chymostatin and hippuryl-histidyl-leucine

Dog, monkey and human aortic tissues contained two distinct types of angiotensin II-generating enzymes; angiotensin converting enzyme (ACE) and chymostatin-sensitive angiotensin II-generating enzyme (CAGE). Endothelium, media and adventitia of canine thoracic aortae were separated using collagenase digestion, and determined for their ACE and CAGE activity. ACE activity was assayed by hippuryl-His-Leu cleavage. CAGE activity was estimated with ANG I as substrate in the presence of inhibitors of ACE and angiotensinases. His-Leu, the common product of both enzyme reactions, was fluorimetrically quantified after o-phthalaldehyde condensation. ACE localized mainly in endothelium, while CAGE distributed predominantly in adventitia. Similar results were obtained with human and monkey aortae. Such a contrasting distribution may indicate the distinct functional role of these two enzymes.

Topics: Aorta; Connective Tissue; Elastic Tissue; Endopeptidases; Endothelium, Vascular; Oligopeptides; Peptidyl-Dipeptidase A; Serine Endopeptidases; Serine Proteinase Inhibitors

An inhibitor of angiotensin I (ANG I) converting enzyme, SA446, reduced the response to ANG I of blood vessels isolated from dogs and monkeys, but did not abolish the response even at high concentrations. The residual action of ANG I in the presence of high concentrations of SA446 could be abolished by (Sar1, Ala8)-ANG II. Vascular strips and crude extracts of vessels and lungs possessed the enzymic activity generating ANG II from ANG I, or hippuric acid from hippuryl-histidyl-leucine (HHL). The HHL-hydrolysing activity of the crude extracts was completely inhibited by SA446 (10(-7) mol/l) and/or Na2-EDTA (10(-3) mol/l). However, the octapeptide generation was not abolished despite the combined treatment with SA446 (5 X 10(-4) mol/l) and Na2-EDTA (5 x 10(-3) mol/l). The residual activity forming ANG II was inhibited by chymostatin and soybean trypsin inhibitor, which however did not affect the HHL-hydrolysis. Combined treatment with SA446 (10(-5) mol/l) and chymostatin (2.5 X 10(-5) mol/l) abolished the vascular action of ANG I but did not alter the action of ANG II. These results strongly suggest that besides the ANG I converting enzyme, another enzyme which generates ANG II is present in vascular tissues and lungs, and may play an important role in the local generation of ANG II, which possibly regulates the regional vascular tone.

Topics: 3-Mercaptopropionic Acid; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Angiotensins; Animals; Blood Vessels; Dogs; Female; In Vitro Techniques; Macaca; Male; Muscle Contraction; Muscle, Smooth, Vascular; Oligopeptides; Sulfhydryl Compounds; Thiazolidines