Enhanced expressions of endothelin-converting enzyme and endothelin receptors in human colonic tissues of Crohn's disease.

Endothelin-1, a powerful vasoconstrictor, forms the endothelin system together with endothelin-converting enzyme and endothelin type A and type B receptors. These endothelin system components are considered to participate in inflammatory and wound healing responses. Previous reports have suggested a role for the endothelin-1 in the pathology of Crohn's disease. In the present study, we immunohistochemically investigated the expressions of the endothelin system components in affected human intestinal tissues of Crohn's disease. Eighteen surgical specimens of colonic tissue obtained from patients with Crohn's disease and 12 normal colonic tissues as controls were examined. Frozen tissue sections cut from the samples were subjected to the immunohistochemical single and double staining. The endothelin system components were expressed mainly in the muscular layers and blood vessels. In diseased colonic tissues, inflammatory infiltration and fibrotic tissue reactions with marked smooth muscle cell proliferation were frequently seen, and were closely associated with increased expressions of the endothelin system components. These results strongly suggest that endothelin-converting enzyme and endothelin type A and type B receptors collectively play a role in the inflammatory and fibrogenic processes of Crohn's disease. Especially, submucosal smooth muscle proliferation, a histological hallmark of strictures, may be attributable to the upregulated endothelin system.

Expression of endothelin-converting enzyme, endothelin-1 and endothelin receptors at the site of percutaneous coronary intervention in humans.

OBJECTIVE: The repair process at the site of injury after percutaneous coronary intervention (PCI) is dominated by neointimal formation composed mainly of smooth muscle cells (SMC). Endothelin-1 (ET-1) is a powerful vasoconstrictor and SMC mitogen. Endothelin-converting enzyme (ECE) is the final key enzyme of endothelin processing. The effects of ET-1 are mediated by binding to endothelin type A (ETA) and endothelin type B (ETB) receptors. The ligand/receptor/ligand-producing system (ET system) could be involved in the pathogenesis of neointimal formation in humans. METHODS: Fifteen post-PCI sites obtained at autopsy and eight atherectomy specimens obtained from restenotic sites were investigated using immunohistochemical single and double staining techniques. Frozen sections were stained with antibodies against ECE, ET-1, ETA and ETB receptors, SMC, macrophages and endothelial cells. RESULTS: At the early stage, less than 3 months after PCI, neointimal SMC were positive for ECE, ET-1, ETA and ETB receptors. The expression of ECE, ET-1, ETA and ETB receptors in these neointimal SMC decreased markedly from 6 months onwards. The ECE, ET-1, ETA and ETB receptor-positive cell areas were significantly (P < 0.005) greater in the first 3 months after PCI compared with 6 months or more after PCI. Atherectomy specimens also showed similar positivity. CONCLUSIONS: These observations strongly suggest that the expression of ECE, ET-1, ETA and ETB receptors is enhanced in neointimal SMC at early stages after PCI injury in human coronary arteries. The increased expression of the ET system may contribute to SMC proliferation/migration and vasoconstriction in human post-PCI coronary lesions.

Expression of the hepatic endothelin system in human cirrhotic livers.

It is considered that endothelin-1 participates in the development of liver cirrhosis and it has been recognized that every component of the endothelin system is upregulated in cirrhotic livers. However, the expression pattern of this system, including interaction between its components, is not fully understood in human livers. In this study, the expression pattern of the endothelin system was examined. Immunohistochemical analysis for endothelin-1, endothelin receptors and endothelin-converting enzyme was performed in 16 cirrhotic and 17 normal human liver tissues. Peptides, proteins, and RNAs extracted from the livers were also investigated using quantitative assays for the components of the hepatic endothelin system. Hepatic endothelin-1 levels were significantly higher in cirrhotic livers (0.084 +/- 0.052 pg/mg wet liver) than in normal livers (0.041 +/- 0.032 pg/mg; p < 0.01), and were closely related to the severity of liver fibrosis and portal hypertension. Immunoreactivity for endothelin-1, endothelin receptors, and endothelin-converting enzyme was detected mainly in fibrous areas and in the hepatic vasculature, and was enhanced in cirrhosis. Although there was a negative correlation between the expression of receptor mRNA and the hepatic endothelin-1 level, the amounts of the mRNAs were greater in cirrhotic livers than in normal livers. However, expression of endothelin-converting enzyme in cirrhotic livers was increased at the protein level but was relatively reduced at the mRNA level. These findings suggest that the hepatic endothelin system is activated in human cirrhotic livers in association with worsening of the disease, but that the regulation of the components of this system in this disorder is complex.

Alterations of endothelin-converting enzyme expression in early and advanced stages of human coronary atherosclerosis.

Endothelin-1 is a potent vasoconstrictor and exhibits a mitogenic activity on vascular smooth muscle cells (SMCs). Endothelin-converting enzyme (ECE) is the final key enzyme of endothelin-1 processing. We studied the immunolocalization of ECE in human coronary atherosclerotic lesions with different disease stages. Frozen sections of normal coronary arteries with diffuse intimal thickening (n=13) and those of coronary arteries with early (n=10) or advanced atherosclerotic plaques (n=13) were studied. Monoclonal antibodies used were directed against SMCs, macrophages, endothelial cells, and ECE. For the identification of cell types that express ECE, double immunostaining analysis was also used. In normal coronary arteries, ECE immunoreactivity was observed in luminal endothelial cells and medial SMCs. Early atherosclerotic plaques, which consisted predominantly of SMCs, showed enhanced ECE expression in luminal endothelial cells and intimal SMCs. In advanced atherosclerotic plaques, distinct ECE expression was found in accumulated macrophages and in endothelial cells of intraplaque microvessels, while luminal endothelial cells showed relatively weak immunoreactivity for ECE. In conclusion, the present study demonstrates that the major cell types expressing ECE within the plaques are different between early and advanced stages of human coronary atherosclerosis. Enhanced ECE expression and possible endothelin-1 generation may contribute to SMC proliferation and vasoconstriction in early atherosclerotic stages, and may promote plaque destabilization in advanced atherosclerotic stages.

Regulation of endothelin-converting enzyme 1 in nephrotic syndrome in rats.

BACKGROUND: Nephrotic syndrome is characterized by severe proteinuria and sodium and water retention. Although endothelin (ET) 1 can cause natriuresis or antinatriuresis, the role played by ET-1 in proteinuria and in sodium retention due to nephrotic syndrome remains unclear. METHODS: We investigated the role played by the ET-1 system in sodium and water retention and in proteinuria in puromycin aminonucleoside induced nephrotic syndrome in rats using microdissected nephron segments, competitive polymerase chain reaction, and Western blot. RESULTS: The expression of prepro ET-1, ET-converting enzyme 1 (ECE-1), and ET A receptor mRNAs, but not ET B receptor mRNA, in the glomeruli was increased in rats with nephrotic syndrome. The cGMP generation in the glomeruli induced by atrial natriuretic peptide and ET-1 was decreased, whereas the ET-3-induced cGMP generation was increased in rats with nephrotic syndrome. ECE-1 mRNA expression was increased not only in the glomeruli, but also in the thick ascending limbs and collecting ducts. The protein expression of ECE-1 was increased in the membrane fraction of the cortex and in the outer and the inner medulla of nephrotic rats. Blockade of ET A and B receptors by bosentan did not inhibit the occurrence of nephrotic syndrome. However, the administration of bosentan increased the urinary sodium excretion. CONCLUSION: These data suggest that an activated ET-1-ET A receptor pathway in glomeruli and/or an increased ECE-1 mRNA expression in distal segments may participate in sodium and water retention, but not in the occurrence of nephrotic syndrome.

Ontogeny of endothelin-converting enzyme gene expression and protein content in the ovine fetal lung.

Endothelin 1 (ET-1) is a potent vasoconstrictor and smooth muscle comitogen that is produced by endothelin-converting enzyme 1 (ECE-1) in endothelium and smooth muscle of the vascular wall in adult pulmonary arteries. However, little is known about the role of ECE-1 in the fetal pulmonary circulation. The ET-1 protein content falls just prior to birth in the fetal lamb lung. We hypothesized that the expression of ECE-1 is developmentally regulated and that ECE-1 levels fall prior to birth. To test this hypothesis, we measured lung ECE-1 mRNA levels and protein content and determined the cellular localization of ECE-1 expression in lung tissues from fetal lambs between 70 and 140 days of gestation (term = 145 days), newborn lambs, and ewes. We found that ECE-1 mRNA expression and protein content were lower in the fetal lamb lung just prior to birth in comparison with the newborn lamb lung. Immunolocalization of ECE-1 protein showed expression of ECE-1 in the vascular endothelium but not in the vascular smooth muscle at all gestational ages. We conclude that ECE-1 is developmentally regulated and that ECE-1 is expressed in the vascular endothelium but not in the smooth muscle of the fetal pulmonary vasculature. We speculate that alterations in ECE-1 contribute to the changes in ET-1 levels during the perinatal period and that the majority of ET-1 produced in the fetal lamb pulmonary vasculature is produced by the vascular endothelium.