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Resident microglia and infiltrating macrophages inflammatory, metabolic and sex-dependent gene-regulatory dynamics in the neonatal hippocampus after hypoxia-ischemia.

ABSTRACT: Neonatal hypoxia-ischemia (HI) is a major cause of perinatal death and long-term disabilities worldwide. Post-ischemic neuroinflammation, characterized by resident and infiltrating immune cells, plays a pivotal role in HI pathophysiology. However, the specific modulatory roles of resident microglia and blood-derived macrophages remain poorly understood. In the present study, we investigated the temporal dynamics of microglia (CX3CR1GFP/+) and infiltrating macrophages (CCR2RFP/+) in the hippocampi of mice subjected to HI at postnatal day (PND) 9. We reveal an increase of CX3CR1GFP+ resident cells in the injured hippocampi soon after HI, peaking at three days post injury. In contrast, infiltration of CCR2RFP/+ cells peaked one day after HI and gradually decreased over time. Transcriptomic signatures of CX3CR1GFP/+ and CCR2RFP/+ cells isolated from hippocampi showed a partial convergence at day 3, through cell type - specific dynamics. Using inflammatory pathway and transcription factor analyses, we identify a distinct post-ischemic response in CCR2RFP/+ cells characterized by differential gene expression in sensome, homeostatic, matrisome, lipid metabolic and inflammatory molecular signatures as consequence of HI. Finally, we identified a sexual dimorphism in CX3CR1GFP/+ cells, where microglia unique genes in males but not in females returned to homeostatic levels comparable to controls three days after HI. In conclusion, our results indicate that CX3CR1GFP/+ and CCR2RFP/+ cells have diverse roles in orchestrating the inflammatory cascade after HI, and that infiltrating macrophages drive post-ischemic inflammation.

ORGANISM(S): Mus musculus

PROVIDER: GSE238220 | GEO | 2023/07/26

REPOSITORIES: GEO

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Project description:Objective: Brain tumors (gliomas) contain large populations of infiltrating macrophages and recruited microglia, which in experimental murine glioma models promote tumor formation and progression. Among the barriers to understanding the contributions of these stromal elements to high-grade glioma (glioblastoma; GBM) biology is the relative paucity of tools to characterize infiltrating macrophages and resident microglia. In this study, we leveraged multiple RNA analysis platforms to identify new monocyte markers relevant to GBM patient outcome. Methods: High-confidence lists of mouse resident microglia- and bone marrow-derived macrophage-specific transcripts were generated using converging RNA-seq and microarray technologies and validated using qRT-PCR and flow cytometry. Expression of select cell surface markers was analyzed in brain-infiltrating macrophages and resident microglia in an induced GBM mouse model, while allogeneic bone marrow transplantation was performed to trace the origins of infiltrating and resident macrophages. Glioma tissue microarrays were examined by immunohistochemistry, and the Gene Expression Omnibus (GEO) database was queried to determine the prognostic value of identified microglia biomarkers in human GBM. Results: We generated a unique catalog of differentially-expressed bone marrow-derived monocyte and resident microglia transcripts, and demonstrated that brain-infiltrating macrophages acquire F11R expression in GBM and following bone-marrow transplantation. Moreover, mononuclear cell F11R expression positively correlates with human high-grade glioma and additionally serves as a biomarker for GBM patient survival, regardless of GBM molecular subtype. Significance: These studies establish F11R as a novel monocyte prognostic marker for GBM critical for defining a subpopulation of stromal cells for future potential therapeutic intervention. Total RNA was isolated from three independently-generated sets of flow sorted bone marrow monocytes (CD11b+ CD45high CD115+ Ly6G- cells) and brainstem microglia (CD11b+ CD45low CD115low Ly6G- cells) for Illumina RNA-Seq, and two additional pools were subsequently generated and submitted for Affymetrix Mouse Exon 1.0ST microarray. Two of the RNA-Seq samples were additionally analyzed by the microarray, for a total of 6 samples (3 monocyte, 3 microglia) in each platform. Data outputs were analyzed by two analysis methods each (RNA-Seq data: ALEXA-Seq and Cufflinks; microarray data: Partek and Aroma). All four lists were merged into a new high-confidence gene list of transcripts that were significantly differentially expressed (DE) in three out of the four analyses. In this dataset, we include exon expression data obtained from flow sorted mouse bone marrow monocytes and brainstem microglia.

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Resident microglia and infiltrating macrophages inflammatory, metabolic and sex-dependent gene-regulatory dynamics in the neonatal hippocampus after hypoxia-ischemia.

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