Runx2 expression in bone metastases from breast cancer patients inversely correlates with Runx2 targeting miRNAs. A, Runx2, Smad5 and pan-cytokeratin proteins were detected by immunohistochemistry in four representative tissue biopsies obtained from bone metastases of breast cancer patients and B, protein by western blot analysis in cell lysates from normal epithelial MCF-10A cells, non-metastatic MCF-7 cells, and metastatic MDA-MB-231 cell lines. C, expression of miRNAs that target Runx2 was analyzed in MCF-10A cells, MCF-7 cells, and in two highly metastatic MDA-MB-231 cell lines by qRT-PCR. U6 expression was used as internal control. D, miRNAs were isolated from paraffin-embedded tissue specimens, reverse transcribed, and expression of miR-135 and miR-203 was quantified by qRT-PCR. U6 was used as an internal control. Scale bar 50 μm. Data are represented as means ± SD of each group. *significant difference (p < 0.05), Tukey’s post-hoc test.