Expression Of E-cadherin In Laryngeal Squamous Cell Carcinoma And Its Relationship With Tumor Invasion And Metastases

Objective:Laryngeal squamous cell carcinoma (LSCC) is one of the commonest tumors in the head and neck, locoregional recurrence, cervical lymph nodes metastases and distant metastases are the factors that significantly affect the prognosis in LSCC patients. An important step in the development of tumor metastasis is the detachment of malignant cells from their original site, which results from the reduced cell-cell adhesiveness. The aim of this study was to investigate the clinical significance of E-cadherin expression in laryngeal squamous cell carcinoma.Methods:The level of E-cadherin expression in 64 tumor tissues and 30 adjacent non-tumor laryngeal tissues were determined by immunohistochemistry. Patients were classified according to age, primary site, T stage, lymph node metastases and histological differentiation. Staining scores were averaged for each group, and mean scores were compared within groups stratified with respect to clinicopathological parameters by one-way ANOVA test. Survival curves were calculated using the Kaplan-Meier method. Differences in 5-year survival rate were analysed by the log-rank test. Multivariate Cox proportional hazards models were used to examine the relative impact of either variable on the disease prognosis.Results:1. The mean staining score for sixty-four carcinomas was 169±68, and the thirty non-tumor laryngeal tissues had a mean staining score of 346±38, the mean score in tumor tissues was significantly lower than the non-tumor laryngeal tissues(P<0.001);2. The expression of E-cadherin were significantly correlated with lymph node metastases (P＜0.001). The differences of E-cadherin expression between the different age (p=0.66), primary site (P=0.445), T stage (P=0.303) and histological differentiation (P=0.159) group were not statistically significant;3. In addition to other known factors such as T-stage and histological grade, E-cadherin staining score was also the statistically significant, independent predictors of lymph node metastases;4. We classified patients as E-cadherin low-expression group and E-cadherin high-expression group according to the mean staining score. The 5-year survival rate in E-cadherin high-expression group were significantly increased than that of E-cadherin low-expression group (Log-rank, P<0.05);5. The results of multivariate Cox proportional hazards model confirmed that only the presence of cervical lymph node metastases was a statistically significant, independent predictor of prognosis (P=0.002). Other parameters, such as primary sit, T-stage, histological grade, E-cadherin expression, cannot predicate disease prognosis separately.Conclusion:1. The expression of E-cadherin is decreased in the tumor tissues, which indicated that E-cadherin may have been involved in the tumor occurrence and development.2. Expression of E-cadherin is an independent predictor of lymph node metastases in laryngeal cancer, it might be a tumor marker for occult lymph node metastases.3. Decreased E-cadherin expression is associated with recurrence and decreased disease survival rate in laryngeal squamous cell carcinoma. Objective:To investigate the impact of epidermal growth factor receptor (EGFR) ligand EGF and small molecule tyrosine kinase inhibitor Erotinib on the proliferation and cell cycle in human laryngeal cancer cell line Hep-2.Methods:Human laryngeal cancer cell line Hep-2 was treated with EGF and Erlotinib, respectively. Then, cell proliferation in different groups was measured by MTT assay. The influence of EGF and Erlotinib on the cell cycle and apoptosis were analysed by Flow Cytometer.Results:1. MTT assay:EGF promoted proliferation of Hep-2 cells as the extension of treatment concentration and time; Erlotinib inhibited proliferation of Hep-2 cells, which induced the decreases in cell number, density and survival rate.2. Flow Cytometer:Hep-2 cells treated with EGF exhibited the decreases in phase G1 and increases in phase S (P<0.05), while the cell apoptosis was not influenced; Treatment of Erlotinib resulted in the increases in phase G1 and decreases in phase S (P＜0.05), and cell apoptosis rate was significantly increased than the control group (P＜0.001).Conclusion:1.EGF accelerates the cell cycle transition from G1 phase to S phase, and promotes Hep-2 cell proliferation.2. Treatment of Erlotinib results in the G1 phase arrest of cell cycle, and induces Hep-2 cell apoptosis, ultimately inhibits Hep-2 cell proliferation. Objective:To investigate the impact of epidermal growth factor receptor (EGFR) ligand EGF and small molecule tyrosine kinase inhibitor Erotinib on the phenotype, cell motility, invasion and metastases in human laryngeal cancer cell line Hep-2.Methods:Human laryngeal cancer cell line Hep-2 was treated with 10ng/ml EGF and 20μmol/L Erlotinib, respectively. The cell phenotype changes were observed using inverted microscope. Changes in the localization and expression of E-cadherin, vimentin were examined by immunocytochemistry and Western blot. Wound healing assay and Transwell invasion assay were used to detect cell motility, invasion and metastases, respectively.Results:1. In the presence of EGF treatment, Hep-2 cells changed its shape to fusiform fibroblastoid phenotype and its colony formation from compact to sparse, while Erlotinib induced the cell morphological changes to round or square, cell grew into cluster;2. Immunocytochemistry:EGF resulted in the down-regulation of E-cadherin protein, especially in the cell membrane; while Erlotinib could up-regulate the E-cadherin expression, in some cell there were recurrent positive membrane expression;3. Western blot:EGF induced down-regulation of E-cadherin expression, and up-regulation of vimentin, p-EGFR in Hep-2 cells; Erlotinib could lead to up-regulation of E-cadherin, down-regulation of vimentin and p-EGFR. Both the two interference had less impact on EGFR expression;4. Wound healing assay:EGF promoted the wound healing, resulted in the increased cell motility and movement; Erlotinib delayed the wound healing, and partly inhibited cell motility and movement. The 24h distance of cell migration in the two experimental group were significantly different than the control group (P<0.05);5. Transwell invasion assay:EGF promoted the invasion and metastases of Hep-2 cells, the invasive numbers of Hep-2 cells with EGF treatment at 24h time point were 58.5±10.3 (P<0.05); Erlotinib inhibited the invasion and metastases of Hep-2 cells, the invasive numbers of Hep-2 cells with Erlotinib treatment at 24h time point were 18.7±4.8, which was significantly less than the control group (P＜0.05).Conclusion:1.EGFR signal modulation regulates E-cadherin expression and cell phenotype, motility, invasion and metastases in human laryngeal cancer cell line Hep-2. 2. Treatment of small molecule tyrosine kinase inhibitor erotinib up-regulates E-cadherin expression, decreases cell motility and movement, inhibits cell invasion and metastases in Hep-2 cells.