摘要
目的:探讨食管癌中铜锌超氧化物歧化酶(SOD1)的表达及其对细胞增殖的影响和机制。方法:选取食管癌组织和癌旁组织各32例,利用实时定量反转录聚合酶链反应(RT-qPCR)、蛋白质印迹法(Western blot)和免疫组织化学法检测SOD1表达。将食管癌细胞系KYSE510随机分为3组:对照组、腺相关病毒(AAV)-绿色荧光蛋白(GFP)组和AAV-SOD1组。AAV-GFP组和AAV-SOD1组细胞分别转染AAV-GFP和AAV-SOD1重组病毒载体,对照组则加磷酸盐缓冲液(PBS)处理。利用噻唑蓝(MTT)法检测细胞活性、二氢乙锭(DHE)探针检测活性氧簇(ROS)产生、Western blot检测Wnt5a和β-连环蛋白(β-catenin)蛋白表达。两组间比较采用t检验,3组间比较采用单因素方差分析并采用Tukey法进行两两比较。结果:与癌旁组织(1.00±0.33)比较,食管癌组织中SOD1 mRNA表达升高(3.76±0.51,t=-25.595,P<0.01)。食管癌中SOD1蛋白表达水平(1.93±0.35)也高于癌旁组织(1.00±0.28,t=-11.734,P<0.01)。转染24、48、72 h后,AAV-SOD1组细胞活性(0.165±0.016、0.283±0.023、0.391±0.036)明显高于对照组(0.133±0.017、0.213±0.028、0.326±0.038)和AAV-GFP组(0.129±0.015、0.209±0.024、0.316±0.032,P<0.01)。转染48 h后,AAV-SOD1组细胞ROS水平(0.757±0.062)明显低于对照组(1.000±0.079)和AAV-GFP组(1.048±0.108,P<0.01),而SOD1、Wnt5a和β-catenin蛋白表达水平则明显高于对照组和AAV-GFP组(P<0.01)。结论:SOD1的表达在食管癌中升高,过表达SOD1通过抑制ROS和调节Wnt信号通路活化诱导细胞增殖。
Objective To investigate the expression of copper/zinc superoxide dismutase(SOD1)in esophageal cancer and its effect and mechanism on cell proliferation.Methods Thirty-two esophageal cancer tissues and adjacent tissues were selected.SOD1 expression was analyzed using real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR),Western blotting and immunohistochemistry methods.Esophageal cancer cell lines were randomly allocated into three groups:control,adeno-associated virus(AAV)-green fluorescent protein(GFP)group and AAV-SOD1 group.Cells in AAV-GFP group and AAV-SOD1 group were transfected with AAV-GFP and AAV-SOD1 recombinant viral vectors,while control was treated with phosphate buffered saline(PBS).Cell viabilities were evaluated with methyl thiazolyl tetrazolium(MTT)assay,reactive oxygen species(ROS)production was detected with dihydroethidium(DHE)probe,and protein expression of Wnt5a andβ-catenin was analyzed with Western blotting.Results Compared with adjacent tissues(1.00±0.33),expression of SOD1 mRNA(3.76±0.51)was enhanced in esophageal cancer tissues(t=-25.595,P<0.01).SOD1 protein expression was also higher in cancer tissues(1.93±0.35)than in adjacent tissues(1.00±0.28,t=-11.734,P<0.01).At 24,48 and 72 h after transfection,cell viabilities in AAV-SOD1 group(0.165±0.016,0.283±0.023,0.391±0.036)were markedly higher than those in control(0.133±0.017,0.213±0.028,0.326±0.038)and AAV-GFP group(0.129±0.015,0.209±0.024,0.316±0.032,P<0.01).At 48 h after transfection,ROS levels in AAV-SOD1(0.757±0.062)were lower than control(1.000±0.079)and AAV-GFP group(1.048±0.108,P<0.01),while protein expression of SOD1,Wnt5a andβ-catenin was higher(P<0.01).Conclusion SOD1 expression was increased in esophageal cancer,and SOD1 overexpression seduced cell proliferation via inhibition of ROS and modulation of Wnt signaling pathway.
作者
张春敭
柳紫阳
盛银良
吴彬
宋亚男
叶贯超
齐宇
Zhang Chunyang;Liu Ziyang;Sheng Yinliang;Wu Bin;Song Ya’nan;Ye Guanchao;Qi Yu(Department of Thoracic Surgery,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Thoracic Surgery,Hami Central Hospital,Hami 839000,China)
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2020年第10期1855-1857,共3页
Chinese Journal of Experimental Surgery
基金
河南省高等学校重点科研项目(19A320053)。
关键词
铜锌超氧化物歧化酶
食管癌
细胞增殖
WNT
活性氧簇
Copper/zinc superoxide dismutase
Esophageal cancer
Cell proliferation
Wnt
Reactive oxygen species