雌激素对宫腔粘连子宫整合素αvβ3及胸苷磷酸化酶表达的影响

Effects of Estrogen on Integrinαvβ3and Thymidine Phosphorylase Expression in Uterine Adhesions

目的揭示整合素αvβ3及胸苷磷酸化酶(thymidine phosphorylase,TYMP)在治疗宫腔粘连(intrauterine adhesion,IUA)中的作用,探索雌激素改善IUA大鼠子宫内膜组织容受性的可能机制,为临床上治疗IUA提供方法。方法将18只SD雌性大鼠适应性喂养后,取6只做体外实验,其余12只随机分为模型组和治疗组,每组6只。所有大鼠于动情期以95%乙醇损伤双侧宫腔造模,造模成功后第14天,模型组开始予以生理盐水2 ml灌胃,治疗组予以雌激素2 ml灌胃。造模28天后处死模型组及治疗组大鼠,取出子宫经甲醛固定后行苏木精-伊红(hematoxylin-eosin,HE)染色,观察各组大鼠子宫内膜组织形态,行免疫组织化学法观察整合素αvβ3及TYMP在各组宫腔中的表达情况。体外实验组大鼠于造模成功后第14天处死,将子宫取出分为2 cm大小一段,将子宫组织置于雌激素培养液中,在处理后的0min、30 min、45 min、60 min行Western blot检测整合素αvβ3及TYMP的蛋白表达水平。选取行宫腔镜检查发现宫腔形态正常的患者及IUA患者各15例,通过宫腔灌洗收集宫腔灌洗液,将采集的临床标本采用实时荧光定量聚合酶链式反应(quantitative real-time-polymerase chain reaction,qPCR)检测临床标本中整合素αvβ3及TYMP mRNA的相对表达量。结果HE染色发现,与模型组相比,治疗组的宫腔形态得到了一定程度的改善,内膜厚度增加,间质中有大量排列整齐的腺体,可见新生血管。免疫组织化学染色结果提示,与模型组相比,治疗组的整合素αvβ3及TYMP的表达水平均明显升高。大鼠子宫体外实验雌激素处理30 min后TYMP的表达显著高于雌激素处理0min,差异具有统计学意义(P<0.05);雌激素处理30 min、45 min、60 min后整合素αvβ3的表达显著高于雌激素处理0min,差异均有统计学意义(P<0.05)。粘连子宫内膜中的整合素αvβ3及TYMP mRNA表达水平明显低于正常宫腔患者,差异均具有统计学意义(P<0.05)。结论整合素αvβ3及TYMP蛋白可作为子宫内膜容受性的评价指标。雌激素可能通过上调整合素αvβ3及TYMP蛋白的表达,促进子宫内膜生长,有效缓解IUA情况,改善子宫内膜容受性。

Objective To reveal the role of integrinαvβ3 and thymidine phosphorylase(TYMP)protein in the treatment of intrauterine adhesion(IUA),to explore the possible mechanisms by which estrogen improves endometrial tissue tolerance in IUA rats,and to provide methods for the clinical treatment of IUA.Methods A total of 6 of 18 SD female rats were used for in vitro experiments after adaptive feeding,and the remaining 12 rats were randomly divided into model group and treatment group,with 6 in each.All rats were damaged with95%ethanol to create membranes in both uterine cavities during the estrous period,on the 14th day after successful modeling,the model group was given saline 2 ml by gavage and the treatment group was given estrogen2 ml by gavage.After 28 days of membrane creation,the rats in the model and treatment groups were executed,and the uterus was removed and fixed in formaldehyde and stained with hematoxylin-eosin(HE)staining to observe the endometrial tissue morphology of each group,the expression of integrinαvβ3 and TYMP in uterine cavity of each group was observed by immunohistochemistry.The rats in the in vitro experimental group were executed on the 14th day after successful membrane formation,and the uterus were divided into a 2 cm section,and the uterine tissues were placed in estrogen culture solution,and the expression levels of integrinαvβ3 and TYMP protein were detected by Western blot at 0min,30 min,45 min and 60 min after experiments.A total of 15 patients with normal uterine morphology and 15 patients with IUA were selected,uterine lavage fluid was collected by uterine lavage,and the relative expression levels of integrinαvβ3 and TYMP mRNA in clinical specimens were detected by quantitative real-time-polymerase chain reac-tion(qPCR).Results The HE staining revealed that the morphology of the uterine cavity was improved to some extent in the treatment group compared with the model group,with increased endometrial thickness,a large number of well-arranged glands in the interstitium and visible neovascularization.Immunohistochemical staining showed that the expression of integrinαvβ3 and TYMP were significantly higher in the treatment group compared with the model group.The expression of TYMP was significantly higher after 30 min of estrogen treatment than after 0min of estrogen treatment in rats uterus in vitro,and the difference was statistically significant(P<0.05);the expression of integrinαvβ3 was significantly higher after 30 min,45 min and 60 min of estrogen treatment than after 0min of estrogen treatment,and4.0 c3 the difference was statistically significant(P<0.05).The expression levels of integrinαvβ3 and TYMP mRNA in the adherent endometrium were significantly lower than those in patients with normal uterine cavity,and the differences were all statistically significant(P<0.05).Conclusion Integrinαvβ3 and TYMP protein can be used as indicators for the evaluation of endometrial tolerability.Estrogen may promote endometrial growth through upregulating the expression of integrinαvβ3 and TYMP protein,effectively relieve IUA and improve endometrial tolerance.