基于大鼠肝星状细胞氧化应激探讨化浊解毒补肾方治疗绝经后女性肝纤维化机制  

Exploring Mechanism of Huazhuo Jiedu Bushen Recipe(化浊解毒补肾方)in Treatment of Liver Fibrosis in Postmenopausal Women Based on H 2O 2-induced Oxidative Stress in Rat Hepatic Stellate Cells

作　　者：郭敏[1] 刘素彤 张永华[1] 张丽慧[1] 徐文倩 杨竹青 刘镇亚 姚翠翠 王璐 GUO Min;LIU Sutong;ZHANG Yonghua;ZHANG Lihui;XU Wenqian;YANG Zhuqing;LIU Zhenya;YAO Cuicui;WANG Lu(Department of Hepatology and Spleen-Stomach,The First Affiliated Hospital of Henan University of Traditional Chinese Medicine,Zhengzhou 450003,Henan,China;The First Clinical Medical College of Henan University of Traditional Chinese Medicine,Zhengzhou 450000,Henan,China)

机构地区：[1]河南中医药大学第一附属医院脾胃肝胆病科,河南郑州450003 [2]河南中医药大学第一临床医学院,河南郑州450000

出　　处：《辽宁中医杂志》2024年第6期183-188,I0009,I0010,共8页Liaoning Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(81904161)。

摘　　要：目的通过研究化浊解毒补肾方含药血清对过氧化氢(H_(2)O_(2))诱导大鼠肝星状细胞(hepatic stellate cells,HSCs)氧化应激的影响,进一步探讨该方治疗绝经后女性肝纤维化的机制。方法制备化浊解毒补肾方含药血清和正常血清。将肝星状细胞随机分为空白组、模型组、空白血清组、氟维司群(ICI 182780,以下简记为ICI)组、中药组、中药+ICI组、雌二醇(estradiol,E2)组、E2+ICI组,随后按照分组处理细胞。采用流式细胞术检测细胞内活性氧(reactive oxygen species,ROS)水平;ELISA法检测细胞中丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxide dismutase,SOD)水平;免疫荧光染色法检测细胞中α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)表达;Western Blot法检测胞浆细胞色素C(cytochrome C)、环氧化酶-Ⅰ(cyclooxygenase-Ⅰ,COX-Ⅰ)、α-SMA、Ⅰ型胶原蛋白(collagenⅠ)表达。结果与空白组比较,模型组细胞中ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均明显升高(P<0.0001,P<0.001),SOD水平、cytochrome C、COX-Ⅰ蛋白表达均明显降低(P<0.0001);与模型组比较,中药组和E2组ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均有明显降低(P<0.0001,P<0.001,P<0.01,P<0.05),SOD水平、cytochrome C、COX-Ⅰ蛋白表达均有明显升高(P<0.0001);与中药组和E2组比较,在各自加入ICI处理后,对应的ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均有明显升高(P<0.0001,P<0.001,P<0.05),SOD水平和cytochrome C、COX-Ⅰ蛋白表达均有明显降低(P<0.001,P<0.05,P<0.01)。结论化浊解毒补肾方含药血清可通过雌激素受体降低ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度,提高SOD水平和cytochrome C、COX-Ⅰ蛋白表达,从而提高抗氧化能力,减轻氧化应激反应,最终起到防治绝经后女性肝纤维化的作用。Objective To study the effect of Huazhuo Jiedu Bushen Recipe(化浊解毒补肾方)containing serum on H_(2)O_(2)-induced oxidative stress in rat hepatic stellate cells(HSCs),and to further explore the mechanism of this prescription in the treatment of liver fibrosis in postmenopausal women.Methods Huazhuo Jiedu Bushen Recipe containing serum and normal serum were prepared.HSCs were randomly divided into the blank group,model group,blank serum group,fulvenstrant(ICI 182780,abbrebiated as ICI)group,traditional Chinese Medicine(TCM)group,TCM+ICI group,estradiol(E2)group,E2+ICI group.Then these HSCs were treated in their respective groups.Reactive oxygen species(ROS)were detected by flow cytometry.The levels of malondialdehyde(MDA)and superoxide dismutase(SOD)were detected by ELISA,the expressions ofα-smooth muscle actin(α-SMA)were detected by immunofluorescence and the expressions of cytochrome C,cyclooxygenase-Ⅰ(COX-Ⅰ),α-SMA and collagenⅠwere detected by Western Blot.Results Compared with those of the blank group,the levels of ROS,MDA,α-SMA and collagenⅠwere increased in the model group(P<0.0001,P<0.001).The levels of SOD,cytochrome C and COX-Ⅰwere decreased(P<0.0001).Compared with those of the model group,the level of ROS,MDA,α-SMA and collagenⅠwere decreased(P<0.0001,P<0.001,P<0.01,P<0.05).The levels of SOD,cytochrome C and COX-Ⅰwere increased in the TCM group and E2 group(P<0.0001).Compared with those of the TCM group and E2 group,the corresponding levels of ROS,MDA,α-SMA and collagenⅠwere increased(P<0.0001,P<0.001,P<0.05)and the levels of SOD,cytochrome C and COX-Ⅰwere decreased after joining ICI(P<0.001,P<0.05,P<0.01).Conclusion Through E2 receptors,Huazhuo Jiedu Bushen Recipe containing serum could reduce the levels of ROS,MDA,α-SMA and collagenⅠ,improve SOD level and the protein expressions of cytochrome C and COX-Ⅰto increase the antioxidant capacity and reduce the oxidative stress,and finally prevent liver fibrosis in postmenopausal women.

关 键 词：化浊解毒补肾方 肝星状细胞 雌激素受体 肝纤维化 氧化应激 

分 类 号：R285[医药卫生—中药学]