes in hyperacute stroke.

Stroke triggers a rapid and complex immune response that is not yet fully understood, especially within hours after an ischemic infarct. Our previous study in stroke patients revealed a significant increase in interferon-gamma (IFN-γ) immediately (hyperacute) and downstream of the ischemic ictus, within the arterial compartment. The present study investigated the source, inciting factors, and role of IFN-γ in a preclinical murine model. Stroke was produced using transient middle cerebral artery occlusion, and immune cells within the arterial vasculature distal to the occlusion (pre- and post-occlusion) were characterized using flow cytometry. Compared with the control samples, the post-occlusion samples presented an increase in IFN-γ and CD69 cells, whereas no significant increase was detected in IL17, IL4, and CD25 cells. Further analysis of the IFN-γ population revealed two novel attributes. First, interrogation of the identity of these IFN-γ cells revealed that the increase in IFN-γ production was largely driven by CD14 cells in the post-occlusion sample, with negligible contributions from other canonical IFN-γ-producing cells (CD4, CD8). Second, the IFN-γ cells exhibited two distinct clusters, an IFN-γ and an IFN-γ population. Further analysis revealed that the IFN-γ population was largely composed of CD14 cells, whereas the IFN-γ population was dominated by CD4 T-cells. To explore the conditions driving IFN-γ production, an ischemia model involving oxygen-glucose deprivation (OGD) was employed. Co-culturing of naïve splenocytes with OGD-treated CNS cells and OGD-derived supernatant resulted in a significant increase in IFN-γCD14 cells, as compared to normoxic controls, an effect that coincided with marked loss of DAPI and NeuNDAPI cells in mixed cortical (neuronal and glial) cultures. In summary, this study identified intra-arterial CD14 monocytes as novel early sources of IFN-γ in the hyperacute phase of stroke, a role traditionally attributed to adaptive immune cells. Using and ischemia models, the findings reveal that injury-associated signals from CNS cells are sufficient to directly induce IFN-γ production in CD14 cells, redefining early stroke immunopathology and uncovering a potential target for timely immunomodulation.