Institute of Biological Sciences, Division of Genetics and Molecular Biology, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
Institute of Biological Sciences, Division of Genetics and Molecular Biology, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia; Centre for Research in Biotechnology for Agriculture (CEBAR), University of Malaya, 50603 Kuala Lumpur, Malaysia.
Lung Cancer. 2014 Feb;83(2):154-62. doi: 10.1016/j.lungcan.2013.11.024. Epub 2013 Dec 6.
Dysregulation in miRNA expression contributes towards the initiation and progression of metastasis by regulating multiple target genes. In this study, variations in miRNA expression profiles were investigated between high and low invasive NSCLC cell lines followed by identification of miRNAs with targets governing NSCLC's metastatic potential.
Two NSCLC sub-cell lines possessing opposing migration and invasion properties were established using serial transwell invasion assays. Global miRNA expression profiles were obtained using microarray followed by RT-qPCR validation. Target prediction and pathway enrichment analyses were conducted on dysregulated miRNAs using DIANA-mirPath, DIANA-microT 4.0 and TargetScan 5.2 softwares. Metastatic effects of dysregulated miRNAs were evaluated using wound healing assay, invasion assay and HUVEC angiogenesis assay following transfection with mimics and inhibitors.
A total of eleven differentially expressed miRNAs were revealed from microarray analyses, with four miRNAs validated through RT-qPCR. Three of these miRNAs were further selected for biological function validations, with only two modulating metastasis. A pathway model describing interactions between miRNAs and metastasis highlighted four major pathways: non-canonical Wnt/PCP, TGF-β, MAPK and integrin-FAK-Src signalling cascade.
These results provide a list of potential candidate metastatic markers during the classification of NSCLCs and a platform for the development of bio-therapeutics targeting these miRNA control elements.
miRNA 表达失调通过调控多个靶基因，促进转移的发生和进展。本研究通过比较高侵袭性和低侵袭性 NSCLC 细胞系之间的 miRNA 表达谱差异，鉴定调控 NSCLC 转移潜能的 miRNA。
采用连续 Transwell 侵袭实验，建立具有相反迁移和侵袭特性的两个 NSCLC 亚系。利用微阵列获得全局 miRNA 表达谱，然后通过 RT-qPCR 进行验证。利用 DIANA-mirPath、DIANA-microT 4.0 和 TargetScan 5.2 软件对失调 miRNA 进行靶预测和通路富集分析。转染 mimics 和抑制剂后，通过划痕愈合实验、侵袭实验和 HUVEC 血管生成实验评估失调 miRNA 的转移效应。
通过微阵列分析共发现 11 个差异表达的 miRNA，其中 4 个 miRNA 通过 RT-qPCR 验证。进一步选择这三个 miRNA 进行生物学功能验证，只有两个调节转移。描述 miRNA 与转移相互作用的通路模型突出了四个主要通路：非经典 Wnt/PCP、TGF-β、MAPK 和整合素-FAK-Src 信号级联。
这些结果为 NSCLC 分类过程中的潜在候选转移标志物提供了一个列表，并为针对这些 miRNA 调控元件的生物治疗药物的开发提供了一个平台。
