| Objective:In this study,the effects of quercetin on the biological behavior of oral cancer cells were studied in vitro.Combined with the previous sequencing results,miR-181a-5p was selected as the target gene,and the function of this gene in the occurrence and development of oral cancer and the possibility of using it as a therapeutic target were analyzed to analyze the anti-tumor effects and regulatory molecular mechanisms of quercetin and miR-181a-5p on oral cancer,and to provide a new strategy for clinical cancer treatment.Methods:1.Tca-8113 and Cal-27 cells were treated with different concentrations of quercetin(Tca-8113: 0,200,250,300,350,400 μg/m L;Cal-27: 0,100,200,300,400,500 μg/m L),4 μg/m L cisplatin was used as a positive control.After cultured for 24 h,48 h and 72 h,the proliferation of oral cancer cells was detected by CCK-8 method,and the optimal concentration and action time of quercetin were screened for subsequent studies.Cell apoptosis and cycle were detected by flow cytometry.Transwell(supplemented with matrix gel)detected cell migration(invasion).The expression level of miR-181a-5p before and after cell treatment was detected by q PCR.The expression of PTEN and BCL2L11 protein in each group was detected by Western Blotting.2.Lipofectamine RNAi MAX reagent was used for transfection of miR-181a-5p mimics and Negative Control,and divided the cells into miR-181a-5p mimics transfection group,miR-181a-5p mimics transfection + quercetin group,Negative Control group,quercetin group and control group.The cell proliferation,migration and invasion ability were detected after transfection;q PCR was used to quantify the m RNA expression of Akt,FOXO1,PTEN,and BCL2L11;Western Blotting was used to determine the protein expression of Akt,p-Akt,FOXO1,p-FOXO1,PTEN,and BCL2L11.Results:1.Effects of different concentrations of quercetin on the biological behavior of oral cancer cells: Compared with the control group,the proliferation,migration and invasion ability of Tca-8113 and Cal-27 cells were significantly weakened after quercetin treatment,apoptosis increased and showed obvious cell cycle arrest;the optimal concentration was 350 μg/m L(TCA-8113)and 400 μg/m L(CAL-27),and the optimal action time was 24h;meanwhile,the expression of miR-181a-5p,PTEN and BCL2L11 increased(P<0.05).2.Effects of quercetin and miR-181a-5p on the expression of PI3K/Akt pathway related proteins: Western Blotting results showed that compared with the control group,PTEN and BCL2L11 m RNA and protein expression levels were up-regulated after quercetin and miR-181a-5p(P<0.05).The expression level of phosphorylated Akt and FOXO1 proteins decreased significantly,but there was no significant difference in Akt and FOXO1 m RNA and protein levels(P>0.05).It shows that quercetin and miR-181a-5p may exert a tumor suppressor effect by affecting the PI3K/Akt pathway.3.Effect of quercetin combined with miR-181a-5p on PI3K/Akt pathway related proteins: After cells transfected miR-181a-5p and given quercetin,PTEN and BCL2L11 m RNA and protein expression levels were up-regulated compared with the control group(P<0.05);phosphorylated Akt and FOXO1 were significantly decreased,while the m RNA and protein levels of Akt and FOXO1 were not significantly different(P>0.05),showing similar trends when quercetin and miR-181a-5P were used alone,but showed stronger effects than when they were used alone(P<0.05).It shows that up-regulation of miR-181a-5p and given quercetin have a stronger effect on PI3K/Akt pathway related proteins.Conclusions:1.Quercetin could inhibit the proliferation,migration and invasion of oral cancer cells,promote cell apoptosis and cause cycle arrest.Its anti-tumor effect may be related to the up-regulation of miR-181a-5p,PTEN and BCL2L11.2.Quercetin may play anti-tumor effect with miR-181a-5p.The mechanism may be that the two jointly promote the expression of PTEN,negatively regulate the PI3K/Akt pathway,reduce the level of FOXO1 phosphorylation,and increase the expression of BCL2L11. |
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