| Project/Area Number |
10671096
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General surgery
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| Research Institution | AKITA UNIVERSITY |
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Principal Investigator |
MINAMIYA Yoshihiro Sch. Of Med., Akita Univ., Associate Professor, 医学部, 助教授 (30239321)
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| Co-Investigator(Kenkyū-buntansha) |
KAWAI Hideki Sch. Of Med., Akita Univ., Research Associate, 医学部, 助手 (20291271)
OGAWA Junichi Sch. Of Med., Akita Univ., Professor, 医学部, 教授 (20112774)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1998: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | Cancer metastasis / endothelial cell / myosin light chain kinase / myosin light chain / hepatic growth factor / myosin light chain kinase / HGF / myosin light chair kinase / myosin light chair |
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| Research Abstract |
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.
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