摘要
目的观察沉默INK4基因座中反义非编码RNA(ANRILA)对人宫颈癌细胞株He La增殖的影响,并探讨其机制。方法取对数生长期He La细胞分为观察组和对照组,分别ANRIL-siRNA、对照siRNA乱序。转染24、48 h时采用CCK-8法观察两组细胞增殖情况(结果以OD450表示)。转染48 h时采用荧光定量PCR法检测两组细胞ANRIL、p21、周期蛋白依赖性激酶6(CDK6)、周期蛋白依赖性激酶2(CDK2)mRNA相对表达量,采用CHIP实验观察两组细胞p21基因H3K27位点三甲基化状态。结果转染24 h观察组、对照组OD450分别为0.55±0.08、0.84±0.11,转染72 h观察组、对照组OD450分别为1.21±0.12、1.82±0.14。转染24、72 h观察组OD450均低于对照组(P<0.05)。转染48 h观察组ANRIL及CDK6、CDK2 mRNA相对表达量低于对照组(P均<0.05),p21 mRNA相对表达量高于对照组(P均<0.05)。转染48 h两组细胞p21基因H3K27位点三甲基化水平观察组、对照组分别为1.19±0.08、5.62±0.21,观察组低于对照组(P<0.05)。结论沉默ANRIL可抑制宫颈癌He La细胞的增殖。其机制是沉默ANRIL可以降低宫颈癌He La细胞p21基因H3K27位点三甲基化水平,促进p21mRNA表达,抑制CDK6、CDK2 mRNA表达,从而抑制宫颈癌He La细胞增殖。
Objective To observe the effect of silencing ANRIL on the proliferation of human cervical cancer cell line HeLa and to explore its mechanism. Methods The HeLa cells in the logarithmic phase were divided into the observation group and the control group,which were transfected with ANRIL-siRNA and control siRNA,respectively. After 24 and 48 h transfection,the proliferation of the two groups was observed by CCK-8 method（ the result was expressed by OD450）. The expression of ANRIL,p21,cell cycle dependent kinase 6（ CDK6） and cyclin-dependent kinase 2（ CDK2） mRNA was detected by fluorescence quantitative PCR at 48 h after transfection,and CHIP assay was used to detect H3K27me3 in p21 gene of the two groups. Results At 24 h after transfection,OD450 of the observation group and the control group were 0. 55± 0. 08 and 0. 84 ± 0. 11,respectively. At 72 h,OD450 of the observation group and the control group were 1. 21 ± 0. 12 and 1. 82 ± 0. 14,respectively. The OD450 of the observation group was lower than that of the control group at 24 and 72 h after transfection（ all P〈0. 05）. The mRNA expression of ANRIL,CDK6,and CDK2 in the observation group was lower than that in the control group at 48 h after transfection（ all P〈0. 05）,while the expression of p21 in the observation group was higher than that in the control group（ P〈0. 05）. H3K27me3 level of p21 gene in the observation group was 1. 19 ±0. 08,which was lower than that（ 5. 62 ± 0. 21） of the control group at 48 h（ P〈0. 05）. Conclusion Silencing ANRIL can inhibit the proliferation of cervical cancer HeLa cells by reducing the H3K27me3 level in p21 gene,and thus promoting the expression of p21 and inhibiting CDK6 and CDK2 mRNA expression.
出处
《山东医药》
CAS
北大核心
2017年第35期12-14,共3页
Shandong Medical Journal
