Figure 6. Microglial cell expression of the CB2R induces an activated phenotype and is not regulated by IL-6 signaling.

(A–C) CB2R^EGFP+ mice were transplanted with B10.BR BM alone or with B10.BR spleen cells. Animals were euthanized 14 days after transplantation. (A) Representative dot plot depicting recipient H-2K^b+ CD45⁺ CD11b⁺ microglial cells. (B) Representative histograms depicting EGFP expression on recipient microglial cells from the amygdala, brain stem, cerebellum, and prefrontal cortex of CB2R^EGFP animals reconstituted with B10.BR BM (BM, red line) or B10.BR BM and spleen cells (GVHD, blue line). (C) Scatterplot showing cumulative median fluorescence intensity (MFI) shifts from 4 replicate experiments. Each data point represents pooled results from 2 mice (n = 8–9 data points). (D) Representative immunofluorescence staining showing CB2R (GFP), microglia (TMEM119), and merged images from the prefrontal cortex of CB2R^EGFP+ mice transplanted with B10.BR BM and spleen cells. Magnified yellow insert box depicts microglial cell that is CB2R^– (blue box) and 1 that expresses the CB2R (green box). Scale bars: 10 μm. (E–G) CB2R^EGFP+ mice were transplanted with B10.BR BM alone or with B10.BR spleen cells and treated with an anti-IL-6R or isotype control antibody. (E) Representative dot plot depicting recipient H-2K^b+ CD45⁺ CD11b⁺ microglial cells. (F) Representative histograms depicting EGFP (CB2R) expression on microglial cells obtained from specified brain regions of CB2R^EGFP animals reconstituted with B10.BR BM only (BM, red line) or with B10.BR BM and spleen cells and treated with an isotype (GVHD, blue line) or anti-IL-6R antibody (GVHD, αIL-6R, green line). (G) Scatterplot data showing cumulative MFI shifts. Each data point represents pooled results from 2 mice. Results are from 5 experiments (n = 10–15 data points). (H–J) CB2R^EGFP+ (KI) or B6 mice were transplanted with B10.BR BM alone or with B10.BR spleen cells. Percentage of microglial cells expressing CB2R (EGFP) (H), and frequency of GFP⁺ (CB2R⁺) and GFP^– (CB2R^–) microglia expressing MHC class II, CD80, and CD86 (I and J). Results are from 2 experiments (n = 3–7 mice/group). Data are presented as mean ± SD. Statistics were performed using a t test with Welch’s correction for pairwise comparisons and a 1-way ANOVA with Tukey’s test for multiple group comparisons. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Source data are provided as a Supporting Data Values file.