葛根素对人成骨样MG-63细胞OPG、RANKL表达的调节作用

Regulatory effect of puerarin on the expression of OPG and RANKL in human osteoblastic MG-63 cells

【目的】观察葛根素对人成骨样MG-63细胞骨吸收调节因子骨保护素(osteoprotegerin,OPG)、NF-κB激活受体配体(receptor activator of nuclear factor kappa-B ligand,RANKL)表达的调节作用,并初步探讨对其可能的作用机制。【方法】选择兼有两种雌激素受体(estrogen receptor,ER)亚型表达的人成骨样MG-63细胞为研究模型,采用RT-PCR及Western blotting技术,检测OPG/RANKL的mRNA和蛋白的表达水平;采用RNA干扰技术特异性的将ERα、ERβ基因沉默,从而检测葛根素是否通过ERα和ERβ介导发挥作用。【结果】葛根素可显著上调成骨细胞OPG mRNA和蛋白的表达水平,下调RANKL mRNA和蛋白的表达水平。ER阻断剂ICI 182,780可完全阻断葛根素诱导的OPG表达水平增高,部分阻断了葛根素对RANKL表达的下调作用。利用RNA干扰技术构建ERα/ERβ不同表达状态的细胞株,结果显示:与对照组MG-63+葛根素、对照组MG-63/scrambled shRNA+葛根素相比,葛根素对MG-63/ERαshRNA和MG-63/ERβshRNA细胞OPG、RANKL的mRNA和蛋白水平的影响无显著性差异。【结论】葛根素显著上调MG-63细胞中OPG的mRNA及蛋白表达水平,同时下调RANKL的mRNA及蛋白表达水平,该作用可经ERα和ERβ共同介导。

【Objective】To observe the regulatory effect of puerarin on the expression of bone-absorption regulatory factors osteoprotegerin(OPG)and receptor activator of NF-κB ligand(RANKL)in human osteoblastic MG-63 cells,and preliminarily explore the possible mechanism.【Methods】Human osteoblastic MG-63 cells containing two estrogen receptor(ER)isoforms were chosen as the suitable model for this study.RT-PCR,Western blot analysis and RNA interference technology were respectively used to detect the effect of puerarin on the mRNA and protein expression of OPG and RANKL in MG-63 cells and explore the relevant mechanisms.【Results】Puerarin could significantly increase the mRNA and protein expression levels of OPG,but decrease the mRNA and protein expression levels of RANKL in osteoblastic MG-63 cells.The ER blocker ICI 182,780 completely blocked the increase of puerarininduced OPG expression level and partially blocked the down-regulation of puerarin on RANKL expression.RNA interference technology suggested the regulatory effect of puerarin on OPG and RANKL expression were mediated by ERαand ERβ.【Conclusion】Puerarin significantly up-regulates OPG expression but down-regulates RANKL expression in osteoblastic MG-63 cells.These effects are mediated via ERαand ERβpathways.