Fig. 5

miR-21-5p promotes VSMCs calcification by targeting the Crim1 gene. A Venn diagram showing potential target genes of miR-21-5p predicted by targetscan, pictar, miRWalk databases. B Schematic illustration of the sequences for miR-21-5p and the WT or mutated 3’-UTR of Crim1 mRNA. C Luciferase reporter assays showing the relative luciferase activity of Crim1 WT or mutated 3’-UTR reporter plasmids in VSMCs co-transfected with miR-21-5p mimics or mimics-NC. Firefly luciferase activity was normalized to renilla luciferase activity. D Western blot analysis of Crim1 levels in VSMCs transfected with miR-21-5p mimics, miR-21-5p inhibitor or their negative controls (NC). E Western blot analysis of the knockdown efficiency of Crim1 siRNAs in VSMCs. F Protein levels of Runx2, BMP2 and BMP4 in VSMCs treated with vehicle, siCrim1^#3 or siRNA-NC under 3.5 mM Pi induction as measured by Western blot. G ALP activity in VSMCs after treatment with vehicle, siCrim1^#3 or siRNA-NC under 3.5 mM Pi induction. Data are the mean ± SD of three independent experiments. *p < 0.05, **p < 0.01