摘要
目的:基于核因子E2相关因子2(Nrf2)-抗氧化反应元件(ARE)信号通路探讨坤泰胶囊联合干细胞治疗对卵巢早衰(POF)大鼠氧化应激损伤的抑制作用。方法:原代培养Sprague-Dawley大鼠骨髓间充质干细胞,培养至第3代备用。雌性Sprague-Dawley大鼠通过腹腔注射环磷酸酰胺建立POF大鼠模型,随机分为POF组、坤泰(Kuntai)组、干细胞(stem cell)组、联合(combination)组及combination+brusatol组,并以注射生理盐水的正常大鼠为对照(control)组,每组10只。干预结束后,分析大鼠体质量增量及卵巢指数变化;ELISA检测血清中黄体生成素(LH)、卵泡刺激素(FSH)、血清雌二醇(E2)、抗缪勒氏管激素(AMH)及丙二醛(MDA)、谷胱甘肽(GSH)、超氧化物歧化酶(SOD)指标;分离卵巢组织,HE染色观察大鼠卵巢病理学变化,RT-qPCR检测抗凋亡BCL-2、促凋亡BAX mRNA表达;Western blot检测Nrf2和亚铁血红素氧化酶1(HO-1)蛋白表达。结果:与control组相比,POF组成熟卵泡较少,卵巢形态结构紊乱,体质量增量、卵巢指数、E2、AMH、SOD和GSH水平、BCL-2 mRNA及Nrf-2和HO-1蛋白表达显著降低,LH、FSH和MDA水平及BAX mRNA表达显著增加(P<0.05);与POF组相比,Kuntai组、stem cell组和combination组卵巢形态得到改善,以combination组改善最为显著,体质量增量、卵巢指数、E2、AMH、SOD和GSH水平、BCL-2 mRNA及Nrf-2和HO-1蛋白表达显著增加,LH、FSH和MDA水平、BAX mRNA表达显著降低,combination与Kuntai组、stem cell组统计学差异显著(P<0.05);与combination组相比,combination+brusatol组成熟卵泡减少,卵泡结构紊乱,体质量增量、卵巢指数、E2、AMH、SOD和GSH水平和BCL-2 mRNA及Nrf-2、HO-1蛋白表达显著降低,LH、FSH和MDA水平及BAX mRNA表达显著增加(P<0.05)。结论:坤泰胶囊联合干细胞可改善POF大鼠卵巢功能,其机制可能与激活Nrf2-ARE信号通路减轻卵巢组织氧化应激有关。
AIM:To investigate the effects of Kuntai capsule combined with stem cell therapy on oxidative stress damage in rats with premature ovarian failure(POF),focusing on the Nrf2-ARE signaling pathway.METHODS:Bone marrow mesenchymal stem cells(BMSCs)from Sprague-Dawley rats were cultured from primary to third generation.Female Sprague-Dawley rats were selected,and mesenchymal stem cells were isolated.A POF rat model was established through intraperitoneal injection of cyclic phosphamide,and the model was randomly divided into five groups:POF group,Kuntai group,stem cell group,combination group,and combination+Brusatol group.Normal rats injected with physio-logical saline served as the control group,with 10 rats in each group.After the intervention,changes in body weight and ovarian index were analyzed.Serum levels of estrogen indicators[luteinizing hormone(LH),follicle-stimulating hormone(FSH),serum estradiol(E2),and anti-Müllerian hormone(AMH)]and oxidative stress markers[malondialdehyde(MDA),glutathione(GSH),and superoxide dismutase(SOD)]were measured.Ovarian tissue was isolated for histologi-cal examination using HE staining and for expression analysis of anti-apoptotic BCL-2 and pro-apoptotic BAX mRNA via RT-qPCR.Western blot analysis was conducted to assess the expression of Nrf2 and heme oxygenase-1(HO-1)proteins.RESULTS:Compared to the control group,the POF group exhibited fewer mature follicles and disorganized ovarian mor-phology,along with significantly reduced body weight gain,ovarian index,E2,AMH,SOD and GSH levels,BCL-2 mRNA,and Nrf2 and HO-1 protein expression.Conversely,LH,FSH,MDA levels,and BAX mRNA expression were significantly increased(P<0.05).In comparison to the POF group,the Kuntai group,stem cell group,and combination group showed improved ovarian morphology,with the combination group demonstrating the most significant improvement.In these groups,body weight gain,ovarian index,E2,AMH,SOD,GSH levels,BCL-2 mRNA,and Nrf2 and HO-1 pro-tein expression were markedly increased,while LH,FSH,MDA levels,and BAX mRNA expression were notably de-creased,with significant statistical differences observed between the combination group and the Kuntai and stem cell groups(P<0.05).In contrast,the combination+Brusatol group exhibited a reduction in mature follicles and disrupted follicular structure compared to the combination group,along with decreased body weight gain,ovarian index,E2,AMH,SOD and GSH levels,BCL-2 mRNA,and Nrf2 and HO-1 protein expression,while LH,FSH and MDA levels,and BAX mRNA expression were significantly increased(P<0.05).CONCLUSION:The combination of Kuntai capsule and stem cell therapy effectively mitigates oxidative stress damage in POF rats,potentially through the activation of the Nrf2-ARE signaling pathway.
作者
张宁
吴晓茜
胡玉芹
田耕
ZHANG Ning;WU Xiaoqian;HU Yuqin;TIAN Geng(Shijiazhuang Fourth Hospital,Shijiazhuang 050011,China)
出处
《中国病理生理杂志》
北大核心
2025年第3期562-568,共7页
Chinese Journal of Pathophysiology
基金
河北省2024年度中医药类科学研究课题计划(No.2024135)。
