Abstract: 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 receptor α and estrogen receptor β in 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 receptor α and β in 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. 

Key words: Gushukang, postmenopausal osteoporosis, estrogen receptor α, estrogen receptor β, estrogen signaling pathway, network pharmacology, traditional Chinese medicine