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Endothelin-Converting Enzyme Activity in Vascular Smooth Muscle Preparations In Vitro

  • Janet J. Maguire
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 206)

Abstract

The synthesis of ET-1, by the specific cleavage of its larger precursor big ET-1, by one or more endothelin-converting enzymes (ECE) has been discussed in Chapter 7. In the human vasculature ET-1, big ET-1 and ECE-1 are localized to secretory and storage granules (1) within endothelial cells (2). ET-1 is released via the constitutive and stimulated pathways, together with big ET-1 (3), and it is possible that additional conversion of this released big ET-1 occurs on the surface of the underlying smooth muscle cells. This is predicted by the observation that big ET-1, infused into the human forearm, significantly increases plasma levels of immunoreactive ET and decreases forearm blood flow in a phosphoramidon-sensitive manner. The hemodynamic response to big ET-1 occurred too quickly for appreciable amounts of phosphoramidon to have penetrated cell membranes, suggesting that the ECE responsible for conversion of infused big ET-1 is probably an ectoenzyme. As endothelial ECE has a predominantly intracellular localization (1), expression of ECE on the surface of human smooth muscle cells may account for the rapid response to exogenous big ET-1. This is consistent with reports that, in isolated vascular preparations, removal of the endothelium does not alter responses to big ET-1 (5-8) implying the presence of a nonendothelial, presumably smooth muscle enzyme. Indeed, ECE activity has been reported in cultured vascular smooth muscle cells (9,10) and one isoform of ECE-1, ECE-1b, has been localized to the smooth muscle cell plasma membrane (11).

Keywords

Fumaric Acid Human Coronary Artery Bathing Medium Penetrate Cell Membrane Human Smooth Muscle Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
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Copyright information

© Humana Press Inc. 2002

Authors and Affiliations

  • Janet J. Maguire
    • 1
  1. 1.Clinical Pharmacology UnitUniversity of CambridgeCambridgeUK

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