氧化应激介导多柔比星导致的大鼠主动脉血管平滑肌细胞损伤  

Doxorubicin-induced thoracic aorta smooth muscle cells injury through oxidative stress pathway in rats

作　　者：张成瑞 谢童 邱会琴 王婵 范彦英 杨彩红 ZHANG Cheng-rui;XIE Tong;QIU Hui-qin;WANG Chan;FAN Yan-ying;YANG Cai-hong(Department of Pharmacology,Basic Medical Sciences Center,Shanxi Medical University,Taiyuan 030001,China)

机构地区：[1]山西医科大学基础医学院药理学教研室,山西太原030001

出　　处：《中国药理学与毒理学杂志》2022年第8期578-583,共6页Chinese Journal of Pharmacology and Toxicology

基　　金：山西省自然科学基金(201901D111198)。

摘　　要：目的探讨氧化应激在多柔比星(Dox)诱导大鼠主动脉血管平滑肌细胞(A7r5细胞)损伤中的作用及其机制。方法A7r5细胞经Dox 0,0.33,1.00和1.33μmol·L^(-1)孵育24 h,MTT法测定细胞存活率;倒置荧光显微镜下观察各组细胞形态;DCFH-DA荧光探针法测定活性氧(ROS)水平;比色法测定过氧化氢酶(CAT)含量;硝酸盐法测定一氧化氮(NO)含量;Western印迹法检测硫氧还蛋白互作蛋白(TXNIP)、还原型烟酰胺腺嘌呤二核苷磷酸(NADPH)氧化酶2(NOX2)、NADPH氧化酶的胞膜催化亚基p22^(phox)、内皮型NO合酶(eNOS)、诱导型NO合酶(iNOS)、蛋白激酶Cα(PKCα)、PKCβ1和PKCβ2蛋白表达水平。结果与对照组细胞相比,Dox 0.33,1.00和1.33μmol·L^(-1)组细胞存活率显著降低(P<0.05,P<0.01),镜下可见A7r5死细胞数目明显增加,不规则型,细胞核呈碎片状;细胞ROS水平显著升高(P<0.01),CAT含量明显下降(P<0.01),NO含量升高(P<0.01);Dox 1.00和1.33μmol·L^(-1)组细胞内TXNIP,p22^(phox),eNOS和iNOS蛋白表达水平显著升高(P<0.01);Dox 1.33μmol·L^(-1)组细胞内PKCα和PKCβ1蛋白表达水平显著升高(P<0.05);Dox 0.33,1.00和1.33μmol·L^(-1)组细胞内PKCβ2蛋白表达水平显著升高(P<0.05),而NOX2蛋白表达水平呈上升趋势,但未达到统计学差异。结论Dox导致A7r5细胞活力下降和氧化应激损伤,其机制可能与PKC激活NADPH氧化酶和NOS活性,进而诱导ROS大量产生有关。OBJECTIVE To investigate the role and mechanism of oxidative stress in thoracic aorta smooth muscle cell(A7r5 cell)injury induced by doxorubicin(Dox)in rats.METHODS A7r5 cells were incubated with Dox 0,0.33,1.00 and 1.33μmol·L^(-1) for 24 h.The cell viability was detected by MTT assay and the cell morphology was observed under an inverted fluorescence microscope.The level of reac⁃tive oxygen species(ROS)was measured by DCFH-DA fluorescence probe.The content of catalase(CAT)was detected by colorimetry and the intracellular nitric oxide(NO)level was evaluated using the nitrate method.The protein expressions of thioredoxin interaction protein(TXNIP),reduced nicotin⁃amide adenine dinucleoside phosphate(NADPH)oxidase 2(NOX2),membrane catalytic subunit P22^(phox) of NADPH oxidase,endothelial NO synthase(eNOS),inducible NOS synthase(iNOS),protein kinase Cα(PKCα),PKCβ1 and PKCβ2 were detected by Western blotting.RESULTS MTT results showed that compared with the cell control group,the survival rate of A7r5 cells was significantly reduced in the Dox 0.33,1.00 and 1.33μmol·L^(-1) groups(P<0.05,P<0.01).Microscopically,the number of A7r5 dead cells increased significantly,showing an irregular pattern and the nucleus was fragmented.Compared with cell control group,ROS levels in Dox 0.33,1.00 and 1.33μmol·L^(-1) groups were signifi⁃cantly increased(P<0.01),CAT content was significantly decreased(P<0.01)and the intracellular NO content was significantly increased(P<0.01).The protein expressions of TXNIP,P22^(phox),eNOS and iNOS were significantly increased in Dox 1.00 and 1.33μmol·L^(-1) groups(P<0.01),the intracellular protein expression of PKCαand PKCβ1 was significantly increased in the Dox 1.33μmol·L^(-1) group(P<0.05),and the protein expression of PKCβ2 was significantly increased in Dox 0.33,1.00 and 1.33μmol·L^(-1) groups(P<0.05),and the protein expression of NOX2 in Dox 0.33,1.00 and 1.33μmol·L^(-1) groups trended up,but the difference was not statistically significant.CONCLUSION Dox can decrea

关 键 词：多柔比星 主动脉血管平滑肌细胞 A7r5细胞 氧化应激 

分 类 号：R979.1[医药卫生—药品] R99[医药卫生—药学]