The Expression Of PAR2 In Esophageal Carcinoma And The Role Of Trypsin For The Proliferation And Migration Of EC109 Cells

Background and objectiveProtease-activated receptor 2 (PAR2) belongs to G protein–coupled receptors (GPCRs) that transmit cellularresponses begun by the actions of extracellular proteases. Aberrant expression of PAR2 has been associated with increased angiogenesis, tumor growth, and metastasis of various cancers. Although PAR2 has been described to play a role in different malignancies, its expression and biological activity in esophageal carcinoma are mostly unknown. We assessed the status of PAR2 expression in esophageal carcinoma,and the role for the tumor growth, and metastasis of esophageal carcinoma.Materials and methods1 Tissue samples: Specimens were obtained from 124 patients who underwent biopsy for esophageal carcinoma at Tumor Hospital of Shantou University Medical College from 2002,2007 to 2010, 49 patients have mucous membranes beside the esophageal carcinoma and faraway from the esophageal carcinoma. All of the patients have therapy records and 75 patients have been observed for 5 years(38 patients of them accepted only esophagectomy and 37 patients of them accepted adjuvant therapy after operation); 58 tissues had Lymph node metastasis and 66 tissues had no Lymph node metastasis; According to pathological classification, 40 tissues were at gradeⅠ,73 tissues at gradeⅡ,11 tissues at gradeⅢ.2 HE stain: Cell morphology was observed by light microscope after HE stain.3 Cell culture:Culture the EC109 cells.4 Immunohistochemistry: The ImmunoCruzTM Staining System three steps method were performed in the study to test the expression of PAR2 in the esophageal carcinoma. Two steps method were performed to test the expression of CD34 in the same patient. 5 Immunofluorescence cell staining :To test the expression of PAR2 in the EC109 cells6 methyl thiazolyl tetrazolium:To test the effect of trypsin on EC109 cells proliferation.7 Cell migration assay: To test the effect of trypsin on EC109 cells migration.Result1 Immunohistochemistry: There were significant overexpressions(65.3%, 81/124) of PAR2 in the cancer cells of the esophageal carcinoma tissues,compared with the expressions of PAR2 on the mucous membranes beside the esophageal carcinoma are 30.6% (15/49) and on the mucous membranes faraway from the esophageal carcinoma are 32.7% (16/49)(P<0.01). And its expression had a significant relationship with histology grade. GradeⅡ(74%) relate with GradeⅠsignificantly, but not with GradeⅢ. MVD of PAR2 expression positive group is 23.59±8.93,of PAR2 expression negative group is 23.27±8.17. PAR2 expression positive patients (28.55±5.43月) had less survival time than negative patients (32.33±6.49月), but the difference was not significant.2 Immunofluorescence cell staining: There was overexpression of PAR2 in the EC109 cells.3 Methyl thiazolyl tetrazolium: There was some complicated effect of trypsin on the proliferation of EC109 cells.4 Cell migration assay: There was some effect of trypsin on the migration of EC109 cells.Conclusion1. The expression rate of PAR2 in esophageal cancer cells is 65.3%, higher than on mucous membranes beside the esophageal carcinoma and mucous membranes faraway from the esophageal carcinoma. And the expression has a significant relationship with histology grade, GradeⅡhas more patients than other grade. It suggested that PAR2 activation has a role on the development of esophageal cancer. 2. The expression of PAR2 in esophageal cancer tissue has no relationship with tumor microvessel density. This suggested that PAR2 expression don't affect the development of cancer through promoting tumor angiogenesis in esophageal cancer.3. PAR2 expression positive patients have less survival time than PAR2 expression negative patients, but the difference isn't significant. We need more tissues to study the relationship between PAR2 expression and survival.4. PAR2 was expressed in esophageal cancer cell line EC109. Trypsin had migration effect on EC109 esophageal cancer cell.