知柏地黄汤治疗绝经后骨质疏松的机制研究  

Mechanism of Zhibai Dihuang decotion in treating postmenopausal osteoporosis

作　　者：江曼亚 叶悦 张琼谊 罗淦清 邓文喆 王萌[1] 栗原博[1] 何剑波 李怡芳[1] 何蓉蓉[1,2] JIANG Man-ya;YE Yue;ZHANG Qiong-yi;LUO Gan-qing;DENG Wen-zhe;WANG Meng;HIROSHI Kurihara;HE Jian-bo;LI Yi-fang;HE Rong-rong(Guangdong Engineering Research Center of Chinese Medicine and Disease Susceptibility,Jinan University,Guangzhou 510632,China;School of Traditional Chinese Medicine,Jinan University,Guangzhou 510632,China;Integration of Traditional Chinese and Western Medicine,Jinan University,Guangzhou 510632,China;The Second Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangdong Provincial Hospital of Chinese Medicine,Guangzhou 510120,China)

机构地区：[1]暨南大学,广东省疾病易感性及中医药研发工程技术研究中心,广东广州510632 [2]暨南大学中医学院,广东广州510632 [3]暨南大学中西医结合博士后流动站,广东广州510632 [4]广州中医药大学第二附属医院广东省中医院,广东广州510120

出　　处：《药学学报》2023年第10期3040-3048,共9页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金青年基金资助项目(82104898);中国博士后科学基金面上资助项目(2021M701442);广州市基础与应用基础研究项目(202201011552);广东省中医药局面上项目(20221180)。

摘　　要：本研究采用摘除卵巢手术(ovariectomy,OVX),建立原发性绝经后的骨质疏松小鼠模型,研究知柏地黄汤对绝经后骨质疏松症(postmenopausal osteoporosis,PMOP)的药效作用及相关机制。实验方案经暨南大学动物实验伦理委员会批准(批准号:20210315-03),所有程序均严格按照动物使用和护理的伦理原则进行。实验分为正常组、OVX组、知柏地黄汤低剂量(32 g·kg^(-1)·day^(-1))和高剂量组(64 g·kg^(-1)·day^(-1))、阳性药阿仑磷酸钠(9.9 mg·kg^(-1)·q3d)组。C57BL/6小鼠造模且给予药物干预8周后,取股骨和胫骨组织检测骨微结构、骨吸收和氧化应激等指标。实验结果显示,给予知柏地黄汤干预后,OVX所致的骨微结构损伤得到缓解,相关参数骨密度(bone mineral density,BMD)、骨体积分数(bonevolume/totalvolume,BV/TV)、骨小梁数目(trabecularnumber,Tb.N)、骨小梁连接密度(connectivity density,Conn.D)均明显增加。同时,TRAP阳性破骨细胞数量明显降低,破骨分化相关蛋白及基因水平降低,说明知柏地黄汤能抑制OVX后增强的破骨细胞活性。之后,利用网络药理学构建知柏地黄汤活性化合物-作用靶点网络,发现其活性成分的靶点基因与氧化应激通路密切相关。最后,骨组织中氧化应激水平检测结果显示,知柏地黄汤治疗后小鼠骨组织氧化应激产物4-羟基壬烯醛(4-hydroxynonenal,4-HNE)和丙二醛(malondialdehyde,MDA)水平显著降低,而抗氧化应激物谷胱甘肽(L-glutathione,GSH)的水平升高。以上结果显示,知柏地黄汤能够调控体内氧化应激水平抑制破骨活性而起到治疗PMOP的作用,将为中医药防治PMOP提供理论依据。In this study,the ovarian surgery(ovariectomy,OVX)was used to establish the osteoporosis mice model of primary menstruation,in order to evaluate the protective effects and mechanisms of Zhibai Dihuang decotion on postmenopausal osteoporosis(PMOP).The animal experimental protocol has been reviewed and approved by Laboratory Animal Ethics Committee of Jinan University(number:20210315-03),in compliance with the Institutional Animal Care Guidelines.C57BL/6 mice were divided into five groups,including Sham group,OVX group,low(32·kg^(-1)·day^(-1))and high dose(64·kg^(-1)·day^(-1))of Zhibai Dihuang decotion groups,positive drug group(alendronate,9.9 mg·kg^(-1)·q3d).After modeling,mice were given medication intervention for 8 weeks,and then femoral and tibial tissues were taken to detect indicators such as bone microstructure,bone resorption,and oxidative stress.The experimental results showed that after Zhibai Dihuang decotion administration,the bone microstructure damage caused by OvX surgery was alleviated,and the relevant parameters bone mineral density(BMD),bone volume/total volume(BV/TV),trabecular number(Tb.N)and connectivity density(Conn.D)both significantly increased.At the same time,the number of TRAP positive osteoclasts decreased significantly,and the levels of proteins and genes related to osteoclast differentiation decreased,indicating that Zhibai Dihuang decoction could inhibit the increased activity of osteoclast caused by OVX.Afterwards,network pharmacology was used to construct the active compound action target network of Zhibai Dihuang decotion,and it was found that the target genes of its active ingredients were closely related to the oxidative stress pathway.Finally,the detection results of oxidative stress levels in bone tissues showed that after treatment with Zhibai Dihuang decotion,the levels of oxidative stress products 4-hydroxynonenal(4-HNE)and malondialdehyde(MDA)in bone tissues of mice significantly decreased,while the levels of antioxidant stress substance L-glutathione(GSH)increased.

关 键 词：绝经后骨质疏松症 氧化应激 破骨细胞 骨吸收 知柏地黄汤 

分 类 号：R966[医药卫生—药理学]