目的 基于大鼠体内实验以及网络药理学探讨白头翁皂苷B₄治疗子宫肌瘤的作用机制。方法 大鼠按体质量随机分为 6组：对照组、模型组、米非司酮（阳性药，1.25 mg·kg^-1）组和白头翁皂苷B₄低、中、高剂量（5、10、20 mg·kg^-1）组，每组 8只。采用雌激素负荷法制备子宫肌瘤模型，于造模成功后ip给药，对照组及模型组每天ip等量0.9%氯化钠溶液。通过对大鼠一般行为学的观察、子宫组织的明场以及病理HE染色切片观察、ELISA试剂盒检测血清与子宫组织中雌二醇、孕酮、雌激素受体的表达水平，明确白头翁皂苷 B₄对大鼠子宫肌瘤是否有治疗作用；通过网络药理学收集白头翁皂苷 B₄与子宫肌瘤的交互靶点，并对其进行蛋白质-蛋白质相互作用（PPI）网络信息的构建以及基因功能富集分析，并将靶点以及药物进行AutoDockVina分子对接分析，确定作用靶点；最终对组织进行相关靶点Bcl-2、Bax、Caspase-3蛋白水平的Western blotting检测进行验证。结果 体内实验结果显示，与模型组比较，白头翁皂苷B₄组大鼠子宫形态明显好转，子宫指数显著降低（P＜0.001）；HE染色显示，白头翁皂苷B₄组大鼠子宫平滑细胞排列整齐，形态正常，色泽均一，肌层炎症细胞浸润减少 ；血清及子宫组织中雌二醇、孕酮、雌激素受体的水平显著降低（P＜0.05、0.01、0.001）。通过Pubchem数据库筛选得到白头翁皂苷B₄相关作用靶点23个，与子宫肌瘤共同作用靶点8个，富集得到69条KEGG信号通路，主要包括细胞凋亡、脂质和动脉粥样硬化、IL-17、NF-κB、小细胞肺癌、MAPK等相关信号通路。AutoDock Vina结果显示白头翁皂苷 B₄与关键靶点 BCL2、BAX、Caspase3结合良好。与模型组比较，白头翁皂苷 B₄中、高剂量组 Bax、Caspase-3蛋白水平显著升高（P＜0.001），Bcl-2蛋白水平显著降低（P＜0.001）。结论 白头翁皂苷B₄可能通过BCL2、BAX、Caspase-3等关键靶点诱导凋亡，同时降低雌二醇、孕酮、雌激素受体的表达，从而治疗子宫肌瘤。

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年广西科技基地和人才专项（2022AC18022）