Chymase-Dependent Generation of Angiotensin II from Angiotensin-(1-12) in Human Atrial Tissue

Abstract

Since angiotensin-(1-12) [Ang-(1-12)] is a non-renin dependent alternate precursor for the generation of cardiac Ang peptides in rat tissue, we investigated the metabolism of Ang-(1-12) by plasma membranes (PM) isolated from human atrial appendage tissue from nine patients undergoing cardiac surgery for primary control of atrial fibrillation (MAZE surgical procedure). PM was incubated with highly purified 125I-Ang-(1-12) at 37°C for 1 h with or without renin-angiotensin system (RAS) inhibitors [lisinopril for angiotensin converting enzyme (ACE), SCH39370 for neprilysin (NEP), MLN-4760 for ACE2 and chymostatin for chymase; 50 µM each]. 125I-Ang peptide fractions were identified by HPLC coupled to an inline γ-detector. In the absence of all RAS inhibitor, 125I-Ang-(1-12) was converted into Ang I (2±2%), Ang II (69±21%), Ang-(1-7) (5±2%), and Ang-(1-4) (2±1%). In the absence of all RAS inhibitor, only 22±10% of 125I-Ang-(1-12) was unmetabolized, whereas, in the presence of the all RAS inhibitors, 98±7% of 125I-Ang-(1-12) remained intact. The relative contribution of selective inhibition of ACE and chymase enzyme showed that 125I-Ang-(1-12) was primarily converted into Ang II (65±18%) by chymase while its hydrolysis into Ang II by ACE was significantly lower or undetectable. The activity of individual enzyme was calculated based on the amount of Ang II formation. These results showed very high chymase-mediated Ang II formation (28±3.1 fmol×min−1×mg−1, n = 9) from 125I-Ang-(1-12) and very low or undetectable Ang II formation by ACE (1.1±0.2 fmol×min−1×mg−1). Paralleling these findings, these tissues showed significant content of chymase protein that by immunocytochemistry were primarily localized in atrial cardiac myocytes. In conclusion, we demonstrate for the first time in human cardiac tissue a dominant role of cardiac chymase in the formation of Ang II from Ang-(1-12).

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.…”

“…Chymases comprise a family of serine peptidases that may form ANG II by hydrolysis of the Phe (3,27,28,89,138). Humans express ␣-chymase, whereas rodents express primarily ␤-chymases, as well as other isoforms (mouse mast cell protease-4 and rat mast cell protease-5) that more closely resemble ␣-chymase in regard to the processing of ANG I to ANG II (27,28,126).…”

“…The ANG II metabolite ANG- (3)(4)(5)(6)(7)(8) or ANG IV has distinct functional properties through the interaction with the insulin-regulated aminopeptidase, although the peptide also stimulates the AT 1 R (4, 80, 150). ANG-(1-9), a potential product of ACE2 processing of ANG I, appears to functionally interact with the AT 2 receptor (101).…”

Involvement of the Renin‐Angiotensin System in Atherosclerosis

“…They are participating in the progression of disease, or contrary, in mechanisms that protect from tissue injury [12]. These components include the (pro) renin receptor [15,16], renin-independent mechanisms of Ang peptide generation from Ang-(1-12) [17,18], intracellular RAS [19], previously mentioned ACE2/Ang-(1-7)/Mas receptor pathway [20] and they all may possess therapeutic potential.…”

New Components of the Renin-Angiotensin-Aldosterone System and Oxidative Stress

“…Ang-(1-12) can be degraded into smaller peptides such as Ang-(1-7) by ACE, neprilysin or chymase [64]. Ang-(1-12) was also reported to bind AT 1 receptors, serving not only as a substrate for smaller active peptides, but also as a ligand [65].…”