Dissecting functional phenotypes of microglia and macrophages in the rat brain after transient cerebral ischemia

Abstract

Ischemic brain injury causes local inflammation, which involves activation of resident microglia, leukocyte, and monocyte infiltration. Involvement of peripheral immune cells in ischemia‐induced damage and repair is debatable. Using flow cytometry, gene expression profiling, and immunocytochemistry, we show that microglia predominate in the ischemic brain and express inflammation mediators at Day 1 after transient middle cerebral artery occlusion (MCAo) in rats. At Day 3, both resident microglia and bone marrow (BM)‐derived macrophages are detected in the ischemic hemispheres and display unique transcriptomic profiles. Functional groups enriched in BM‐macrophages are indicative of the pro‐regenerative, immunosuppressive phenotype. Transient depletion of peripheral macrophages with clodronate‐filled liposomes reduced the number of Arg1+ Iba1+ expressing cells in the ischemic brain. The analysis of microglia and macrophage signature genes shows that each cell type maintains the expression of their identity genes, even if gene expression is modified in a response to environmental clues. At Day 7, infiltrating BM‐macrophages exhibit the reduced expression of Arg1, the elevated expression of iNos and many inflammatory genes, as shown by RNA sequencing. This is consistent with their switch toward a pro‐inflammatory phenotype. We propose that BM‐macrophages recruited to the injured brain early after ischemia could contribute to functional recovery after stroke, but they switch toward a pro‐inflammatory phenotype in the ischemic parenchyma. Our results point to the detrimental role of microglia in an ischemic brain and the primarily pro‐regenerative role of infiltrating BM‐macrophages.

Protective Microglial Subset in Development, Aging, and Disease: Lessons From Transcriptomic Studies

“…Investigation of the genes enriched in microglia three days after middle cerebral artery occlusion compared to sham controls, however, revealed Spp1, Gpnmb, Lgals3, Fabp5, and Axl among others, fitting with the potential presence of CD11c+ microglia-like cells in this context, diluted among other microglia. Consistent with this, Ccl2 mRNA was found to be increased in microglia and macrophages at this time point (132), an aspect that has been associated with the emergence of CD11c+ microglia (81). Another study, conducted in a model of phototrombic stroke on whole tissue, actually showed upregulation of Gpnmb, Itgax, and Clec7a in a cluster associated with early response (133), which the authors related to the DAM phenotype (74).…”

“…Studying microglia in context of inflammation can get quite complicated due to massive infiltration of peripheral immune cells, notably monocytes and macrophages, occurring subsequently to TBI (128), SCI (129), and stroke (127,130,131). In a study comparing the transcriptomics of microglia and macrophages after ischemia in rats, it was reported that microglia played a detrimental role and macrophages played a beneficial role with regard to recovery, based on their expression of classical inflammation markers (132). Investigation of the genes enriched in microglia three days after middle cerebral artery occlusion compared to sham controls, however, revealed Spp1, Gpnmb, Lgals3, Fabp5, and Axl among others, fitting with the potential presence of CD11c+ microglia-like cells in this context, diluted among other microglia.…”

FTY720 Exerts Anti-Glioma Effects by Regulating the Glioma Microenvironment Through Increased CXCR4 Internalization by Glioma-Associated Microglia

“…Rat primary microglial cells were generated from 1-day-old postnatal Sprague-Dawley rats as described previously (24). Briefly, microglia were isolated from cerebral cortices by mechanical dissociation, 0.25% trypsin/EDTA (Gibco, Grand Island, NY, USA), and plated into poly-D-lysine-coated (0.1 mg/ml; Sigma Chemical, St. Louis, MO, USA) T25 culture flasks.…”

Crosstalk between stressed brain cells: direct and indirect effects of ischemia and aglycemia on microglia

“…Increasing evidence suggests that microglia may even adapt disease-specific phenotypes, e.g., in Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, multiple sclerosis, and traumatic brain injury [5][6][7]. After a stroke, microglia develop a specific phenotype that can be distinguished from macrophages [8][9][10]. In post-ischemic inflammation, microglia have both beneficial and detrimental effects.…”