黄芩苷对多囊卵巢综合征大鼠颗粒细胞增殖和凋亡的影响

Effects of baicalin on proliferation and apoptosis of granulosa cells in rats with polycystic ovary syndrome

目的探讨黄芩苷对多囊卵巢综合征(polycystic ovary syndrome,PCOS)大鼠颗粒细胞增殖和凋亡的影响及对磷脂酰肌醇3-激酶(phosphatidylinositol-3-kinase,PI3K)/蛋白激酶B(protein kinase B,Akt)信号通路的调节作用。方法48只大鼠成功建立PCOS模型,并随机分为模型组、黄芩苷低剂量组、黄芩苷高剂量组和阳性对照组,每组12只,另取12只大鼠作为对照组。黄芩苷低剂量组、黄芩苷高剂量组大鼠分别灌胃25、50 mg∙kg^(−1)黄芩苷,阳性对照组大鼠灌胃0.3392 mg∙kg^(−1)炔雌醇环丙孕酮片(达英-35),对照组和模型组大鼠灌胃等体积生理盐水,每日1次,连续3周。用酶联免疫吸附试验(enzyme linked immuno⁃sorbent assay,ELISA)检测黄体生成素(luteinizing hormone,LH)、卵泡刺激素(follicle stimulating hormone,FSH)、睾酮(testos⁃terone,T)以及雌二醇(estradiol,E2)含量;用HE染色观察卵巢组织病理损伤,细胞增殖检测试剂盒(cell counting kit-8,CCK-8)检测颗粒细胞存活率;用流式细胞仪检测细胞凋亡率,蛋白质印迹法检测PI3K、磷酸化磷脂肌醇3-激酶(phospho PI3K,p-PI3K)、Akt以及p-Akt蛋白相对表达量。结果对照组大鼠可见黄体,未见囊状扩张卵泡;模型组大鼠卵巢体积变大,少见黄体,可见囊状扩张卵泡;黄芩苷低剂量组、黄芩苷高剂量组及阳性对照组大鼠卵巢体积变小,可见黄体,囊状卵泡扩增不明显。与对照组比较,模型组FSH和E2水平、颗粒细胞存活率以及p-PI3K和p-Akt蛋白相对表达量降低,LH、T水平和颗粒细胞凋亡率升高(P<0.05);与模型组比较,黄芩苷低剂量组、黄芩苷高剂量组和阳性对照组FSH、E2水平,颗粒细胞存活率及p-PI3K和p-Akt蛋白相对表达量升高,LH、T水平及颗粒细胞凋亡率降低,其中阳性对照组各指标水平变化最显著,其次是黄芩苷高剂量组(P<0.05)。结论黄芩苷可促进PCOS大鼠卵巢颗粒细胞增殖,抑制凋亡,改善卵巢组织病理损伤,其可能是通过激活PI3K/Akt信号通路发挥作用的。

Objective To investigate the effects of baicalin on the proliferation and apoptosis of granulosa cells in rats with polycystic ovary syndrome(PCOS),and the regulation of phosphatidylinositol-3-kinase(PI3K)/protein kinase B(Akt)signaling pathway.Methods Forty-eight rats were successfully established PCOS model and randomly divided into model group,baicalin low-dose group,baicalin high-dose group and positive control group,12 rats in each group.Another 12 rats were selected as control group.Baicalin low-dose and baicalin high-dose groups were given baicalin 25 mg∙kg^(−1)and 50 mg∙kg^(−1),respectively.The positive control group was given Ethinylestradiol and Cyproterone Acetate Tablets(Diane-35)0.3392 mg∙kg^(−1).The control group and model group were given equal volume of normal saline once a day,for consecutive 3 weeks.The contents of luteinizing hormone(LH),follicle stimulating hormone(FSH),testosterone(T)and estradiol(E2)were detected by enzyme linked immunosorbent assay(ELISA),the pathological damage of ovarian tissue was observed by HE staining,the proliferation rate of granulosa cells was detected by CCK-8,the apoptosis rate was detected by flow cytometry,the relative expression levels of PI3K,p-PI3K,Akt and p-Akt were detected by Western blotting.Results In the control group,luteum was observed,but expanded follicles were not observed.In the model group,the ovarian volume was enlarged,luteum was rare,and cystic dilated follicles were visible.In the low and high dose baicalin groups and the positive control group,the ovarian volume was reduced,luteum was visible,and cystic follicle amplification was not obvious.Compared with control group,the levels of FSH and E2 and the proliferation rate of granulosa cells and the relative expression of p-PI3K and p-Akt protein in model group were decreased,while the levels of LH and T,the apoptosis rate of granulosa cells were increased(P<0.05).Compared with model group,the levels of FSH and E2 and the proliferation rate of granulosa cells and relative expression of p-PI3K and p-Akt protein were increased in baicalin low-dose and high-dose groups and positive control group,while the levels of LH and T,apoptosis rate of granulosa cells decreased,and the changes of all indicators in positive control group were the most significantly.The second group was baicalin high-dose group and baicalin low-dose group(P<0.05).Conclusion Baicalin can promote the proliferation of ovarian granulosa cells in PCOS rats,inhibit apoptosis and improve pathological damage of ovarian tissue,possibly by inhibiting PI3K/Akt signaling pathway.