Fig. 6

miR-34a suppresses IL-6R expression and M2-like macrophage polarization. (a and b) MiR-34a levels were examined by qRT-PCR analysis in MCF-10A, MDA-MB-231 and A549 cells with different MCT-1 expression conditions. (c) The mature miR-34a levels were compared in the MDA-MB-231 cells (control vs. MCT-1) after transfection of pre-miR-34a and scramble-miR oligonucleotides. (d and e) IL-6R mRNA levels were compared in different MCT-1 backgrounds (control vs. MCT-1; scramble vs. shMCT-1#3) after transfection of pre-miR-34a and scramble-miR oligonucleotides. (f) IL-6R mRNA levels were compared in different MCT-1 backgrounds (control vs. MCT-1; scramble vs. shMCT-1#3) after transfection of pLemiR-antago-miR-34a or pLemiR-scramble. The MDA-MB-231 cells were virally transfected with pLemiR-scramble or pLemiR-pre-miR-34a, and then were co-incubated with THP-1 for 48 h to analyze the macrophage differentiation markers. (g) Pan-macrophages (indicated by F4/80). (h) M1-like macrophages (shown by CD86). (i and j) M2-like macrophages (revealed by CD163 and CD206). The results are expressed as the mean ± SD (n = 3). One-way ANOVA with a post hoc two-tailed t-test was used to calculate the statistical significance of pairwise comparisons. (*p < 0.05; **p < 0.01; *** p < 0.001). (k) THP-1 differentiation into M1 (CD86) or M2 (CD163 and CD206) macrophages were examined by qRT-PCR after culture with the MDA-MB-231 cells (control vs. MCT-1) introduced with pLemiR-antago-miR-34a or pLemiR-scramble. (l) The amounts of Snail, IL-10 and Stat3 were inspected in the THP-1 cells. Different blots with normalized internal control were used to present the data of the experiment. (m) Implications of MCT-1-targeted therapies. The oncogenic MCT-1 pathway in TNBC can be retracted by MCT-1 inhibitor(s), IL-6R immunotherapy or miR-34a expression, together they can further prevent EMT progression, cancer stemness, M2 macrophage polarization and tumor progression