目的 利用網(wǎng)絡(luò)藥理學(xué)方法，探究暖宮七味丸治療痛經(jīng)的作用機(jī)制并采用痛經(jīng)模型大鼠對(duì)其進(jìn)行實(shí)驗(yàn)驗(yàn)證。方法 使用中藥系統(tǒng)藥理學(xué)數(shù)據(jù)庫(kù)與分析平臺(tái)（TCMSP）、中醫(yī)藥整合藥理學(xué)研究平臺(tái)（TCMIP）和網(wǎng)絡(luò)藥理學(xué)在線數(shù)據(jù)庫(kù)（TCMID）結(jié)合文獻(xiàn)報(bào)道確定暖宮七味丸的活性成分，通過(guò)Swiss Target Prediction在線平臺(tái)得到活性成分對(duì)應(yīng)的靶點(diǎn)。利用DisGeNET數(shù)據(jù)庫(kù)和GeneCard數(shù)據(jù)庫(kù)確定原發(fā)性痛經(jīng)的相關(guān)靶點(diǎn)，手動(dòng)篩選化合物-疾病的共同靶點(diǎn)，將潛在共同靶點(diǎn)蛋白上傳至Biogrid平臺(tái)，獲取藥物-疾病靶蛋白相互作用（PPI）網(wǎng)絡(luò)，利用Cytoscape 3.7.2軟件對(duì)PPI網(wǎng)絡(luò)進(jìn)行網(wǎng)絡(luò)圖優(yōu)化和拓?fù)鋵W(xué)分析。利用DAVID 6.8數(shù)據(jù)庫(kù)對(duì)潛在靶點(diǎn)進(jìn)行京都基因與基因組百科全書(shū)（KEGG）通路富集分析及基因本體論（GO）注釋分析。動(dòng)物實(shí)驗(yàn)驗(yàn)證采用縮宮素致痛經(jīng)大鼠模型觀察暖宮七味丸高、中、低劑量（2.0、1.0、0.5 g·kg^-1）的鎮(zhèn)痛作用，采用Western blotting方法觀察暖宮七味丸高、低劑量（2.0、0.5 g·kg^-1）對(duì)痛經(jīng)模型大鼠子宮組織環(huán)氧合酶-2（COX-2）蛋白表達(dá)的影響，并測(cè)定各組大鼠子宮組織中前列腺素F2α（PGF2α）水平。通過(guò)大鼠離體子宮肌張力實(shí)驗(yàn)觀察暖宮七味丸水溶液（22.5、11.25 mg·mL^-1）對(duì)大鼠子宮收縮幅度和收縮頻率的影響。結(jié)果 網(wǎng)絡(luò)藥理學(xué)預(yù)測(cè)結(jié)果顯示，暖宮七味丸治療痛經(jīng)的潛在靶點(diǎn)有68個(gè)，PPI核心靶點(diǎn)有IL-6、PTGS2、TNF和TP53等。涉及到的通路有炎癥相關(guān)的信號(hào)通路、癌癥通路和雌激素信號(hào)通路等，涉及到的生物功能有炎癥反應(yīng)、一氧化氮的生物調(diào)節(jié)過(guò)程和對(duì)激素的反應(yīng)和對(duì)血管的調(diào)節(jié)功能等。動(dòng)物實(shí)驗(yàn)結(jié)果顯示，與模型組比較，暖宮七味丸高、中、低劑量組和陽(yáng)性藥組大鼠的扭體反應(yīng)次數(shù)顯著減少（P＜0.01、0.001），同時(shí)扭體反應(yīng)的潛伏期均明顯延長(zhǎng)（P＜0.01、0.001）；暖宮七味丸高、低劑量組痛經(jīng)模型大鼠子宮組織中過(guò)氧化物合成酶2（PTGS2）相關(guān)蛋白COX-2表達(dá)顯著下調(diào)（P＜0.001）），同時(shí)子宮組織中PGF2α水平顯著降低（P＜0.001）。暖宮七味丸高、低劑量水提液對(duì)大鼠子宮收縮幅度和頻率均有顯著的抑制作用（P＜0.001）。結(jié)論 暖宮七味丸可以通過(guò)下調(diào)PTGS2靶點(diǎn)進(jìn)而調(diào)控子宮肌張力起到治療痛經(jīng)的效果。

Objective To explore the mechanism of Nuangong Qiwei Pill in treatment of primary dysmenorrhea by network pharmacological methods and to verify the prediction results by animal experiments. Methods The active components of Nuangong Qiwei Pills were screened by using the traditional Chinese Medicine System Pharmacology database and analysis platform (TCMSP), traditional Chinese medicine integrated pharmacology research platform (TCMIP) and online pharmacology database (TCMID) combined with literature reports, and the corresponding targets of the active components were obtained through the Swiss target prediction online platform. The relevant targets of primary dysmenorrhea were obtained by using the database of DisGeNET and GeneCard, the common targets of compounds and diseases were manually screened, the potential common target proteins were uploaded to the Biogrid platform. The protein-protein interaction (PPI) network of drug-disease target was obtained, and the network diagram optimization and topology analysis of PPI network were carried out by using the software of Cytoscape 3.7.2. David 6.8 database was used to analyze the enrichment of Kyoto Encyclopedia of genes and genomes (KEGG) pathway and Gene Ontology (GO) of potential targets. The rat model of dysmenorrhea caused by oxytocin was used to observe the analgesic effect of high, medium and low doses (2.0, 1.0, 0.5 g·kg^-1) of Nuangong Qiwei Pills, and the effect of high and low doses (2.0, 0.5 g·kg^-1) of Nuangong Qiwei Pills on the expression of cyclooxygenase-2 (COX-2) protein in the uterus of dysmenorrhea rats was observed by Western blotting method, and prostaglandin F2α (PGF2α) level in the uterus of rats in each group was measured. The effects of the aqueous solution of Nuangong Qiwei Pills (22.5, 11.25 mg·mL^-1) on the amplitude and frequency of uterine contraction in rats were observed by the tension test of isolated uterine muscles in rats. Results The predicted results of network pharmacology showed that there were 68 potential targets of Nuangong Qiwei Pills in treatment of dysmenorrhea, and the core targets of PPI were IL-6, PTGS2, TNF and TP53. The pathways involved include inflammation related signaling pathways, cancer pathways and estrogen signaling pathways, and the biological functions involved include inflammatory response, biological regulation process of nitric oxide, response to hormones and regulation function of blood vessels. The results of animal experiments showed that compared with the model group, the number of writhing reaction of rats in the high, medium and low dose of Nuangong Qiwei Pills groups and positive drug groups was significantly reduced (P < 0.01, 0.001), and the incubation period of writhing reaction was significantly prolonged (P < 0.01, 0.001). The expression of PTGS2 related protein COX-2 in the uterus of dysmenorrhea model rats in the high, medium and low dose groups of Nuangong Qiwei Pills groups was significantly down regulated (P < 0.001), and the level of PGF2α in the uterus decreased significantly (P < 0.001). The high and low doses of water extract of Nuangong Qiwei Pills had significant inhibitory effects on the amplitude and frequency of uterine contraction in rats (P < 0.001). Conclusion Nuangong Qiwei Pills have therapeutic effect on dysmenorrhea by down regulating PTGS2 target and then regulating uterine muscle tension.

R285.5

國(guó)家重點(diǎn)研發(fā)計(jì)劃（2018YFC1708203）；中央高校基本科研業(yè)務(wù)費(fèi)專項(xiàng)（3332021064）；內(nèi)蒙古自治區(qū)科技計(jì)劃（2020GG0199）