补肾安胎方及其拆方对控制性超排卵小鼠胚胎着床干预的比较

Comparative Study of Bushenantai Recipe and Its Two Components on Blastocyst Implantation of Controlled Ovarian Hyperstimulation

目的观察补肾安胎方及其补肾、活血组分对控制性超排卵(controlled ovarian hyperstimulation,COH)小鼠胚泡围着床期类肝素结合表皮生长因子(heparin-binding epidermal growth factorlike growthfactor,HB-EGF)及其受体的影响,探讨补肾安胎方及其拆方对着床环节的作用机制。方法将雌性小鼠随机分为正常组(N)、模型组(M)、补肾安胎组(BH)、补肾组(BS)和活血组(HX)。观察妊娠第1天阴栓率、妊娠第6天妊娠率和孕鼠着床位点数;采用反转录聚合酶链反应(RT-PCR)方法测定妊娠第5天、第6天子宫内膜HB-EGF mRNA的表达,采用免疫组织化学法测定同期子宫内膜表皮生长因子受体(epidermal growth factor receptor,EGFR)蛋白的表达。结果模型组阴栓率、妊娠率较正常组显著降低(P<0.01),着床位点数显著升高(P<0.01);中药各组治疗后,阴栓率、妊娠率均较模型组显著升高(P<0.05,P<0.01),但着床位点数均显著降低(P<0.05,P<0.01),以补肾安胎组作用最为明显;活血组阴栓率高于补肾组(P<0.05),着床位点数较补肾组降低(P<0.01),妊娠率两组比较,差异无统计学意义。正常组HB-EGF mRNA表达最高,模型组显著低于正常组(P<0.01),各中药组均显著高于模型组(P<0.01);各中药组HB-EGF mRNA表达比较,补肾安胎组明显高于活血组和补肾组(P<0.05),活血组高于补肾组(P<0.05);妊娠第6天,HB-EGF mRNA表达量较妊娠第5天降低。EGFR蛋白在围着床期子宫内膜腺体和基质细胞胞质中表达,各组小鼠EGFR蛋白表达空间分布差异无统计学意义。正常组EGFR蛋白表达丰富;模型组表达较正常组显著减少(P<0.01),部分缺失;中药各组EGFR表达量均高于模型组(P<0.01);其中补肾活血组高于活血组和补肾组(P<0.05,P<0.01),活血组高于补肾组(P<0.05,P<0.01)。结论补肾安胎方及补肾、活血组分均能够改善控制性超排卵小鼠的着床率和妊娠率,促进胚泡生长和HB-EGF及其受体的表达,其中以补肾安胎全方作用最佳,补肾、活血组分在改善胚泡着床过程中具有协同作用。

Objective To investigate the role of Bushenantai recipe and its two components on the expression of heparin-binding epidermal growth factor-like growth factor（HB-EGF）and its receptor during blastocyst implantation of mice with controlled ovarian hyperstimulation（COH）,and to probe the mechanisms of Bushenantai recipe and its two components on blastocyst implantation.Methods All mice were randomly divided into normal group（N）,model group（M）,Bushenantai group（BH）,Bushen group（BS）and Huoxue group（HX）.Except for the normal group,the other four groups were intraperitoneal injected with pregnant mare serum gonadotropin（PMSG）and human chorionic gonadotropin（HCG）for inducing COH.All groups mated with fertile males.The vaginal plug rate on pregnancy day 1,and pregnancy rate and the number of implantation sites on pregnancy day 6 were observed.The uterus was collected on pregnancy day 5 and 6.The expression of HB-EGF mRNA of the uterus was detected by RT-PCR.The protein expression of epidermal growth factor receptor（EGFR）was analyzed using immunohistochemistry.Results The vaginal plug and pregnancy rates in the model group were significantly lower（P0.01）whereas the number of implantation sites in the model group increased significantly（P0.01）when compared to the normal group.The vaginal plug rate and pregnancy rate in the TCM groups（including Bushenantai group,Bushen group and Huoxue group）were all significantly higher than those in the model group（P0.05,P0.01）;the vaginal plug rate in Bushenantai group was higher than the Bushen and Huoxue groups（P0.05,P0.01）.The vaginal plug rate in the Huoxue group was higher than the Bushen group（P0.05）,but the number of implantation sites showed lower（P0.01）.The expressions of HB-EGF mRNA and EGFR protein in the endometrium of model group were significantly lower than those in the normal group（P0.01）.The expressions of HB-EGF mRNA and EGFR protein in the TCM groups were higher than those in the model group（P0.01）.HB-EGF mRNA and EGFR protein in the Huoxue group showed better than the Bushen group（P0.05,P0.01）,but lower than the Bushenantai group（P0.05,P0.01）.Conclusion Bushenantai recipe and its Bushen,Huoxue components could improve the vaginal plug rates and pregnancy rates of COH mice,improve the blastocyst growth,and up-regulated the expression of HB-EGF mRNA and EGFR protein.Bushen and Huoxue components had some cooperative effects on blastocyst implantation.