Adult Plasma Research Articles

Fresh frozen plasma transfusion – a risk factor for pulmonary hemorrhage in extremely low birth weight infants?

To evaluate risk factors for pulmonary hemorrhage (PH) in extremely low birth weight infants (ELBW) taking into consideration coagulation screens, platelet counts, transfusion of fresh frozen plasma (FFP), and platelet concentrates prior to PH. A retrospective case-control study consisting of 20 ELBW infants with PH and 40 matched controls. Coagulation screens, platelet counts at birth and at onset of PH, and transfusion frequencies prior to PH were compared to case-controls at birth and 24-96 h after birth. While the initial platelet counts, fibrinogen concentrations, and international normalized ratios were similar in PH infants and controls, the activated partial prothrombin time was prolonged (P=0.05). Compared to 28% of case controls (P<0.05), 55% of infants with later PH received FFP prior to PH. Platelet counts were significantly lower at onset of PH (median 81/nL; range: 37-236/nL) compared to controls (166/nL; 27-460/nL; P<0.005). Multivariate analysis indicated a lack of antenatal steroids, supplemental oxygen, and transfusion of FFP as independent risk factors for PH. Prolonged activated partial thromboplastin time (aPTT) might be associated with PH. PH does not primarily depend upon severe thrombocytopenia. A developmental mismatch in hemostasis by transfusion of adult donor plasma should be considered a risk factor for PH.

Effect of Ichthyophonus on blood plasma chemistry of spawning Chinook salmon and their resulting offspring in a Yukon River tributary.

Ichthyophonus is a protozoan parasite of Alaska Chinook salmon Oncorhynchus tshawytscha. In this study, we determined whether spawning Chinook salmon in the Yukon River drainage exhibited a measurable stress response (i.e. elevated plasma cortisol concentrations) and detectable changes in selected blood plasma chemistry parameters when infected with Ichthyophonus. The resulting alevin were also analyzed for any differences in blood plasma chemistry caused by parental infection with Ichthyophonus. In 2010, 2011, and 2012, spawning adult Chinook salmon were collected from the Salcha River, Alaska, USA, and the prevalence of Ichthyophonus in these fish was 7.8, 6.3, and 8.3%, respectively. Fish with no clinical signs of Ichthyophonus and Ichthyophonus-positive parents were cross-fertilized to investigate potential second-generation effects as a result of Ichthyophonus infection. We found no significant difference in cortisol concentrations in blood plasma between Ichthyophonus-positive and -negative adults or between alevin from Ichthyophonus-positive and -negative parents. There were no significant differences in blood plasma parameters (e.g. alanine aminotransferase, creatine kinase, glucose) of Ichthyophonus-negative and -positive adults, with the exception of aspartate aminotransferase, which was significantly higher in plasma of Ichthyophonus-negative adults. All clinical chemistry parameters for alevin resulting from both Ichthyophonus-negative and -positive parents were not significantly different. Based on this study, which has a limited sample size and low prevalence of Ichthyophonus, offspring of Chinook salmon appear to suffer no disadvantage as a result of Ichthyophonus infection in their parents on the Salcha River.

Fetuin-A: A Biomarker for Type II Diabetes Mellitus

Fetuins are blood glycoproteins manufactured in the liver and secreted into the bloodstream. These glycoproteins make up a large group of binding proteins which mediates the transit and presence of a myriad of substances in the blood stream. Serum albumin, which is the most abundant protein in adult animal plasma is best known as the representative of these carrier proteins. In bone homeostasis, it is the circulating glycoprotein that plays a critical role. It plays major roles also in prevention of vascular calcification, disruption of adipocyte function and impairment of insulin signaling. Even with its major role, it is highly dependable in preventing and/or amplifying of the above disease processes. Certain diseases have been associated with high levels of Fetuin A. Although low levels of it in the plasma assist potentially in the protective effect of artery calcification in non-Chronic Kidney Disease (CKD) patients, high levels of the glycoprotein are the greatest concern in patients at higher risk of cardiovascular disease (CVD) and diabetes. The range of serum fetuin A in healthy adults is between 0.4 and 1mg/ml serum. The most studied function of fetuin A is mediated by the D1 domain. But the domain that binds with insulin receptor is yet to be known. The B chain consists of 27 amino acid residues which are distributed unevenly among the charged and neutral portion. Fetuins are proteins highly expressed in the liver blood plasma. They bear post translational modifications in proteolytic processing, phosphorylation, complex glycosylation and sulfation. The precussors of Fetuin A in humans are single-chained to the mature circulating double-chain form. In septicaemia and bovine, human fetuin A is perceptible to further proteolytic cleavage. Since the discovery of Fetuin A as a glycoprotein that inhibits vascular calcification in early 1990s, the biologic attributed roles has increased and still increasing exponentially. Apart from the roles it plays in type II diabetes mellitus and cardiovascular disease, other roles have been noted. Other effects and roles of Fetuin A are still being researched on.

Cell-Free DNA Is Elevated after Acute Arterial Injury in Infants

Purpose: Circulating cell-free DNA in blood may originate from cell necrosis, apoptosis, or NETosis. NETosis is a recently described inflammatory process in which neutrophils release a meshwork of DNA fibers called neutrophil extracellular traps (NETs) to attack microorganisms. Higher levels of cell-free DNA have been observed in patients with sepsis and malignancy, and it may be linked to arterial thrombosis as well. We examined changes in cell-free DNA in infants who experienced acute, iatrogenic thrombosis of the femoral artery after arterial catheterization to better understand the role of cell-free DNA in clot formation.Methods: Plasma samples were collected from infants < 1 year of age with congenital heart disease during cardiac catheterization. Post-procedurally, femoral artery thrombus formation at the site of sheath insertion was detected by ultrasound. Cell-free DNA was quantified at two time points (before and after arterial catheterization) in three infants who developed femoral artery thrombus after catheterization and three age-matched controls. Cell-free DNA was also quantified from plasma of normal adults as a comparison.Results: At baseline,cell-free DNA level for the infant cohort was similar to that in normal adults (13.9 ng/mL vs. 10.3 ng/mL, p = 0.20, Figure). After arterial catheterization, cell-free DNA was increased from baseline in all infants (51.0 ng/mL vs. 13.9 ng/ml, p = 0.004). However, infants who developed femoral artery thrombus after catheterization did not demonstrate higher cell-free DNA after arterial injury, compared to the control group (47.1 ng/mL vs. 54.9 ng/ml, p = 0.83).Conclusions: Cell-free DNA is significantly and consistently elevated in infants after arterial catheterization. As part of NETosis, cell-free DNA may serve as an inflammatory mediator in infants who experience arterial injury. We plan to confirm the findings of this pilot study in a larger human infant cohort as well as in a piglet model of vascular injury. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Bleeding Problems in Extremely Low Birth Weight Neonates: Quick (and Wintrobe) Thinking Needed

Bleeding complications occur all too commonly among extremely low birth weight (ELBW) neonates. Although sometimes the bleeding is minimal and transient, some hemorrhages are life-altering or life-ending events. Compared with term neonates, ELBW neonates typically have lower platelet counts, reduced platelet function (when measured with in vitro testing), and prolonged coagulation times. These laboratory findings have led to attempts to “correct” the “immature” hemostatic systems of ELBW neonates by prophylactically transfusing adult donor platelets and/or adult plasma to nonbleeding neonates in the hope of reducing their bleeding risk. Although well-meaning and consistent with laboratory normal values, this approach has been ineffectual. In this review, we seek to teach basic philosophies used by 2 pioneers of hematology, Drs Quick and Wintrobe. We apply their principles to the population of ELBW infants. We also review practical steps that neonatologists can take to reduce the risk of hemorrhagic problems, particularly intracranial hemorrhages, that occur in ELBW neonates.

Diagnostic and prognostic significance of systemic alkyl quinolones for P. aeruginosa in cystic fibrosis: A longitudinal study

BackgroundPulmonary P. aeruginosa infection is associated with poor outcomes in cystic fibrosis (CF) and early diagnosis is challenging, particularly in those who are unable to expectorate sputum. Specific P. aeruginosa 2-alkyl-4-quinolones are detectable in the sputum, plasma and urine of adults with CF, suggesting that they have potential as biomarkers for P. aeruginosa infection. AimTo investigate systemic 2-alkyl-4-quinolones as potential biomarkers for pulmonary P. aeruginosa infection. MethodsA multicentre observational study of 176 adults and 68 children with CF. Cross-sectionally, comparisons were made between current P. aeruginosa infection using six 2-alkyl-4-quinolones detected in sputum, plasma and urine against hospital microbiological culture results. All participants without P. aeruginosa infection at baseline were followed up for one year to determine if 2-alkyl-4-quinolones were early biomarkers of pulmonary P. aeruginosa infection. ResultsCross-sectional analysis: the most promising biomarker with the greatest diagnostic accuracy was 2-heptyl-4-hydroxyquinoline (HHQ). In adults, areas under the ROC curves (95% confidence intervals) for HHQ analyses were 0.82 (0.75–0.89) in sputum, 0.76 (0.69–0.82) in plasma and 0.82 (0.77–0.88) in urine. In children, the corresponding values for HHQ analyses were 0.88 (0.77–0.99) in plasma and 0.83 (0.68–0.97) in urine.Longitudinal analysis: Ten adults and six children had a new positive respiratory culture for P. aeruginosa in follow-up. A positive plasma HHQ test at baseline was significantly associated with a new positive culture for P. aeruginosa in both adults and children in follow-up (odds ratio (OR)=6.67;-95% CI:-1.48–30.1;-p=0.01 and OR=70; 95% CI: 5–956;-p<0.001 respectively). ConclusionsAQs measured in sputum, plasma and urine may be used to diagnose current infection with P. aeruginosa in adults and children with CF. These preliminary data show that plasma HHQ may have potential as an early biomarker of pulmonary P. aeruginosa. Further studies are necessary to evaluate if HHQ could be used in clinical practice to aid early diagnosis of P. aeruginosa infection in the future.

Homocysteine metabolism is associated with cerebrospinal fluid levels of soluble amyloid precursor protein and amyloid beta.

Disturbed homocysteine metabolism may contribute to amyloidogenesis by modulating the amyloid precursor protein (APP) production and processing. The objective of this study was to investigate the relationships between cerebral amyloid production and both blood and cerebrospinal fluid (CSF) markers of the homocysteine metabolism. We assessed CSF concentrations of soluble APPα, soluble APPβ, and amyloid β1-42 (Aβ1-42), as well as plasma levels of homocysteine (Hcys), total vitamin B12, and folate, and CSF concentrations of homocysteine (Hcys-CSF), 5-methyltetrahydrofolate (5-MTHF), S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) in 59 subjects with normal cognition. Linear regression analyses were performed to assess associations between homocysteine metabolism parameters and amyloid production. The study was approved by the Ethical Committee of the University of Bonn. After controlling for age, gender, APOEe4 status, and albumin ratio (Qalb), higher Aβ1-42 CSF levels were associated with high Hcys and low vitamin B12 plasma levels as well as with high Hcys, high SAH, and low 5-MTHF CSF levels. Higher CSF concentrations of sAPPα and sAPPβ were associated with high SAH levels. The results suggest that disturbed homocysteine metabolism is related to increased CSF levels of sAPP forms and Aβ1-42, and may contribute to the accumulation of amyloid pathology in the brain. Disturbed homocysteine metabolism may contribute to amyloidogenesis by modulating the amyloid precursor protein (APP) production and processing. We found associations between CSF levels of soluble APP forms and Aβ1-42, and markers of the homocysteine metabolism in both plasma and CSF in adults with normal cognition. Disturbed homocysteine metabolism may represent a target for preventive and early disease-modifying interventions in Alzheimer's disease.

Patient age does not affect mefloquine concentrations in erythrocytes and plasma during the acute phase of falciparum malaria

ObjectiveTo evaluate whether patient age has a significant impact on mefloquine concentrations in the plasma and erythrocytes over the course of treatment for uncomplicated falciparum malaria. MethodsA total of 20 children aged between 8 and 11 years and 20 adult males aged between 22 and 41 years with uncomplicated falciparum malaria were enrolled in the study. Mefloquine was administered to patients in both age groups at a dose of 20mgkg−1. The steady-state drug concentrations were measured by reversed-phase high performance liquid chromatography. ResultsAll patients had an undetectable mefloquine concentration on day 0. In adults, the plasma mefloquine concentrations ranged from 770 to 2930ngmL−1 and the erythrocyte concentrations ranged from 2000 to 6030ngmL−1. In children, plasma mefloquine concentrations ranged from 881 to 3300ngmL−1 and erythrocyte concentrations ranged from 3000 to 4920ngmL−1. There was no significant correlation between mefloquine concentrations in the plasma and erythrocytes in either adults or children. ConclusionIn the present study, we observed no effect of patient age on the steady-state concentrations of mefloquine in the plasma and erythrocytes. We found that the mefloquine concentration in the erythrocytes was approximately 2.8-times higher than in the plasma. There were no significant correlations between mefloquine concentrations in the erythrocytes and plasma for either age group.

Blood biomarkers indicate mild neuroaxonal injury and increased amyloid β production after transient hypoxia during breath-hold diving

Objective: To determine whether transient hypoxia during breath-hold diving causes neuronal damage or dysfunction or alters amyloid metabolism as measured by certain blood biomarkers.Design: Sixteen divers competing in the national Swedish championship in breath-hold diving and five age-matched healthy control subjects were included. Blood samples were collected at baseline and over a course of 3 days where the divers competed in static apnea (STA), dynamic apnea without fins (DYN1) and dynamic apnea with fins (DYN2).Main outcomes: Biomarkers reflecting brain injury and amyloid metabolism were analysed in serum (S-100β, NFL) and plasma (T-tau, Aβ42) using immunochemical methods.Results: Compared to divers’ baseline, Aβ42 increased after the first event of static apnea (p = 0.0006). T-tau increased (p = 0.001) in STA vs baseline and decreased after one of the dynamic events, DYN2 (p = 0.03). Further, T-tau correlated with the length of the apneic time during STA (ρ = 0.7226, p = 0.004) and during DYN1 (ρ = 0.66, p = 0.01).Conclusion: The findings suggest that transient hypoxia may acutely increase the levels of Aβ42 and T-tau in plasma of healthy adults, further supporting that general hypoxia may cause mild neuronal dysfunction or damage and stimulate Aβ production.

Postprandial Inflammatory Responses and Free Fatty Acids in Plasma of Adults Who Consumed a Moderately High-Fat Breakfast with and without Blueberry Powder in a Randomized Placebo-Controlled Trial

Background: Saturated fatty acids (FAs) released from triglyceride-rich lipoproteins (TGRLs) activate Toll-like receptor 2 (TLR-2) and induce the expression of proinflammatory cytokines in monocytes. Certain plant polyphenols inhibit TLR-mediated signaling pathways.Objective: We determined whether plasma free FAs (FFAs) after a moderately high-fat (MHF, 40% kcal from fat) breakfast modulate the inflammatory status of postprandial blood, and whether blueberry intake suppresses FFA-induced inflammatory responses in healthy humans.Methods: Twenty-three volunteers with a mean ± SEM age and body mass index (in kg/m2) of 30 ± 3 y and 21.9 ± 0.4, respectively, consumed an MHF breakfast with either a placebo powder or 2 or 4 servings of blueberry powder in a randomized crossover design. The placebo powder was provided on the first test day and the blueberry powder doses were randomized with a 2-wk washout period. Plasma concentrations of lipids, glucose, and cytokines were determined. To determine whether FFAs derived from TGRL stimulate monocyte activation, and whether this is inhibited by blueberry intake, whole blood was treated with lipoprotein lipase (LPL).Results: The median concentrations of FFAs and cytokines [tumor necrosis factor-α, interleukin (IL)-6 and IL-8] in postprandial plasma (3.5 h) decreased compared with fasting plasma regardless of the blueberry intake (P < 0.001 for FFAs and P < 0.05 for cytokines). However, concentrations of FFAs and cytokines including IL-1β increased in LPL-treated whole blood compared with untreated blood samples from participants who consumed the placebo powder. Blueberry intake suppressed IL-1β and IL-6 production in LPL-treated postprandial blood compared with the placebo control when fasting changes were used as a covariate.Conclusions: The plasma FFA concentration may be an important determinant affecting inflammatory cytokine production in blood. Supplementation with blueberry powder did not affect plasma FFA and cytokine concentrations; however, it attenuated the cytokine production induced by ex vivo treatment of whole blood with LPL. This trial was registered at clinicaltrials.gov as NCT01594008.

Postbooster Antibodies from Humans as Source of Diphtheria Antitoxin.

Diphtheria antitoxin for therapeutic use is in limited supply. A potential source might be affinity-purified antibodies originally derived from plasma of adults who received a booster dose of a vaccine containing diphtheria toxoid. These antibodies might be useful for treating even severe cases of diphtheria.

Maternally Administered Cyclic Glycine-Proline Increases Insulin-Like Growth Factor-1 Bioavailability and Novelty Recognition in Developing Offspring.

Cyclic glycine-proline (cGP), a metabolite of IGF-1, is an endogenous neuropeptide that improves memory in adult rats. The presence and concentrations of endogenous cGP, and its association with IGF-1 and IGF binding protein-3 (IGFBP-3) in rat milk and plasma, were evaluated during postnatal development. Maternal-infantile transfer of cGP during lactation and its efficacy on the memory of developing offspring were also investigated. Dams were gavaged with either cGP (3 mg/kg) or saline daily from postnatal days 8-22. Concentrations of cGP were measured in dams' milk, and concentrations of cGP, IGF-1, and IGFBP-3 were measured in the plasma of dams, pups, and young adults. The recognition memory, locomotor function, and anxiety-like behavior of offspring were evaluated using behavioral tests. Endogenous cGP was detected in rat milk, and its concentration was higher during peak lactation compared with late lactation. Comparisons within control groups showed low endogenous IGF-1 and IGFBP-3 and high endogenous cGP concentrations in the plasma of male pups. The reduced IGFBP-3 and increased cGP may be a response to increase the bioavailability of IGF-1 during infancy. Exogenous cGP showed oral bioavailability and effective maternal-infantile transfer through milk. Maternally transferred cGP also led to improved recognition memory in the developing offspring, possibly through increased IGF-1 bioavailability, with no effect on locomotor activity and anxiety-like behavior. These results show that cGP is an essential endogenous peptide during early postnatal development as it improves the bioavailability of IGF-1 during infancy. Furthermore, maternal cGP supplementation offers an effective and natural route of administration for improving memory in the developing offspring.

Chemometric analysis of attenuated total reflectance infrared spectral data for quantitation of immunoglobulin G in equine plasma and serum

Immunoglobulin G (IgG) is a crucial antibody to protect animals from invasion by microorganisms. Although there exist several methods in veterinary medicine to measure IgG levels for diagnostic and therapeutic purposes, these methods suffer from various weaknesses. Infrared (IR) spectroscopy coupled with chemometric tools such as principal component regression has been widely employed for the measurement of compounds in mixtures with the advantages that include simplicity, quickness and low test cost. Earlier investigation for IgG assay based on transmission IR spectroscopy using laboratory grade equipment has been conducted, but it is not readily transferrable to the clinic, hospital or small laboratory setting. More robust attenuated total reflectance (ATR) IR spectroscopy platforms have recently been developed for a range of roles in the field. This study investigated the possibility of using ATR-IR spectroscopy to determine the IgG concentrations in foal serum and adult horse plasma samples. The results of this work showed that immunoglobulin G concentrations predicted by ATR-IR spectroscopy with chemometric analysis had good agreement with those obtained from the radial immunodiffusion (RID) reference method. The precision of this approach was most compatible to RID method when the IgG concentration was high, but poorer for lower IgG concentrations. It was also showed that building a united calibration model for serum and plasma samples is likely. The results of this work indicate that ATR-IR spectroscopy coupled with chemometric analysis is a promising technique to measure the equine serum and plasma IgG concentrations in the veterinary clinical or hospital environment.

Differential Contributions of Intrinsic and Extrinsic Pathways to Thrombin Generation in Adult, Maternal and Cord Plasma Samples.

BackgroundThrombin generation (TG) is a pivotal process in achieving hemostasis. Coagulation profiles during pregnancy and early neonatal period are different from that of normal (non-pregnant) adults. In this ex vivo study, the differences in TG in maternal and cord plasma relative to normal adult plasma were studied.MethodsTwenty consented pregnant women and ten consented healthy adults were included in the study. Maternal and cord blood samples were collected at the time of delivery. Platelet-poor plasma was isolated for the measurement of TG. In some samples, anti-FIXa aptamer, RB006, or a TFPI inhibitor, BAX499 were added to elucidate the contribution of intrinsic and extrinsic pathway to TG. Additionally, procoagulant and inhibitor levels were measured in maternal and cord plasma, and these values were used to mathematically simulate TG.ResultsPeak TG was increased in maternal plasma (393.6±57.9 nM) compared to adult and cord samples (323.2±38.9 nM and 209.9±29.5 nM, respectively). Inhibitory effects of RB006 on TG were less robust in maternal or cord plasma (52% vs. 12% respectively) than in adult plasma (81%). Likewise the effectiveness of BAX499 as represented by the increase in peak TG was much greater in adult (21%) than in maternal (10%) or cord plasma (12%). Further, BAX499 was more effective in reversing RB006 in adult plasma than in maternal or cord plasma. Ex vivo data were reproducible with the results of the mathematical simulation of TG.ConclusionNormal parturient plasma shows a large intrinsic pathway reserve for TG compared to adult and cord plasma, while TG in cord plasma is sustained by extrinsic pathway, and low levels of TFPI and AT.

Increased levels of hyper-stable protein aggregates in plasma of older adults.

Proteins that misfold into hyper-stable/degradation-resistant species during aging may accumulate and disrupt protein homeostasis (i.e., proteostasis), thereby posing a survival risk to any organism. Using the method diagonal two-dimensional (D2D) SDS-PAGE, which separates hyper-stable SDS-resistant proteins at a proteomics level, we analyzed the plasma of healthy young (<30years) and older (60-80years) adults. We discovered the presence of soluble SDS-resistant protein aggregates in the plasma of older adults, but found significantly lower levels in the plasma of young adults. We identified the inflammation-related chaperone protein haptoglobin as the main component of the hyper-stable aggregates. This observation is consistent with the growing link between accumulations of protein aggregates and aging across many organisms. It is plausible higher amounts of SDS-resistant protein aggregates in the plasma of older adults may reflect a compromise in proteostasis that may potentially indicate cellular aging and/or disease risk. The results of this study have implications for further understanding the link between aging and the accumulation of protein aggregates, as well as potential for the development of aging-related biomarkers. More broadly, this novel application of D2D SDS-PAGE may be used to identify, quantify, and characterize the degradation-resistant protein aggregates in human plasma or any biological system.

Tracking isoflavones in whole soy flour, soy muffins and the plasma of hypercholesterolaemic adults

Information on changes in soy isoflavones during processing is needed to better understand the relationship between isoforms, food matrix, bioavailability and health. The abundance and transformation of isoflavones were tracked in soy flour, a baked soy muffin and plasma of hypercholesterolaemic adults (n = 142) who consumed muffins daily for 6 weeks. Isoflavones were identified and quantified in soy flour and muffins by HPLC-UV and in plasma by LC-MS/MS. Equol status was determined and used to assess the relationship between plasma isoflavones and dietary factors. Baking soy flour altered the relative proportion of isoflavone isoforms by promoting a conversion of malonyl- to β-glucosides in soy muffins (P < 0.001). Daily soy consumption resulted in >3-fold increase in plasma isoflavones with a doubling of dose (P < 0.001). Dietary factors did not correlate with plasma isoflavones. These findings may be used to guide the development of soy-based functional foods.

Hepatitis B Virus Surface Antigen Activates Myeloid Dendritic Cells via a Soluble CD14-Dependent Mechanism.

Hepatitis B virus (HBV) infection can cause chronic liver disease, which is associated with increased risk of liver cirrhosis, liver failure, and liver cancer. Clearance of HBV infection requires effective HBV-specific immunity; however, the immunological mechanisms that determine the development of effective HBV-specific immunity are poorly understood. Dendritic cells (DC) play a pivotal role in the regulation of antiviral immunity. Here, we investigated the interaction between HBV surface antigen (HBsAg), the main envelope glycoprotein of HBV, and BDCA1(+) myeloid dendritic cells (mDC). Exposure of peripheral blood-derived BDCA1(+) mDC to HBsAg resulted in strong DC maturation, cytokine production, and enhanced capacity to activate antigen-specific cytotoxic T cells (CTLs). By using neutralizing antibodies, crucial roles for CD14 and Toll-like receptor 4 (TLR4) in HBsAg-mediated BDCA1(+) mDC maturation were identified. Concordantly, HBsAg-mediated DC maturation required fetal calf serum (FCS) or human plasma, naturally containing soluble CD14 (sCD14). Intriguingly, HBsAg-induced DC maturation was significantly reduced in umbilical cord blood plasma, which contained less sCD14 than adult plasma, indicating that sCD14 is an important host factor for recognition of HBsAg by DC and subsequent DC activation. A direct interaction between sCD14 and HBsAg was demonstrated by using enzyme-linked immunosorbent assay (ELISA). Moreover, sCD14-HBsAg complexes were detected both in vitro and in sera of HBV-infected patients. The abundance of sCD14-HBsAg complexes varied between chronic HBV disease stages and correlated with activation of BDCA1(+) mDC in vivo We conclude that HBsAg activates BDCA1(+) DC via an sCD14-dependent mechanism. These findings provide important novel insights into the initiation of HBV-specific immunity and facilitate development of effective immunotherapeutic interventions for HBV. Hepatitis B virus (HBV) infection is a significant health problem, as it causes progressive liver injury and liver cancer in patients with chronic HBV infection, which affects approximately 250 million individuals worldwide. Some of the infected adults and the majority of neonates fail to mount an effective immune response and consequently develop chronic infection. The viral and host factors involved in the initiation of effective HBV-specific immune responses remain poorly understood. Here we identified CD14 and TLR4 as receptors for HBsAg, the main HBV envelope antigen. HBsAg induced strong maturation of dendritic cells (DC), which have a central role in regulation of virus-specific immunity. These results provide essential novel insights into the mechanisms underlying the initiation of HBV-specific immunity. Intriguingly, since neonates have naturally low sCD14, the finding that serum-derived sCD14 is a crucial host factor for recognition of HBsAg by DC may have implications for immunity of neonates to HBV infection.

Live imaging of hepatic and intestinal intracellular ApoA‐I in zebrafish

Elevated high‐density lipoprotein (HDL) cholesterol is incontrovertibly correlated with protection from cardiovascular disease in humans. The main structural component of HDL isapolipoprotein A‐I (APOA‐I). Cellular APOA‐I dynamics (e.g., transport, degradation, recycling) directly influence circulating APOA‐I and HDL levels. APOA‐Icell biology has remained largely unexplored, despite the strong link between cellular APOA‐I, circulating HDL, and cardiovascular disease risk. Furthermore, both the liver and the intestine synthesize HDL and APOA‐I, but the biological significance, and whether APOA‐I cell biology differs by tissue, is not currently understood. The optically clear larval zebrafish presents an excellent opportunity to visualize global and cellular APOA‐I dynamics in a live animal. Zebrafish have two ApoA‐I proteins; our insitu hybridization analysis shows ApoA‐Ia is expressed strongly in the intestine and weakly in the liver, and that ApoA‐Ib is expressed in the liver. We created transgenic zebrafish expressing fluorescently labeled human orzebrafish APOA‐I (APOA‐I‐mCherry) driven by liver‐ or intestine‐specific promoters to visualize APOA‐I of hepatic vs. intestinal origin in vivo. Agarose gel electrophoresis of adult zebrafish plasma indicates that the human and zebrafish APOA‐I‐mCherryfusion proteins are incorporated into HDL particles. Confocal microscopy of live larvae reveals that both secreted APOA‐I‐mCherry fusions localize to specific tissues and subcellular domains. We hypothesized that human APOA‐I‐mCherrylocalizes to the endosomal transport system of enterocytes and hepatocytes, and that due to the unique biology of hepatocytes and enterocytes, this trafficking may vary by cell type. To investigate this hypothesis, the human APOA‐I transgenic larvae were studied in genetic backgrounds that express Rab‐GFP fusion proteins, which mark specific endosomal compartments, on the ubiquitous, inducible HSP70 promoter. Using live imaging, we found that human APOA‐I‐mCherry co‐localizes with markers of early(GFP‐Rab5c), recycling (GFP‐Rab11a), and late endosomes/lysosomes (GFP‐Rab7, Lysotracker). This data also suggests that whether APOA‐I‐mCherry co‐localizes with these endosomal markers is influenced by both the tissue APOA‐I‐mCherry isderived from, and the tissue it is taken up by; for example, APOA‐I‐mCherry secreted from the intestine co‐localizes with recycling endosomes in the liver, but APOA‐I‐mCherry secreted from the liver does not co‐localize with recycling endosomes in the intestine. On‐going experiments will examine the route of intracellular APOA‐I transport and how it differs by tissue. In conclusion, we harnessed the larval zebrafish to visualize apolipoprotein dynamics at both the multi‐organ and subcellular levels. To our knowledge, this is the first time tissue‐specific apolipoprotein transport has been visualized in vivo.Support or Funding InformationThis research was supported by the NIH (DK093399 and NIDDK F32DK096786 (JO)), Carnegie Institution of Washington Endowment, and G. Harold and Leila Y. Mathers Charitable Foundation.

First steps in the harmonisation of adult plasma amino acid reference intervals in Australasia

First steps in the harmonisation of adult plasma amino acid reference intervals in Australasia

Pharmacokinetic study of amaranth extract in healthy humans: A randomized trial

ObjectiveNitric oxide (NO) is one of the most important signaling molecules produced within the body. Continuous generation of NO is essential for the integrity of the cardiovascular system. The aim of this study was to assess whether oral intake of a nitrate (NO3ˉ)-rich dietary supplement (amaranth extract) is able to increase NO3ˉ and nitrite (NO2ˉ) levels in blood plasma and saliva of healthy adults. MethodsIn the present study, bioavailability and pharmacokinetics of NO3ˉ and NO2ˉ from amaranth extract (2 g as single dose) was studied in 16 healthy individuals and compared with placebo in a crossover design. The NO3ˉ and NO2ˉ levels in plasma as well as saliva were measured up to 24 h. ResultsAfter administration of amaranth extract, the NO3ˉ levels in plasma as well as saliva were found to be significantly (P < 0.001) higher than in the placebo group. The NO2ˉ level in plasma was slightly higher (P < 0.05) in the amaranth group (test group) compared with that in the placebo group, whereas the saliva NO2ˉ level was significantly high (P < 0.001) in the amaranth extract–treated group than the placebo group. ConclusionsThese results clearly indicate that a single oral dose of amaranth extract is able to increase the NO3ˉ and NO2ˉ levels in the body for at least 8 h. The increase in NO3ˉ and NO2ˉ levels can help to improve the overall performance of people involved in vigorous physical activities or sports.