【摘要】 目的评价解郁育胞方对慢性应激大鼠着床窗期外周血Thl7、Treg细胞及着床窗子宫内膜RORγt、Foxp3的表达,从免疫炎症角度探讨其改善母胎界面微环境的作用机制。方法 SPF级雌性SD大鼠40只,按体质量分层法随机分为4组:正常对照组、应激组、应激+泼尼松组、应激+解郁育胞方组,每组10只。除正常对照组外,其余3组大鼠建立慢性应激模型。于刺激第15天开始,每天9:00应激+泼尼松组给予波尼松溶液灌胃,正常对照组、应激组灌服等体积纯化水;应激+解郁育胞方组灌服解郁育胞方溶液。4组大鼠分别于第28天16:00按照雌∶雄=2∶1合笼,经阴道涂片确认妊娠后第4天停药,当日16:00处死大鼠,釆用流式细胞技术检测着床窗外周血Thl7、Treg细胞;同时采用免疫组化技术检测着床窗子宫内膜RORγt、Foxp3的表达。结果①Foxp3的表达:与正常对照组比较,应激组Foxp3的表达显著降低(P<0.05),应激+解郁育胞方组差异无统计学意义(P>0.05)。②RORγt的表达:与正常对照组比较,应激组子宫内膜RORγt的表达显著增加(P<0.05),应激+解郁育胞方组差异无统计学意义(P>0.05)。③外周血Th17、Treg亚群比例及比值比较:与应激组比较,其余3组Th17亚群细胞比例均明显降低(P<0.05);与应激组比较,应激+泼尼松组、应激+解郁育胞方组Treg亚群细胞比例明显增多(P<0.05)。正常对照组与应激+解郁育胞方组Th17/Treg比值差异无统计学意义(P>0.05);与应激组比较,其余3组Th17/Treg比值均有明显差异(P<0.05)。④相关性分析:Th17/Treg比值与RORγt表达呈正相关(r=0.608,P<0.05),Th17/Treg比值和T Foxp3的表达呈负相关(r=-0.389,P<0.05)。结论①解郁育胞方能降低慢性应激大鼠外周血Th17/Treg比值,降低应激对机体的炎症损伤,增加机体抗应激能力。②大剂量解郁育胞方能明显提高慢性应激大鼠着床窗子宫内膜中Foxp3表达水平,降低RORγt的表达。③解郁育胞方可能通过平衡体内由于紧张应激导致的Th17/Treg失衡,改善子宫内膜炎症反应,从而改善着床微环境,提高妊娠率。更多还原

【Abstract】 Objective To evaluate the effect of Jieyuyubao recipe on expression of RORγt and Foxp3 in peripheral blood Thl7,Treg cells in endometrium of chronically stressed rats during the embryo implantation stage,and to reveal the effect of improving the microenvironment of maternal-fetal interface from the perspective of immune inflammation mechanism. Methods Forty specific-pathogen-free( SPF) quality female Sprague Dawley( SD) rats were randomly divided into 4 groups( 10 in each groups) according to the body mass stratification method: normal control group,stress group,stress + prednisone group,and stress + Jieyuyubao group. Expect for the normal control group,the chronic stress model was established in the other three groups.On the 15 th day after stimulation,the stress + prednisone group was administered with prednisone solution at 9: 00 daily; normal control group,stress group irrigation serve an equal volume of purified water; the stress + Jieyuyubao group serve Jieyuyubao solution. The male and female rats( male/female ratio was 1∶2) mated since 16: 00 of 28 th day in all 4 groups. The vaginal smear was used to confirm pregnancy,and irrigation was stopped on the 4 th day after the pregnancy. The rats were sacrificed at16: 00 on the same day,and the flow cytometry was used to detect the Thl7 and Treg cells in the peripheral blood. The expression of RORγt and Foxp3 in the endometrium was detected by immunohistochemistry. Results ①Compared with the normal control group,the expression of Foxp3 was significantly decreased in the stress group( P < 0. 05). There was no significant difference between the stress + Jieyuyubao group and the normal control group( P>0.05). ②Compared with the normal control group,the expression of RORγt in the endometrium of the stress group was significantly increased( P < 0. 05). There was no significant difference between the stress + Jieyuyubao group and the normal control group( P>0.05). ③Compared with the ratio of Th17 cell in the stress group,the ratio in the normal control group,Stress + prednisone group,and the stress + Jieyuyubao group was significantly lower( P<0.05). In addition,the ratio of Treg cells in the stress + prednisone group and the stress + Jieyuyubao group increased significantly comparing to the stress group( P<0.05). There was no significant difference in Th17/Treg between the control group and the stress + Jieyuyubao group( P>0.05). Compared with the stress group,the Th17/Treg of the remaining three groups was significantly different( P<0.05).④Th17/Treg ratio was positively correlated with RORγt expression( r = 0.608,P< 0. 05),and Th17/Treg ratio was negatively correlated with T Foxp3 expression( r =-0. 389,P < 0. 05). Conclusion Jieyuyubao recipe can effectively reduce the ratio of Th17/Treg in peripheral blood of rats with chronic stress,reduce the inflammatory damage,and increase the anti-stress ability of the body. High dose of Jieyuyubao recipe can significantly increase Foxp3 and reduce RORγt of endometrium in implantation window. We speculate that Jieyuyubao improves the implantation of the endometrial inflammatory response by balancing the imbalance of Th17/Treg caused by stress,thereby improving the implantation microenvironment and increasing the pregnancy rate.更多还原