骨疏康颗粒对去卵巢小鼠骨质疏松模型骨丢失和破骨细胞凋亡的影响  

Effects of Gushukang Granules on bone loss in ovariectomized mice and apoptosis of osteoclasts

作　　者：郑志坚[1] 舒冰[2,3] 赵世天 倪国栋 马增斌 王拥军[2,3] 赵东峰[2,3] ZHENG Zhi-jian;SHU Bing;ZHAO Shi-tian;NI Guo-dong;MA Zeng-bin;WANG Yong-jun;ZHAO Dong-feng(Department of Acupuncture and Massage,Beijing Hospital/National Center of Gerontology/Institute of Geriatric Medicine,Chinese Academy of Medical Sciences,Beijing 100730,China;Longhua Hospital,Shanghai University of Chinese Medicine,Shanghai 200032,China;Key Laboratory of Ministry of Education(Therapy of Muscles and Bones),Shanghai 200032,China)

机构地区：[1]北京医院针灸按摩科,国家老年医学中心,中国医学科学院老年医学研究院,北京100730 [2]上海中医药大学附属龙华医院,上海200032 [3]教育部筋骨理论与治法重点实验室,上海200032

出　　处：《中华中医药杂志》2020年第7期3647-3651,共5页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家自然科学基金重点项目(No.81730107);国家自然科学基金面上项目(No.81673991);国家自然科学基金青年科学基金项目(No.81804116);教育部创新团队计划项目(No.IRT1270);科技部重点领域创新团队计划项目(No.2015RA4002);国家重点研发计划项目(No.2018YFC1704300);上海市进一步加快中医药事业发展三年行动计划[No.ZY(2018-2020)-CCCX-3003]。

摘　　要：目的:通过构建体内去卵巢小鼠骨质疏松模型和体外骨疏康颗粒含药血清诱导破骨细胞分化和凋亡模型,明确骨疏康颗粒诱导破骨细胞凋亡的作用,为阐明骨疏康颗粒治疗绝经后骨质疏松症提供新的作用机制。方法:120只3月龄雌性C57BL/6小鼠随机选择20只行假手术造模(Con)。其余100只实施去卵巢(OVX)手术,造模1周内随机分为5组,每组20只:OVX(0.9%氯化钠溶液)、CAL(钙尔奇D)、GSKL(骨疏康颗粒2g·kg^-1·d^-1)、GSKM(骨疏康颗粒4g·kg^-1·d^-1)、GSKH(骨疏康颗粒8g·kg^-1·d^-1),连续灌胃3个月后收取小鼠腰椎(L3~L5),进行抗酒石酸盐酸性磷酸酶(TRAP)检测,观察其对破骨细胞数量的影响;同时采用TUNEL和TRAP双染法,观察破骨细胞的凋亡情况。体外实验部分,25只2月龄SD大鼠随机分为5组:生理盐水组(Con)、钙尔奇组(CAL)、GSKL、GSKM和GSKH,每组5只。灌胃完成后制备含药血清。取材C57BL/6小鼠单核巨噬细胞(BMMs)诱导破骨细胞的分化(4d),然后加入骨疏康颗粒制备的含药血清(5%),继续诱导2d后进行TRAP染色,观察含药血清对破骨细胞凋亡的影响。结果:体内研究发现,与对照组比较,OVX组小鼠破骨细胞的数量显著增多(P<0.05),骨疏康颗粒干预组小鼠破骨细胞数量显著下降(P<0.05)。TRAP和TUNEL双染色显示骨疏康颗粒促进小鼠体内破骨细胞的凋亡(P<0.05)。体外研究发现骨疏康颗粒含药血清能够诱导BMMs来源的破骨细胞的凋亡(P<0.05)。结论:骨疏康颗粒能够促进体内和体外破骨细胞凋亡,从而降低破骨细胞的骨吸收,延缓骨丢失和治疗骨质疏松症。Objective:Establishing an ovariectomized mice model in vivo of osteoporosis and building a model of osteoclast differentiation and apoptosis in vitro induced by serum containing Gushukang Granules(GSK)to clarify the role of Gushukang Granules in inducing osteoclast apoptosis and provide a new mechanism for the treatment of osteoporosis.Methods:A total of 1203-month clean female C57BL/6 mice,20 mice were randomly selected for sham surgery(Con).The remaining 100 mice underwent OVX surgery,and randomly divided into 5 groups(20 mice in each group)within 1 week of modeling:OVX(saline),CAL(calcium chi D),GSKL(GSK 2g·kg^-1·d^-1),GSKM(GSK 4g·kg^-1·d^-1),GSKH(GSK 8g·kg^-1·d^-1),after 3 months of intervention,the lumbar vertebrae of mice in each group were collected for tissue anti-tartrate acid phosphatase(TRAP),to observe the effect on the density of osteoclasts;the left tibia was collected and the apoptosis of osteoclasts was observed by TUNEL and TRAP double staining.In the part of in vitro experiment,252-month old SD rats were randomly divided into 5 groups for gavage:normal saline group(Con),calcine group(CAL),GSKL,GSKM and GSKH,5 mice in each group.Finally,C57BL/6 mice mononuclear macrophages(BMMs)were collected to induce osteoclast differentiation(4 days),and add the drug-containing serum prepared by GSK(5%),and the TRAP staining was conducted two days later to observe the effect of drug-containing serum on osteoclast apoptosis.Results:Comparing to the control group,the OVX mice showed the increased density of osteoclasts according to the in vivo data(P<0.05).However,the numbers of osteoclast in GSK-intervened mice were significantly decreased(P<0.05).TUNEL and TRAP double staining showed that GSKs promoted the apoptosis of osteoclasts in GSK-intervened mice(P<0.05).For in vitro osteoclastogenesis,the GSK-containing serum induced the increased apoptosis of osteoclasts from bone marrow macrophages(BMMs)(P<0.05).Conclusion:GSK promote the apoptosis osteoclasts both in vivo and in vitro,and reduces the bone abso

关 键 词：骨疏康颗粒 去卵巢小鼠 破骨细胞凋亡 骨吸收 绝经后骨质疏松症 

分 类 号：R285.5[医药卫生—中药学]