Abstract P727: Single-Cell RNA Sequencing Identifies Ifi27l2a as a Novel Gene Involved in Regulating Microglial Inflammation in the Context of Aging and Stroke

Abstract

Microglial cells (MG) serve as resident immune cells in the brain and play a critical role in the acute response and chronic recovery to stroke. However, how the transcriptional signature of these cells is altered in aging and stroke remains poorly defined. Here, we compared the MG transcriptomic response to stroke in young and aged mice by single-cell RNA sequencing (scRNAseq). Our unbiased approach identified Ifi27l2a (a member of the interferon signaling family) as one of the most highly upregulated genes in MG with aging or ischemic stroke. Further analyses suggested that this gene may be a novel regulator of MG inflammation in aging and stroke. Methods: Permanent distal middle cerebral artery occlusion (pdMCAO) or sham surgery was performed in young and aged mice of both sexes (3 & 20 months). scRNAseq was performed to explore the transcriptional signature in brains at 14-days post-stroke and sham controls. We used qRT-PCR to validate our scRNAseq findings and compare regional transcriptional changes. Human microglial cells (HMC3; subjected to OGD) and human autopsy brain sections (stroke versus non-stroke) were used to examine IFI27l2 protein expression. Results: Using comprehensive scRNAseq analysis, we identified Ifi27l2a as the most highly up-regulated gene in aged MG following stroke. Further analysis showed a positive correlation between Ifi27l2a and multiple “MG activation” genes. In contrast, a negative correlation was shown with genes involved in promoting the phagocytic MG phenotype (e.g. Spp1, Cst7, Lpl, and Itgax). Our qRT-PCR results validated the central scRNAseq findings and further showed that Ifi27l2a was significantly up-regulated in thalamus of aged brains (p = 0.0412) and trending in cortex (p = 0.23) (n=4 young/aged). Following stroke, Ifi27l2a and other markers of MG activation were significantly increased in both brain regions (n= 4-6). In human tissues, we observed an increase in IFI27l2 protein in HMC3 cells with OGD and in brain of stroke patients. Conclusions: We identified Ifi272la as a highly up-regulated gene in MG in aging and following stroke. Our findings suggest that Ifi27l2a may be a novel regulator of neuroinflammation in MG, and thus provide the basis for a new therapeutic target to reduce inflammation in aging and stroke.