The Development of a Vascular Renin Extraction Procedure and Method of Radioimmunoassay to Measure Changes in Vascular Renin-Like Activity following DOCA Administration in the Pig.

Abstract

Methods for a tissue renin extraction and indirect RIA for angiotensin I were developed and evaluated to maximize the renin-like activity in the pig aorta. Miniature Hormel pigs were administered DOCA for periods of up to six weeks to decrease PRA. The change in VRA before and after DOCA implantation was measured by the modified extraction and RIA procedures. Vascular renin was concentrated and partially purified in an extraction process designed to eliminate destructive angiotensinases. This preparation of vascular renin permitted the determination of its activity over a 24 hour incubation interval. The increased generation of Al over 24 hours facilitated its detection by the RIA. An evaluation of inhibitors, substrate concentrations, and incubation times permitted maximization of renin-like activity for the RIA. The highest renin-like activity was demonstrated when extract samples were incubated in the presence of 8-0H/Na2-FDTA inhibitors and 44% renin substrate volume (maximum allowable volume under assay conditions) up to 24 hours. An incubation temperature of 37°C was employed and a pH of 7.4 to minimize Al generation by other proteolytic enzymes. The results demonstrated a parallel change between the PRA and the VRA in the thoracic, renal, and iliac segments; the renal segment showing the highest VRA.