益肾蠲痹法含药血清对大鼠成骨和破骨细胞OPG/RANKL/RANK信号通路的影响  

Effect of serum containing Yishen Juanbi method on OPG/RANKL/RANK signaling pathway in osteoblasts and osteoclasts of rats

作　　者：沈丽英 李涯松[2] 张莹莹[2] 邹玉琼[2] 郝桂锋[2] 王天乐 SHEN Li-ying;LI Ya-song;ZHANG Ying-ying;ZOU Yu-qiong;HAO Gui-feng;WANG Tian-le(The Second Clinical Medical College,Zhejiang Chinese Medical University,Hangzhou,Zhejiang 310053,China)

机构地区：[1]浙江中医药大学第二临床医学院,浙江杭州310053 [2]浙江省人民医院杭州医学院附属人民医院风湿免疫科,浙江杭州310014

出　　处：《中华全科医学》2020年第2期197-201,共5页Chinese Journal of General Practice

基　　金：浙江省自然科学基金项目(LY14H270017);浙江省中医药管理局重点资助项目(2016ZZ006)。

摘　　要：目的探讨益肾蠲痹法含药血清对骨代谢信号通路的影响。方法正常SD大鼠随机分为对照组和益肾蠲痹组,分别经生理盐水和益肾蠲痹汤灌胃给药,获取含药血清;分离大鼠成骨细胞和破骨细胞,利用碱性磷酸酶和Van kossa染色鉴定成骨细胞,TRAP和甲苯胺蓝染色鉴定破骨细胞;将成骨和破骨细胞各分为4组:对照组、益肾蠲痹法含药血清低、中和高浓度组;通过免疫印迹检测成骨细胞骨保护蛋白(OPG)、核因子κB受体活化因子配体(RANKL)和破骨细胞核因子κB受体活化因子(RANK)的表达。结果低、中、高浓度含药血清诱导成骨细胞OPG的表达分别为对照组的97%(P=0.710)、122%(P=0.005)和155%(P<0.001),RANKL表达分别为对照组的81%(P<0.001)、63%(P<0.001)和53%(P<0.001),表明OPG和RANKL表达与含药血清浓度分别呈正、负相关;含药血清使破骨细胞RANK表达分别调节对照组的75%(P<0.001)、50%(P<0.001)和46%(P<0.001),表明RANK表达与含药血清剂量呈负相关。结论益肾蠲痹法含药血清通过调节OPG/RANKL/RANK信号轴可能在抗骨丢失过程中发挥作用。Objective To explore the effect of serum containing Yishen Juanbi method on the signal pathway of bone metabolism. Methods SD rats were randomly divided into control groups and Yishen Juanbi Decoction groups. The two group rats were given saline solution and Yishen Juanbi Decoction by gavage, respectively, and the serum containing Yishen Juanbi decoction was collected. The osteoblasts were isolated and verified by Van kossa staining and ALP staining, and the osteoclasts were isolated and verified by TRAP and toluidine blue staining. Both cell types were separated into four groups: control group, low, medium and high doses of serum containing Yishen Juanbi decoction groups. The immunoblotting assay was utilized to measure the expression of osteoprotegerin(OPG), receptor activator of nuclear factor-κB ligand(RANKL) and osteoclast receptor activator of nuclear factor-κB(RANK). Results Compared to control group, the expression of OPG and RANKL in osteoblasts induced by low, medium and high concentrations of serum containing Yishen Juanbi decoction were 97%(P=0.710), 122%(P=0.005) and 155%(P<0.001) respectively, and the expression of RANKL was 81%(P<0.001), 63%(P<0.001) and 53%(P<0.001) respectively. The results indicated that the expression of OPG was positively correlated with the concentration of serum and the expression of RANKL was negatively correlated with the concentration of serum. Meanwhile, the expression of RANK in osteoclasts was regulated to 75%(P<0.001), 50%(P<0.001) and 46%(P<0.001) compared to the control group, suggesting that the expression of RANK was negatively correlated with the concentration of serum. Conclusion The serum containing Yishen Juanbi decoction may play a prevention role in the process of bone loss by regulating OPG/RANKL/rank signal axis.

关 键 词：益肾蠲痹法 骨保护蛋白/核因子κB受体激活蛋白配体/核因子κB受体激活蛋白 成骨细胞 破骨细胞 

分 类 号：R593.23[医药卫生—内科学] R243[医药卫生—临床医学]