电针对宫腔粘连大鼠子宫内膜修复的效应观察

Observation on the effect of electroacupuncture on the regeneration of endometrium in rats with intrauterine adhesion

目的:观察电针对宫腔粘连(IUA)大鼠子宫内膜纤维化程度及炎性反应的影响,探讨电针修复子宫内膜的可能机制。方法:雌性SD大鼠随机分为空白组、模型组、电针组,每组15只。采用机械搔刮联合脂多糖感染双重损伤法建立IUA大鼠模型。电针组予“关元”针刺、双侧“子宫”“三阴交”电针治疗,15min/次,1次/d,连续干预2个动情周期。每组5只大鼠于动情期取材,HE染色法观察子宫内膜形态及腺体数目变化,Masson染色法观察子宫内膜纤维化面积,免疫组织化学法检测子宫内膜组织中Ⅰ型胶原蛋白(Col-Ⅰ)、转化生长因子-β1(TGF-β1)蛋白阳性表达,Westernblot法检测子宫组织中整合素αγβ3(Integrinαγβ3)的蛋白表达量,ELISA法检测大鼠子宫组织中白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)的含量;每组剩余10只大鼠于妊娠第8天取出子宫组织,比较各组大鼠子宫内胚胎着床数目。结果:空白组大鼠动情期的子宫组织结构完整,内膜层清晰,宫腔通畅,形态规则,腺体密集;模型组大鼠动情期的子宫内膜层被破坏,宫腔明显狭窄呈粘连状态,腺体稀疏;电针组大鼠动情期的子宫组织结构基本完整,部分区域新生腺体较多,趋近于正常。与空白组相比,模型组大鼠子宫内膜腺体数量明显减少(P<0.01),子宫内膜纤维化面积比值升高(P<0.01),子宫内膜组织中Col-Ⅰ、TGF-β1蛋白阳性表达升高(P<0.01),子宫组织Integrinαγβ3蛋白表达水平降低(P<0.01),IL-1β、TNF-α含量升高(P<0.01),损伤侧子宫胚胎着床数明显减少(P<0.01);与模型组相比,电针组大鼠子宫内膜腺体数目显著增多(P<0.01),子宫内膜纤维化面积比值明显降低(P<0.01),子宫内膜组织中Col-Ⅰ、TGF-β1蛋白阳性表达降低(P<0.01,P<0.05),子宫组织Integrinαγβ3蛋白表达水平升高(P<0.01),IL-1β、TNF-α含量降低(P<0.01),损伤侧子宫胚胎着床数明显增加(P<0.05)。结论:电针能够改善IUA大鼠子宫内膜纤维化,降低IUA大鼠子宫组织炎性反应,增强子宫内膜容受性,促进子宫内膜修复,有利于胚胎着床。

Objective To observe the effect of electroacupuncture(EA)on the degree of endometrial fibrosis and inflammatory response in the rat model of intrauterine adhesion(IUA),so as to explore the possible mechanism of EA underlying improving IUA and promoting endometrium regeneration.Methods Forty-five female SD rats were randomly divided into blank,model and EA groups,with 15 rats in each group.The IUA model was established by mechanical scratching combined with lipopolysaccharide infection.EA was applied to bilateral“Zigong”(EX-CA1)and“Sanyinjiao”(SP6),with acupuncture applied to“Guanyuan”(CV4)for rats in the EA group,started from the 2(nd)day after modeling,15 minutes every time,once a day for 2 consecutive estrous cycles.Samples from 5 rats in each group were collected during estrus period.Changes of endometrial histopathology and number of glands were observed after HE staining.The area of endometrial fibrosis was observed and calculated after Masson staining.The positive expressions of collagen type I(Col-I)and transforming growth factorβ1(TGF-β1)proteins in endometrial tissue were detected by immunohistochemistry method.The protein expression of integrinαγβ3 in uterine tissue was detected by Western blot.The contents of interleukin(IL)-1βand tumor necrosis factorα(TNF-α)in uterine tissue were detected by ELISA.Samples from remaining 10 rats in each group were collected on the 8(nd)day after modeling,15 minutes every time,once a day for 2 consecutive estrous cycles.Samples from 5 rats in each group were collected during estrus period.Changes of endometrial histopathology and number of glands were observed after HE staining.The area of endometrial fibrosis was observed and calculated after Masson staining.The positive expressions of collagen type I(Col-I)and transforming growth factorβ1(TGF-β1)proteins in endometrial tissue were detected by immunohistochemistry method.The protein expression of integrinαγβ3 in uterine tissue was detected by Western blot.The contents of interleukin(IL)-1βand tumor necrosis factorα(TNF-α)in uterine tissue were detected by ELISA.Samples from remaining 10 rats in each group were collected on the 8(th)day of gestation for calculation of the embryo implantation numbers of the rats.Results HE staining showed complete uterine tissue structure of the rats in the blank group during estrus period,with clear endometrial layer,unobstructed and regular uterine cavity,and dense glands.Destroyed endometrial layer,narrowed and adhered uterine cavity,and sparse glands of the rats were seen in the model group,which was relatively milder in the EA group.Following modeling,the number of endometrial glands,the protein expression of Integrinαγβ3,the number of implanted uterine embryos on the injured side of the model group were significantly decreased(P<0.01),while the area of endometrial fibrosis,the positive expressions of Col-I and TGF-β1 proteins,and the contents of IL-1βand TNF-αin the uterine tissue were significantly increased(P<0.01)in comparison with those in the blank group.After intervention,the number of endometrial glands,the protein expression of Integrinαγβ3,the number of implanted uterine embryos on the injured side of the EA group were significantly increased(P<0.01,P<0.05),while the area of endometrial fibrosis,the positive expressions of Col-I and TGF-β1 proteins,and the contents of IL-1βand TNF-αin the uterine tissue were significantly decreased(P<0.01,P<0.05)compared with the model group.Conclusion EA can enhance endometrial receptivity,and promote endometrial regeneration,be conducive to embryo implantation in IUA model rats,which may be related to its effect in alleviating endometrial fibrosis and reducing inflammatory response.