Mann-Whitney U test was used to analyze the association between mRNA expression levels and the clinical histopathological parameters of the patients. The survival of ��-Nicotinamide order patients with ESCC after surgery was examined using the Kaplan-Meier method, and the survival times were compared using the log-rank test. Univariate analysis and multivariate analysis was performed using the Cox’s regression model. P-values were

considered significant at p < 0.05. Results Quantitative RT-PCR of VEGF-C in cell lines We first investigated the expression of VEGF-C in 12 esophageal cancer cell lines (KYSE30, KYSE50, KYSE70, KYSE110, KYSE140, KYSE150, KYSE180, KYSE270, KYSE410, KYSE450, KYSE510, KYSE520), and in the Het-1A cell line. In most of the KYSE series of cell lines, especially KYSE410, high levels of VEGF-C were detected, yet in Het-1A, VEGF-C was not detected at

all (Fig. 1). Figure 1 The expression of VEGF-C in esophageal cell lines. Most KYSE cell lines S3I-201 supplier express VEGF-C. Het-1A cells do not express VEGF-C. Quantitative RT-PCR of VEGF-C in clinical specimens We next examined VEGF-C expression in 106 pairs of resected ESCC tumors and in corresponding noncancerous esophageal mucosal tissue find more specimens. Our data reveals that VEGF-C expression in cancerous tissue is higher than in corresponding noncancerous esophageal mucosa (Fig. 2a). We also examined the relationship between the clinico-pathological factors and the expression of VEGF-C in ESCC. The expression of VEGF-C was found to be higher in Stage2B-4A tumors than in Stage0-2A tumors (Table 1, Fig. 2b). We also examined the relationship between the expression of VEGF-C and the survival data. The patients were divided into two groups according to the expression of VEGF-C. The cut off value was median expression of VEGF-C (high expression group of 53 cases and a low expression group of 53 cases). The patients in the high VEGF-C expression group had significantly shorter survival after surgery than the patients in the low expression group (p = 0.0065 by log-rank test; Fig. 3). Univariate analysis showed that, among the clinico-pathological factors, the extent of the primary

tumor, lymph node metastasis, and high expression of VEGF-C were all statistically significant prognostic factors (Table 2). Multivariate analysis showed that the extent of the primary tumor and lymph node metastasis ROS1 were independent prognostic factor (Table 3). Figure 2 Comparison of mRNA expression of VEGF-C in cancer and corresponding noncancerous esophageal mucosa (a) and in Stage0-2A patients and Stage2B-4A patients (b). The VEGF-C expression in ESCC tumors is significantly higher than in the corresponding noncancerous esophageal mucosa (a). The VEGF-C expression is higher in Stage2B-4A patients than in Stage0-2A patients (b). Figure 3 Survival rate of patients with ESCC according to the mRNA expression of VEGF-C. Patients with high expression of VEGF-C have significantly shorter survival after surgery (p = 0.