MicroRNA-1 And MicroRNA-2 Inhibition In Lung Cancer Cell Invasion And Adhesion Of Biological Research Role

Background and objective MicroRNAs (miRNAs) are small noncoding RNA gene products approximately19-22nucleotide single stranded RNAs that are found in diverse organisms regulating genes by either inducing mRNA degradation or inhibiting translation,which hence have been implicated in several cellular processes including proliferation, differentiation, apoptosis and development. To date, both in vivo and in vitro studies of lung cancer demonstrate a dysregulation of miRNA expression and suggest its importance in the pathogenesis of lung cancer. Recently, in small cell lung cancer(SCLC) patients, we have identified that patients with high-expression level of miR-1and miR-2have better overall survivals and progressive-free survivals compared to the low-expresion level patients, and this2miRNA signature is an independent predictor of prognosis for SCLC patients. This work was aimed to investigate the effect of overexpressed miR-1and miR-2on the phenotype of lung cancer cell lines in vitro.Methods SCLC cell line NCI-H446and NSCLC cell line NCI-H1299were transient transfected with chemically synthesized mature miR-1and miR-2mimic oligonucleotides by lipofecatamine2000. At48h after transfection, we carried out the validation of expression level for each miRNA in cell lines by real-time RT-PCR. At24h after transfection, the proliferation assay was carried out by manual cell counting; the pre-coated matrigel transwell model was used for the invasion assay; the cell-matrix adhesion assay was used to compare the adhesive ability between each cell line and corresponding control with pre-coated fibemectin96well plate; flow cytometry was used to analyze the distribution of cell cycle with PI staining. At24h after tarnsfection, these cells were treated with cisplatin for24h,and then flow cytometry with PI staining was employed to analyze cell apoptosis.Results Here we show that at48h after transfection,the expression level of miR-1and miR-2in NCI-H446and NCI-H1299cell lines were all significantly upregulated. In in vitro invasion assay, the percentage of invaded cell number from NCI-H446and NCI-H1299cells with transient overexpressed miR-1,miR-2or miR-1/miR-2relative to the invaded number from corresponding control cells was respectively55.0%±8.5%、65.7%±8.5%、71.0%±8.5%and73.6%±3.5%、61.8%±11.1%、83.7%±8.3%; in in vitro adhesion assay, overexpressed miR-1,miR-2or miR-1/miR-2all inhibited cell adhesion to matrix in NCI-H446and NCI-H1299cell lines;and overexpressed miR-2induced cell cycle arrest in H1299cell line, by increasing the percentage of the cells in G2/M phase;however,neither of these miRNAs had significant effect on cell proliferation and apoptosis induced by cisplatin in NCI-H446and NCI-H1299cell lines.Conclusions These findings indicate cancer-protective effect of miR-1and miR-2in SCLC may come from suppression of invasive and adhesive ability of lung cancer cells.