2003 Fiscal Year Final Research Report Summary
Analysis of metastasis mechanism in experimental lung metastasis with heparanase transgenic tumor cells
| Project/Area Number |
14571278
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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 |
Thoracic surgery
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| Research Institution | Keio University |
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Principal Investigator |
HORINOUCHI Hirohisa Keio University, School of Medicine, Assistant Prof., 医学部, 講師 (60173647)
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| Co-Investigator(Kenkyū-buntansha) |
KOBAYASHI Koichi Keio University, School of Medicine, Professor, 医学部, 教授 (80051704)
WATANABE Masazumi Keio University, School of Medicine, Instructor, 医学部, 助手 (90201227)
YAMAMOTO Manabu Keio University, School of Medicine, Instructor, 医学部, 助手 (10317159)
KIMURA Yoshishige Keio University, School of Medicine, Instructor, 医学部, 助手 (80327535)
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| Project Period (FY) |
2002 – 2003
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| Keywords | heparanase / lung metastasis / Heparan sulphate proteoglycan / Interstitial matrix / angiogenesis |
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| Research Abstract |
To establish metastasis to the lung, several steps are considered necessary. Among them, destruction of the existing architecture is thought necessary for tumor cell growth. MMPs (Matrix metalloproteinase) are the most studied molecule which is a key factor of invasion. In existing matrix, heparin sulfate proteoglycan is forming network of skeleton and it attract various heparin binding proteins such as bFGF,PDGF,VEGF which are tightly bound to cell growth and angiogenesis. When bounded they don't influence their activity to the cells around but when released by enzymatic cleavage, their functions are activated. Key enzyme of this mechanism is heparanase.
The aim of this study is visualizing the newly generated tumor circulation and for the aid of visualization tumor cells are transected with GFP protein coding plasmid. Transfection of GFP gene was only successful within several days after transfection. Expression of the protein observed as fluorescence faded after tumor cell began to grow.
To establish lung metastasis model, 6-10 days' subcutaneous inoculation of LY80 cells and mince the subcutaneous tumor followed by intravenous injection was necessary. Following this method, after 14-16 days, lung metastasis was found.
After 14-16 days, rats were euthanized and lung metastasis were dissected, weighed, and stored in -80 degree. Heparanase activity was studied using heparin sulphate degradation kit.
Results : In small lung metastasis (less than 3mm in diameter) showed relatively high value of heparanase activity, while large metastasis (more than 3mm in diameter) showed low heparanase activity compare to the normal lung tissue. Additional
These result indicate that we may suspect in early stage of metastasis, heparanase may play an important role in degradation of existing heparin sulphate proteoglycan network in the matrix.
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