Detection by bioassay and specific enzyme-linked immunosorbent assay of phosphoramidon-inhibitable endothelin-converting activity in brain and endothelium.
The endothelin-converting enzyme (ECE) activity present in endothelial cells and rat and human brains was characterized using a selective and rapid bioassay for endothelin-1 (ET-1) or endothelin-3 (ET-3) together with a sensitive enzyme-linked immunosorbent assay. We found that ECE activity was predominantly in the membrane fraction of endothelial cells from which it could be extracted by treatment with detergent. In rat brain tissue, the ECE activity was in the membrane fraction and was not solubilized by detergent treatment. Further dissection of the brain revealed that there was a strong localization of ECE activity in the hypothalamus, midbrain, and medulla oblongata in agreement with earlier observations of ET-like immunoreactivity and binding sites. Experiments with human brain tissue also showed the presence of ECE activity. In conclusion, our studies confirmed the presence of ECE activity within endothelial cells, and showed ECE to be localized in brain tissue in sites consistent with the selective distribution of the ET-1 synthetic pathway. In all tissues studied, the ECE activity was significantly inhibited by phosphoramidon or ethylenediaminetetra-acetate.
D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism*; D001681:Biological Assay; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D019332:Endothelin-1; D000072396:Endothelin-Converting Enzymes; D016232:Endothelins / Q000378:metabolism / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000201:enzymology*; D004797:Enzyme-Linked Immunosorbent Assay; D006020:Glycopeptides / Q000494:pharmacology*; D007031:Hypothalamus / Q000201:enzymology; D008526:Medulla Oblongata / Q000201:enzymology; D008636:Mesencephalon / Q000201:enzymology; D008666:Metalloendopeptidases; D015260:Neprilysin / Q000037:antagonists & inhibitors*; D011498:Protein Precursors / Q000378:metabolism
D019332:Endothelin-1; D016232:Endothelins; D006020:Glycopeptides; D011498:Protein Precursors; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; D015260:Neprilysin; D000072396:Endothelin-Converting Enzymes; D006152:Cyclic GMP; C008890:phosphoramidon