A Low Selenium Level Is Associated with Lung and Laryngeal Cancers

Purpose

It has been suggested that selenium deficiency is a risk factor for several cancer types. We conducted a case-control study in Szczecin, a region of northwestern Poland, on 95 cases of lung cancer, 113 cases of laryngeal cancer and corresponding healthy controls.

Methods

We measured the serum level of selenium and established genotypes for four variants in four selenoprotein genes (GPX1, GPX4, TXNRD2 and SEP15). Selenium levels in the cases were measured after diagnosis but before treatment. We calculated the odds of being diagnosed with lung or laryngeal cancer, conditional on selenium level and genotype.

Results

Among lung cancer cases, the mean selenium level was 63.2 µg/l, compared to a mean level of 74.6 µg/l for their matched controls (p<0.0001). Among laryngeal cancer cases, the mean selenium level was 64.8 µg/l, compared to a mean level of 77.1 µg/l for their matched controls (p<0.0001). Compared to a serum selenium value below 60 µg/l, a selenium level above 80 µg/l was associated with an odds ratio of 0.10 (95% CI 0.03 to 0.34; p = 0.0002) for lung cancer and 0.23 (95% CI 0. 09 to 0.56; p = 0.001) for laryngeal cancer. In analysis of four selenoprotein genes we found a modest evidence of association of genetic variant in GPX1 with the risk of lung and laryngeal cancers.

Conclusion

A selenium level below 60 µg/l is associated with a high risk of both lung and laryngeal cancer.     

Serum Uric Acid Concentrations in Meat Eaters,Fish Eaters,Vegetarians and Vegans: A Cross-Sectional Analysis in the EPIC-Oxford Cohort

Introduction

Circulating concentrations of uric acid may be affected by dietary components such as meat, fish and dairy products, but only a few studies have compared uric acid concentrations among individuals who exclude some or all of these foods from their diet. The aim of this study was to investigate differences in serum uric acid concentrations between meat eaters, fish eaters, vegetarians and vegans.

Subjects and Methods

A sample of 670 men and 1,023 women (424 meat eaters, 425 fish eaters, 422 vegetarians and 422 vegans, matched on age and sex) from the European Prospective Investigation into Cancer and Nutrition Oxford cohort were included in this cross-sectional analysis. Diet was assessed using a semi-quantitative food frequency questionnaire and serum concentrations of uric acid were measured. Mean concentrations of uric acid by diet group were calculated after adjusting for age, body mass index, calcium and alcohol intake.

Results

In both men and women, serum uric acid concentrations differed significantly by diet group (p<0.0001 and p = 0.01, respectively). The differences between diet groups were most pronounced in men; vegans had the highest concentration (340, 95% confidence interval 329–351 µmol/l), followed by meat eaters (315, 306–324 µmol/l), fish eaters (309, 300–318 µmol/l) and vegetarians (303, 294–312 µmol/l). In women, serum uric acid concentrations were slightly higher in vegans (241, 234–247 µmol/l) than in meat eaters (237, 231–242 µmol/l) and lower in vegetarians (230, 224–236 µmol/l) and fish eaters (227, 221–233 µmol/l).

Conclusion

Individuals consuming a vegan diet had the highest serum concentrations of uric acid compared to meat eaters, fish eaters and vegetarians, especially in men. Vegetarians and individuals who eat fish but not meat had the lowest concentrations of serum uric acid.     

Serum Lipopolysaccharide Binding Protein Levels Predict Severity of Lung Injury and Mortality in Patients with Severe Sepsis

Background

There is a need for biomarkers insuring identification of septic patients at high-risk for death. We performed a prospective, multicenter, observational study to investigate the time-course of lipopolysaccharide binding protein (LBP) serum levels in patients with severe sepsis and examined whether serial serum levels of LBP could be used as a marker of outcome.

Methodology/Principal Findings

LBP serum levels at study entry, at 48 hours and at day-7 were measured in 180 patients with severe sepsis. Data regarding the nature of infections, disease severity, development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), and intensive care unit (ICU) outcome were recorded. LBP serum levels were similar in survivors and non-survivors at study entry (117.4±75.7 µg/mL vs. 129.8±71.3 µg/mL, P = 0.249) but there were significant differences at 48 hours (77.2±57.0 vs. 121.2±73.4 µg/mL, P<0.0001) and at day-7 (64.7±45.8 vs. 89.7±61.1 µg/ml, p = 0.017). At 48 hours, LBP levels were significantly higher in ARDS patients than in ALI patients (112.5±71.8 µg/ml vs. 76.6±55.9 µg/ml, P = 0.0001). An increase of LBP levels at 48 hours was associated with higher mortality (odds ratio 3.97; 95%CI: 1.84–8.56; P<0.001).

Conclusions/Significance

Serial LBP serum measurements may offer a clinically useful biomarker for identification of patients with severe sepsis having the worst outcomes and the highest probability of developing sepsis-induced ARDS.     

Rosuvastatin improves impaired endothelial function,lowers high sensitivity CRP,complement and immuncomplex production in patients with systemic sclerosis - a prospective case-series study

Introduction

We studied the effect of rosuvastatin on endothelial and macrovascular function, cardiovascular risk factors and the complement pathway in patients with systemic sclerosis (SSc).

Methods

Altogether 28 patients with SSc underwent laboratory and complex vascular assessments before and after six months of 20 mg rosuvastatin treatment. Flow-mediated dilation (FMD) of the brachial artery, as well as carotid artery intima-media thickness (ccIMT), carotid-femoral and aorto-femoral pulse wave-velocity (PWV) were analyzed by ECG-synchronized ultrasound. Ankle-brachial index (ABI) was determined by Doppler, and forearm skin microcirculation was assessed by Laser Doppler perfusion monitoring.

Results

Brachial artery FMD significantly improved upon rosuvastatin therapy (2.2% ± 3.3% before versus 5.7% ± 3.9% after treatment, P = 0.0002). With regard to patient subsets, FMD significantly improved in the 21 lcSSc patients (from 2.1% to 5.6%, P = 0.001). In the seven dcSSc patients, we observed a tendency of improvement in FMD (from 3% to 6%, P = 0.25). Changes in PWV, ccIMT and ABI were not significant. Mean triglyceride (1.7 ± 0.97 versus 1.3 ± 0.46 mmol/l, P = 0.0004), total cholesterol (5.3 ± 1.6 mmol/l versus 4.2 ± 1.3 mmol/l, P = 0.0003), low density lipoprotein cholesterol (3.0 ± 1.3 versus 2.2 ± 1.0 mmol/l, P = 0.005) and C-reactive protein levels (CRP) (5.1 ± 5.2 versus 3.4 ± 2.7, P = 0.01) levels significantly decreased after rosuvastatin treatment. Mean C3, C4 and IC levels also decreased significantly as compared to pretreatment values.

Conclusions

Six-month rosuvastatin therapy improves endothelial function and lowers CRP, C3, C4 and IC levels indicating possible favourable effects of this statin on the cardiovascular and immune system in SSc.     

Serum Iron Concentration,but Not Hemoglobin,Correlates with TIMI Risk Score and 6-Month Left Ventricular Performance after Primary Angioplasty for Acute Myocardial Infarction

Objective

Anemia is associated with high mortality and poor prognosis after acute coronary syndrome (ACS). Increased red cell distribution width (RDW) is a strong independent predictor for adverse outcomes in ACS. The common underlying mechanism for anemia and increased RDW value is iron deficiency. It is not clear whether serum iron deficiency without anemia affects left ventricular (LV) performance after primary angioplasty for acute myocardial infarction (AMI). We investigated the prognostic value of serum iron concentration on LV ejection fraction (EF) at 6 months and its relationship to thrombolysis in myocardial infarction (TIMI) risk score in post MI patients.

Methods

We recruited 55 patients who were scheduled to undergo primary coronary balloon angioplasty after AMI and 54 age- and sex-matched volunteers. Serum iron concentration and interleukin-6 levels were measured before primary angioplasty. LVEF was measured by echocardiography at baseline and after 6 months. TIMI risk score was calculated for risk stratification.

Results

Serum iron concentration was significantly lower in those in whom LVEF had not improved ≥10% from baseline (52.7±24.1 versus 80.8±50.8 µg/dl, P = 0.016) regardless of hemoglobin level, and was significantly lower in the AMI group than in the control group (62.5±37.7 versus 103.0±38.1 µg/dl, P<0.001). Trend analysis revealed that serum iron concentration decreased as TIMI risk score increased (P = 0.002). In addition, lower serum iron concentrations were associated with higher levels of inflammatory markers. Multiple linear regression showed that baseline serum iron concentration can predict LV systolic function 6 months after primary angioplasty for AMI even after adjusting for traditional prognostic factors.

Conclusion

Hypoferremia is not only a marker of inflammation but also a potential prognostic factor for LV systolic function after revascularization therapy for AMI, and may be a novel biomarker for therapeutic intervention.     

The Sonic Hedgehog Signaling Pathway Induces Myopic Development by Activating Matrix Metalloproteinase (MMP)-2 in Guinea Pigs

Purpose

To investigate whether the Sonic hedgehog (Shh) signaling induces myopic development by increasing the expression of matrix metalloproteinase (MMP)-2 in guinea pigs.

Methods

A translucent diffuser was glued onto the right eye to induce form-deprivation myopia (FDM) in 10 guinea pigs. Four guinea pigs were served as a control group. The other 100 guinea pigs were subdivided into 5 groups (20 per group) and received a 10 µl intravitreal injection every 2 days for 4 times. Two groups were injected with 20 or 50 µg/ml Shh amino-terminal peptide (Shh-N) into the right eye and 0.1% bovine serum albumin into the other. FDM was induced in the right eyes of the three cyclopamine-treated groups and both eyes were injected with 50, 100, or 200 µg/ml cyclopamine. Retinoscopic refraction and eye dimensions were assessed on Day 14 of treatment. MMP-2 protein expression was determined in both scleras by western blotting.

Results

Both concentrations of Shh-N stimulated myopic development and axial growth as compared with control eyes. Myopia and axial elongation were significantly greater in the 50 µg/ml than in the 20 µg/ml Shh-N group (P<0.001 and P = 0.0019, respectively). All three doses of cyclopamine significantly attenuated myopic development compared with the FDM group (P<0.0001). Cyclopamine at 100 or 200 µg/ml significantly reduced axial elongation compared with the FDM group (P = 0.044 and P = 0.001, respectively). FDM-induced myopia and axial elongation were significantly greater in the 50 µg/ml than in the 200 µg/ml cyclopamine group (P<0.0001 and P = 0.008, respectively). MMP-2 expression was significantly greater in Shh-N–treated eyes than in the control eyes, and was lower in the cyclopamine plus FDM groups than in the FDM group.

Conclusions

The Shh signaling pathway induces myopic development by activating MMP-2 in guinea pigs.     

Intravitreal Docosahexaenoic Acid in a Rabbit Model: Preclinical Safety Assessment

Purpose

The purpose of the present study was to evaluate the retinal toxicity of a single dose of intravitreal docosahexaenoic acid (DHA) in rabbit eyes over a short-term period.

Methods

Sixteen New Zealand albino rabbits were selected for this pre-clinical study. Six concentrations of DHA (Brudy Laboratories, Barcelona, Spain) were prepared: 10 mg/50 µl, 5 mg/50 µl, 2’5 mg/50 µl, 50 µg/50 µl, 25 µg/50 µl, and 5 µg/50 µl. Each concentration was injected intravitreally in the right eye of two rabbits. As a control, the vehicle solution was injected in one eye of four animals. Retinal safety was studied by slit-lamp examination, and electroretinography. All the rabbits were euthanized one week after the intravitreal injection of DHA and the eyeballs were processed to morphologic and morphometric histological examination by light microscopy. At the same time aqueous and vitreous humor samples were taken to quantify the concentration of omega-3 acids by gas chromatography. Statistical analysis was performed by SPSS 21.0.

Results

Slit-lamp examination revealed an important inflammatory reaction on the anterior chamber of the rabbits injected with the higher concentrations of DHA (10 mg/50 µl, 5 mg/50 µl, 2''5 mg/50 µ) Lower concentrations showed no inflammation. Electroretinography and histological studies showed no significant difference between control and DHA-injected groups except for the group injected with 50 µg/50 µl.

Conclusions

Our results indicate that administration of intravitreal DHA is safe in the albino rabbit model up to the maximum tolerated dose of 25 µg/50 µl. Further studies should be performed in order to evaluate the effect of intravitreal injection of DHA as a treatment, alone or in combination, of different retinal diseases.     

Improved Methods of Carnivore Faecal Sample Preservation,DNA Extraction and Quantification for Accurate Genotyping of Wild Tigers

Background

Non-invasively collected samples allow a variety of genetic studies on endangered and elusive species. However due to low amplification success and high genotyping error rates fewer samples can be identified up to the individual level. Number of PCRs needed to obtain reliable genotypes also noticeably increase.

Methods

We developed a quantitative PCR assay to measure and grade amplifiable nuclear DNA in feline faecal extracts. We determined DNA degradation in experimentally aged faecal samples and tested a suite of pre-PCR protocols to considerably improve DNA retrieval.

Results

Average DNA concentrations of Grade I, II and III extracts were 982pg/µl, 9.5pg/µl and 0.4pg/µl respectively. Nearly 10% of extracts had no amplifiable DNA. Microsatellite PCR success and allelic dropout rates were 92% and 1.5% in Grade I, 79% and 5% in Grade II, and 54% and 16% in Grade III respectively. Our results on experimentally aged faecal samples showed that ageing has a significant effect on quantity and quality of amplifiable DNA (p<0.001). Maximum DNA degradation occurs within 3 days of exposure to direct sunlight. DNA concentrations of Day 1 samples stored by ethanol and silica methods for a month varied significantly from fresh Day 1 extracts (p<0.1 and p<0.001). This difference was not significant when samples were preserved by two-step method (p>0.05). DNA concentrations of fresh tiger and leopard faecal extracts without addition of carrier RNA were 816.5pg/µl (±115.5) and 690.1pg/µl (±207.1), while concentrations with addition of carrier RNA were 49414.5pg/µl (±9370.6) and 20982.7pg/µl (±6835.8) respectively.

Conclusions

Our results indicate that carnivore faecal samples should be collected as freshly as possible, are better preserved by two-step method and should be extracted with addition of carrier RNA. We recommend quantification of template DNA as this facilitates several downstream protocols.     

Iron,Hepcidin and Inflammatory Status of Young Healthy Overweight and Obese Women in Australia

Background and Aims

Evidence suggests obesity-related inflammation alters iron metabolism potentially increasing the risk of iron deficiency. This cross-sectional study aimed to investigate iron, hepcidin and inflammatory status in young, healthy overweight and obese women.

Methods

114 young (18–25 years), healthy comorbidity-free women with a body mass index (BMI) ≥27.5 kg/m² were recruited. Biochemical data were analysed using mean ± standard deviation or median (interquartile range) and multivariate modelling. Biochemical markers were also stratified according to varying degrees of overweight and obesity.

Results

Anaemia (haemoglobin <120 g/l) and iron deficiency (serum ferritin <15.0 µg/l) were prevalent in 10% and 17% of participants respectively. Mean/median soluble transferrin receptor was 1.61±0.44 mg/l; hepcidin 6.40 (7.85) ng/ml and C-reactive protein (CRP) 3.58 (5.81) mg/l. Multivariate modelling showed that BMI was a significant predictor of serum iron (coefficient = -0.379; standard error = 0.139; p = 0.008), transferrin saturation (coefficient = -0.588; standard error = 0.222; p = 0.009) and CRP (coefficient = 0.127; standard error = 0.024; p<0.001). Stratification of participants according to BMI showed those with ≥35.0 kg/m² had significantly higher CRP (p<0.001) than those in lower BMI categories.

Conclusions

Increasing obesity was associated with minor disturbances in iron metabolism. However, overall outcomes indicated simple iron deficiency (hypoferritinaemia) was the primary iron-related abnormality with no apparent contribution of inflammation or hepcidin, even in those with BMI >35.0 kg/m². This indicates that obesity alone may not be sufficient to induce clinically significant disturbances to iron metabolism as previously described. This may be attributed to the lack of comorbidity in this cohort.     

Crovirin,a Snake Venom Cysteine-Rich Secretory Protein (CRISP) with Promising Activity against Trypanosomes and Leishmania

Background

The neglected human diseases caused by trypanosomatids are currently treated with toxic therapy with limited efficacy. In search for novel anti-trypanosomatid agents, we showed previously that the Crotalus viridis viridis (Cvv) snake venom was active against infective forms of Trypanosoma cruzi. Here, we describe the purification of crovirin, a cysteine-rich secretory protein (CRISP) from Cvv venom with promising activity against trypanosomes and Leishmania.

Methodology/Principal Findings

Crude venom extract was loaded onto a reverse phase analytical (C8) column using a high performance liquid chromatographer. A linear gradient of water/acetonitrile with 0.1% trifluoroacetic acid was used. The peak containing the isolated protein (confirmed by SDS-PAGE and mass spectrometry) was collected and its protein content was measured. T. cruzi trypomastigotes and amastigotes, L. amazonensis promastigotes and amastigotes and T. brucei rhodesiense procyclic and bloodstream trypomastigotes were challenged with crovirin, whose toxicity was tested against LLC-MK₂ cells, peritoneal macrophages and isolated murine extensor digitorum longus muscle. We purified a single protein from Cvv venom corresponding, according to Nano-LC MS/MS sequencing, to a CRISP of 24,893.64 Da, henceforth referred to as crovirin. Human infective trypanosomatid forms, including intracellular amastigotes, were sensitive to crovirin, with low IC₅₀ or LD₅₀ values (1.10–2.38 µg/ml). A considerably higher concentration (20 µg/ml) of crovirin was required to elicit only limited toxicity on mammalian cells.

Conclusions

This is the first report of CRISP anti-protozoal activity, and suggests that other members of this family might have potential as drugs or drug leads for the development of novel agents against trypanosomatid-borne neglected diseases.     

Amiodarone Inhibits Apamin-Sensitive Potassium Currents

Background

Apamin sensitive potassium current (I _KAS), carried by the type 2 small conductance Ca²⁺-activated potassium (SK2) channels, plays an important role in post-shock action potential duration (APD) shortening and recurrent spontaneous ventricular fibrillation (VF) in failing ventricles.

Objective

To test the hypothesis that amiodarone inhibits I _KAS in human embryonic kidney 293 (HEK-293) cells.

Methods

We used the patch-clamp technique to study I _KAS in HEK-293 cells transiently expressing human SK2 before and after amiodarone administration.

Results

Amiodarone inhibited I_KAS in a dose-dependent manner (IC₅₀, 2.67±0.25 µM with 1 µM intrapipette Ca²⁺). Maximal inhibition was observed with 50 µM amiodarone which inhibited 85.6±3.1% of I_KAS induced with 1 µM intrapipette Ca²⁺ (n = 3). I_KAS inhibition by amiodarone was not voltage-dependent, but was Ca²⁺-dependent: 30 µM amiodarone inhibited 81.5±1.9% of I _KAS induced with 1 µM Ca²⁺ (n = 4), and 16.4±4.9% with 250 nM Ca²⁺ (n = 5). Desethylamiodarone, a major metabolite of amiodarone, also exerts voltage-independent but Ca²⁺ dependent inhibition of I _KAS.

Conclusion

Both amiodarone and desethylamiodarone inhibit I _KAS at therapeutic concentrations. The inhibition is independent of time and voltage, but is dependent on the intracellular Ca²⁺ concentration. SK2 current inhibition may in part underlie amiodarone''s effects in preventing electrical storm in failing ventricles.     

Relationships between Levels of Serum IgE,Cell-Bound IgE,and IgE-Receptors on Peripheral Blood Cells in a Pediatric Population

Background

Elevated serum immunoglobulin (Ig) E is a diagnostic marker of immediate-type allergic reactions. We hypothesize that serum IgE does not necessarily reflect total body IgE because in vivo IgE can be bound to cell surface receptors such as FcεRI and FcεRII (CD23). The aim of this study was to analyze the relationships between levels of serum IgE, cell-bound IgE, and IgE-receptors on peripheral blood cells in a pediatric population.

Methodology

Whole blood samples from 48 children (26 boys, 22 girls, mean age 10,3±5,4 years) were analyzed by flow cytometry for FcεRI, CD23, and cell-bound IgE on dendritic cells (CD11c+MHC class II+), monocytes (CD14+), basophils (CD123+MHC class II-) and neutrophils (myeloperoxidase+). Total serum IgE was measured by ELISA and converted into z-units to account for age-dependent normal ranges. Correlations were calculated using Spearman rank correlation test.

Principal Findings

Dendritic cells, monocytes, basophils, and neutrophils expressed the high affinity IgE-receptor FcεRI. Dendritic cells and monocytes also expressed the low affinity receptor CD23. The majority of IgE-receptor positive cells carried IgE on their surface. Expression of both IgE receptors was tightly correlated with cell-bound IgE. In general, cell-bound IgE on FcεRI+ cells correlated well with serum IgE. However, some patients carried high amounts of cell-bound IgE despite low total serum IgE levels.

Conclusion/Significance

In pediatric patients, levels of age-adjusted serum IgE, cell-bound IgE, and FcεRI correlate. Even in the absence of elevated levels of serum IgE, cell-bound IgE can be detected on peripheral blood cells in a subgroup of patients.     

CD4 Cell Count and the Risk of AIDS or Death in HIV-Infected Adults on Combination Antiretroviral Therapy with a Suppressed Viral Load: A Longitudinal Cohort Study from COHERE

Background

Most adults infected with HIV achieve viral suppression within a year of starting combination antiretroviral therapy (cART). It is important to understand the risk of AIDS events or death for patients with a suppressed viral load.

Methods and Findings

Using data from the Collaboration of Observational HIV Epidemiological Research Europe (2010 merger), we assessed the risk of a new AIDS-defining event or death in successfully treated patients. We accumulated episodes of viral suppression for each patient while on cART, each episode beginning with the second of two consecutive plasma viral load measurements <50 copies/µl and ending with either a measurement >500 copies/µl, the first of two consecutive measurements between 50–500 copies/µl, cART interruption or administrative censoring. We used stratified multivariate Cox models to estimate the association between time updated CD4 cell count and a new AIDS event or death or death alone. 75,336 patients contributed 104,265 suppression episodes and were suppressed while on cART for a median 2.7 years. The mortality rate was 4.8 per 1,000 years of viral suppression. A higher CD4 cell count was always associated with a reduced risk of a new AIDS event or death; with a hazard ratio per 100 cells/µl (95% CI) of: 0.35 (0.30–0.40) for counts <200 cells/µl, 0.81 (0.71–0.92) for counts 200 to <350 cells/µl, 0.74 (0.66–0.83) for counts 350 to <500 cells/µl, and 0.96 (0.92–0.99) for counts ≥500 cells/µl. A higher CD4 cell count became even more beneficial over time for patients with CD4 cell counts <200 cells/µl.

Conclusions

Despite the low mortality rate, the risk of a new AIDS event or death follows a CD4 cell count gradient in patients with viral suppression. A higher CD4 cell count was associated with the greatest benefit for patients with a CD4 cell count <200 cells/µl but still some slight benefit for those with a CD4 cell count ≥500 cells/µl. Please see later in the article for the Editors'' Summary     

Serum Testosterone Levels and Androgen Receptor CAG Polymorphism Correlate with Hepatitis B Virus (HBV)-Related Acute Liver Failure in Male HBV Carriers

Background

Augmentation of androgen/androgen receptor (AR) pathway may influence chronic hepatitis B (CHB) more likely in males. AR activity is modulated by a polymorphic CAG repeat sequence in AR exon 1. This study aimed to investigate the relationship between serum testosterone levels, CAG repeat numbers and hepatitis B virus (HBV)-related acute liver failure (ALF).

Methods

Three hundred and seventy eight male CHB patients with ALF and 441 asymptomatic HBV carriers (AsCs) were recruited. AR CAG repeats numbers were analyzed. The serum testosterone levels of AsCs, ALFs and patients with hepatitis B flare groups, and sequential serum samples, were assessed quantitatively.

Results

The median CAG repeat (M-CAG) frequency was significantly higher in ALF patients than AsCs (P<0.001). Patients with M-CAG alleles (P<0.001, OR 3.0, 95% CI 2.1–4.2) had the highest risk for ALF. Serum testosterone levels were significantly higher (P<0.001) at hepatitis flare point (8.2±3.0 ng/mL) than inactive phase (6.4±2.0 ng/mL). CHB (8.30±2.71 ng/mL, P = 7.6×10⁻⁶) and ALF group (2.61±1.83 ng/mL, P = 1.7×10⁻¹⁷) had significantly different levels of testosterone in comparison with AsCs group (6.56±2.36 ng/mL). The serum testosterone levels sharply decreased from hepatitis flare phase to liver failure phase, and tended to be normal at the recovery phase. Male AsCs with M-CAG alleles had significantly lower serum testosterone levels (P<0.05).

Conclusions

There was a serum testosterone fluctuation during hepatitis B flare and HBV-related ALF, and the median CAG repeats in AR gene exon 1 were associated with lower serum testosterone levels in asymptomatic HBV carriers and an increased susceptibility to HBV-related ALF.     

A Novel Immunological Assay for Hepcidin Quantification in Human Serum

Background

Hepcidin is a 25-aminoacid cysteine-rich iron regulating peptide. Increased hepcidin concentrations lead to iron sequestration in macrophages, contributing to the pathogenesis of anaemia of chronic disease whereas decreased hepcidin is observed in iron deficiency and primary iron overload diseases such as hereditary hemochromatosis. Hepcidin quantification in human blood or urine may provide further insights for the pathogenesis of disorders of iron homeostasis and might prove a valuable tool for clinicians for the differential diagnosis of anaemia. This study describes a specific and non-operator demanding immunoassay for hepcidin quantification in human sera.

Methods and Findings

An ELISA assay was developed for measuring hepcidin serum concentration using a recombinant hepcidin25-His peptide and a polyclonal antibody against this peptide, which was able to identify native hepcidin. The ELISA assay had a detection range of 10–1500 µg/L and a detection limit of 5.4 µg/L. The intra- and interassay coefficients of variance ranged from 8–15% and 5–16%, respectively. Mean linearity and recovery were 101% and 107%, respectively. Mean hepcidin levels were significantly lower in 7 patients with juvenile hemochromatosis (12.8 µg/L) and 10 patients with iron deficiency anemia (15.7 µg/L) and higher in 7 patients with Hodgkin lymphoma (116.7 µg/L) compared to 32 age-matched healthy controls (42.7 µg/L).

Conclusions

We describe a new simple ELISA assay for measuring hepcidin in human serum with sufficient accuracy and reproducibility.     

Serum Interleukin-18 Levels Are Associated with Physical Activity in Japanese Men

Objective

To investigate the link between serum interleukin-18 (IL-18) levels and physical activity in Japanese men.

Methods

A total of 81 men (45.7±17.6 years old) was enrolled in this cross-sectional investigation study. We assessed anthropometric and body composition parameters. Serum IL-18 levels, physical activity by uniaxial accelerometers, peak oxygen uptake and metabolic risk parameters were also evaluated.

Results

Serum IL-18 levels were 179.4±84.7 pg/mL. Physical activity evaluated by Σ[metabolic equivalents × h per week (METs⋅h/w)]was significantly and negatively correlated with serum IL-18 levels (r = −0.252, p = 0.0235). These associations remained even after adjusting for age, peak oxygen uptake and other confounding factors.

Conclusion

Serum IL-18 levels were closely associated with physical activity independent of peak oxygen uptake in Japanese men.     

A Pro-Inflammatory Role of C5L2 in C5a-Primed Neutrophils for ANCA-Induced Activation

Background

The complement system is crucial for the development of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In particular, C5a and its receptor on neutrophils, CD88, play a central role. The functional role of the second receptor of C5a, C5L2, remains unclear. In the current study, we investigated the role of C5L2 in C5a-primed neutrophils for ANCA-induced activation.

Methods

The effect of blocking C5L2 by anti-human C5L2 blocking antibody were tested on respiratory burst and degranulation of C5a-primed neutrophils activated with ANCA, as well as on membrane-bound proteinase 3 (mPR3) and concentration of myeloperoxidase (MPO) in supernatant of C5a-primed neutrophils. An antagonist for CD88 was also employed.

Results

Blocking C5L2 resulted in a significantly decreased MPO concentration in the supernatant of C5a-primed neutrophils. mPR3 expression increased from 209.0±43.0 in untreated cells to 444.3±60.8 after C5a treatment (P<0.001), and decreased to 375.8±65.44, 342.2±54.3 and 313.7±43.6 by pre-incubating blocking C5L2 antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-priming group, P<0.001, P<0.001, and P<0.001), respectively. In C5a-primed neutrophils, subsequently activating with MPO-ANCA-positive IgG, the MFI value was 425.8±160.6, which decreased to 292.8±141.2, 289.7±130.0 and 280.3±136.4 upon pre-incubation with mouse anti-human C5L2 blocking antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-primed neutrophils, for MPO-ANCA-positive IgG-induced activation, P<0.05, P<0.05, and P<0.05), respectively. Blocking C5L2 also resulted in significantly decreased C5a-primed neutrophils for PR3-ANCA-positive IgG-induced activation. Moreover, the lactoferrin concentration in the supernant significantly decreased in pre-incubation with anti-human C5L2 blocking antibody, compared with C5a-primed neutrophils induced by PR3- or MPO-ANCA-positive IgG.

Conclusions

C5L2 may be implicated in the pro-inflammatory role in C5a-primed neutrophils for ANCA-induced activation.     

Ambient Stable Quantitative PCR Reagents for the Detection of Yersinia pestis

Background

Although assays for detecting Yersinia pestis using TaqMan probe-based real-time PCR have been developed for years, little is reported on room-temperature-stable PCR reagents, which will be invaluable for field epidemic surveillance, immediate response to public health emergencies, counter-bioterrorism investigation, etc. In this work, a set of real-time PCR reagents for rapid detection of Y. pestis was developed with extraordinary stability at 37°C.

Methods/Principal Findings

TaqMan-based real-time PCR assays were developed using the primers and probes targeting the 3a sequence in the chromosome and the F1 antigen gene caf1 in the plasmid pMT1of Y. pestis, respectively. Then, carbohydrate mixtures were added to the PCR reagents, which were later vacuum-dried for stability evaluation. The vacuum-dried reagents were stable at 37°C for at least 49 days for a lower concentration of template DNA (10 copies/µl), and up to 79 days for higher concentrations (≥10² copies/µl). The reagents were used subsequently to detect soil samples spiked with Y. pestis vaccine strain EV76, and 5×10⁴ CFU per gram of soil could be detected by both 3a- and caf1-based PCR reagents. In addition, a simple and efficient method for soil sample processing is presented here.

Conclusions/Significance

The vacuum-dried reagents for real-time PCR maintain accuracy and reproducibility for at least 49 days at 37°C, indicating that they can be easily transported at room temperature for field application if the machine for performing real-time PCR is available. This dry reagent is of great significance for routine plague surveillance.     

Physical and Perceptual Cooling with Beverages to Increase Cycle Performance in a Tropical Climate

Purpose

This study compares the effects of neutral temperature, cold and ice-slush beverages, with and without 0.5% menthol on cycling performance, core temperature (T_co) and stress responses in a tropical climate (hot and humid conditions).

Methods

Twelve trained male cyclists/triathletes completed six 20-km exercise trials against the clock in 30.7°C±0.8°C and 78%±0.03% relative humidity. Before and after warm-up, and before exercise and every 5 km during exercise, athletes drank 190 mL of either aromatized (i.e., with 0.5 mL of menthol (5 gr/L)) or a non-aromatized beverage (neutral temperature: 23°C±0.1°C, cold: 3°C±0.1°C, or ice-slush: −1°C±0.7°C). During the trials, heart rate (HR) was continuously monitored, whereas core temperature (T_co), thermal comfort (TC), thermal sensation (TS) and rate of perceived exertion (RPE) were measured before and after warm-up, every 5 km of exercise, and at the end of exercise and after recovery.

Results

Both the beverage aroma (P<0.02) and beverage temperature (P<0.02) had significant and positive effects on performance, which was considerably better with ice-slush than with a neutral temperature beverage, whatever the aroma (P<0.002), and with menthol vs non-menthol (P<0.02). The best performances were obtained with ice-slush/menthol and cold/menthol, as opposed to neutral/menthol. No differences were noted in HR and T_co between trials.

Conclusion

Cold water or ice-slush with menthol aroma seems to be the most effective beverage for endurance exercise in a tropical climate. Further studies are needed to explore its effects in field competition.