摘要
目的:探讨八珍汤合归肾丸加减方通过调节1型辅助性T细胞与2型辅助性T细胞(Th1/Th2)免疫平衡对胚胎着床障碍模型小鼠的助孕机制。方法:将80只ICR雌鼠于受孕第1天按随机数字表法分为正常组、模型组、西药组和中药组,每组20只。除正常组外,各组小鼠在受孕第4天灌胃米非司酮溶液(0.2 mg/只)制备胚胎着床障碍模型。中药组灌胃八珍汤合归肾丸加减方水煎液(20.8 g·kg^(-1));西药组灌胃地屈孕酮溶液(3.9 mg·kg^(-1));正常组与模型组灌胃等体积生理盐水,并于受孕第1天开始给药,每日1次,干预至取材前1 d。通过肉眼观察小鼠着床点数并计算受孕率;称量各组小鼠体质量;苏木素-伊红(HE)染色观察子宫内膜病理形态;蛋白免疫印迹法(Western blot)检测子宫组织T盒子转录因子(T-bet)、GATA结合蛋白3(GATA3)、γ干扰素(IFN-γ)及白细胞介素-4(IL-4)蛋白表达;实时荧光定量聚合酶链式反应(Real-time PCR)检测子宫组织T-bet、GATA3、IFN-γ、IL-4 mRNA表达;酶联免疫吸附测定法(ELISA)检测血清IFN-γ和IL-4水平;计算T-bet/GATA3、IFN-γ/IL-4值,评估Th1/Th2免疫平衡。结果:与正常组比较,模型组受孕率变化差异无统计学意义,平均着床点数及体质量显著降低(P<0.01);子宫内膜腺体稀少,间质致密,核下未见空泡;子宫组织中T-bet、IFN-γ表达明显升高(P<0.05,P<0.01),GATA3、IL-4表达明显降低(P<0.05,P<0.01);血清中IFN-γ水平显著升高(P<0.01),IL-4水平显著降低(P<0.01);Th1/Th2比值(T-bet/GATA3、IFN-γ/IL-4)显著升高(P<0.01)。与模型组比较,各给药组受孕率无显著变化,平均着床点数及体质量显著升高(P<0.01);子宫内膜腺体密度增加,间质水肿,核下可见空泡;子宫组织中T-bet、IFN-γ表达显著降低(P<0.01),GATA3、IL-4表达明显升高(P<0.05,P<0.01);血清中IFN-γ水平明显降低(P<0.05,P<0.01),IL-4水平明显升高(P<0.05);Th1/Th2比值(T-bet/GATA3、IFN-γ/IL-4)显著降低(P<0.01)。结论:八珍汤合归肾丸加减可显著提高胚胎着床障碍小鼠的着床点数,其机制可能与调控特异性转录因子T-bet、GATA3表达,纠正Th1/Th2免疫失衡及改善子宫内膜容受性有关。
Objective:To investigate the mechanism of the modified of Bazhentang combined with Guishentang in improving pregnancy outcomes in mouse models of embryo implantation dysfunction by regulating T helper 1/T helper 2(Th1/Th2)immune balance.Methods:Eighty ICR female mice were randomly divided into four groups(n=20 per group)on gestational day 1(GD1):control,model,western medicine,and traditional Chinese medicine(TCM)groups.Except for the control group,all mice received mifepristone solution(0.2 mg/mouse)via oral gavage on GD4 to induce embryo implantation dysfunction.The TCM group received a water decoction of the modified of Bazhentang combined with Guishentang(20.8 g·kg^(-1)),with the western medicine group administered dydrogesterone(3.9 mg·kg^(-1)),and the control/model groups given equal volumes of saline.All treatments were administered once daily from GD1 until one day before sample collection.Outcomes included implantation site counts(macroscopic observation),pregnancy rates,body weight,endometrial histopathology(hematoxylineosin staining),uterine expression of T-box expressed in T cells(T-bet),GATA-binding protein 3(GATA3),interferon gamma(IFN-γ),and interleukin-4(IL-4)at protein(Western blot)and mRNA(real-time polymerase chain reaction,Real-time PCR)levels,serum IFN-γand IL-4 levels(enzyme-linked immunosorbent assay,ELISA),and Th1/Th2 immune balance evaluated by calculating T-bet/GATA3 and IFN-γ/IL-4 ratios.Results:Compared to the control group,the model group showed no significant change in pregnancy rate but exhibited a marked reduction in average implantation sites and body weight(P<0.01).Histopathological analysis revealed endometrial abnormalities,including decreased glandular density,stromal compaction,and absence of nucleolar vacuoles.At the molecular level,uterine tissue in the model group demonstrated significantly upregulated expression of T-bet and IFN-γ(P<0.05,P<0.01),alongside markedly downregulated GATA3 and IL-4 expression(P<0.05,P<0.01).Serum analysis confirmed markedly elevated IFN-γ(P<0.01)and reduced IL-4 levels(P<0.01),resulting in significantly increased T-bet/GATA3 and IFN-γ/IL-4 ratios(P<0.01).Compared to the model group,pregnancy rates in all treatment groups showed no significant change.Implantation sites and body weight increased substantially(P<0.01),with restored endometrial morphology characterized by enhanced glandular density,stromal edema,and reappearance of nucleolar vacuoles.Significant downregulation of T-bet and IFN-γ(P<0.01)and upregulation of GATA3 and IL-4(P<0.05,P<0.01)in uterine tissue were observed.Serum IFN-γlevels were significantly reduced(P<0.05,P<0.01),while IL-4 levels were significantly elevated(P<0.05).The Th1/Th2 ratios were significantly decreased(P<0.01).Conclusion:The modified of Bazhentang combined with Guishentang significantly enhances the number of embryo implantation sites in mice with embryo implantation dysfunction,potentially through modulating T-bet/GATA3 expression,restoring Th1/Th2 immune balance,and improving endometrial receptivity.
作者
邓强
吴锋英
尹璐
王珺
叶朝阳
黄佳梅
金志春
DENG Qiang;WU Fengying;YIN Lu;WANG Jun;YE Zhaoyang;HUANG Jiamei;JIN Zhichun(Hubei University of Chinese Medicine,Wuhan 430065,China;Maternal and Child Health Hospital of Hubei Province,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430070,China)
出处
《中国实验方剂学杂志》
北大核心
2025年第18期68-76,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
全国名老中医药专家传承工作室建设项目(国中医药人教函[2022]75号)
湖北省中医药管理局联合基金项目(ZY2025L191)
湖北省自然科学基金妇幼创新发展联合基金项目(2025AFD684)。
