Immunoreactivity and Guanosine 3′,5′-Cyclic Monophosphate Activating Actions of Various Molecular Forms of Human B-Type Natriuretic Peptide

Abstract

Abstract-Recent studies support the speculation that different molecular forms of the cardiac hormone BNP with differential biological activity may circulate in heart failure and be detected by conventional assays. In the current study we determined the ability of 3 widely used conventional assays to detect these different forms thought to circulate in heart failure. We also evaluated the ability of pro-BNP (1-108), N-terminal peptide (NT)-pro-BNP (1-76), and BNP 3-32, the latter a cleavage product of BNP 1-32 by dipeptidyl peptidase IV, on an equimolar basis to activate cGMP in cultured cardiac fibroblasts and cardiomyocytes compared with the biologically active mature BNP 1-32. Specifically, we observed that the Roche NT-pro-BNP assay detected both NT-pro-BNP 1-76 and pro-BNP 1-108 and that Biosite Triage and Shionogi detected both mature BNP 1-32 and the shortened BNP 3-32. Moreover, in cultured cardiac fibroblasts and cardiomyocytes, BNP 1-32 (10 Ϫ6 mol/L) activated cGMP. BNP 3-32 demonstrated a similar cGMP activating property in both cardiac cell types. In contrast, the cGMP response to pro-BNP 1-108 and NT-pro-BNP 1-76 was not significantly greater than no treatment alone. We conclude that widely used commercial assays for NT-pro-BNP 1-76 and BNP 1-32 cannot differentiate among pro-, processed, or degraded forms and, thus, may not thoroughly identify circulating BNP forms in heart failure patients. These findings also demonstrate differential cGMP activating properties of BNP forms and, importantly, that pro-BNP 1-108 and NT-pro-BNP 1-76 have reduced cGMP activity in vitro that may have biological relevance to human heart failure.

Processing of Pro–B-Type Natriuretic Peptide: Furin and Corin as Candidate Convertases2

“…Unprocessed proBNP has markedly reduced, if any, hormonal activity (35,36 ). The presence of unprocessed prohormone in the blood of HF patients suffering from insufficient natriuresis and diuresis remains an unexplained phenomenon.…”

Natriuretic Peptides and Analytical Barriers

“…ProBNP is a poor stimulator of guanylyl cyclase. There are differential cGMP activating properties of BNP forms and, notably, proBNP 1-108 and NT-proBNP 1-76 have reduced cGMP activity in vitro (29 ). Some speculate that unprocessed proBNP is a circulating "reserve" of BNP since in vivo cleavage of proBNP produces BNP.…”

“…While the first generation of NT-proBNP immunoassays uses polyclonal antibodies to amino acids 1-21 and 39 -50, the second generation uses monoclonal antibodies (MAb) recognizing the central region of NTproBNP: amino acids 22-28 and 42-46 (27)(28)(29)(30)(31). Many studies have demonstrated the negative effect of glycosylation for antibody recognition to the middle area of the NT-proBNP molecule.…”

Corin Is Present in the Normal Human Heart, Kidney, and Blood, with Pro–B-Type Natriuretic Peptide Processing in the Circulation

“…Increasing attention on proBNP has occurred based on reports that it is increased in the plasma of humans with advanced heart failure (HF) (11,12 ). Further, recent studies have reported that circulating corin is reduced in human HF, advancing the concept that increased proBNP in human HF, which is recognized by conventional BNP assays and has reduced cGMP actions, is a result of a corin deficiency (12)(13)(14).…”