摘要
目的:探讨肾缺血再灌注(ischemia-reperfusion,I/R)损伤诱导肾小管上皮细胞发生Parthanatos及激活该死亡途径的上游信号。方法:构建小鼠肾I/R损伤模型和细胞缺氧复氧损伤模型,并使用聚腺苷二磷酸核糖聚合酶1(Poly ADP-ribose polymerase 1,PARP-1)抑制剂3-氨基苯甲酰胺(3-aminobenzamide,3AB)和抗氧化剂N-乙酰-L-半胱氨酸(N-acetyl-L-cysteine,NAC)预处理。苏木精和伊红(hematoxylin and eosin,HE)染色观察小鼠肾组织病理损伤,检测血清肌酐(creatinine,Cre)和尿素氮(blood urea nitrogen,BUN)评估肾功能,细胞计数试剂盒-8(cell counting kit-8,CCK-8)和乳酸脱氢酶(lactate dehydrogenase,LDH)检测细胞存活率与死亡率,2′,7′-二氯荧光素二醋酸盐(2′,7′-dichlorodihydrofluorescein diacetate,DCFH-DA)检测细胞内活性氧(reactive oxygen species,ROS)水平,利用相应试剂盒检测细胞超氧化物歧化酶(superoxide dismutase,SOD)活性和丙二醛(malondialdehyde,MDA)水平,应用酶联免疫吸附测定和Western blot检测DNA损伤标记物及Parthanatos相关蛋白表达。结果:3AB预处理减轻I/R导致的肾组织损伤和肾功能障碍(P<0.05),减少缺氧复氧损伤导致的细胞死亡(P<0.05)。NAC预处理降低缺氧复氧细胞内ROS和MDA水平(P<0.05)、提高SOD活性(P<0.05)、抑制DNA损伤和Parthanatos通路激活(P<0.05)、减少细胞死亡(P<0.05),以及减轻肾损伤和肾功能障碍(P<0.05)。结论:I/R导致的氧化应激能够引起DNA的损伤并激活PARP-1,进而促使肾小管上皮细胞发生Parthanatos,这为有效防治肾I/R损伤提供了新的思路。
Objective:To investigate renal ischemia-reperfusion(I/R)injury inducing Parthanatos in renal tubular epithelial cells and the upstream signals that initiate this death pathway′s activation.Methods:A mouse model of renal I/R injury and a hypoxia/reoxygenation injury model of renal tubular epithelial cells were established and pretreated with Poly ADP-ribose polymerase 1(PARP-1)inhibitor 3-aminobenzamide(3AB)and antioxidant N-acetyl-L-cysteine(NAC).Hematoxylin and eosin(HE)staining was used to observe renal tissue pathological damage.Serum creatinine(Cre)and blood urea nitrogen(BUN)were detected to evaluate renal function.The cell counting kit-8(CCK-8)and lactate dehydrogenase(LDH)assays were used to detect cell viability and mortality.2′,7′-dichlorodihydrofluorescein diacetate(DCFH-DA)was used to detect intracellular reactive oxygen species(ROS).The activities of superoxide dismutase(SOD)and the levels of malondialdehyde(MDA)in cells were detected using corresponding kits.Enzyme-linked immunosorbent assay(ELISA)and Western blot were applied to detect DNA damage markers and the expression of Parthanatos-related proteins.Results:3AB pretreatment alleviated I/R-caused renal tissue damage and renal dysfunction(P<0.05),and reduced hypoxia-reoxygenation-induced cell death(P<0.05).NAC pretreatment decreased the levels of intracellular ROS and MDA in hypoxia-reoxygenation cells(P<0.05),increased the activity of SOD(P<0.05),inhibited DNA damage and Parthanatos pathway activation(P<0.05),reduced cell death(P<0.05),and alleviated renal injury and renal dysfunction(P<0.05).Conclusion:I/R-caused oxidative stress can lead to DNA damage and activate PARP-1,which in turn promotes the occurrence of Parthanatos in renal tubular epithelial cells.This provides new ideas for the effective prevention and treatment of renal I/R injury.
作者
谷宇
傅文婷
郑曦
魏巍
王艳玲
尧永华
GU Yu;FU Wenting;ZHENG Xi;WEI Wei;WANG Yanling;YAO Yonghua(Department of Anesthesiology,Guangzhou Institute of Cancer Research,the Affiliated Cancer Hospital,Guangzhou Medical University,Guangzhou 510000,Guangdong,China;Department of Anesthesiology,the Third Affiliated Hospital of Sun Yat-sen University,Guangzhou 510000,Guangdong,China)
出处
《暨南大学学报(自然科学与医学版)》
北大核心
2025年第1期13-23,共11页
Journal of Jinan University(Natural Science & Medicine Edition)
基金
国家自然科学基金项目(82072218)
广东省自然科学基金项目(2023A1515010288)
广州市科技计划项目(202201010791)
广州市科学技术局市校联合项目(2024A03J0648)
广州医科大学科研能力提升项目。
