BACKGROUND B-type natriuretic peptide (BNP), which is activated in heart failure (HF), is processed to an active form by corin. The corin gene is expressed in the human heart and kidney, but corin protein expression in the heart, kidney, and circulation, along with whether proBNP is processed by circulating corin, remains unknown. METHODS We examined corin protein expression by immunostaining and Western blot in human heart and kidney, and we assessed the circulating corin concentration by ELISA. We examined histidine-tagged (His-tag) proBNP1–108 processing in serum and plasma by immunoprecipitation and Western blot and sequenced the processed form. RESULTS Normal human heart and kidney displayed the presence of corin, especially in cells around the vasculature. Both corin and proBNP1–108 were present in the plasma of healthy human subjects, with circulating corin significantly higher in men than women (P < 0.0001) and a positive correlation of corin to age (P = 0.0497, r = 0.27). In fresh normal plasma and serum, His-tag proBNP1–108 was processed to a lower molecular weight form confirmed to be BNP. Processed BNP was higher in men than women (P = 0.041) and was positively correlated to plasma corin concentrations (P = 0.041, r = 0.65). CONCLUSIONS Our results support the concept that proBNP1–108 may be processed outside of the heart in the circulation where the proprotein convertase is present. Moreover, sex may impact this process, since corin concentrations are higher in men. These findings may have important physiologic and pathophysiologic implications for the proBNP/corin system in the human.

Myocardial aging is characterized by LV fibrosis leading to diastolic and systolic dysfunction. Studies have established the potent anti-fibrotic and anti-proliferative properties of C-type natriuretic peptide (CNP), however the relationship between circulating CNP, LV fibrosis and associated changes in LV function with natural aging are undefined. Accordingly, we characterized the relationship of plasma CNP with LV fibrosis and function in 2, 11 and 20 month old male Fischer rats. Further in vitro, we establish the anti-proliferative actions of CNP and the participation of the clearance receptor using adult human cardiac fibroblasts. Here we establish for the first time that a progressive decline in circulating CNP characterizes natural aging and is strongly associated with a reciprocal increase in LV fibrosis which precedes impairment of diastolic and systolic function. Additionally we demonstrate in cultured adult human cardiac fibroblasts that the direct anti-proliferative actions of high dose CNP may involve a non-cGMP pathway via the clearance receptor. Taken together these studies provide new insights into myocardial aging and the relationship to the anti-fibrotic and anti-proliferative peptide CNP.

Objectives Our objective was to determine the prognostic value of plasma N-terminal pro-B-type natriuretic peptide (NT-proBNP) for death and cardiovascular events among subjects without risk factors for heart failure (HF), which we term healthy normal. Background Previous studies report that plasma NT-proBNP has prognostic value for cardiovascular events in the general population even in the absence of HF. It is unclear if NT-proBNP retains predictive value in healthy normal subjects. Methods We identified a community-based cohort of 2,042 subjects in Olmsted County, Minnesota. Subjects with symptomatic (stage C/D) HF were excluded. The remaining 1,991 subjects underwent echocardiography and NT-proBNP measurement. We further defined healthy normal (n = 703) and stage A/B HF (n = 1,288) subgroups. Healthy normal was defined as the absence of traditional clinical cardiovascular risk factors and echocardiographic structural cardiac abnormalities. Subjects were followed for death, HF, cerebrovascular accident, and myocardial infarction with median follow-up of 9.1, 8.7, 8.8, and 8.9 years, respectively. Results NT-proBNP was not predictive of death or cardiovascular events in the healthy normal subgroup. Similar to previous reports, in stage A/B HF, plasma NT-proBNP values greater than age-/sex-specific 80th percentiles were associated with increased risk of death, HF, cerebrovascular accident, and myocardial infarction (p < 0.001 for all) even after adjustment for clinical risk factors and structural cardiac abnormalities. Conclusions These findings do not support the use of NT-proBNP as a cardiovascular biomarker in healthy normal subjects and have important implications for NT-proBNP–based strategies for early detection and primary prevention of cardiovascular disease.

Objective We sought to determine the physiologic actions and potential therapeutic applications of mutant atrial natriuretic peptide (mANP). Background The cardiac hormone atrial natriuretic peptide (ANP) is a 28 amino acid (AA) peptide which consists of a 17 AA ring structure together with a 6 AA N-terminus and a 5 AA C-terminus. In a targeted scan for sequence variants within the human ANP gene, a mutation was identified which results in a 40 AA peptide consisting of native ANP(1-28) and a C-terminal extension of 12 AA. We have termed this peptide mutant ANP. Methods In vitro 3′,5′-cyclic guanosine monophosphate (cGMP) activation in response to mANP was studied in cultured human cardiac fibroblasts known to express natriuretic peptide receptor A. The cardiorenal and neurohumoral properties of mANP compared to ANP were assessed in vivo in normal dogs. Results We observed an incremental in vitro cGMP dose response with increasing concentrations of mANP. In vivo with high dose mANP (33 pmol/kg/min) we observed significantly greater plasma cGMP activation, diuretic, natriuretic, glomerular filtration rate enhancing, renin-angiotensin-aldosterone system inhibiting, cardiac unloading and blood pressure lowering properties when compared to native ANP. Low dose mANP (2 pmol/kg/min) has natriuretic and diuretic properties without altering systemic hemodynamics compared to no natriuretic or diuretic response with low dose native ANP. Conclusions These studies establish that mANP activates cGMP in vitro and exerts greater and more sustained natriuretic, diuretic, glomerular filtration rate, and renal blood flow enhancing actions than native ANP.

Background-Diastolic dysfunction associated with high blood pressure (BP) leads to cardiac remodeling and fibrosis and progression to congestive heart failure. B-type natriuretic peptide (BNP) has BP-lowering, antifibrotic, and antihypertrophic properties, which makes BNP an attractive agent for attenuating the adverse cardiac remodeling associated with hypertension. In the current study, we tested the effects of sustained cardiac proBNP gene delivery on BP, cardiac function, and remodeling in spontaneously hypertensive rats (SHR). Methods and Results-We used the myocardium-tropic adeno-associated virus serotype 9 (AAV9) vector to achieve continuously enhanced cardiac rat proBNP expression. In SHR, a single systemic administration of AAV9 vector allowed long-term cardiac BNP overexpression, resulting in reductions in systolic and diastolic BP for 9 months after injection. Left ventricular (LV) thickness, LV end-systolic dimensions, and LV mass were reduced, whereas ejection fraction was significantly increased, in BNP-treated compared with untreated SHR. Circumferential systolic strain and strain rate of the early phase of diastole were improved in BNP-treated compared with untreated SHR. Noncardiac overexpression of BNP via AAV2 vector was not associated with changes in BP and plasma BNP in SHR. Furthermore, normal Wistar rats injected with AAV9 proBNP vector showed significantly reduced heart weights 4 weeks after injection without BP reduction. Conclusions-AAV9 vector facilitates sustained cardiac proBNP overexpression and improves LV function in hypertensive heart disease. Long-term proBNP delivery improved both systolic and diastolic function. The effects on cardiac structure and function occurred independently of BP-lowering effects in normal Wistar rats. (Circulation. 2011;123:1297-1305.)

Abstract-Recent studies report that, in the absence of heart failure and renal failure, plasma B-type natriuretic peptide (BNP) has prognostic value for mortality. We sought to confirm and extend these previous studies to assess BNP, measured by 3 distinct assays, as a biomarker for mortality in a strategy to enhance efforts at primary prevention and to better understand the clinical phenotype of such subjects at risk. We used a community-based cohort of 2042 subjects from Olmsted County, Minn, and individuals with heart or renal failure were excluded. BNP was assessed using 3 assays including Biosite and Shionogi for mature, biologically active BNP and the Roche assay for apparently nonbiologically active amino-terminal pro-BNP (NT-proBNP). Thorough echocardiographic and clinical data were recorded for all of the participants. Median follow-up for mortality was 5.6 years. BNP by all 3 of the assays was predictive of mortality. NT-proBNP and Biosite assays remained significant even after adjustment for traditional clinical risk factors and echocardiographic abnormalities including left ventricular hypertrophy and diastolic dysfunction. Echocardiography documented widespread structural changes in those with increasing BNP levels yet below levels observed in heart failure. We report in a large, well-characterized community-based cohort, free of heart failure, the first study to compare 3 distinct BNP assays as biomarkers for mortality in the same cohort. Our findings confirm the potential use of NT-proBNP and BNP biomarkers for future events and underscore that these peptides may also serve as biomarkers for underlying cardiac remodeling secondary to diverse cardiovascular disease entities. Key Words: natriuretic peptide Ⅲ hypertrophy T he cardiac hormone B-type natriuretic peptide (BNP) has proved useful in the diagnosis of human heart failure (HF). 1-3 Recently, studies have provided compelling data that plasma BNP, even in the absence of HF, has prognostic value for future cardiovascular events. 4 -6 Specifically, Wang et al 4 from the Framingham Heart Study reported in a prospective investigation in the general population without HF or renal failure that with each 1 SD increase in log BNP levels, there were significant increases in the risk of death, HF, atrial fibrillation, stroke or TIA, and first cardiovascular event over 5.2 mean years of follow-up. The reported plasma values associated with increased risk were well below the HF diagnosis threshold of 100 pg/mL. These important results suggest that, in the absence of HF, before the development of overt cardiac disease, modest elevations in plasma BNP are meaningful. Whereas this previous study provided significant information regarding the prognostic implications of BNP values below those seen in HF, the Framingham Heart Study 4 and others 5,7 have asked for validation of these findings in additional large, community-based studies.The current study was, therefore, designed to validate, as well as extend these recent seminal reports. We used the Prevalence ...

Impaired renal function with loss of nephron number in chronic renal disease (CKD) is associated with increased cardiovascular morbidity and mortality. However, the structural and functional cardiac response to early and mild reduction in renal mass is poorly defined. We hypothesized that mild renal impairment produced by unilateral nephrectomy (UNX) would result in early cardiac fibrosis and impaired diastolic function, which would progress to a more global left ventricular (LV) dysfunction. Cardiorenal function and structure were assessed in rats at 4 and 16 wk following UNX or sham operation (Sham); (n = 10 per group). At 4 wk, blood pressure (BP), aldosterone, glomerular filtration rate (GFR), proteinuria, and plasma B-type natriuretic peptide (BNP) were not altered by UNX, representing a model of mild early CKD. However, UNX was associated with significantly greater LV myocardial fibrosis compared with Sham. Importantly, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining revealed increased apoptosis in the LV myocardium. Further, diastolic dysfunction, assessed by strain echocardiography, but with preserved LVEF, was observed. Changes in genes related to the TGF-β and apoptosis pathways in the LV myocardium were also observed. At 16 wk post-UNX, we observed persistent LV fibrosis and impairment in LV diastolic function. In addition, LV mass significantly increased, as did LVEDd, while there was a reduction in LVEF. Aldosterone, BNP, and proteinuria were increased, while GFR was decreased. The myocardial, structural, and functional alterations were associated with persistent changes in the TGF-β pathway and even more widespread changes in the LV apoptotic pathway. These studies demonstrate that mild renal insufficiency in the rat results in early cardiac fibrosis and impaired diastolic function, which progresses to more global LV remodeling and dysfunction. Thus, these studies importantly advance the concept of a kidney-heart connection in the control of myocardial structure and function.

. Lack of activation of molecular forms of the BNP system in human grade 1 hypertension and relationship to cardiac hypertrophy. Am J Physiol Heart Circ Physiol 291: H1529 -H1535, 2006. First published April 28, 2006 doi:10.1152/ajpheart.00107.2006.-We evaluated relationships among two circulating molecular forms of brain natriuretic peptide (BNP32 and NT-proBNP), severity of hypertension (HTN), and cardiac hypertrophy in subjects with mild, moderate, and severe HTN. We prospectively studied 78 patients (43 males; mean age 51.4 Ϯ 11 yr) with essential HTN and 28 age-and sex-matched controls. BNP32 and NT-proBNP were measured by radioimmunoassay. In grade 1 HTN, BNP32 was not elevated and NT-proBNP was reduced (P ϭ 0.030) compared with controls. However, log-transformed values of BNP32 and NT-proBNP were both increased with severity of HTN from grade 1 to 3 (P Ͻ0.0001 and P ϭ 0.003, respectively). By multivariate analysis, log BNP32 was independently predicted by age (␤ ϭ 0.210, P ϭ 0.026) and HTN grade (␤ ϭ 0.274, P ϭ 0.004), whereas log NT-proBNP was independently predicted by sex (␤ ϭ 0.235, P ϭ 0.012) and HTN grade (␤ ϭ 0.218, P ϭ 0.0023). Two forms of BNP were measured in normal subjects and patients with essential HTN. In grade 1 HTN, BNP32 was unchanged and NTproBNP was significantly reduced compared with controls. As severity increased in humans with grade 1 to 3 HTN, both BNP32 and NT-proBNP levels were increased while not being affected by the presence of left ventricular hypertrophy. The lack of activation of BNP32 together with the reduction of NT-proBNP in grade 1 HTN may represent an impaired response of the BNP system in the early phase of HTN. The later activation of both forms of BNP may be a late compensatory effect, because it correlates with severity of HTN rather than cardiac hypertrophy/remodeling. natriuretic peptide; brain natriuretic peptide; amino-terminal pro-brain natriuretic peptide; left ventricular hypertrophy BRAIN NATRIURETIC PEPTIDE (BNP) is synthesized in cardiac myocytes as the precursor proBNP, a 108-amino acid peptide (4, 27, 33), and is converted to biologically active peptide BNP32 via a proteolytic cleavage by corin, a type II transmembrane cardiac serine protease (22,38,39). BNP is present in the circulation in two different forms, the biologically active BNP32 that elicits different beneficial cardiorenal and hormonal actions (1,5,8,11,25,26,40) and the amino-terminal portion of BNP (NT-proBNP) that consists of 76 amino acids.In congestive heart failure (CHF) as well as in hypertension (HTN), ventricular cardiomyocytes are progressively recruited to synthesize BNP (24). Whereas initially a successful defensive endocrine response by the heart to preserve cardiorenal homeostasis might occur, a hyporesponsiveness to cardiac peptides, which might contribute in turn to the progression of hypertension, could follow later. Although BNP32 has emerged as an important diagnostic and prognostic biomarker of CHF (1, 12, 37), its levels are markedly variable in HTN. Therefore, its b...