Genetic analyzes to clarify the pathophysiological role of adrenomedullin in cardiovascular system
| Project/Area Number |
08670840
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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 |
Circulatory organs internal medicine
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| Research Institution | Dokkyo University School of Medicine |
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Principal Investigator |
ISHIMITSU Toshihiko Dokkyo University School of Medicine Department of Medicine, Associate Professor, 医学部, 助教授 (80232346)
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| Co-Investigator(Kenkyū-buntansha) |
KANGAWA Kenji National Cardiovascular Center Research Institute Department of Biochemistry, Di, 生化学部, 部長 (00112417)
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| Project Period (FY) |
1996 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥2,500,000 (Direct Cost: ¥2,500,000)
Fiscal Year 1997: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1996: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | adrenomedullin / gene expression / vascular endothelial cells / transcription factors / 遺伝子 / プロモーター領域 |
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| Research Abstract |
The purpose of this study is to elucidate the pathophysiological role of adrenomedullin (AM), a hypotensive peptide, in cardiovascular system. Because vascular cells are supposed to be the main source of AM,the luciferase vectors containing promotor region DNA of human AM gene in various length were transfected into human aortic endothelial cells (HAEC) using liposome, and the regulatory mechanism human AM gene expression in vascular endothelial cells were examined. Functional cis-elements were ascertained by gel shift assay using HAEC nuclear extract. Northern blot analysis revealed considerable AM mRNA expression in cultured HAEC,and 2 transcription start sites were recognized at 21 and 25 bp downstream from the TATA box. Serial deletion of the inserted 5'-flanking region DNA length resulted in reduction in promoter activity by 41% between 110 and 90 bp and further reduction by 42% between 66 and 29 bp. The 19 bp DNA fragment without TATA box had almost no promoter activity. Gel shift assay revealed presence of HAEC nuclear proteins specifically bind to nuclear factor for interleukin-6 expression (NF-IL6) consensus sequence existing from -85 to -93 bp of the AM gene 5'-flanking region and activator protein 2 (AP-2) consensus sequence clustered from -33 to -68 bp. Furthermore, mutation of NF-IL6 consensus sequence in the 5'-flanking region resulted in 42% reduction in the expression of luciferase activity. These findings suggest that the TATA box is essential for the expression of AM gene and that NF-IL6 and AP-2 sites in the promoter region are the functional elements in the transcriptional regulation of human AM gene in vascular endothelial cells.
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Report
(3 results)
Research Products
(5 results)
