环状RNAcirc-ccnb1通过抑制细胞周期蛋白核转位对食管鳞癌增殖和迁移的影响

Circular RNA affect the proliferation and migration of esophageal squamous cell carcinoma by inhibiting cell cycle protein nuclear translocation

目的探讨环状RNA circ-ccnb1对细胞周期蛋白(cyclinb1、ccnb1)的作用,以及对食管鳞癌增殖和迁移的影响。方法①收集60例食管鳞癌组织及癌旁正常组织标本,按癌组织及正常组织分为两组,通过免疫组化检测两组组织中ccnb1的表达,并分析食管鳞癌中ccnb1表达与临床病理因素(淋巴结转移、病理分期)之间的相关性。②培养食管鳞癌细胞,并分为两组:正常食管上皮细胞(Het1-A)和食管鳞癌细胞109(EC109)。采用蛋白免疫印迹法(WB)检测两组中ccnb1蛋白的表达。③用携带circ-ccnb1的质粒转染EC109,检测其中ccnb1的核转位情况,将细胞分为3组:未转染组(Blank组)、转染空载体组(Vector组)及转染circ-ccnb1组(cir-ccnb1组)。转染成功后进行核桨分离,采用WB检测转移至胞核内的ccnb1蛋白量的变化,细胞增殖实验(CCK-8法)及划痕实验检测细胞增殖率、迁移率。结果食管鳞癌组织中ccnb1蛋白的表达高于正常组织,EC109细胞中ccnb1蛋白的表达高于Het1-A细胞(均P<0.05)。与Blank组相比,cir-ccnb1组细胞中ccnb1蛋白的核转位水平明显降低,细胞增值率、迁移率降低(均P<0.05)。结论cir-ccnb1可以通过抑制ccnb1在食管鳞癌细胞中的核转位进而影响食管鳞癌的发展,即抑制癌细胞增殖和迁移。

Objective To investigate the relationship between circular RNA circ-ccnb1 and cyclinb1 and its effect on the development(proliferation and migration)of esophageal cancer.Methods 60 cases of esophageal squamous cell carcinoma and adjacent normal tissues were collected and divided into two groups.The expression of ccnb1 in the two groups was detected by immunohistochemistry,and the correlation between the expression of ccnb1 and clinicopathological factors(lymph node metastasis,pathological stage)was analyzed.Esophageal squamous cell carcinoma cells were cultured and divided into two groups:normal esophageal epithelial cells(Het1-A)and esophageal squamous cell carcinoma cells 109(Ec109).The expression of ccnb1 protein was detected by Western blot.The cells were divided into three groups:blank group,vector group and cir ccnb1 group.WB was used to detect the change of ccnb 1 protein transferred into the nucleus.Cell proliferation rate(CCK-8 method)and cell migration rate were detected by cell proliferation assay(CCK-8 method)and scratch test.Results The expression level of ccnb1 protein in esophageal squamous cell carcinoma tissues was higher than that in normal tissues(P<0.05),and the expression level of ccnb1 protein in EC109 cells was higher than that in Het1-A cells(P<0.05).Compared with the control group,the nuclear translocation level of ccnb1 protein in circ-ccnb1 cells was significantly lower(P<0.001),and the cell proliferation rate and migration rate were both lower(P<0.05).Conclusion Circ-ccnb1 can affect the development of esophageal squamous cell carcinoma by inhibiting the nuclear translocation of ccnb1 in esophageal squamous cell carcinoma cells,that is,inhibiting proliferation and migration.