6366

CCL21

Secondary lymphoid tissue chemokine (SLC/CCL21) and its receptor CCR7 have been implicated in lymph node metastasis, whereas the mechanism of which remains unclear. epithelial-mesenchymal transition (EMT) plays an important role in invasion and migration of cancer cells. We presumed that CCL21/CCR7 axis activates EMT process to induce cancer cell invasion and metastasis. Firstly, the expressions of CCR7 and EMT markers were examined by immunohistochemical staining in the primary breast carcinoma tissues from 60 patients who underwent radical mastectomy. Then, we investigated whether CCL21/CCR7 induces EMT process during mediating cancer cell invasion or migration in vitro. By immunohistolochemistry, high expressions of CCR7, Slug and N-cadherin were seen in 60, 65, and 76.67 % of tumors, respectively, and significantly associated with lymph node metastases as well as clinical pathological stage. Furthermore, the CCR7 expression was significantly correlated to Slug and N-cadherin. In vitro, stimulating breast cancer cell lines 1428, MCF-7 and MDA-MB-231 with CCL21, the invasion and migration of tumor cells were promoted, and simultaneously, EMT phenotype of tumor cells was enhanced, including down-regulation of E-cadherin, up-regulation of Slug, Vimentin and N-cadherin at both protein and mRNA levels. Inversely, knockdown of CCR7 by shRNA suppressed tumor cell invasion, migration and EMT phenotype induced by CCL21. These results indicated that CCL21/CCR7 axis could activate EMT process during chemotaxis of breast carcinoma cells.

In recent decades, the CXC chemokine receptor 7 (CCR7) [corrected] and its ligand CCL21 have been extensively reported to be associated with tumorigenesis. Meanwhile, Slug signaling induces the epithelial-mesenchymal transition (EMT) process in chondrosarcoma development. In the present study, we explored the functions of CCL21/CCR7 [corrected] in Slug-mediated EMT in the chondrosarcoma. We analyzed protein expression of CCR7 [corrected] and Slug in human chondrosarcoma samples. Effects of CCR7 [corrected] on chondrosarcoma cells were assessed by in vitro assays. Additionally, CCR7 [corrected] pathways were further investigated by pharmacological and genetic approaches. We found that the altered CCR7 [corrected] (81.7 %) and Slug (85.0 %) expression in human chondrosarcoma tissues were significantly associated with grade, recurrence, and 5-year overall survival. According to in vitro assays, CCL21 stimulation induced the expression of phosph-ERK, phosph-AKT, Slug and N-cadherin in SW1353 cells, while the expression of E-cadherin was down-regulated. Furthermore, Slug signaling modulated E- to N-cadherin switch, which was influenced by the kinase inhibitor PD98059 and LY294002. In addition, the genetic silencing of Slug inhibited the capacity of migration and invasion of SW1353 cells. In conclusion, CCL21/CCR7 [corrected] pathway activates ERK and PI3K/AKT signallings to up-regulate Slug pathway, leading to the occurrence of EMT process in human chondrosarcoma. This study lays a new foundation for molecule-targeted therapy of human chondrosarcoma.

OBJECTIVE: To investigate the mechanism underlying that chemokine (C-C motif) ligand 21 (CCL21) promotes the metastasis ability of human pancreatic cancer Panc-1 cells. METHODS: Transwell(TM) was used to access the chemotaxis effect of CCL21 on Panc-1 cells. Real-time quantitative PCR was performed to detect the expression of C-C chemokine receptor type 7 (CCR7) mRNA in the upper and lower chambers. Immunofluorescence staining and Western blotting were employed to examine the expressions of the epithelial-mesenchymal transition (EMT)-related proteins and CD133 of Panc-1 cells in the lower chamber, which were compared with those of the upper chamber as the control. RESULTS: The numbers of the Panc-1 cells induced by 0, 50, 100, 200 ng/mL CCL21 were 13.00 +/- 3.00, 78.00 +/- 9.00, 161.00 +/- 11.00, 281.00 +/- 17.00, respectively; with the increase of the concentration of CCL21, there were more cells migrating from the upper to the lower chamber; and the cells in the lower chamber expressed higher level of CCR7 mRNA than the ones staying in the upper chamber. The relative protein expressions of MMP-9, vimentin, E-cadherin and CD133 in the lower chamber were 0.42 +/- 0.04, 0.36 +/- 0.03, 0.12 +/- 0.02, 0.46 +/- 0.03, respectively, which were statistically significantly different from those in the upper chamber (0.15 +/- 0.02, 0.25 +/- 0.02, 0.25 +/- 0.03, 0.13 +/- 0.02, respectively). CONCLUSION: CCL21/CCR7 axis maybe play an important role in the metastasis of pancreatic cancer stem cells by EMT and up-regulation of MMP-9.

tumor cells frequently disseminate through the lymphatic system during metastatic spread of breast cancer and many other types of cancer. Yet it is not clear how tumor cells make their way into the lymphatic system and how they choose between lymphatic and blood vessels for migration. Here we report that mammary tumor cells undergoing epithelial-mesenchymal transition (EMT) in response to transforming growth factor-beta (TGF-beta1) become activated for targeted migration through the lymphatic system, similar to dendritic cells (DCs) during inflammation. EMT cells preferentially migrated toward lymphatic vessels compared with blood vessels, both in vivo and in 3D cultures. A mechanism of this targeted migration was traced to the capacity of TGF-beta1 to promote CCR7/CCL21-mediated crosstalk between tumor cells and lymphatic endothelial cells. On one hand, TGF-beta1 promoted CCR7 expression in EMT cells through p38 MAP kinase-mediated activation of the JunB transcription factor. Blockade of CCR7, or treatment with a p38 MAP kinase inhibitor, reduced lymphatic dissemination of EMT cells in syngeneic mice. On the other hand, TGF-beta1 promoted CCL21 expression in lymphatic endothelial cells. CCL21 acted in a paracrine fashion to mediate chemotactic migration of EMT cells toward lymphatic endothelial cells. The results identify TGF-beta1-induced EMT as a mechanism, which activates tumor cells for targeted, DC-like migration through the lymphatic system. Furthermore, it suggests that p38 MAP kinase inhibition may be a useful strategy to inhibit EMT and lymphogenic spread of tumor cells.

C-C chemokine receptor type 7 (CCR7) has been implicated in lymph node metastasis of various cancers. Previous studies have revealed that epithelialmesenchymal transition (EMT) is involved in the chemotactic process mediated by CCR7 and its ligands in various types of carcinoma. However, the underlying mechanism of this process remains to be fully elucidated. The present study investigated whether chemokine (CC motif) ligand 21 (CCL21)/CCR7 may activate EMT of lung cancer cells and their associated signaling pathways. A549 and H520 lung cancer cell lines were examined in vitro in the present study. The results indicated that A549 and H520 expressed CCR7, but reduced levels of CCL21. Following stimulation of lung cancer cell lines with CCL21, the expression of the epithelial marker Ecadherin was downregulated, and the mesenchymal markers Vimentin/Slug and extracellular signalregulated kinase (ERK) were upregulated. In addition, the ERK inhibitor PD98059 may inhibit EMT caused by CCL21, and decreased cell migration and invasion initiated by CCL21. Furthermore, lung adenocarcinoma tissues from 50 patients who underwent lung cancer operations were investigated by immunohistochemistry. The findings revealed that CCR7, Slug and Vimentin were highly expressed in lung carcinoma tissues, and were significantly associated with lymph node metastasis and clinical pathological stages, respectively. CCR7 expression was correlated positively with expression levels of Slug and Vimentin. CCL21 was expressed positively in the endothelium of lymphatic vessels adjacent to cancer cells, and weakly in lung cancer cells. Collectively, these results demonstrated that CCL21/CCR7 may activate EMT in lung cancer cells via the ERK1/2 signaling pathway. The current study provides evidence that a close interaction exists between CCL21/CCR7chemotaxis and EMT procedures in lung cancer metastasis, providing a basis for the development of therapeutic targets.