lncRNAGAS5通过下调miR-223的表达提高结肠癌细胞的放射敏感性

IncRNA GAS5 enhances the radiosensitivity of colon cancer cells by down-regulating miR-223expression

目的 探究生长特异抑制物5（lncRNA growth arrest-specific 5，lncRNA GAS5）通过靶向调控miR-223的表达对结肠癌细胞的放射敏感性的影响。方法 采用荧光定量PCR（qPCR）检测多种结肠癌细胞系中lncRNA GAS5的表达量，选择表达量较低的作为后续研究对象；细胞克隆实验检测过表达lncRNA GAS5对结肠癌SW480细胞放射敏感性的影响；通过生物信息学数据库starBase以及双荧光素酶报告基因实验预测以及验证lncRNA GAS5的靶基因miR-223；qPCR检测miR-223在多种结肠癌细胞系中的表达量，以及过表达lncRNA GAS5对SW480细胞中miR-223表达量的影响。结果 与人正常结肠上皮细胞（NCM460）相比，lncRNA GAS5在结肠癌SW480、LOVO、HT-29、SW620细胞系中的表达量显著降低（t=15.25、8.69、14.42、11.62，P〈0.05），其中在SW480细胞中的表达量最低；过表达lncRNA GAS5或者下调miR-223显著降低细胞的存活分数（8 Gy时lncRNA GAS5：t=13.51，P〈0.05；anti-miR-223：t=14.93，P〈0.05），促进凋亡（lncRNA GAS5：t=8.30，P〈0.05；anti-miR-223：t=7.32，P〈0.05），增加结肠癌细胞放射敏感性；生物信息学分析显示，lncRNA GAS5 3＇端序列中包含与miR-223的结合位点，过表达/下调lncRNA GAS5后，miR-223的表达量下降/上升。结论 lncRNA GAS5通过靶向调控miR-223的表达促进结肠癌细胞凋亡、抑制其存活，从而提高结肠癌细胞的放射敏感性。

Objective To explore the effect of lncRNA of growth arrest-specific 5 （lncRNA GAS5） on the radiosensitivity of colon cancer cells by targeting miR-223. Methods The expressions of lncRNA GAS5 in a few of colon cancer cell lines were detected by real-time quantitative PCR （qPCR）. The cell lines with low expression level of lncRNA GAS5 were selected for subsequent study. The effect of overexpression of lncRNA GAS5 on the radiosensitivity of colon cancer SW480 cells was detected by cell cloning experiments. The target gene miR-223 of lncRNA GAS5 was predicted and validated by the bioinformatics database starBase and dual luciferase reporter assays. qPCR was used to detect the expression of miR-223 in various colon cancer cell lines and the influence of lncRNA GAS5 overexpression on the expression of miR-223 in SW480 cells. Results Compared with normal human colonic epithelial cells （NCM460）, the expressions of lncRNA GAS5 in the colon cancer SW480, LOVO, HT-29 and SW620 cell lines were significantly lower（t=15.25, 8.69, 14.42, 11.62, P〈0.05）, with the lowest level in SW480 cells. Both overexpression of lncRNA GAS5 and down-regulation of miR-223 significantly increased the radiosensitivity of colon cancer cells by decreasing cell survival fraction（at 8 Gy, lncRNA GAS5, t=13.51, P〈0.05;anti-miR-223, t=14.93, P〈0.05）and promoting apoptosis（lncRNA GAS5, t=8.30, P〈0.05;anti-miR-223, t=7.32, P〈0.05）. Bioinformatics analysis showed that the 3＇sequence of lncRNA GAS5 contained the binding sites with miR-223. After overexpression or downregulation of lncRNA GAS5, the expression of miR-223 was enhanced or reduced. Conclusions The lncRNA GAS5 promotes the apoptosis of colon cancer cells and inhibits its survival by targeting miR-223 expression, thereby increases the radiosensitivity of colon cancer cells.