摘要
目的探究褪黑素通过抑制PTEN/AKT/FOXO3a通路在小鼠卵巢中的激活遏制慢性应激下小鼠卵泡发育不良的机制。方法选取60只6周龄雌性C57BL/6N小鼠,随机分为正常对照组、慢性应激组和褪黑素组,每组各20只。使用实验室称重仪记录小鼠的身体和卵巢的质量。通过酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)试剂盒检测小鼠血清激素水平。通过组织学分析计算小鼠卵巢不同阶段的卵泡数量。通过ELISA和实时定量聚合酶链式反应(real-timefluorescence quantitative polymerase chain reaction,RT-qPCR)检测小鼠血清抗米勒管激素(anti-Müllerianhormone,AMH)的表达。通过蛋白印迹法检测磷脂酰肌醇-3-激酶(phosphatidylinositol-3-kinase)/蛋白激酶B(protein kinase B,AKT)/叉头框转录因子3a(forkhead box transcription factor 3a,FOXO3a)信号通路的表达。通过PCR检测小鼠卵巢PI3K/AKT/FOXO3a的转录水平。统计学方法采用LSD-t检验、χ^(2)检验、单因素方差分析或重复测量资料方差分析。结果慢性应激组与正常对照组的初级卵泡数量[(174±30)个与(107±17)个,t=-148.098]、磷酸化张力蛋白同源物(p-phosphatase tensinhomolog deleted on chromosome ten,p-PTEN)蛋白(2.03±0.19与1.26±0.16,t=-32.207)、p-AKT蛋白(1.99±0.18与1.07±0.05,t=-70.021)、p-FOXO3a蛋白(2.18±0.20与1.18±0.11,t=-64.621)、皮质醇稳定蛋白(cortistatin,CORT)(137±12与83±7,t=-124.235)、促卵泡激素(follicle-stimulating hormone,FSH)[(13.1±1.9)IU/L与(8.2±1.6)IU/L,t=-25.388]比较,慢性应激组水平高于正常对照组(P值均<0.05)。慢性应激组与正常对照组的小鼠身体质量[(22.5±2.5)g与(28.4±4.7)g,t=12.906]、卵巢质量[(9±3)mg与(23±5)mg,t=45.302]、雌二醇水平[(36±8)μg/L与(75±10)μg/L,t=88.937]、雄激素水平[(0.13±0.01)μg/L与(0.20±0.02)μg/L,t=23.123]、促黄体生成素(luteinizing hormone,LH)水平[(3.2±0.3)IU/L与(4.6±0.5)IU/L,t=31.170]以及原始卵泡数量[(87±9)个与(143±2
Objective To explore the mechanism of melatonin inhibiting chronic stress-induced folliculardysplasia by inhibiting the activation of PTEN/AKT/FOXO3a pathway in ovaries of mice.Method Sixty6-week-old female C57BL/6N mice were randomly divided into normal control group,chronic stress group,and melatonin group,20 mouse in each group.The body mass and ovarian mass of the mice were recordedusing a laboratory balance.Enzyme-linked immunosorbent assay(ELISA)kits were used to measure theserum hormone levels in mice.Histological analysis was performed to determine the number of folliclesat different stages in ovaries of mice.ELISA and real-time fluorescence quantitative polymerase chainreaction(RT-qPCR)were used to detect the expression of anti-Müllerian hormone(AMH)in serum of mice.Western blotting analysis was conducted to examine the expression of the phosphatidylinositol 3 kinase(PI3K)/protein kinase B(AKT)/Forkhead box transcription factor 3a(FOXO3a)signaling pathway.PCR was used to measure the transcription levels of PI3K/AKT/FOXO3a in ovaries of mice.Statistical methodswere used for LSD t-test,χ^(2) tests,one-way analysis of variance(ANOVA)or repeated measurement data analysisof variance.Result Chronic stress group were higher than those with normal control group in number ofprimary follicles(174±30 vs 107±17,t=-148.098),p-phosphatase and tensin homolog deleted on chromosometen(p-PTEN)proteins(2.03±0.19 vs 1.26±0.16,t=-32.207),p-AKT proteins(1.99±0.18 vs 1.07±0.05,t=-70.021),p-FOXO3a proteins(2.18±0.20 vs 1.18±0.11,t=-64.621),cortistatin(CORT)(137±12 vs83±7,t=-124.235),follicle-stimulating hormone(FSH)[(13.1±1.9)IU/L vs(8.2±1.6)IU/L,t=-25.388](allP<0.001).Chronic stress group were lower than those with normal control group in body mass[(22.5±2.5)gvs(28.4±4.7)g,t=12.906],ovarian mass[(9±3)mg vs(23±5)mg,t=45.302],estradiol levels[(36±8)μg/Lvs(75±10)μg/L,t=88.937],androgen levels[(0.13±0.01)μg/L vs(0.20±0.02)μg/L,t=23.123],luteinizinghormone(LH)levels[(3.2±0.3)IU/L vs(4.6±0.5)IU/L,t=31.170
作者
席红
王艳秋
余红琴
董薇
Xi Hong;Wang Yanqiu;Yu Hongqin;Dong Wei(Department of Gynaecology,Panjin Liaoyou Baoshihua Hospital,Liaoning,Panjin 124010,China)
出处
《发育医学电子杂志》
2024年第3期210-216,223,共8页
Journal of Developmental Medicine (Electronic Version)
