高压氧联合纳米二氧化硅粒子对结肠癌细胞Colon-26的抑制效应

Inhibitory effect of hyperbaric oxygen combined with nano-silica particles on colon cancer cell Colon-26

目的探讨高压氧(HBO)联合纳米二氧化硅粒子(Nano-SiO_(2))对结肠癌细胞Colon-26的抑制效应,并探讨可能的分子机制。方法将结肠癌细胞Colon-26分为对照组、Nano-SiO_(2)处理组、HBO组及HBO与Nano-SiO_(2)联合处理组,采用Lipofectamine 2000转染特定的质粒(miR-218-5p mimics、miR-218-5p inhibitor)或microRNA于细胞内,以MTT法测定不同分组下的细胞量、微列阵检测法测定不同分组下的整体microRNA表达差异、逆转录定量聚合酶链反应及免疫印迹测定不同分子的表达情况。结果HBO组与对照组比较,细胞存活率差异无统计学意义(P>0.05);Nano-SiO_(2)处理可使细胞存活率降至对照组的60%,而联合处理可使细胞存活率降至对照组的30%,差异均有统计学意义,且2种处理组间差异也有统计学意义(P<0.05)。Nano-SiO_(2)处理组及联合处理组有27个microRNA的表达存在明显差异,其中联合处理组细胞中miR-218-5p显著高于HBO组和Nano-SiO_(2)组,差异有统计学意义(P<0.05)。双荧光素酶报告基因系统证实miR-218-5p对CDK6的3’非翻译区具有明显的抑制作用(P<0.05)。microRNA阴性对照转染细胞中,联合处理组较Nano-SiO_(2)组细胞数量显著降低,差异有统计学意义(P<0.05);而miR-218-5p inhibitor转染组中,联合处理组与Nano-SiO_(2)组的细胞数量差异无统计学意义(P>0.05)。结论HBO和Nano-SiO_(2)联合处理可通过miR-218-5p/CDK6信号轴上调抑制结肠癌细胞Colon-26的增殖。

Objective To explore the inhibitory effect of hyperbaric oxygen(HBO)combined with silica nanoparticles(Nano-SiO_(2))on colon cancer cell Colon-26 and the possible molecular mechanism.Methods The colon cancer cells Colon-26 were divided into control group,Nano-SiO_(2) treatment group,HBO group,and combined treatment group of HBO and Nano-SiO_(2).Lipofectamine 2000 was used to transfect specific plasmids or microRNA into cells.MTT method was used to determine the number of cells in different groups.Meanwhile,the overall microRNA expression differences in the different groups were detected by microarray detection,and the expressions of different molecules were determined by qRT-PCR and western blotting.Results There was no significant difference in cell survival rate between the HBO group and the control group(P>0.05).The treatment of Nano-SiO_(2) can reduce the cell survival rate to 60%of the control group,while the combined treatment of HBO and Nano-SiO_(2) can reduce the cell survival rate to 30%of the control group,with statistical significance compared with the control group,and the difference between the two groups was also statistically significant(P<0.05).There were significant differences in the expressions of 27 microRNA between the Nano-SiO_(2) treatment group and the combined treatment group of HBO and Nano-SiO_(2),among which miR-218-5p in the combined treatment group was significantly higher than those in the HBO group and the Nano-SiO_(2) group,and the differences were statistically significant(P<0.05).Dual luciferase reporter gene system confirmed that miR-218-5p had significant inhibitory effect on 3'UTR of CDK6(P<0.05).The number of miR-NC transfected cells in the combined treatment group of HBO and Nano-SiO_(2) was significantly decreased compared with that in the Nano-SiO_(2) group,and the difference was statistically significant(P<0.05).However,comparing the miR-218-5p inhibitor transfection,there was no statistically significant difference in the number of cells between the combined treatment group of HBO and Nano-SiO_(2) and the Nano-SiO_(2) group(P>0.05).Conclusion The combined treatment with HBO and Nano-SiO_(2) can inhibit the proliferation of colon cancer cell lines by up-regulating mir-218-5p/CDK6 signal axis.