The half-life of endothelium-derived relaxing factor released from bovine aortic endothelial cells in culture

Abstract

1. The half-life of endothelium-derived relaxing factor (EDRF) in Krebs solution was determined by bioassay in vitro. 2. A column of bovine aortic endothelial cells grown on microcarrier beads in suspension culture was perfused with Krebs solution. EDRF was released from these cells by sequential treatment with increasing concentrations of bradykinin (0.01-100 nM). EDRF was detected by the relaxation of an endothelium-denuded ring segment of dog coronary artery. 3. Complete bradykinin concentration-relaxation curves were determined in the absence or presence of coils of tubing that increased the transit time (delay) between the cell column and the assay tissue. An estimate of the falls in concentration, and hence of the half-life of EDRF, was obtained from the shift of the bradykinin concentration-relaxation curves. 4. Mass-action equations were used to model the relationship between the indirectly acting agonist bradykinin and the relaxation via EDRF. The modelling adequately predicted the consequences of different transit delay times (0-4 half-lives). 5. This new analysis of half-life of an active intermediate emphasizes the measurement of changes in concentration with increasing transit time rather than a fall in tissue response. 6. The half-life of EDRF in Krebs solution is 41 s.