Angiotensin II formation in dog heart is mediated by different pathways in vivo and in vitro

Abstract

Angiotensin-converting enzyme (ACE) inhibitors (I) have beneficial effects that are presumably mediated by decreased angiotensin II (ANG II) production. However, in vitro assays in human heart extracts have demonstrated that > 75% of ANG II-forming enzyme activity was not inhibited by captopril (Cap) and therefore did not appear to be related to ACE but was inhibited by chymostatin, suggesting that it was predominantly chymase-like activity. Previous work in our laboratory has demonstrated a similar relative contribution of ACE and chymase-like activity toward ANG II formation in vitro in dog heart tissue extracts. Accordingly, we compared Cap-inhibitable ANG II formation in vitro in heart tissue of five adult mongrel dogs to the in vivo Cap-inhibitable, ANG II-forming activity across the myocardial bed in four openchest, adult mongrel dogs. In vitro studies demonstrated that only 6 +/- 2% of ANG II formation was inhibited by Cap from heart tissue extracts of the left ventricular midwall. In in vivo studies, ANG I (0.5 nmol/min) followed by ANG I plus the ACE inhibitor Cap (0.1 mumol/min) was infused into the left anterior descending artery, and ANG II was assayed in the proximal aorta and coronary sinus. The arterial-venous (A-V) difference of ANG II across the myocardial circulation increased significantly during ANG I infusion (-13.4 +/- 23.5 to 142.8 +/- 71.4 pg/ml; P < 0.03). Subsequent coinfusion of Cap with ANG I significantly decreased the myocardial A-V difference of ANG II by 60 +/- 18% (P < 0.05). Thus, in contrast to the in vitro situation, ANG II formation in vivo is inhibited significantly by Cap in the normal dog heart. This comparison of in vivo and in vitro conversion of ANG I to ANG II by ACE and chymase-like activity suggests that in vitro assays may underestimate the functional contribution of ACE to intracardiac ANG II formation.

Biochemical evaluation of the renin-angiotensin system: the good, bad, and absolute?

“…Danser and colleagues (113,139) suggest that urinary renin may be an alternative measure of an active RAS within the kidney. Quenched fluorescent substrates based on the ANG- (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14) sequence are available to measure renin. Although this approach does not rely on the detection of ANG I or an exogenous source of angiotensinogen, the sensitivity and specificity of the peptide substrates are not comparable with angiotensinogen.…”

Evidence for Angiotensin-Converting Enzyme– and Chymase-Mediated Angiotensin II Formation in the Interstitial Fluid Space of the Dog Heart In Vivo

“…4,8 We hypothesized that because chymase is present in the interstitium, chymase-mediated Ang II formation catalyzed by chymase should be substantial in the ISF space of the dog heart in vivo. Thus, our intent was to reproduce our in vitro assay conditions established in heart tissue extracts by infusion of Ang I substrate alone and Ang I plus inhibitors into the ISF space of the dog heart in vivo.…”

“…We chose 15 mol/L Ang I as the initial dose to infuse into the microdialysis probes, which was 40-fold lower than the 600 mol/L Ang I dose used in our in vitro experiments. 4,8 Our choices for doses of captopril and chymostatin ISF infusions were also based on our in vitro studies. 4,8 For the in vivo experiments, we chose chymostatin 1 mmol/L and captopril 2.5 mmol/L, 10 and 25 times the in vitro doses of chymostatin and captopril, respectively.…”

“…3 Studies in dog and human heart have demonstrated that chymase accounts for Ͼ90% of Ang II formation in heart tissue extracts in vitro, 4 -6 whereas ACE is responsible for Ͼ80% of Ang II formation across the coronary vascular bed in vivo. 7,8 This difference in intracardiac Ang II formation in vitro and in vivo may be related to the distribution and compartmentalization of chymase and ACE in the heart. In situ hybridization and electron microscope-immunocytochemical studies in the human heart have demonstrated that mast cells and other interstitial cell types synthesize and store chymase and that active enzyme is present in the extracellular matrix of the myocardium and vessel walls.…”

Role For Chymase in Heart Failure

“…In striking contrast, experiments in vivo with normal 5,6 or failing dog hearts 7,8 have demonstrated that most (Ͼ70%) of the Ang II formation can be inhibited by an ACE inhibitor, indicating that, under these conditions, the major Ang IIforming enzyme is ACE. Moreover, in normal human hearts, Zisman et al 9 have demonstrated that, in vivo, the major Ang II-forming enzyme is ACE, as 90% of the Ang II formation could be inhibited by an ACE inhibitor.…”