下调血管内皮生长因子A对食管癌ECA-109细胞的辐射增敏作用  

Down-regulation of VEGFA increases the radio sensitivity of esophageal cancer ECA-109 cell

作　　者：翁小坤 胡莉钧 孙菲 汪建林 谢凯 孔泽 周梦耘 高留刚 于静萍 Weng Xiaokun;Hu Lijun;Sun Fei;Wang Jianlin;Xie Kai;Kong Ze;Zhou Mengyun;Gao Liugang;Yu Jingping(Department of Radiotherapy,Changzhou No.2 People′s Hospital,Affiliated Hospital of Nanjing Medical University,213003,China)

机构地区：[1]南京医科大学附属常州第二人民医院放疗科,213003

出　　处：《中华放射医学与防护杂志》2020年第11期813-819,共7页Chinese Journal of Radiological Medicine and Protection

基　　金：国家自然科学基金(11705095)江苏省"333工程"第二层次人才科研资助项目(BRA2019025)。

摘　　要：目的下调食管癌ECA-109细胞的血管内皮生长因子A(VEGFA)表达,观察其辐射敏感性的改变并初步探讨其发生机制。方法将食管癌ECA-109细胞分成VEGFA转染组、空载组、X射线组和VEGFA转染组联合X射线组。运用qPCR法检测VEGFA基因的表达;Western blot法检测VEGFA蛋白的表达;细胞增殖实验(CCK8法)测定细胞的增殖变化;克隆形成法分析不同照射剂量(0、2、4、6、8 Gy)细胞的辐射敏感性;流式细胞术分析细胞凋亡变化;免疫荧光法检测γ-H2AX焦点的数量。结果ECA-109细胞VEGFA转染组与空载组比较,VEGFA基因表达明显减少(t=11.98,P<0.05)、VEGFA蛋白表达显著下调(t=12.38,P<0.05);ECA-109转染组细胞相较于空载组细胞,其增殖(A450值)明显受到抑制(t=2.78、7.25、21.93、13.21,P<0.05);与空载组比较,VEGFA转染组ECA-109细胞的D0、Dq、SF2值下降(t=5.83、8.56、7.68,P<0.05),放射增敏比SERD0、SERDq分别为1.41、2.09,凋亡比例明显增加、且联合X射线后ECA-109细胞的凋亡比例进一步增加(t=17.63、22.48、33.87,P<0.05);ECA-109细胞VEGFA转染组和空载组在X射线照射后2 h内细胞核形成的γ-H2AX焦点数量均明显增加,24 h后空载组细胞核的γ-H2AX焦点数量恢复照射前水平,而转染组中的γ-H2AX焦点数量仍高于照射前水平(t=7.00,P<0.05)。结论下调VEGFA能抑制食管癌细胞的增殖,减少细胞集落形成并促进其凋亡,增加食管癌ECA-109细胞的辐射敏感性,其机制与DNA损伤修复密切相关。Objective To observe the effect and underlying mechanism of down-regulation of VEGFA on the radiosensitivity of esophageal cancer ECA-109 cells.Methods Esophageal cancer cells were divided into four groups:sh-VEGFA group,vector control group,X-ray plussh-VEGFA group and X-ray plus vector group.The expressions of VEGFA gene and protein were detected by qPCR and Western blot,respectively.Cell proliferation and survival was measured by CCK8 assay and cloning formation,respectively.Cell apoptosis was detected by flow cytometry,andγ-H2AX foci were detected by immune-fluorescence assay.Results Compared with the vector group,the expression of VEGFA gene was decreased in sh-VEGFA group(t=11.98,P<0.05),and the expression of VEGFA protein was also reduced(t=12.38,P<0.05).After VEGFA being down-regulated,the cell proliferation(A450)was obviously inhibited(t=2.78,7.25,21.93,13.21,P<0.05),and the cell clone formation of the sh-VEGFA group was significantly decreased so that D0,Dqand SF2 of sh-VEGFA group were decreased(t=5.83,8.56,7.68,P<0.05),and SERD0and SERDqwere increased.Compared with the vector group,the apoptosis rate in the sh-VEGFA group and the X-ray group was significantly increased and further increased in the sh-VEGFA plus X-ray group(t=17.63,22.48,33.87,P<0.05),and the number ofγ-H2AX foci in both sh-VEGFA and vector groups were significantly increased within 2 h after X-ray irradiation.At 24 h after irradiation,the number ofγ-H2AX foci returned to normal level in the vector group but remained at a higher level in the sh-VEGFA group(t=7.00,P<0.05).Conclusions Down-regulation of VEGFA inhibits the proliferation and colony formation,promotes apoptosis and hence increases the radiosensitivity of esophageal carcinoma cells via a pathway related to DNA damage repair.

关 键 词：食管癌 血管内皮生长因子A 辐射敏感性 DNA损伤修复 

分 类 号：R735.1[医药卫生—肿瘤]