Processing Of proBNP Research Articles

Exposing the High Heterogeneity of Circulating Pro B-Type Natriuretic Peptide Fragments in Healthy Individuals and Heart Failure Patients.

The high molecular complexity of variably O-glycosylated and degraded pro B-type natriuretic peptide (proBNP) derived molecular forms challenges current immunoassays. Antibodies used show pronounced differences in cross-reactivities with these circulating fragments, which still need to be better characterized on a molecular level. To pave the way for advanced quantitative assays in the future, it is critical to fully understand these circulating forms. Plasma samples were collected from 8 heart failure (HF) patients and 2 healthy controls. NT-proBNP and proBNP were purified by immunoprecipitation and analyzed by nano-flow liquid chromatography coupled to high-resolution mass spectrometry. Fragments formed during proteolysis in solution digestion were distinguished from naturally occurring peptides by using an 18O stable isotope labeling strategy. We detected 16 previously unknown circulating fragments of proBNP peptides (9 of which are located in the N-terminal and 7 in the C-terminal region), revealing a more advanced state of degradation than previously known. Two of these fragments are indicative of either unidentified processing modes or a far-reaching C-terminal degradation (or a combination thereof) of the precursor proBNP. Our results further restrict ideal target epitopes for immunoassay antibodies and expand the current thinking of diversity, degradation, and processing of proBNP, as well as the distribution of circulating forms.

ProBNP processing is decreased by obesity in patients with heart failure

We thank Doctors Parcha and Arora (1), and Nishikimi and Nakagawa (2) for their interest in our study reporting that proBNP not glycosylated at threonine 71 (T71) was decreased in obese patients with chronic heart failure (3). We appreciate these authors taking the time to provide well-constructed overviews of the current literature relating to processing of proBNP and the known links between NT-proBNP and BNP and obesity. While BNP and NT- proBNP measurement is mandated in the diagnosis and risk stratification of heart failure (4), levels of these peptides are decreased with obesity, complicating their use in the clinical assessment of heart failure in obese individuals (5). The reasons for this decrease have not been adequately explained although many theories have been debated (2,3).

150 ProBNP glycosylation of threonine 71 is increased with obesity in patients with heart failure

Abstract Funding Acknowledgements The National Heart Foundation of New Zealand Background Heart failure (HF) is a leading cause of morbidity and mortality worldwide. Plasma levels of B-type natriuretic peptide (BNP) or its amino terminal congener (NT-proBNP) are used for HF diagnosis and risk stratification. Circulating levels of both BNP and NT-proBNP are reduced by obesity and this phenomenon is one of the key weaknesses of the diagnostic performance of the natriuretic peptides in HF. Formation of BNP from enzymatic cleavage of proBNP1-108 between residues 76 and 77 by corin and/or furin is influenced by the degree of proBNP glycosylation, therefore we investigated the relationship between proBNP glycosylation, plasma NT-proBNP and body mass index (BMI) in HF patients. Methods Three assays were developed to distinguish between total proBNP (glycosylated plus non-glycosylated proBNP), proBNP not glycosylated at threonine 71 (NG-T71) and proBNP not glycosylated in the central region (NG-C). Intra and inter-assay CVs were <15%, limits of detection were <2 pmol/L and samples diluted in parallel. Results Applying these assays and an NT-proBNP assay to plasma samples from 106 healthy volunteers and 238 patients with HF determined that concentrations (median(IQR)) of proBNP, NG-T71 and NT-proBNP were greater in HF patients compared to controls (24.9 (3.6-55), 9.4 (1.5-21) and 212 (104-409) pmol/L vs 3.0 (1.5-19), 3.0 (1.5-14.5) and 4.7 (2-8) pmol/L respectively, all p < 0.012). NG-C was undetectable in most samples. ProBNP levels in HF patients with BMI above and below 30 kg/m2 were not different (21.9 (2.6-70) pmol/L and 25.7 (3.9-53) pmol/L respectively, p = 0.85), whereas HF patients with BMI > 30 had lower NT-proBNP and NG-T71 levels (121 (64-248) and 3 (1.5-16) pmol/L verse 271 (178-486) and 13.5 (1.5-24.2) pmol/L respectively, p < 0.003) and higher proBNP:NT-proBNP and proBNP:NG-T71 ratios (p = 0.001 and p = 0.02 respectively) than those with BMI < 30. Discussion and Conclusion Using three new assays specific for different glycosylated forms of proBNP we have shown that the processing of proBNP is dysregulated in heart failure compared to controls due to increased glycosylation at threonine 71 of proBNP. Obese patients with HF have even greater dysregulation, demonstrated by decreased concentrations of proBNP that is not glycosylated at T71 (NG-T71), and concomitant decreases in NT-proBNP. Thus, we have shown for the first time that increased BMI is associated with increased proBNP glycosylation at T71 in patients with heart failure. Glycosylation-induced impairment of proBNP processing explains, at least in part, the reduction in plasma concentrations of B-type cardiac natriuretic peptides observed in obesity. Using these assays to evaluate the proBNP profile of larger patient cohorts will further develop understanding of the relationships between BNP production, BMI and heart failure pathogenesis, which would be expected to lead to increased diagnostic performance.

Cutting Edge of Brain Natriuretic Peptide (BNP) Research - The Diversity of BNP Immunoreactivity and Its Clinical Relevance.

Brain (or B-type) natriuretic peptide (BNP) is a cardiac hormone produced in the heart and an established biochemical marker for heart failure (HF) because the level in plasma increases in proportion to disease severity. Recently, the diversity of BNP molecular forms in the peripheral circulation, which includes mature BNP (BNP1-32) and its metabolites (BNP3-32, BNP4-32, and BNP5-32), was demonstrated. Moreover, studies showed that unprocessed BNP prohormone (proBNP) is also secreted from the heart, and its secretion is increased in patients with HF. Interestingly, BNP1-32, its metabolites, and proBNP are all detected as immunoreactive BNP by the currently available BNP assay system. Current N-terminal proBNP (NT-proBNP) assay systems also can react to both NT-proBNP and proBNP. In addition, the N-terminal region of proBNP and NT-proBNP are often O-glycosylated, which may result in underestimation of total NT-proBNP level, which includes both glycosylated and non-glycosylated NT-proBNP, by the NT-proBNP assay system. More recently, we have shown that miR30-GALNT-dependent O-glycosylation in the N-terminal region of proBNP affects the processing of proBNP and contributes to its secretion from the heart. The level of proBNP relative to BNP (proBNP/BNP ratio) in the coronary sinus is higher in patients with more severe HF. The proBNP/BNP ratio and the deglycosylated NT-proBNP level may be new and clinically useful biomarkers of HF.

Brain Natriuretic Peptide and Its Biochemical, Analytical, and Clinical Issues in Heart Failure: A Narrative Review.

Heart failure (HF) is a primary cause of morbidity and mortality worldwide. As the most widely studied and commonly applied natriuretic peptide (NP), B-type natriuretic peptide (BNP) has the effects of diuresis, natriuresis, vasodilation, anti-hypertrophy, and anti-fibrosis and it inhibits the renin-angiotensin-aldosterone and sympathetic nervous systems to maintain cardiorenal homeostasis and counteract the effects of HF. Both BNP and N-terminal pro B-type natriuretic peptide (NT-proBNP) are applied as diagnostic, managing, and prognostic tools for HF. However, due to the complexity of BNP system, the diversity of BNP forms and the heterogeneity of HF status, there are biochemical, analytical, and clinical issues on BNP not fully understood. Current immunoassays cross-react to varying degrees with pro B-type natriuretic peptide (proBNP), NT-proBNP and various BNP forms and cannot effectively differentiate between these forms. Moreover, current immunoassays have different results and may not accurately reflect cardiac function. It is essential to design assays that can recognize specific forms of BNP, NT-proBNP, and proBNP to obtain more clinical information. Not only the processing of proBNP (corin/furin) and BNP (neprilysin), but also the effects of glycosylation on proBNP processing and BNP assays, should be targeted in future studies to enhance their diagnostic, therapeutic, and prognostic values.

Effects of Sacubitril/Valsartan on cardiovascular biomarkers in chronic heart failure patients

Sacubitril/valsartan that combines a neprilysin (NEP) inhibitor and an angiotensin receptor blocker (ARB) was shown superior to enalapril for the treatment of heart failure with reduced ejection fraction (HFrEF) (1). The inhibition of sNEP by Sacubitril on the processing of proBNP remains unclear. The aim was to evaluate the evolution of cardiovascular biomarkers in patients who were switched from angiotensin converting enzyme inhibitor or ARB to sacubitril/valsartan and to evaluate the possible implication of the T71 proBNP glycosylation. We included 90 consecutive HFrEF patients who were treated with sacubitril/valsartan according to the current ESC guidelines in two university hospital. Plasma samples were collected before the first intake of sacubitril/valsartan, at day 30 (D30) and at day 90 (D90) for 60 patients who were selected for further paired analysis. sNEP activity decreased from D30 onward and being the lowest at D90. In contrast, there was no change in sNEP concentration. BNP and NT-proBNP plasma levels remained unchanged while high-sensitive troponin I decrease and GLP-1 increase from D90. We observed a gradual increase in T71 proBNP glycosylation. To assess the effect of T71 proBNP glycosylation on the processing of proBNP, we plotted the slope of the linear regression between BNP and NT-proBNP versus the median values of T71 proBNP glycosylation at each time points; the direct correlation observed ( R 2 = 0.99) strongly suggest that the independent evolution of BNP and NT-proBNP plasma levels in patients receiving sacubitril/valsartan results directly from the increase in T71 proBNP glycosylation. The results showed that while sacubitril/valsartan decreased sNEP activity without no effect on the mNEP release. The increase in T71 proBNP glycosylation being responsible for the discrepancies between BNP and NT-proBNP plasma levels.

Abstract 405: Unprocessed ProBNP Secretion is Regulated by miR30-GALNTs Axis Dependent O-glycosylation in the Pathological Myocardium

Background: It has been reported that biological inactive pro-brain natriuretic peptide(proBNP), precursor peptide of bioactive BNP, is increased in patients with heart failure, as well as mature BNP-32. However, underlying mechanisms of increased plasma proBNP levels in heart failure remain unclear. Methods: We used the conventional BNP assay system, which detects both proBNP and BNP-32, and our recently developed specific human proBNP assay system. We assessed cardiac production of proBNP and BNP(=proBNP+BNP-32) in patients with heart failure. To elucidate the molecular mechanisms underlying the processing and secretion of proBNP, we generated lentivirus vectors carrying several mutations in glycosylation sites of human proBNP. We evaluated the proBNP/BNP ratio, as an efficacy of processing of proBNP to BNP-32. Results: In coronary sinus, plasma proBNP/BNP ratios were significantly elevated in heart failure patients in accordance with the disease severity. In conditioned medium of cultured ventricular myocytes expressing wild type human proBNP, proBNP/BNP ratio was 37%, while in those of myocytes expressing glycosylation-resistant mutant of proBNP, proBNP/BNP ratio was only 3%, demonstrating the essential contribution of glycosylation to the secretion of proBNP. Among 7 glycosylation sites, we found that glycosylation of Thr48 and Thr71 in proBNP is critical for the secretion of proBNP from ventricular myocytes by attenuating proBNP processing. GalNAc-transferase(GALNT) is the largest glycosyltransferase enyme family, which controll O-glycosylasion. We identified GALNT 1 and 2 mediated the glycosylation-regulated increase in cardiac proBNP secretion by screening with siRNAs. Indeed, GALNT1 and 2-expression was increased in FCS-treated ventricular myocytes and failing hearts. We further found that GALNT1 and 2 expression is suppressed by miR-30 family, and that miR-30s expression is decreased in FCS-treated ventricular myocytes and failing hearts. Conclusions: We have elucidated novel molecular pathways that miR-30-GALNTs axis-mediated O -glycosylation in proBNP contributes to increased proportion of proBNP among secreted BNP in the pathological myocardium.

Effect of Corin Deficiency and Mutation on Cardiac Natriuretic Peptide Processing in the Intact Heart Under Cardiac Pressure-Overload Conditions

Natriuretic peptides ProANP and Pro BNP are translated as biologically-inactive pro-hormones that require proteolytic cleavage into biologically active carboxyl-terminal fragments by the serine protease Corin. Corin gene mutations are associated with risk for hypertension and left-ventricular hypertrophy in Blacks. However, the relative importance of Corin and the Corin T555I/Q568P mutations to proBNP processing in the heart is lacking. We adapted the Langendorff ex vivo cardiac perfusion model to allow dynamic assessment of cardiac processing of cardiac-derived proBNP using immunoassays specific for the murine molecular forms of unprocessed BNP1-95 and the active, carboxyl-terminus fragment murine BNP45. We also developed an inducible, cardiomyocyte-restricted Corin knockout mouse and a Corin T623I/Q636P homozygous knockin mouse to study the importance of cardiac Corin and the human Corin T555I/Q568P mutations to cardiac processing of proBNP. Results. As compared to WT controls, cardiomyocyte-restricted Corin knockout mice demonstrated significantly decreased BNP processing, and reduced myocardial cGMP and PKG kinase activity. The Corin623I(636P) knockin mice also demonstrated less severe but significant impairment in proBNP processing and reduced cardiac cGMP and PKG activity consistent with reduced GC-A activity. Cardiomyocyte Corin is essential to adequate processing of cardiac-derived proBNP and the autocrine/paracrine NPS. The murine Corin 623I(636P) heterozygous knockin model demonstrates impaired processing and reduced NPS autocrine/paracrine signaling during acute cardiac distention suggesting that Blacks heterozygous for the Corin I555(P568) haplotype may have reduced cardiac protection form the influence of chronic hypertension.

Secretion of Prohormone of B-Type Natriuretic Peptide, proBNP1-108, Is Increased in Heart Failure

Using a novel, specific assay for proBNP(1-108), this study tested the hypotheses that proBNP(1-108) is secreted by both nonfailing and failing human hearts and that proBNP(1-108) secretion is increased in failing hearts. The prohormone of B-type natriuretic peptide (proBNP(1-108)) is a 108-amino acid peptide produced primarily by the heart and cleaved into biologically active BNP(1-32) and the biologically inactive NT-proBNP(1-76). It is unknown to what extent increased cardiac proBNP1-108 secretion compared to reduced peripheral processing is responsible for elevated proBNP(1-108) levels in patients with heart failure (HF) compared to subjects without HF. The transcardiac gradient of proBNP(1-108) was determined by collecting arterial blood and blood from the coronary sinus (CS). Samples from subjects without overt heart disease (n = 9) were collected during cardiac catheterization after coronary artery disease had been excluded. Samples from HF patients (n = 21) were collected during implantation of a biventricular pacemaker. ProBNP(1-108) was measured with a new assay. Values are medians (25th/75th percentiles). The gradient of proBNP(1-108) across the nonfailing hearts was 8 (2/20) ng/l (aorta: 15 [1/25] ng/l; CS: 24 [8/41] ng/l; p = 0.018). The transcardiac gradient of proBNP(1-108) in the failing hearts was 326 (96/482) ng/l (arterial: 381 [201/586] ng/l; CS: 709 [408/1,087] ng/l; p<0.001). The transcardiac gradient was greater in failing than nonfailing hearts (p = 0.001). ProBNP(1-108) is secreted by nonfailing and failing human hearts, but more so in the latter. It remains to be established where peripheral processing of proBNP(1-108) occurs and how this is affected by disease.

Glycosylation and processing of pro-B-type natriuretic peptide in cardiomyocytes

B-type natriuretic peptide (BNP) and its related peptides are biomarkers for the diagnosis of heart failure. Recent studies identified several O-glycosylation sites, including Thr-71, on human pro-BNP but the functional significance was unclear. In this study, we analyzed glycosylation and proteolytic processing of pro-BNP in cardiomyocytes. Human pro-BNP wild-type (WT) and mutants were expressed in HEK 293 cells and murine HL-1 cardiomyocytes. Pro-BNP and BNP were analyzed by immunoprecipitation and Western blotting. Glycosidases and glycosylation inhibitors were used to examine carbohydrates on pro-BNP. The effects of furin and corin expression on pro-BNP processing in cells also were examined. We found that in HEK 293 cells, recombinant pro-BNP contained significant amounts of O-glycans with terminal oligosialic acids. Mutation at Thr-71 reduced O-glycans on pro-BNP and increased pro-BNP processing. In HL-1 cardiomyocytes, residue Thr-71 contained little O-glycans, and pro-BNP WT and T71A mutant were processed similarly. In HEK 293 cells, pro-BNP was processed by furin. Mutations at Arg-73 and Arg-76, but not Lys-79, prevented pro-BNP processing. In HL-1 cardiomyocytes, which express furin and corin, single or double mutations at Arg-73, Arg-76 and Lys-79 did not prevent pro-BNP processing. Only when all these three residues were mutated, was pro-BNP processing completely blocked. Our data indicate that pro-BNP glycosylation in cardiomyocytes differed significantly from that in HEK 293 cells. In HEK 293 cells, furin cleaved pro-BNP at Arg-76 whereas in cardiomyocytes corin cleaved pro-BNP at multiple residues including Arg-73, Arg-76 and Lys-79.

Human Pro–B-Type Natriuretic Peptide Is Processed in the Circulation in a Rat Model

The appearance of B-type natriuretic peptide (BNP) in the blood is ultimately caused by proteolytic processing of its precursor, proBNP. The mechanisms leading to the high plasma concentration of unprocessed proBNP are still poorly understood. The goals of the present study were to examine whether processing of proBNP takes place in the circulation and to evaluate the clearance rate of proBNP and proBNP-derived peptides. We studied the processing of human proBNP in the circulation and the clearance rate of proBNP and proBNP-derived peptides (BNP and N-terminal fragment of proBNP, NT-proBNP) in rats by injecting the corresponding peptides and analyzing immunoreactivity at specific time points. Glycosylated and nonglycosylated proBNP and NT-proBNP were used in the experiments. We applied immunoassays, gel filtration, and mass spectrometry (MS) techniques to analyze the circulation-mediated processing of proBNP. ProBNP was effectively processed in the circulation into BNP (1-32) and various truncated BNP forms as confirmed by gel filtration and MS analysis. Glycosylation of proBNP close to the cleavage-site region suppressed its processing in the circulation. The terminal half-life for human glycosylated proBNP was 9.0 (0.5) min compared with 6.4 (0.5) min for BNP. For NT-proBNP, the terminal half-lives were 15.7 (1.4) min and 15.5 (1.3) min for glycosylated and nonglycosylated forms, respectively. In rats, processing of human proBNP to active BNP occurs in the circulation. The clearance rate of proBNP is quite similar to that of BNP. These observations suggest that peripheral proBNP processing may be an important regulatory step rather than mere degradation.

Current biochemistry, molecular biology, and clinical relevance of natriuretic peptides

The mammalian natriuretic peptide family consists of atrial (ANP), brain [B-type; BNP] and C-type natriuretic peptide (CNP) and three receptors, natriuretic receptors-A (NPR-A), -B (NPR-B) and -C (NPR-C). Both ANP and BNP are abundantly expressed in the heart and are secreted mainly from the atria and ventricles, respectively. By contrast, CNP is mainly expressed in the central nervous system, bone and vasculature. Plasma concentrations of both ANP and BNP are elevated in patients with cardiovascular disease, though the magnitude of the increase in BNP is usually greater than the increase in ANP. This makes BNP is a clinically useful diagnostic marker for several pathophysiological conditions, including heart failure, ventricular remodeling and pulmonary hypertension, among others. Recent studies have shown that in addition to BNP-32, proBNP-108 also circulates in human plasma and that levels of both forms are increased in heart failure. Furthermore, proBNP-108 is O-glycosylated and circulates at higher levels in patients with severe heart failure. In this review we discuss recent progress in our understanding of the biochemistry, molecular biology and clinical relevance of the natriuretic peptide system.

Comparison of Mass Spectrometry and Clinical Assay Measurements of Circulating Fragments of B-Type Natriuretic Peptide in Patients With Chronic Heart Failure

Multiple B-type natriuretic peptide (BNP) fragments circulate in patients with heart failure (HF) but the types and relative quantities, particularly in relation to bioactive BNP 1-32, remain poorly defined. The purpose of the study was to relate clinically available BNP values with quantitative information on the concentration of pre-secretion and post-processed fragments of BNP detected by mass spectrometry. Seventy Class I-IV patients were prospectively enrolled with blood drawn into tubes containing a preservative to protect against BNP degradation. Samples were analyzed by quantitative mass spectrometry (MS) immunoassay for intact BNP 1-32 and its fragments. Clinical BNP 1-2 was measured by standard clinical laboratory methods. ProBNP 1-108, corin, and clinically measured BNP levels were elevated, but MS BNP 1-32 levels were low and differed from clinical BNP (P=0.01). Intact MS BNP 1-32 correlated modestly with clinical BNP (r=0.46, P<0.001). MS BNP fragments 3-32, 4-32, and 5-32 demonstrated the best associations with clinical BNP; fragment 5-32 with a correlation coefficient of r=0.81 (P<0.001). ProBNP 1-108 is measured by clinical BNP assays and contributes to the cumulative results of the BNP assay. However, the observation that clinically measured BNP correlates best with MS degradation fragments and relatively poorly with MS BNP 1-32 suggests that a significant component of circulating clinical BNP is composed of such fragments that are known to demonstrate little biological activity. There appear to be multiple pathways involved in the dysregulation of proBNP in HF, and both the processing of proBNP and the downstream degradation to BNP 1-32 appear to be critical.

A New Signal From B-Type Natriuretic Peptide in ST-Elevation Myocardial Infarction

A New Signal From B-Type Natriuretic Peptide in ST-Elevation Myocardial Infarction

Blood N-Terminal proBNP as a Potential Indicator of Cardiac Preload in Patients with High Volume Load

Stroke volume is mainly determined by preload, afterload and contractility. Accordingly, measuring cardiac preload provides essential information for treatment of hemodynamically unstable patients. Hemodynamic monitoring is widely used to measure cardiac preload, but the monitoring method is time-consuming and invasive. It is therefore important to establish a simple and non-invasive test for monitoring cardiac preload. Brain natriuretic peptide (BNP), which lowers systemic vascular resistance, is synthesized as proBNP in response to myocardial wall stretch, and blood BNP has been used as an indicator of preload. Here, we measured blood level of N-terminal proBNP (NT-proBNP), which is generated during processing of proBNP, because NT-proBNP is stable and easily measured at the bedside. To assess the correlation between blood NT-proBNP and preload, we also measured the global end-diastolic volume index (GEDVI) that reflects the cardiac preload. GEDVI was calculated with the volumes in all chambers of the heart at the time of end-diastole. Eight male patients (57.6 +/- 25.3 years old) with high volume load (1,000 ml within 4 hours) were included in the present study: 3 subjects with burn, 3 subjects with sepsis, a patient with alcoholic ketoacidosis and a resuscitated patient. Blood levels of NT-proBNP were 1,316.3 +/- 1,154.5 pg/ml (47 blood samples from the eight patients; the normal range, < 125 pg/ml). Notably, the increase in the NT-proBNP levels was associated with the increased GEDVI (r = 0.61, p < 0.0001). Therefore, blood NT-proBNP may be a good indicator of cardiac preload in patients with high volume load.

Biochemistry of B-type natriuretic peptide – where are we now?

The discovery of the cardiac endocrine function more than 25 years ago was a breakthrough for the implementation of a routine laboratory marker of heart failure. The heart secretes two different natriuretic peptides with similar biological effects. B-type natriuretic peptide (BNP) and N-terminal proBNP (NT-proBNP) emerged as the superior diagnostic markers compared to circulating A-type natriuretic peptide (ANP) and proANP derived peptide forms. However, the biochemistry with the processing and metabolism of proBNP, NT-proBNP and BNP with their impact on BNP quantification are still incompletely understood. Published data and own results were used. This review shows that the present understanding of the biochemistry of BNP and its circulating forms is far from complete. The regulation of BNP secretion occurs mainly at the level of gene transcription with only minor stores of proBNP and processed BNP 1-32 within cardiomyocytes. proBNP is believed to be split into BNP 1-32 and NT-proBNP 1-76 mainly upon secretion, although limited amounts of processed BNP have also been described in the secretory granules of atrial cardiomyocytes. proBNP can be also detected in the circulation and exists as a monomer. There is no evidence for relevant processing of proBNP into BNP 1-32 and NT-proBNP in the circulation. Only small amounts of the intact hormone BNP 1-32 appear to circulate in plasma, there are no known BNP binding proteins. The major circulating BNP forms appear to be split products of BNP 1-32, but are still not sufficiently characterized. Dipeptidyl-peptidase IV degrades BNP 1-32 to BNP 3-32 at its N-terminal end. NT-proBNP appears to be rapidly truncated at both ends as well. proBNP and NT-proBNP are glycosylated to a variable degree. BNP and NT-proBNP are extracted by the kidneys to a comparable extent. In contrast to BNP, the biological half-life of NT-proBNP in humans is still not known. The current commercial BNP assays do not significantly cross-react with NT-proBNP and vice versa. However, BNP as well as NT-proBNP assays cross-react with proBNP to a varying extent. The better characterization of circulating proBNP, BNP, and NT-proBNP forms is necessary to define the relevant standard for assay standardization. Until then, assays should be designed to detect the total amount of the circulating analyte to achieve optimal sensitivity. The identification of possible disease effects on proBNP, BNP, and NT-proBNP metabolism could pave the way for the development of more disease specific assays in the clinical setting.

Natriuretic Peptides and Myocardial Structure

It has been more than 2 decades since the seminal report by DeBold demonstrating the existence of atrial natriuretic peptide (ANP) as a cardiac hormone that links the heart and kidney in cardiorenal homeostasis.1 Since that discovery, a family of structurally similar but genetically distinct peptides have been reported that function via well-characterized particulate guanylyl cyclase receptors linked to cGMP. These include ANP and B-type naturetic peptide (BNP), which are ligands for the naturetic peptide receptor A, and C-type naturetic peptide, which binds to the naturetic peptide receptor B.2 In the current issue, Rame et al3 focus attention on the processing of the cardiac peptide prohormones to mature biologically active peptides by the protease corin. This study was predicated by the work by Yan et al,4 which established corin as the natriuretic peptide-converting enzyme for ANP and presumably BNP (Figure). This cleavage of the prohormone to the mature peptide is essential for biological activity exemplified by a novel murine model of corin deletion in which the phenotype is hypertension and cardiac hypertrophy.5 Processing of pro-ANP and pro-BNP. This figure illustrates the processing of these 2 procardiac natriuretic peptides by corin to biologically active mature cardiac natriuretic peptides ANP and BNP. These peptides mediate their …

The Role of the Circulation in Processing pro-Brain Natriuretic Peptide (proBNP) to Amino-Terminal BNP and BNP-32

Hunt, P. J., E. A. Espiner, G. M. Nicholls, A. M. Richards and T. G. Yandle. The role of the circulation in processing pro-brain natriuretic peptide (proBNP) to amino-terminal BNP and BNP-32. Peptides 18(10) 1475–1481, 1997.—Human proBNP (purified from cardiac tissue) was incubated at 37°C in whole blood, serum and plasma and the products analyzed by size exclusion high pressure liquid chromatography and radioimmunoassay (RIA). In addition to RIAs for BNP-32 and NT-proBNP(1-13), a newly developed RIA for proBNP(62-76) was also used to identify the peptides. Incubation with serum resulted in the formation of a 9 kDa and a 3 kDa peptide, consistent with the N-terminal and the C-terminal peptides of the propeptide. Minimal processing of proBNP was seen in blood or plasma, suggesting that the circulation does not play a major role in the activation of proBNP. Analysis of degradation products of human proBNP using site directed specific antisera indicates that removal of N-terminal amino acids from proBNP occurs in serum. These findings support the view that the “high molecular weight BNP-32” previously identified in human plasma comprises amino-terminal deleted forms, and is unlikely to be intact proBNP(1-108).