人牙周膜细胞骨保护因子和破骨细胞分化因子蛋白的表达及1α,25(OH)_2维生素D_3的调节

Expression of OPG and RANKL at protein level in human periodontal ligament cells and the effect of l_α,25(OH)_2 vitamin D_3 on the secretion of OPG protein in vitro

目的：研究人牙周膜细胞骨保护因子(osteoprotegerm，OPG)和破骨细胞分化因子(receptor activator nucle-ar factor kappa B ligand，RANKL)蛋白的表达，以及骨吸收促进因子1α，25(OH)_2维生素 D_3对 OPG 蛋白表达的影响。方法：用系列酶消化法体外培养人牙周膜细胞，用免疫细胞化学检测细胞上表达的 RANKL 蛋白，以酶联免疫吸附实验检测无外源性刺激以及1α，25(OH)_2维生素 D_3诱导2、4、6 d 时细胞分泌到培养基中的 OPG 蛋白。结果：人牙周膜细胞胞膜和胞浆上表达 RANKL 蛋白，分泌 OPG 蛋白到培养基中。1α，25(OH)_2维生素 D_3降低 OPG蛋白的分泌。结论：人牙周膜细胞表达 OPG 和 RANKL，1α，25(OH)_2维生素 D_3影响 OPG 的表达，推测其通过OPG/RANKL 通路参与骨改建。这一结论为进一步研究牙周膜细胞参与骨改建的机制打下基础。

Objective:To study the expression of osteoprotegerin(OPG)and receptor activator nuclearfactor kappa B ligand(RANKL)at protein level in human periodontal ligament cells(HPDLCs),and theeffect of 1α,25(OH)_2 vitamin D_3[1,25(OH)_2 vitD_3] on the secretion of OPG protein in vitro.Meth-ods:HPDLCs were harvested in vitro by sequential digestion with trypsin and collagenase.The expressionof RANKL in HPDLCs at protein level was tested by immunocyto-chemistry.Enzyme-linked immuno-adsordent assay(ELISA)was used to detect the OPG protein which was secreted into the culture mediumby HPDLCs cultured with and without 10^(-8) mol/L 1α,25(OH)_2 vitD_3 on the 0,2nd,4th,and 6th days,respectively.Results:RANKL protein was detected on the membrane and plasma of HPDLCs,and OPGprotein was secreted in the culture medium.The secretion of OPG protein was down-regulated by 10^(-8)mol/L 1α,25(OH)_2 vitD_3.Conclusion:HPDLCs have the bone metabolism system of OPG/RANKL,which works during the process of 1α,25(OH)_2 vitD_3 inducing HPDLCs.The conclusion has laid thegroundwork for the study on bone remodelling mechanisms of HPDLCs.