目的 基于大鼠體內(nèi)實(shí)驗(yàn)以及網(wǎng)絡(luò)藥理學(xué)探討白頭翁皂苷B₄治療子宮肌瘤的作用機(jī)制。方法 大鼠按體質(zhì)量隨機(jī)分為 6組：對(duì)照組、模型組、米非司酮（陽性藥，1.25 mg·kg^-1）組和白頭翁皂苷B₄低、中、高劑量（5、10、20 mg·kg^-1）組，每組 8只。采用雌激素負(fù)荷法制備子宮肌瘤模型，于造模成功后ip給藥，對(duì)照組及模型組每天ip等量0.9%氯化鈉溶液。通過對(duì)大鼠一般行為學(xué)的觀察、子宮組織的明場(chǎng)以及病理HE染色切片觀察、ELISA試劑盒檢測(cè)血清與子宮組織中雌二醇、孕酮、雌激素受體的表達(dá)水平，明確白頭翁皂苷 B₄對(duì)大鼠子宮肌瘤是否有治療作用；通過網(wǎng)絡(luò)藥理學(xué)收集白頭翁皂苷 B₄與子宮肌瘤的交互靶點(diǎn)，并對(duì)其進(jìn)行蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）網(wǎng)絡(luò)信息的構(gòu)建以及基因功能富集分析，并將靶點(diǎn)以及藥物進(jìn)行AutoDockVina分子對(duì)接分析，確定作用靶點(diǎn)；最終對(duì)組織進(jìn)行相關(guān)靶點(diǎn)Bcl-2、Bax、Caspase-3蛋白水平的Western blotting檢測(cè)進(jìn)行驗(yàn)證。結(jié)果 體內(nèi)實(shí)驗(yàn)結(jié)果顯示，與模型組比較，白頭翁皂苷B₄組大鼠子宮形態(tài)明顯好轉(zhuǎn)，子宮指數(shù)顯著降低（P＜0.001）；HE染色顯示，白頭翁皂苷B₄組大鼠子宮平滑細(xì)胞排列整齊，形態(tài)正常，色澤均一，肌層炎癥細(xì)胞浸潤(rùn)減少 ；血清及子宮組織中雌二醇、孕酮、雌激素受體的水平顯著降低（P＜0.05、0.01、0.001）。通過Pubchem數(shù)據(jù)庫(kù)篩選得到白頭翁皂苷B₄相關(guān)作用靶點(diǎn)23個(gè)，與子宮肌瘤共同作用靶點(diǎn)8個(gè)，富集得到69條KEGG信號(hào)通路，主要包括細(xì)胞凋亡、脂質(zhì)和動(dòng)脈粥樣硬化、IL-17、NF-κB、小細(xì)胞肺癌、MAPK等相關(guān)信號(hào)通路。AutoDock Vina結(jié)果顯示白頭翁皂苷 B₄與關(guān)鍵靶點(diǎn) BCL2、BAX、Caspase3結(jié)合良好。與模型組比較，白頭翁皂苷 B₄中、高劑量組 Bax、Caspase-3蛋白水平顯著升高（P＜0.001），Bcl-2蛋白水平顯著降低（P＜0.001）。結(jié)論 白頭翁皂苷B₄可能通過BCL2、BAX、Caspase-3等關(guān)鍵靶點(diǎn)誘導(dǎo)凋亡，同時(shí)降低雌二醇、孕酮、雌激素受體的表達(dá)，從而治療子宮肌瘤。

Objective To investigate the therapeutic mechanism of Pulsatilla saponin B₄ on leiomyomas in rats based on in vivo experiments and network pharmacology.Methods Rats were randomly divided into six groups according to body weight: control group, model group, mifepristone (positive drug, 1.25 mg·kg^-1) group, and Pulsatilla saponin B₄ low, medium, and high dose (5, 10, 20 mg·kg^-1) groups, with eight rats in each group. The leiomyoma model was established by hormone loading, and ip injection was given after the model was established. The control group and model group were injected with an equal volume of 0.9% sodium chloride solution daily. The general behavior of rats was observed, the uterine tissue was observed under bright field, and the pathological HE staining sections were observed. The expression levels of estradiol, progesterone, and estrogen receptor in serum and uterine tissue were detected by ELISA kits, to determine whether Pulsatilla saponin B₄ had a therapeutic effect on rat leiomyomas. The interactive targets of Pulsatilla saponin B₄ and leiomyoma were collected by network pharmacology, and the protein interaction network information was constructed and gene function enrichment analysis was performed. The targets and drugs were subjected to AutoDock Vina molecular docking analysis to determine the action targets. Finally, the protein levels of related targets Bcl-2, Bax, and Caspase-3 in the tissue were detected by Western blotting, and validated.Results The in vivo experiment results showed that compared with the model group, the uterine morphology of rats treated with Pulsatilla saponin B₄ improved significantly, and the uterine coefficient was significantly lower (P < 0.001); HE staining showed that rat uterine smooth cells were arranged in a regular pattern, with normal morphology, uniform color, and reduced inflammatory cell infiltration in the muscle layer. Serum and uterine tissue levels of estradiol, progesterone, and estrogen receptors were significantly lower (P < 0.05, 0.01, 0.001). Using the Pubchem database, 23 targets related to Pulsatilla saponin B₄ were screened, 8 of which were common targets with leiomyoma. Sixty-nine KEGG signaling pathways were enriched, including apoptosis, lipid and atherosclerosis, IL-17, NF-κB, small cell lung cancer, and MAPK related signaling pathways. The AutoDock Vina results showed that Pulsatilla saponin B₄ bound well to key targets BCL2, BAX, and Caspase3. Compared with the model group, the middle and high dose groups of Pulsatilla saponin B₄ showed significantly increased levels of Bax and Caspase-3 protein (P < 0.001), and significantly decreased levels of Bcl-2 protein (P < 0.001).Conclusion Pulsatilla saponin B₄ may induce apoptosis through key targets such as BCL2, BAX, and Caspase3, while reducing the expression of estrogen, progesterone, and estrogen receptors, thereby treating leiomyoma.

R285.5

2022年廣西科技基地和人才專項(xiàng)（2022AC18022）