长链非编码RNA赖氨酰氧化酶样蛋白1反义RNA在结直肠癌中的表达及对结直肠癌细胞增殖、侵袭和迁移能力的影响

The expression of long non-coding RNA LOXL1-AS1 in colorectal cancer and its effect on the proliferation,invasion and migration of colorectal cancer cells

目的 探究长链非编码RNA(lncRNA)赖氨酰氧化酶样蛋白1反义RNA(LOXL1-AS1)在结直肠癌中的表达及对结直肠癌细胞增殖、侵袭及迁移能力的影响和可能的作用机制。方法 选取2016年1月至2018年12月在河南省人民医院行结直肠癌根治术的56例病人新鲜癌组织,采用实时定量聚合酶链反应(qRT-PCR)检测结直肠癌组织样本及结直肠癌细胞系SW480、SW620、HCT116和Lovo细胞中lncRNA LOXL1-AS1的表达水平,以配对癌旁组织及人正常结直肠黏膜细胞FHC作正常对照;构建靶向LOXL1-AS1序列的小干扰RNA(si-LOXL1-AS1)并转染SW480和Lovo细胞,采用细胞计数试剂盒(CCK-8)、平板克隆实验、划痕实验和Transwell小室法检测沉默lncRNA LOXL1-AS1表达对结直肠癌细胞增殖、侵袭和迁移能力的影响。采用蛋白质印迹法(Western blotting)检测沉默LOXL1-AS1表达后磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路相关蛋白及上皮-间质转化(EMT)相关蛋白的表达情况。结果 LOXL1-AS1在结直肠癌组织中的相对表达水平显著高于癌旁组织(1.37±0.05比0.87±0.05,P<0.05),其表达水平与肿瘤分化程度、TNM分期、肝转移和MSI状态显著相关(均P<0.05);LOXL1-AS1在结直肠癌细胞系(SW480、SW620、HCT116、Lovo)中的表达水平显著高于FHC细胞(1.93±0.10、1.32±0.04、1.50±0.07、1.78±0.09比1.10±0.07,P<0.05),沉默lncRNA LOXL1-AS1表达可抑制SW480细胞和Lovo细胞的增殖、侵袭及迁移能力,E-钙黏蛋白(E-cadherin)表达含量升高,N-钙黏蛋白(N-cadherin)、磷酸化脂酰肌醇3-激酶(p-PI3K)和磷酸化蛋白激酶B(p-Akt)表达含量降低(均P<0.05)。结论 LOXL1-AS1在结直肠癌组织中高表达并与临床病理特征相关,沉默LOXL1-AS1可显著降低结直肠癌细胞的增殖、侵袭及迁移能力,该机制可能与PI3K/Akt信号通路及EMT途径有关。

Objective To explore the expression of long non-coding RNA(lncRNA) Lysyl Oxidase Like Protein 1 antisense RNA 1(LOXL1-AS1) in colorectal cancer(CRC),and to investigate the influence of lncRNA LOXL1-AS1 on the proliferation,invasion and migration of CRC cells and the possible mechanism.Methods CRC tissue samples were taken from 56 patients who underwent colorectal cancer surgery in Henan Provincial People's Hospital from January 2016 to December 2018.The expression of lncRNA LOXL1-AS1 in 56 CRC tissue samples and CRC cell lines SW480,SW620,HCT116 and Lovo were detected by quantitative real-time PCR(qRT-PCR),and the para-carcinoma tissues and human intestinal epithelial cell line FHC were paired as normal controls.Small interfering RNA(si-LOXL1-AS1) targeting the LOXL1-AS1 sequence was constructed and transfected into SW480 and Lovo cells.The effects of silencing lncRNA LOXL1-AS1 expression on the proliferation,invasion and migration of CRC cells were respectively detected by cell counting kit-8(CCK8),colony formation assays,wound-healing and Transwell assay.Western blotting was used to detect the expression of phosphatidylinositol 3-kinase/protein kinase B(PI3K/Akt) signaling pathway related proteins and epithelial-mesenchymal transformation(EMT) related proteins.Results The relative expression level of LOXL1-AS1 in CRC tissues was significantly higher than that in adjacent normal tissues(1.37±0.05 vs.0.87±0.05,P0.05),which was closely related to tumor differentiation degree,TNM staging,liver metastasis and MSI status of CRC(all P0.05).The expressions of lncRNA LOXL1-AS1 in CRC cell lines SW480,SW620,HCT116 and Lovo were significantly higher than that of FHC cells(1.93±0.10,1.32±0.04,1.50±0.07,1.78±0.09 vs.1.10±0.07,P0.05).The expression of silencing lncRNA LOXL1-AS1 could inhibit the proliferation,invasion and migration of SW480 cells and Lovo cells,up-regulate the expression of E-cadherin,down-regulate the expression of N-cadherin,P-PI3K and p-Akt(all P0.05).Conclusions LncRNA LOXL1-AS1 is highly expressed in CRC tissues and is correlated with clinicopathological features of CRC patients.Silencing LOXL1-AS1 can significantly inhibit the proliferation,invasion and metastasis of CRC cells,which may be related to EMT and PI3K/Akt signaling pathways.