© 1998 by European Society of Cardiology
Copyright © 1998, European Society of Cardiology
The discovery of endothelins
National Cardiovascular Center, Fujishirodai 5-7-1, Suita, Osaka 565, Japan
Received 18 March 1998; accepted 13 May 1998
It is generally accepted that the vascular endothelium is not a simple barrier between the blood stream and vascular bed. It has many functions in the regulation of vascular function that are mediated by various factors released from the endothelium which act on itself or other vascular cells in autocrine and paracrine manners. This concept emerged after the discovery of the endothelium-derived relaxing factor (EDRF) in 1980. The EDRF was later characterized as nitric oxide in 1987. Before the discovery of the EDRF, prostacycline was discovered, which is also a relaxing factor produced by the endothelium. These two relaxing factors are released from endothelial cells in response to a variety of vasoactive substances, and are thought to regulate vascular tone.
In contrast to endothelium-derived relaxing factors, less progress had been made in the field of endothelium-derived constricting factors such as thromboxane, a strong vasoconstrictor which is produced in the endothelium.
In 1985, Hickey et al. reported the existence of a vasoconstricting factor in a conditioned medium of cultured bovine endothelial cells [1]. They suggested that the factor was a peptide. In May 1987, we started purification and identification of this factor. We collected a large amount of conditioned medium of cultured bovine endothelial cells which contained the factor, and were able to quickly isolate the factor and determine the whole sequence of the peptide. We organized an excellent group to promote this project, and Yanagisawa, Kuribara, Goto and Kimura were involved. Kimura is an expert of peptide chemistry, Goto is a pharmacologist, and both Yanagisawa and Kurihara are excellent active investigators. They finished the purification of the factor by the end of July, which was followed by amino acid sequence analysis and cDNA cloning of the peptide. Both experiments were quickly finished, and the results of the amino acid sequence analysis by automatic peptide sequencer and cDNA analysis were the same. The factor was a peptide which consisted of 21 amino acid residues, 2492 daltons in size including four cysteine residues. The four cysteine residues were found to form two intramolecular disulfide bonds. Then, we asked Dr. Fujino of Takeda Pharmaceutical Company to synthesize the peptide. Both the synthesized and native peptides were chemically and pharmacologically the same. We named the peptide ‘endothelin (ET)’ and submitted a report to Nature in October, 1987. It was published in the early spring of 1988 [2]. Fortunately, many new experimental techniques for modern biology were developing at that time such as culture techniques of endothelial cells, high performance liquid chromatography, high performance automatic peptide sequencer and new molecular biological methods. We made use of these techniques.
ET was the most potent vasoconstrictive peptide so far reported and was characterized by its long-lasting action. No amino acid sequence similar to that of ET had been previously reported. Therefore, numerous scientists were interested in the peptide due to its characteristic structural and pharmacological properties. Indeed, discovery of the peptide induced an explosion of worldwide research. Initially, the sustained increase in blood pressure elicited by the peptide appeared to suggest involvement of the peptide in mechanisms of hypertension or maintenance of blood pressure. Recent reports demonstrated that endogenous ET conferred basal constrictor tone of the peripheral vascular bed and played a fundamental physiological role in the maintenance of blood pressure in humans [3]. However, a great number of subsequent reports demonstrated that the mechanisms were not so simple. In addition, numerous reports demonstrated that ET had a variety of pharmacological actions not only in cardiovascular but also in non-cardiovascular systems. Thus, a number of scientists were interested in this peptide. As this fact was found at the initial step of the research, we intended to expedite the progress extensively by distributing synthetic ET to investigators who were interested in the peptide before the publication of the first report. Consequently, we clarified within a very short period its chemical structure, biological activity, isoforms, receptor structure and biosynthetic pathway using various techniques of modern biology.
Following the initial publication of the peptide, analysis of the human ET gene revealed the existence of two other ET-like peptide genes (Fig. 1) [4]. Later it was reported that these peptides were expressed in various tissues and cells. We named them ET-2 and ET-3 in addition to ET-1 which was the first ET found in the conditioned medium of cultured endothelial cells. Endothelial cells predominately produce ET-1. These endogenous ETs are also expressed with different patterns in a wide variety of cell types. ET-1 exists in cardiac myocytes, vascular smooth muscle cells, kidney etc. ET-2 is expressed in the kidney. ET-3 is expressed densely in the intestine and adrenal gland. The results suggest a variety function of ETs in different tissues and organs. In addition, a group of very similar ET-like peptides, the sarafotoxins was reported shortly after the initial discovery of ET. Sarafotoxins are rare snake venoms. Structurally, the sarafotoxins are very similar to ETs and act on the ET receptor. Four sarafotoxins (STXa, STXb, STXc and STXd) which have similar structures to that of ET have been reported (Fig. 1).
|
Sequence analysis of ET cDNA revealed that ET is produced by a precursor, named preproendothelin. After the signal peptide is removed, the precursor is cleaved by an endopeptidase specific for a pair of dibasic amino acids, resulting in an intermediate form, big ET (Fig. 2). Recent studies demonstrated that this protease is a furin-like enzyme and that the cleavage of the precursor by this protease is an essential step before the cleavage of big ET-1. Big ET must be cleaved by an unknown enzyme to produce ET, which we named ET converting enzyme (ECE). Many investigators started to search for this enzyme. Recently, it was purified and the cDNA clone for the enzyme was isolated [5, 6]. It has a similar structure to neutral endopeptidase, and there are several isoforms of the enzyme. The human ECE-1 gene expressed in two different isoforms, named ECE-1
/a and ECE-1 β/b [7]which differ only in the N-terminal amino acid sequences. Another enzyme termed ECE-2 was also identified [8]. Both ECE-1 and ECE-2 cleaved big ET-1 efficiently than either big ET-2 or big ET-3. The former is expressed ubiquitously with highest expression in endothelium, lung, ovary, testis and adrenal medulla, while the latter is expressed in neural tissues. Very recently, existence of the third isoform named ECE-3 was demonstrated in bovine iris, which selectively cleaved big ET-3 [9]. ECE was located at the cell surface and on intracellular vesicles [10]. Recently, several inhibitors specific for the ET converting enzyme have been reported.
|
One of the important discoveries following the early step was the discovery of two ET receptors, ETA and ETB [11, 12]. Both belong to a family of heptahelical G-protein coupled receptors (Table 1). There is 68% amino acid identity between the two receptor subtypes. In the early stage of ET research, a great number of pharmacological studies suggested that the responses to ETs could be divided into two groups according to the pharmacological potency of the three peptides. Indeed, these two receptors, ETA and ETB, are distinct in their ligand binding affinity and distribution in tissues and cells. ETA has a high affinity to ET-1 and ET-2, but a low affinity to ET-3. ETB has equally potent affinities to all three endogenous ETs. ETA exists on smooth muscle and mediates vasoconstriction. In contrast, ETB exists on endothelium and mediates the release of relaxing factors such as nitric oxide and prostacycline. However, several reports demonstrated that ETB on some vascular smooth muscle also mediated vasoconstriction.
|
Cloning of the ET receptor facilitated the development of ET-receptor antagonists. In 1991, a fermentation product of BE-18257B was recognized as an antagonist for the ET-receptor, which was followed by synthesis of BQ-123 and FR139317, two derivatives of BE-18257B, at the initial step of development of ET antagonists. Then, many selective and non-selective antagonists for ETA and ETB emerged.
Initially, this peptide aroused interest of many cardiologists due to its unique actions on the cardiovascular system. However, the physiological and pathophysiological significance of ET is still unclear. To elucidate the mechanism, ET receptor antagonists are helpful. Accumulating evidence demonstrates that ET plays an important pathophysiological role in many disorders. At the fifth International Conference on ET held in Kyoto in 1997, a large part of the discussion was devoted to the use of antagonists. ET antagonists demonstrated significant beneficial effects in pathological conditions, including congestive heart failure, pulmonary hypertension, cerebrovascular spasm after subarachnoid haemorrhage, acute renal failure and essential hypertension. Recently several kinds of ET or ET receptor knockout mice were produced. Observation of these mice has provided us with important information regarding the physiological and pathophysiological significance of ET. ET-1-deficient mice had craniofacial and cardiac abnormalities at birth and die of respiratory failure soon after the birth [13]. In contrast, ET-3 or ETB receptor-deficient mice exhibited aganglionic megacolon associated with coat color spotting, resembling a hereditary syndrome of humans: i.e. Hirschsprung's disease [14]. In addition to those unexpected results, those knockout mice gave us important knowledge of cardiovascular actions of ETs. It is interesting that adult ETB-deficient mice and rats exhibit significantly elevated blood pressure under healthy, baseline conditions. In these mice, the hypertension was shown to be salt-sensitive and resistant to ETA blockade, suggesting the function of the ETB receptor as a physiologically relevant natriuretic receptor in the kidney.
Thus, the discovery of ET has created a new scientific field. A large amount of information has been accumulated during the past decade. Now, it is generally recognized that ET induces both beneficial and harmful effects in the living body. Physiological roles of ET will be clarified in the near future, as well as antagonists for treatment of several disorders.
At the last International Conference on ET, about 300 presentations were given. This field is still growing. It is interesting that questions raised at the initial steps of ET research still continue to be valid in this field ten years after the discovery.
Time for primary review 30 days
 |
References |
|---|
 Top References |
|---|
- Hickey K.A., Rubanyi G., Paul R.J., Highemith R.F. Characterization of a coronary vasoconstrictor produced by cultured endothelial cells. Am J Physiol (1985) 248:C550.[Web of Science][Medline]
- Yanagisawa M., Kurihara H., Kimura S., et al. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature (1988) 332:411.[CrossRef][Medline]
- Haynes W.H., Ferro O.J., O'kane K.P.J. Systemic endothelin receptor blockade decreases peripheral vascular resistance and blood pressure in humans. Circulation (1996) 93:1860.
[Abstract/Free Full Text] - Inoue A., Yanagisawa M., Kimura S., et al. The human endothelin family: three structurally and pharmacologically distinct isopeptides predicted by three separate genes. Proc Natl Acad Sci USA (1989) 86:2863.
[Abstract/Free Full Text] - Shimada K., Takahashi M., Tanzawa K. Cloning and functional expression of endothelin-converting enzyme from rat endothelial cells. J Biol Chem (1994) 269:18274–18278.
- Xu D., Emoto N., Giaid A. ECE-1: A membrane-bound metalloprotease that catalyzes the proteolytic activation of big endothelin-1. Cell (1994) 78:473–485.[CrossRef][Web of Science][Medline]
- Shimada K., Takahashi M., Ikeda M., Tanzawa K. Identification and characterization of two isoforms of an endothelin-converting enzyme-1. FEBS Lett (1995) 371:140–144.[CrossRef][Web of Science][Medline]
- Emoto N., Yanagisawa M. Endothelin-converting enzyme-2 is a membrane-bound phosphoramidon-sensitive metalloprotease with acidic pH optimum. J Biol Chem (1995) 279:16262–16268.
- Hasegawa H., Hiki K., Sawamura T., et al. Purification of a novel endothelin-converting enzyme specific for big endothelin-3. FEBS Lett (1998) 428:304–308.[CrossRef][Web of Science][Medline]
- Barnes K., Brown C., Turner A.J. Endothelin-converting enzyme: Ultrastructural localization and its recycling from the cell surface. Hypertension (1998) 31:3–9.
[Abstract/Free Full Text] - Arai H., Hori S., Aramori I., Ohkubo H., Nakanishi S. Cloning and expression of a cDNA encoding an endothelin receptor. Nature (1990) 348:730.[CrossRef][Medline]
- Sakumi T., Yanagisawa M., Takuwa Y., et al. Cloning of a cDNA encoding a non-isopeptide-selective subtype of the endothelin receptor. Nature (1990) 348:782.
- Kurihara Y., Kurihara H., Suzuki H., et al. Elevated blood pressure and craniofacial abnormalities in mice deficient in endothelin-1. Nature (1994) 368:703–710.[CrossRef][Medline]
- Puffenberger E.G., Hosoda K., Washington S.S., et al. A missense mutation of the endothelin-B-receptor gene in multigenic Hirschspring's disease. Cell (1994) 79:1257–1266.[CrossRef][Web of Science][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  P. J. Troy and A. B. Waxman Review: Portopulmonary hypertension: challenges in diagnosis and management Therapeutic Advances in Gastroenterology, September 1, 2009; 2(5): 281 - 286. [Abstract] [PDF]
|
|
|
|
 |
|
 W. M Maharsy, L. N Kadi, N. G Issa, K. M Bitar, A. H Der-Boghossian, R. Abrahamian, and A. B Bikhazi Cross-talk related to insulin and angiotensin II binding on myocardial remodelling in diabetic rat hearts Journal of Renin-Angiotensin-Aldosterone System, June 1, 2007; 8(2): 59 - 65. [Abstract] [PDF]
|
|
|
|
 |
|
 J. Y. C. Tsang, W. J. E. Lamm, B. Neradilek, N. L. Polissar, and M. P. Hlastala Endothelin receptor blockade does not improve hypoxemia following acute pulmonary thromboembolism J Appl Physiol, February 1, 2007; 102(2): 762 - 771. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. A. Sarafidis and G. L. Bakris Insulin and Endothelin: An Interplay Contributing to Hypertension Development? J. Clin. Endocrinol. Metab., February 1, 2007; 92(2): 379 - 385. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. S. Hassan, T. M. Williams, P. G. Frank, and M. P. Lisanti Caveolin-1-deficient aortic smooth muscle cells show cell autonomous abnormalities in proliferation, migration, and endothelin-based signal transduction Am J Physiol Heart Circ Physiol, June 1, 2006; 290(6): H2393 - H2401. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. S. Hassan, S. Mukherjee, F. Nagajyothi, L. M. Weiss, S. B. Petkova, C. J. de Almeida, H. Huang, M. S. Desruisseaux, B. Bouzahzah, R. G. Pestell, et al. Trypanosoma cruzi Infection Induces Proliferation of Vascular Smooth Muscle Cells Infect. Immun., January 1, 2006; 74(1): 152 - 159. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. N. Channick, O. Sitbon, R. J. Barst, A. Manes, and L. J. Rubin Endothelin receptor antagonists in pulmonary arterial hypertension J. Am. Coll. Cardiol., June 16, 2004; 43(12_Suppl_S): 62S - 67S. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Galie, A. Manes, and A. Branzi The endothelin system in pulmonary arterial hypertension Cardiovasc Res, February 1, 2004; 61(2): 227 - 237. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. H.S. Kim and L. J. Rubin Endothelin in Health and Disease: Endothelin Receptor Antagonists in the Management of Pulmonary Artery Hypertension Journal of Cardiovascular Pharmacology and Therapeutics, March 1, 2002; 7(1): 9 - 19. [Abstract] [PDF]
|
|
|
|
 |
|
 T. F. Luscher and M. Barton Endothelins and Endothelin Receptor Antagonists : Therapeutic Considerations for a Novel Class of Cardiovascular Drugs Circulation, November 7, 2000; 102(19): 2434 - 2440. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Barnes and A. J Turner Endothelin converting enzyme is located on {alpha}-actin filaments in smooth muscle cells Cardiovasc Res, June 1, 1999; 42(3): 814 - 822. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||