补肾中药骨灵片对人成骨细胞功能和p38活化原蛋白激酶通路的影响

目的探讨含补肾中药骨灵片大鼠血清对人成骨细胞功能和对p38活化原蛋白激酶通路的影响。方法取人髂骨松质骨，采用改良胰酶一胶原酶消化法分离培养人成骨细胞；根据血清药理学的方法制备不同浓度（低、中、高剂量）的骨灵片血清，并以生理盐水处理的大鼠血清为阴性对照，将制备好的血清加入第二代的人成骨细胞培养液中培养。采用Western—blot及Real timeRT—PCR的方法观察各组成骨细胞护骨素和护骨素配体基因表达和磷酸化p38含量，用MTT法测定各组成骨细胞增殖率。利用茜素红染色法测定矿化结节形成的数量。结果含骨灵片（低、中、高剂量）药液血清处理的大鼠血清均能上调成骨细胞护骨素基因表达。下调护骨素配体基因表达，刺激成骨细胞的增殖，增加矿化结节形成的数量，刺激p38磷酸化。结论补肾中药骨灵片可能通过p38活化原蛋白激酶通路调控护骨素以及护骨素配体的表达，促进成骨细胞增殖以及矿化结节形成，抑制骨破坏。维持骨平衡。

Gu Ling Pian, A Kidney-Tonifying Traditional Chinese Medicine, Regulates the Functions of Osteoblasts and the p38 MAPK Pathway in Vitro

Objective To investigate the effect of kdney-tonifying tranditional Chinese medicine,Gu Ling Pian(GLP)on regulating human osteoblasts function and the osteoprotegerin(OPG)/receptor activator of nuclear factor kappa B ligand(RANKL)expression via p38 MAPK pathway in vitro.Methods Human osteoblast were cultured with serum obtained from rat fed with high,medium,or low dose GLP.The expression levels of the OPG and RANKL mRNA were determined by real-time RT-PCR.Total p38 and phosphorylated p38 were detected by western blot.Mineralized bone nodule was detected by alizarin red staining method.Results GLP serum promoted osteoblasts differentiation,increased the formation of mineralized bone nodule,up-regulated the OPG expression while RANKL expression decreased and increased the total protein of phosphorylated p38.Conclusion Kdney-tonifying traditional Chinese medicine,Gu Ling Pian,induced osteoblasts differentiation and synthesis of OPG/RANKL probably through p38 MAPK pathway.