背景:卵巢储备功能下降(DOR)被认为是一种难治性生殖内分泌疾病,对女性生殖产生负面影响。养精中玉汤(YJZYD)具有治疗不孕症的作用。然而,关于YJZYD保留卵巢储备的机制的研究很少。
目的:通过UPLC-ESI-MS/MS探讨YJZYD抗DOR的可能机制,网络药理学,和实验验证。
方法:用UPLC-ESI-MS/MS测定YJZYD的化学物质。通过ETCM数据库确定YJZYD和DOR的相关靶标,GeneCards数据库,和PubMed数据库。共同目标与DAVID数据库一起使用,并用PPI网络可视化。进行GO和KEGG富集分析以探索生物学进展和途径。体内实验,通过ATP评估能量产生,用TUNEL法分析细胞凋亡率。血清FSH,AMH,通过ELISA评估E2水平。免疫印迹法和免疫组化法检测SIRT1、PGC1α、NRF1,COXIV,FSHR,CYP19A1,PI3K,p-Akt,Akt,Bcl-2和Bax。
结果:通过UPLC-ESI-MS/MS鉴定了YJZYD中的132个成分。从YJZYD和DOR中提取149个重叠目标,前20个常见目标包括AKT1和CYP19A1。ATP结合参与GO分析。在KEGG富集分析中,代谢途径是最高的,并纳入PI3K-Akt信号通路。体内实验,YJZYD改善卵巢指数和组织形态学。YJZYD治疗后,血清FSH,E2和AMH被很好地调节,ATP含量上调。此外,Bax在卵巢组织中的表达受到抑制,而SIRT1、PGC1α、NRF1,COXIV,FSHR,CYP19A1,PI3K,Bcl-2和p-Akt/Akt增强。
结论:YJZYD可通过介导类固醇生成减轻体内生殖内分泌紊乱和卵巢病变,能量代谢,和细胞凋亡。本研究揭示了YJZYD抗DOR的机制,为进一步研究提供理论依据。
BACKGROUND: Diminished ovarian reserve (DOR) was considered a refractory reproductive endocrine condition that negatively affected female reproductivity. Yangjing Zhongyu Decoction (YJZYD) had effects on treating infertility. However, there were few studies on the mechanisms of YJZYD preserving ovarian reserve.
OBJECTIVE: To explore the possible mechanisms of YJZYD against DOR by UPLC-ESI-MS/MS, network pharmacology, and experimental validation.
METHODS: The chemicals of YJZYD were measured by UPLC-ESI-MS/MS. The correlating targets of YJZYD and DOR were identified by the ETCM database, GeneCards database, and PubMed database. The common targets were employed with the DAVID database and visualized with the PPI network. GO and KEGG enrichment analyses were carried out to explore biological progression and pathways. In vivo experiments, energy production was assessed by ATP, and apoptosis rate was analyzed by TUNEL. The serum FSH, AMH, and E2 levels were evaluated by ELISA. Western blotting and immunohistochemistry were used to measure the expression of SIRT1, PGC1α, NRF1, COX IV, FSHR, CYP19A1, PI3K, p-Akt, Akt, Bcl-2, and Bax.
RESULTS: 132 components in YJZYD were identified by UPLC-ESI-MS/MS. 149 overlapped targets were extracted from YJZYD and DOR, and the top 20 common targets included AKT1 and CYP19A1. ATP binding was involved in GO analysis. In the KEGG enrichment analysis, the metabolic pathway was the top, and the PI3K-Akt signaling pathway was included. In vivo experiments, YJZYD improved ovarian index and histomorphology. After YJZYD treatment, serum FSH, E2, and AMH were well-modulated, and the content of ATP was up-regulated. Besides, the expression of Bax was suppressed in ovarian tissue, while the expressions of SIRT1, PGC1α, NRF1, COX IV, FSHR, CYP19A1, PI3K, Bcl-2, and p-Akt/Akt were enhanced.
CONCLUSIONS: YJZYD could attenuate reproductive endocrine disturbance and ovarian lesions in vivo by mediating steroidogenesis, energy metabolism, and cell apoptosis. This study uncovered the mechanisms of YJZYD against DOR, providing a theoretical basis for further study.