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Objective To evaluate the effect of microRNA-182-5p (miR-182-5p) on proliferation and apoptosis of non-small cell lung cancer (NSCLC) A549 cells by targeting forkhead box O3a (FOXO3a).Methods The difference of miR-182-Sp expression between human normal lung epithelial cells BEAS-2B and NSCLC cells A549 was compared.The A549 cells were chosen,and miR-182-Sp mimic (miR-182-Sp mimic group),miR-182-Sp inhibitor (miR-182-5p inhibitor group),negative control mimic (NC mimic group) and negative control inhibitor (NC inhibitor group) were transfected respectively.The expression of miR-182-Sp was detected by reverse transcription-polymerase chain reaction (RT-PCR).The protein expression of FOXO3a was detected by Western blotting.The cell proliferation activity was detected by methyl thiazolyl tetrazolium (MTT) method.The cell apoptosis was detected by flow cytometry.The targeted relationship between miR-182-5p and FOXO3a was detected by dual-luciferase experiment.Results The miR-182-5p expression of A549 cells and BEASo2B cells respectively was 3.21 ±0.24 and 1.01 ±0.11,and the difference was statistically significant (t =14.209,P＜0.001).The miR-182-5p expression of NC mimic group,miR-182-5p mimic group,NC inhibitor group and miR-182-5p inhibitor group respectively was 1.09 ± 0.20,12.80 ± 1.10,1.03 ± 0.11and 0.47 ± 0.08,and the difference was statistically significant (F =87.872,P ＜ 0.001).The FOXO3a expression of the above four groups respectively was 118.34 ± 16.71,50.89 ± 11.58,125.33 ± 20.87 and 289.26 ± 34.51,and the difference was statistically significant (F =62.125,P ＜ 0.001).The 72 h proliferation activity of the four groups respectively was 1.12 ± 0.13,1.70 ± 0.14,1.07 ± 0.13 and 0.71 ± 0.11,and the difference was statistically significant (F =31.336,P ＜ 0.001).The proliferation activity of miR-182-5p mimic group was significantly higher than that of NC mimic group (P ＜ 0.05),and the proliferation activity of miR-182-5p inhibitor group was significantly lower than that of NC inhibitor group (P ＜0.05).The apoptosis rate of the four groups respectively was (5.51 t±1.80)％,(1.41 ±0.50)％,(6.24 ± 1.71)％ and (47.93 ± 5.12) ％,and the difference was statistically significant (F =211.081,P ＜ 0.001).The apoptosis rate of miR-182-5p mimic group was significantly lower than that of NC mimic group (P ＜ 0.05),and the apoptosis rate of miR-182-5p inhibitor group was significantly higher than that of NC inhibitor group (P ＜0.001).The miRNA target genes prediction software test results showed that miR-182-5p could act on FOXO3a 3' untranslated region (UTR).Compared with transfection NC mimic,co-transfection miR-182-5p mimic and FOXO3a-Wt could make luciferase activity of A549 significantly decreased (1.20 ±0.14 vs.0.62 ±0.10;t =5.839,P =0.004).Conclusion miR-182-5p can enhance proliferation and inhibit apoptosis of A549 cell by targeting FOXO3a.
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Objective The up-regulated expression of miR-182 is associated with poor prognosis of triple-negative breast cancer. This study explored the biological function of the miR-182/MTSS1 signaling pathway in three-negative breast cancer (TNBC) and the mechanism of its regulation. Methods The relative quantitative expressions of miR-182 and MTSS1 were detected in the cancerous and adjacent tissues of 30 cases of TNBC, the influence of miR-182 and MTSS1 on the proliferation and invasiveness of the cells evaluated by cell function tests, the potential binding sites of miR-182 to MTSS1 predicted with the Targetscan software, and MTSS1 confirmed to be the target gene by the dual luciferase reporter system. After transfection of miR-182 into the MCF-7 cells, RT-PCR and Western blot were used to determine the gene and protein expressions of MTSS1 and verify the regulatory effect of miR-182 targeting MTSS1. Results The expression of miR-182 was significantly higher (t=-8.409, P=0.000), while that of MTSS1 lower in the cancerous than in the adjacent tissue (t=2.961, P=0.006). The over expression of miR-182 and silenced expression of MTSS1 markedly enhanced the proliferation and migration of the MCF-7 cells compared with those of the control (P<0.01), while inhibition of miR-182 and over expression of MTSS1 remarkably suppressed their proliferation and migration of the MDA-MB-231 cells (P<0.01). The base sequences of 1083-1089 of the MTSS1 gene were confirmed to be the target binding sites of miR-182. After transfection of miR-182, the expression of MTSS1 in the MCF-7 cells was significantly down-regulated as compared with that in the control (t=-5.918, P= 0.027). Conclusion Target binding of miR-182 to MTSS1 down-regulates the expression of MTSS1 and promotes cell proliferation and migration, which may play an important biological role in the metastasis of TNBC.
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AIM:To investigate the effect of cepharanthine on the growth of human lung carcinoma A549 cells and the underlying mechanism. METHODS:After A549 cells were treated with cepharanthine,the growth inhibitory rate was detected by MTT assay. The cell morphological changes were observed under light microscope. The apoptosis of the A549 cells was analyzed by flow cytometry. The expression of microRNA (miR)-150, miR-182, p53 mRNA and FOXO1 mRNA were detected by real-time PCR. The downstream target genes were predicted by software,and the expression of p53 and FOXO1 was determined by Western blot. RESULTS:After cepharanthine treatment,the growth of A549 cells was in-hibited,the apoptosis rate was significantly increased,and the expression levels of miR-150 and miR-182 were significantly decreased. With cepharanthine treatment at 10 μmol/L,the expression levels of p53 and FOXO1 were elevated;however, with cepharanthine at 30 μmol/L,the expression levels of p53 and FOXO1 were decreased. After transfection with miR-150,the expression of p53 was significantly decreased, while the expression of FOXO1 was significantly decreased after transfection with miR-182. CONCLUSION:Cepharanthine inhibits the growth of A549 cells and promotes the apoptosis of A549 cells by inhibiting the expression of miR-150 and miR-182. miR-150 and miR-182 may down-regulate the expression of p53 and FOXO1,respectively.