乌鸡白凤片对 4-去氧乙烯基环已烯所致小鼠早发性卵巢功能不全的影响

Effect of Wuji Baifeng Tablets on premature ovarian insufficiency in mice induced by VCD

关键词:  乌鸡白凤片  4-去氧乙烯基环已烯  早发性卵巢功能不全  促卵泡生长素  促黄体生成素

Key words:  Wuji Baifeng Tablets  VCD  primary ovarian insufficiency  FSH  LH

摘要:

[目的] 运用 4-去氧乙烯基环已烯（VCD）建立早发性卵巢功能不全（POI）小鼠模型， 探究乌鸡白凤片对VCD 致 POI 小鼠的治疗作用。[方法] 1）建立 POI 小鼠模型： 8 周龄雌性昆明小鼠 60 只， 随机分为正常（ CON）组30 只， 模型（MOD）组 30 只。MOD 组腹腔注射 160 mg/ （kg· d）VCD 复制 POI 模型， CON 组注射同体积芝麻油。造模期间每日阴道涂片观察动情周期； 造模第 15~65 天， 每 5 d 进行眼眶取血， 酶联免疫吸附（ELISA）法检测血清促卵泡生长素（FSH）、促黄体生成素（LH）水平； 造模第 45~65 天， 每 5 d 取小鼠子宫卵巢， 称质量后计算脏器指数， 苏木精-伊红（HE）染色观察卵巢形态学改变。2）乌鸡白凤片对 POI 小鼠的治疗作用： 按照上述造模方法再选取 8 周龄雌性昆明小鼠 60 只， 造模成功后随机分为 MOD 组、戊酸雌二醇（E2）组[0.3 mg/ （kg· d）]、乌鸡白凤片低、中、高剂量组 [410、820、1640 mg/ （kg· d）]， 每组 10 只。CON 组和 MOD 组予以等体积羧甲基纤维素钠（CMC-Na）灌胃。每日 1 次， 连续给药 45 d， 给药期间观察小鼠动情周期。于给药第 46 天处死小鼠， ELISA 法检测血清 FSH、LH 水平， 取小鼠子宫卵巢，称质量后计算脏器指数， HE 染色观察卵巢形态学改变。[结果] 1）小鼠 POI 模型的建立： 与 CON 组相比， MOD 组小鼠第 55 天动情周期紊乱持续 4 周， VCD 造模第 50、55 天 MOD 组小鼠血清 FSH、LH 显著升高 2 次， 子宫、卵巢湿质量及脏器指数显著下降， 生长卵泡数量显著减少。2）乌鸡白凤片对 POI 小鼠的治疗作用： 灌胃给药 45 d 后， 与 CON 组相比， MOD 组小鼠动情周期紊乱， 血清 FSH、LH 显著升高， 子宫、卵巢湿质量及脏器指数显著下降， 生长卵泡数量显著减少。与 MOD 组相比， 乌鸡白凤片中、高剂量组小鼠动情周期全部恢复正常， 且乌鸡白凤片各剂量组小鼠血清FSH、LH 下降， 子宫、卵巢湿质量及脏器指数显著升高， 卵巢内原始卵泡数量明显增多。[结论] 160 mg/ （kg· d） VCD 腹腔注射 15 d 后， 观察 40 d， 总计造模 55 d 后可制备出 POI 小鼠模型； 乌鸡白凤片可能通过调节小鼠下丘脑-垂体-卵巢轴（HPOA）下调血清 FSH、LH 水平， 调节小鼠动情周期， 促进卵泡发育从而治疗 VCD 所致小鼠 POI。

Abstract:

[Objective] Establish the mouse model of primary ovarian insufficiency（POI） with 4-deoxyvinylcyclohexene（VCD） and explore the therapeutic effect of Wuji Baifeng Tablets on VCD-induced POI mice. [Methods] 1）The POI mouse model was established：sixty 8 week old Kunming female mice were randomly divided into 30 control （CON） mice and 30 model （MOD） mice. In MOD group，the POI model was reproduced by intraperitoneal injection of 160 mg/（kg·d） VCD，and CON was injected with sesame oil of the same volume. During the modeling period，the estrus cycle was observed by vaginal smear every day；from day 15 to day 65 of modeling，blood was taken from orbit every 5 days，and serum follicle stimulating hormone （FSH） and luteinizing hormone （LH） levels were detected by ELISA. At 45 to 65 days after modeling，the uterus and ovary of mice were taken every 5 days，and the organ index was calculated after weighing，and morphological changes in the ovaries were observed by hematoxylin-eosin （HE） staining. 2）Therapeutic effect of Wuji Baifeng Tablets on POI mice：Another sixty 8 week old female Kuming mice were selected according to the above modelling method. After successful modeling，they were randomly divided into MOD group，estradiol valerate （E2） group [0.3 mg/（kg·d）]，and low，medium and high dose Wuji Baifeng Tablets groups[410，820，1640 mg/（kg·d）]，with 10 mice in each group. Once daily for 45 consecutive days，the estrus cycle of mice was observed during administration. The mice were killed on day 46 of administration，and serum FSH and LH levels were detected by ELISA. The uterus and ovaries of mice were taken，and the viscera index was calculated after weighing. The morphological changes in ovaries were observed by HE staining. [Results] 1）Establishment of mouse POI model：compared to the CON group，estrous cycle disorder in the MOD group lasted for 4 weeks on day 55. At day 50 and day 55 of VCD modeling，serum FSH and LH in MOD mice increased significantly twice. The wet weight and organ index of uterus and ovary decreased significantly，and the number of growing follicles decreased significantly. 2）Therapeutic effect of Wuji Baifeng Tablets on POI mice：45 days after intragastric administration，compared with the CON group，the estrous cycle of mice in the MOD group was disordered，and serum FSH and LH were significantly increased. Compared with the MOD group，the estrus cycle of mid-dose and high-dose Wuji Baifeng Tablets returned to normal，and serum FSH and LH of mice in each Wuji Baifeng Tablets dose group decreased significantly. The wet weight and organ index of uterus and ovary decreased significantly，and the number of primordial follicles in the ovary increased significantly. [Conclusion] After 15 days of intraperitoneal injection of 160 mg/（kg·d） VCD，a mouse model of POI could be prepared after 40 days of observation，and a total of 55 days of modeling. Wuji Baifeng Tablets may reduce serum FSH and LH levels by regulating the hypothalamus pituitary ovary axis （HPOA） in mice，regulating the estrous cycle of mice，and promoting follicular development to treat mouse POI caused by VCD.