Hugl-1基因在人大肠癌组织中的表达及其对LS174细胞增殖和迁移能力的影响

Analysis of Hugl-1 gene expression and the biological effects in colorectal cancer

目的研究Hugl-1基因在人大肠癌组织中的表达及其与临床病理的相关性,探讨外源Hugl-1基因转染对大肠癌LS174细胞增殖和迁移能力的影响。方法采用RT-PCR及Western-blot法检测人大肠癌组织及相应癌旁正常组织中mRNA和蛋白质的表达水平。以成功构建的分别能稳定表达外源pcDNA3.0-Hugl-1、空载体pcDNA3.0的大肠癌细胞系和未转染的LS174细胞为研究对象,通过软琼脂集落形成、划痕修复、细胞黏附和Transwell细胞侵袭等一系列细胞学功能实验,观察稳定转染细胞株Hugl-1的表达差异对肿瘤细胞增殖、黏附、运动和侵袭的影响。结果 Hugl-1基因在大肠癌组织中表达下调或缺失,表达水平低于癌旁正常组织。Hugl-1表达降低与肿瘤分期及淋巴结转移相关。RT-PCR、Western-blot显示重组体pcDNA3.0-Hugl-1转染细胞株中目的基因Hugl-1在mRNA和蛋白质水平上表达均较空载体转染细胞株和未转染细胞株增强(P<0.05);对转染前后的细胞观察表明,重组体转染组细胞增殖能力无明显改变,但是其迁移运动和侵袭能力降低,细胞黏附能力增加。结论研究表明,Hugl-1表达下调与大肠癌的发生发展有关,Hugl-1基因表达下调可能使肿瘤细胞的结构稳定性丧失,促使肿瘤细胞扩散。

Objective To determine the expression of Hugl-1 gene in colorectal cancer and the biological behaviors of LS174 before and after transfection in vitro.Methods Reverse transcriptionpolymerase chain reaction （RT-PCR） and Western blots methods were used to detect the expression level of mRNA and protein of Hugl-1 gene in colorectal cancer tissues and according para-cancer normal tissues.RTPCR and Western-blot showed respectively that the Hugl-1 gene were expressed higher in cell lines transfected by pcDNA3.0-Hugl-1 than in cell lines un-transfected and transfected by an empty vector both on mRNA and protein lever.Soft agar colony formation,wound-healing experiment,adhesion assay and Matrigel invasion assays were used in vitro to study the effects of Hugl-1 expression on transfected cells proliferation,adhesion,movement and invasion.Results Hugl-1 was downregulated or lost in most of the tumor samples analysed,and that these losses were associated with advanced stage and particularly with lymph node metastases.Transfected with Hugl-1 eukaryotic expression vector could not affect cell proliferation,but could significantly increase cell adhesion and decrease cell movement and invasion capabilities compared with the untransfected cells.Conclusions Loss of Hugl-1 expression contributes to colorectal cancer progression.Reduced Hugl-1 expression may destabilize epithelial structures and contribute to cancer cell dissemination