白头翁汤正丁醇提取物通过调控自噬与凋亡缓解外阴阴道念珠菌病

n-butanol extract of Pulsatilla decoction alleviates vulvovaginal candidiasis by inducing autophagy and inhibiting apoptosis

摘要:

外阴阴道念珠菌病(vulvovaginal candidiasis, VVC)是女性生殖道常见的真菌感染性疾病。尽管针对VVC的传统治疗手段已相对成熟，但仍存在一定局限性。白头翁汤作为中国医学的经典方剂在临床上已被证实对VVC具有明确疗效，但其具体作用机制尚未完全阐明。目的 通过网络药理学及动物实验阐明中药白头翁汤正丁醇提取物(n-butanol extract of Pulsatilla decoction, BEPD)对VVC的作用机制。方法 构建VVC小鼠模型，评估BEPD对VVC的疗效；通过网络药理学筛选出BEPD对VVC潜在的作用信号通路，并采用蛋白印迹法、免疫荧光、免疫组化、实时荧光定量PCR检测小鼠阴道黏膜组织中自噬、凋亡及其相关通路蛋白的变化水平。结果 网络药理学分析显示，白头翁汤发挥抗真菌活性、治疗VVC的关键靶点包括PIK3RA和AKT1等；KEGG分析结果表明，白头翁汤可能通过调控PI3K-Akt信号通路发挥治疗VVC的作用。动物实验证实，相对于VVC模型组，BEPD治疗后，PI3K、p-Akt、p-mTOR蛋白表达下降，自噬相关蛋白LC3B和ATG5表达显著增加，凋亡相关蛋白Bax和Cleaved-Caspase-3的表达则明显下调，抗凋亡相关蛋白Bcl-2表达明显上调。结论 BEPD可能通过抑制PI3K-Akt-mTOR信号通路促进阴道上皮细胞自噬并抑制凋亡，从而恢复阴道黏膜上皮屏障稳态，缓解VVC。

Abstract:

Vulvovaginal candidiasis (VVC) is a prevalent fungal infection affecting the female reproductive tract. Although conventional therapeutic approaches for VVC are relatively well-established, they still exhibit certain limitations. Pulsatilla decoction, a classic traditional Chinese medicine formula, has demonstrated significant clinical efficacy in the treatment of VVC. However, its precise mechanism of action remains incompletely elucidated.Objective To clarify the therapeutic mechanism of the n-butanol extract of Pulsatilla decoction (BEPD) on VVC through network pharmacology and animal experiments.Methods A mouse model of VVC was established and the therapeutic effect of BEPD on VVC was evaluated. Network pharmacology was employed to screen the potential signaling pathways of BEPD on VVC. Western blotting, immunofluorescence, immunohistochemistry, and real-time fluorescence quantitative PCR were employed to measure the changes in autophagy, apoptosis, and related pathway proteins in the vaginal mucosa of mice.Results Network pharmacology analysis identified PIK3R1 and AKT1 as key targets of Pulsatilla decoction in exerting antifungal activity against VVC. KEGG pathway enrichment analysis indicated that Pulsatilla decoction exerted its therapeutic effects on VVC by regulating the PI3K-Akt signaling pathway. Animal experiments confirmed that compared with the VVC model group, the BEPD treatment down-regulated the expression of PI3K, p-Akt, and p-mTOR, significantly up-regulated the expression of autophagy-related proteins LC3B and ATG5, significantly inhibited the expression of apoptosis-related proteins Bax and Cleaved-Caspase-3, and significantly promoted the expression of anti-apoptosis-related protein Bcl-2.Conclusion BEPD may promote autophagy and inhibit apoptosis of vaginal epithelial cells by inhibiting the PI3K-Akt-mTOR signaling pathway, thereby restoring the homeostasis of the vaginal mucosal epithelial barrier and alleviating VVC.