Moreover, gain- and loss-of-function experiments and western blotting analysis revealed that transient transfection of miR-515-3p mimic or miR-515-3p-expressing plasmid not only markedly reduced vimentin and MMP3 expression at mRNA and protein levels, but also decreased fibronectin and N-cadherin expression in ESCC cell lines (Fig. 4h and Supplementary Fig. S6a, b), whereas silencing of miR-515-3p with inhibitor had opposite effect on vimentin, MMP3 and EMT markers (Fig. 4i and Supplementary Fig. S6c). This evidence concerns the gene MMP3 and esophageal squamous cell carcinoma.