ABSTRACT
BACKGROUND: This study sought to study the expression of the long non-coding RNA (lncRNA) taurine-upregulated gene 1 (TUG1) in tongue squamous cell carcinoma (TSCC) and reveal its possible function. METHODS: qRT-PCR was used to evaluate 27 samples of fresh TSCC tissues and adjacent normal tongue tissues. siRNA technology was employed to downregulate TUG1 expression in CAL-27 and SCC-9 cell lines. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was utilized to assess cell proliferation ability; apoptosis and cell-cycle phases were analysed via flow cytometry. RESULTS: qRT-PCR findings indicated that the lncRNA TUG1 was upregulated in TSCC tissues compared with adjacent normal tongue tissues (P<.05). After TUG1 expression was downregulated using siRNA technology, cell proliferation was significantly inhibited (P<.05), and the number of cells in S phase was reduced (P<.05). CONCLUSION: The lncRNA TUG1 may represent a potential oncogene in TSCC.
Subject(s)
Carcinoma, Squamous Cell/genetics , Gene Expression Regulation, Neoplastic , Head and Neck Neoplasms/genetics , RNA, Long Noncoding/genetics , Tongue Neoplasms/genetics , Humans , Squamous Cell Carcinoma of Head and Neck , Tissue Culture Techniques , Up-RegulationABSTRACT
OBJECTIVE: To construct an eukaryotic coexpression vector containing Mycobacterium tuberculosis heat shock protein 70 (mtHSP70) and enhanced green fluorescent protein (EGFP) controlled by cytomegalovirus promoter using pIRES-EGFP vector. METHODS: The mtHSP70 gene fragment was amplified by PCR from pVAX-mtHSP70-HSV2gD using specific primers. The PCR product was cloned into the vector pMD 18-T vector, and the correct clone was selected according to DNA sequence analysis. The interested mtHSP70 gene fragment was subcloned into pCMV-IRES-EGFP vector with XhoI and EcoR I digestion. The recombinant plasmid was transfected into mouse melanoma B16 cell line, and the green fluorescent cells were detected by fluorescence microscopy and mtHSP70 expression was detected by Western blotting. RESULTS: The recombinant plasmid obtained was confirmed by enzyme digestion. The transfected mouse melanoma B16 cells exhibited green fluorescence under fluorescence microscopy and expressed mtHSP70 protein as demonstrated by Western blotting. CONCLUSION: The eukaryotic coexpression vector PCMV-mtHSP70-IRES-EGFP has been established to allow further investigation of the role of mtHSP70 gene in tumor immunotherapy.
Subject(s)
Genetic Vectors/genetics , Green Fluorescent Proteins/biosynthesis , HSP70 Heat-Shock Proteins/biosynthesis , Mycobacterium tuberculosis/metabolism , Animals , Base Sequence , Cancer Vaccines , Cell Line, Tumor , Cytomegalovirus/genetics , Cytomegalovirus/metabolism , Genetic Vectors/biosynthesis , Green Fluorescent Proteins/genetics , HSP70 Heat-Shock Proteins/genetics , Mice , Molecular Sequence Data , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/genetics , Sequence Analysis, DNAABSTRACT
OBJECTIVE: To investigate the effect of inducible nitric oxide synthase (iNOS) on vascular endothelial growth factor (VEGF) expression. METHODS: The vector containing short hairpin RNA of iNOS was transfected into Tca8113 cells using the RNA interference (RNAi) technique. The gene and protein expression of iNOS and VEGF was examined by RT-PCR and Western blot. RESULTS: The expression of iNOS, VEGF gene in Tca8113 cells was significantly different between the experimental and control groups 24 h and 48 h after transfection (P < 0.05). The protein expression of iNOS was different between the two groups 36 h and 48 h after transfection, and of VEGF was also different between the two groups 48 h after transfection (P < 0.05). CONCLUSIONS: The expression of VEGF could be down-regulated by silencing the iNOS gene in Tca8113 cells.