| Background and objectives Tongue squamous cell carcinoma(TSCC) is one of the most common malignant tumors in head and neck. It has the features of strong infiltration, rapid growth,high malignancy and the risk of cervical lymph nodal metastases early, which are related to the stage and are significantly affected treatment. In recent years, Although comprehensive treatments of malignant tumor have made rapid progress, the recurrence rate and mortality of patients with tongue squamous carcinoma is still high due to its strong aggressivity and high lymph node metastasis rate. The abilities of invasiveness and distant metastasis on the prognosis of Tongue squamous Carcinoma, Therefore, the study of invasion and metastasis mechanism and how to block this process to improve the survival rate of the patients of Tongue squamous Carcinoma is very important. In recent years, molecular targeted drug therapy as an important component of biological therapy has received increasing attention, and the Biological Therapy for cancer has become the fourth treatment methods after surgery, radiotherapy and chemotherapy. The mechanism of growth, metastasis and invasion of tongue squamous cell carcinoma has received more and more attentions by scholars, especially in cell and molecular level. Versican is a large extracellular matrix proteoglycan and and is a key ingredient of the extracellular matrix. It`s one of a member of the aggregating proteoglycans(PGs) family. Versican is composed of three domains: the amino terminal G1 domain, he carboxyl terminal G3 domain and the core protein contains the GAG attachment region and the chondroitin sulphate chains. The G1 domain is comprised of an immunoglobulin-like motif, followed by two proteoglycan tandem repeats, and the G3 domain contains two epidermal growth factor-like repeats, a carbohydrate recognition domain and complement binding protein-like subdomains. Higher expression of versican correlates with worse prognosis.The interaction of versican with its binding partners plays diverse roles in cell adhesion, proliferation, migration and angiogenesis. Versican can increase the abilities of motility, proliferation and metastasis of tumor cells has been confirmed by many studies. First, to detect the expressional information of Versican in the tongue squamous carcinomas and normal tongue tissue through immunohistochemistry(SP), and by comparison to study the relationship between Versican and the differentiation degree and the TNM stage of the tumor and local lymph nodes transfer. Second, to cultivate Tca8113 cells in vitro, and to construct the cellular model of silencing Versican gene though transfected the tongue squamous cell carcinoma cell line Tea-8113 cells with Versican-si RNA,and then observe the transfection efficiency. To observe the change of proliferation, adhesion, invasion and migration of Tca-8113 cell after transfection, and to discuss the roles of versican in the biological behavior tongue squamous cell carcinoma cell line Tca-8113 cells.Based on these results, potential therapeutic targets are suggested.This study includes the following two parts:Part?: The expression and significance of Versican in tongue squamous cell carcinomas.Materials and methods 1, Clinical materials: 120 cases of surgical resection specimen of Department of Stomatology of the First Affiliated Hospital of Zhengzhou University in January 2012 to January 2015, which confirmed as the tongue squamous cell carcinoma by pathological examination after surgery. All patients had definite pathological diagnosis and the complete clinical data. Female: 51 cases,Male:69 cases; Aged 28-75; Patients with lymph transfer: 38 cases, and without lymph transfer 82 cases; According to the international cancer UICC2002 TNM stages: period?-?79 cases, period?-?41 cases; Good differentiation(G1) :65 cases, moderately differentiation(G2) :40 cases, poorly differentiation(G3):15 cases. 2, Method: Immunohistochemical staining method is applied to detect Versican expression in 120 patients with tongue squamous carcinoma and 10 cases of normal tongue tissue.Results: Immunohistochemical staining showed that Versican positive expression appears in the tumor stroma and cytoplasm of a few cancer cells. Versican weakly expressed in vascular weeks in 10 normal tongue tissue; Versican in 120 patients with tongue squamous carcinoma is mainly expressed in tumor stroma( high expression rate was 56.67%), and the positive rate of the cancer cells was 8.33%. The high expression rate of versican is associated with the histological grading of tongue squamous carcinoma,(p < 0.05); The Versican high expression rate of TSCC tissue is related to its TNM stage; Also the versican positive rate has a relationship with lymph nodes transfer, and the high expression rate of tongue cancer with lymphatic transfer is 92.11%(p < 0.05).Brief summary: Versican expression in tongue carcinoma tissues are up-regulated compared to the nomal tongue tissue, and the expression of Versican may be related to the metastasis and prognosis of tongue squamous cell carcinoma.Part?: Cell transfection and the effects of Versican gene silence on the biological behavior of Tca-8113 cells.Materials and methods 1,Cell lines: Human tongue squamous carcinoma cells Tca-8113 bought in Benjing TIANDZ biotechnology company. 2,Methods: 1) Regular culture of Tca-8113 cell lines. 2) Transfected Tca-8113 cells with Versican-si RNA to constructing the cellular model of silencing Versican gene. 3) The experiment is divided into three groups: experimental group, blank control group and negative control group. 4)Transfection efficiencies of versican-si RNA were analyzed by fluorescent microscop; q RT-PCR was used to detect the changes of m RNA level of Versican in Tca-8113 cells after versican silencing; At last, MTT assay detected the influences on the proliferation and Scratch test was used to examine the cell migratory of Tca-8113 cells after versican silence in vitro.Cell adherence and transwell assay were used to check the influence of versican silence on the adherent and invasive ability of Tca-8113 in vitro respectively.Results 1)The high-efficiency transfection( above 70%) were obtained. QRT-PCR indicated that the m RNA of Versican was down-regulated in the experimental group compared to the controls(P < 0.05). 2) MTT assay, Scratch test and Transwell assay demonstrated that Versican silencing reduced the proliferative, invasive and migratory abilities of Tca-8113 cells in vitro(P < 0.05); and adhesion ability of Tca-8113 cells was more stronger assayed by cell adhesion teset in vitro(P < 0.05).Brief Summary Versican-si RNA can effectively interfere the expression of Versican in the tongue squamous carcinoma Tca-8113 cells in m RNA level. Silencing the expression of Versican Can up-regulate the ability of adhesion and down-regulate the abilities of proliferation, invasion and migration in vitro of Tca8113 cells.Conclusion 1) Versican expression in tongue carcinoma tissues are up-regulated compared to the nomal tongue tissue, and the expression of Versican may be related to the metastasis and prognosis of tongue squamous cell carcinoma. 2) Versican-si RNA can effectively interfere the expression of Versican in the tongue squamous carcinoma Tca-8113 cells in m RNA level. 3) Silencing the expression of Versican Can up-regulate the ability of adhesion and down-regulate the abilities of proliferation, invasion and migration in vitro of Tca8113 cells. |
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