The Study Of Extracellular Signal-Regulated Kinase And PTEN Protein Expression In Oral Squamous Cell Carcinoma

Introduction and ObjectiveOral squamous cell carcinoma ( OSCC) is one of the most commonly oral cancer encountered in clinic that is a serious threat to human health with a high morbidity and lymph node transfer. It has been estimated that the incidence of OSCC is 90% of all human oral mucosal malignant tumor and 40% of the head and neck tumor.Mitogen activated protein kinase ( MAPK) is one of the most important membrance - to - nucleus signaling mechanisms. Extracellular signal - regulated kinase (ERK) pathway including ERK1 and ERK2 is a subfamily of MAPK. ERK signal pathway is a key and classic pathway of the system, and mainly participates in adjusting cell - cycle and promotes cell proliferation and differentiation. It has been indicated that ERK activation is closely related to carcinogenesis and development and malignancy potential of head and neck cancer.PTEN gene is a tumor suppressor gene, encoding a dual - specificity phosphatase. PTEN protein participates in adjusting many kinds of cell signal -transduction pathway and is closely related to cellular growth, differentiation, and carcinogenesis. PTEN protein negative expression has been commonly identified in many tumor cell lines and primary tumors. Most of scholars considered that PTEN protein could inversely regulate and control the activation of ERK/ MAPK signal pathway in many ways.Strating from mRNA and protein level, the aim of this research was to investigate the relationship between ERK1/2 signal - transduction pathway activation and carcinogenesis and progression of OSCC. We also investigate the rela-tionship between PTEN protein absent and ERKl/2 protein activation, the influence of ERK to Tca8113 cell biological characters, in order to study carcinogenesis and treatment mechanism of OSCC in clinic.Materials and MethodsFresh material was obtained from 32 patients with squamous cell carcinoma and 13 patients with oral mucosa. Reverse transcriptase polymetase chain reaction (RT-PCR) was applied to detect ERKl/2mRNA; Western blot was applied to detect ERKl/2 protein expression; Immunohistochemistry was applied to detect PTEN protein expression; and in the meanwhile the change of ERKl/ 2 activity was examined to measure its efficiency of phosphate transferase; Human Tca8113 cell line were grown as monolayer in RPMI1640 medium supplemented with 10% fetal calf serum. Meanwhile Tca8113 cell line were grown in RPMI1640 medium supplemented with 10% fetal call serum and 40μmol/L PD98059. Light microscope was applied to detect morphy exchange of ERK to Tca8113 cell;MTT colormetric assay and Flow cytometry were applied to detect cell proliferation, cell cycle and cell apoptosis of ERK to Tca8113 cell.ResultsIn OSCC, ERK1 mRNA, ERK2mRNA expression level and ERKl/2 activity were higher than oral normal mucosa (P <0. 05) ; ERKlmRNA expression appeared to be correlated with ERK2mRNA (r =0. 388 , P =0. 028) , and giving priority to ERK2mRNA expression. ERKlmRNA and ERK2mRNA expression were not correlated with clinical stages, lymph node metastasis, differentiations and regions of OSCC (P > 0. 05). ERK 1/2 activity was correlated with neither ERK1mRNA nor ERK2mRNA expression levels ( r = 0.024, P = 0. 895; r =0. 116, P =0. 528). ERK1/2 activity was higher in clinical Ⅲ -Ⅳ stages and lymph node metastasis cases than in clinical Ⅰ - Ⅱ stages and no lymph node metastasis cases (t=2.761, P=0.010; t=3.013, P=0.005). ERKl/2 activity was also not correlated with differentiations and regions of OSCC (F = 1.829, P = 0.179; t = 1.371, P=0.181).The positive expression of PTEN protein was 62. 5% of OSCC cases. The positive expression rate of PTEN protein in clinical Ⅰ — Ⅱ stages and clinical Ⅲ -Ⅳ stages has no significant difference (P =0.387). The positive expression rate of PTEN protein in lymph node metastasis cases was higher than in no lymph node metastasis cases (P=0.016); PTEN protein expression was not correlated with differentiations and regions and clinical stages of OSCC (P =0. 503; P=0.447; P=0.387) ; In OSCC, ERKl protein expression appeared to be correlated with ERK2 protein expression (r=0.412, P = 0.019) , and giving priority to ERK2 protein expression. ERKl, ERK2 protein expression were not correlated with clinical stages, lymph node metastasis, differentiations and regions of OSCC (P>0.05). ERKl/2 activity was correlated with neither ERKl nor ERK2 protein expression (r=0.119, P = 0. 515; r = 0. 024, P = 0. 895); In the cases of PTEN protein expression deletion, the ERKl/2 activity was higher than in the cases of PTEN protein positive expression (t=2. 299, P = 0. 029) , but there was no significant difference in ERKl and ERK2 protein expression (t=0.837, P=0.409; t=0.659, P =0.515).After treatment with 40μmol/L PD98059, ERKl/2 activity of Tca8113 cell was decreased significantly (P <0. 05). The inhibition rates were 12. 16% , 35. 75% , 50.92% and 57.62% of 24h, 48h, 72h and 96h after treatment with PD98059. The inhibition rate increased significantly (F = 19. 60, P =0. 021). Cell cycle analysis indicated that the ratios of GO/Gl phase cell was increased significantly compared with normal cultured cell (F = 68. 86, P = 0. 003) , the ratio of GO/Gl phase cell decreased but there was no significant difference (F = 1.77, P=0. 274) , the ratio of G2/M phase cell decreased significantly (F = 104.63, P =0. 002). The ratios of apoptotic cells treated with PD98059 were 2. 35% , 3. 79% and 5. 60% for 24h, 48h and 72h,were decreased significantly compared with normal cultured cell (F = 30.02, P=0.012).ConclusionsThese data suggest that ERKl/2 signal - transduction pathway is activating...