circ_0009910对结直肠癌细胞增殖、凋亡的影响及机制

Effect and mechanism of circ_0009910 on proliferation and apoptosis of colorectal cancer cells

目的探讨环状RNA_0009910(circ_0009910)对结直肠癌细胞(SW620)增殖、凋亡的影响及机制。方法常规培养SW620细胞,将细胞随机分组并进行转染,si-NC组转染si-NC、si-circ_0009910组转染si-circ_0009910、pcDNA组转染pcDNA、pcDNA-circ_0009910组转染pcDNA-circ_0009910、miR-NC组转染miR-NC、miR-198组转染miR-198 mimic、si-circ_0009910+anti-miR-NC组共转染si-circ_0009910和anti-miR-NC、si-circ_0009910+anti-miR-198组共转染si-circ_0009910和miR-198 inhibitor。转染48 h,用qRT-PCR检测细胞中circ_0009910、miR-198表达,用MTT法检测细胞增殖能力,用Annexin V-FITC/PI双染法检测细胞凋亡能力,用Western blotting法检测细胞核相关抗原Ki-67(Ki-67)、B细胞淋巴瘤/白血病-2(Bcl-2)蛋白、Bcl-2相关X蛋白(Bax)表达,采用双荧光素酶报告实验验证circ_0009910的靶基因。结果与si-NC组比较,si-circ_0009910组细胞增殖能力低,细胞凋亡率高,Ki-67、Bcl-2蛋白相对表达量低,Bax蛋白相对表达量高(P均<0.05)。Circular RNA Interactome软件预测出circ_0009910的部分碱基序列可与miR-198形成互补配对。与miR-NC组比较,miR-198组转染WT-circ_0009910后荧光素酶活性低(P<0.05);而两组转染MUT-circ_0009910后荧光素酶活性比较差异无统计学意义(P>0.05)。与pcDNA组比较,pcDNA-circ_0009910组miR-198相对表达量低(P<0.05);与si-NC组比较,si-circ_0009910组miR-198相对表达量高(P<0.05);与miR-NC组比较,miR-198组SW620细胞增殖能力低,细胞凋亡率高,Ki-67、Bcl-2蛋白相对表达量低,Bax蛋白相对表达量高(P均<0.05);与si-circ_0009910+anti-miR-NC组比较,si-circ_0009910+anti-miR-198组细胞增殖能力高,细胞凋亡率低,Ki-67、Bcl-2蛋白相对表达量高,Bax蛋白相对表达量低(P均<0.05)。结论circ_0009910低表达通过靶向调控miR-198的表达抑制SW620细胞增殖,促进其促凋亡。

Objective To investigate the effect and mechanism of circular RNA_0009910(circ_0009910)on the proliferation and apoptosis of colorectal cancer cells(SW620).Methods SW620 cells were cultured routinely and randomly grouped and transfected.The cells in the si-NC group were transfected with si-NC,si-circ_0009910 group with si-circ_0009910,the pcDNA group with pcDNA,pcDNA-circ_0009910 group with pcDNA-circ_0009910,miR-NC group with miR-NC,and miR-198 group with miR-198 mimic,and the cells in the si-circ_0009910+anti-miR-NC group were co-transfected with si-circ_0009910 and anti-miR-NC,and the si-circ_0009910+anti-miR-198 group with si-circ_0009910 and miR-198 inhibitor.After transfection for 48 h,the expression of circ_0009910 and miR-198 in the cells was detected by qRT-PCR,the cell proliferation was measured by MTT,apoptosis was determined by Annexin V-FITC/PI double staining,and the expression levels of nuclei-related antigen Ki-67(Ki-67),B-cell lymphoma/leukemia-2(Bcl-2)protein,Bcl-2 related X protein(Bax)were detected by Western blotting.Using dual luciferase report experiment to determine the target genes of circ_0009910.Results Compared with the si-NC group,the si-circ_0009910 group had lower cell proliferation,higher apoptosis rate,lower relative expression of Ki-67 and Bcl-2 protein,and higher relative expression of Bax protein(all P<0.05).Circular RNA Interactome software predicted that part of the base sequence of circ_0009910 could form a complementary pair with miR-198.Compared with the miR-NC group,the luciferase activity of the miR-198 group after transfection with WT-circ_0009910 was lower(P<0.05);no significant difference in luciferase activity after transfection with MUT-circ_0009910 was found between the two groups(P>0.05).Compared with the pcDNA group,the relative expression of miR-198 in the pcDNA-circ_0009910 group was lower(P<0.05);compared with the si-NC group,the relative expression of miR-198 in the si-circ_0009910 group was higher(P<0.05).Compared with the miR-NC group,the SW620 cell proliferation ability of the miR-198 group was lower,the apoptosis rate was higher,the relative expression of Ki-67 and Bcl-2 protein was lower,and the relative expression of Bax protein was higher(all P<0.05).Compared with the si-circ_0009910+anti-miR-NC group,the si-circ_0009910+anti-miR-198 group had higher cell proliferation ability,lower apoptosis rate,higher relative expression of Ki-67 and Bcl-2 protein,and lower relative expression of Bax protein(all P<0.05).Conclusion The low expression of circ_0009910 inhibits the proliferation of SW620 cells and promotes its apoptosis through targeted regulation of miR-198 expression.