Objective: The present study aimed to investigate the mechanism of genistein, a tyrosine kinase inhibitor, regulating the differentiation of vascular smooth muscle cells (VSMCs) into osteoblasts via the OPG/RANKL (Osteoprotegerin/Receptor Activator of Nuclear Factor-κB Ligand) pathway.
Methods: The mouse VSMCs were isolated, purified and cultured. We constructed the LV5-Tnfrsf11b overexpression lentiviral vector and LV3-OPG-309 interference lentiviral vector. The OPG overexpression was induced and the growth of VSMCs infected with the lentiviral vector was observed. The VSMC calcification and control group were treated with different doses of genistein. The mRNA and protein expression levels of OPG, α-SM-actin (smooth muscle actin), ALP (alkaline phosphatase) and OPN (osteopontin) were detected in VSMCs after treatment using RT-PCR and Western Blot.
Result: We induced OPG overexpression and performed lentiviral vector infection of the VSMCs to suppress OPG expression, respectively, which was followed by treatment with genistein. The results showed that the relative expression of OPG was the highest in the VSMC calcification +genistein +OPG overexpression-inducing treatment group. It was the lowest in the VSMC calcification +OPG expression-suppressing treatment group. The relative expression of ALP was the highest in the VSMC calcification +OPG expression-suppressing treatment group, and the lowest in the VSMCs+genistein treatment group.
Conclusion: OPG gene plays an important regulatory role in the growth of VSMCs, by suppressing the calcification of VSMCs. Genistein could regulate the differentiation of VSMCs into osteoblasts via the OPG/RANKL pathway in a dose-dependent manner.
Keywords: OPG/RANKL pathway; VSMCs; genistein.