| Project/Area Number |
14570826
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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 |
Dermatology
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| Research Institution | St.Marianna University School of Medicine |
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Principal Investigator |
MIZOGUCHI Masako St.Marianna University School of Medicine, Department of Dermatology, Professor & Director, 医学部, 教授 (30010250)
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| Co-Investigator(Kenkyū-buntansha) |
KAWAKAMI Tamihiro St.Marianna Univ., Dept of Dermatol., Lecurer, 医学部, 講師 (20297659)
KAWA Yoko St.Marianna Univ., Dept of Dermatol., Assistanat, 医学部, 助手 (10082273)
鹿島 眞人 聖マリアンナ医科大学, 医学部, 講師 (50169421)
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| Project Period (FY) |
2002 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2003: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2002: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Keywords | differentiation of melanocyte / 1.25(OH)2D3 / endothelin B receptor / MITF / elastin / elastin receptor / neural crest cell line / 悪性黒色腫の治療 / 色素異常症 / 活性型ビタミンD_3 |
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| Research Abstract |
The effects of 1.25-dihydroxyvitamin D3(1.25(OH)2D3) on the differentiation of immature melanocyte precursors were studied. The NCCmelb4 is an immature melanocyte cell lie established from mouse neural crest cells(NCC). 1,25(OH)2D3 inhibited the growth of NCCmelb4 cells at concentrations higher than 10-8M. That growth inhibition was accompanied by the induction of tyrosinase and a change in DOPA reactivity from negative to positive. Electron microscopy demonstrated that melanosomes were in more advanced stages after 1,25(OH)2D3 treatment. In primary cultures of murine NCC, DOPA-positive cells were increased after 1.25(OH)2D3 treatment. These findings indicate that 1.25(OH)2D3 stimulates the differentiation of immature melanocyte precursors. Moreover, immunostaining and RT-PCR analysis revealed that endothelin B receptor (EDNRB) expression was induced in NCCmelb4 cells following treatment with 1.25(OH)2D3. The induction of EDNRB by 1.25(OH)2D3 was also demonstrated in neural crest prima
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ry culture, but not in immature melanocytes. The expression of microphthalmia-associated transcription factor (MITF) was induced in NCCmelb4 cells treated with 1.25(OH)2D3 and endothelin 3, but not by 1.25(OH)2D3 alone, suggesting that endothelin 3 may stimulate the expression of the MITF gene after binding to the EDNRB induced by 1.25(OH)2D3. These findings suggest a regulatory role for vitamin D3 in melanocyte development and melanogenesis.
Using the same cell line, the effects of elastin on the melanocyte differenciation were studied. We identified the 67 kDa elastin receptor and its functions for the first time of murine melanocyte precursors of both neural culture explants and the cell line. Confocal microscopy showed the elastin receptors located in the cell membrane and cytoplasm. mRNA analysis also confirmed the expression of the receptor. We cultured neural crest explants on elastin peptide (VGVAPG)-coated and non-coated plates and found that elastin significantly stimulated melanocyte precursor's DOPA reaction, migration, and dendrite formation during melanocyte lineage development of the explants. An electron microscopic examination revealed that elastin increase melanosome maturation. However, elastin did not change significantly the number of TRP-1-positive melanocytes in the explants. The alamar blue assay revealed that 10-5 -10-9 M of elastin peptide did not proliferate NCC melb4 cells. Taken together, elastin accelerates differentiation, migration and dendrite formation of melanocyte precursors without significant proliferation. Elastin, the major component of elastic fiber, may play a previously unappreciated role in regulating the behavior of immature melanocytes. Less
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