毓麟珠对卵巢早衰大鼠SIRT1-FoxO1-自噬通路的调控作用  

Regulatory effect of Yulinzhu on SIRT1-FoxO1-autophagy pathway in rats with premature ovarian failure

作　　者：何啸兰[1] 王鹍鹏[2] 周月希 吴尚蓉 胡雅君[1] HE Xiao-lan;WANG Kun-peng;ZHOU Yue-xi;WU Shang-rong;HU Ya-jun(Department of Reproductive Medicine,Wuhan Hospital of Integrated Traditional Chinese and Western Medicine,Wuhan 430030,China;Wuhan Hospital of Traditional Chinese Medicine,Wuhan 430030,China)

机构地区：[1]武汉市中西医结合医院生殖医学科,湖北武汉430030 [2]武汉市中医医院,湖北武汉430030

出　　处：《中国病理生理杂志》2022年第6期1091-1104,共14页Chinese Journal of Pathophysiology

基　　金：湖北省卫健委中医药科研项目(No.ZY2019M005)。

摘　　要：目的:探讨毓麟珠对卵巢早衰(POF)大鼠自噬的影响,并分析其防治POF的作用机制。方法:雌性SD大鼠采用随机数字表法选取16只为正常组,其余大鼠通过腹腔注射环磷酰胺建立POF模型。造模成功大鼠分为POF组、戊酸雌二醇(0.09 mg/kg)组(简称雌二醇组)、低剂量(7.56 g/kg)毓麟珠组、高剂量(15.12 g/kg)毓麟珠组和毓麟珠+沉默信息调节因子1(SIRT1)抑制剂EX527组(简称毓麟珠+EX527组),每组16只。低、高剂量毓麟珠组大鼠分别灌胃相应剂量的毓麟珠药液,雌二醇组大鼠给予戊酸雌二醇溶液灌胃,毓麟珠+EX527组大鼠在给予15.12 g/kg毓麟珠药液灌胃的同时腹腔注射10 mg/kg EX527,正常组和POF组大鼠给予等体积的生理盐水干预,每天1次,连续给药3周。给药结束后,记录各组大鼠的双侧卵巢重量和体重,计算卵巢指数;ELISA检测各组大鼠血清雌二醇(E2)、促卵泡激素(FSH)和抗缪勒管激素(AMH)水平;检测各组大鼠卵巢组织丙二醛(MDA)、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)水平;苏木精-伊红(HE)染色和TUNEL染色观察各组大鼠卵巢组织病理变化,计数各级卵泡数,并分析窦卵泡中颗粒细胞凋亡水平;免疫组织化学法检测各组大鼠卵巢组织微管相关蛋白1轻链3B(LC3B)蛋白表达;RT-qPCR检测各组大鼠卵巢组织LC3B、p62、beclin-1、Atg5和Atg7的mRNA表达;Western blot检测各组大鼠卵巢组织LC3A/B、p62、beclin-1、Atg5、Atg7、SIRT1、叉头框蛋白O1(FoxO1)和乙酰化FoxO1(Ac-FoxO1)的蛋白水平;评估POF大鼠的生育能力。结果:与正常组相比,POF组大鼠卵巢指数,血清E2和AMH水平,卵巢组织SOD和CAT活性,卵巢原始卵泡、初级卵泡、次级卵泡和窦卵泡数量,p62 mRNA和蛋白水平,SIRT1和FoxO1蛋白水平,以及受孕率和胚胎数均显著降低,而血清FSH水平,卵巢组织MDA水平,闭锁卵泡数量,颗粒细胞凋亡水平,卵巢LC3B蛋白阳性表达,LC3B、beclin-1、Atg5和Atg7 mRNA水平,beclin-1、Atg5�AIM:To investigate the effect of Yulinzhu on autophagy of premature ovarian failure(POF)rats and analyze its mechanism of preventing and treating POF.METHODS:Female SD rats were randomly divided into nor⁃mal group,POF group,estradiol valerate group(estradiol group),low-dose Yulinzhu group,high-dose Yulinzhu group,and Yulinzhu+silent information regulator 1(SIRT1)inhibitor EX527 group(Yulinzhu+EX527 group),with 16 rats in each group.The rats in low-and high-dose Yulinzhu groups were given Yulinzhu liquid by gavage at 7.56 and 15.12 g/kg,respectively.The rats in estradiol group were given estradiol valerate solution by gavage,and the rats in Yulinzhu+EX527 group were given 15.12 g/kg of Yulinzhu liquid by gavage and 10 mg/kg EX527 by intraperitoneal injection.The rats in normal group and POF group were given an equal volume of normal saline.These treatments were conducted once per day for 3 weeks.After the administration,the weight of both ovaries and body weight were recorded,and the ovarian index was calculated.ELISA was used to detect the serum levels of estradiol(E2),follicle-stimulating hormone(FSH)and anti-Müllerian hormone(AMH)of the rats in each group.The levels of malondialdehyde(MDA),superoxide dis⁃mutase(SOD)and catalase(CAT)in the ovarian tissue of rats in each group were detected.Hematoxylin-eosin(HE)staining and TUNEL staining were used to observe the pathological changes of the ovarian tissue of rats in each group.The numbers of follicles at various levels were counted,and the level of granulosa cell apoptosis in antral follicles was ana⁃lyzed.Immunohistochemical method was used to detect the expression of microtubule-associated protein 1 light chain 3B(LC3B)protein in ovarian tissues of rats in each group.RT-qPCR was used to detect the mRNA expression of LC3B,p62,beclin-1,Atg5 and Atg7 in ovarian tissues of rats in each group.Western blot was used to detect the protein levels of LC3A/B,p62,beclin-1,Atg5,Atg7,SIRT1,forkhead box protein O1(FoxO1)and acetylated FoxO1(Ac-FoxO1)in ovarian tissues

关 键 词：毓麟珠 卵巢早衰 氧化应激 自噬 SIRT1-FoxO1信号通路 

分 类 号：R711.75[医药卫生—妇产科学] R363.2[医药卫生—临床医学]