摘要
目的研究EFNB2对膀胱癌细胞增殖、凋亡、侵袭和迁移等生物学功能的影响,探讨其作为免疫治疗的可能性。方法以膀胱癌细胞系T24、5637、J82和UM-UC-3细胞为研究对象,选用人膀胱上皮永生化细胞系SV-HUC-1细胞作为阴性对照。通过慢病毒载体转染shRNA-EFNB2进行敲低实验,然后分别采用qRT-PCR和Western blot检测细胞中EFNB2的mRNA和蛋白表达水平;CCK8和克隆形成实验检测细胞增殖活力;流式细胞术检测细胞周期和细胞凋亡率;Transwell和细胞划痕实验检测细胞侵袭和迁移能力;Western blot检测Bcl2、Bax、Vimentin和E-cadherin的蛋白表达水平。结果与人膀胱上皮永生化细胞系SV-HUC-1细胞相比,膀胱癌细胞系中EFNB2的表达水平显著增高,其中以T24细胞表达最高(P<0.01)。敲低T24细胞系EFNB2基因表达后,T24细胞增殖活力显著下降(P<0.05);细胞周期虽然尚未有显著变化,但细胞凋亡率显著增高(P<0.05),并且抗凋亡蛋白Bcl2表达显著下降(P<0.05),促凋亡蛋白Bax表达显著升高(P<0.01);T24细胞侵袭(P<0.05)和迁移(P<0.05)能力均显著下调,并且对EMT有抑制作用的分子E-cadherin表达升高而促进EMT的分子Vimentin表达则下降,差异均有统计学意义(P<0.01)。结论EFNB2在膀胱癌细胞系T24细胞的增殖、侵袭和迁移等生物学功能中发挥重要作用,为膀胱癌免疫治疗的潜在靶标。
This study was designed to investigate the effect of ephrin-B2(EFNB2)on the proliferation,apoptosis,invasion and migration of bladder cancer cell,and explore the possibility of EFNB2 as an immunotherapeutic target.Bladder cancer cell lines T24,5637,J82 and UM-UC-3 were introduced as experimental subjects,and human bladder epithelial immortalized cell line SV-HUC-1 was used as negative control.ShRNAEFNB2 was transfected with lentivirus vector for knockdown test,and the mRNA and protein expression levels of EFNB2 were detected by qRT-PCR and Western blotting,respectively.CCK8 and clone formation assay were used to detect cell proliferation activity.Cell cycle and apoptosis rate were detected by flow cytometry.Transwell and cell scratch assay were used to detect cell invasion and migration;Western blot was used to detect protein expression levels of Bcl2,Bax,Vimentin and E-cadherin.Data showed that compared with cell line SV-HUC-1,the expression level of EFNB2 in bladder cancer cell line was significantly increased,and T24 cell was the highest.After knockdown of EFNB2 gene expression in T24 cell line,T24 cell proliferation activity decreased significantly.Although the cell cycle was not significantly changed,the apoptosis rate was significantly increased,the expression of anti-apoptotic protein Bcl2 was significantly decreased,and the expression of pro-apoptotic protein Bax was significantly increased.The invasion and migrationabilities of T24 cells were significantly down-regulated,and the expression of E-cadherin,which inhibits EMT,was increased,while Vimentin,which promotes EMT,was decreased.In conclusion,EFNB2 plays an important role in the proliferation,invasion,migration and otherbiological functions of bladder cancer cell line T24 cells,and could be a potential target for immunotherapy ofbladder cancer.
作者
郭江
张克勤
GUO Jiang;ZHANG Keqin(Urinary Nephropathy Center,Second Affiliated Hospital of Chongqing Medical University,Chongqing 400065,China)
出处
《免疫学杂志》
CAS
CSCD
北大核心
2022年第3期250-256,共7页
Immunological Journal
基金
国家自然科学基金面上项目(81670684)
重庆市集成示范计划(CSTC2015jcsf0054)。
