Head-to-head comparison of 10 natriuretic peptide assays.

BACKGROUND: The aim of the study was to compare NT-proBNP and BNP levels in fresh samples from heart failure (HF) patients measured using 10 immunoassays and to assess their agreement. METHODS: NT-proBNP (CobasH232(®), Elecsys(®), Vidas(®), Vista(®), XPand(®), Vitros(®)) and BNP (Triage(®), Access(®), CentaurXP(®), Architect(®)) levels were measured in 39 heparin and 19 EDTA samples, respectively. RESULTS: The Pearson correlation coefficient ranged between 0.929 (Triage(®-)Centaur(®)) and 0.994 (Access(®)-Architect(®)) for BNP assays and between 0.972 (Vidas(®)-Cobas H232(®)) and 0.999 (Vitros(®)-Vidas(®)) for NT-proBNP assays. Passing Bablok regression analyses showed a significant difference in the slopes [0.80 (Centaur(®)-Triage(®)) to 1.84 (Architect(®)-Centaur(®))] and intercepts [-55 ng/L (Architect(®)-Centaur(®)) to 48 ng/L (Access(®)-Triage®)] for BNP assays, and a lower heterogeneity between NT-proBNP assays [0.83 (Vidas(®)-Elecsys(®)) to 1.20 (Vitros(®)-Vidas(®)) and -97 ng/L (XPand(®)-CobasH232(®)) to 51 ng/L (CobasH232(®)-Elecsys(®)) for slopes and intercepts, respectively]. The concordance correlation coefficient revealed a poor (ρc<0.90) to moderate (ρc=0.90-0.95) agreement in 4/6 pairs of BNP assays and an almost perfect (ρc>0.99) agreement in 5/15 pairs of NT-proBNP assays. The acceptable difference limit reflecting the number of individual discrepant results between two assays, ranged between 15.1% (Access(®)-CentaurXP(®)) and 34.5% (Architect(®)-Triage(®)) for BNP assays, and between 10.9% (Vidas(®)-Vitros(®)) and 55% (CobasH232(®)-Xpand(®)) for NT-proBNP assays. CONCLUSIONS: This study stresses the lack of transferability of the results obtained using different techniques to measure BNP and NT-proBNP levels in fresh samples. Individual reference ranges and HF diagnostic cut-offs should be assessed for each commercial NP immunoassay. We recommend to systematically monitoring HF patients using the same assay (BNP or NT-proBNP) over the time.

[Inventory of the use of natriuretic peptides in France]. / État des lieux de l'utilisation des peptides natriurétiques en France.

Since the introduction of routine assay for natriuretic peptides (NP), there is an increasing number of clinical applications for these assays. Due to the continuously increasing number of prescription of those tests, a reappraisal of the use of natriuretic peptide assays, namely BNP and NT-proBNP in France was necessary. This was achieved through a national survey to obtain a detailed description of NP prescription and realization by French laboratories. A questionnaire was sent in April 2010 to hospital and private clinical chemists. Statistical analysis of results concerned 584 answers. This survey demonstrated an equivalent use of BNP and NT-proBNP both in public or private laboratories together with a huge heterogeneity of tests used within labs. Medical prescription heterogeneity both in public or private sectors confirms the large implication of those tests in clinical diagnosis. These assays are not yet standardized, so clinicians and biologists should be very careful when interpreting the results for diagnostic or therapeutic monitoring.

Is heparin plasma suitable for the determination of B-type natriuretic peptide on the Beckman-Coulter Access 2?

BACKGROUND: The use of heparin as an alternative to EDTA in the production of plasma samples is of particular interest for B-type natriuretic peptide (BNP) measurements. Lithium heparin is now widely used for the determination of biochemical parameters, including cardiac markers. The goal of this study was to determine the feasibility of measuring BNP using heparin plasma instead of EDTA plasma with the Access 2 system (Beckman-Coulter). METHODS: BNP was determined in heparin plasma and EDTA plasma from 24 patients within 1 h of blood collection. Additional measurements were performed with heparin plasma, every hour for the first 4 h, and then 8 h after the collection of blood that was stored at room temperature. RESULTS: At H(0), the observed BNP concentrations in heparin plasma were much higher (mean values 65% higher) than those in EDTA plasma. Using predetermined thresholds, this difference would lead to 30% discordance between samples in heparin and EDTA. BNP stability decreased over time in heparin plasma: immunoreactivity decreased approximately by 30% during the first 2 h and by 60% after 8 h. CONCLUSIONS: Heparin plasma does not seem to be a suitable alternative to EDTA plasma for measurement of BNP using the Access 2 system, even if measurements are performed immediately after blood sampling.

The neuropeptide substance P regulates aldosterone secretion in human adrenals.

Aldosterone, produced by the adrenals and under the control of plasma angiotensin and potassium levels, regulates hydromineral homeostasis and blood pressure. Here we report that the neuropeptide substance P (SP) released by intraadrenal nerve fibres, stimulates aldosterone secretion via binding to neurokinin type 1 receptors (NK1R) expressed by aldosterone-producing adrenocortical cells. The action of SP is mediated by the extracellular signal-regulated kinase pathway and involves upregulation of steroidogenic enzymes. We also conducted a prospective proof-of-concept, double blind, placebo-controlled clinical trial aimed to investigate the impact of the NK1R antagonist aprepitant on aldosterone secretion in healthy male volunteers (EudraCT: 2008-003367-40, ClinicalTrial.gov: NCT00977223). Participants received during two 7-day treatment periods aprepitant (125 mg on the 1st day and 80 mg during the following days) or placebo in a random order at a 2-week interval. The primary endpoint was plasma aldosterone levels during posture test. Secondary endpoints included basal aldosterone alterations, plasma aldosterone variation during metoclopramide and hypoglycaemia tests, and basal and stimulated alterations of renin, cortisol and ACTH during the three different stimulatory tests. The safety of the treatment was assessed on the basis of serum transaminase measurements on days 4 and 7. All pre-specified endpoints were achieved. Aprepitant decreases aldosterone production by around 30% but does not influence the aldosterone response to upright posture. These results indicate that the autonomic nervous system exerts a direct stimulatory tone on mineralocorticoid synthesis through SP, and thus plays a role in the maintenance of hydromineral homeostasis. This regulatory mechanism may be involved in aldosterone excess syndromes.

Clinical Value of Natriuretic Peptides in Predicting Time to Dialysis in Stage 4 and 5 Chronic Kidney Disease Patients.

BACKGROUND: Anticipating the time to renal replacement therapy (RRT) in chronic kidney disease (CKD) patients is an important but challenging issue. Natriuretic peptides are biomarkers of ventricular dysfunction related to poor outcome in CKD. We comparatively investigated the value of B-type natriuretic peptide (BNP) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) as prognostic markers for the risk of RRT in stage 4 and 5 CKD patients, and in foretelling all-cause mortality and major cardiovascular events within a 5-year follow-up period. METHODS: Baseline plasma BNP (Triage, Biosite) and NT-proBNP (Elecsys, Roche) were measured at inclusion. Forty-three patients were followed-up during 5 years. Kaplan-Meier analysis, with log-rank testing and hazard ratios (HR), were calculated to evaluate survival without RRT, cardiovascular events or mortality. The independent prognostic value of the biomarkers was estimated in separate Cox multivariate analysis, including estimated glomerular filtration rate (eGFR), creatininemia and comorbidities. RESULTS: During the first 12-month follow-up period, 16 patients started RRT. NT-proBNP concentration was higher in patients who reached endpoint (3221 ng/L vs 777 ng/L, p = 0.02). NT-proBNP concentration > 1345 ng/L proved significant predictive value on survival analysis for cardiovascular events (p = 0.04) and dialysis within 60 months follow-up (p = 0.008). BNP concentration > 140 ng/L was an independent predictor of RRT after 12 months follow-up (p<0.005), and of significant predictive value for initiation of dialysis within 60 months follow-up. CONCLUSIONS: Our results indicate a prognostic value for BNP and NT-proBNP in predicting RRT in stage 4 and 5 CKD patients, regarding both short- and long-term periods. NT-proBNP also proved a value in predicting cardiovascular events. Natriuretic peptides could be useful predictive biomarkers for therapeutic guidance in CKD.

Osmotic stress, a proinflammatory signal in Caco-2 cells.

Hyper- (450 mOsm/l) and hypoosmotic exposure (150 mOsm/l) of Caco-2 cells, a human intestinal epithelial cell line, induced a twofold- and a fivefold increase in the production of IL-8, a constitutively expressed cytokine, respectively. This was observed both in the presence or in the absence of added proinflammatory cytokines and the stimulatory effect of osmotic stress was additive to that induced by the cytokines. Thus, IL-8 production appeared minimal around isoosmolarity, i.e. 300 mOsm/l. Concerning the signalling pathway involved, specific inhibition of p38- or p42/44 MAP kinases decreased the IL-8 production by about 30% independently of the osmotic condition used. Inhibition of c-jun-NH2-terminal kinase (JNK) by using both dicoumarol and SP600125 totally inhibited the stimulatory effect of hypoosmolarity. Moreover, hypoosmolarity induced an about threefold increase in JNK activity demonstrating that JNK was specifically involved in the effect of hypoosmolarity on IL-8 production. This is not the case for hyperosmolarity. Such an effect of osmotic stress was not restricted to IL-8, but was also observed on the production of IL-6, a non-constitutively expressed cytokine. Again, IL-6 production appeared minimal in isoosmotic condition. Taken together, these results demonstrate that osmotic stress is a proinflammatory signal in Caco-2 cells and suggest that an osmosensor might specifically exist in intestinal epithelial cells.

Effects of vitamin A on in vitro maturation of pre-pubertal mouse spermatogonial stem cells.

Testicular tissue cryopreservation is the only potential option for fertility preservation in pre-pubertal boys exposed to gonadotoxic treatment. Completion of spermatogenesis after in vitro maturation is one of the future uses of harvested testicular tissue. The purpose of the current study was to evaluate the effects of vitamin A on in vitro maturation of fresh and frozen-thawed mouse pre-pubertal spermatogonial stem cells in an organ culture system. Pre-pubertal CD1 mouse fresh testes were cultured for 7 (D7), 9 (D9) and 11 (D11) days using an organ culture system. Basal medium was supplemented with different concentrations of retinol (Re) or retinoic acid (RA) alone or in combination. Seminiferous tubule morphology (tubule diameter, intra-tubular cell type), intra-tubular cell death and proliferation (PCNA antibody) and testosterone level were assessed at D7, D9 and D11. Pre-pubertal mouse testicular tissue were frozen after a soaking temperature performed at -7 °C, -8 °C or -9 °C and after thawing, were cultured for 9 days, using the culture medium preserving the best fresh tissue functionality. Retinoic acid at 10(-6)M and retinol at 3.3.10(-7)M, as well as retinol 10(-6)M are favourable for seminiferous tubule growth, maintenance of intra-tubular cell proliferation and germ cell differentiation of fresh pre-pubertal mouse spermatogonia. Structural and functional integrity of frozen-thawed testicular tissue appeared to be well-preserved after soaking temperature at -8 °C, after 9 days of organotypic culture using 10(-6)M retinol. RA and Re can control in vitro germ cell proliferation and differentiation. Re at a concentration of 10(-6)M maintains intra-tubular cell proliferation and the ability of spermatogonia to initiate spermatogenesis in fresh and frozen pre-pubertal mouse testicular tissue using a soaking temperature at -8 °C. Our data suggested a possible human application for in vitro maturation of cryopreserved pre-pubertal testicular tissue.

Evaluation of endocrine testing of Leydig cell function using extractive and recombinant human chorionic gonadotropin and different doses of recombinant human LH in normal men.

BACKGROUND: The functional testing of endocrine testis uses extractive human chorionic gonadotropin (ehCG). Recombinant human hCG (rhCG), avoiding any contamination, should replace ehCG. Moreover, a functional evaluation with recombinant human LH (rhLH) would be closer to physiology than a pharmacological testing with hCG. METHODS: The study was conducted in normal men. We first evaluated the dose-effect of ehCG on plasma testosterone and estradiol levels, before and after injection of either hCG or vehicle. Secondly, the responses to the optimal dose of ehCG were compared with those of rhCG. Thirdly, we investigated the dose-effect of rhLH, on steroid hormone secretion. LH, testosterone, and estradiol plasma levels were measured after the injection of either rhLH or placebo. RESULTS: ehCG induced dose-dependent increases in plasma estradiol and testosterone levels. They respectively peaked at 24 and 72 h after the injection. The most potent dose of ehCG (5000 IU) induced results similar to those observed with 250 microg (6500 IU) rhCG. By comparison with placebo, rhLH induced a significant and dose-dependent increase in plasma testosterone levels 4 h after the injection. Peak response of testosterone to rhLH and rhCG was significantly correlated. rhLH did not induce significant change in plasma estradiol level. CONCLUSIONS: In normal men, a single i.v. injection of 150 IU rhLH induces a 25% rise in plasma testosterone levels by comparison with placebo. At the moment, the dynamic evaluation using hCG remains the gold standard test to explore the Leydig cell function. The use of 250 microg rhCG avoiding any contamination should be recommended.