KITAMURA Kazuo Miyazaki Medical College, Faculty of Medicine, Assistant Professor, 医学部, 助手 (50204912)
NAKAZATO Masamitu Miyazaki Medical College, Faculty of Medicine, Assistant Professor, 医学部, 助手 (10180267)
|Budget Amount *help
¥6,400,000 (Direct Cost : ¥6,400,000)
Fiscal Year 1993 : ¥2,200,000 (Direct Cost : ¥2,200,000)
Fiscal Year 1992 : ¥4,200,000 (Direct Cost : ¥4,200,000)
In a survey for unknown bioactive peptides in mammalian cell lines, we detected the most potent chick rectum relaxant activity in the culture medium of phorbol ester-treated human monocytic cell line, THP-1. Thus, we started purification from the culture medium of THP-1 and isolated four peptides exhibiting strong relaxant effects on chick rectum. By microsequence analysis, amino acid sequences of these peptides were deduced to be a series of human C-type natriuretic peptides (CNPs) with several amino acid elongations from the N-terminus of CNP-22. Furthermore, CNP mRNA was detected in the THP-1 cells, and expression of CNP gene and CNP concentration in the culture medium were shown to be greatly increased by the stimulation of phorbol ester.
The demonstration of CNP production in THP-1 monocytic cell line suggests that CNP also functions as a local chemical mediator in the immune and vascular system, although CNP has been recognized to function as a neuropeptide exclusively in the cent
ral nervous system.
We have also been searching for peptides which may be relevant to circulation control, using an assay system which monitors the elevating activty of platelet cAMP.In the present study, we discovered a novel hypotensive peptide in human pheochromocytoma extract. Since this peptide is abundant in normal adrenal medulla as well as in pheochoromocytoma tissue arising from adrenal medulla, it was designated "adrenomedullin". The peptide, consisting of 52 amino acids, has one intramolecular disulfide bond and shows slight homology with calcitonin gene related peptide. It was found to elicit a potent and long lasting hypotensive effect in anesthetized rat.
Since adrenomedullin was abundant in pheochromocytoma tissue, the cDNA library of pheochromocytoma was constructed, and the cDNA clone concoding an adrenomedullin precursor was isolated and sequencd. The precursor for human adrenomedullin (human preproadrenomedullin) is 185 amino acids in length, including an adrenomedullin sequence, followed by a glycine (the amide donor). Proadrenomedullin (proAM) contains a unique twenty amino acid sequence followed by Gly-Lys-Arg in the N- terminal region. It is possible that a novel 20 residues peptide "proadrenomedullin N- terminal 20 peptide" (proAM-N20) with an amidated carboxy teminus is processed from the precursor. By RNA blot anaylysis, human adrenomedullin mRNA was found to be highly expressed in several tissues including adrenal medulla, ventricle, lung and kidney as well as pheochromocytoma.
Adrenomedullin was widely distributed in several tissues and circulates in blood in a considerable concentration. These data suggest that adrenomedullin is a new hormone participating in circulation control. Occurrence of adrenomedullin indicates the possible existence of a novel system for circulation control. Less