依帕司他对放射性肺炎小鼠线粒体氧化应激损伤的保护作用  

Protective efficacy of epalrestat on mitochondrial oxidative stress damage for radiation pneumonitis in mice

作　　者：李泽朋 顾文强 陈晓 王银华 李先伟 LI Zepeng;GU Wenqiang;CHEN Xiao;WANG Yinhua;LI Xianwei(Department of Pharmacology,Wannan Medical College,Wuhu 241002,Anhui,China;Department of Oncology,Wuhu No.2 People's Hospital,Wuhu 241000,Anhui,China)

机构地区：[1]皖南医学院药学院药理学教研室,安徽芜湖241002 [2]芜湖市第二人民医院肿瘤放疗科,安徽芜湖241000

出　　处：《中国临床药理学与治疗学》2024年第7期809-818,共10页Chinese Journal of Clinical Pharmacology and Therapeutics

基　　金：安徽省高校自然科学研究重大项目(KJ2021ZD0106);安徽省卫生健康委科研重点项目(AHWJ2021a033)。

摘　　要：目的:探讨依帕司他(Epa)对放射性肺炎(RP)小鼠线粒体氧化应激损伤的影响及其机制。方法:C57BL/6小鼠随机对照(CON)组、放射(IR)组、IR联合10 mg/kg Epa处理组及IR联合20 mg/kg Epa处理组,每组各16只。采用6MV X线直线加速器全胸单次照射15 Gy建立放射性肺炎模型。照射后连续灌胃给药6~8周。HE染色观察肺组织病理变化。透射电镜观察肺组织线粒体结构。ELISA检测血浆白细胞介素6(IL-6)、肿瘤坏死因子-α(TNF-α)及转化生长因子-β1(TGF-β1)的水平。免疫组化检测肺组织醛糖还原酶(AR)的表达。比色法检测肺组织丙二醛(MDA)及4-羟基壬烯醛(4-HNE)含量。制备肺组织单细胞悬液,使用DCFH-DA探针检测细胞内活性氧(ROS)水平。实时定量PCR检测AR、IL-6、TNF-α和TGF-β1 mRNA的表达。Western blot法检测AR、IL-6、TNF-α、TGF-β1、BAX、Bcl2、Cleaved Caspase-3、8-氧鸟嘌呤DNA糖基化酶1(OGG1)及沉默信号调节因子3(SIRT3)的蛋白水平。结果:与CON组相比,IR组肺泡水肿,肺泡间隔增厚并伴大量炎症细胞浸润;炎症因子IL-6、TNF-α和TGF-β1的水平明显升高(P<0.01);Bcl2的表达明显下调而BAX、Cleaved Caspase-3的表达明显上调(P<0.05,P<0.01)。与IR组相比,Epa连续给药6~8周后,小鼠肺组织炎症损伤明显减轻,炎症因子IL-6、TNF-α和TGF-β1的水平明显降低(P<0.05,P<0.01),细胞凋亡程度明显减轻(Bcl2的表达上调而BAX、Cleaved Caspase-3的表达下调)。与CON组相比,IR组AR的表达明显升高,ROS、MDA及4-HNE的水平明显增加(P<0.01),OGG1和SIRT3的表达明显降低(P<0.01),线粒体损伤明显加剧。而与IR组相比,Epa连续给药6~8周后,IR组AR的表达明显下调(P<0.05,P<0.01),ROS、MDA及4-HNE的水平明显降低,OGG1和SIRT3的表达明显增加,线粒体损伤明显减轻(P<0.05,P<0.01)。结论:Epa对放射性肺炎具有保护作用,其作用可能与其抑制AR的表达,减轻线粒体氧化应激损伤,抑制炎症反应和细胞凋亡有关。AIM:To investigate the effects of epalrestat(Epa)on the mitochondrial oxidative stress damage of radiation pneumonitis(RP)mice and to explore its possible mechanism.METHODS:C57BL/6 mice were randomly divided into control(CON),Irradiation(IR),IR combined with Epa(10 mg/kg)and IR combined with Epa(20 mg/kg)group,16 mice in each group.Mouse models of RP were established by whole thorax irradiation at a dose of 15Gy using a 6-MV linear accelerator.Continuous intragastric administration after IR for 6 or 8 weeks.Lung histopathology was analyzed by HE staining.The expression of aldose reductase(AR)was determined by immunohistochemistry.Mitochondrial morphology of lung tissues was observed by transmission electron microscopy.The levels of inflammatory cytokines(IL-6,TNF-αand TGF-β1)in plasma were detected by ELISA.The contents of Malondialdehyde(MDA)and 4-hydroxynonenal(4-HNE)in lung tissues were determined by colorimetry.Single cell suspension of lung tissues was prepared and reactive oxygen species(ROS)levels in the cells was examined using a DCFH-DA fluorescent probe.Real-time quantitative PCR was used to determine the expression of AR,IL-6,TNF-αand TGF-β1.The protein levels of AR,IL-6,TNF-α,TGF-β1,BAX,Bcl2,Cleaved Caspase-3,8-oxoguanine DNA glycosylase 1(OGG1)and silent information regulator 3(SIRT3)were detected by Western blot analysis.RESULTS:Compared with the CON group,the alveolar hyperplasia,alveolar septum thickening and inflammatory cell infiltration were observed in the IR group.Moreover,the content of inflammatory factors such as IL-6,TNF-αand TGF-β1 and the expression of BAX and Cleaved Caspase-3 were significantly increased,and the expression of Bcl2 was obviously decreased after irradiation.Compared with the IR group,Epa robustly alleviated RP.Meanwhile,Epa down-regulated inflammatory cells infiltration and the expression of inflammatory cytokines,such as IL-6,TNF-αand TGF-β1.In addition,Epa could down-regulate the expression of BAX and Cleaved Caspase-3,and up-regulate Bcl2 in lung tissues.Co

关 键 词：依帕司他 放射性肺炎 醛糖还原酶 线粒体 氧化应激 小鼠 

分 类 号：R965.2[医药卫生—药理学]