OBJECTIVES: Tongue squamous cell carcinoma (TSCC) is a common cancer in the oral and maxillofacial region, which seriously endangers people's life and health.Heterogeneous nuclear ribonucleoprotein A2/B1(hnRNP A2/B1) is an RNA-binding protein that regulates the expression of a variety of genes and participates in the occurrence and development of a variety of cancers. This study aims to investigate the role of hnRNP A2/B1 in TSCC progression. METHODS: The differential expression of hnRNP A2/B1 in oral squamous cell carcinoma (OSCC) and normal oral mucosa cells and tissues was analyzed based on the gene expression profiles of GSE146483 and GSE85195 in the Gene Expression Omnibus (GEO) database. The correlation between hnRNP A2/B1 expression and disease-free survival of TSCC patients was analyzed based on TSCC related chip of GSE4676. TSCC cancer and paracancerous tissue samples of 30 patients were collected in Hunan Cancer Hospital from July to December 2021. Real-time RT-PCR and Western blotting were used to verify the mRNA and protein expression of hnRNP A2/B1 in TSCC patients'samples, respectively. Human TSCC Tca-8113 cells were transfected with hnRNP A2/B1 empty vector (a sh-NC group), knockdown plasmid (a sh-hnRNP A2/B1 group), empty vector overexpression plasmid (an OE-NC group) and overexpression plasmid (an OE-hnRNP A2/B1 group), respectively. The knockdown or overexpression efficiency of hnRNP A2/B1 was detected by Western blotting. The proliferation activity of Tca-8113 cells was detected by cell counting kit-8 (CCK-8), and the apoptosis rate of Tca-8113 cells was detected by flow cytometry. RESULTS: Based on the analysis of OSCC-related chips of GSE146483 and GSE85195 in the GEO database, it was found that hnRNP A2/B1 was differentially expressed in the OSCC and normal oral mucosa cells and tissues (all P<0.01). Meanwhile, the analysis of TSCC related chip GSE4676 confirmed that the expression of hnRNP A2/B1 was negatively correlated with the disease-free survival of TSCC patients (P=0.006). The results of real-time RT-PCR and Western blotting showed that the relative expression levels of hnRNP A2/B1 mRNA and protein in TSCC tissues were significantly up-regulated compared with those in adjacent tissues (all P<0.01). The results of Western blotting showed that the expression level of hnRNP A2/B1 in Tca-8113 cells was significantly inhibited or promoted after knockdown or overexpression of hnRNP A2/B1 (all P<0.01). The results of CCK-8 and flow cytometry showed that inhibition of hnRNP A2/B1 expression in Tca-8113 cells reduced cell proliferation activity (P<0.05) and increased cell apoptic rate (P<0.01). Overexpression of hnRNP A2/B1 in Tca-8113 cells significantly increased cell proliferation (P<0.05) and decreased cell apoptosis (P<0.01). CONCLUSIONS HnRNP A2/B1 is a key factor regulating the proliferation and apoptosis of TSCC cells. Inhibition of hnRNP A2/B1 expression can reduce the proliferation activity of TSCC cells and promote the apoptosis of TSCC cells.
Humans ; Carcinoma, Squamous Cell/genetics* ; Sincalide/metabolism* ; Tongue Neoplasms/genetics* ; Mouth Neoplasms ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism* ; RNA, Messenger ; Tongue/metabolism* ; Cell Line, Tumor

OBJECTIVETo investigate the expression of LRG-1 in clinical specimens and Tca8113 cell line of tongue carcinoma and analyze the relationship between LRG-1 expression and the clinicopathological parameters.

METHODSLRG-1 expression was detected in 40 tongue squamous cell carcinoma (TSCC) tissues and paired normal adjacent tissues, 20 atypical hyperplasia tissues of the tongue, and 20 tissues of tongue cancer in situ using immunohistochemical method. The expression of LRG-1 in Tca8113 cell line was detected using flow cytometry. The expression of LRG-1 was also detected in human TSCC tissues and Tca8113 cells with Western blotting. The effect of LRG-1 on the proliferation of HUVECs was determined using MTT assay, and its effect on angiogenesis was evaluated with Matrigel tube formation assays.

RESULTSHuman TSCC tissues had a significantly higher rate of positive expression for LRG-1 (85%, 34/40) than the adjacent tissues (10%, 4/40), invasive tongue cancer (30%, 6/20), and tongue cancer in situ (50%, 10/20) (P<0.05). LRG-1 expression was correlated with the degree of tumor differentiation, clinical stage and lymph node metastasis of the tumor (P<0.05) but not with the patients' age or gender. In the in vitro experiment, LRG-1 promoted HUVEC proliferation and angiogenesis.

CONCLUSIONAbnormal LRG-1 expression is present in the human TSCC tissue and Tca8113 cells. LRG-1 can promote HUVEC proliferation and angiogenesis in vitro, suggesting its possible role in promoting tumor angiogenesis.

Carcinoma, Squamous Cell ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Glycoproteins ; genetics ; metabolism ; Human Umbilical Vein Endothelial Cells ; Humans ; Lymphatic Metastasis ; Tongue ; metabolism ; pathology ; Tongue Neoplasms ; genetics ; metabolism

OBJECTIVETo study the effect of kallikrein-related peptidase 10 (KLK10) on the proliferation and invasiveness of human tongue cancer cell line Tca8113.

METHODSThe eukaryotic expression vector harboring KLK10 gene (pIRES2-EGFP-KLK10) was transfected in Tca8113 cells and the stable cell lines were selected by G418 screening. The mRNA and protein expression of KLK10 in transfected Tca8113 cells were assayed by RT-PCR and Western blotting, respectively, and the proliferation and invasiveness of the cells were evaluated by MTS cell growth assay and Transwell chamber invasion experiments.

RESULTSA stable Tca8113 cell line with high KLK10 expression was obtained, which showed significantly increased mRNA and protein expression levels of KLK10 and obviously attenuated proliferation and invasiveness compared with control and empty vector-transfected cells (P<0.05).

CONCLUSIONEnhancing KLK10 gene expression can decrease the proliferation and invasiveness of human tongue cancer cells in vitro.

Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Humans ; Kallikreins ; genetics ; Tongue Neoplasms ; genetics ; pathology

OBJECTIVES: This study aims to investigate the effect of RhoE expression on the migration and invasion of tongue squamous cell carcinoma (TSCC). METHODS: Forty-eight TSCC cases were selected from the Maxillofacial Surgery Center of Qingdao Municipal Hospital from 2017 to 2019. The expression of RhoE in the specimens (TSCC and adjacent tissues) was detected by immunohistochemistry, and RhoE mRNA and protein were extracted to further detect the expression of RhoE. SCC-4 and CAL-27 cells were selected for RESULTS: The expression level of RhoE in TSCC was significantly lower than that in adjacent tissues ( CONCLUSIONS RhoE expression is low in TSCC. Over expression RhoE in TSCC can significantly decrease its migration and invasion abilities. Hence, RhoE may play an important role in regulating the metastasis and invasion of TSCC and provide a new target for gene therapy.
Carcinoma, Squamous Cell ; Cell Line, Tumor ; Humans ; Matrix Metalloproteinase 2 ; Neoplasm Invasiveness ; Tongue ; Tongue Neoplasms ; rho GTP-Binding Proteins/genetics* ; rho-Associated Kinases

OBJECTIVETo investigate the inhibition of telomerase activity and cellular proliferation in tongue cancer TCCA-8113 cell lines by antisense human tankyrase-1 RNA treatment, and explore the possibility of the tankyrase-1 as a target of gene therapy for tongue cancer.

METHODSThe replication deficient retrovirus expressing tankyrase-1 antisense RNA was constructed to infect the TCCA-8113 cells. Tankyrase-1 expression was examined by RT-PCR. Telomerase activity was assayed by telomerase repeat amplification protocol (TRAP). Cell proliferation was investigated by cellular growth curve. Cellular apoptosis was detected by flow cytometry method and invert microscope.

RESULTSTankyrase-1 expression and telomerase activity of tongue cancer TCCA-8113 cells were significantly inhibited. There was G(1)-S phase arrest when TCCA-8113 cells were treated with antisense tankyrase-1 transduction. Cellular proliferation was arrested, and cellular apoptosis occurred after antisense tankyrase transduction.

CONCLUSIONSThe transduction of antisense tankyrase-1 by retroviral vector can significantly inhibit the tankyrase-1 expression and telomerase activity of tongue cancer TCCA-8113 cell lines, and arrest the cellular proliferation and promote cellular apoptosis. The tankyrase may be a potential target of gene therapy for tongue cancer.

Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Genetic Therapy ; Genetic Vectors ; genetics ; Humans ; RNA, Antisense ; genetics ; Retroviridae ; genetics ; Tankyrases ; genetics ; Telomerase ; metabolism ; Tongue Neoplasms ; genetics ; metabolism ; pathology ; therapy ; Transfection

OBJECTIVES: A study was conducted to investigate the molecular mechanism of chromodomain helicase/ATPase DNA binding protein 1-like gene (CHD1L) influencing the invasion and metastasis of tongue squamous cell carcinoma and to provide a new target for clinical inhibition of invasion and metastasis of tongue squamous cell carcinoma. METHODS: Ualcan website was used to analyze the expression of CHD1L in normal epithelial tissue and primary head and neck squamous cell carcinoma and to analyze the effect of lymph node metastasis on the expression of CHD1L in tissues with head and neck squamous cell carcinoma. The relationship between CHD1L expression and the survival rate of patients with head and neck squamous cell carcinoma was tested by the GEPIA website. Western blot was used to quantify the levels of CHD1L protein in human tongue squamous cell carcinoma CAL27 and immortalized human skin keratinocyte cell HaCaT. After knocking down CAL27 in human tongue squamous cell carcinoma cells with an RNA interference plasmid, the cells were designated as SiCHD1L/CAL27 and Scr/CAL27. Western blot was utilized to detect the expression of CHD1L in each group of cells. The change in CAL27 cell proliferation ability was tested by EdU proliferation test after CHD1L knockdown. The change of cell migration ability of each group cells was tested through the wound healing assay. Western blot was used to detect epithelial-mesenchymal transition (EMT) marker E-cadherin and Vimentin protein expression levels. RESULTS: Ualcan database showed that the expression of CHD1L in primary head and neck squamous cell carcinoma tissues was higher than in normal epithelial tissues and in head and neck squamous cell carcinoma tissues with lymph node metastasis. GEPIA website analysis showed that the overall survival rate of patients with head and neck squamous cell carcinoma with high expression of CHD1L was significantly lower than that of patients with low expression. Western blot results showed that CHD1L expression in human tongue squamous carcinoma cells CAL27 was higher than that of human normal skin cells HaCaT. CHD1L expression in SiCHD1L/CAL27 cells was much lower than that in Scr/CAL27 cells. Results of EdU proliferation experiments showed the significant reduction in the cell proliferation ability of the SiCHD1L/CAL27 cells. Results of the wound healing experiments showed the reduction in the migration capacity of the SiCHD1L/CAL27 cells. The expression of E-cadherin increased, whereas that of Vimentin decreased, in SiCHD1L/CAL27 cells. CONCLUSIONS CHD1L promoted the EMT, proliferation, migration, and invasion ability of tongue squamous cell carcinoma cells.
Adenosine Triphosphatases ; Carcinoma, Squamous Cell/genetics* ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; DNA Helicases ; DNA-Binding Proteins ; Epithelial-Mesenchymal Transition ; Gene Expression Regulation, Neoplastic ; Head and Neck Neoplasms ; Humans ; Neoplasm Invasiveness/genetics* ; Tongue ; Tongue Neoplasms/genetics*

OBJECTIVEA functional VEGF-C cDNA was cloned from a patient with squamous cell carcinoma (SCC) of tongue in order to study the important role of vascular endothelial growth factor (VEGF)-C in lymphatic dissemination of malignancies in gene level.

METHODSRT-PCR was employed to clone the human VECF-C encoded cDNA from a surgical specimen of a lingual SCC patient. Then it was subcloned into plasmid vector pCRII and sequenced.

RESULTSA 1.1 kb human VEGF-C cDNA fragment was amplified from the lingual SCC. The sequencing results of the fragment demonstrated that it had 99.6% similarity with the reported human VEGF-C cDNA (representing the 559-1,611 bp according the sequence of Genbank Entry X94216).

CONCLUSIONAn encoded fragment VEGF-C cDNA was successfully cloned from a lingual SCC and provided a necessary material for further study.

Carcinoma, Squamous Cell ; genetics ; Cloning, Molecular ; DNA, Complementary ; genetics ; Humans ; Sequence Analysis, DNA ; Tongue Neoplasms ; genetics ; Vascular Endothelial Growth Factor C ; genetics

OBJECTIVETo determine whether the human telomerase reverse transcriptase (hTERT) gene silencing could be effectively induced by PCR-derived siRNA expression cassettes (SEC) transfected by the fifth generation polyamidoamine dendrimer (G5 PAMAM-D) in Tca8113 cells.

METHODSFour SEC were rationally designed and constructed based on a two-step PCR reaction. The SEC were then transferred into Tca8113 cells using G5 PAMAM-D, and hTERT expression was investigated by real-time fluorescence-quantitative reverse transcriptase-PCR and western blot analysis.

RESULTSThe RNA interference effects of the SEC targeted for varying hTERT mRNA positions showed a significant disparity. Among them, SEC-A revealed the most potent inhibitory effects (above 95% of reduction), followed by SEC-D and SEC-C, and SEC-B had no effect on hTERT expression (P > 0.05). That the endogenous hTERT gene silencing induced by G5 PAMAM dendrimer-mediated SEC-A was highly sequence-specific, and multiple transfection as well as properties of the vectors were routinely attributable to the specific suppression.

CONCLUSIONSSpecific inhibition of endogenous hTERT expression by use of a PCR-based short hairpin siRNA technique and dendrimer transfer system may serve as a novel strategy for treatment of tongue cancers expressing hTERT in vitro.

Carcinoma, Squamous Cell ; enzymology ; genetics ; Cell Line, Tumor ; Gene Expression ; Genetic Vectors ; Humans ; RNA, Small Interfering ; genetics ; Telomerase ; genetics ; Tongue Neoplasms ; enzymology ; genetics ; Transfection

OBJECTIVEThe aim of this study was to investigate suppression effects of the transfection of human tumor necrosis factor-alpha (hTNF-alpha) gene on tongue carcinoma cells.

METHODSThe shuttle plasmid containing hTNF-alpha gene was extracted and purified, then it was transferred into Tca8113 tongue carcinoma cells with cationic liposome DOSPER. The control group was only given equivalent liposomes, except the plasmid. After culturing for 24, 48, 72 and 96 hours, the expression of hTNF-alpha gene in Tca8113 cells was analysed by ELISA and, the survival rate of transferred cells was assayed by MTT enzymatic labeling technique.

RESULTSThe transferred Tca8113 cells displayed significantly overexpression of hTNF-alpha (P < 0.05). The survival rate of the transferred Tca8113 cells was decreased significantly (P < 0.05).

CONCLUSIONTransfection of hTNF-alpha gene in vitro mediated by cationic liposomes can induce the overexpression of hTNF-alpha and inhibit the growth of tongue carcinoma cells.

Carcinoma, Squamous Cell ; genetics ; pathology ; Cell Division ; Gene Transfer Techniques ; Genetic Vectors ; Humans ; Plasmids ; genetics ; Tongue Neoplasms ; genetics ; pathology ; Transfection ; Tumor Cells, Cultured ; Tumor Necrosis Factor-alpha ; genetics

BACKGROUND: The association between miR-532-3p and tongue squamous cell carcinoma (TSCC) has been examined in the literature to improve the survival rate of patients with this tumor. However, further studies are needed to confirm the regulatory roles of this microRNA (miRNA) in TSCC. The objective of this study was to investigate the roles played by and the underlying mechanism used by the miR-532-3p/podoplanin (PDPN) axis in TSCC development. METHODS: Western blotting and quantitative real-time reverse transcription-polymerase chain reaction (RT-qPCR) were performed to evaluate the PDPN expression level in TSCC tissues and cells. The proliferative, adhesive, and migratory capabilities of TSCC cells (CAL-27 and CTSC-3) were examined using cell counting kit-8 (CCK-8), cell adhesion, and wound-healing assays, respectively. The dual-luciferase reporter (DLR) assay was later conducted to confirm the relationship between miR-532-3p and PDPN. RESULTS: The results indicated that PDPN expression was enriched in TSCC tissues and cells, and that the expression of PDPN was associated with some clinicopathological parameters of TSCC, including lymph node metastasis (P = 0.001), tumor-node-metastasis (TNM) staging (P = 0.010), and grading (P = 0.010). Further analysis also showed that PDPN knockdown inhibited the viability, adhesive ability, and migratory capacity of CAL-27 and CTSC-3 cells, effects that could be reversed by the application of a miR-532-3p inhibitor. Additionally, PDPN was found to be a direct target of miR-532-3p. CONCLUSIONS This research suggested that by targeting PDPN, miR-532-3p could inhibit cell proliferation viability, adhesion, and migration in TSCC. Findings also revealed that the miR-532-3p/PDPN axis might provide more insights into the prognosis and treatment of TSCC.
Carcinoma, Squamous Cell/genetics* ; Cell Line, Tumor ; Cell Proliferation/genetics* ; Gene Expression Regulation, Neoplastic ; Humans ; Membrane Glycoproteins ; MicroRNAs/genetics* ; Tongue Neoplasms/genetics*