葛根对成骨细胞OPG、RANKL mRNA 表达的影响_药学论文.docVIP

葛根对成骨细胞OPG、RANKL mRNA 表达的影响_药学论文 葛根对成骨细胞OPG、RANKL mRNA 表达的影响_药学论文 作者：吴乃中，李红丽，崔淑云 【摘要】 目的 本研究用RT-PCR的方法观察葛根中药血清对成骨细胞细胞因子：OPG、RANKL mRNA表达的影响，力求从细胞水平和分子水平对葛根抗骨质疏松的机制进行探讨，为葛根应用于临床防治骨质疏松提供有力的实验依据。方法 将(250±10)g的SD大鼠36只，随机分为两组：对照组灌服生理盐水，中药组灌服2g/ml的葛根10ml/kg·d，1日2次灌胃，连续3天，最后一次灌胃后中药组和对照组分别1h、2h、4h无菌操作股动脉采血，收集血清备用。出生24h内的SD大鼠，无菌取颅盖骨，做成骨细胞培养。第三代成骨细胞 5×104/ml细胞悬液，24h待细胞贴壁后加入实验因素，6天后用异硫氰酸胍一步法提取总RAN，然后用RT-PCR检测OPG、RANKL mRNA的表达。结果 最后一次用药后2h采血的中药血清可明显上调成骨细胞OPGmRNA表达，且明显下调成骨细胞RANKL mRNA的表达。随血清添加量的增加，对成骨细胞OPG mRNA表达上调作用越强，对成骨细胞RANKL mRNA的表达无影响。结论 实验表明，葛根可能通过上调成骨细胞OPG mRNA的表达，下调成骨细胞RANKL mRNA的表达，抑制破骨细胞的产生和功能，为葛根防治骨质疏松提供了有力的实验依据。 【关键词】 葛根；成骨细胞；OPG；RANKL 【Abstract】 Objective To explore the effects of roots of kudzu vine on the mRNA expression of OPG and RANKL of osteoblasts using RT-PCR. To provide the experimental base for its preventing and treating the OP in clinic， it is urgent to explore the mechanism of roots of kudzu vine anti-osteoporosis in cell and molecular level.Methods 36 SD rats weighting (250±10)g，are randomly divided into two groups. Control group and herb group were administered orally with physiological salt solution and with epimedium sagittatum maxim（2g/ml）twice a day at 10ml/kg body weight per day， respectively for three days. At the third day the serum was collected from the femoral artery at one hour， two hours and four hours after the last administration of medicine. Primary rat osteoblasts were isolated from newborn mouse calvariae. The third passage osteoblasts were plated at the density of 5×104/ml in 75ml culture flask， twenty four hours later treatment factors were added. Total RAN was extracted from osteoblasts using guanidine， and mRNA expression of OPG and RANKL were examined using RT-PCR.Results Serum of roots of kudzu vine can up-regulate mRNA expression of OPG of osteoblasts and down-regulate mRNA expression of RANKL of osteoblasts.The effect of serum of medicine harvested at 2h after the last medicine administration is the most strong and the eff