雌激素受体在骨疏康治疗绝经后骨质疏松症中的作用与机制

The role and mechanism of estrogen receptor in the treatment of postmenopausal osteoporosis by Gushukang

背景:骨疏康作为治疗绝经后骨质疏松症肾虚血瘀证的中成药,其具体作用机制需深入研究。目的:研究骨疏康对去卵巢大鼠血清雌激素、骨微结构和雌激素受体的影响。方法:首先进行网络药理学分析,分别获取骨疏康有效成分、作用靶点和绝经后骨质疏松症的靶点,利用Cytoscape构建骨疏康有效成分-靶点网络,利用STRING数据库和Cytoscape进行蛋白互作分析(PPI)及核心靶点的筛选,利用DAVID数据库对交集靶点进行基因本体论(GO)和京都基因与基因组百科全书(KEGG)富集分析。然后建立去卵巢SD大鼠动物模型,骨疏康灌胃3个月,ELISA检测血清雌激素水平,microCT检测骨微结构,Western blot检测骨组织雌激素受体ERα和ERβ蛋白表达。结果与结论:①网络药理学分析显示,骨疏康中共有132个有效成分,150个作用靶点,与1155个绝经后骨质疏松症靶点取交集,获得87个骨疏康治疗绝经后骨质疏松症的作用靶点。②构建骨疏康有效成分-靶点网络图发现处于核心位置的有效成分有槲皮素、山柰酚、木犀草素、柚皮素和异鼠李素,处于核心位置的靶点有NCOA2,ESR2,AR,F2,ESR1,PTGS1。③PPI分析及筛选后最终获得的靶点有MAPK8,ESR1,JUN,R3C1,RELA和FOS,其中ESR1为两次分析获得的共同核心靶点。④随后的KEGG富集分析显示雌激素、TNF、凋亡等信号通路,因此动物实验重点关注骨疏康对雌激素信号通路中不同亚型的雌激素受体的影响。⑤动物实结果显示,骨疏康组骨微结构明显改善,血清雌激素无明显变化,但骨组织中ERα和ERβ蛋白量明显升高,说明骨疏康治疗绝经后骨质疏松症可能与其雌激素样作用和提高雌激素受体蛋白的表达有关。

BACKGROUND:The specific mechanism of Gushukang,as a Chinese traditional patent medicine for the treatment of postmenopausal osteoporosis of kidney deficiency and blood stasis,needs further studies.OBJECTIVE:To investigate the effect of Gushukang on serum sex hormones,bone microstructure and estrogen receptor in postmenopausal osteoporosis.METHODS:Firstly,network pharmacological analysis was performed.The active ingredients and action targets of Gushukang and the targets of postmenopausal osteoporosis were obtained respectively.Cytoscape was used to construct the active ingredient-target network.STRING database and Cytoscape were used for protein-protein interaction analysis and screening of core targets.DAVID database was used for Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses of intersection targets.Then the ovariectomized Sprague-Dawley rats were used in the animal experiment.Gushukang was administered by gavage for 3 months.The serum estrogen level was detected by ELISA,the bone microstructure was detected by microCT,and the protein expression of estrogen receptorαand estrogen receptorβin bone tiusse was detected by western blot.RESULTS AND CONCLUSION:The network pharmacological research results identified 132 active ingredients and 150 targets of Gushukang and 1155 targets of postmenopausal osteoporosis.After intersections with 1155 postmenopausal osteoporosis targets,87 targets of active ingredients of Gushukang against postmenopausal osteoporosis were obtained.By constructing the active ingredient-target network,it was found that the active ingredients at the core were quercetin,kaempferol,luteolin,naringin and isorhamnetin,and the targets at the core were NCOA2,ESR2,AR,F2,ESR1 and PTGS1.The final targets obtained after the protein-protein interaction analysis and screening included MAPK8,ESR1,JUN,R3C1,RELA and FOS,of which ESR1 was the common core target obtained from the two analyses.KEGG enrichment analysis showed estrogen,tumor necrosis factor,apoptosis and other signaling pathways.Therefore,animal experiments focused on the effect of Gushukang on different subtypes of estrogen receptors in the estrogen signaling pathway.The results showed that in the Gushukang group,bone microstructure was significantly improved,serum estrogen level had no significant change,but the protein expression of estrogen receptorαandβin bone tissue was significantly increased.All the findings indicate that the mechanism of Gushukang in the treatment of postmenopausal osteoporosis may be related to its hormone-like effect and the enhancement of estrogen receptor expression.