基因间长链非编码RNA152靶向微小RNA?103a?3p对非小细胞肺癌细胞增殖和侵袭迁移的影响

目的探讨基因间长链非编码RNA 152(LINC00152)靶向调控微小RNA-103a-3p(miR-103a-3p)表达及对非小细胞肺癌(NSCLC)细胞增殖和侵袭迁移的影响。方法采用实时定量PCR(QPCR)检测正常肺上皮细胞BEAS-2B及NSCLC细胞(ANIP-973、NCI-H157、A549和NCI-H1975)的LINC00152水平。选取LINC00152水平最高的细胞分别转染LINC00152特异性小干扰RNA(si-LINC00152组)或无关序列(si-NC组),另设未转染细胞为对照组。QPCR检测LINC00152水平,活细胞计数CCK-8法、Transwell小室和划痕实验测定细胞增殖、侵袭和迁移能力,Western blotting检测基质金属蛋白酶(MMP)-2、MMP-9和第10号染色体缺失的磷酸酶及张力蛋白同源基因(PTEN)的水平;荧光素酶报告实验验证LINC00152靶向结合miR-103a-3p的能力。结果NSCLC细胞的LINC00152水平均高于BEAS-2B细胞(P<0.05),尤其是NCI-H1975细胞的最高。si-LINC00152组的LINC00152水平为0.352±0.087,低于对照组的1.058±0.219和si-NC组的1.126±0.139(P<0.05)。与si-NC组和对照组相比,si-LINC00152组NCI-H1975细胞转染48、72 h的增殖活力下降(P<0.05);si-LINC00152组的划痕愈合率和穿膜细胞数分别为(27.386±2.428)%和(78.840±5.031)个,低于si-NC组的(77.675±4.803)%和(179.208±13.264)个及对照组的(76.371±5.385)%和(174.003±15.678)个(P<0.05);与si-NC组和对照组相比,si-LINC00152组的MMP-2和MMP-9水平均降低,而PTEN水平升高(P<0.05)。对照组和si-NC组上述指标的差异无统计学意义(P>0.05)。双荧光素酶报告分析证实,miR-103a-3p模拟物降低了野生型LINC00152的荧光素酶活性(P<0.05),但对突变型无影响(P>0.05)。结论LINC00152在NSCLC细胞中高表达并发挥促癌作用,与NSCLC的迁移侵袭密切相关,LINC00152与miR-103a-3p间的相互作用在NSCLC靶向治疗中有一定潜能。

Targeted regulation of LINC00152 on microRNA-103a-3p and its effect on proliferation,invasion and migration of non-small cell lung cancer cells

Objective To investigate the targeted regulation of long intergenic non-coding RNA 152(LINC00152)on the expression of microRNA-103a-3p(miR-103a-3p)and its effect on the proliferation,invasion and migration of non-small cell lung cancer(NSCLC)cells.Methods Real-time quantitative PCR(QPCR)was used to detect the LINC00152 levels of normal pulmonary epithelial cells BEAS-2B and different NSCLC cells including ANIP-973,NCI-H157,A549 and NCI-H1975.Cells with the highest level of LINC00152 were transfected with LINC00152 specific small interfering RNA(si-LINC00152 group)or unrelated sequence(si-NC group),and cells without transfection were used as control group.The LINC00152 level was detected by QPCR.The abilities of cell proliferation,invasion and migration were investigated by CCK-8,Transwell chamber and scratch experiments.Levels of matrix metalloproteinase(MMP)-2,MMP-9 and phosphatase and tensin homology deleted on chromosome 10(PTEN)were detected by Western blotting.Targeted binding of LINC00152 on miR-103a-3p was verified by luciferase reporter gene experiment.Results LINC00152 levels of NSCLC cells especially NCI-H1975 were higher than that of BEAS-2B cells(P<0.05).The level of LINC00152 in si-LINC00152 group was 0.352±0.087,lower than 1.058±0.219 in control group and 1.126±0.139 in si-NC group(P<0.05).Compared with si-NC group and control group,the proliferation activity of NCI-H1975 cells in si-LINC00152 group decreased after 48 and 72 h-transfection(P<0.05).The scratch healing rate and number of membrane-piercing cells in si-LINC00152 group were(27.386±2.428)%and 78.840±5.031,lower than(77.675±4.803)%and 179.208±13.264 in si-NC group and(76.371±5.385)%and 174.003±15.678 in control group(P<0.05).Compared with si-NC group and control group,decreased levels of MMP-2 and MMP-9 but enhanced level of PTEN were observed in si-LINC00152 group(P<0.05).No significant difference were observed on the above indices between si-NC group and control group(P>0.05).Luciferase analysis showed that miR-103a-3p decreased the luciferase activity of wild-type LINC00152(P<0.05),but had no effect on mutant one(P>0.05).Conclusion LINC00152 is highly expressed in NSCLC cells and plays a role in promoting cancer,which is closely related to the migration and invasion of NSCLC.The interaction between LINC00152 and miR-103a-3p has a certain potential in the targeted therapy of NSCLC.