Experimental Study Of Puerarin On Osteoblastic Proliferation And Its Signalling Pathway In Vitro

Purpose:This study aims to investigatethe the molecule mechanism of puerarin induced osteoblast proliferation and the dose-dependent relationship. By inhibiting ERK5 activity, as a mitogen-activated protein kinase (MAPK) family, we observed the markers of osteoblastic proliferation and downstream signaling proteins of ERK5. Eventually, our study will elucidate the mechanism of puerarin induced osteoblast proliferation and will provide a theoretical basis for the treatment of puerarin for osteoporosis.Methods:This study is performed to investigate the mechanism of puerarin induced osteoblast proliferation and for the osteoporosis treatment using molecular biology methods, from the cellular and molecular level. Before experiment, the culture medium containing puerarin is prepared. This experiment is divided into control group, puerarin groups (0.01 umol/L、0.1umol/L、1umol/L、10umol/L、 100umol/L) with different time (one to five days). MTT assay is used to assess the activity of osteoblast proliferation and examine the best dose of puerarin for osteoblast proliferation. Trypan blue assay is used to investigate the best dose of osteoblast survival by puerarin. We also examine the effect of puerarin on the osteoblastic cell cycle using Flow Cytometry. Besides, we observe the protein expression of G1/S phase and ERK5 expression, and investigate the role of ERK5 signalling pathway in the regulations of osteoblast proliferation.Conclusions:In summary, we find that (1) puerarin with the dose of lumol/L 10umol/L �'� 100umol/L significantly promotes the osteoblast proliferation, of which, puerarin in the five day is the most effective; (2) different dose of puerarin has different effect on osteoblast proliferation; (3) The main phase of osteoblast proliferation induced by puerarin is G1/S1 phase. (4) ERK5 signalling pathway may play an important role in the regulation of osteoblast proliferation by puerarin.