Systematic search on a smooth muscle stimulant peptide in mammalian kidney.

在对哺乳动物细胞系中未知生物活性肽的调查中，我们在phorbol酯处理的人单核细胞系THP-1培养基中检测到最强的鸡直肠松弛剂活性。因此，我们从THP-1的培养基开始纯化，分离出4个对鸡直肠有强松弛作用的肽。通过微序列分析，这些肽的氨基酸序列推断为一系列人类c型利钠肽（CNPs），从CNP-22的n端延伸出几个氨基酸。此外，在THP-1细胞中检测到CNP mRNA，并且在佛波酯的刺激下，CNP基因的表达和培养基中CNP的浓度显著增加。在THP-1单核细胞系中产生CNP的实验表明，尽管CNP被认为仅在中枢神经系统中作为神经肽起作用，但CNP在免疫和血管系统中也作为局部化学介质起作用。我们也一直在寻找可能与循环控制有关的肽，使用一种监测血小板cAMP升高活性的测定系统。在本研究中，我们从人嗜铬细胞瘤提取物中发现了一种新的降压肽。由于这种肽在正常肾上腺髓质以及由肾上腺髓质产生的嗜铬细胞瘤组织中含量丰富，因此被称为“肾上腺髓质素”。该肽由52个氨基酸组成，具有1个分子内二硫键，与降钙素基因相关肽具有轻微的同源性。在麻醉的大鼠中发现它能产生持久有效的降压作用。由于嗜铬细胞瘤组织中肾上腺髓质素含量丰富，我们构建了嗜铬细胞瘤cDNA文库，分离并测序了编码肾上腺髓质素前体的cDNA克隆。人肾上腺髓质素前体（人肾上腺髓质素前体）的长度为185个氨基酸，包括一个肾上腺髓质素序列，然后是甘氨酸（酰胺供体）。原肾上腺髓质素（proAM）含有独特的20个氨基酸序列，其后是N端Gly-Lys-Arg。有可能由前体加工出一种新的20残基肽“肾上腺髓质素N-末端20肽”（proAM-N20），其羧基端经过修饰。通过RNA印迹分析，发现人肾上腺髓质mRNA在肾上腺髓质、脑室、肺、肾及嗜铬细胞瘤等组织中均有高表达。肾上腺髓质素广泛分布于多种组织中，并以相当高的浓度在血液中循环。这些数据提示肾上腺髓质素是一种参与循环控制的新激素。肾上腺髓质素的出现表明可能存在一种新的循环控制系统。少

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 … More 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