Puerarin inhibits TRPM3/miR-204 to promote MC3T3-E1 cells proliferation, differentiation and mineralization

Puerarin is an isoflavonoid phytoestrogen extracted from the root of Radix Pueraria, has attracted increasing attention because of its beneficial effects on anti-osteoporosis, but the molecular mechanisms underlying its actions on osteoblasts are not fully understood. The current study aimed to investigate the effect of puerarin on MC3T3-E1 osteoblastic cells proliferation, differentiation and mineralization, in vitro and its underlying mechanisms. The results indicated that puerarin significantly promoted the osteoblasts proliferation, enhanced alkaline phosphatase activity and increased the formation of mineralized nodules. Following treatment with puerarin, the expression levels of transient receptor potential Melastatin 3 (TRPM3) and microRNA-204 (miR-204) were decreased, whereas that of Runt-related transcription Factor 2 (Runx2) increased. TRPM3-small interfering RNA and 2-aminoethoxydiphenyl borate (2-APB, inhibitor of TRPM3) promoted the expression of Runx2 and thus improved the development of osteoblasts, but pregnenolone sulfate, which is the agonist of TRPM3, inhibited the effects. In addition, puerarin induced the changes of intracellular Ca²⁺ concentration ([Ca²⁺ ]_i ) and extracellular Ca²⁺ concentration ([Ca²⁺ ]₀ ) through TRPM3 might be involved in the biological process of MC3T3-E1 cells. These results suggested that puerarin may promote MC3T3-E1 cell proliferation, differentiation and mineralization, which may be related to the downregulation of TRPM3/miR-204 and following regulating [Ca²⁺ ]_i and [Ca²⁺ ]₀ , and activation of Runx2.