Four-hour infusions of synthetic atrial natriuretic peptide in normal volunteers.

Two doses of synthetic atrial natriuretic peptide (0.5 and 5.0 micrograms/min) and its vehicle were infused intravenously for 4 hours in eight salt-loaded normal volunteers, and the effect on blood pressure, heart rate, renal hemodynamics, solute excretion, and secretion of vasoactive hormones was studied. The 0.5 micrograms/min infusion did not alter blood pressure or heart rate, whereas the 5.0 micrograms/min infusion significantly reduced the mean pressure by 20/9 mm Hg after 2.5 to 3 hours and increased the heart rate slightly. Inulin clearance was not significantly changed, but the mean p-aminohippurate clearance fell by 13 and 32% with the lower and higher doses, respectively. Urinary excretion of sodium and chloride increased slightly with the lower dose. With the higher dose, a marked increase in urinary excretion of sodium, chloride, and calcium was observed, reaching a peak during the second hour of the infusion. Potassium and phosphate excretion did not change significantly. A brisk increase in urine flow rate and fractional water excretion was seen only during the first hour of the high-dose infusion. Signs and symptoms of hypotension were observed in two subjects. No change in plasma renin activity, angiotensin II, or aldosterone was observed during either infusion, but a marked increase occurred after discontinuation of the high-dose infusion. In conclusion, the 5 micrograms/min infusion induced a transient diuretic effect, delayed maximal natriuretic activity, and a late fall in blood pressure, with no change in inulin clearance but a dose-related decrease in p-aminohippurate clearance. Despite large amounts of sodium excreted and blood pressure reduction, no counterregulatory changes were observed in the renin-angiotensin-aldosterone system or plasma vasopressin levels during the infusion.

Impact of orthologous melan-A peptide immunizations on the anti-self melan-A/HLA-A2 T cell cross-reactivity.

In HLA-A2 individuals, the CD8 T cell response against the differentiation Ag Melan-A is mainly directed toward the peptide Melan-A26-35. The murine Melan-A24-33 sequence encodes a peptide that is identical with the human Melan-A26-35 decamer, except for a Thr-to-Ile substitution at the penultimate position. Here, we show that the murine Melan-A24-33 is naturally processed and presented by HLA-A2 molecules. Based on these findings, we compared the CD8 T cell response to human and murine Melan-A peptide by immunizing HLA-A2 transgenic mice. Even though the magnitude of the CTL response elicited by the murine Melan-A peptide was lower than the one elicited by the human Melan-A peptide, both populations of CTL recognized the corresponding immunizing peptide with the same functional avidity. Interestingly, CTL specific for the murine Melan-A peptide were completely cross-reactive against the orthologous human peptide, whereas anti-human Melan-A CTL recognized the murine Melan-A peptide with lower avidity. Structurally, this discrepancy could be explained by the fact that Ile32 of murine Melan-A24-33 created a larger TCR contact area than Thr34 of human Melan-A26-35. These data indicate that, even if immunizations with orthologous peptides can induce strong specific T cell responses, the quality of this response against syngeneic targets might be suboptimal due to the structure of the peptide-TCR contact surface.

Adjuvant cisplatin-based combined chemotherapy for lymph node (LN)-positive urothelial carcinoma of the bladder (UCB) after radical cystectomy (RC): a retrospective international study of >1500 patients.

OBJECTIVE: To compare outcomes of patients with lymph node (LN)-positive urothelial carcinoma of the bladder (UCB) treated with or without cisplatin-based combined adjuvant chemotherapy (AC) after radical cystectomy (RC). PATIENTS AND METHODS: We retrospectively analysed 1523 patients with LN-positive UCB, who underwent RC with bilateral pelvic LN dissection. All patients had no evidence of disease after RC. AC was administered within 3 months. Competing-risks models were applied to compare UCB-related mortality. RESULTS: Of the 1523 patients, 874 (57.4%) received AC. The cumulative 1-, 2- and 5-year UCB-related mortality rates for all patients were 16%, 36% and 56%, respectively. Administration of AC was associated with an 18% relative reduction in the risk of UCB-related death (subhazard ratio 0.82, P = 0.005). The absolute reduction in mortality was 3.5% at 5 years. The positive effect of AC was detectable in patients aged ≤70 years, in women, in pT3-4 disease, and in those with a higher LN density and lymphovascular invasion. This study is limited by its retrospective and non-randomised design, selection bias, the absence of central pathological review and lack in standardisation of LN dissection and cisplatin-based protocols. CONCLUSION: AC seems to reduce UCB-related mortality in patients with LN-positive UCB after RC. Younger patients, women and those with high-risk features such as pT3-4 disease, a higher LN density and lymphovascular invasion appear to benefit most. Appropriately powered prospective randomised trials are necessary to confirm these findings.

Liquid-crystalline ordering of antimicrobial peptide-DNA complexes controls TLR9 activation.

Double-stranded DNA (dsDNA) can trigger the production of type I interferon (IFN) in plasmacytoid dendritic cells (pDCs) by binding to endosomal Toll-like receptor-9 (TLR9; refs , , , , ). It is also known that the formation of DNA-antimicrobial peptide complexes can lead to autoimmune diseases via amplification of pDC activation. Here, by combining X-ray scattering, computer simulations, microscopy and measurements of pDC IFN production, we demonstrate that a broad range of antimicrobial peptides and other cationic molecules cause similar effects, and elucidate the criteria for amplification. TLR9 activation depends on both the inter-DNA spacing and the multiplicity of parallel DNA ligands in the self-assembled liquid-crystalline complex. Complexes with a grill-like arrangement of DNA at the optimum spacing can interlock with multiple TLR9 like a zipper, leading to multivalent electrostatic interactions that drastically amplify binding and thereby the immune response. Our results suggest that TLR9 activation and thus TLR9-mediated immune responses can be modulated deterministically.

Antagonistic peptide technology for functional dissection of CLE peptides revisited.

In the Arabidopsis thaliana genome, over 1000 putative genes encoding small, presumably secreted, signalling peptides can be recognized. However, a major obstacle in identifying the function of genes encoding small signalling peptides is the limited number of available loss-of-function mutants. To overcome this, a promising new tool, antagonistic peptide technology, was recently developed. Here, this antagonistic peptide technology was tested on selected CLE peptides and the related IDA peptide and its usefulness in the context of studies of peptide function discussed. Based on the analyses, it was concluded that the antagonistic peptide approach is not the ultimate means to overcome redundancy or lack of loss-of-function lines. However, information collected using antagonistic peptide approaches (in the broad sense) can be very useful, but these approaches do not work in all cases and require a deep insight on the interaction between the ligand and its receptor to be successful. This, as well as peptide ligand structure considerations, should be taken into account before ordering a wide range of synthetic peptide variants and/or generating transgenic plants.

The development of decision limits for the GH-2000 detection methodology using additional insulin-like growth factor-I and amino-terminal pro-peptide of type III collagen assays.

The GH-2000 and GH-2004 projects have developed a method for detecting GH misuse based on measuring insulin-like growth factor-I (IGF-I) and the amino-terminal pro-peptide of type III collagen (P-III-NP). The objectives were to analyze more samples from elite athletes to improve the reliability of the decision limit estimates, to evaluate whether the existing decision limits needed revision, and to validate further non-radioisotopic assays for these markers. The study included 998 male and 931 female elite athletes. Blood samples were collected according to World Anti-Doping Agency (WADA) guidelines at various sporting events including the 2011 International Association of Athletics Federations (IAAF) World Athletics Championships in Daegu, South Korea. IGF-I was measured by the Immunotech A15729 IGF-I IRMA, the Immunodiagnostic Systems iSYS IGF-I assay and a recently developed mass spectrometry (LC-MS/MS) method. P-III-NP was measured by the Cisbio RIA-gnost P-III-P, Orion UniQ? PIIINP RIA and Siemens ADVIA Centaur P-III-NP assays. The GH-2000 score decision limits were developed using existing statistical techniques. Decision limits were determined using a specificity of 99.99% and an allowance for uncertainty because of the finite sample size. The revised Immunotech IGF-I - Orion P-III-NP assay combination decision limit did not change significantly following the addition of the new samples. The new decision limits are applied to currently available non-radioisotopic assays to measure IGF-I and P-III-NP in elite athletes, which should allow wider flexibility to implement the GH-2000 marker test for GH misuse while providing some resilience against manufacturer withdrawal or change of assays. Copyright © 2015 John Wiley & Sons, Ltd.

Multi-institutional validation of the prognostic value of Ki-67 labeling index in patients treated with radical prostatectomy.

OBJECTIVE: Several smaller single-center studies have reported a prognostic role for Ki-67 labeling index in prostate cancer. Our aim was to test whether Ki-67 is an independent prognostic marker of biochemical recurrence (BCR) in a large international cohort of patients treated with radical prostatectomy (RP). METHODS: Ki-67 immunohistochemical staining on prostatectomy specimens from 3,123 patients who underwent RP for prostate cancer was retrospectively performed. Univariable and multivariable Cox regression models were used to assess the association of Ki-67 status with BCR. RESULTS: Ki-67 positive status was observed in 762 (24.4 %) patients and was associated with lymph node involvement (LNI) (p = 0.039). Six hundred and twenty-one (19.9 %) patients experienced BCR. The estimated 3-year biochemical-free survivals were 85 % for patients with negative Ki-67 status and 82.1 % for patients with positive Ki-67 status (log-rank test, p = 0.014). In multivariable analysis that adjusted for the effects of age, preoperative PSA, RP Gleason sum, seminal vesicle invasion, extracapsular extension, positive surgical margins, lymphovascular invasion, and LNI, Ki-67 was significantly associated with BCR (HR = 1.19; p = 0.019). Subgroup analysis revealed that Ki-67 is associated with BCR in patients without LNI (p = 0.004), those with RP Gleason sum 7 (p = 0.015), and those with negative surgical margins (p = 0.047). CONCLUSION: We confirmed Ki-67 as an independent predictor of BCR after RP. Ki-67 could be particularly informative in patients with favorable pathologic characteristics to help in the clinical decision-making regarding adjuvant therapy and optimized follow-up scheduling.

The predictive value of preoperative B-type natriuretic peptide in children undergoing cardiac surgery

Introduction: B-type natriuretic peptide (BNP) is a biomarker of myocardial stress. In children, the value of preoperative BNP on postoperative outcome is unclear. The aim of this study was to determine the predictive value of preoperative NT-proBNP on postoperative outcome in children after congenital heart surgery. Results: Ninety-seven patients were included in the study with a median age of 3.3 years [0.7-5.2]. Preoperative median NT-proBNP was 412 pg/ml [164-1309]. NT-proBNP was above the P95 reference value for age in 56 patients (58%). Preoperative NT-proBNP was significantly higher in patients who had mechanical ventilation duration of more than 2 days (1156 pg/ml [281-1951] vs. 267 pg/ml [136-790], p=0.003) and who stayed more than 6 days in the pediatric intensive care unit (727 pg/ml [203-1951] vs. 256 pg/ml [136-790], p=0.007). However, preoperative NT-proBNP was not significantly higher in patients with an increased inotropic score, a prolonged cardiopulmonary bypass time or an increased surgical risk category. Conclusions: An elevated preoperative NT-proBNP reflects hemodynamic status and cardiac dysfunction, and therefore is a valuable adjunct in predicting a complicated postoperative course. ___________________________________ Introduction: Le peptide natriurétique type B (BNP) est un marqueur reflétant le stress myocardique. Dans la population pédiatrique, la signification des valeurs préopératoire de BNP, en particulier sur l'évolution postopératoire, n'est pas clairement établie. Le but de l'étude est de déterminer la valeur prédictive de la partie NT sérique du BNP (NT-proBNP) sur l'évolution post opératoire d'enfants porteur d'une cardiopathie congénitale et ayant eu une chirurgie cardiaque. Résultats: Nonante-sept enfants ont été inclus dans l'étude, avec un âge médian de 3.3 ans [0.7-5.2]. La valeur médiane du NT-proBNP préopératoire était de 412 pg/ml [164-1309]. Le NT-proBNP préopératoire était supérieur au P95 des valeurs de référence pour l'âge chez 56 patients (58%). Le NT-proBNP préopératoire était significativement plus élevé chez les patients ayant eu plus de deux jours de ventilation mécanique dans la période postopératoire (1156 pg/ml [281-1951] vs. 267 pg/ml [136-790], p=0.003) et ayant été hospitalisés plus de 6 jours dans l'unité de soins intensifs pédiatrique (727 pg/ml [203-1951] vs. 256 pg/ml [136-790], p=0.007). Par contre, le NT-proBNP préopératoire n'était pas significativement plus élevé chez les patients ayant eu un score d'inotrope élevé pendant leur hospitalisation aux soins intensifs, un temps de circulation extracorporelle prolongé ou ayant subi une chirurgie avec un risque chirurgical élevé. Conclusions: Un NT-proBNP sérique élevé en préopératoire reflète l'importance du stress myocardique induit par l'hémodynamique et la dysfonction myocardique, il est un marqueur qui permet d'améliorer l'identification des patients à risque d'avoir une évolution post opératoire compliquée.

The role of adjuvant chemotherapy for lymph node-positive upper tract urothelial carcinoma following radical nephroureterectomy: a retrospective study.

OBJECTIVE: To evaluate the effect of adjuvant chemotherapy (AC) on mortality after radical nephroureterectomy (RNU) for upper tract urothelial carcinoma (UTUC) with positive lymph nodes (LNs) and to identify patient subgroups that are most likely to benefit from AC. PATIENTS AND METHODS: We retrospectively analysed data of 263 patients with LN-positive UTUC, who underwent full surgical resection. In all, 107 patients (41%) received three to six cycles of AC, while 156 (59.3%) were treated with RNU alone. UTUC-related mortality was evaluated using competing-risks regression models. RESULTS: In all patients (Tall N+), administration of AC had no significant impact on UTUC-related mortality on univariable (P = 0.49) and multivariable (P = 0.11) analysis. Further stratified analyses showed that only N+ patients with pT3-4 disease benefited from AC. In this subgroup, AC reduced UTUC-related mortality by 34% (P = 0.019). The absolute difference in mortality was 10% after the first year and increased to 23% after 5 years. On multivariable analysis, administration of AC was associated with significantly reduced UTUC-related mortality (subhazard ratio 0.67, P = 0.022). Limitations of this study are the retrospective non-randomised design, selection bias, absence of a central pathological review and different AC protocols. CONCLUSIONS: AC seems to reduce mortality in patients with pT3-4 LN-positive UTUC after RNU. This subgroup of LN-positive patients could serve as target population for an AC prospective randomised trial.

Synthetic long peptide booster immunization in rhesus macaques primed with replication-competent NYVAC-C-KC induces a balanced CD4/CD8 T-cell and antibody response against the conserved regions of HIV-1.

The Thai trial (RV144) indicates that a prime-boost vaccine combination that induces both T-cell and antibody responses may be desirable for an effective HIV vaccine. We have previously shown that immunization with synthetic long peptides (SLP), covering the conserved parts of SIV, induced strong CD4 T-cell and antibody responses, but only modest CD8 T-cell responses. To generate a more balanced CD4/CD8 T-cell and antibody response, this study evaluated a pox-vector prime/SLP boost strategy in rhesus macaques. Priming with a replication-competent NYVAC, encoding HIV-1 clade C gag, pol and nef, induced modest IFNγ T-cell immune responses, predominantly directed against HIV-1 Gag. Booster immunization with SLP, covering the conserved parts of HIV-1 Gag, Pol and Env, resulted in a more than 10-fold increase in IFNγ ELISpot responses in four of six animals, which were predominantly HIV-1 Pol-specific. The animals showed a balanced polyfunctional CD4 and CD8 T-cell response and high Ab titres.