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@#[Abstract] Objective: To detect the expression of miR-126 in oral squamous cell carcinoma (OSCC) and to analyze its correlation with clinicopathological features and prognosis of patients, as well as to explore the effect of miR-126 over-expression on the malignant biological behaviors of Tca8113 cells. Methods: A total of 62 pairs of cancer and para-cancerous tissue specimens from OSCC patients who were surgically treated in the First Affiliated Hospital of Zhengzhou University from June 2016 to June 2018 were collected for this study; in addition, human tongue squamous carcinoma Tca8113 cell line and human mouth keratinocyte HOK cell line were also selected for this study. The expression of miR-126 in cancer tissues and cells was detected by qPCR, and the relationship between miR-126 expression and clinicopathological features and prognosis of the patients was analyzed. miR-126 mimics and miR-NC plasmids were respectively transfected into Tca8113 cells by liposome transfection technology. Cell proliferation, apoptosis, migration and invasion were detected by MTT method, Flow cytometry and Transwell chamber method, respectively; and the expressions of apoptosis, migration and invasion related proteins were detected by Western blotting. Results: The expression level of miR-126 in OSCC tissues and Tca8113 cells was significantly lower than that in para-cancerous tissues and HOK cells (all P<0.01). The expression of miR-126 was associated with TNM stage and lymph node metastasis (all P<0.05), and patients with high miR-126 expression had significantly better overall survival rate than patients with low expression (P<0.05). After transfection with miR-126 mimics, the cell proliferation, migration and invasion ability significantly decreased (P<0.05 or P<0.01) while the apoptosis rate significantly increased in Tca8113 cells (P<0.01), the expression levels of Bcl-2, N-cadherin and vimentin in Tca8113 cells significantly decreased (all P<0.01), and expression levels of Bax and E-cadherin significantly increased (all P<0.01). Conclusion: miR-126 is low expressed in OSCC tissues and Tca8113 cells. Up-regulation of miR-126 inhibits cell proliferation, migration and invasion and promotes apoptosis of Tca8113 cells.
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@# Objective: To investigate the molecular mechanisms of lncRNA XIST/miR-34a-5p/SIRT6 axis regulating the proliferation and metastasis of oral squamous cell carcinoma (OSCC) cells. Methods: Specimens of tumor tissues and paracancer tissues of 47 patients with OSCC, who visited the Qingdao Stomatological Hospital from March 2013 to March 2018, were enrolled in this study. qPCR was performed to measure the mRNA expressions of lncRNA XIST, miR-34a-5p and SIRT6 in OSCC tissues and cell lines. WB was performed to measure the protein levels of SIRT6, Ki67, pcDNA, cleaved-caspase3, cleaved-caspase8, E-cadherin and vimentin in OSCC tissues and cell lines. CCK-8 assay was performed to observe the effect of lncRNA XIST knockdown on proliferation of Cal-27 and Tca-8113 cells; Tanswell assay was performed to detect migration and invasion of Cal-27 and Tca-8113 cells; flow cytometry was performed to detect the apoptosis of Cal-27 and Tca-8113 cells; and dual luciferase reporter assay was performed to verify the relationship between lncRNAXIST and miR-34a, or miR-34a and SIRT6. Results: Expressions of lncRNAXIST and SIRT6 were up-regulated in OSCC tissues and cell lines (all P<0.05), reversely, miR-34a-5p was down-regulated in OSCC tissues and cell lines (P<0.01). lncRNA XIST knockdown significantly suppressed OSCC cells proliferation, migration and invasion, and induced apoptosis of OSCC cells (all P<0.01); however, simultaneous transfection with miR-34a-5p inhibitor or pcDNA-SIRT6 vector exerted opposite effect. lncRNA XIST knockdown significantly increased cell proliferation and metastasis related protein expression in OSCC cells (all P< 0.01), and simultaneous transfection with miR-34a-5p inhibitor or pcDNA-SIRT6 vector exerted opposite effect. In addition, lncRNA XIST directly targeted miR-34a-5p, and miR-34a-5p targeted SIRT6, lncRNA XIST upregulated SIRT6 expression via inhibiting miR34a-5p (P<0.01). Conclusion: Taken together, lncRNA XIST/miR-34a-5p/SIRT6 axis regulates the proliferation and metastasis of OSCC cells and provides potential therapeutic targets for OSCC.
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Objective To compare the growth of three different cancer cell lines on chick chorioallantoic membrane (CAM), to select the best transplanted cancer cell line for establishing a transplanted tumor model and to observe the biological characteristics.Methods The human lung cancer cell line A549, human tongue cancer cell line TCA8113 and human liver cancer cell line QGY7703 were respectively inoculated into CAM at the 7th day of age.The chick embryo survival rate, tumor survival rate, tumor formation rate and induced angiogenesis were detected and the growth characteristics of the transplanted tumor model were observed.Results Compared with the groups inoculated with A549 cells and QGY7703 cells, the tumor formation rate of TCA8113 cells was the highest (P < 0.05), to be the best cancer cell line for transplanted tumor.The optimal inoculated number of cells was 8.0×106/chick embryo, the optimal growth period of the tumor was 4~8 d, and the best experiment time was 7 d after inoculation.Conclusion The TCA-CAM transplanted tumor model of tongue squamous cell cancer is successfully established for further study of the biological characteristics and mechanisms of tumor growth, angiogenesis, invasion and metastasis, and provide a good experimental animal model for anti-tumor drug screening.
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AIM:To investigate the effect of genistein on the proliferation of human oral cancer TCA 8113 cells and to explore the underlying mechanisms .METHODS:The cell proliferation was examined by MTT assay , cell counting and colony formation assay .Western blotting was employed to examine the protein levels of vascular endothelial growth fac -tor (VEGF), extracellular signal-regulated kinase (ERK) and p-ERK.RESULTS: Genistein significantly inhibited the proliferation of TCA8113 cells in a concentration-dependent fashion .Moreover , genistein dose-dependently decreased the protein levels of VEGF, ERK and p-ERK.The expression of VEGF was also blunted by U 0126, a specific inhibitor of ERK.U0126 and axitinib, a VEGF receptor antagonist , both significantly inhibited the proliferation of TCA 8113 cells. CONCLUSION:Genistein inhibits the proliferation of TCA8113 cells, which may be related to its inhibitory effect on ERK expression and activation , thus subsequently decreasing the expression of VEGF .
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Aim To investigate the effects of sodium phenylbutyrate on cell proliferation,apoptosis and its expression of p21 and survivin genes in human tongue squamous cancer Tca8113 cell line.Methods The cellular proliferation inhibitory ratio was evaluated by MTT assay and the apoptosis and cell cycle of the Tca8113 cell line was detected by FCM.The expression of p21 and survivin genes was analyzed with Western blot and RT-PCR.Results Sodium phenylbutyrate could inhibit the Tca8113 cells proliferation,promote cell apoptosis and arrest the cells at G_1/G_0 phase.The expression of p21 gene in Tca8113 cell line treated by sodium phenylbutyrate was increased,and one of survivin gene was decreased.Conclusions Sodium phenylbutyrate induces up-regulation of p21 gene and down-regulation of survivin gene,which inhibits Tca8113 cell proliferation and induces its apoptosis and arrests the cells at G_1/G_0 phase.And the increase of p21 mRNA expression is negatively correlated with the decrease of survivin mRNA expression(r_s=-0.548,P<0.01),and so is its protein expression(r_s=-0.514,P<0.01).
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Objective:To study the effects of hydroxycamptothecine(HC PT ) and pingyangmycin(PYM) alone or in combination(HCPT/PYM) on the proliferation and telomerase activity of human tongue carcinoma Tca 8113 cells.Methods :The growth inhibitary effects of HCPT or PYM on Tca 8113 cells were stu died by MTT assay and the IC 30 values of the two drugs were obtained. Then the cells were exposed to HCPT or PYM at IC 30 , or to their combination. T he clonogenecity, cell cycle distribution, morphological change and telomerase a ctivity were studied by clonogenic assay, flow cytometry,transmision electron mi croscopy and telomeric repeat amplification protocol (TRAP) respectively.Results:IC 30 (ng/ml) of HCPT and PYM was 109 and 416 respectivel y, when in combination(HCPT/PYM) the IC 30 of HCPT and PYM was 35 and 100 r espectively. The clonogenecity(%) of the control, HCPT,PYM and HCPT/PYM treated cells was 31.7,11.6,15.2 and 4.1 respectively. PYM decreased S and G 2 phase ce lls,increased G 1 phase cells.HCPT or HCPT/PYM decreased G 1 phase cells and i ncreased S and G 2 phase cells.Drug treatment resulted in cell organ degenerati on and decrese of telomerase activity.The A value at A 450 of telomerase in control,HCPT,PYM and HCPT/PYM treated cells was 1.89?0.03,0.82?0.02,0.77?0.0 2 and 0.53?0.02 respectively(P
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objective: To study the effects of hydroxycampothecin(HCPT), pingyangmycin(PYM), cis platinum (DDP) or their combination on the proliferation of human tongue carcinoma Tca8113 cells. Methods: MTT assay, cell counting,clonogenic assay and HE staining were used to study the effects of HCPT, PYM, DDP or their combination on the proliferation of human tongue carcinoma Tca8113 cells. Results: ①Synergistic inhibiton was observed when HCPT at the concentration (ng/ml) of 10～100 combined with PYM at 32～320 or DDP at 32～320, antagonistic effect was observed when PYM at 32～320 combined with DDP at 32～320; ②After treatment with PYM, HCPT or PYM plus HCPT at IC 30 the clonogenecity (%) of the cells was 15.2, 11.6 and 4.1 respectively, while that of untreated cells 31.7. Conclusion: The combination of HCPT with PYM may be synergistic in the inhibition of tongue carcinoma cell proliferation.
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Objective: To study the effects of BMPs signals on the proliferation of tongue cancer Tca8113 cells. Methods: Th e cDNA of truncated BMP-II receptor was transfected into Tca8113 cells by usin g FuGENE6 transfection kit, the transfected cells were named Tca8113ZR. The pro liferation and DNA synthesis of Tca8113 and Tca8113ZR cells were investigated b y MTT assay,FCM and BrdU analysis. Results: In MTT assay the A value of Tca8113 and Tca8113ZR cells was 0.47?0.01 and 0.35?0.01 (P0.05).Conclus ion: BMPs might be involved in the development of squamous cell carc inoma of tongue.
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Objective:To study the differences of gene expression between Tca8113 and Tca8113/CBP tissues in nude mice. Methods:Tca8113 cells were injected subcutaneously in both sides of armpits of nude mice at the concentration of 5?106 cells/0.1 ml. Two weeks after injection, Carboplatin was used subcutaneously around the tumor 0.01 mg/g (weight) each day in Tca8113/CBP group while Tca8113 group was injected with physiologic saline as control. Mice were sacrificed 10 weeks after drug injection. The two kinds of tissues were investigated by human 16k cDNA v2.1 SBC-R-HC-100-21 gene chip. Results:Among the 16 000 target genes, there were 719 genes whose expression levels showed differences between the two kinds of tumor tissues. Conclusion:Microarray technique can simultaneously screen different genes from above-mentioned two kinds of tissues. Further analysis of the obtained genes will be helpful to understand the molecular mechanism of multidrug resistance.
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Objective: To investigate the expression of heat shock response protein(HSP_ 70 ) in human squamous tongue cancer Tca8113 cells during the recovery periods of heat shock.Methods:Tca8113 cells were subjected to heat shock at 43 ℃ for 30 min, then the cells were cultured for 2,4,6,8,12,24 and 48 h respectively. The expression of HSP_ 70 in the cells was examined with immunohistochemical method, Quantitative analysis was performed by FCM and the cell vitality was detected by MTT method.Results:Heat shock induced HSP_ 70 expression in Tca8113 cells at 43 ℃ for 30 min and the maximum proportion of the positive cells were observed and HSP_ 70 reached the maximum value at 12 h after heat shock(P