The role of endothelial cell on cancer cell metastasis

点击翻译按钮获取中文摘要

Chemotaxis is a key step in the process of cancer cell invasion and metastasis. We studied the role of nonmuscle myosin light chain kinase (nm-MLCK) in cancer cell chemotaxis toward hepatocyte growth factor (HGF) using A549 cells, a model cell line derived from lung adenocarcinoma. Our analysis entailed characterizing expression of nm-MLCK using Western blot ; examining chemotaxis toward HGF in Boyden chambers equipped with 8 μm pore polycarbonate membranes ; examining myosin II filament formation by immunolabeling cells with anti-myosin II antiserum ; and examining myosin light chain (MLC) phosphorylation by myosin II immunoprecipitation, followed by immunoblotting with anti-phosphoserine and anti-phosphotheonine antibodies. To assess the dependency on intracellular CaィイD12+ィエD1 and MLCK activity, experiments were performed in presence and absence of the CaィイD12+ィエD1 chelator, BAPTA, or the specific MLCK antagonist, ML-7. A549 cells were found to express a 214kDa nm-MLCK, which in the presence of HGF, phosphorylated MLC on both serine and threonine residues. HGF also elicited myosin II filament formation and chemotaxis which was maximal at 24 h in the presence of 30 ng/ml HGF. All of the aforementioned reactions were inhibited by both BAPTA and ML-7. Thus, it appears that CaィイD12+ィエD1-dependent nm-MLCK activity regulates HGF-induced A549 cell chemotaxis.