| Project/Area Number |
11839029
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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 Institution | University of Occupational Environmental Health |
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Principal Investigator |
FUJIHIRA Takashi UOEH, School of Med., First Dept.of Int.Med., Assistant professor, 医学部, 講師 (40159124)
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| Co-Investigator(Kenkyū-buntansha) |
江藤 澄哉 産業医科大学, 医学部, 教授 (90010347)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Keywords | endocrine disruptor / protein phosphorylation / two dimensional gel electrophoresis / signal transduction / 二次元電気泳動 / 細胞内情報伝達 |
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| Research Abstract |
Recently importance of environmental endocrine disruptors has been notified in many countries. These small chemical compounds influenced human endocrine system, especially reproductive system. Various testing methods for evaluating endocrine disrupting effect are being developed. Bisphenol A (BPA) is one of endocrine disruptors and acts like estrogen. BPA is used as coating materials in infant feeding bottles, inside coating of canned food, polycarbonated devices for eating and prosthodontics. We focused on intracellular protein phosphorylation and tried to develop its detecting system by two dimensional gel electrophoresis (2D-PAGE). Primary cultured thyroid cells from patients with Graves' disease and human neuroblastoma cell line (MSN cells) were used as a sample. Cultured cells were stimulated by various concentration of BPA and extracted protein sample was separated by 2D-PAGE and western blot analysis was performed by anti-phosphothyrosin, anti-phosphoserine and anti-phosphothreonine antibodies and was detected by enhanced chemiluminescence (ECL) reaction. BPA increased protein expression of 80 kDa spot in human thyroid follicular cells and facilitated thyrosin phosphorylation of 34 kDa spot in MSN cells on dose-dependent manner. BPA promoted heat shock protein 27 (HSP27) expression in MSN cells. HSP27 is thought to be important for regulation of cell function in estrogen responsive cells. Our detected system for endocrine disruptors is not specific method for intracellular individual signalling step, however, to target protein phosphorylation is very useful for broad screening system of signal transduction abnormalities. Moreover, significant detected spot on 2D-PAGE is going to be analysed by proteome databases on the Internet. We are trying to accumulate spot data influenced by other endocrine disruptors.
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