Expression Of EphB1 And EphrinB1 In Human Esophageal Carcinoma And Their Relationship With Clinical Parameters

Esophageal carcinoma is one of the most malignant tumours of digestive tract, and it is the eighth most common cancer worldwide, and the sixth most common cause of death from cancer. In the worldwide, the high incidence areas of esophageal cancer include:China, Southern Africa, Eastern Africa and so on. However, according to the latest report, the highest region of esophageal standardized mortality is still Henan Province (32.95/100,000). It is very important to further research the related genes of esophageal cancer for early diagnosis of esophageal cancer and find the effective gene therapeutic targets.Eph/Ephrin family is one of the largest known RTKs family. Eph receptor and ephrin ligands mediate bidirectional signaling participate in physiology, homeostasis and diseases and tumorigenesis. At present, the relationship between Eph/Ephrin expression and esophagus cancer has been rarely reported. Therefore, this research assayed the expression levels of EphB1 and EphrinB1 in esophageal cancers tissues and their matched normal tissues, and analyzed their relationship with clinical and pathological parameters of esophageal cancers. And further investigate the function of EphrinB1 in esophageal cancer cell lines.Material and methods1 Subjects, samples and clinical dataSubjects were hospitalized patients diagnosed as esophageal cancer waiting for surgery in the Tumor Hospital of Linzhou City in Henan Province from 2007 to 2008. According to pathological identification, the cancer tissues and matched normal esophageal tissues of 92 subjects were taken during operation. The general information and clinical information of the subjects were collected.2 Determination of EphBl and EphrinBl mRNA in sampleReal time PCR was used to determine the mRNA expression levels of EphB1 and EphrinBl in the esophageal cancer tissues and matched normal esophageal tissues,β-actin was used as an internal reference gene.3 Determination of EphBl and EphrinBl protein in samples The expression of EphB1 and EphrinB1 protein in the esophageal cancer tissues and the corresponding normal esophageal tissues was detected by western blot and immunohistochemistry. All samples were observed under the three strongest focuses at×200 magnification. Calculating the number of positive cells and the percentage of cells expressing target protein. The percentage ratio of positive cells of esophageal cancer tissues and that of the matched normal esophageal tissues was the index of protein expression level.4 Cell transfectionTransfer the carrier of EphrinB1 expression and pcDNA3.1 empty carrier into TE-12 cells respectively. Detecting EphrinB1 protein level by Western, and analysis the influence of the growth of TE-12 cells by MTT method.5 Data analysisThe data were processed by SPSS16.0 statistical software. Analysis mRNA expression in 92 pairs of esophageal cancer tissues and matched normal tissues by paired test,X2-test was adopted to study the relationship between the EphB1/ EphrinB1 mRNA and protein expression levels and its clinical parameters and other related factors in 92 pairs of esophageal cancer samples. The level of statistical significance was 0.05.Results1 The expression of EphBl/EphrinBl mRNA in esophageal cancer and the relationship with the individual clinical and pathological characteristics1.1 The expression of EphBl and EphrinB1 mRNA in esophageal cancerReal time PCR results showed that up-regulation of EphB1 and EphrinB1 mRNA expression was respectively found in 66.3% (61/92 cases) and 64.2 %(59/92 cases) esophageal cancer tissues compared with the paired normal esophagus tissues. Paired t-est indicated that the mRNA expression levels of EphB1 and EphrinB1 in esophageal cancer tissues and matched normal esophageal tissues are statistically significant (t=8.555, P＜0.001;t=7.488, P＜0.001). Pearson's analysis shows, EphBl and EphrinB1 mRNA expression is highly relevant in esophageal cancer tissues (r= 0.625, P＜0.001). 1.2 The relationship between EphBl/EphrinBl mRNA expression and the individual clinical and pathological characteristics in esophageal cancerThe expression level of EphB1 mRNA in patients with tumor family history was also significantly up-regulation compared with that of the ones without tumor family history (X2=6.458, P＜0.05). The expression level of EphB1 mRNA in patients with lymph node metastasis was significantly higher than that of the ones without lymph node metastasis (X2=7.422, P＜0.01).The expression level of EphrinB 1 mRNA of smokers was significantly higher than that of non-smokers (X2=4.394, P<0.05). The expression level of EphrinB1 mRNA of the patients with lymph node metastasis was significantly up-regulation compared with that of the ones without lymph node metastasis (X2=9.159, P＜0.01). EphrinB1 mRNA expression with infiltrating depth was significant correlation (x2= 6.438, P＜0.05).2 The expression of EphBl/EphrinBl protein in esophageal cancer and the relationship with the individual clinical and pathological characteristics2.1 Western blotThe expression levels of EphB 1 and EphrinB 1 protein in tumor tissues were up-regulation compared with the normal tissues.2.2 The expression of EphBl and EphrinBl protein in esophageal cancerThe expression of EphB1 and EphrinB1 protein respectively in 63.0% (58/92 cases) and 60.9% (56/92 cases) esophageal cancer tissues were positive compared with the paired normal esophagus tissues, and the results were consistent with real time PCR results.2.3 The relationship between EphBl/EphrinBl protein expression and the individual clinical and pathological characteristics in esophageal cancerThe expression level of EphB1 protein in patients with tumor family history was significantly higher than that of the ones without tumor family history (X2=4.147, P<0.05). The expression level of EphB1 protein in patients with lymph node metasta-sis was significantly higher than that of the ones without lymph node metastasis(X2= 5.733, P<0.05). EphB1 protein expression with infiltrating depth was significant co-rrelation (X2=5.674, P< 0.05). EphrinB1 protein expression of smokers was significantly higher than that of non-smokers (X2=5.787, P<0.05). EphrinB1 protein expression of the patients with tumor family history was significantly up-regulation compared with that of the ones without tumor family history (X2=5.376, P<0.05). EphrinB1 protein expression of the patients with lymph node metastasis was significantly higher than that of the ones without lymph node metastasis (X2=7.928, P<0.01). The expression level of EphrinB1 protein of the patients with tumors of deeper infiltrating depth was significantly high-er than that of the ones with tumors of thinner infiltrating depth (X2= 9.113, P< 0.01).3 Cell transfection resultsHuman esophageal cancer cell lines after transfection EphrinB1 enhanced the proliferative ability. With the growth of EphrinB 1 expression level, the proliferative capacity of the cancer cells gradually increased.Conclusion1 The expression level of EphB 1 was up-regulated in esophageal cancer tissues, compared with the paired normal tissues. The expression levels of EphB 1 mRNA and protein were closely correlated with lymph node metastasis and family history.2 The expression level of EphrinB1 was up-regulated in esophageal cancer tissues compared with the paired normal tissues. The expression levels of Ephrin B1 mRNA and protein were significantly correlated lymph node transfer and infiltrating depth.3 The expression of EphB 1 and EphrinB 1 in esophageal cancer was highly relevant, and both maybe interaction in the process of tumorigenesis and development, and common regulating tumor angiogenesis, tumor cell invasion and metastasis.4 The overexpressed EphrinB1 may enhanced the growth and proliferative capacity of human esophageal cancer cells.