Apigenin Inhibits Proliferation, Migration, and Invasion of Human Tongue Carcinoma Tca8113 Cells Through Regulating the MAPK Signaling Pathways.

OBJECTIVE: This study aimed to examine the effects of apigenin (API) on the proliferation, migration, and invasion of human tongue squamous cell carcinoma Tca8113 cells and explore its probable mechanisms. METHODS: After treating Tca8113 cells with API, the cell proliferation, migration, and invasive capacities were identified by tetrazolium salt colorimetry (MTT) assay, cell scratch test, and Transwell chamber test. Cellular immunofluorescence staining was used to localize mitogen-activated protein kinase 1 (MAPK1) and extracellular regulated protein kinase (ERK) 1/2 proteins. Western blot was used to detect the variations of the related protein expression levels. RESULTS: 1)Through the MTT assay, API significantly inhibited cell proliferation (P<0.01). 2) In the cell scratch test, the distance of lateral migration after the API treatment was significantly shorter compared to the control group (P<0.01). 3) The invasion rate in the lower chamber of the Transwell chamber was lower in the API group (P<0.01). 4) Cellular immunofluorescence staining presented that the total-MEKK1 was localized in the cytoplasm, p-MEKK1 was localized in the nuclear membrane and cytoplasm, and p-ERK1/2 was localized in the cytoplasm and nucleus. ⑤ After API was applied to cells, the expressions of p-MEKK1 and p-ERK1/2 proteins significantly reduced (P<0.01). CONCLUSION: Apigenin (API) significantly inhibits the proliferation, migration, and invasion of Tca8113 cells and its mechanism may be associated with the MAPK signaling pathway.

Expression of miR-126 in oral squamous cell carcinoma tissues and its clinical significance / 中国肿瘤生物治疗杂志

@#[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.

Study on mechanism of down-regulating ikca1 molecule affecting the increment of oral squamous cell carcinoma.

This research aimed to explore the mechanism of mediating the down-regulation of the calcium-activated potassium channel (IKCa1) gene expression in human oral squamous cell carcinoma Tca-8113 cells, thereby affecting cell proliferation and apoptosis. The expression level of IKCa1 in Tca-8113 cell line (oral squamous cell carcinoma) and HOEC cell line (human normal oral epithelial cell) was detected by RT-PCR. Then, after IKCa1 was knocked down in Tca-8113 cell line and HOEC cell line by RNA interference, and then cell proliferation levels were detected by cell counting kit 8 (CCK-8) method. Cell cycle distribution was detected by flow cytometry. Apoptosis was detected by membrane linked protein V-FITC/propidium iodide (PI) double-staining apoptosis detection kit. The protein expression level of IKCa1 was detected by Western Blot method. According to RT-PRC results, IKCa1 was significantly more expressed in Tca-8113 cell line than in HOEC cell line (P< 0.01).  In addition, the mRNA expression levels in the normal oral epithelium and oral squamous cell carcinoma showed the same trend. After knocking down IKCa1 in Tca-8113 cell line, the IKCa1siRNA group significantly inhibited cell proliferation compared with the siNC control group. The results of flow cytometry showed that the proportion of apoptotic Tca-8113 cells transfected with IKCa1siRNA was significantly increased. The ratio of early apoptosis and late apoptosis of Tca-8113 cells increased (P< 0.05). To investigate the effect of IKCa1 on apoptosis, we tested the expression levels of apoptosis-related proteins. The results showed that the mRNA level of IKCa1siRNA group was significantly decreased by 44.41% compared with the control group (p< 0.01). Meanwhile, the mRNA level of Bax was significantly increased by 36.0% (p< 0.05). Our results showed that knocking down IKCa1 in Tca-8113 cells could induce cell cycle arrest and apoptosis to produce an anti-proliferation effect, thus inhibiting the expression of IKCa1 has an anti-cancer effect in oral squamous cell carcinoma.

lncRNA XIST/miR-34a-5p/SIRT6 axis regulates proliferation and metastasis of oral squamous cell carcinoma cells / 中国肿瘤生物治疗杂志

@# 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.

Effect of TET1 regulating MGMT on chemotherapy resistance of oral squamous cell carcinoma stem cells.

The study was to evaluate the effect of ten-eleven translocation 1 (TET1) regulating o6-methylguanine-DNA methyltransferase (MGMT) in chemotherapy resistance of oral squamous cell carcinoma (OSCC) stem cells. OSCC stem cells were divided into the blank, negative control (NC), TET1-siRNA, TET1-siRNA + MGMT-OE, and MGMT-OE groups. Methylation-specific polymerase chain reaction (MSP), qRT-PCR and Western blotting were conducted to detect the methylation status of MGMT, expressions of TET1, MGMT, ABCG2, and Oct-4. Cell proliferation, cisplatin chemosensitivity, and cell cycle and apoptosis, were detected using CCK8 and flow cytometry. A chromatin immunoprecipitation (ChIP) assay was employed for detecting the link between TET1 and MGMT gene promoters. In comparison to the NC group, the TET1-siRNA group exhibited increased levels of MGMT methylation, the number of apoptotic cells and cisplatin chemosensitivity consisting of varying concentrations, however, decreased levels of mRNA and protein expressions of TET1 as well as MGMT, cell viability, the number of cells in the S phase, and protein expressions of ABCG2 and Oct-4 were all have diminished amounts. The TET1-siRNA + MGMT-OE and MGMT-OE groups had higher MGMT mRNA and protein expression, as well as increased protein expressions of ABCG2 and Oct-4, greater cell activity, higher number of cells in the S phase, decreased apoptotic rates in cells and decreased cisplatin chemosensitivity with different concentrations. Our study provided evidence that low-expression of TET1 in OSCC stem cells may stimulate MGMT promoter methylation, while inhibiting MGMT mRNA expression, this ultimately strengthens the sensitivity of OSCC stem cells in regards to chemotherapeutics.

Contrastive Analysis of three transplanted tumor models in chick embryo chorioallantoic membrane / 中国比较医学杂志

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.

Genistein inhibits proliferation of human oral cancer TCA8113 cells through suppression of VEGF expression / 中国病理生理杂志

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 .

The effect of hydroxycamptothecin and pingyangmycin on human squamous cell carcinoma of the tongue.

The purpose of this study was to test hydroxycamptothecin (HCPT) and pingyangmycin (PYM) for their ability to inhibit the squamous cells of tongue carcinoma (Tca8113 cells). The effect of these compounds was tested using the MTT assay in vitro, clonogenic assays, flow cytometry, morphological observation, telomeric repeat amplification protocol (TRAP), transplantation of tumors into athymic mice and TUNEL staining. Treatment with HCPT and PYM, alone or in combination, inhibited the tumor cells and showed a greater inhibition when the drugs were combined. The cloning efficiency of Tca8113 cells was decreased. The microstructure and cell cycle of the cells changed significantly as a result of treatment. Telomerase activity was significantly inhibited in a time-dependent manner. By appearing to promote apoptosis, the drugs demonstrated a significant level of inhibition of the tumor cells in an athymic mouse model, promoting prolonged survival. HCPT and PYM have a marked cytotoxic effect on Tca8113 cells which is improved when used in combination.

The effects of sodium phenylbutyrate(SPB)on the expression of p21 and survivine genes on human tongue squamous cancer cell line Tca8113 / 中国药理学通报

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).

Inhibitary effects of hydroxycampothecin, pingyangmycin, cis-platinum or their combination on the human tongue carcinoma cells / 实用口腔医学杂志

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.

Effects of hydroxycamptothecine and pingyangmycin alone or in combination on the proliferation and telomerase activity of human tongue carcinoma cells / 实用口腔医学杂志

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

Analysis of gene expression profiles in Tca8113/CBP nude mice by microarray technique / 实用口腔医学杂志

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.

The effects of transfection of a truncated BMP-II receptor on the prolife ration of Tca8113 cells / 实用口腔医学杂志

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.

Expression of heat shock response protein induced by heat shock in human Tca8113 cells / 实用口腔医学杂志

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