succinyl-trialanine-4-nitroanilide and Arteriosclerosis

To elucidate the nature of the abnormality of elastin metabolism in arteriosclerosis, we determined the elastase-type activity in the human radial artery of patients with chronic renal failure due to glomerular disease and diabetes. Elastase-type activity was determined by HPLC analysis of the hydrolyzed products of succinyl-trialanine-4-nitroanilide, a sensitive synthetic substrate for elastase. Three kinds of hydrolyzed products, (L-Ala)2-NA, L-Ala-NA and NA, were found after incubation of the substrate with human radial artery in the presence of amastatin (an inhibitor of aminopeptidases). We assumed the activity that liberates NA to be an elastase-type activity because purified human aorta elastase liberates NA from the substrate. The pH optima of the human artery and rat aorta activities were 6.0 and 6.8, respectively. The elastase activity in human radial artery and rat aorta was inhibited by diisopropyl phosphofluoridate, a serine protease inhibitor, and by elastatinal, an elastase inhibitor. The elastase-type activity in the radial artery of patients with chronic renal failure was significantly lower than that of the control group, and the decrease was especially marked in the patients with juvenile onset diabetes. These results suggest that the elastin metabolism is abnormal in the radial artery in diseases that tend to cause atherosclerosis.

Topics: Adult; Age Factors; Aged; Animals; Anti-Bacterial Agents; Aorta; Arteries; Arteriosclerosis; Chromatography, High Pressure Liquid; Female; Forearm; Humans; Kidney Failure, Chronic; Kinetics; Male; Middle Aged; Oligopeptides; Pancreatic Elastase; Peptides; Rats; Rats, Inbred Strains

Human serum was found to contain enzyme activities hydrolyzing succinyl trialanine paranitroanilide and 3H-kappa-elastin Sepharose substrates. Both types of activities could be partly abolished by serine active site titrants (phenylmethanesulfonylfluoride, diisopropylphosphorofluoridate) and partly by neutral chelating agents (EDTA; 1-10-phenanthroline). The combination of phenylmethanesulfonylfluoride and EDTA gave a complete inhibition of human serum elastase-type activities indicating the presence of at least two different types of elastases (serine and metalloproteases) in human serum. In nonsmokers, the average serum elastase-type activity on succinyl trialanine paranitroanilide was found equal to 78.1 ng/ml porcine pancreatic elastase equivalents and on 3H-kappa-elastin sepharose beads equal to 688.8 ng/ml. No statistically significant differences were observed in elastase levels in the sera of individuals presenting clinical symptoms of atherosclerosis. The sera of patients suffering from chronic obstructive lung diseases contained, however, higher amounts of elastase-type activities, respectively equal to 237.2 ng/ml on succinyl trialanine paranitroanilide and 1,096 ng/ml on 3H-kappa-elastin Sepharose beads and was quantitatively significant when compared with control subjects.

Topics: Adult; Aged; Arteriosclerosis; Elastin; Humans; Lung Diseases, Obstructive; Middle Aged; Oligopeptides; Pancreatic Elastase; Sepharose; Substrate Specificity