LincRNA-p21 Inhibits The Progression Of Non-Small Cell Lung Cancer Via Targeting MiR-17-5p

BackgroundNon-small-cell lung cancer(NSCLC)is well established as one of the major subtypes of human lung cancer.NSCLC is characterized by a high incidence rate and poor patient prognosis.Previous studies have identified that long intergenic non-coding RNA-p21(lincRNA-p21)serves a key role in the development of tumor and malignant metastasis.However,the majority of the underlying mechanisms for lincRNA have not been completely elucidated.Objective1.To measure the expression of LincRNA-p21 in NSCLC tumor tissues in comparison to adjacent healthy tissues.2.To investigate the effect of miR-17-5p and LincRNA-p21 in cancer cell proliferation,migration,invasion and,apoptosis.3.To checkout the relationship between miR-17-5p and LincRNA-p21.Methods(1)Here,the expression of LincRNA-p21 in NSCLC tumor tissues in comparison to adjacent healthy tissues was measured by qRT-PCR.And the deregulation of lincRNA-p21 in NSCLC tumor tissues in comparison to adjacent healthy tissues was examined using reverse transcription-quantitative polymerase chain reaction.(2)Furthermore,the effect of lincRNA-p21 over expression and knockdown on different NSCLC cell lines was further investigated in vitro.Then we got four groups:1?original A549;2?A549+pcDNA-LincRNA-p21;3?original PC9;4?PC9+si-LincRNA-p21.The cell proliferation was detected by using a Cell Counting Kit-8 assay,and transwell chambers were used to analyze the cells invading ability.An Annexin V-FITC and propidium iodide(PI)staining kit was used to detect the cell apoptosis.(3)Bionformatics analysis was used to predict targets of miR-17-5p.To verify whether miR-17-5p was a direct gene target of lincRNA-p21,and pmirGLO was used to construct a lincRNA-p21 luciferase reporter with binding site for miR-17-5p.The influence of miR-17-5p on different NSCLC cell lines were also studied in vitro using miR-17-5p mimics and inhibitors.Then we had six groups:1-4;5?A549+pcDNA-LincRNA-p21+miR-17-5p mimic;6?PC9+si-LincRNA-p21 + miR-17-5p inhibitor.Cell Counting Kit-8 assay,transwell chambers and Annexin/PI were used again.(4)Western blot analysis for Bcl-2 and MMP9 protein expression levels in different cell groups following treatments with miR-17-5p mimics or inhibitor.(5)The relationship between SMOC1 and miR-17-5p was analyzed by bioinformatics and luciferase reporter analysis.(6)In order to decteced the effect of lincRNA-p21 on lung tumor growth in vivo,18 BALB/c female nude mice were treated by tumor cells.Results(1)The results demonstrated a significant low-expression of LincRNA-p21 in NSCLC tumor tissues.(2)LincRNA-p21 presented efficient inhibition on the progression of lung cancer cells by suppressing cell proliferation,migration and invasion and promoting cell apoptosis.(3)An evident negative correlation between LincRNA-p21 and miR-17-5p expressions was observed,and the inhibition effect of overexpressed LincRNA-p21 on lung cancer cells could be counteracted by miR-17-5p.Bioinformatics and luciferase reporter analysis results confirmed that miR-17-5p is a direct target for LincRNA-p21.This study provides the first evidence for LincRNA-p21 to inhibit the progression of NSCLC via direct targeting miR-17-5p associated signaling pathway.(4)The western blot analysis results demonstrate that the overexpressed lincRNA-p21 significantly suppressed the expression levels of Bcl-2 and MMP9 compared with the control group,and following the addition of miR-17-5p mimics,the expression levels of Bcl-2 and MMP9 were partially recovered,whereby the expression levels of Bcl-2 and MMP9 were upregulated.Furthermore,the knockdown of si-lincRNA-p21 resulted in a significant increase in Bcl-2 and MMP9 levels compared with the NC group,whereas the addition of miR-17-5p inhibitors caused a reduction in the expression levels of Bel-2 and MMP9 compared with that in the si-lincRNA-p21+miR-17-5p NC group.These results indicated that lincRNA-p21 affected lung cancer cells through altering the expression of Bcl-2 and MMP9.(5)The bioinformatics analysis results identified the recognition sequence position 127-133 of SMOC1 for miR-17-5p.The dual-luciferase reporter analysis results provided evidence that SMOC1 was a direct gene target for miR-17-5p.(6)LincRNA-p21 inhibits lung tumor growth in vivo.Conclusion1.The lincRNA-p21 exhibited a significantly higher expression level in normal tissues compared with tumor tissues.2.Overexpression of LincRNA-p21 can significantly inhibit the proliferation and invasion of tumor cells.3.There is a pathway of LincRNA-p21?miR-17-5p?SMOC1 which can affects the progression of non-small cell lung cancer.