摘要
目的:研究CD155通过调控Ras/Erk通路对肺腺癌细胞活性、增殖、凋亡的影响,并探讨其作用的分子机制。方法:通过慢病毒转染法构建过表达或敲低CD155的人肺腺癌H23和H522细胞株,qRT-PCR检测肺腺癌细胞中CD155 mRNA表达水平,CCK-8法检测细胞活力,免疫荧光法测定细胞增殖和凋亡水平;Western blot检测PCNA、caspase-3、Ras、Erk1蛋白的表达及活化水平;小分子药物FTI 277或FR 180204处理过表达CD155的H23和H522细胞抑制Ras及Erk通路,检测对细胞活性、增殖、凋亡的影响;采用慢病毒共转染的方法构建CD155与SPRY2共同过表达的H23细胞系,免疫共沉淀法检测细胞中SPRY2、CD155蛋白间的结合关系,以同样的方法检测对细胞活性、增殖、凋亡的影响。结果:慢病毒转染肺腺癌H23和H522细胞可上调或下调细胞中CD155的表达,差异具有统计学意义(P <0. 05);CD155过表达可促进肺腺癌细胞活性、增殖及PCNA蛋白的表达,抑制细胞凋亡及caspase-3的活化,敲低CD155则表现出相反的作用,差异具有统计学意义(P <0. 05);CD155过表达可激活Ras和Erk1,敲低CD155则抑制Ras和Erk1的激活,差异具有统计学意义(P <0. 05);抑制Ras和Erk通路可抑制过表达CD155引起的细胞增殖及活性的促进作用,缓解过表达CD155诱导的细胞凋亡,差异具有统计学意义(P <0. 05);SPRY2过表达逆转CD155过表达对H23细胞增殖及活性的促进作用,抑制对Ras/Erk通路激活的促进作用,差异具有统计学意义(P <0. 05)。结论:CD155可通过提高Ras/Erk通路的激活程度,促进肺腺癌细胞在体外的增殖及活性,其作用机制可能是部分由CD155与SPRY2蛋白发生相互作用,降低了SPRY2蛋白对Ras/Erk通路的抑制作用引起的。
Objective:To investigate the effects of CD155 on the activity,proliferation and apoptosis of lung adenocarcinoma cells by regulating Ras/Erk pathway,and to explore its molecular mechanism.Methods:Human lung adenocarcinoma H23 and H522 cell lines overexpressing or knocking down CD155 expression were constructed by lentiviral transfection.The expression of CD155 mRNA in lung adenocarcinoma cells was detected by qRT-PCR.Cell viability were detected by CCK-8 assay.Cell proliferation and apoptosis were measured by immunofluorescence.PCNA,caspase-3,Ras,Erk1 protein expression and activation levels were detected by Western blot.Small molecule drug FTI 277 or FR 180204 treated CD155-expressing H23 and H522 cells to inhibit Ras and Erk pathways.The effect of cell activity,proliferation and apoptosis was detected.The H23 cell line overexpressing CD155 and SPRY2 was constructed by co-transfection of lentivirus.The binding relationship between SPRY2 and CD155 protein was detected by immunoprecipitation.The same method detected the effects on cell activity,proliferation,and apoptosis.Results:Lentivirus-transfected lung adenocarcinoma H23 and H522 cells up-regulated or down-regulated the expression of CD155 in the cells,and the difference was statistically significant(P<0.05).CD155 overexpression promoted the activity,proliferation and expression of PCNA protein in lung adenocarcinoma cells,inhibited apoptosis and activation of caspase-3.Knockdown of CD155 showed the opposite effect,and the difference was statistically significant(P<0.05).CD155 overexpression activated Ras and Erk1.Knockdown of CD155 inhibited Ras and Erk1.The difference was statistically significant(P<0.05).Inhibition of Ras and Erk pathways inhibited cell proliferation and activity induced by overexpression of CD155,and alleviated overexpression of CD155-induced apoptosis.The difference was statistically significant(P<0.05).SPRY2 overexpression reversed the effect of CD155 overexpression on the proliferation and activity of H23 cells,and inhibited the activation of Ras/Erk pathway,the difference was statistically significant(P<0.05).Conclusion:CD155 can promote the proliferation and activity of lung adenocarcinoma cells in vitro by increasing the activation of Ras/Erk pathway.The mechanism of action may be due to the interaction between CD155 and SPRY2 protein,which reduces the inhibition of the Ras/Erk pathway by SPRY2 protein.
作者
白桦
张松
肖鹏
栗敏
Bai Hua;Zhang Song;Xiao Peng;Li Min(Department of Oncology,People's Hospital of Zhengzhou,Henan Zhengzhou 450000,China;Department of Radiology,the First Affiliated Hospital of Zhengzhou University,Henan Zhengzhou 450000,China)
出处
《现代肿瘤医学》
CAS
2019年第20期3580-3586,共7页
Journal of Modern Oncology
