Fig. 8
Tumor-derived miR-375 is taken up by monocytes/MΦ and facilitates their infiltration. a–e 1 × 107 MCF-7 control or decoy cells were injected subcutaneously in the right and left flank of female NMRI-Foxn1nu mice, which were pre-treated with 17β-estradiol pellets for 1 week. After 35 days or a maximum tumor volume of 1.5 cm3 tumors were collected for flow cytometry and cell sorting. a Experimental scheme. b The tumor growth was monitored by measuring tumor volume (n = 5-6 per group). c Tumors were analyzed for the number of infiltrating MΦ and monocytes by flow cytometry as the number of cells in 100 mg tumor (n ≥ 5). d, e Infiltrating murine MΦ were sorted out of MCF-7 tumors and analyzed for the miR-375 content d and Pxn and Tns3 expression e by qPCR (n = 6). f C57BL/6 mice bone marrow-derived MΦ were cocultured for 48 h with E0771 murine breast cancer cells and Pxn and Tns3 mRNA expression was analyzed in MΦ (n = 6). g E0771 cells were stably transfected with miR-375 decoy (decoy) or empty vector (control) and miR-375 content was measured by qPCR (n = 7). h–k 50,000 E0771 control or decoy cells were injected in mammary gland 3 and 8 of 8-week-old female C57BL/6 mice. After 14 days, blood, bone marrow, spleen, and tumors were collected for flow cytometry and cell sorting. h Experimental scheme. i Single-cell suspensions of tumors were analyzed for the number of infiltrating MΦ and monocytes as the number of cells in 100 mg tumor. RNA from j plasma as well as k monocytes and MΦ from blood, bone marrow, spleen, and tumors were analyzed by qPCR for the abundance of miR-375 (n = 4–5 per group). Data are means ± SEM. P-values of i–k were calculated using nonparametric two-tailed Student’s t-test (b), two-tailed Student’s t-test (c–e, i–k) and one-sample t-test (f, g). *p < 0.05, **p < 0.01, ***p < 0.001
