文献类型：期刊文献

中文题名：基于MDM4/p53/GPX4信号通路探讨加味少腹逐瘀汤对子宫内膜异位症大鼠异位子宫内膜组织铁死亡的影响

英文题名：Effect of Modified Shaofu Zhuyutang on Ferroptosis in Ectopic Endometrial Tissues of Rats with Endometriosis Based on MDM4/p53/GPX4 Signaling Pathway

作者：张作良[1];林祥羽[1];王婉润[1];王家兴[1];杨亚玲[2];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：39

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;甘肃省人民医院院内科研基金项目(24GSSYE-12)。

语种：中文

中文关键词：子宫内膜异位症;铁死亡;加味少腹逐瘀汤;小鼠双体4/肿瘤抑制因子p53/谷胱甘肽过氧化物酶4(MDM4/p53/GPX4)信号通路

外文关键词：endometriosis;ferroptosis;modified Shaofu Zhuyutang;murine double minute 4/tumor suppressor p53/glutathione peroxidase 4(MDM4/p53/GPX4)signaling pathway

摘要：目的:探讨加味少腹逐瘀汤通过小鼠双体4/肿瘤抑制因子p53/谷胱甘肽过氧化物酶4(MDM4/p53/GPX4)信号通路诱导子宫内膜异位症大鼠异位子宫内膜组织铁死亡的作用机制。方法:将70只SPF级雌性SD大鼠随机分为空白组(n=10)、假手术组(n=10)和造模组(n=50),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)),加味少腹逐瘀汤高、中、低剂量组(30、15、7.5 g·kg^(-1)),每组10只。给药4周后,处死大鼠取材。透射电镜观察大鼠子宫内膜组织超微结构改变,普鲁士蓝染色观察子宫内膜组织铁离子沉积,酶联免疫吸附测定法(ELISA)检测各组大鼠血清中白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)含量,比色法检测大鼠子宫内膜组织铁离子(Fe2+)、丙二醛(MDA)、谷胱甘肽(GSH)含量,免疫荧光法(IF)检测子宫内膜组织GSH、GPX4相对荧光强度,蛋白免疫印迹法(Western blot)检测子宫内膜组织中磷脂酰肌醇3-激酶(PI3K)、磷酸化(p)-PI3K、蛋白激酶B(Akt)、p-Akt、MDM4、p53、GPX4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PI3K、Akt、MDM4、p53、GPX4 mRNA表达。结果:与空白组及假手术组比较,模型组大鼠超微结构可见铁死亡损伤减轻,普鲁士蓝染色可见铁离子聚集数量减少,铁离子阳性表达面积下降,血清IL-6、IL-1β、TNF-α含量升高,子宫内膜组织Fe^(2+)、MDA相对含量降低,GSH相对含量升高,GSH、GPX4荧光相对强度增强(P<0.01),PI3K、p-PI3K、Akt、p-Akt、MDM4、GPX4蛋白及mRNA表达量上升,p53蛋白及mRNA表达量下降(P<0.01)。与模型组比较,孕三烯酮组及加味少腹逐瘀汤高、中剂量组异位子宫内膜组织,透射电镜超微结构可见铁死亡损伤不同程度加重,普鲁士蓝染色可见铁离子聚集数量增加,铁离子阳性表达面积上升,血清IL-6、IL-1β、TNF-α含量下降,子宫内膜组织Fe^(2+)、MDA相对含量升高,GSH相对含量下降,GSH、GPX4荧光相对强度减弱,PI3K、p-PI3K、Akt、p-Akt、MDM4、GPX4蛋白及mRNA表达量下降,p53蛋白及mRNA表达量上升(P<0.05,P<0.01)。结论:加味少腹逐瘀汤可能通过干预MDM4/p53/GPX4信号通路关键蛋白,诱导异位子宫内膜组织铁死亡,减轻炎性反应,从而对子宫内膜异位症起到治疗作用。
Objective:To investigate the mechanism of modified Shaofu Zhuyutang in inducing ferroptosis in ectopic endometrial tissues of rats with endometriosis through the murine double minute 4(MDM4)/tumor suppressor p53/glutathione peroxidase 4(GPX4)signaling pathway.Methods:Seventy SPF-grade female SD rats were randomly divided into a blank group(n=10),a sham-operated group(n=10),and a modeling group(n=50).The sham-operated group underwent laparotomy,while the modeling group was subjected to the autotransplantation method to establish an endometriosis model.After successful modeling,the animals were randomly assigned to the model group,progesterone group(0.25 mg·kg^(-1)),and modified Shaofu Zhuyutang high-,medium-,and low-dose groups(30,15,and 7.5 g·kg^(-1),respectively),with 10 rats per group.After four weeks of drug administration,the rats were euthanized for sample collection.The weight and volume of ectopic endometrial tissues were recorded for each group.Transmission electron microscopy(TEM)was employed to observe ultrastructural changes in endometrial tissues,while Prussian blue staining was used to assess iron ion deposition.Serum levels of interleukin-6(IL-6),interleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α)were measured by enzyme-linked immunosorbent assay(ELISA).The relative levels of Fe^(2+),malondialdehyde(MDA),and glutathione(GSH)in endometrial tissues were determined by colorimetric assay.Immunofluorescence(IF)was used to detect the relative fluorescence intensities of GSH and GPX4 in endometrial tissues.The relative expression levels of phosphatidylinositol 3-kinase(PI3K),phosphorylated PI3K(p-PI3K),protein kinase B(Akt),phosphorylated Akt(p-Akt),MDM4,p53,and GPX4 proteins were detected by Western blot.Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)was used to assess the mRNA expression of PI3K,Akt,MDM4,p53,and GPX4.Results:Compared with the blank and sham-operated groups,the model group exhibited reduced ferroptotic damage in ultrastructural observations,decreased ferroptotic aggregates and positive iron ion expression area on Prussian blue staining,elevated serum IL-6,IL-1β,and TNF-αlevels,reduced Fe^(2+)and MDA content,increased GSH content in endometrial tissues,and enhanced GSH and GPX4 fluorescence intensities(P<0.01).The protein and mRNA expression levels of PI3K,p-PI3K,Akt,p-Akt,MDM4,and GPX4 were elevated,while those of p53 were decreased(P<0.01).Compared with the model group,in the progesterone group and the modified Shaofu Zhuyutang high-and medium-dose groups,ferroptotic damage in ultrastructural observations was exacerbated to varying degrees by TEM,and ferroptotic aggregates and positive iron ion expression areas were increased on Prussian blue staining.Serum IL-6,IL-1β,and TNF-αlevels decreased,Fe^(2+)and MDA content increased,and GSH content decreased in endometrial tissues.GSH and GPX4 fluorescence intensities weakened,while the protein and mRNA expression levels of PI3K,p-PI3K,Akt,p-Akt,MDM4,and GPX4 decreased,and those of p53 increased(P<0.05,P<0.01).Conclusion:modified Shaofu Zhuyutang may exert therapeutic effects in endometriosis by inducing ferroptosis in ectopic endometrial tissues,alleviating inflammatory responses,and modulating key proteins in the MDM4/p53/GPX4 signaling pathway.