Fig. 5

Gene networks targeted by miR-130a & miR-145. a Identification of miRNA targeted genes from TargetScan mouse database, which was intersected with mRNA expression microarray data comparing tumor Gr-1⁺CD11b⁺ cells with those from healthy control mice. Seven targets were common for miR-130a and -145. b IPA analysis of gene networks targeted by miR-130a (purple), miR-145 (blue), or both (orange) involving TGFβ and IGF pathways. c Validation of the major pathway mediators comparing tumor-associated myeloid cells with those from healthy controls, qRT-PCR (left) and Western blot (right). d qRT-PCR (left) and Western blot (right) from Gr-1⁺CD11b⁺ cells ex vivo treated with miR-130a or -145 or control mimics. e Immunofluorescence images of TβRII (Green), IGF1R (red), and DAPI (blue) in Gr-1⁺CD11b⁺ cells from the spleen of 4T1 tumor-bearing mice. scale bar: 10 μm. f M1/M2 cytokine ratio post restorations of TβRII, IGF1R, and IRS1 in Gr-1⁺CD11b⁺ cells that overexpress miR-130a or miR-145. The ratio of M1/M2 cytokines was calculated by dividing each M1 cytokine (TNFα, IL-12, GM-CSF) to M2 cytokine (IL-10, IL-4) as described in Material and Methods. g–l metastasis reduction by IGF1R inhibitor NT157: g The number of metastatic nodules of 4T1 tumor-bearing Tgfbr2^MyeKO and WT mice treated with NT157 (n = 7–10). h The number of metastatic nodules in 4T1 tumor-bearing mice that received IGF1R inhibitor NT157. i qRT-PCR of ARG1, TGFβ1, and IL-10 in Gr-1⁺CD11b⁺ cells from 4T1 tumor-bearing mice treated with NT157 (n = 5). j The number of metastatic nodules from mice bearing E0771 tumor treated with NT157 (n = 5–6). k qRT-PCR of ARG1, TGFβ1, and IL-10 in Gr-1⁺CD11b⁺ cells from E0771 tumor-bearing mice treated with NT157 (n = 4). l The number of metastatic nodules of 4T1 tumor-bearing mice treated with NT157 with CD8α neutralizing antibody or IgG as control (n = 10). Mice with similar tumor weight were used for metastatic nodule counts (Supplementary Fig. 5j). Data are presented as mean±SEM, and Student’s t test was performed. *p < 0.05, **p < 0.01, ***p < 0.001