Figure S4 from JMJD6 Shapes a Pro-tumor Microenvironment via ANXA1-Dependent Macrophage Polarization in Breast Cancer
Jmjd6-mediated ANXA1 secretion promotes M2 macrophage differentiation in vitro and ex vivo. A) Mean fluorescence intensity (MFI) of M1 and M2 markers in BMDM differentiated into M0, M1 and M2.** p<0,001, *** means p<0,0001, **** p<0,00001 without conditioned medium. B) Real time PCR showing the expression M1 (CD40 and NOS2) and M2 (CD206 and ARG1) markers in BMDMs differentiated into M0, M1 and M2 macrophages without CM. Datapoints show technical replicates.** p<0,001, *** p<0,0001. C) Gating strategies for FACS analysis on ex vivo mouse samples of Figure 6D. CD45+ leukocytes were gated on live cells, excluding dead cells and debris. In CD45+ population, myeloid cells were analyzed by the expression of CD11b, Gr-1, Ly6C and F4/80 markers. Monocyte-like myleloid-derived immunosuppressive cells (M-MDSC) were identified as Ly6Chigh Gr-1int expression. Granulocyte-like myeloid-derived immunosuppressive cells (G-MDSC) were identified as Gr-1high Ly6Cint cells. M2 macrophages were identified as CD206+ cells in the F4/80 gate. G-MDSC were identified as Gr-1high Ly6Cint cells. M2 macrophages were identified as CD206+ cells in the F4/80 gate.