摘要
目的:探讨敲减泛素结合酶E2T(UBE2T)的表达对肺癌细胞A549生长、侵袭和裸鼠成瘤性的影响,初步阐明其作用机制。方法:靶向UBE2T基因的慢病毒载体(shRNA1、shRNA2、shRNA3)及阴性对照慢病毒载体(shRNA-NC)转染人非小细胞肺癌(NSCLC)A549细胞系,筛选干扰效果较好的shRNA3进行后续实验。CCK-8法、流式细胞术、Transwell小室法检测细胞的增殖、凋亡及侵袭能力,Western blot法检测相关蛋白表达。检测小鼠体内的成瘤质量,免疫组化法检测瘤体组织中Ki67、VEGF蛋白表达情况。结果:与shRNA-NC组相比,UBE2T-shRNA3组各时间点的细胞增殖率、穿膜细胞数及Ki67、VEGF、N-cad蛋白表达明显下降(P<0.05),细胞凋亡率、p21、cleaved cas3/cas3、cleaved cas9/cas9明显上升(P<0.05)。体内实验结果显示,与shRNA-NC组相比,UBE2T-shRNA3组小鼠的瘤体质量及Ki67、VEGF蛋白阳性表达率明显下降(P<0.05)。结论:敲减UBE2T的表达可抑制肺癌细胞A549的增殖、侵袭,促进细胞凋亡,抑制裸鼠成瘤。
Objective:To explore the effects of knocking-down expression of ubiquitin-conjugating enzyme E2T(UBE2T)on growth,invasion and nude mouse tumorigenicity of lung cancer cells A549,and preliminarily clarify its action mechanism.Methods:The lentiviral vectors targeting UBE2T gene(shRNA1,shRNA2,shRNA3)and lentiviral vector of negative control(shRNA-NC)were transfected into human non-small cell lung cancer(NSCLC)A549 cell lines.The shRNA3 with better interference effect was screened out for follow-up experiments.The proliferation,apoptosis and invasion abilities of cells were detected by CCK-8,flow cytometry and Transwell chamber assay.The expressions of related proteins were detected by Western blot.The mass of formation tumors in mice was detected.The expressions of Ki67 and VEGF proteins in tumor tissues were detected by immunohistochemical method.Results:Compared with shRNA-NC group,cells proliferation rate,number of penetrating cells,and expressions of Ki67,VEGF and N-cad proteins were significantly decreased in UBE2T-shRNA3 group(P<0.05),while apoptosis rate,p21,cleaved cas3/cas3 and cleaved cas9/cas9 were significantly increased(P<0.05).The results of in vivo experiments showed that compared with shRNA-NC group,tumor mass,and the positive expression rates of Ki67 and VEGF proteins were significantly decreased in UBE2T-shRNA3group(P<0.05).Conclusion:Knocking-down the expression of UBE2T can inhibit the proliferation and invasion of lung cancer cells A549,promote apoptosis,and inhibit nude mouse tumorigenicity.
作者
段方方
赵晓丽
侯小霞
王留晏
陈露
周寒丽
张玉洁
孔天东
DUAN Fangfang;ZHAO Xiaoli;HOU Xiaoxia;WANG Liuyan;CHEN Lu;ZHOU Hanli;ZHANG Yujie;KONG Tiandong(Department of Oncology,the Third People's Hospital of Zhengzhou,Henan Province,Zhengzhou 450000,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2022年第20期2490-2494,2500,共6页
Chinese Journal of Immunology
基金
河南省医学科技攻关计划项目(2018020748)。
关键词
泛素结合酶E2T
人非小细胞肺癌A549细胞系
肺癌
增殖
凋亡
侵袭
Ubiquitin-conjugating enzyme E2T
Human non-small cell lung cancer A549 cell line
Lung cancer
Proliferation
Apoptosis
Invasion
