6). In accordance with flow cytometry data (Fig. 2C), gene expression analysis of MHCII, a molecule thought to be on both M1 and M2 cells, revealed that the Arg1− macrophage population as a whole expressed much higher levels of MHCII transcripts (not shown)

and higher FDA-approved Drug Library solubility dmso levels of Ciita (class II, MHC, transactivator) than the Arg1+ macrophages (Fig. 5). The MHCII+ Arg1− macrophages may thus have increased capacity to present antigen to CD4+ T cells. Taken together, we conclude that Arg1+ and Arg1− macrophages each have mixed expression of M2 and M1 properties, and under the conditions of TBI Arg1 cannot be used as a marker for conventional M2 cells. To further compare Arg1+ and Arg1− TBI brain macrophages with M1 and M2 macrophages, we performed a meta-analysis of genes differentially expressed between Arg1+ and Arg1− TBI brain macrophages compared with genes differentially expressed between IFN-γ- or IL-4-stimulated bone marrow derived macrophages (BMDMs) stimulated in vitro with IFN-γ or with IL-4, representing M1 and M2 cells, respectively [38]. Arg1+ and Arg1− macrophages each upregulated a variety of genes that were also expressed

by BMDMs in response to either IFN-γ or IL-4 (Fig. 7). Thus, Arg1+ and Arg1− TBI brain macrophage subsets have features of both M1 and M2 phenotypes (Fig. 7). There are at least two explanations for these findings, not mutually exclusive: (i) individual brain macrophages may have features of both M1 and M2 cells (including cells Sirolimus cell line that are incompletely polarized or are in transition from between different states of polarization and (ii) there may be subsets of cells within the Arg1+ and Arg1− cells that have different expression of M1 and M2 markers. Regardless, the gene expression profiles demonstrate that Arg1+ and Arg1− macrophages

differ by many genes other than just Arg1. The most striking and novel differences between Arg1+ and Arg1− macrophages were in their unique chemokine profiles. Arg1+ macrophages MYO10 preferentially expressed a chemokine repertoire that included Ccl24 (which is also secreted by M2 cells; 6.2-fold), Cxcl7 (5.4-fold), Cxcl4 (2.4-fold), Cxcl3 (4.5-fold), Cxcl1 (3.6−fold), Cxcl14 (2.4-fold), and Ccl8 (2.3-fold) (Fig. 5). Arg1− macrophages, in contrast, preferentially upregulated Ccl17 (6.8-fold), Ccl5 (4.4-fold), Ccl22 (3.7-fold), and Ccr7 (tenfold) (Fig. 5). Although the gene profile of the Arg1+ macrophages suggests that they are not typical or homogeneously polarized M2 cells, they may have a role in promoting wound healing and in suppressing inflammation. Thus, Arg1+ macrophages preferentially expressed Spry2 (sprouty2; 2.4-fold), Cd9 (2.2-fold), Cd38, and Mt2 (metallothionein-2; 4.2-fold, Fig. 5). Sprouty2 and CD9 have protective roles in wound healing in skin injury models [39, 40]. Mt2 and Cd38 have been implicated in neuroprotection during brain injury [41, 42].