目的 探究制首乌水提物（PPMAE）通过调控NOD样受体热蛋白结构域相关蛋白3（NLRP3）/半胱氨酸天冬氨酸蛋白酶-1（Caspase-1）/消皮素D（GSDMD）信号通路抑制细胞焦亡，从而改善早发性卵巢功能不全（POI）的作用机制。方法 动情周期规律（卵巢功能正常）的雌性SD大鼠随机分为对照组、模型组、戊酸雌二醇(EV,0.18 mg·kg^-1，阳性药)组和PPMAE低、高剂量(1、4 g·kg^-1)^[11]组，每组6只。每天上午，模型组及各药物组ig雷公藤多苷混悬液(75 mg·kg^-1)诱导POI模型，对照组ig等体积0.9%氯化钠溶液。每天下午则分别ig相应药物，对照组与模型组ig 0.9%氯化钠溶液。阴道涂片检测动情周期，检测卵巢指数；苏木素-伊红（HE）染色与透射电镜分别观察卵泡发育及颗粒细胞超微结构；ELISA检测血清卵泡刺激素（FSH）、雌二醇（E₂）水平及卵巢组织乳酸脱氢酶（LDH）释放量；免疫荧光检测卵巢组织ASC斑点形成；实时荧光定量PCR（qRT-PCR）和Western blotting检测卵巢组织NLRP3、Caspase-1、GSDMD水平；免疫组织化学染色检测卵巢组织白细胞介素（IL）-1β、IL-18含量。结果 与模型组比较，PPMAE能显著改善POI大鼠动情周期紊乱、升高卵巢指数（P<0.05、0.01），改善卵巢病理损伤，增加原始卵泡储备并减少闭锁卵泡（P<0.01、0.001）；显著降低LDH释放，降低血清FSH水平，升高血清E2水平（P<0.05、0.01、0.001）。分子机制显示，PPMAE可显著抑制ASC斑点形成，下调NLRP3、Caspase-1、GSDMD的转录与蛋白表达，并降低下游分子IL-1β和IL-18含量（P<0.05、0.001）。结论 PPMAE可能通过抑制NLRP3/Caspase-1/GSDMD信号通路介导的细胞焦亡，从而改善POI。

Objectives This study investigates whether aqueous extract from processed Polygonum multiflorum（PPMAE） alleviates premature ovarian insufficiency（POI） by inhibiting pyroptosis via the NOD-like receptor pyrin domain-containing protein 3（NLRP3）/cysteinyl aspartate specific proteinase-1（Caspase-1）/gasdermin D（GSDMD）pathway. Methods Female SD rats with regular estrous cycles（normal ovarian function） were randomly divided into a control group, a model group, an estradiol valerate(EV, 0.18 mg·kg^-1, positive drug) group, and low-and high-dose PPMAE(1 and 4 g·kg^-1) groups, with 6 rats in each group. Every morning, the model group and each drug group were ig administered tripterygium glycosides suspension(75 mg·kg^-1) to induce the POI model, while the control group was ig administered an equal volume of 0.9% sodium chloride solution. Every afternoon, the corresponding drugs were ig administered, and the control group and the model group were ig administered 0.9% sodium chloride solution. Vaginal smears were used to detect estrous cycles, and the ovarian index was measured. Hematoxylin-eosin（HE） staining and transmission electron microscopy were used to observe follicular development and granulosa cell ultrastructure. ELISA was used to detect serum folliclestimulating hormone（FSH）, estradiol（E₂） levels, and lactate dehydrogenase（LDH） release in ovarian tissue. Immunofluorescence was used to detect ASC speck formation in ovarian tissue. Real-time fluorescence quantitative PCR（qRT-PCR） and Western blotting were used to detect the expression of NLRP3, Caspase-1, and GSDMD in ovarian tissue. Immunohistochemistry was used to detect the content of interleukin（IL）-1β and IL-18 in ovarian tissue. Results Compared with the model group, PPMAE significantly improved estrous cycle disorders in POI rats, increased the ovarian index（P ＜0.05, 0.01）, improved ovarian pathological damage, increased the reserve of primordial follicles, and reduced atretic follicles（P ＜0.01, 0.001）; significantly reduced LDH release, decreased serum FSH levels, and increased serum E2 levels（P ＜0.05, 0.01, 0.001）. Molecular mechanisms showed that PPMAE could significantly inhibit ASC speck formation, down-regulate the transcription and protein expression of NLRP3, Caspase-1, and GSDMD, and reduce the content of downstream molecules IL-1β and IL-18（P ＜0.05, 0.001）. Conclusion PPMAE may ameliorate POI by inhibiting pyroptosis through the NLRP3/Caspase-1/GSDMD pathway.

R965

国家自然科学基金资助项目(82360952)