文献类型：期刊文献

中文题名：基于TRL4/NF-κB/NLRP3信号通路介导的细胞焦亡探讨加味少腹逐瘀汤拮抗异位子宫内膜组织纤维化机制

英文题名：Mechanism of Modified Shaofu Zhuyutang in Antagonising Ectopic Endometrial Tissue Fibrosis Based on Cellular Pyroptosis Mediated by TRL4/NF-κB/NLPR3 Signaling Pathway

作者：张作良[1];王家兴[1];王婉润[1];林祥羽[1];岳斌[1];张志瑞[1];王亦楠[1];杨亚玲[2];魏冬青[2];黄灿灿[1];武权生[1]

第一作者：张作良

机构：[1]甘肃中医药大学中医临床学院,兰州730000;[2]甘肃中医药大学附属医院,兰州730030

第一机构：甘肃中医药大学中医临床学院

年份：2025

卷号：31

期号：4

起止页码：19

中文期刊名：中国实验方剂学杂志

外文期刊名：Chinese Journal of Experimental Traditional Medical Formulae

收录：;北大核心:【北大核心2023】；

基金：国家自然科学基金项目(82260949);甘肃省自然科学基金项目(23JRRA1199);甘肃省中医药科研项目(GZKP-2022-22);兰州市科技计划项目(2021-1-97,2023-ZD-21);甘肃中医药大学研究生创新基金项目。

语种：中文

中文关键词：子宫内膜异位症;纤维化;细胞焦亡;加味少腹逐瘀汤;Toll样受体4/核转录因子-κB/NOD样受体蛋白3(TRL4/NF-κB/NLRP3)信号通路;NLRP3炎性小体

外文关键词：endometriosis;fibrosis;pyroptosis;modified Shaofu Zhuyutang;Toll-like receptor 4/nuclear factor-κB/NODlike receptor protein 3(TRL4/NF-κB/NLPR3)signaling pathway;NLPR3 inflammasomes

摘要：目的:探讨加味少腹逐瘀汤通过Toll样受体4/核转录因子-κB/NOD样受体蛋白3(TRL4/NF-κB/NLRP3)信号通路拮抗子宫内膜异位症异位子宫内膜组织细胞焦亡及纤维化的作用机制。方法:将72只SPF级雌性SD大鼠随机分为假手术组(n=12)和造模组(n=60),假手术组大鼠行剖腹探查术,造模组大鼠按照自体移植法构建子宫内膜异位症疾病模型。造模成功后随机分为模型组,孕三烯酮组(0.25 mg·kg^(-1)),加味少腹逐瘀汤低、中、高剂量组(7.5、15、30 g·kg^(-1)),每组12只。给药4周后,观察大鼠自主活动能力及热痛潜伏期,处死大鼠取材,马松(Masson)染色观察子宫内膜组织病理变化,酶联免疫吸附测定法(ELISA)检测各组大鼠血清中白细胞介素-18(IL-18)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、转化生长因子-β(TGF-β)含量,免疫组化法(IHC)检测子宫内膜组织肿瘤坏死因子相关因子6(TRAF6)、NLRP3蛋白表达面积,免疫荧光法(IF)检测子宫内膜组织胱天蛋白酶-1(Caspase-1)、消皮素D(GSDMD)相对荧光强度,蛋白免疫印迹法(Western blot)检测子宫内膜组织中TRL4、髓样分化因子88(MyD88)、TRAF6、NF-κB p65、磷酸化核转录因子-κB p65(p-NF-κB p65)、NLRP3蛋白相对表达量,实时荧光定量聚合酶链式反应(Real-time PCR)检测子宫内膜组织TRL4、MyD88、TRAF6、NF-κB、NLRP3 mRNA表达。结果:与假手术组比较,模型组大鼠自主活动能力减弱,热痛潜伏期缩短;血清IL-18、IL-1β、TNF-α、TGF-β含量升高,TRAF6、NLRP3蛋白相对表达面积上升,Caspase-1、GSDMD荧光相对强度增强,TRL4、MyD88、TRAF6、NF-κB p65、p-NF-κB p65、NLRP3蛋白相对表达量及TRL4、MyD88、TRAF6、NF-κB、NLRP3 mRNA表达量上升(P<0.01)。与模型组比较,孕三烯酮组及加味少腹逐瘀汤中、高剂量组大鼠自主活动能力增强,热痛潜伏期延长;子宫内膜组织纤维化程度减轻;血清IL-18、IL-1β、TNF-α、TGF-β含量降低,TRAF6、NLRP3蛋白相对表达面积下降,Caspase-1、GSDMD荧光相对强度减弱,TRL4、MyD88、TRAF6、p-NF-κB p65、NLRP3蛋白相对表达量及TRL4、MyD88、TRAF6、NF-κB、NLRP3 mRNA表达量下降(P<0.05,P<0.01)。结论:加味少腹逐瘀汤可能通过干预TRL4/NF-κB/NLRP3信号通路关键蛋白,减轻NLRP3炎性小体诱导的细胞焦亡,拮抗异位子宫内膜组织纤维化进程,改善盆腔炎性微环境,缓解疼痛,从而对子宫内膜异位症起到治疗作用。
Objective:To investigate the mechanism of action of modified Shaofu Zhuyutang in antagonizing cellular pyroptosis and fibrosis in ectopic endometrial tissues of endometriosis through the Toll-like receptor 4/nuclear factor-κB/NOD-like receptor protein 3(TRL4/NF-κB/NLPR3)signaling pathway.Methods:Seventy-two SPF-grade female SD rats were randomly divided into a sham-operated group(n=12)and a modeling group(n=60).The rats in the sham-operated group underwent a caesarean section,while the rats in the modeling group were used to establish an endometriosis model through the autotransplantation method.After successful modeling,the animals were randomly divided into the model group,progesterone group(0.25 mg·kg^(-1)),and modified Shaofu Zhuyutang low-,medium-,and high-dose groups(7.5,15,30 g·kg^(-1)),with 12 animals in each group.After 4 weeks of drug administration,voluntary activity and heat pain latency were observed.The rats were sacrificed for tissue collection,and Masson staining were used to observe histopathological changes in the endometrial tissues.Enzyme-linked immunosorbent assay(ELISA)was used to measure serum levels of interleukin-18(IL-18),interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),and transforming growth factor-β(TGF-β).Immunohistochemistry(IHC)was used to detect the protein expression area of tumor necrosis factor-related factor 6(TRAF6)and NLPR3 in the endometrial tissues.Immunofluorescence(IF)was used to detect the relative fluorescence intensity of Caspase-1 and gasdermin D(GSDMD)in the endometrial tissues.Western blot was employed to measure the relative expression of TRL4,myeloid differentiation factor 88(MyD88),TRAF6,NF-κB p65,phosphorylated NF-κB p65(p-NF-κB p65),and NLPR3 proteins in endometrial tissues.Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)was used to detect the mRNA expression of TRL4,MyD88,TRAF6,NF-κB,and NLPR3 in the endometrial tissues.Results:Compared with the sham-operated group,rats in the model group showed reduced voluntary activity and shorter heat pain latency.Serum levels of IL-18,IL-1β,TNF-α,and TGF-βwere elevated.The relative expression areas of TRAF6 and NLPR3 proteins were increased,and the relative fluorescence intensity of Caspase-1 and GSDMD was enhanced.The relative expression of TRL4,MyD88,TRAF6,NF-κB p65,p-NF-κB p65,and NLPR3 proteins,along with the expression of TRL4,MyD88,TRAF6,NF-κB,and NLPR3 mRNA,were significantly increased(P<0.01).Compared with the model group,rats in the progesterone group and the modified Shaofu Zhuyutang medium-and highdose groups exhibited improved voluntary activity,longer heat pain latency,the fibrosis of endometrial tissue is alleviated.Serum levels of IL-18,IL-1β,TNF-α,and TGF-βwere decreased.The relative expression areas of TRAF6 and NLPR3 proteins decreased,and the relative fluorescence intensity of Caspase-1 and GSDMD weakened.The relative expression of TRL4,MyD88,TRAF6,p-NF-κB p65,NLPR3 proteins,and TRL4,MyD88,TRAF6,NF-κB,and NLPR3 mRNA expression were reduced(P<0.05,P<0.01).Conclusion:Modified Shaofu Zhuyutang may play a therapeutic role in endometriosis by interfering with key proteins in the TRL4/NF-κB/NLPR3 signaling pathway,reducing NLRP3 inflammasome-induced cellular pyroptosis,antagonizing the fibrosis process in ectopic endometrial tissues,improving the inflammatory microenvironment in the pelvic cavity,and alleviating pain.