长链非编码RNA CCAT2对肝癌细胞增殖和侵袭迁移能力的影响及其机制探讨

Effects of lncRNA CCAT2 on proliferation,invasion,and migration of hepatocellular carcinoma cells

目的观察长链非编码RNA CCAT2(lncRNA CCAT2)对肝细胞癌(HCC)细胞增殖和侵袭迁移能力的影响,并探讨其机制。方法将对数生长期的HCC细胞Hep G2随机分为A、B、C组,采用Lipofectamine3000分别瞬时转染对照质粒、小干扰RNA质粒(si-CCAT2)、si-CCAT2+多梳蛋白EZH2过表达质粒。采用Real-time PCR法检测细胞中的lncRNA CCAT2、EZH2 mRNA,Western blotting法检测细胞中的EZH2蛋白,MTT法观察培养24、48、72、96 h细胞增殖能力(OD490),细胞划痕及Transwell实验分别观察细胞的迁移、侵袭能力。结果与A组比较,B组细胞中lncRNA CCAT2和EZH2的mRNA和蛋白表达水平均下降(P均<0.05);与B组比较,C组胞中EZH2的mRNA和蛋白表达水平均增加(P均<0.05)。与A组同时点比较,B组细胞增殖的OD490值降低(P均<0.05);与B组同时点比较,C组细胞增殖的OD490值升高(P均<0.05)。与A组比较,B组细胞划痕愈合百分比、穿膜细胞数均下降(P均<0.05);与B组比较,C组细胞划痕愈合百分比、穿膜细胞数均增加(P均<0.05)。结论 lncRNA CCAT2可通过上调EZH2的表达促进HCC细胞的增殖、侵袭和迁移,进而促进HCC的发生发展。

Objective To observe the role and mechanism of lncRNA CCAT2 in proliferation,invasion and migration of hepatocellular carcinoma（HCC） cells. Methods Hep G2 cells in logarithmic phase were randomly divided into groups A,B,and C,which were transiently transfected with control plasmids,small interfering RNA（si-CCAT2）,and si-CCAT2＋ EZH2 overexpression plasmid by using Lipofectamine~3000. The real-time PCR was used to detect the expression of lncRNA CCAT2 and EZH2 mRNA. Western blotting was used to detect the protein level of EZH2. MTT method was used to observe the proliferation of cells at 24,48,72 and 96 h（OD490）. Scratch test and Transwell assay were used to detect cell migration and invasion. Results Compared with group A,the mRNA and protein expression of lncRNA CCAT2 and EZH2 decreased in the group B（both P〈0. 05）. Compared with group B,the mRNA and protien levels of EZH2 increased in the group C（both P〈0. 05）. The OD490 value representing cell proliferation was lower in the group B than that of the group A,and the OD490 value in the group C was higher than that of the group B（all P〈0. 05）. The percentage of wound healing and the number of penetrating cells in the group B were lower than those of the group A（both P〈0. 05）; compared with group B,the percentage of wound healing and the number of penetrating cells were higher in the group C than in the group B（both P〈0. 05）. Conclusions LncRNA CCAT2 can promote the proliferation,invasion,and migration of HCC cells by up-regulating the expression of EZH2,and thus promote the occurrence and development of HCC.