1988 Fiscal Year Final Research Report Summary
Development of serum-free and protein-free cell culture methods for a long-term of mammalian tissue cells including human cells and their application to medical research
| Project/Area Number |
60870018
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| Research Category |
Grant-in-Aid for Developmental Scientific Research
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| Allocation Type | Single-year Grants |
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| Research Field |
Experimental pathology
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| Research Institution | Kyoto University |
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Principal Investigator |
ADACHI Toshihiro Assistant, Department of pathology, Faculty of Medicine Kyoto University, 医学部, 助手 (70025609)
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| Co-Investigator(Kenkyū-buntansha) |
能勢 尚志 鐘紡薬品研究所, 所長
OKAMOTO Eiichi Assistant, Department of Pathology, Faculty of medicine, Kyoto University, 医学部, 助手 (30115810)
OKADA Shigeru Lecture, Department of pathology, Faculty of Medicine Kyoto University, 医学部, 講師 (20033201)
MIDORIKAWA Osamu Professor, Department of Pathology, Faculty of Medicine, Kyoto University, 医学部, 教授 (40025515)
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| Project Period (FY) |
1985 – 1987
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| Keywords | Serum-free culture method / fibronectin / 70K cell-spreading protein / Monoclonal antibodies immunihistochemistry / 70K細胞接着蛋白質 / 免疫組織化学 / 細胞接着阻止蛋白質 |
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| Research Abstract |
The first purpose of this experiment was to establish various kinds of cell lines growing in synthetic, defined medium and the second purpose was to detect bioactive substances (factors) in the medium and their application for medical research. Various kinds of cell lines were established to grow continuously in modified Ham F12 medium without any addition of serum or proteins. The cell lines includ mous fibroblast cell line (m cells, F cells), so-called reticulum cell lines (MR cells and SP cells), human fibrosarcoma cell line (HT1080-SF2), human teratocarcinoma cell line (HMTcells) and human kidny adenocarcinoma cell line. The second purpose is to detect bioactive factors (proteins) from the serum-free medium. The cell lines growing in the serum-free medium were morphologically classified into two groups, adherent and non-adherent cells. Adherent cells (m cells) produce fibronectin which is related to microfilament activation and cell growth. This cell line also produced transferrin
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in the medium. On the other hand, non-adherent cells (MR cells and SP cells) did not produce fibronectin, but these cell lines produced fibronectin-receptorsin serum-free culture medium and when fibronectin is added in the medium, these cells spread on th culture wessels. Human plasma contains adhesive proteins, that is, fibronectin and 70K cell-spreading protein. As far as the cell lines established in our laboratories did not produce 70K cell-spreading proteins in vitro. Monoclonal antibodies were produced against humam 70K cell-spreading protein. By immunohistochemical studies, we found that the 70K cell-spreading protein is mainly produced from epithelial cells such as liver and pancreatic duct cells. By the appication of serum-free culture method, we found that the cells produce cell-spreading inhibitory protein and the protein(s) were also founf in the human plasmaand were purified. This cell-spreadin inhibitory protein is basic protein with molecular-weight of 50-65K and bind to both the fibronectin and 70K cell-spreading protein.
The cell lines reported here are very usuful for the studies of cytokines produced from the cells and is also useful for the application to medical research. Less
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