下调VPS4A表达对食管鳞癌恶性生物学行为影响的实验研究  

The effect of down-regulated VPS4A expression on malignant biological behavior of esophageal squamous cell carcinoma

作　　者：徐冰[1] 陈婷婷[1] 陈慧 孙新臣 成红艳 XU Bing;CHEN Tingting;CHEN Hui;SUN Xinchen;CHENG Hongyan(Department of Oncology, Northern Jiangsu People's Hospital, Yangzhou 225001,China)

机构地区：[1]江苏省苏北人民医院肿瘤科,江苏扬州225001 [2]南京医科大学第一附属医院放疗科,210009 [3]南京医科大学第一附属医院全科医学科,210009

出　　处：《临床肿瘤学杂志》2021年第10期865-870,共6页Chinese Clinical Oncology

基　　金：国家自然青年科学基金资助项目(8210102316);国家自然科学基金资助项目(82073344)。

摘　　要：目的探讨VPS4A对食管鳞癌进展及放射敏感性的影响。方法转染靶向VPS4A的短发夹RNA(shRNA)慢病毒干扰载体至人食管鳞癌细胞Kyse150和Eca109,构建食管鳞癌细胞VPS4A低表达模型,采用实时荧光定量PCR(qPCR)和Western blotting验证转染效率。将食管鳞癌分为对照组(NC组)和干扰组(shVPS4A),并且联合辐照,通过平板克隆形成试验检测各组细胞增殖情况;通过流式细胞术以及transwell实验检测VPS4A表达对细胞凋亡和侵袭迁移能力的影响;裸鼠皮下瘤模型用于实验结果的体内验证。结果与NC组比较,shVPS4A组VPS4A基因和蛋白表达水平明显下降(P<0.05);shVPS4A组在0、2、4、6、8 Gy剂量点的克隆集落数目均少于NC组,差异有统计学意义(P<0.05);对照组Eca109细胞0 Gy和8 Gy凋亡率为(4.00±0.17)%、(13.60±0.53)%,而下调VPS4A表达后0 Gy和8 Gy凋亡率为(8.03±0.17)%、(18.84±0.58)%;对照组Kyse150细胞0 Gy和8 Gy凋亡率为(3.95±0.13)%、(14.82±0.32)%,而下调VPS4A表达后0 Gy和8 Gy凋亡率分别为(7.81±0.29)%、(18.94±0.34)%。shVPS4A组Eca109细胞和Kyse150细胞迁移、侵袭细胞数均低于NC组,差异具有统计学意义(P<0.05)。与NC组比较,shVPS4A组E-cadherin表达升高,N-cadherin和Vimentin表达降低。裸鼠体内成瘤实验结果显示shVPS4A组瘤体质量明显小于NC组。结论下调VPS4A表达可以提高食管鳞癌细胞的辐射敏感性,促进细胞凋亡以及抑制其侵袭迁移能力,VPS4A有潜力成为食管癌治疗新靶标。Objective To investigate the effect of vacuolar protein sorting 4 homolog A(VPS4A)on the progression and radiosensitivity of esophageal squamous cell carcinoma.Methods Human esophageal squamous cell carcinoma cells Kyse150 and Eca109 were transfected with short hair pin RNA(shRNA)interference vector targeting VPS4A to construct esophageal squamous cell carcinoma cell model with low VPS4A expression.The transfection efficiency was verified by real-time quantitative PCR(qPCR)and Western blotting.Esophageal squamous cell carcinoma was divided into control group(NC group)and interference group(shVPS4A group).The proliferation of each group was detected by plate clonogenesis test.The effects of VPS4A on apoptosis,invasion and migration were detected by flow cytometry and Transwell assay.A nude mouse model of subcutaneous tumor was used to verify the results in vivo.Results Compared with NC group,the expression of VPS4A mRNA and protein in shVPS4A group were significantly decreased(P<0.05).The number of clone colonies in shVPS4A group was significantly lower than that in NC group at the dose points of 0,2,4,6 and 8 Gy,with statistical significance(P<0.05).The apoptotic rates of Eca109 cells in NC group were(4.00±0.17)%and(13.60±0.53)%at 0 Gy and 8 Gy,but were(8.03±0.17)%and(18.84±0.58)%at 0 Gy and 8 Gy after VPS4A downregulation.The apoptotic rates of 0 Gy and 8 Gy Kyse150 cells in the control group were(3.95±0.13)%and(14.82±0.32)%,while the apoptotic rates of 0 Gy and 8 Gy Kyse150 cells after VPS4A downregulation were(7.81±0.29)%and(18.94±0.34)%,respectively.The number of migration and invasion cells of Eca109 and Kyse150 cells in shVPS4A group was lower than that in NC group,and the differences were statistically significant(P<0.05).Compared with NC group,e-cadherin expression was increased in shVPS4A group,while N-cadherin and Vimentin expression was decreased.The results of tumor formation experiment in nude mice showed that the tumor mass of shVPS4A group was significantly lower than that of NC group.Concl

关 键 词：食管鳞癌 VPS4A 放射敏感性 凋亡 侵袭 迁移 

分 类 号：R735.1[医药卫生—肿瘤]