Re-visit the Concept of M1 versus M2 Phenotypes of BV2 Microglia and Test Their Effects on Stroke Outcome in Mice

Abstract Backgrounds: Whether there are distinctive macrophage functional phenotypes of M1 versus M2 has been debated. We re-examined them by studying M1/M2 gene and protein expressions in cultured BV2 microglial cells and their effects on stroke outcomes in vivo. Methods: BV2 microglia cells were cultured and polarized with lipopolysaccharide (LPS) and interleukin-4 (IL-4) to produce M (LPS) and M (IL-4) phenotypes, which were originally defined as M1 and M2 phenotypes, respectively. Typical M1 and M2 gene or protein expression patterns were analyzed in M (LPS) and M (IL-4) phenotypes and their distinctive effects on stroke outcomes were compared. Results: M (LPS) and M (IL-4) had distinctive morphologies. M (IL-4) had significantly higher gene expressions of the typical M2 markers and other anti-inflammatory genes, while M (LPS) had higher gene expression of typical M1 markers and other pro-inflammatory genes. Nevertheless, M2 gene expressions were also enhanced in M (LPS), and M1 gene expressions were increased in M (IL-4), although with relatively lower levels. Adoptive transfer of M (IL-4) reduced infarction and improved neurological scores, while M (LPS) macrophages generated the opposite effect. Fluorescence activated cell sorting (FACS) and confocal studies suggest that M (IL-4) inhibited, while M (LPS) promoted the infiltration of monocyte-derived macrophages and iNOS-positive cells. Conclusions: M (LPS) and M (IL-4) from cultured BV2 cells indeed are distinctive functional phenotypes, but it is inaccurate to simply classify them into M1 and M2 phenotypes based on a few typical gene and protein markers.