不同浓度青娥丸含药血清对体外诱导破骨细胞MMP3OPN通路蛋白表达及其凋亡机制的影响  

Effect of Qinge Pills on expression of MMP3/OPN pathway proteins and osteoclast apoptosis of ovariectomized rats

作　　者：王度 戴燚[2] 范彦博[3] 

机构地区：[1]武汉市第三医院骨科,湖北武汉430060 [2]武汉市中医医院骨伤科,湖北武汉430014 [3]武汉市中医医院药理学实验室,湖北武汉430014

出　　处：《中国医院药学杂志》2016年第24期2173-2178,共6页Chinese Journal of Hospital Pharmacy

基　　金：武汉市科技局应用基础研究计划项目(编号:2015060101010061);武汉市中青年医学骨干人才项目(编号:2014-41);湖北省卫生厅青年科技人才项目(编号:QJX2010-48);武汉市青年科技晨光计划项目(编号:200950431215);武汉市卫计委临床医学项目(编号:WZ10C02)

摘　　要：目的:观察不同浓度复方青娥丸含药血清对去势鼠破骨细胞MMP3-OPN-P53通路蛋白表达及其凋亡的影响,探讨青娥丸抑制骨丢失的作用机制。方法:将20只SD大鼠去势饲养12周,处死选取股骨骨髓,进行体外OC培养。另用含2 g青娥丸生药的药液喂养20只SD大鼠,随机均分为空白对照(A)组(给纯化水1 mL),低浓度(B)组(给药0.5 mL),中浓度(C)组(给药1 mL),高浓度(D)组(给药2 mL),提取各组含药血清干预OC,运用图像系统测定TRAP阳性染色数目和骨磨片陷窝面积,流式细胞仪测定OC凋亡水平,RT-PCR法测定OC通路蛋白MMP3、OPN和P53的mRNA表达水平。结果:(1)术后12周模型组大鼠腰椎及股骨骨密度为(0.159±0.011)g·cm^(-2)和(0.158±0.011)g·cm^(-2),假手术组为(0.330±0.012)g·cm^(-2)和(0.298±0.016)g·cm^(-2),2组有明显差异(P<0.05),表明大鼠骨质疏松的模型建立成功。(2)加入不同药物干预后,D组中MMP3含量明显低于其余各组(P<0.01),OPN和P53含量高于其余各组(P<0.05)。B、C、D是随着含药血清依次增加的3组,MMP3表达依次减少,D组最少,而OPN和P53表达依次增加,其中D组在含药血清药物添加后明显增加,明显不同于A组(P<0.01)。(3)空白对照组流式细胞仪检测无亚G1峰,G0~G1期细胞占23%。低浓度组、中浓度组和高浓度组流式细胞仪均检测在G1期前出一陡立的凋亡峰(亚G1峰),占细胞总数的38%,42%和58%。结论:研究发现青娥丸含药血清可以通过MMP3-OPN-P53信号通路激活OC凋亡,抑制骨丢失,改善骨代谢,防治绝经后骨质疏松症。OBJECTIVE To investigate effects of compound Chinese medicine Qinge Pills on osteoclasts（OC）apoptosis and expression of MMP3-OPN-p53 protein pathway and its mechanism.METHODS Twenty rats were castrated for 12 weeks and killed,bone marrow was selected for OC culture in vitro.Another 20 SD rats were fed with solution containing2 g of raw drug of Qinge Pills,and were evenly and randomly divided into blank group（A）（1 mL of distilled water）,low dose group（B）（0.5mL of drug solution）,medium dose group（1 mL of drug solution）,high dose group（D）（2 mL of drug solution）.Drug serum was extracted from different groups to intervene OC.Image processing system was used to count staining with positive TRAP and lacunea area of bone slices,flow cytometry was used to determine OC apoptosis.RT-PCR was used to determine mRNA levels of OC pathway proteins including MMP3,OPN and P53.RESULTS Mineral densities of lumbar vertebrae and femur 12 weeks post operation were（0.159±0.011）and（0.158±0.011）g·cm^（-2） in model group,（0.330±0.012）and（0.298±0.016）g·cm^（-2） in sham operation group（P〈0.05）,indicating that rat osteoporosis model was successfully established.After different drugs were added,Compared with other groups,MMP3 content was obviously lower（P〈0.01）,OPN and P53 were higher in group D（P〈0.05）.In group B,C and D,MMP3 expression was reduced in order as drug in serum increased,the lowest value was observed in group D.But OPN and P53 expression were increased in order,group D had significantly increased value compared with group A（P〈0.01）.Flow cytometry in blank control group showed no sub G1 peak,and 23%cells at G0-G1 phase.Flow cytometry showed a sharp apoptosis peak before G1 phase in low,medium and high dose groups,accounting for 38%,42% and 58% of total cells.CONCLUSION Through MMP3-OPN-P53 signaling pathway,Qinge Pill medicated serum can activate OC apoptosis,inhibit bone loss,improve bone metabolism,and prevent and treat postmenopausal osteoporosi

关 键 词：基质金属蛋白酶3 骨桥蛋白受体 中药复方 信号通路 破骨细胞 

分 类 号：R285.5[医药卫生—中药学]