Fig. 5

MiR-30c-5p inhibits VSMC differentiation from adventitial Sca-1⁺ progenitor cells through targeting OPG. Adventitial Sca-1⁺ progenitor cells were transfected with mimic control/miR-30c-5p mimic/inhibitor control/miR-30c-5p inhibitor for 12–24 h. The expression of OPG was measured by RT-qPCR (a, b) and western blot (c). Adventitial Sca-1⁺ progenitor cells were induced by TGF-β1. The expression of OPG was detected by RT-qPCR at different time points (0 h, 2 h, 6 h, 10 h) (d). Adventitial Sca-1⁺ progenitor cells were transfected with siRNA control and siRNA-OPG 4–6 h before the TGF-β1 induced differentiation and the cells were collected after another 72 h. The expressions of OPG, SMαA, SM22α, SMMHC, and h1-caponin were measured by RT-qPCR (e). Adventitial Sca-1⁺ progenitor cells were co-transfected with siR-OPG and miR-30c-5p mimic/miR-30c-5p inhibitor 4–6 h before the TGF-β1 induced differentiation and the cells were collected after another 72 h. The expression of SMαA, SM22α, SMMHC, and h1-caponin were detected by RT-qPCR and western blot (f–i). HEK-293T cells were co-transfected with psiCHECK2-OPG-3′UTR (j) or psiCHECK2-OPGmut-3′UTR (k) and mimic control/miR-30c-5p mimic/inhibitor control/miR-30c-5p inhibitor, and the luciferase and renilla activity were detected by luminometer and the relative luciferase activity was calculated (*P < 0.05, **P < 0.01, ns P ≥ 0.05)