一贯煎通过调节Parp-1的翻译后修饰保护肝细胞DNA  

Yiguanjian decoction inhibits hepatocytic DNA damage by regulating post-translational modification of Parp-1

作　　者：张宇佳 叶杰[1] 卢艳琳 闫晓风[1] 李华[1] 王晓玲[1] ZHANG Yu-jia;YE Ting-jie;LU Yan-lin;YAN Xiao-feng;Li Hua;WANG Xiao-ling(Dept of Biology,School of Basic Medical Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Dept of Clinical Medicine,Suzhou Vocational Health College,Suzhou,Jiangsu 215009,China;Dept of tumor,Longhua Hospital Affiliated to Shanghai University of Chinese Medicine,Shanghai 200032,China)

机构地区：[1]上海中医药大学基础医学院生物教研室,上海201203 [2]苏州卫生职业技术学院临床医学院,江苏苏州215009 [3]上海中医药大学附属龙华医院肿瘤科,上海200032

出　　处：《中国药理学通报》2023年第2期373-379,共7页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金青年项目(No 81703832)。

摘　　要：目的观察一贯煎(Yiguanjian decoction,YGJ)影响Parp-1修复H_(2)O_(2)诱导肝细胞DNA损伤的机制。方法培养小鼠胚胎肝细胞BNL CL.2,建立H_(2)O_(2)细胞损伤模型并用YGJ预处理,CCK-8检测细胞活力,Annexin V/PI流式细胞术检测细胞死亡,EDU掺入检测细胞增殖,8-羟基脱氧鸟苷比色法和γ-H_(2)AX细胞荧光染色检测DNA损伤。免疫印迹法检测Parp-1、Parp-1的Par化和乙酰化。结果H_(2)O_(2)作用后,肝细胞凋亡率增加;细胞增殖被明显抑制;DNA损伤持续存在于整个实验过程。Parp-1在6 h升高后维持高表达,Parp-1的Par化和乙酰化均先升高随后降低。Sirt-1的抑制剂EX527可明显提高Parp-1的Par化,而其激动剂SRT1720可明显降低Parp-1的Par化。YGJ作用后细胞凋亡率和DNA损伤明显减少,增殖细胞数明显增加,Parp-1高表达时间点提前,Parp-1的Par化明显提高,乙酰化时间点推迟,且持续时间更长。结论YGJ可降低H_(2)O_(2)诱导的肝细胞DNA损伤,其作用机制与Sirt-1对Parp-1去乙酰化维持其含量、负调控Parp-1的Par化维持一定的活性有关。Aim To investigate the mechanism of yiguanjian decoction(YGJ)protecting H_(2)O_(2)-induced DNA damage in hepatocytes by regulating Parp-1 activity.Methods DNA damage model was established by H_(2)O_(2)in cultured mouse embryonic hepatocytes BNL CL2,and cells were treated with YGJ.Cell viability was determined by CCK8,apoptotic rates were determined by flow cytometry with Annexin V/PI staining,cell proliferation was determined by EDU incorporation,DNA damage was measured with 8-hydroxydeoxyguanosine colorimetric method and fluorescence staining of γ-H_(2)AX,and parylation and acetylation level of Parp-1,Parp-1 protein expression were determined by immunoblotting.Results H_(2)O_(2)stress significantly inhibited hepatocyte proliferation in the whole process,and DNA damage persisted throughout the experimental process.Parp-1 expression was maintained high level from 6 h to 24 h.Parylation and acetylation level of Parp-1 both increased first and then decreased.EX527,an inhibitor of Sirt-1,significantly improved the parylation modification of Parp-1,while its agonist SRT1720 significantly reduced the parylation modification of Parp-1.After YGJ treatment,the apoptotic rates of apoptosis and DNA damage were significantly reduced,numbers of proliferating cells significantly increased,and the time point of Parp-1higher expression was early.Parylation of Parp-1 significant increased and its acetylation was delayed and lasted longer.Conclusions YGJ reduces H_(2)O_(2)-induced DNA damage in hepatocytes,and the mechanism is related to the maintenance of deacetylation of Parp-1 through Sirt-1 and while negatively regulating the parylation of Parp-1.

关 键 词：肝细胞 PARP-1 Sirt-1 翻译后修饰 DNA损伤 H_(2)O_(2) 

分 类 号：R-332[医药卫生] R289.5R322.47R342.3R345.5