LncRNA SNHG25在直肠癌细胞中的表达及其调节miR-330-3p/SMYD3轴对直肠癌细胞迁移及侵袭的影响

Expression of lncRNA SNHG25 in rectal cancer cells and its influences on the migration and invasion of rectal cancer cells by regulating the miR-330-3p/SMYD3 axis

目的探究长链非编码RNA(lncRNA)小核仁RNA宿主基因25(SNHG25)对直肠癌(RC)细胞迁移、侵袭的影响及微小RNA(miR)-330-3p/组蛋白甲基化转移酶3(SMYD3)轴在此过程中所发挥的作用。方法使用Real-Time PCR检测SNHG25在不同细胞系的表达及其沉默效率,Transwell实验检测下调SNHG25对RC细胞迁移和侵袭的影响。再次培养细胞,并依实验目的分为NC组(不作任何处理)、miR-330-3p mimics组(转染miR-330-3p模拟物)、miR-330-3p inhibitor组(转染miR-330-3p抑制物)、sh-SNHG25-1+miR-330-3p inhibitor组(转染sh-SNHG25-1+miR-330-3p抑制物)、SMYD3组(转染pcDNA-SMYD3)、si-SMYD3(转染si-SMYD3)、SMYD3+miR-330-3p mimics(转染pcDNA-SMYD3+miR-330-3p模拟物),Real-Time PCR检测各组细胞中miR-330-3p与SMYD3 mRNA表达;Transwell实验检测各组RC细胞迁移、侵袭能力。结果与人正常肠黏膜上皮细胞系(FHC)细胞比较,RC细胞(SW837、SW1463及HR8348)中SNHG25的表达增高,且在HR8348细胞中表达明显高于其他RC细胞系(3.12±0.17、1.96±0.15、4.87±0.22比1.00±0.03,P<0.05),选择HR8348细胞进行后续实验。与sh-NC比较,sh-SNHG25-1/2/3组细胞中SNHG25表达均被下调,且sh-SNHG25-1下调效率最高(1.02±0.01比0.21±0.02、0.52±0.03、0.36±0.04,P<0.05);sh-SNHG25-1组细胞中RC细胞迁移数量和侵袭数量均减少(205.13±21.23比120.34±11.98和89.92±8.15比30.27±3.59,P<0.05)。双荧光素酶报告基因实验显示,SNHG25与miR-330-3p、miR-330-3p与SMYD3均存在负向调控关系。与NC组比较,miR-330-3p mimics组细胞中miR-330-3p表达增加(1.03±0.01比1.67±0.15,P<0.05),迁移细胞数量和侵袭细胞数量减少(210.08±12.96比131.36±10.45和77.45±8.69比21.74±2.04,P<0.05);miR-330-3p inhibitor组细胞中miR-330-3p表达降低(1.03±0.01比0.43±0.05,P<0.05),迁移细胞数量和侵袭细胞数量增多(210.08±12.96比246.95±16.34和77.45±8.69比122.03±9.51,P<0.05);与miR-330-3p inhibitor比较,sh-SNHG25-1+miR-330-3p inhibitor组细胞中miR-330-3p表达增高(0.43±0.05比0.76±0.04,P<0.05),细胞侵袭和迁移数量减少(246.95±16.34比167.54±11.79和122.03±9.51比52.88±5.07,P<0.05)。与NC组相比,SMYD3组细胞中SMYD3 mRNA表达增高(1.02±0.03比2.34±0.19,P<0.05),细胞的迁移和侵袭数量增多(215.26±15.38比351.04±20.96和86.33±9.65比129.87±11.52,P<0.05);si-SMYD3组细胞中SMYD3 mRNA表达降低(1.02±0.03比0.31±0.04,P<0.05),细胞的迁移和侵袭数量减少(215.26±15.38比128.54±15.47和86.33±9.65比28.92±4.91,P<0.05)。与SMYD3组相比,SMYD3+miR-330-3p mimics组细胞中SMYD3 mRNA表达降低(2.34±0.19比0.72±0.07,P<0.05),细胞的迁移和侵袭数量减少(351.04±20.96比168.08±13.44和129.87±11.52比63.01±5.18,P<0.05)。结论下调SNHG25表达可能通过调控miR-330-3p/SMYD3轴抑制RC细胞侵袭及迁移。

Objective To explore the effects of long non-coding RNA(lncRNA)small nucleolar RNA host gene 25(SNHG25)on the migration and invasion of rectal cancer(RC)cells and the role of the microRNA(miR)-330-3p/SET and MYND domain containing protein 3(SMYD3)axis in this process.Methods Real-Time PCR was used to detect the expression of SNHG25 in different cell lines and its silencing efficiency,Transwell experiment was used to detect the effects of down-regulation of SNHG25 on the migration and invasion of RC cells.The cells were cultured and divided into NC group(without any treatment),miR-330-3p mimics group(transfected with miR-330-3p mimic),miR-330-3p inhibitor group(transfected with miR-330-3p inhibitor),sh-SNHG25-1+miR-330-3p inhibitor group(transfected with sh-SNHG25-1+miR-330-3p inhibitor),SMYD3 group(transfected with pc DNA-SMYD3),si-SMYD3(transfected with si-SMYD3),and SMYD3+miR-330-3p mimics(transfected with pc DNA-SMYD3+miR-330-3p mimics)according to the experimental purpose.Real-Time PCR was used to detect the expression of miR-330-3p and SMYD3 mRNA in each group of cells;Transwell experiment was used to detect the migration and invasion abilities of RC cells in each group.Independent sample t-test was used for comparison between the two groups,single factor analysis of variance was used for comparison between multiple groups,and SNK-q test was used for comparison between the two groups.Results Compared with FHC cells,the expression of SNHG25 in SW837,SW1463,and HR8348(3.12±0.17,1.96±0.15,4.87±0.22 vs 1.00±0.03,P<0.05)were increased,and the expression of SNHG25 in HR8348 cells was significantly higher than that in other RC cell lines,HR8348 cells were to conduct followup experiments.Compared with sh-NC,the expression of SNHG25 in the cells of the sh-SNHG25-1/2/3 group was down-regulated(1.02±0.01 vs 0.21±0.02,0.52±0.03,0.36±0.04,P<0.05),and the down-regulation efficiency of sh-SNHG25-1 was the highest;The number of migration and invasion of RC cells in the sh-SNHG25-1 group decreased(205.13±21.23 vs 120.34±11.98and 89.92±8.15 vs 30.27±3.59,P<0.05).The dual-luciferase reporter gene experiment showed that SNHG25 and SMYD3 all have a negative regulatory relationship with miR-330-3p.Compared with the NC group,the expression of miR-330-3p in the miR-330-3p mimics group was increased(1.03±0.01 vs 1.67±0.15,P<0.05),while the number of migrating cells and invading cells were decreased(210.08±12.96 vs 131.36±10.45 and 77.45±8.69 vs 21.74±2.04,P<0.05);the expression of miR-330-3p in miR-330-3p inhibitor group was decreased(1.03±0.01 vs 0.43±0.05,P<0.05),while the number of migrating cells and invasion were increased(210.08±12.96 vs 246.95±16.34 and 77.45±8.69 vs 122.03±9.51,P<0.05).Compared with miR-330-3p inhibitor,the expression of miR-330-3p in sh-SNHG25-1+miR-330-3p inhibitor group was increased(0.43±0.05 vs 0.76±0.04,P<0.05),and the number of cell invasion and migration in the sh-SNHG25-1 group were decreased(246.95±16.34 vs 167.54±11.79 and122.03±9.51 vs 52.88±5.07,P<0.05).Compared with the NC group,the expression of SMYD3 mRNA in the SMYD3 group was increased(1.02±0.03 vs 2.34±0.19,P<0.05),as well as the number of cell migration and invasion were increased(215.26±15.38 vs 351.04±20.96 and 86.33±9.65 vs 129.87±11.52,P<0.05);the expression of SMYD3 mRNA in the si-SMYD3 group wes decreased(1.02±0.03 vs 0.31±0.04,P<0.05),as well as the number of cell migration and invasion were decreased(215.26±15.38 vs 128.54±15.47 and 86.33±9.65 vs 28.92±4.91,P<0.05).Compared with the SMYD3 group,the expression of SMYD3 mRNA in the SMYD3+miR-330-3p mimics group was decreased(2.34±0.19 vs 0.72±0.07,P<0.05),as well as the number of cell migration and invasion were decreased(351.04±20.96 vs 168.08±13.44 and 129.87±11.52 vs 63.01±5.18,P<0.05).Conclusion Down-regulating the expression of SNHG25 may inhibits the invasion and migration of RC cells by regulating the miR-330-3p/SMYD3 axis.