Critical Role of Neprilysin in Kidney Angiotensin Metabolism

Abstract

Rationale: Kidney homeostasis is critically determined by the coordinated activity of the renin-angiotensin system (RAS) including the balanced synthesis of its main effector peptides angiotensin (Ang) II and Ang-(1-7). The condition of enzymatic overproduction of Ang II relative to Ang- (1-7) is termed RAS dysregulation, and leads to cellular signals, which promote hypertension and organ damage, and ultimately progressive kidney failure. Angiotensin-converting enzyme 2 (ACE2) and neprilysin (NEP) induce the 'alternative', and potentially reno-protective axis by enhancing Ang-(1-7) production. However, their individual contribution to baseline RAS balance and whether their activities change in chronic kidney disease (CKD), has not yet been elucidated. Objective: To examine whether NEP-mediated Ang-(1-7) generation exceeds Ang II formation in the healthy kidney compared to diseased kidney. Methods and Results: In this exploratory study, we used mass spectrometry (LC-MS/MS) to measure Ang II and Ang-(1-7) synthesis rates of ACE, chymase and NEP, ACE2, prolyl-endopeptidase (PEP), prolyl-carboxypeptidase (PCP) in kidney biopsy homogenates in 11 healthy living kidney donors and 12 patients with CKD. Spatial expression of RAS enzymes was determined by immunohistochemistry. Healthy kidneys showed higher NEP-mediated Ang-(1-7) synthesis than Ang II formation, thus displaying a strong preference towards the reno-protective alternative RAS axis. In contrast, in CKD kidneys higher levels of Ang II were recorded, which originated from mast cell chymase activity. Conclusions: Ang-(1-7) is the dominant RAS peptide in healthy human kidneys with NEP rather than ACE2 being essential for its generation. Severe RAS dysregulation is present in CKD dictated by high chymase-mediated Ang II formation. Kidney RAS enzyme analysis might lead to novel therapeutic approaches for CKD.

Mast Cell Chymase and Kidney Disease

“…The analysis of Ang II in human kidney lysates using mass spectrometry in the presence of an ACE inhibitor further showed that in normal kidneys more than 80% of Ang II generation was ACE-dependent, while this proportion was reversed in kidneys from CKD patients, supporting a predominant role in pathology [59] (Figure 1). The inhibitors of both ACE and chymase blunted almost all Ang II generation [59]. While these data are completely in line with significant renal infiltration by chymase-positive MCs, such as observed in rejected kidneys [45], IgA nephropathy [60,61], diabetic [58,62] and hypertensive nephropathy [63], as well as in autosomal polycystic kidney disease [64], they may simply reflect the relative amount of ACE and chymase in tissues, without being functionally relevant.…”

“…The possibility to have ACE-independent Ang II generation launched a series of studies evoking a possible role of chymase-dependent Ang II formation in various human tissues, such as the vasculature [51], heart [12] and kidneys after a high salt intake [57] and in patients with diabetic nephropathy [58]. The analysis of Ang II in human kidney lysates using mass spectrometry in the presence of an ACE inhibitor further showed that in normal kidneys more than 80% of Ang II generation was ACE-dependent, while this proportion was reversed in kidneys from CKD patients, supporting a predominant role in pathology [59] (Figure 1). The inhibitors of both ACE and chymase blunted almost all Ang II generation [59].…”

ACE2, the Counter-Regulatory Renin–Angiotensin System Axis and COVID-19 Severity

“…There is severe RAS dysregulation in CKD [65]. In normal subjects, overall kidney ANG (1-7) generation exceeds ANG II generation by 2.6-fold and is 3.9 times higher than in CKD, indicating preponderance of the counter-regulatory RAS axis in healthy kidneys (Figure 3).…”

“…Although ACE2 has been described as a crucial player in the enzymatic conversion of ANG II to ANG (1-7), this may not be the case in the kidney, in which neprilysin (NEP) seems to be the major source of renal ANG (1-7) [64]. Indeed, it is estimated that in the healthy human kidneys, ACE2, prolylcarboxypeptidase (PCP), and POP together contribute less than 15% of total ANG (1-7) production [65].…”

Cyclotides Isolated From Violet Plants of Cameroon Are Inhibitors of Human Prolyl Oligopeptidase

“…Arg-vasopressin, substance P, oxytocin, bradykinin and neurotensin have been identified as substrates of POP ( López et al, 2011 ). Further, angiotensin II conversion to angiotensin (1-7) is also POP dependent and thus, POP may be relevant in renal diseases, including hypertension and cardiovascular disorders ( Kaltenecker et al, 2020 ; Serfozo et al, 2020 ). POP was also being discussed as drug target in dementia, schizophrenia and amnesia; in this regard, POP inhibitors were studied in clinical trials ( López et al, 2011 ; López et al, 2013 ; Mannisto and Garcia-Horsman, 2017 ), without any drug being translated into clinical use, yet.…”