The Effects And Mechanism Of Silent Circadian Gene PER2 On Cell Biological Behavior Of Human Oral Squamous Cell Carcinoma SCC15 Cells

Object: To investigate the effects and mechanism of circadian gene PER2 on cell proliferation,apoptosis,metastasis and invasion of human oral squamous cell carcinoma SCC15 cells.Methods: RNA interference technology was used to silence PER2 gene in SCC15 cells.Three sequences of the sh RNA(short hairpin RNA)to effectively silence the PER2 gene were inserted into the lentiviral vectors.Human oral squamous cell carcinoma(OSCC)SCC15 cells were infected by the packaged shRNA lentiviral vector excluded PER2 sequences were acted as the control group,and cells with no treatment were acted as the blank group.Real-time fluorescent quantitative PCR was used to determine changes of mRNA expression of Ki-67,murine double minute2(MDM2),P53,Bcl-2,Bax,c-Myc,metalloproteinase 2(MMP-2),tissue inhibitor of metalloproteinase 2(Timp-2)and vascular endothlial growth factor(VEGF).The CCK-8 assays was used to evaluate SCC15 cells growth in vitro,colony formation assay was performed to explore cellclone formation rate,flow cytometry was performed to detect cell proliferation、apoptosis and cell cycle distribution after silence PER2 gene,and transwell assay was used to determine changes of ability of cell migration and invasion,tumorigenesis in vivo assay was performed to detect cell growth in vivo.Result: Three groups of PER2-shRNA lentiviral vectors were constructed successfully,and Real time RT-PCR and Western blot demonstrsted that the PER2-shRNA-I group had the most effective silenteffects(experimental group).PER2 downregulation increased the mRNA expression level of Ki-67,MDM2,Bcl-2,c-Myc and MMP-2,and decreased the mRNA expression level of p53、Bax and Timp-2(P<0.05 for all).After silence PER2 gene in SCC15 cells,the cell apoptosis index(AI)was significantly reduced(P<0.05),and cell proliferation index(PI),cell growth,cell migration and invasion ability were significantly increased(P<0.05 for all).The in vivo experiments also proved that the tumorigenicity of SCC15 cells was significantly enhanced after PER2silence(P<0.05).Conclusion: Clock gene PER2 through the regulation of the numerous important downstream tumor-related genes,thus playing a major role in tumor suppression,and it may be a novel molecular target for cancer treatment.