Abstract
Lymph node metastasis in patients with urinary bladder cancer (UBC) is always associated with poor prognosis and is the determinant for tumor staging and the development of treatment regimens; however, its underlying mechanisms remain to be studied. Immunohistochemical staining of tumor sections from 62 UBC patients was performed using CCR7, D2-40 and CD34 antibodies. We showed that increased CCR7 expression was significantly associated with positive lymph node status (P=0.008), pT3-T4 tumor stage (P=0.015), tumor grade (P=0.010) and worse overall survival (OS, P<0.001) and that both CCR7 expression and lymph node metastasis were independent prognostic factors for OS (P=0.031 and P=0.001, respectively) based on multivariate analysis. We found that there was a significant association between MLVD and lymph node status (P=0.006), but this relation was not observed for MVD. Furthermore, we showed that increased CCR7 expression correlated significantly with higher MLVD (P=0.014) and MVD (P=0.002). Wound-healing and matrigel transwell assays indicated that activation of CCR7 with CCL21 significantly enhanced the invasion and migration abilities of UM-UC-3 cells, and this enhanced effect was significantly abrogated by CCR7 knockdown using siRNA. Western blot analysis revealed that the phospho-ERK1/2 level was markedly increased when UM-UC-3 cells were treated with CCL21 and significantly decreased when the CCR7 gene was silenced. MEK/ERK1/2 inhibition with PD98059 significantly suppressed the migration and invasion abilities of UM-UC-3 cells and also significantly abrogated the effects of CCL21/CCR7 on cell migration and invasion. Based on these results, we conclude that activation of the CCL21/CCR7 chemoaxis promotes lymph node metastasis of UBC in at least two ways. Firstly, although CCR7 is a promoting factor that induces both lymphangiogenesis and angiogenesis, it may promote lymph node metastasis through its lymphangiogenic effect rather than through its angiogenic effect. Secondly, the CCL21/CCR7 chemoaxis promotes the migration and invasion of UBC cells via the MEK/ERK1/2 signaling pathway rather than the PI3K/AKT pathway.