电针对多囊卵巢综合征大鼠卵巢细胞分泌功能及kisspeptin/kiss1r系统的影响

Effects of electroacupuncture on the secretion function of ovarian cells and kisspeptin/kiss1r system in rats with polycystic ovarian syndrome

目的:观察电针对多囊卵巢综合征(PCOS)大鼠卵巢颗粒细胞与膜细胞分泌激素功能及kisspeptin与kiss1r表达的影响,探讨电针改善PCOS大鼠卵巢功能障碍的机制。方法:雌性SD大鼠随机分为对照组、模型组、电针组和氟他胺组,每组12只。采用0.1 mg/mL来曲唑(10 mL·kg^(-1)·d^(-1))灌胃复制PCOS大鼠模型。电针组电针大鼠“关元”,每次20 min,氟他胺组给予氟他胺溶液(50 mg·kg^(-1)·d^(-1))灌胃,两组均每日治疗1次,共治疗14 d。造模后和治疗后称量大鼠体质量及卵巢质量并计算卵巢指数;HE染色法观察大鼠卵巢组织形态改变;ELISA法检测血清睾酮(T)、促黄体生成素(LH)、雌二醇(E2)含量,卵巢颗粒细胞及膜细胞培养液中E2、T含量及卵巢组织kisspeptin的含量;免疫组织化学法检测大鼠卵巢中kisspeptin阳性表达;Western blot法检测卵巢组织kiss1r蛋白表达量。结果:与对照组比较,模型组大鼠体质量、卵巢质量及卵巢指数,血清中T、LH及膜细胞培养液中T含量,卵巢中kisspeptin含量及阳性表达均升高(P<0.01,P<0.05),颗粒细胞培养液中E2含量降低(P<0.01)。与模型组比较,电针组与氟他胺组大鼠体质量、卵巢质量及卵巢指数、血清中T与LH、膜细胞培养液中T、卵巢中kisspeptin含量与阳性表达均降低(P<0.01,P<0.05),颗粒细胞培养液中E2含量升高(P<0.05,P<0.01)。结论:电针可调节PCOS大鼠血清性激素水平、卵巢颗粒细胞和膜细胞的分泌功能及卵巢kisspeptin蛋白表达,表现出与雄激素受体阻断干预类似的效应,提示电针对PCOS大鼠卵巢功能障碍的改善作用可能与kisspeptin/kiss1r系统有关。

Objective To observe the effects of electroacupuncture(EA) on hormone secretion function of ovarian granulosa cells and theca cells,as well as the expression changes of kisspeptin and kiss1r in rats with polycystic ovarian syndrome(PCOS),so as to explore the mechanism of EA for relieving ovarian dysfunction in PCOS rats.Methods Forty-eight SD female rats were randomly divided into control group,model group,EA group and flutamide group,with 12 rats in each group.PCOS rat model was replicated with the gavage of letrozole(0.1 mg/mL,10 mL·kg^(-1)·d^(-1)).In the EA group,EA(2 Hz,2 mA) was used to stimulate “Guanyuan”(CV4) for 20 min each time.In the flutamide group,flutamide solution(50 mg·kg^(-1)·d^(-1)) was administrated by gavage.The treatments were given once daily for 14days in each group.After the modeling and treatment,the body and ovarian weights of the rats were measured,and the ovarian index was calculated.Using HE staining,the morphological changes of ovary were observed.ELISA was adopted to detect the contents of testosterone(T),luteinizing hormone(LH) and estradiol(E2) in serum,the contents of E2 and T in the culture medium of ovarian granulosa cells and theca cells,as well as the content of kisspeptin in the ovarian tissue.The positive expression of kisspeptin in ovary was observed by immunohistochemical method,and the protein expression of its receptor kiss1r was detected by Western blot.Results Compared with the control group,the body and ovarian weights,ovarian index,the contents of T and LH in serum and that of T in the culture medium of theca cells,as well as the content and positive expression of kisspeptin in ovary were all increased(P<0.01,P<0.05);and the content of E2 in the culture medium of granulosa cells was decreased(P<0.01) in the model group.When compared with the model group,in the EA and flutamide groups,the body and ovarian weights,ovarian index,the contents of T and LH in serum and that of T in the culture medium of theca cells,as well as the content and expression of kisspeptin in ovary were all decreased(P<0.01,P<0.05);and the content of E2 in the culture medium of granulosa cells was increased(P<0.05,P<0.01).Conclusion EA regulates the serum sex hormone levels,the secretion function of the ovarian granulosa cells and theca cells,and the ovarian kisspeptin/kiss1r protein expression in PCOS rats,showing the similar effect as receptor blockade intervention.It is suggested that the improvement of EA in ovarian dysfunction of PCOS rats may be related to the kisspeptin/kiss1r system.