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Sample records for plasma soluble cd14

  1. Kinetics of plasma procalcitonin, soluble CD14, CCL2 and IL-10 after a sublethal infusion of lipopolysaccharide in horses.

    PubMed

    Bonelli, Francesca; Meucci, Valentina; Divers, Thomas J; Wagner, Bettina; Intorre, Luigi; Sgorbini, Micaela

    2017-02-01

    Endotoxemia represents a significant clinical and economic problem for the equine industry. This study assesses the kinetics of soluble CD14 (sCD14), chemokine (CC motif) ligand 2 (CCL2), interleukin 10 (IL-10) and plasma procalcitonin (PCT) in healthy horses after the intravenous infusion of lipopolysaccharide (LPS). The aim was to contribute to the basic understanding of the equine species-specific kinetics of these molecules in response to LPS exposure, which could support further findings in clinical studies and identify valuable inflammatory biomarkers for equine practice. Eleven healthy horses were involved in this experimental in vivo study. Horses were classified as healthy before the LPS infusion. After the pre-infusion blood collection (T0), all horses received an infusion of E. coli endotoxin (30ng/kg over 30min). Data and samples were collected 1h (T1), 2 (T2), 3 (T3) and 24h (T24) after infusion. Plasma sCD14, CCL2 and IL-10 were evaluated with a fluorescent bead-based assay, while PCT was evaluated with an equine PCT ELISA assay. A one-way ANOVA test was performed between each blood-sampling time for PCT, sCD14 and IL-10, and a Friedman test was performed for CCL2. Plasma PCT, IL-10 and CCL2 concentrations increased statistically significantly at T1, T2 and T3 compared to T0. No statistically significant differences were found between plasma IL-10 and CCL2 concentrations between T0 vs T24, although plasma PCT values remained high 24h after LPS infusion. Plasma sCD14 concentration showed no statistically significant differences for any of sampling times. Our results demonstrate that LPS injection into healthy horses results in PCT, CCL2 and IL-10 increases in plasma without an increase in sCD14. The increases in PCT, CCL2 and IL-10 are related to the inflammatory response induced by circulating lipopolysaccharide. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Interaction of CD14 haplotypes and soluble CD14 on pulmonary function in agricultural workers.

    PubMed

    LeVan, Tricia D; Smith, Lynette M; Heires, Art J; Mikuls, Ted R; Meza, Jane L; Weissenburger-Moser, Lisa A; Romberger, Debra J

    2017-03-16

    Agricultural environments are contaminated with organic dusts containing bacterial components. Chronic inhalation of organic dusts is implicated in respiratory diseases. CD14 is a critical receptor for gram-negative lipopolysaccharide; however, its association with respiratory disease among agricultural workers is unknown. The objective of this study was to determine if serum soluble CD14 (sCD14) levels are associated with lung function among agricultural workers and if this association is modified by genetic variants in CD14. This cross-sectional study included 584 veterans with >2 years of farming experience and that were between the ages of 40 and 80 years. Participants underwent spirometry and were genotyped for four tagging CD14 polymorphisms (CD14/-2838, rs2569193; CD14/-1720, rs2915863; CD14/-651, rs5744455; and CD14/-260, rs2569190). Serum sCD14 was assayed by ELISA. Subjects were 98% white males with a mean age 64.5 years. High soluble CD14 levels (> median sCD14) were associated decreased lung function (FEV1/FVC, p = 0.011; % predicted FEV1, p = 0.03). When stratified by COPD (yes/no) and smoking status (ever/never), high sCD14 levels (> median sCD14) were associated with low lung function among ever smokers with COPD (% predicted FEV1, padj = 0.0008; FEV1/FVC, padj = 0.0002). A similar trend was observed for never smokers with COPD; however, results did not reach statistical significance due to small sample size. There was a significant sCD14 x COPD/smoking interaction with lung function (% predicted FEV1, pinter = 0.0498; FEV1/FVC, pinter = 0.011). Regression models were adjusted for age, body mass index, education, sex, race and years worked on a farm. No association was found between CD14 polymorphisms/haplotypes (CD14/-2838; CD14/-1720; CD14/-651; CD14/-260) and sCD14 levels. The final model included the variables sCD14 and haplotypes and a haplotype x sCD14 interaction term. Individuals with the GTTG haplotype (CD14

  3. Lipopolysaccharide binding protein and soluble CD14 catalyze exchange of phospholipids.

    PubMed Central

    Yu, B; Hailman, E; Wright, S D

    1997-01-01

    Lipopolysaccharide binding protein (LBP) is a plasma protein known to facilitate the diffusion of bacterial LPS (endotoxin). LBP catalyzes movement of LPS monomers from LPS aggregates to HDL particles, to phospholipid bilayers, and to a binding site on a second plasma protein, soluble CD14 (sCD14). sCD14 can hasten transfer by receiving an LPS monomer from an LPS aggregate, and then surrendering it to an HDL particle, thus acting as a soluble "shuttle" for an insoluble lipid. Here we show that LBP and sCD14 shuttle not only LPS, but also phospholipids. Phosphatidylinositol (PI), phosphatidylcholine, and a fluorescently labeled derivative of phosphatidylethanolamine (R-PE) are each transferred by LBP from membranes to HDL particles. The transfer could be observed using recombinant LBP and sCD14 or whole human plasma, and the plasma-mediated transfer of PI could be blocked by anti-LBP and partially inhibited by anti-CD14. sCD14 appears to act as a soluble shuttle for phospholipids since direct binding of PI and R-PE to sCD14 was observed and because addition of sCD14 accelerated transfer of these lipids. These studies define a new function for LBP and sCD14 and describe a novel mechanism for the transfer of phospholipids in blood. In further studies, we show evidence suggesting that LBP transfers LPS and phospholipids by reciprocal exchange: LBP-catalyzed binding of R-PE to LPS x sCD14 complexes was accompanied by the exit of LPS from sCD14, and LBP-catalyzed binding of R-PE to sCD14 was accelerated by prior binding of LPS to sCD14. Binding of one lipid is thus functionally coupled with the release of a second. These results suggest that LBP acts as a lipid exchange protein. PMID:9006000

  4. Comparison of lipopolysaccharides and soluble CD14 measurement between clinically endotoxaemic and nonendotoxaemic horses.

    PubMed

    Fogle, J; Jacob, M; Blikslager, A; Edwards, A; Wagner, B; Dean, K; Fogle, C

    2017-03-01

    Clinically useful biomarkers are needed for early identification of endotoxaemic horses. Soluble CD14 (sCD14) is amplified early in response to inflammatory signals, including bacterial lipopolysaccharide (LPS), and may prove a useful biomarker for clinical endotoxaemia. The aim of this study was to determine if sCD14 could serve as a more reliable biomarker of the clinical signs of endotoxaemia, compared to measuring LPS alone. Prospective observational study in horses at a veterinary teaching hospital. Plasma samples were collected from 20 healthy horses and 35 horses presenting for emergency evaluation. Horses were classified as clinically endotoxaemic, using previously established criteria, if they had a heart rate >70 beats/min, packed cell volume >45% and/or a lesion likely to result in endotoxaemia. Soluble CD14 was measured using a cytometric bead-based assay and LPS was measured using a Limulus amoebocyte lysate (LAL) assay. Soluble CD14 was higher in horses classified as clinically endotoxaemic (median 1102 ng/ml, interquartile range 439 ng/ml), compared to clinically nonendotoxaemic (median 692 ng/ml, interquartile range 455 ng/ml, P = 0.03. There was no difference in LPS concentrations between clinically nonendotoxaemic (median 5.4 endotoxin units [EU]/ml, interquartile range 5 EU/ml) and endotoxaemic horses (median 7.2 EU/ml, interquartile range 17 EU/ml, P = 0.2). There was no correlation between sCD14 and LPS values in paired serum samples. LPS and sCD14 values were used to generate a receiver operating characteristic curve. The area under the curve for LPS and sCD14 was <0.7, suggesting that sCD14 and LPS were poor predictors of clinical endotoxaemia for the horses in this study. Further investigation is warranted to assess the utility of sCD14 measurement as a clinically useful biomarker to identify endotoxaemia in horses. © 2016 EVJ Ltd.

  5. Soluble CD14 as a Diagnostic and Prognostic Biomarker in Hematological Patients with Febrile Neutropenia

    PubMed Central

    Intke, Carina; Hämäläinen, Sari; Jantunen, Esa; Juutilainen, Auni

    2017-01-01

    Objective Elevated levels of a cell surface glycoprotein, soluble cluster of differentiation 14 (sCD14), have been observed in patients with sepsis. Only scarce data are available on sCD14 in hematological patients with chemotherapy-induced febrile neutropenia. The study aim was to investigate sCD14 as an early biomarker in febrile neutropenia after intensive chemotherapy to detect a rapidly deteriorating clinical course early enough to avoid serious infectious complications. Patients and Methods This prospective study included 87 adult hematological patients at the start of febrile neutropenia after intensive chemotherapy for acute myeloid leukemia or after autologous stem cell transplantation. The study endpoints were septic shock, severe sepsis, and positive blood culture findings. sCD14 was analyzed from day 0 to day 2, and its prognostic capacity was compared to that of C-reactive protein and procalcitonin. Results Plasma level of sCD14 predicted the development of septic shock on day 1 (p = 0.001) and day 2 but not the development of severe sepsis or blood culture positivity in hematological patients with chemotherapy-induced febrile neutropenia. Conclusions Soluble CD14 did not predict an overall complicated course at the early stages of febrile neutropenia. However, it was helpful in predicting the progression of the clinical course of neutropenic fever to septic shock. PMID:28845081

  6. Soluble CD14 enriched in colostrum and milk induces B cell growth and differentiation.

    PubMed

    Filipp, D; Alizadeh-Khiavi, K; Richardson, C; Palma, A; Paredes, N; Takeuchi, O; Akira, S; Julius, M

    2001-01-16

    Induction of resting B cell growth and differentiation requires a complex series of temporally coordinated signals that are initiated on contact with activated helper T cells. These signals complement one another, each rendering the B cell susceptible to factors supporting progressive activation. Here, we demonstrate that soluble CD14 (sCD14) bypasses the physiological sequelae of events that limit B cell activation. B cell growth and differentiation in vitro is induced by both native and recombinant forms of sCD14 at nanomolar concentrations. sCD14-mediated cellular activation does not require membrane CD14 expression, depends on a region of CD14 that is not involved in lipopolysaccharide binding, and requires functional Toll-like receptor 4. Consistent with biological activity of sCD14 in vitro, its administration to neonatal mice enhances Ig secretion. The results presented establish sCD14 as a naturally occurring soluble B cell mitogen of mammalian origin.

  7. Associations of Soluble CD14 and Endotoxin with Mortality, Cardiovascular Disease, and Progression of Kidney Disease among Patients with CKD

    PubMed Central

    Poesen, Ruben; Ramezani, Ali; Claes, Kathleen; Augustijns, Patrick; Kuypers, Dirk; Barrows, Ian R.; Muralidharan, Jagadeesan; Evenepoel, Pieter; Meijers, Björn

    2015-01-01

    Background and objectives CD14 plays a key role in the innate immunity as pattern-recognition receptor of endotoxin. Higher levels of soluble CD14 (sCD14) are associated with overall mortality in hemodialysis patients. The influence of kidney function on plasma sCD14 levels and its relationship with adverse outcomes in patients with CKD not yet on dialysis is unknown. This study examines the associations between plasma levels of sCD14 and endotoxin with adverse outcomes in patients with CKD. Design, setting, participants, & measurements We measured plasma levels of sCD14 and endotoxin in 495 Leuven Mild-to-Moderate CKD Study participants. Mild-to-moderate CKD was defined as presence of kidney damage or eGFR<60 ml/min per 1.73 m2 for ≥3 months, with exclusion of patients on RRT. Study participants were enrolled between November 2005 and September 2006. Results Plasma sCD14 was negatively associated with eGFR (ρ=–0.34, P<0.001). During a median follow-up of 54 (interquartile range, 23–58) months, 53 patients died. Plasma sCD14 was predictive of mortality, even after adjustment for renal function, Framingham risk factors, markers of mineral bone metabolism, and nutritional and inflammatory parameters (hazard ratio [HR] per SD higher of 1.90; 95% confidence interval [95% CI],1.32 to 2.74; P<0.001). After adjustment for the same risk factors, plasma sCD14 was also a predictor of cardiovascular disease (HR, 1.30; 95% CI, 1.00 to 1.69; P=0.05). Although plasma sCD14 was associated with progression of CKD, defined as reaching ESRD or doubling of serum creatinine in models adjusted for CKD-specific risk factors (HR, 1.24; 95% CI, 1.01 to 1.52; P=0.04), significance was lost when adjusted for proteinuria (HR, 1.19; 95% CI, 0.96 to 1.48; P=0.11). There was neither correlation between plasma endotoxin and sCD14 (ρ=–0.06, P=0.20) nor was endotoxin independently associated with adverse outcome during follow-up. Conclusions Plasma sCD14 is elevated in patients with

  8. Soluble CD14 subtype presepsin (sCD14-ST) and lipopolysaccharide binding protein (LBP) in neonatal sepsis: new clinical and analytical perspectives for two old biomarkers.

    PubMed

    Mussap, Michele; Noto, Antonio; Fravega, Marco; Fanos, Vassilios

    2011-10-01

    Several biochemical markers have been proposed over the past years to manage critically ill newborns with acute inflammation and sepsis. The state of the art in diagnosing and monitoring neonatal sepsis, severe sepsis and septic shock consists of the measurement of plasma C-reactive protein (CRP) and procalcitonin (PCT) at the onset and in the course of the disease. CRP and PCT in combination are clinically significant in diagnosing and monitoring septic newborns; however, CRP and PCT have a very limited value for risk stratification and in predicting outcome. The availability of commercial methods for the automated measurement of the soluble CD14 subtype presepsin (sCD14-ST) and lipopolysaccharide binding protein (LBP) represent a challenge for the evaluation in clinical practice of reliable markers of neonatal sepsis, specifically for the very early diagnosis, the classification into class of severity, and the prediction of complications and death.

  9. Recombinant soluble CD14 reduces severity of intramammary infection by Escherichia coli.

    PubMed

    Lee, Jai-Wei; Paape, Max J; Elsasser, Theodore H; Zhao, Xin

    2003-07-01

    The interaction among gram-negative bacteria, the innate immune system, and soluble CD14 (sCD14) has not been well documented. The effect of recombinant bovine sCD14 (rbosCD14) on milk somatic cell count (SCC), bacterial clearance, and cytokine production was investigated by using a bovine intramammary Escherichia coli infection model. We first determined whether rbosCD14 would increase the SCC during a lipopolysaccharide (LPS) challenge. Three quarters of each of six healthy lactating cows were injected with either 0.3 microg of LPS, 0.3 microg of LPS plus 100 micro g of rbosCD14, or saline. In comparison with quarters injected with LPS alone, the SCC was twofold higher (P < 0.05) in quarters injected with LPS plus rbosCD14 after the challenge. We therefore hypothesized that when E. coli bacteria invade the mammary gland, sCD14 in milk would interact with LPS and rapidly recruit neutrophils from the blood to eliminate the bacteria before establishment of infection. To test this hypothesis, two quarters of each of nine healthy cows were challenged with either 50 CFU of E. coli plus saline or 50 CFU of E. coli plus 100 microg of rbosCD14. Quarters challenged with E. coli plus rbosCD14 had a more rapid recruitment of neutrophils, which was accompanied by a faster clearance of bacteria, lower concentrations of tumor necrosis factor alpha and interleukin-8 in milk, and milder clinical symptoms, than challenged quarters injected with saline. Results indicate that increasing the concentration of sCD14 in milk may be a potential strategy with which to prevent or reduce the severity of infection by coliform bacteria.

  10. Recombinant bovine soluble CD14 sensitizes the mammary gland to lipopolysaccharide.

    PubMed

    Wang, Yan; Zarlenga, Dante S; Paape, Max J; Dahl, Geoffrey E

    2002-05-01

    Standard therapies including administration of potent antibiotics, aggressive fluid resuscitation and metabolic support have not been successful in relieving symptoms and reducing mortality associated with acute coliform mastitis. It is important to understand the pathophysiological response of the mammary gland to coliform infections when designing preventive or therapeutic regimens for controlling coliform mastitis. Our laboratory has previously shown that macrophages and polymorphonuclear neutrophils in milk express CD14 on their cell surface. In this study, we found that soluble CD14 (sCD14) is present in milk whey as a 46kDa protein reacted with anti-ovine CD14 antibody. Additional functional studies found that: (1) under serum-free condition, complexes of LPS-recombinant bovine soluble CD14 (rbosCD14) induced activation of mammary ductal epithelial cells (as measured by changes in interleukin-8 (IL-8) mRNA level by competitive RT-PCR) at low concentrations of LPS after 6 or 24h incubation (1-1000ng/ml), whereas LPS alone did not induce activation of mammary ductal epithelial cells at the same concentrations, and (2) intramammary injection of low concentrations of LPS did not increase concentration of leukocytes in milk. In contrast, LPS-rbosCD14 complex containing the same concentration of LPS increased the concentration of leukocytes in the injected mammary gland at 12 and 24h post-injection. These results indicate that rbosCD14 sensitizes mammary epithelial cells to low concentrations of LPS in vitro and in vivo. Endogenous sCD14 in milk may be important in initiating host responses to Gram-negative bacterial infections.

  11. Soluble CD14 Enhances the Response of Periodontal Ligament Stem Cells to P. gingivalis Lipopolysaccharide

    PubMed Central

    Andrukhov, Oleh; Andrukhova, Olena; Özdemir, Burcu; Haririan, Hady; Müller-Kern, Michael; Moritz, Andreas; Rausch-Fan, Xiaohui

    2016-01-01

    Periodontal ligament stem cells (PDLSCs) are lacking membrane CD14, which is an important component of lipopolysaccharide (LPS) signaling through toll-like receptor (TLR) 4. In the present study we investigated the effect of soluble CD14 on the response of human PDLSCs to LPS of Porphyromonas (P.) gingivalis. Human PDLSCs (hPDLSCs) were stimulated with P. gingivalis LPS in the presence or in the absence of soluble CD14 (sCD14) and the production of interleukin (IL)-6, chemokine C-X-C motif ligand 8 (CXCL8), and chemokine C-C motif ligand 2 (CCL2) was measured. The response to P. gingivalis LPS was compared with that to TLR4 agonist Escherichia coli LPS and TLR2-agonist Pam3CSK4. The response of hPDLSCs to both P. gingivalis LPS and E. coli LPS was significantly enhanced by sCD14. In the absence of sCD14, no significant difference in the hPDLSCs response to two kinds of LPS was observed. These responses were significantly lower compared to that to Pam3CSK4. In the presence of sCD14, the response of hPdLSCs to P. gingivalis LPS was markedly higher than that to E. coli LPS and comparable with that to Pam3CSK4. The response of hPdLSCs to bacterial LPS is strongly augmented by sCD14. Local levels of sCD14 could be an important factor for modulation of the host response against periodontal pathogens. PMID:27504628

  12. Elevated milk soluble CD14 in bovine mammary glands challenged with Escherichia coli lipopolysaccharide.

    PubMed

    Lee, J W; Paape, M J; Elsasser, T H; Zhao, X

    2003-07-01

    The purpose of this study was to determine whether soluble CD14 (sCD14) in milk was affected by stage of lactation, milk somatic cell count (SCC), presence of bacteria, or lipopolysaccharide (LPS)-induced inflammation. Milk samples from 100 lactating cows (396 functional quarters) were assayed for sCD14 in milk to determine effects of stage of lactation, SCC, and intramammary infection. The concentration of sCD14 was highest in transitional milk (0 to 4 d postpartum) and in milk with high SCC (> 750,000 cells/ml). Most of the infected quarters (> 80%) were infected by coagulase-negative staphylococci and yeast. No difference was found between noninfected and infected quarters. One quarter of six healthy lactating cows was challenged with 100 microg LPS in order to study the kinetics of sCD14 during an LPS-induced inflammation. Milk samples were collected at various intervals until 72 h after injection. Rectal temperature, milk tumor necrosis factor-alpha, and interleukin-8 increased immediately after challenge. The increase in sCD14 paralleled the increase in SCC, peaked at 12 h, and started to decline after 24 h. Serum leakage, as characterized by the level of bovine serum albumin in milk, peaked at 4 h and then gradually decreased. All parameters remained at basal levels in control quarters throughout the study. In vitro experiments indicated that neutrophils released sCD14 in response to LPS in a dose-dependent manner. The results indicate that the concentration of sCD14 was significantly increased in milk after LPS challenge. The increase was not likely due to serum leakage. Instead, infiltrated neutrophils might be the main source of increased sCD14 in milk during inflammation.

  13. Soluble CD14 in cerebrospinal fluid is associated with markers of inflammation and axonal damage in untreated HIV-infected patients: a retrospective cross-sectional study.

    PubMed

    Jespersen, Sofie; Pedersen, Karin Kæreby; Anesten, Birgitta; Zetterberg, Henrik; Fuchs, Dietmar; Gisslén, Magnus; Hagberg, Lars; Trøseid, Marius; Nielsen, Susanne Dam

    2016-04-21

    HIV-associated cognitive impairment has declined since the introduction of combination antiretroviral treatment (cART). However, milder forms of cognitive impairment persist. Inflammation in the cerebrospinal fluid (CSF) has been associated with cognitive impairment, and CSF neurofilament light chain protein (NFL) and CSF neopterin concentrations are increased in those patients. Microbial translocation in HIV infection has been suggested to contribute to chronic inflammation, and lipopolysaccharide (LPS) and soluble CD14 (sCD14) are markers of microbial translocation and the resulting monocyte activation, respectively. We hypothesised that microbial translocation contributes to inflammation and axonal damage in the central nervous system (CNS) in untreated HIV infection. We analyzed paired samples of plasma and CSF from 62 HIV-infected, untreated patients without cognitive symptoms from Sahlgrenska University Hospital, Gothenburg, Sweden. Measurements of neopterin and NFL in CSF were available from previous studies. Plasma and CSF sCD14 was measured using ELISA (R&D, Minneapolis, MN), and plasma and CSF LPS was measured using LAL colorimetric assay (Lonza, Walkersville, MD, USA). Univariate and multivariate regression analyses were performed. LPS in plasma was associated with plasma sCD14 (r = 0.31, P = 0.015), and plasma sCD14 was associated with CSF sCD14 (r = 0.32, P = 0.012). Furthermore, CSF sCD14 was associated with NFL (r = 0.32, P = 0.031) and neopterin (r = 0.32, P = 0.012) in CSF. LPS was not detectable in CSF. In a multivariate regression model CSF sCD14 remained associated with NFL and neopterin after adjusting for age, CD4+ cell count, and HIV RNA in CSF. In a group of untreated, HIV-infected patients LPS was associated with sCD14 in plasma, and plasma sCD14 was associated CSF sCD14. CSF sCD14 were associated with markers of CNS inflammation and axonal damage. This suggest that microbial translocation might be a driver

  14. Ingested soluble CD14 from milk is transferred intact into the blood of newborn rats

    PubMed Central

    Ward, Tonya L.; Spencer, William J.; Davis, Laura D. R.; Harrold, JoAnn; Mack, David R.; Altosaar, Illimar

    2016-01-01

    Background Milk contains immunological constituents that comprise an edible immune system conveyed from mother to newborn. Soluble Cluster of Differentiation 14 (sCD14) is a protein found in significant quantities in human milk (~8–29 μg/ml). At a tenfold lower concentration in the blood (~3 μg/ml), the most notable role of sCD14 is to sequester lipopolysaccharide of Gram-negative bacteria from immune cells. Methods To explore the pharmacodynamics of this milk protein and its biological fate, the biodistribution of radiolabeled sCD14 (14C, 125I) was monitored in 10 d old rat pups. Results Up to 3.4 ± 2.2% of the radiolabeled-sCD14 administered was observed, intact, in the pup blood for up to 8 h post-ingestion. Additionally, 30.3 ± 13.0% of the radiolabeled-sCD14 administered was observed degraded in the stomach at 8 h post-ingestion. A reservoir of intact, administered sCD14 (3.2 ± 0.3%), however, remained in the stomach at 8 h post-ingestion. Intact sCD14 was observed in the small intestine at 5.5 ± 1.6% of the dose fed at 8h post-ingestion. Conclusions The presence of intact sCD14 in the blood and gastrointestinal tract of newborns post-ingestion has implications in the development of allergies, obesity and other inflammation-related pathogeneses later in life. PMID:24232637

  15. Increased Risk of Radiographic Emphysema in HIV Is Associated With Elevated Soluble CD14 and Nadir CD4

    PubMed Central

    Attia, Engi F.; Akgün, Kathleen M.; Wongtrakool, Cherry; Goetz, Matthew Bidwell; Rodriguez-Barradas, Maria C.; Rimland, David; Brown, Sheldon T.; Soo Hoo, Guy W.; Kim, Joon; Lee, Patty J.; Schnapp, Lynn M.; Sharafkhaneh, Amir; Justice, Amy C.

    2014-01-01

    BACKGROUND: The association between HIV and emphysema remains incompletely understood. We sought to determine whether HIV is an independent risk factor for emphysema severity and whether markers of HIV severity and systemic biomarkers of inflammation (IL-6), altered coagulation (D-dimer), and immune activation (soluble CD14) are associated with emphysema. METHODS: We performed a cross-sectional analysis of 114 participants with HIV infection and 89 participants without HIV infection in the Examinations of HIV-Associated Lung Emphysema (EXHALE) study. Participants underwent chest CT imaging with blinded semiquantitative interpretation of emphysema severity, distribution, and type. We generated multivariable logistic regression models to determine the risk of HIV for radiographic emphysema, defined as > 10% lung involvement. Similar analyses examined associations of plasma biomarkers, HIV RNA, and recent and nadir CD4 cell counts with emphysema among participants with HIV infection. RESULTS: Participants with HIV infection had greater radiographic emphysema severity with increased lower lung zone and diffuse involvement. HIV was associated with significantly increased risk for > 10% emphysema in analyses adjusted for cigarette smoking pack-years (OR, 2.24; 95% CI, 1.12-4.48). In multivariable analyses restricted to participants with HIV infection, nadir CD4 < 200 cells/μL (OR, 2.98; 95% CI, 1.14-7.81), and high soluble CD14 level (upper 25th percentile) (OR, 2.55; 95% CI, 1.04-6.22) were associated with increased risk of > 10% emphysema. IL-6 and D-dimer were not associated with emphysema in HIV. CONCLUSIONS: HIV is an independent risk factor for radiographic emphysema. Emphysema severity was significantly greater among participants with HIV infection. Among those with HIV, nadir CD4 < 200 cells/μL and elevated soluble CD14 level were associated with emphysema, highlighting potential mechanisms linking HIV with emphysema. PMID:25080158

  16. Soluble CD14 subtype (sCD14-ST) presepsin in premature and full term critically ill newborns with sepsis and SIRS.

    PubMed

    Mussap, Michele; Puxeddu, Elisabetta; Puddu, Melania; Ottonello, Giovanni; Coghe, Ferdinando; Comite, Paola; Cibecchini, Francesco; Fanos, Vassilios

    2015-12-07

    Neonatal sepsis still remains a major cause of morbidity and mortality in neonatal intensive care unit (NICU). Recently, soluble CD14 subtype (sDC14-ST) also named presepsin, was proposed as an effective biomarker for diagnosing, monitoring, and assessing the risk of neonatal sepsis and septic shock. The aim of this study was to investigate the diagnostic accuracy of sCD14-ST presepsin in diagnosing neonatal bacterial sepsis and in discriminating non-bacterial systemic inflammatory response syndrome (SIRS) from bacterial sepsis. This study involved 65 critically ill full-term and preterm newborns admitted to the neonatal intensive care unit (NICU), divided into three groups: 25 newborns with bacterial neonatal sepsis (group A); 15 newborns with a diagnosis of non-bacterial SIRS and with no localizing source of bacterial infection (group B); and 25 babies with no clinical or bacteriological signs of systemic or local infection receiving routine NICU care, most of them treated with phototherapy for neonatal jaundice (group C). A total of 102 whole blood samples were collected, 40 in group A, 30 in group B and 32 in group C. In 10 babies included in group A, sCD14-ST presepsin was also measured in an additional second blood sample collected 3 days after the start of antibiotic treatment. sCD14-ST presepsin was measured by a commercially available chemiluminescent enzyme immunoassay (CLEIA) optimized on an automated immunoassay analyzer. Statistical analysis was performed by means of MedCalc® statistical package; receiver operating characteristic (ROC) analysis was computed, and the area under the ROC curve (AUC) was used to evaluate the ability of sCD14-ST to discriminate neonatal bacterial sepsis from non-bacterial SIRS. Blood sCD14-ST presepsin levels were found significantly higher in bacterial sepsis when compared with controls (p<0.0001); similarly, they were higher in non-bacterial SIRS when compared with controls (p<0.0001). However, no statistically

  17. Soluble CD14 acts as a shuttle in the neutralization of lipopolysaccharide (LPS) by LPS-binding protein and reconstituted high density lipoprotein

    PubMed Central

    1995-01-01

    We have recently shown that lipopolysaccharide (LPS)-binding protein (LBP) is a lipid transfer protein that catalyzes two distinct reactions: movement of bacterial LPS (endotoxin) from LPS micelles to soluble CD14 (sCD14) and movement of LPS from micelles to reconstituted high density lipoprotein (R-HDL) particles. Here we show that LBP facilitates a third lipid transfer reaction: movement of LPS from LPS- sCD14 complexes to R-HDL particles. This action of LBP is catalytic, with one molecule of LBP enabling the movement of multiple LPS molecules into R-HDL. LBP-catalyzed movement of LPS from LPS-sCD14 complexes to R-HDL neutralizes the capacity of LPS to stimulate polymorphonuclear leukocytes. Our findings show that LPS may be transferred to R-HDL either by the direct action of LBP or by a two- step reaction in which LPS is first transferred to sCD14 and subsequently to R-HDL. We have observed that the two-step pathway of LPS transfer to R-HDL is strongly favored over direct transfer. Neutralization of LPS by LBP and R-HDL was accelerated more than 30- fold by addition of sCD14. Several observations suggest that sCD14 accelerates this reaction by serving as a shuttle for LPS: addition of LBP and sCD14 to LPS micelles resulted in LPS-sCD14 complexes that could diffuse through a 100-kD cutoff filter; LPS-sCD14 complexes appeared transiently during movement of LPS to R-HDL facilitated by purified LBP; and sCD14 could facilitate transfer of LPS to R-HDL without becoming part of the final LPS-R-HDL complex. Complexes of LPS and sCD14 were formed transiently when LPS was incubated in plasma, suggesting that these complexes may play a role as intermediates in the neutralization of LPS under physiological conditions. These findings detail a new activity for sCD14 and suggest a novel mechanism for lipid transfer by LBP. PMID:7536794

  18. Soluble CD14 acts as a DAMP in human macrophages: origin and involvement in inflammatory cytokine/chemokine production.

    PubMed

    Lévêque, Manuella; Simonin-Le Jeune, Karin; Jouneau, Stéphane; Moulis, Solenn; Desrues, Benoit; Belleguic, Chantal; Brinchault, Graziella; Le Trionnaire, Sophie; Gangneux, Jean-Pierre; Dimanche-Boitrel, Marie-Thérèse; Martin-Chouly, Corinne

    2017-05-01

    The innate immune system is able to detect bacterial LPS through the pattern recognition receptor CD14, which delivers LPS to various TLR signaling complexes that subsequently induce intracellular proinflammatory signaling cascades. In a previous study, we showed the overproduction of the soluble form of CD14 (sCD14) by macrophages from patients with cystic fibrosis (CF). CF is an autosomal recessive disorder that is caused by mutations in the gene that encodes the CFTR protein and is characterized by persistent inflammation. Macrophages play a significant role in the initial stages of this disease due to their inability to act as suppressor cells, leading to chronic inflammation in CF. In this work, we investigated the origin of sCD14 by human macrophages and studied the effect of sCD14 on the production of inflammatory cytokine/chemokine. Our data indicate that sCD14 stimulate proinflammatory cytokine/chemokine production in a manner that is independent of LPS but dependent on the TLR-4/CD14 membrane complex, NF-κB, and the inflammasome. Therefore, sCD14, overproduced by CF macrophages, originates primarily from the endocytosis/exocytosis process and should be considered to be a danger-associated molecular pattern. This elucidation of the origin and inflammation-induced mechanisms associated with sCD14 contributes to our understanding of maintained tissue inflammation.-Lévêque, M., Simonin-Le Jeune, K., Jouneau, S., Moulis, S., Desrues, B., Belleguic, C., Brinchault, G., Le Trionnaire, S., Gangneux, J.-P., Dimanche-Boitrel, M.-T., Martin-Chouly, C. Soluble CD14 acts as a DAMP in human macrophages: origin and involvement in inflammatory cytokine/chemokine production. © FASEB.

  19. Neopterin and Soluble CD14 Levels as Indicators of Immune Activation in Cases with Indeterminate Pattern and True Positive HIV-1 Infection.

    PubMed

    Uysal, Hayriye Kırkoyun; Sohrabi, Pari; Habip, Zafer; Saribas, Suat; Kocazeybek, Emre; Seyhan, Fatih; Calışkan, Reyhan; Bonabi, Esad; Yuksel, Pelin; Birinci, Ilhan; Uysal, Omer; Kocazeybek, Bekir

    2016-01-01

    We aimed to evaluate the roles of the plasma immune activation biomarkers neopterin and soluble CD14 (sCD14) in the indirect assessment of the immune activation status of patients with the indeterminate HIV-1 (IHIV-1) pattern and a true HIV-1-positive infection (PCG). This cross-sectional and descriptive study included eighty-eight patients with the IHIV-1 pattern, 100 patients in the PCG, and 100 people in a healthy control group (HCG). Neopterin and sCD14 levels were determined by competitive and sandwich ELISA methods, respectively. Mean neopterin and sCD14 levels among those with the IHIV-1 pattern were significantly lower than among the PCG (p < 0.001 and p = 0.001, respectively), but they were similiar to those in the HCG (p = 0.57 and p = 0.66, respectively. Mean neopterin and sCD14 levels among the PCG were found to be significantly higher than among those with the IHIV-1 pattern (p < 0.001 and p = 0.001, respectively) and among those in the HCG (p = 0.001, p < 0.001, respectively). Neopterin did not have adequate predictive value for identifying those in the PCG (area under the curve [AUC] = 0.534; 95% CI, 0.463-0.605; p = 0.4256); sCD14 also had poor predictive value but high specificity (100%) for identifying those in the PCG (AUC = 0.627; 95% CI, 0.556-0.694; p = 0.0036). While low levels of these two biomarkers were detected among those with the IHIV-1 pattern, they were found in high levels among those in the PCG. These two markers obviously cannot be used as a sceening test because they have low sensitivies. Taken together, we suggest that neopterin and sCD14 may be helpful because they both have high specificity (92%-100%) as indirect non-specific markers for predicting the immune activation status of individuals, whether or not they have true positive HIV-1.

  20. Neopterin and Soluble CD14 Levels as Indicators of Immune Activation in Cases with Indeterminate Pattern and True Positive HIV-1 Infection

    PubMed Central

    Uysal, Hayriye Kırkoyun; Sohrabi, Pari; Habip, Zafer; Saribas, Suat; Kocazeybek, Emre; Seyhan, Fatih; Calışkan, Reyhan; Bonabi, Esad; Yuksel, Pelin; Birinci, Ilhan; Uysal, Omer; Kocazeybek, Bekir

    2016-01-01

    Background We aimed to evaluate the roles of the plasma immune activation biomarkers neopterin and soluble CD14 (sCD14) in the indirect assessment of the immune activation status of patients with the indeterminate HIV-1 (IHIV-1) pattern and a true HIV-1-positive infection (PCG). Methods This cross-sectional and descriptive study included eighty-eight patients with the IHIV-1 pattern, 100 patients in the PCG, and 100 people in a healthy control group (HCG). Neopterin and sCD14 levels were determined by competitive and sandwich ELISA methods, respectively. Results Mean neopterin and sCD14 levels among those with the IHIV-1 pattern were significantly lower than among the PCG (p < 0.001 and p = 0.001, respectively), but they were similiar to those in the HCG (p = 0.57 and p = 0.66, respectively. Mean neopterin and sCD14 levels among the PCG were found to be significantly higher than among those with the IHIV-1 pattern (p < 0.001 and p = 0.001, respectively) and among those in the HCG (p = 0.001, p < 0.001, respectively). Neopterin did not have adequate predictive value for identifying those in the PCG (area under the curve [AUC] = 0.534; 95% CI, 0.463–0.605; p = 0.4256); sCD14 also had poor predictive value but high specificity (100%) for identifying those in the PCG (AUC = 0.627; 95% CI, 0.556–0.694; p = 0.0036). Conclusions While low levels of these two biomarkers were detected among those with the IHIV-1 pattern, they were found in high levels among those in the PCG. These two markers obviously cannot be used as a sceening test because they have low sensitivies. Taken together, we suggest that neopterin and sCD14 may be helpful because they both have high specificity (92%-100%) as indirect non-specific markers for predicting the immune activation status of individuals, whether or not they have true positive HIV-1. PMID:27031691

  1. Soluble alpha-enolase activates monocytes by CD14-dependent TLR4 signalling pathway and exhibits a dual function

    PubMed Central

    Guillou, Clément; Fréret, Manuel; Fondard, Emeline; Derambure, Céline; Avenel, Gilles; Golinski, Marie-Laure; Verdet, Mathieu; Boyer, Olivier; Caillot, Frédérique; Musette, Philippe; Lequerré, Thierry; Vittecoq, Olivier

    2016-01-01

    Rheumatoid arthritis (RA) is the most common form of chronic inflammatory rheumatism. Identifying auto-antigens targeted by RA auto-antibodies is of major interest. Alpha-enolase (ENO1) is considered to be a pivotal auto-antigen in early RA but its pathophysiologic role remains unknown. The main objective of this study was to investigate the in vitro effects of soluble ENO1 on peripheral blood mononuclear cells (PBMC) from healthy donors and RA patients in order to determine the potential pathogenic role of ENO1. ELISA, transcriptomic analysis, experiments of receptor inhibition and flow cytometry analysis were performed to determine the effect, the target cell population and the receptor of ENO1. We showed that ENO1 has the ability to induce early production of pro-inflammatory cytokines and chemokines with delayed production of IL-10 and to activate the innate immune system. We demonstrated that ENO1 binds mainly to monocytes and activates the CD14-dependent TLR4 pathway both in healthy subjects and in RA patients. Our results establish for the first time that ENO1 is able to activate in vitro the CD14-dependent TLR4 pathway on monocytes involving a dual mechanism firstly pro-inflammatory and secondly anti-inflammatory. These results contribute to elucidating the role of this auto-antigen in the pathophysiologic mechanisms of RA. PMID:27025255

  2. Chronic immune activation in common variable immunodeficiency (CVID) is associated with elevated serum levels of soluble CD14 and CD25 but not endotoxaemia.

    PubMed

    Litzman, J; Nechvatalova, J; Xu, J; Ticha, O; Vlkova, M; Hel, Z

    2012-12-01

    Common variable immunodeficiency (CVID), the most frequent symptomatic immunoglobulin primary immunodeficiency, is associated with chronic T cell activation and reduced frequency of CD4(+) T cells. The underlying cause of immune activation in CVID is unknown. Microbial translocation indicated by elevated serum levels of lipopolysaccharide and soluble CD14 (sCD14) has been linked previously to systemic immune activation in human immunodeficiency virus/acquired immune deficiency syndrome (HIV-1/AIDS), alcoholic cirrhosis and other conditions. To address the mechanisms of chronic immune activation in CVID, we performed a detailed analysis of immune cell populations and serum levels of sCD14, soluble CD25 (sCD25), lipopolysaccharide and markers of liver function in 35 patients with CVID, 53 patients with selective immunoglobulin (Ig)A deficiency (IgAD) and 63 control healthy subjects. In CVID subjects, the concentration of serum sCD14 was increased significantly and correlated with the level of sCD25, C-reactive protein and the extent of T cell activation. Importantly, no increase in serum lipopolysaccharide concentration was observed in patients with CVID or IgAD. Collectively, the data presented suggest that chronic T cell activation in CVID is associated with elevated levels of sCD14 and sCD25, but not with systemic endotoxaemia, and suggest involvement of lipopolysaccharide-independent mechanisms of induction of sCD14 production. © 2012 The Authors Clinical and Experimental Immunology © 2012 British Society for Immunology.

  3. Chronic immune activation in common variable immunodeficiency (CVID) is associated with elevated serum levels of soluble CD14 and CD25 but not endotoxaemia

    PubMed Central

    Litzman, J; Nechvatalova, J; Xu, J; Ticha, O; Vlkova, M; Hel, Z

    2012-01-01

    Common variable immunodeficiency (CVID), the most frequent symptomatic immunoglobulin primary immunodeficiency, is associated with chronic T cell activation and reduced frequency of CD4+ T cells. The underlying cause of immune activation in CVID is unknown. Microbial translocation indicated by elevated serum levels of lipopolysaccharide and soluble CD14 (sCD14) has been linked previously to systemic immune activation in human immunodeficiency virus/acquired immune deficiency syndrome (HIV-1/AIDS), alcoholic cirrhosis and other conditions. To address the mechanisms of chronic immune activation in CVID, we performed a detailed analysis of immune cell populations and serum levels of sCD14, soluble CD25 (sCD25), lipopolysaccharide and markers of liver function in 35 patients with CVID, 53 patients with selective immunoglobulin (Ig)A deficiency (IgAD) and 63 control healthy subjects. In CVID subjects, the concentration of serum sCD14 was increased significantly and correlated with the level of sCD25, C-reactive protein and the extent of T cell activation. Importantly, no increase in serum lipopolysaccharide concentration was observed in patients with CVID or IgAD. Collectively, the data presented suggest that chronic T cell activation in CVID is associated with elevated levels of sCD14 and sCD25, but not with systemic endotoxaemia, and suggest involvement of lipopolysaccharide-independent mechanisms of induction of sCD14 production. PMID:23121673

  4. Bone Anabolic Effects of Soluble Si: In Vitro Studies with Human Mesenchymal Stem Cells and CD14+ Osteoclast Precursors

    PubMed Central

    Costa-Rodrigues, J.; Reis, S.; Castro, A.; Fernandes, M. H.

    2016-01-01

    Silicon (Si) is indispensable for many cellular processes including bone tissue metabolism. In this work, the effects of Si on human osteogenesis and osteoclastogenesis were characterized. Human mesenchymal stem cells (hMSC) and CD14+ stem cells, as osteoblast and osteoclast precursors, were treated with a wide range of Si concentrations, covering the physiological plasma levels. Si promoted a dose-dependent increase in hMSC proliferation, differentiation, and function, at levels similar to the normal basal plasma levels. Additionally, a decrease in the expression of the osteoclastogenic activators M-CSF and RANKL was observed. Also, Si elicited a decrease in osteoclastogenesis, which became significant at higher concentrations, as those observed after meals. Among the intracellular mechanisms studied, an upregulation of MEK and PKC signalling pathways was observed in both cell types. In conclusion, Si appears to have a direct positive effect on human osteogenesis, at basal plasma levels. On the other hand, it also seemed to be an inhibitor of osteoclastogenesis, but at higher concentrations, though yet in the physiological range. Further, an indirect effect of Si on osteoclastogenesis may also occur, through a downregulation of M-CSF and RANKL expression by osteoblasts. Thus, Si may be an important player in bone anabolic regenerative approaches. PMID:26798359

  5. Unchanged Levels of Soluble CD14 and IL-6 Over Time Predict Serious Non-AIDS Events in HIV-1-Infected People.

    PubMed

    Sunil, Meena; Nigalye, Maitreyee; Somasunderam, Anoma; Martinez, Maria Laura; Yu, Xiaoying; Arduino, Roberto C; Utay, Netanya S; Bell, Tanvir K

    2016-12-01

    HIV-1-infected persons have increased risk of serious non-AIDS events (SNAEs) despite suppressive antiretroviral therapy. Increased circulating levels of soluble CD14 (sCD14), soluble CD163 (sCD163), and interleukin-6 (IL-6) at a single time point have been associated with SNAEs. However, whether changes in these biomarker levels predict SNAEs in HIV-1-infected persons is unknown. We hypothesized that greater decreases in inflammatory biomarkers would be associated with fewer SNAEs. We identified 39 patients with SNAEs, including major cardiovascular events, end stage renal disease, decompensated cirrhosis, non-AIDS-defining malignancies, and death of unknown cause, and age- and sex-matched HIV-1-infected controls. sCD14, sCD163, and IL-6 were measured at study enrollment (T1) and proximal to the event (T2) or equivalent duration in matched controls. Over ∼34 months, unchanged rather than decreasing levels of sCD14 and IL-6 predicted SNAEs. Older age and current illicit substance abuse, but not HCV coinfection, were associated with SNAEs. In a multivariate analysis, older age, illicit substance use, and unchanged IL-6 levels remained significantly associated with SNAEs. Thus, the trajectories of sCD14 and IL-6 levels predict SNAEs. Interventions to decrease illicit substance use may decrease the risk of SNAEs in HIV-1-infected persons.

  6. Septin: a factor in plasma that opsonizes lipopolysaccharide-bearing particles for recognition by CD14 on phagocytes

    PubMed Central

    1992-01-01

    We have previously reported that lipopolysaccharide (LPS) binding protein (LBP) opsonizes endotoxin (LPS) for recognition by CD14 on phagocytes. Here we show that normal human plasma contains high titers of an activity that also binds LPS (Re, 595) and mediates recognition by CD14. Opsonization of LPS-coated particles with plasma enables the particles to be bound by phagocytes. Further, opsonization with plasma also enables subnanogram-per-milliliter concentrations of LPS to induce dramatic alterations in the function of leukocyte integrins on polymorphonuclear leukocytes and to induce secretion of tumor necrosis factor by monocytes, suggesting that opsonization by factors in plasma may be important in responses of cells to endotoxin. The opsonic activity in plasma appears distinct from LBP since it is not blocked by neutralizing antibodies against LBP. Surprisingly, the opsonic activity of plasma is not present in a single protein species, but at least two species must be combined to observe activity. Further, the opsonic activity of plasma for LPS is blocked by addition of protease inhibitors, suggesting that proteolytic activity or activities are required for opsonization. These properties are suggestive of the action of a protease cascade, but opsonic activity of plasma is not affected by blockade or depletion of either the complement or clotting cascades. We propose the name "septin" to describe this novel LPS- opsonizing activity in plasma. PMID:1380975

  7. Serum Soluble CD14 Is a Potential Prognostic Indicator of Recurrence of Human Breast Invasive Ductal Carcinoma with Her2-Enriched Subtype

    PubMed Central

    He, Weifeng; Tong, Yifan; Wang, Ying; Liu, Jingjing; Luo, Gaoxing; Wu, Jun; Zhang, Jin

    2013-01-01

    In clinical practice, breast cancers with lymph node positive, ER/PR-negative and overexpressed human epidermal growth factor receptor 2 (LN+ER/PR-Her2+) have high risk of recurrence, but the effective biomarkers of prognostic for this type tumor are still lacking. Since breast cancers with LN+ER/PR-Her2+ is at higher risk of recurrence than those with LN-ER/PR+Her2-. The differential proteins between those two groups could be related to the risk of recurrence. Herein, we report that serum soluble CD14 (sCD14) was revealed as the stable differential protein between LN+ER/PR-Her2+ (n=50) and LN-ER/PR+Her2- (n=50) breast cancer patients by proteomics analysis. To validate sCD14 as a biomarker for predicting recurrence of breast cancer, 90 breast cancer patients with LN+ER/PR-Her2+ and 93 patients with LN-ER/PR+Her2- were recruited. The patients with higher level of serum sCD14 at primary surgery showed to be at significantly lower risk of relapse in 3 years follow-up than those with lower level of serum sCD14 at primary surgery. The levels of serum sCD14 at primary surgery were significantly correlated to the risk of 3-year recurrence of LN+ER/PR-Her2+ breast cancer and the corresponding AUC of the ROC curve was 0.833 (95% CI, and 0.742 to 0.920). Therefore, we surmise that serum sCD14 could be a potential biomarker for predicting the prognosis of breast invasive ductal carcinoma with LN+ER/PR-Her2+. PMID:24086515

  8. Breastfeeding, soluble CD14 concentration in breast milk and risk of atopic dermatitis and asthma in early childhood: birth cohort study.

    PubMed

    Rothenbacher, D; Weyermann, M; Beermann, C; Brenner, H

    2005-08-01

    Breast milk contains a variety of bioactive substances, among them, soluble CD14 (sCD14), which plays an important role in innate immunity. We analysed data of a large prospective birth cohort study to examine the determinants of sCD14 in breast milk, and investigated whether breastfeeding practice and sCD14 concentrations in breast milk are determinants of the risk of atopic dermatitis (AD) and asthma in children. Eight hundred and three mothers and their newborns were included in this analysis. We measured sCD14 concentrations in breast milk samples collected 6 weeks post-partum. During a 2-year follow-up the cumulative incidences of AD and asthma were recorded. Overall, AD was reported for 20.6% of the 2-year-olds and asthma was reported for 19.6%. We found the lowest incidence of physician-reported AD in children of mothers without a history of atopic diseases if breastfed for 6 to less than 9 months. Furthermore, we found an inverse association between duration of breastfeeding and risk of asthma, which was especially evident in children with mothers without a history of atopic disease (P=0.01). These patterns persisted after control for other factors by multivariate analysis methods. The protective effect of breastfeeding seemed to be synergistic with sCD14 concentrations in breast milk (P for trend 0.0005). The results of this prospective birth cohort study suggest that a longer duration of breastfeeding does decrease the risk for asthma in early childhood, especially in children of mothers without a history of atopic disease. The beneficial effects of breastfeeding might be further supported by high levels of sCD14 in breast milk.

  9. Soluble CD14 and toll-like receptor-2 are potential salivary biomarkers for oral lichen planus and burning mouth syndrome.

    PubMed

    Srinivasan, Mythily; Kodumudi, Krithika N; Zunt, Susan L

    2008-01-01

    Oral lichen planus (OLP) and burning mouth syndrome (BMS) are chronic conditions affecting the oral mucosa characterized by pain and burning sensation. Saliva plays a significant role in the maintenance of physical and functional integrity of normal oral mucosa. Identification of potential "salivary biomarkers" for early diagnosis and/or monitoring of human diseases is being explored. We investigated the soluble forms of innate immune associated proteins CD14 and toll-like receptor-2 in unstimulated whole saliva (UWS) as potential biomarkers for OLP and BMS. Our results suggest that the levels of sCD14 and sTLR-2 in UWS were upregulated in OLP and BMS respectively. In addition, oral epithelial cells in the saliva of patients with OLP and BMS exhibited elevated levels of CD14 mRNA and decreased levels of TLR-2 mRNA. Interestingly, presence of co-existent oral candidiasis nullified these changes.

  10. Comparative Analysis of Hepatic CD14 Expression between Two Different Endotoxin Shock Model Mice: Relation between Hepatic Injury and CD14 Expression

    PubMed Central

    Hozumi, Hiroyasu; Tada, Rui; Murakami, Taisuke; Adachi, Yoshiyuki; Ohno, Naohito

    2013-01-01

    CD14 is a glycoprotein that recognizes gram-negative bacterial lipopolysaccharide (LPS) and exists in both membrane-bound and soluble forms. Infectious and/or inflammatory diseases induce CD14 expression, which may be involved in the pathology of endotoxin shock. We previously found that the expression of CD14 protein differs among the endotoxin shock models used, although the reasons for these differences are unclear. We hypothesized that the differences in CD14 expression might be due to liver injury, because the hepatic tissue produces CD14 protein. We investigated CD14 expression in the plasma and liver in the carrageenan (CAR)-primed and D-galN-primed mouse models of endotoxin shock. Our results showed that severe liver injury was not induced in CAR-primed endotoxin shock model mice. In this CAR-primed model, the higher mRNA and protein expression of CD14 was observed in the liver, especially in the interlobular bile duct in contrast to D-galN-primed-endotoxin shock model mice. Our findings indicated that the molecular mechanism(s) underlying septic shock in CAR-primed and D-galN-primed endotoxin shock models are quite different. Because CD14 expression is correlated with clinical observations, the CAR-primed endotoxin shock model might be useful for studying the functions of CD14 during septic shock in vivo. PMID:23308276

  11. Circulating classical CD14++CD16- monocytes predict shorter time to initial treatment in chronic lymphocytic leukemia patients: Differential effects of immune chemotherapy on monocyte-related membrane and soluble forms of CD163.

    PubMed

    Lapuc, Izabela; Bolkun, Lukasz; Eljaszewicz, Andrzej; Rusak, Malgorzata; Luksza, Ewa; Singh, Paulina; Miklasz, Paula; Piszcz, Jaroslaw; Ptaszynska-Kopczynska, Katarzyna; Jasiewicz, Malgorzata; Kaminski, Karol; Dabrowska, Milena; Bodzenta-Lukaszyk, Anna; Kloczko, Janusz; Moniuszko, Marcin

    2015-09-01

    Three main monocyte subsets: classical CD14++CD16-, intermediate CD14++CD16+ and non-classical CD14+CD16++, differentially regulate tumor growth and survival. Thereby, in the present study we aimed to determine the role of distinct monocyte subsets in the prognostication of chronic lymphocytic leukemia (CLL). Moreover, we set out to analyze the effects of standard immune chemotherapy on different monocyte subsets and levels of membrane-associated and soluble forms of CD163, a monocyte/macrophage-related immunomodulatory protein. We demonstrated that the number of peripheral blood classical CD14++CD16- monocytes assessed at the time of diagnosis was negatively correlated with lymphocytosis and was decreased in the CLL patients who required immediate treatment as opposed to patients who qualified to 'watch and wait' strategy. Notably, lower baseline levels of classical CD14++CD16- monocytes in CLL patients who were qualified for 'watch and wait' therapy were associated with shorter time to initial treatment. Notably, therapy with rituximab, cyclophosphamide and fludarabine resulted in a significant reduction in the number of non-classical CD14+CD16++ monocytes and soluble form of CD163 but upregulation of membrane-associated monocyte CD163. Our data indicate that distinct monocyte subsets and two forms of CD163 are differentially modulated by both CLL and immune chemotherapy. Moreover, we proposed that quantification of classical monocytes at the time of diagnosis contributes to better prognostication of CLL patients.

  12. Soluble CD83 Inhibits T Cell Activation by Binding to the TLR4/MD-2 Complex on CD14+ Monocytes

    PubMed Central

    Horvatinovich, Joe M.; Grogan, Elizabeth W.; Norris, Marcus; Steinkasserer, Alexander; Lemos, Henrique; Mellor, Andrew L.; Tcherepanova, Irina Y.; Nicolette, Charles A.

    2017-01-01

    The transmembrane protein CD83, expressed on APCs, B cells, and T cells, can be expressed as a soluble form generated by alternative splice variants and/or by shedding. Soluble CD83 (sCD83) was shown to be involved in negatively regulating the immune response. sCD83 inhibits T cell proliferation in vitro, supports allograft survival in vivo, prevents corneal transplant rejection, and attenuates the progression and severity of autoimmune diseases and experimental colitis. Although sCD83 binds to human PBMCs, the specific molecules that bind sCD83 have not been identified. In this article, we identify myeloid differentiation factor-2 (MD-2), the coreceptor within the TLR4/MD-2 receptor complex, as the high-affinity sCD83 binding partner. TLR4/MD-2 mediates proinflammatory signal delivery following recognition of bacterial LPSs. However, altering TLR4 signaling can attenuate the proinflammatory cascade, leading to LPS tolerance. Our data show that binding of sCD83 to MD-2 alters this signaling cascade by rapidly degrading IL-1R–associated kinase-1, leading to induction of the anti-inflammatory mediators IDO, IL-10, and PGE2 in a COX-2–dependent manner. sCD83 inhibited T cell proliferation, blocked IL-2 secretion, and rendered T cells unresponsive to further downstream differentiation signals mediated by IL-2. Therefore, we propose the tolerogenic mechanism of action of sCD83 to be dependent on initial interaction with APCs, altering early cytokine signal pathways and leading to T cell unresponsiveness. PMID:28193829

  13. Stable transduction of bovine TLR4 and bovine MD-2 into LPS-nonresponsive cells and soluble CD14 promote the ability to respond to LPS.

    PubMed

    Sauter, Kay-Sara; Brcic, Marija; Franchini, Marco; Jungi, Thomas W

    2007-07-15

    The interaction of bovine cells with lipopolysaccharide (LPS) was explored using human embryo kidney (HEK) 293 cell line stably transduced with bovine toll-like receptor-4 (TLR4) alone or in combination with bovine MD-2. These lines and mock-transduced HEK293 cells were tested by flow cytometry for LPS-fluorescein isothiocyanate (LPS-FITC) binding, nuclear factor kappa B (NFkappaB) activation, interleukin-8 (IL-8) production and interferon-beta mRNA expression/interferon (IFN) type I production. Whereas bovine TLR4 was sufficient to promote binding of high concentrations of LPS-FITC, both bovine TLR4 and MD-2 were required for activation by LPS, as assessed by NFkappaB activation and IL-8 production. Induction of IFN bioactivity was not observed in doubly transduced HEK293 cells, and no evidence for IFN-beta mRNA induction in response to LPS was obtained, although cells responded by IFN-beta mRNA expression to stimulation by Sendai virus and poly-inosinic acid-poly-cytidylic acid (poly(I:C)). Cells stably transduced with both bovine TLR4 and bovine MD-2 responded to LPS by IL-8 production, in decreasing order, in the presence of fetal bovine serum (FCS), of human serum, and of human serum albumin (HSA). The reduced activity in the presence of HSA could be restored by the addition of soluble CD14 (sCD14) but not of LPS binding protein (LBP). This is in contrast to macrophages which show a superior response to LPS in the presence of HSA when compared with macrophages stimulated by LPS in the presence of FCS. This suggests that macrophages but not HEK293 cells express factors rendering LPS stimulation serum-independent. Stably double-transduced cells reacted, in decreasing order, to LPS from Rhodobacter sphaeroides, to LPS from Escherichia coli, to synthetic lipd-IVa (compound 406), to diphosphoryl-lipid-A (S. minnesota) and to monophosphoryl-lipid-A (S. minnesota). They failed to react to the murine MD-2/TLR4 ligand taxol. This resembles the reactivity of bovine

  14. Presepsin (soluble CD14 subtype) and procalcitonin levels for mortality prediction in sepsis: data from the Albumin Italian Outcome Sepsis trial

    PubMed Central

    2014-01-01

    Introduction Sepsis, a leading cause of death in critically ill patients, is the result of complex interactions between the infecting microorganisms and the host responses that influence clinical outcomes. We evaluated the prognostic value of presepsin (sCD14-ST), a novel biomarker of bacterial infection, and compared it with procalcitonin (PCT). Methods This is a retrospective, case–control study of a multicenter, randomized clinical trial enrolling patients with severe sepsis or septic shock in ICUs in Italy. We selected 50 survivors and 50 non-survivors at ICU discharge, matched for age, sex and time from sepsis diagnosis to enrollment. Plasma samples were collected 1, 2 and 7 days after enrollment to assay presepsin and PCT. Outcome was assessed 28 and 90 days after enrollment. Results Early presepsin (day 1) was higher in decedents (2,269 pg/ml, median (Q1 to Q3), 1,171 to 4,300 pg/ml) than in survivors (1,184 pg/ml (median, 875 to 2,113); P = 0.002), whereas PCT was not different (18.5 μg/L (median 3.4 to 45.2) and 10.8 μg/L (2.7 to 41.9); P = 0.31). The evolution of presepsin levels over time was significantly different in survivors compared to decedents (P for time-survival interaction = 0.03), whereas PCT decreased similarly in the two groups (P = 0.13). Presepsin was the only variable independently associated with ICU and 28-day mortality in Cox models adjusted for clinical characteristics. It showed better prognostic accuracy than PCT in the range of Sequential Organ Failure Assessment score (area under the curve (AUC) from 0.64 to 0.75 vs. AUC 0.53 to 0.65). Conclusions In this multicenter clinical trial, we provide the first evidence that presepsin measurements may have useful prognostic information for patients with severe sepsis or septic shock. These preliminary findings suggest that presepsin may be of clinical importance for early risk stratification. PMID:24393424

  15. Elevated soluble CD14 and lower D-dimer are associated with cigarette smoking and heavy episodic alcohol use in persons living with HIV (PLWH)

    PubMed Central

    Cioe, Patricia A.; Baker, Jason; Kojic, E.; Onen, N.; Hammer, John; Patel, Pragna; Kahler, Christopher W.

    2015-01-01

    Background Persons living with HIV (PLWH) are at increased risk for cardiovascular disease in part due to persistent inflammation and coagulation activation. Methods We examined whether smoking and heavy episodic alcohol use (defined as 5 or more drinks on one occasion) were associated with greater monocyte activation (sCD14) and coagulation (D-dimer) in participants in the Study to Understand the Natural History of HIV and AIDS in the Era of Effective Therapy (the ‘SUN’ Study), a prospective observational cohort. Results Using regression analysis (n = 689), current smoking compared with nonsmoking was associated with significantly elevated sCD14 (B= 135.57, 95%CI (84.95, 186.19), p < .001); whereas heavy alcohol use compared to non-heavy use was associated with significantly lower D-dimer levels (B= −.059, 95%CI (−.102, −.016), p = .007). Conclusions Smoking cessation should be encouraged by HIV care providers to improve mortality outcomes from all causes of death, particularly cardiovascular disease. PMID:26181818

  16. Viable head and neck tumor spheroids stimulate in vitro autologous monocyte MCP-1 secretion through soluble substances and CD14/lectin-like receptors.

    PubMed

    Olsnes, Carla; Heimdal, John-Helge; Kross, Kenneth W; Olofsson, Jan; Aarstad, Hans Jørgen

    2005-12-01

    Biopsies from carcinoma tissue and benign control mucosa from head and neck squamous cell carcinoma (HNSCC) patients were used to establish fragment (F)-spheroids in vitro. We have previously shown that autologous monocytes co-cultured with F-spheroids in vitro augment their secretion of monocyte chemotactic protein-1 (MCP-1). Presently, the aims of the present work were to study whether the metabolic activity, secreted products and/or specific receptor/ligand on the surface of the F-spheroids and monocytes are necessary for stimulation of the monocyte MCP-1 secretion upon F-spheroid co-culture. Actinomycin D (1 mug/ml for 24 h) pre-treatment of the F-spheroids abolished the monocyte MCP-1 co-culture response. Co-culture of monocytes and F-spheroids separated by a semi-permeable membrane showed a decreased, but still present, monocyte MCP-1 co-culture response. Conditioned medium from F-spheroids stimulated allogenous monocytes to secrete MCP-1. The addition of glucose or galactose, but not mannose, to co-cultures partially inhibited the monocyte MCP-1 co-culture response. The addition of anti-CD14 antibody diminished the MCP-1 co-culture response. In conclusion, the monocyte MCP-1 co-culture response is dependent on metabolically active spheroids, secreted stimuli, and is augmented by direct contact with F-spheroids, possibly via lectin-like receptors and the CD14 receptor.

  17. Brief Report: CD14brightCD16- monocytes and sCD14 level negatively associate with CD4-memory T-cell frequency and predict HCV-decline on therapy.

    PubMed

    Judge, Chelsey J; Sandberg, Johan K; Funderburg, Nicholas T; Sherman, Kenneth E; Butt, Adeel A; Kang, Minhee; Landay, Alan L; Lederman, Michael M; Anthony, Donald D

    2016-11-01

    During HIV+ hepatitis C virus (HCV)+ coinfection CD14CD16 monocytes produce soluble immune-activation markers that predict disease progression and poor response to interferon (IFN)-α treatment. We evaluated relationships among immune activation, monocyte phenotype, CD4-memory T cells, and HCV-, cytomegalovirus-, and cytomegalovirus/Epstein-Barr virus/influenza-specific IFN-γ-response before and during IFN-α treatment. Effector-memory and central-memory CD4 T-cell frequencies were lower in HCV+ HIV+ donors than in uninfected donors and correlated negatively with HCV level, CD14CD16 monocytes, and plasma sCD14. sCD14 and CD14CD16 monocytes negatively correlated with IFN-α-dependent HCV decline. CD4 effector-memory T cells positively associated with cytomegalovirus/Epstein-Barr virus/influenza(CEF)-specific IFN-γ response, while sCD14 negatively associated with both CD4 effector-memory T cells and CEF-specific IFN-γ response. These data support a role for memory-CD4 T cells in HCV containment and link immune activation and CD14CD16-monocyte frequency to the failure of IFN-dependent HCV clearance.

  18. The predictive value of soluble biomarkers (CD14 subtype, interleukin-2 receptor, human leucocyte antigen-G) and procalcitonin in the detection of bacteremia and sepsis in pediatric oncology patients with chemotherapy-induced febrile neutropenia.

    PubMed

    Urbonas, Vincas; Eidukaitė, Audronė; Tamulienė, Indrė

    2013-04-01

    Prediction of bacteremia/sepsis in childhood oncology patients with febrile neutropenia still remains a challenge for the medical community due to the lack of reliable biomarkers, especially at the beginning of infectious process. The objective of this study was to evaluate diagnostic value of soluble biomarkers (CD14 subtype, interleukin-2 receptor, HLA-G) and procalcitonin (PCT) in the identification of infectious process at the beginning of a febrile episode in pediatric oncology patients. A total of 62 episodes of febrile neutropenia in 37 childhood oncology patients were enrolled in this study. Serum samples were collected at presentation after confirmation of febrile neutropenia and analyzed according to recommendations of manufacturers. Patients were classified into bacteremia/sepsis and fever of unknown origin groups. Median of PCT and sIL-2R were considerably higher in bacteremia/sepsis group compared to fever of unknown origin group, whereas median of sHLA-G and presepsin levels between investigated groups did not differ sufficiently. PCT and sIL-2R determination might be used as an additional diagnostic tool for the detection of bacteremia/sepsis in childhood oncology patients with febrile neutropenia. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. CD14 and IL18 gene polymorphisms associated with colorectal cancer subsite risks among atomic bomb survivors.

    PubMed

    Hu, Yiqun; Yoshida, Kengo; Cologne, John B; Maki, Mayumi; Morishita, Yukari; Sasaki, Keiko; Hayashi, Ikue; Ohishi, Waka; Hida, Ayumi; Kyoizumi, Seishi; Kusunoki, Yoichiro; Tokunaga, Katsushi; Nakachi, Kei; Hayashi, Tomonori

    2015-01-01

    Colorectal cancer (CRC) is a common malignancy worldwide, and chronic inflammation is a risk factor for CRC. In this study, we carried out a cohort study among the Japanese atomic bomb (A-bomb) survivor population to investigate any association between immune- and inflammation-related gene polymorphisms and CRC. We examined the effects of six single-nucleotide polymorphisms of CD14 and IL18 on relative risks (RRs) of CRC. Results showed that RRs of CRC, overall and by anatomic subsite, significantly increased with increasing radiation dose. The CD14-911A/A genotype showed statistically significant higher risks for all CRC and distal CRC compared with the other two genotypes. In addition, the IL18-137 G/G genotype showed statistically significant higher risks for proximal colon cancer compared with the other two genotypes. In phenotype-genotype analyses, the CD14-911A/A genotype presented significantly higher levels of membrane and soluble CD14 compared with the other two genotypes, and the IL18-137 G/G genotype tended to be lower levels of plasma interleukin (IL)-18 compared with the other two genotypes. These results suggest the potential involvement of a CD14-mediated inflammatory response in the development of distal CRC and an IL18-mediated inflammatory response in the development of proximal colon cancer among A-bomb survivors.

  20. CD14 and IL18 gene polymorphisms associated with colorectal cancer subsite risks among atomic bomb survivors

    PubMed Central

    Hu, Yiqun; Yoshida, Kengo; Cologne, John B; Maki, Mayumi; Morishita, Yukari; Sasaki, Keiko; Hayashi, Ikue; Ohishi, Waka; Hida, Ayumi; Kyoizumi, Seishi; Kusunoki, Yoichiro; Tokunaga, Katsushi; Nakachi, Kei; Hayashi, Tomonori

    2015-01-01

    Colorectal cancer (CRC) is a common malignancy worldwide, and chronic inflammation is a risk factor for CRC. In this study, we carried out a cohort study among the Japanese atomic bomb (A-bomb) survivor population to investigate any association between immune- and inflammation-related gene polymorphisms and CRC. We examined the effects of six single-nucleotide polymorphisms of CD14 and IL18 on relative risks (RRs) of CRC. Results showed that RRs of CRC, overall and by anatomic subsite, significantly increased with increasing radiation dose. The CD14–911A/A genotype showed statistically significant higher risks for all CRC and distal CRC compared with the other two genotypes. In addition, the IL18–137 G/G genotype showed statistically significant higher risks for proximal colon cancer compared with the other two genotypes. In phenotype–genotype analyses, the CD14–911A/A genotype presented significantly higher levels of membrane and soluble CD14 compared with the other two genotypes, and the IL18–137 G/G genotype tended to be lower levels of plasma interleukin (IL)-18 compared with the other two genotypes. These results suggest the potential involvement of a CD14-mediated inflammatory response in the development of distal CRC and an IL18-mediated inflammatory response in the development of proximal colon cancer among A-bomb survivors. PMID:27081544

  1. Sevelamer Does Not Decrease Lipopolysaccharide or Soluble CD14 Levels But Decreases Soluble Tissue Factor, Low-Density Lipoprotein (LDL) Cholesterol, and Oxidized LDL Cholesterol Levels in Individuals With Untreated HIV Infection

    PubMed Central

    Sandler, Netanya G.; Zhang, Xinyan; Bosch, Ronald J.; Funderburg, Nicholas T.; Choi, Andrew I.; Robinson, Janet K.; Fine, Derek M.; Coombs, Robert W.; Jacobson, Jeffrey M.; Landay, Alan L.; Douek, Daniel C.; Tressler, Randall; Read, Sarah W.; Wilson, Cara C.; Deeks, Steven G.; Lederman, Michael M.; Gandhi, Rajesh T.

    2014-01-01

    Abnormal levels of inflammation are associated with cardiovascular disease and mortality in human immunodeficiency virus (HIV)–infected patients. Microbial translocation, which may cause inflammation, is decreased by sevelamer in patients undergoing hemodialysis. In this single-arm study, we evaluated the effects of 8 weeks of sevelamer therapy on 36 HIV-infected subjects who were not receiving antiretroviral therapy. Sevelamer did not significantly change markers of microbial translocation, inflammation, or T-cell activation. During sevelamer treatment, however, levels of soluble tissue factor, low-density lipoprotein (LDL) cholesterol, and oxidized LDL cholesterol decreased significantly, whereas D-dimer levels increased. Thus, in this study population, sevelamer did not reduce microbial translocation but may have yielded cardiovascular benefits. Clinical Trials Registration. NCT 01543958. PMID:24864123

  2. Polymorphisms in the CD14 and TLR4 genes independently predict CD4+ T-cell recovery in HIV-infected individuals on antiretroviral therapy

    PubMed Central

    YONG, Yean K; SHANKAR, Esaki M; SOLOMON, Ajantha; SPELMAN, Tim; FAIRLEY, Christopher K; ELLIOTT, Julian; HOY, Jennifer; CAMERON, Paul; KAMARULZAMAN, Adeeba; LEWIN, Sharon R

    2016-01-01

    Background Chronic HIV infection leads to marked depletion of CD4+ T cells in the gastrointestinal (GI) tract and increased microbial translocation measured by an increase in circulating lipopolysaccharide (LPS) levels. Here, we hypothesised that single-nucleotide polymorphisms (SNPs) in genes encoding the Toll-like receptor 4 (TLR4) and CD14, the principal receptors for LPS, were associated with CD4+ T-cell recovery post-antiretroviral therapy (ART). Methods Prospective study of predominantly Caucasian HIV-infected subjects receiving suppressive ART for at least 12 months. We analysed the CD14 SNPs C-260T and the TLR4 SNPs A+896G, C+1196T. We also determined the levels of LPS and soluble CD14 (sCD14) in plasma samples collected pre- and post-ART initiation. CD4+ T-cell recovery was assessed by linear mixed models. Results Following ART, individuals with a TT genotype compared to a CT or CC genotype for CD14 C-260T SNP showed higher levels of sCD14 (p=0.008 and 0.003 respectively). The CC genotype for the CD14 C-260T SNP, compared to CT or TT and the TLR4 SNP (AC/GT) compared to the homozygous genotype (AA/CC) were both independently associated with enhanced long-term CD4+ T-cell recovery (>3 months; p<0.001). Conclusions Polymorphisms in CD14 and TLR4 are independently associated with long-term CD4+ T-cell recovery in HIV-infected individuals post-ART. PMID:27281059

  3. Polymorphisms in the CD14 and TLR4 genes independently predict CD4+ T-cell recovery in HIV-infected individuals on antiretroviral therapy.

    PubMed

    Yong, Yean K; Shankar, Esaki M; Solomon, Ajantha; Spelman, Tim; Fairley, Christopher K; Elliott, Julian H; Hoy, Jennifer; Cameron, Paul U; Kamarulzaman, Adeeba; Lewin, Sharon R

    2016-09-10

    Chronic HIV infection leads to marked depletion of CD4 T cells in the gastrointestinal tract and increased microbial translocation measured by an increase in circulating lipopolysaccharide (LPS) levels. Here, we hypothesized that single-nucleotide polymorphisms (SNPs) in genes encoding the Toll-like receptor 4 (TLR4) and CD14, the principal receptors for LPS, were associated with CD4 T-cell recovery postantiretroviral therapy (ART). Prospective study of predominantly white HIV-infected participants receiving suppressive ART for at least 12 months. We analysed the CD14 SNPs C-260T and the TLR4 SNPs A+896G, C+1196T. We also determined the levels of LPS and soluble CD14 in plasma samples collected pre-ART and post-ART initiation. CD4 T-cell recovery was assessed by linear mixed models. Following ART, individuals with a TT genotype compared with a CT or CC genotype for CD14 C-260T SNP showed higher levels of soluble CD14 (P = 0.008 and 0.003, respectively). The CC genotype for the CD14 C-260T SNP, compared with CT or TT, and the TLR4 SNP (AC/GT), compared with the homozygous genotype (AA/CC), were both independently associated with enhanced long-term CD4 T-cell recovery (>3 months; P < 0.001). Polymorphisms in CD14 and TLR4 are independently associated with long-term CD4 T-cell recovery in HIV-infected individuals post-ART.

  4. Human CD14 mediates recognition and phagocytosis of apoptotic cells.

    PubMed

    Devitt, A; Moffatt, O D; Raykundalia, C; Capra, J D; Simmons, D L; Gregory, C D

    1998-04-02

    Cells undergoing programmed cell death (apoptosis) are cleared rapidly in vivo by phagocytes without inducing inflammation. Here we show that the glycosylphosphatidylinositol-linked plasma-membrane glycoprotein CD14 on the surface of human macrophages is important for the recognition and clearance of apoptotic cells. CD14 can also act as a receptor that binds bacterial lipopolysaccharide (LPS), triggering inflammatory responses. Overstimulation of CD14 by LPS can cause the often fatal toxic-shock syndrome. Here we show that apoptotic cells interact with CD14, triggering phagocytosis of the apoptotic cells. This interaction depends on a region of CD14 that is identical to, or at least closely associated with, a region known to bind LPS. However, apoptotic cells, unlike LPS, do not provoke the release of pro-inflammatory cytokines from macrophages. These results indicate that clearance of apoptotic cells is mediated by a receptor whose interactions with 'non-self' components (LPS) and 'self' components (apoptotic cells) produce distinct macrophage responses.

  5. Serum CD14 levels in polytraumatized and severely burned patients.

    PubMed Central

    Krüger, C; Schütt, C; Obertacke, U; Joka, T; Müller, F E; Knöller, J; Köller, M; König, W; Schönfeld, W

    1991-01-01

    Recently it has been demonstrated that the CD14 molecule which is expressed on monocytes and macrophages serves as a receptor for lipopolysaccharide (LPS) bound to LPS-binding protein (LBP) and thus mediates LPS-induced tumour necrosis factor (TNF) production. Here we report that CD14 is found as a soluble (s) molecule in serum. In healthy volunteers sCD14 levels (mean +/- s.e.m.) were 3.7 +/- 0.05 micrograms/ml (n = 30, 25-50 years of age) as determined by ELISA (detection limit 20 ng/ml serum) using two monoclonal antibodies in a sandwich technique. In polytraumatized patients (n = 16) significantly decreased levels (1.7 +/- 0.3) were detected immediately after the trauma, which increased to 4.9 +/- 0.3 micrograms/ml within the first 6 days post trauma. sCD14 remained elevated during the first 14 days post trauma in patients with the most severe injuries (injury severity score greater than 45 points), whereas a return to normal levels was observed in patients with an injury score of less than 45 points. In addition, the levels of the high-density lipoproteins that partially inactivate free endotoxin are significantly decreased post trauma. No correlation between parameters of inflammation (C3a and neopterin levels, leucocyte counts, amount of band cells), liver function and sCD14 levels was established. Comparable to polytraumatized patients, increased sCD14 serum levels were observed in five patients with burn trauma (burned area greater than 35%) within the second week post trauma when clinical signs of septicaemia were evident. PMID:1713813

  6. The N-terminal half of membrane CD14 is a functional cellular lipopolysaccharide receptor.

    PubMed

    Viriyakosol, S; Kirkland, T N

    1996-02-01

    CD14, a glycosylphosphatidylinositol-anchored protein on the surface of monocytes, macrophages, and polymorphonuclear leukocytes, is a receptor for lipopolysaccharide (LPS). It was recently reported that an N-terminal 152-amino-acid fragment of soluble CD14 was an active soluble lipopolysaccharide receptor (T. S. -C. Juan, M. J. Kelley, D. A. Johnson, L. A. Busse, E. Hailman, S. D. Wright, and H. S. Lichenstein, J. Biol. Chem. 270:1382-1387, 1995). To determine whether the N-terminal half of the membrane CD14 was a functional LPS receptor on the cell membrane, we engineered a chimeric gene coding for amino acids 1 to 151 of CD14 fused to the C-terminal region of decay-accelerating factor and expressed it in Chinese hamster ovary cells and 70Z/3 cells. We found that the chimeric, truncated CD14 is a fully functional LPS receptor in both cell lines.

  7. CD14 Directs Adventitial Macrophage Precursor Recruitment: Role in Early Abdominal Aortic Aneurysm Formation

    PubMed Central

    Blomkalns, Andra L.; Gavrila, Daniel; Thomas, Manesh; Neltner, Bonnie S.; Blanco, Victor M.; Benjamin, Stephanie B.; McCormick, Michael L.; Stoll, Lynn L.; Denning, Gerene M.; Collins, Sean P.; Qin, Zhenyu; Daugherty, Alan; Cassis, Lisa A.; Thompson, Robert W.; Weiss, Robert M.; Lindower, Paul D.; Pinney, Susan M.; Chatterjee, Tapan; Weintraub, Neal L.

    2013-01-01

    Background Recruitment of macrophage precursors to the adventitia plays a key role in the pathogenesis of abdominal aortic aneurysms (AAAs), but molecular mechanisms remain undefined. The innate immune signaling molecule CD14 was reported to be upregulated in adventitial macrophages in a murine model of AAA and in monocytes cocultured with aortic adventitial fibroblasts (AoAf) in vitro, concurrent with increased interleukin‐6 (IL‐6) expression. We hypothesized that CD14 plays a crucial role in adventitial macrophage precursor recruitment early during AAA formation. Methods and Results CD14−/− mice were resistant to AAA formation induced by 2 different AAA induction models: aortic elastase infusion and systemic angiotensin II (AngII) infusion. CD14 gene deletion led to reduced aortic macrophage infiltration and diminished elastin degradation. Adventitial monocyte binding to AngII‐infused aorta in vitro was dependent on CD14, and incubation of human acute monocytic leukemia cell line‐1 (THP‐1) monocytes with IL‐6 or conditioned medium from perivascular adipose tissue (PVAT) upregulated CD14 expression. Conditioned medium from AoAf and PVAT induced CD14‐dependent monocyte chemotaxis, which was potentiated by IL‐6. CD14 expression in aorta and plasma CD14 levels were increased in AAA patients compared with controls. Conclusions These findings link CD14 innate immune signaling via a novel IL‐6 amplification loop to adventitial macrophage precursor recruitment in the pathogenesis of AAA. PMID:23537804

  8. Derivation of multipotent progenitors from human circulating CD14+ monocytes.

    PubMed

    Seta, Noriyuki; Kuwana, Masataka

    2010-07-01

    Circulating CD14(+) monocytes are originated from hematopoietic stem cells in the bone marrow and believed to be committed precursors for phagocytes, such as macrophages. Recently, we have reported a primitive cell population termed monocyte-derived multipotential cells (MOMCs), which has a fibroblast-like morphology in culture and a unique phenotype positive for CD14, CD45, CD34, and type I collagen. MOMCs are derived from circulating CD14(+) monocytes, but circulating precursors for MOMCs still remain undetermined. Comparative analysis of gene expression profiles of MOMCs and other monocyte-derived cells has revealed that embryonic stem cell markers, Nanog and Oct-4, are specifically expressed by MOMCs. In vitro generation of MOMCs requires binding to fibronectin and exposure to soluble factors derived from activated platelets. MOMCs contain progenitors with capacity to differentiate into a variety of nonphagocytes, including bone, cartilage, fat, skeletal and cardiac muscle, neuron, and endothelium, indicating that circulating monocytes are more multipotent than previously thought. In addition, MOMCs are capable of promoting ex vivo expansion of human hematopoietic progenitor cells through direct cell-to-cell contact and secretion of a variety of hematopoietic growth factors. These findings obtained from the research on MOMCs indicate that CD14(+) monocytes in circulation are involved in a variety of physiologic functions other than innate and acquired immune responses, such as repair and regeneration of the damaged tissue.

  9. Pet keeping and tobacco exposure influence CD14 methylation in childhood.

    PubMed

    Munthe-Kaas, Monica Cheng; Bertelsen, Randi J; Torjussen, Tale Maehre; Hjorthaug, Hanne Sagsveen; Undlien, Dag E; Lyle, Robert; Gervin, Kristina; Granum, Berit; Mowinckel, Petter; Carlsen, Kai Håkon; Carlsen, Karin C Lødrup

    2012-12-01

    Several CD14 gene-environment interactions in relation to the development of allergic diseases have been reported, but the underlying biological mechanisms are unclear. We recently showed that CD14 methylation increased during childhood, parallelling a decreased impact of CD14 polymorphisms on soluble CD14 levels. Here, we aim to explore whether environmental stimuli during childhood affects CD14 methylation, thereby providing a biological mechanism through which environment may modulate genetic effect. CD14 methylation levels were quantified in 157 children from the prospective Environment and Childhood Asthma birth cohort at ages 2 and 10. Associations between CD14 methylation levels and house dust levels of endotoxin, β(1,3)-glucans (at 2 yr only), allergens (dog, cat, and house dust mite), pet keeping and tobacco smoke exposure (TSE; questionnaire data) at 2 and 10 yr were explored. Children in homes without pets had larger increases in CD14 methylation through childhood (2-10 yr) compared with children with pets (2.1% increase (p = 0.003) vs. 0.4% decrease (n.s.), global p = 0.04). At 10 yr of age, lower CD14 methylation values were found in children with pets compared with children without pets at both 2 and 10 yr (5.4% vs. 7.5% [p = 0.02]). A similar trend was detected for TSE; children not exposed show larger increases in CD14 methylation, most pronounced in school-age girls exposed vs. not exposed to tobacco (5.5% vs. 7.5% methylation, p = 0.037). Pet keeping and TSE appears to limit increase in CD14 methylation from 2 to 10 yr of age. This may partly explain the diverging CD14 allele associations with allergic diseases detected in different environments. © 2012 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

  10. Enhanced host immune recognition of mastitis causing Escherchia coli in CD-14 transgenic mice.

    PubMed

    Wall, R; Powell, A; Sohn, E; Foster-Frey, J; Bannerman, D; Paape, M

    2009-01-01

    Escherchia coli causes mastitis, an economically significant disease in dairy animals. E. coli endotoxin (lipopolysaccharide, LPS) when bound by host membrane proteins such as CD-14, causes release of proinflammatory cytokines recruiting neutrophils as an early, innate immune response. Excessive proinflammatory cytokines causes tissue damage, compromising mammary function. If present, soluble CD-14 (sCD-14) might out compete membrane bound CD-14, lessening the severity of the inflammatory response. To test this hypothesis transgenic mice, expressing sCD-14 in their milk (31 to 316 microg/ml), were evaluated. A cell culture study demonstrated, in the presence of LPS, milk from transgenic mice increased secretion of cytokine IL-8 compared to milk from nontransgenic littermates (18 +/- 3 vs. 35 +/- 2 ng/mL, p < 0.001). To assess protection afforded by the transgene, glands were infused with E. coli. Recovery of bacteria showed no clear relationship between sCD14 concentration and the number of organisms recovered; however, there was a strong relationship between sCD14 concentration and edema (r(2) = 0.999, p < 0.001), as measured by weight of fluid in harvested glands. Highest expressing lines had the least edema, suggesting the presence of sCD14 had an effect on reducing the inflammatory response to E. coli, thus, possibly protecting against gland tissue damage.

  11. TLR4/CD14 Variants-Related Serologic and Immunologic Dys-Regulations Predict Severe Sepsis in Febrile De-Compensated Cirrhotic Patients

    PubMed Central

    Fan, Wen-Chien; Liu, Chih-Wei; Ou, Shuo-Ming; Huang, Chia-Chang; Li, Tzu-Hao; Lee, Kuei-Chuan; Huang, Shiang-Fen; Yang, Ying-Ying; Hsieh, Yun-Cheng; Hsieh, Shie-Liang; Hou, Ming-Chih; Lin, Han-Chieh

    2016-01-01

    Genetic variants and dysfunctional monocyte had been reported to be associated with infection susceptibility in advanced cirrhotic patients. This study aims to explore genetic predictive markers and relevant immune dysfunction that contributed to severe sepsis in febrile acute de-compensated cirrhotic patents. Polymorphism analysis of candidate genes was undergone in 108 febrile acute de-compensated cirrhotic patients and 121 healthy volunteers. Various plasma inflammatory/regulatory cytokines, proportion of classical (CD 16-, phagocytic) and non-classical (CD16+, inflammatory) monocytes, lipopolysaccharide (LPS)-stimulated toll-like receptor 4 (TLR4) and intracellular/extracellular cytokines on cultured non-classical monocytes, mCD14/HLA-DR expression and phagocytosis of classical monocytes were measured. For TLR4+896A/G variant allele carriers with severe sepsis, high plasma endotoxin/IL-10 inhibits HLA-DR expression and impaired phagocytosis were noted in their classical monocyte. In the same group, increased non-classical monocyte subset, enhanced LPS-stimulated TLR4 expression and TNFα/nitrite production, and systemic inflammation [high plasma soluble CD14 (sCD14) and total nitric oxide (NOx) levels] were noted. For CD14-159C/T variant allele carriers with severe sepsis, persist endotoxemia inhibited mCD14/HLA-DR expression and impaired phagocytosis of their classical monocyte. In the same group, increased non-classical monocyte subset up-regulated TLR4-NFκB-iNOS and p38MAPK pathway, stimulated TNFα/nitrite production and elicited systemic inflammation. In febrile acute de-compensated cirrhotic patients, TLR4+896A/G and CD14-159C/T polymorphisms-related non-classical and classical monocytes dysfunction resulted in increased severe sepsis risk. Malnutrition, high plasma endotoxin and sCD14 levels, single TLR4+896A/G or CD14-159C/T variant allele carriers and double variant allele carriers are significant predictive factors for the development of severe

  12. The CD14 C-260T single nucleotide polymorphism (SNP) modulates monocyte/macrophage activation in treated HIV-infected individuals.

    PubMed

    Rajasuriar, Reena; Kong, Yong Yean; Nadarajah, Reshika; Abdullah, Noor Kamila; Spelman, Tim; Yuhana, Muhamad Yazli; Ponampalavanar, Sasheela; Kamarulzaman, Adeeba; Lewin, Sharon R

    2015-01-27

    HIV-infected individuals have an increased risk of cardiovascular disease (CVD). T-allele carriers of the CD14 C-260T single-nucleotide polymorphism (SNP) have reported increased expression of the LPS-binding receptor, CD14 and inflammation in the general population. Our aim was to explore the relationship of this SNP with monocyte/macrophage activation and inflammation and its association with sub-clinical atherosclerosis in HIV-infected individuals. Patients with no pre-existing CVD risk factors on suppressive antiretroviral therapy were recruited from University Malaya Medical Centre, Malaysia (n = 84). The CD14 C-260T and TLR4 SNPs, Asp299Gly and Thr399Ile were genotyped and soluble(s) CD14 and sCD163 and high-sensitivity C-reactive protein, hsCRP were measured in plasma. Subclinical atherosclerosis was assessed by measuring carotid intima media thickness (cIMT). The association between CD14 C-260T SNP carriage and cIMT was assessed in a multivariable quantile regression model where a p-value of <0.05 was considered significant. We found the CD14 C-260T T-allele in 56% of the cohort and evidence of subclinical atherosclerosis in 27%. TT genotype was associated with higher sCD163 (p = 0.009) but only marginally higher sCD14 (p = 0.209) and no difference in hsCRP (p = 0.296) compared to CC/CT. In multivariable analysis, only Framingham risk score was independently associated with higher cIMT while lower sCD163 was trending towards significance. No association was found in TT-genotype carriers and cIMT measurements. The CD14 C-260T SNP was associated with increased monocyte activation but not systemic inflammation or cIMT in this HIV-infected cohort with low CVD risk profile.

  13. Renal excretion of plasma soluble melanins by healthy human adults.

    PubMed

    Hegedus, Z L; Nayak, U

    1993-01-01

    The soluble melanins of blood plasma form in vivo and in vitro from dopa, catecholamines, catechol, hydroquinone, homogentisic acid, 3-hydroxykynurenine, 3-hydroxyanthranilic acid, p-aminophenol, p-phenylenediamine and other structurally related end(ex)ogenous compounds by oxidative polymerization. The mean quantity of natural melanins in normal plasma is 1.61 +/- 0.10 (standard deviation) mg/ml, (n = 20) and in uraemic plasma 2.72 +/- 0.38 mg/ml, (n = 16). The plasma melanins (approximately 3%), are associated with proteins (approximately 85%), mucoproteins (approximately 0.25%), lipids (approximately 0.4%), as soluble lipofuscins, and probably are associated with proteins without lipids as soluble melanoproteins. Fluorescence, UV-VIS and IR spectroscopies and the melanin isolation method show the presence of soluble melanins in the urine of healthy people. Soluble melanins can also be formed in vitro in the urine by oxidative polymerization of the precursors. In most of the urine samples we studied, melanins were present in larger amounts than the urinary proteins, indicating that the kidneys can selectively excrete the melanin components of the lipofuscins, and that the solubility of melanins does not depend upon combination with proteins. The quantities of purified melanins precipitated with 6 N HCl at 110 degrees C during 72 h from urine samples collected during 24 h periods ranged from 0.1460 g to 3.7627 g (mean 1.1303 +/- 1.1739 g, n = 8) and the plasma clearance rates ranged from 0.06 ml/min to 1.56 ml/min (mean 0.48 +/- 0.48 ml/min, n = 8). From the individual 24 h urine samples we obtained from 9 to 216 mg/dl of precipitated melanins while the individual plasma samples contained from 145 to 175 mg/dl.

  14. Combined Plasma Elevation of CRP, Intestinal-Type Fatty Acid-Binding Protein (I-FABP), and sCD14 Identifies Older Patients at High Risk for Healthcare-Associated Infections.

    PubMed

    Elena, Paillaud; Sylvie, Bastuji-Garin; Anne, Plonquet; Emile, Foucat; Bénédicte, Fournier; Emmanuelle, Boutin; Aurélie, Le Thuaut; Yves, Levy; Sophie, Hue

    2017-06-03

    We hypothesized that low-grade inflammation was driven by microbial translocation and associated with an increased risk of healthcare-associated infections (HAIs). We included 121 patients aged 75 years or over in this prospective cohort study. High-sensitivity C-reactive protein (hs-CRP), I-FABP, and sCD14 -- as markers for low-grade inflammation, intestinal epithelial barrier integrity, and monocyte activation, respectively -- were measured at admission. HAIs occurred during hospitalization in 62 (51%) patients. Elevated hs-CRP (≥6.02 mg/L, i.e., the median) was associated with a significantly higher HAI risk when I-FABP was in the highest quartile (odds ratio [OR], 4; 95% confidence interval [95%CI], 1.39-11.49; P=0.010). In patients with hs-CRP elevation and highest-quartile I-FABP, sCD14 elevation (≥0.65 pg/mL, i.e., the median) was associated with an 11-fold higher HAI risk (OR, 10.8; 95%CI, 2.28-51.1; P=0.003). Multivariate analyses adjusted for invasive procedures and comorbidities did not change the associations linking the three markers to the HAI risk. Increased levels of hs-CRP, I-FABP, and sCD14 may reflect loss of intestinal epithelial barrier integrity with microbial translocation leading to monocyte activation and low-grade inflammation. In our cohort, these markers identified patients at high risk for HAIs.

  15. CD14, a Receptor for Complexes of Lipopolysaccharide (LPS) and LPS Binding Protein

    NASA Astrophysics Data System (ADS)

    Wright, Samuel D.; Ramos, Robert A.; Tobias, Peter S.; Ulevitch, Richard J.; Mathison, John C.

    1990-09-01

    Leukocytes respond to lipopolysaccharide (LPS) at nanogram per milliliter concentrations with secretion of cytokines such as tumor necrosis factor-α (TNF-α). Excess secretion of TNF-α causes endotoxic shock, an often fatal complication of infection. LPS in the bloodstream rapidly binds to the serum protein, lipopolysaccharide binding protein (LBP), and cellular responses to physiological levels of LPS are dependent on LBP. CD14, a differentiation antigen of monocytes, was found to bind complexes of LPS and LBP, and blockade of CD14 with monoclonal antibodies prevented synthesis of TNF-α by whole blood incubated with LPS. Thus, LPS may induce responses by interacting with a soluble binding protein in serum that then binds the cell surface protein CD14.

  16. Establishment of the cell line, HeLa-CD14, transfected with the human CD14 gene.

    PubMed

    Ning, Bo-Tao; Tang, Yong-Min

    2012-04-01

    CD14 is the pivotal molecule in the diagnosis and therapy of CD14-associated diseases, and is important in bacteremia. The HeLa cell line is regarded as immortal due to its prolific character. The HeLa cell line is derived from human cervical cancer cells and has been widely used in cancer research and gene transfection. In the present study, we established the expression plasmid pcDNA3.1(+)-CD14, and transfected it into the human cervical cancer cell line HeLa to establish a stable cell line (HeLa-CD14) expressing human CD14 antigen on the membrane. After the human CD14 gene was cloned and sequenced through RT-PCR and T-A cloning techniques, the eukaryotic expression vector pcDNA3.1(+)-CD14 was constructed by cleaving with double restriction endonucleases and ligating with T4 ligase. HeLa cells were transfected with the pcDNA3.1(+)-CD14 recombinant plasmid using Superfect transfection reagent. The cells were selected using G418 and the expression of human CD14 on the transfectant was confirmed by RT-PCR and immunohistochemistry. The expression of CD14 mRNA was significantly different between the blank pcDNA3.1(+)-transfected cell group and the pcDNA3.1(+)-CD14-transfected cell group (p<0.01). The fluorescence was significantly stronger on the established stable cell line than on the transiently transfected HeLa cells, and no visible fluorescence was observed in blank pcDNA3.1(+)-transfected cells. In this study, the human CD14 transfectant, stable cell line HeLa-CD14, was successfully established, which may be used to study CD14 and cervical cancer in vitro and in vivo.

  17. CNS invasion by CD14+/CD16+ peripheral blood-derived monocytes in HIV dementia: perivascular accumulation and reservoir of HIV infection.

    PubMed

    Fischer-Smith, T; Croul, S; Sverstiuk, A E; Capini, C; L'Heureux, D; Régulier, E G; Richardson, M W; Amini, S; Morgello, S; Khalili, K; Rappaport, J

    2001-12-01

    Increases in circulating CD14+/CD16+ monocytes have been associated with HIV dementia; trafficking of these cells into the CNS has been proposed to play an important role in the pathogenesis of HIV-induced neurological disorders. This model suggests that events outside the CNS leading to monocyte activation initiate the process leading to HIV dementia. To investigate the role of this activated monocyte subset in the pathogenesis of HIV dementia, we examined brain specimens from patients with HIV encephalopathy (HIVE), HIV without encephalopathy, and seronegative controls. An accumulation of perivascular macrophages was observed in HIVE. The majority of these cells identified in microglial nodules and in the perivascular infiltrate were CD14+/CD16+. P24 antigen colocalized with both CD14 and CD16 suggesting that the CD14+/CD16+ macrophage is a major reservoir of HIV-1 infection in CNS. Using CD45/LCA staining, the perivascular macrophage was distinguished from resident microglia. In addition to perivascular and nodular localizations, CD16 also stained ramified cells throughout the white matter. These cells were more ramified and abundant than cells positive for CD14 in white matter. Double staining for p24 and CD16 suggests that these cells were often infected with HIV-1. The prominent distribution of CD14+ cells in HIVE prompted our analysis of soluble CD14 levels in cerebrospinal fluid. Higher levels of soluble CD14 (sCD14) were observed in patients with moderate-to-severe HIV dementia, suggesting the utility of sCD14 as a surrogate marker. CD14+/CD16+ monocytes may play a role in other neurological disorders and sCD14 may be useful for evaluating these conditions.

  18. Circulating levels of the innate and humoral immune regulators CD14 and CD23 are associated with adult glioma.

    PubMed

    Zhou, Mi; Wiemels, Joseph L; Bracci, Paige M; Wrensch, Margaret R; McCoy, Lucie S; Rice, Terri; Sison, Jennette D; Patoka, Joseph S; Wiencke, John K

    2010-10-01

    Allergy history has been consistently inversely associated with glioma risk. Two serologic markers, soluble CD23 (sCD23) and soluble CD14 (sCD14), are part of the innate and adaptive humoral immune systems and modulate allergic responses in opposite directions, with sCD23 enhancing and sCD14 blunting inflammatory responses. We measured sCD23 and sCD14 in serum from blood that was drawn at a single time point from 1,079 glioma patients postdiagnosis and 736 healthy controls. Glioma was strongly associated with high sCD14 [highest versus lowest quartile odds ratio (OR), 3.94; 95% confidence interval (95% CI), 2.98-5.21] and low sCD23 (lowest versus highest quartile OR, 2.5; 95% CI, 1.89-3.23). Results were consistent across glioma histologic types and grades, but were strongest for glioblastoma. Whereas temozolomide treatment was not associated with either sCD14 or sCD23 levels among cases, those taking dexamethasone had somewhat lower sCD23 levels than those not taking dexamethasone. However, sCD23 was associated with case status regardless of dexamethasone treatment. These results augment the long-observed association between allergies and glioma and support a role for the innate and adaptive humoral functions of the immune system, in particular immunoregulatory proteins, in gliomagenesis.

  19. Soluble form of the endothelial adhesion molecule CD146 binds preferentially CD16+ monocytes.

    PubMed

    Garibaldi, Silvano; Barisione, Chiara; Ghigliotti, Giorgio; Spallarossa, Paolo; Barsotti, Antonio; Fabbi, Patrizia; Corsiglia, Luca; Palmieri, Daniela; Palombo, Domenico; Brunelli, Claudio

    2012-06-01

    The cell adhesion molecule CD146 is normally located at the endothelial cell-to-cell junction and colocalizes with actin cytoskeleton. The soluble form of CD146 (sCD146) has been identified in the endothelial cell supernatant and in normal human plasma, and is increased in pathologic conditions with altered endothelial function. Soluble CD146 binding to monocytes promotes their transendothelial migration, which represents a central step in the development of atherosclerotic plaque. Since peripheral blood monocytes are characterized by a phenotypic and functional heterogeneity, with different transendothelial migration capacity, we hypothesized that monocyte subsets differently bind sCD146. Based on surface CD14 and CD16 expression monocytes were distinguished by flow cytometry (FACS) into three subsets: CD14++/CD16-, CD14++/CD16+ and CD14+/CD16+. CD16+ monocytes have been found to possess higher transendothelial migration ability. FACS analysis on blood monocytes from 30 healthy subjects revealed that higher percentages of CD14++/CD16+ (median, first and third quartile: 2.26, 1.62-3.87) and of CD14+/CD16+ (2.59, 1.28-4.80) were positive for CD146 (both p < 0.01), in comparison to CD14++/CD16- (0.66, 0.47-1.01). Moreover, in vitro treatment of ficoll separated monocytes with recombinant CD146 showed that both CD16+ subsets increased their percentage of CD146-positive events compared to CD16- monocytes (p < 0.01). Soluble CD146 levels were evaluated by ELISA in plasma samples of subjects from our study group and showed a correlation with percentage of CD146-positive CD14+/CD16+ monocyte subset. In this work we have demonstrated that monocyte subsets behave differently with regard to their sCD146 binding activity; because binding of CD146 influences transendothelial migration of monocytes, modulation of monocyte-CD146 interaction may represent a potential target to limit atherosclerotic plaque development.

  20. A polymorphism in CD14 modifies the effect of farm milk consumption on allergic diseases and CD14 gene expression.

    PubMed

    Bieli, Christian; Eder, Waltraud; Frei, Remo; Braun-Fahrländer, Charlotte; Klimecki, Walt; Waser, Marco; Riedler, Josef; von Mutius, Erika; Scheynius, Annika; Pershagen, Göran; Doekes, Gert; Lauener, Roger; Martinez, Fernando D

    2007-12-01

    Consumption of farm milk in early life is associated with less asthma and allergies. We hypothesized that genetic variation in the innate immunity receptor CD14 might modify the association between farm milk consumption and asthma and atopy. Questionnaire data, serum IgE levels, and genotypes for 4 single nucleotide polymorphisms in CD14 were assessed in farmers' and nonfarmers' children from 2 European populations (Allergy and Endotoxin study, n = 576; Prevention of Allergy Risk factors for Sensitization in children related to Farming and Anthroposophic Lifestyle study, n = 1539). In a subsample (n = 222) CD14 gene expression was measured in peripheral blood leukocytes. The effects of farm milk and CD14 genotypes on asthma, allergies, and CD14 expression and their interactions were investigated. We found a significant interaction between genetic variation in CD14/-1721 and farm milk consumption. Adjusted odds ratios for the association between farm milk and asthma varied between the genotypes: AA, 0.18 (95% CI, 0.07-0.47); AG, 0.47 (95% CI, 0.26-0.86); and GG, 0.98 (95% CI, 0.46-2.08). Similar patterns were observed for symptoms of allergic rhinoconjunctivitis and pollen sensitization. CD14/-1721 also modified the association between farm milk and CD14 gene expression (adjusted geometric means ratios: AA, 1.61 (95% CI, 0.98-2.66); AG, 1.11 (95% CI, 0.71-1.72); and GG, 0.76 (95% CI, 0.39-1.48). The protective effect of farm milk consumption on allergic diseases is stronger in children carrying the A allele in CD14/-1721 than in children homozygous for the G allele. This might be mediated through farm milk-induced upregulated CD14 gene expression. Our results support the hypothesis that the inverse association between farm milk consumption and allergic diseases is mediated by CD14-activated innate immune mechanisms.

  1. Modulation of CD14 and TLR4.MD-2 activities by a synthetic lipid A mimetic

    PubMed Central

    Cighetti, Roberto; Ciaramelli, Carlotta; Sestito, Stefania Enza; Zanoni, Ivan; Kubik, Łukasz; Ardá-Freire, Ana; Calabrese, Valentina; Granucci, Francesca; Jerala, Roman; Martín-Santamaría, Sonsoles; Jiménez-Barbero, Jesus

    2014-01-01

    Monosaccharide lipid A mimetics composed by a glucosamine core linked to two fatty acid chains and bearing one or two phosphates have been synthesized. While compounds 1 and 2, with one phosphate group, were practically inactive in inhibiting LPS-induced TLR4 signaling and cytokine production in HEK-blue™ cells and murine macrophages, compound 3 with two phosphates was found to be active in efficiently inhibiting TLR4 signal in both cell types. The direct interaction of molecule 3 with MD-2 co-receptor has been investigated by means of NMR and molecular modeling/docking analysis. This compound also interacts directly with CD14 receptor, stimulating its internalization by endocytosis. Experiments on macrophages show that the effect on CD14 reinforces the activity on MD-2.TLR4, because compound 3 activity is higher when CD14 is important for TLR4 signaling i,e, at low LPS concentration. The dual MD-2 and CD14 targeting, accompanied by good solubility in water and lack of toxicity, suggests the use of monosaccharide 3 as a lead compound to develop drugs directed against TLR4-related syndromes. PMID:24339336

  2. Maternal Plasma Soluble TRAIL is Decreased in Preeclampsia

    PubMed Central

    Chaemsaithong, Piya; Chaiworapongsa, Tinnakorn; Romero, Roberto; Korzeniewski, Steven J.; Stampalija, Tamara; Than, Nandor Gabor; Dong, Zhong; Miranda, Jezid; Yeo, Lami; Hassan, Sonia S

    2014-01-01

    Objective Preeclampsia (PE) is characterized by systemic intravascular inflammation. Women who develop PE are at an increased risk for cardiovascular disease in later life. Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) has anti-atherosclerotic effects in endothelial cells and can mediate neutrophil apoptosis. Low soluble TRAIL (sTRAIL) and high C-reactive protein (CRP) concentrations are associated with an increased risk of future cardiovascular disease in non-pregnant individuals. The aim of this study was to determine whether maternal plasma concentrations of sTRAIL and CRP differ between women with PE and those with uncomplicated pregnancies. Methods This cross-sectional study included women with an uncomplicated pregnancy (n=93) and those with PE (n=52). Maternal plasma concentrations of sTRAIL and CRP concentrations were determined by ELISA. Results 1) The median plasma sTRAIL concentration (pg/mL) was significantly lower and the median plasma CRP concentration was significantly higher in women with PE than in those with an uncomplicated pregnancy (25.55 vs. 29.17; p = 0.03 and 8.0 vs. 4.1; p=0.001, respectively); 2) the median plasma concentration sTRAIL/CRP ratio was twofold lower in women with PE than in those with an uncomplicated pregnancy (p<0.001); and 3) women with plasma sTRAIL and CRP ratio in the lowest quartile were eight times more likely to have PE than women with concentrations in the upper three quartiles (OR 8.9; 95% CI: 2.8–27.8). Conclusion Maternal plasma sTRAIL concentrations are lower (while those of CRP are higher) in women with PE than in those with uncomplicated pregnancies. These findings are consistent with the evidence of intravascular inflammation in this disorder. PMID:23688319

  3. CD14 mediates binding of high doses of LPS but is dispensable for TNF-α production.

    PubMed

    Borzęcka, Kinga; Płóciennikowska, Agnieszka; Björkelund, Hanna; Sobota, Andrzej; Kwiatkowska, Katarzyna

    2013-01-01

    Activation of macrophages with lipopolysaccharide (LPS) involves a sequential engagement of serum LPS-binding protein (LBP), plasma membrane CD14, and TLR4/MD-2 signaling complex. We analyzed participation of CD14 in TNF-α production stimulated with 1-1000 ng/mL of smooth or rough LPS (sLPS or rLPS) and in sLPS binding to RAW264 and J744 cells. CD14 was indispensable for TNF-α generation induced by a low concentration, 1 ng/mL, of sLPS and rLPS. At higher doses of both LPS forms (100-1000 ng/mL), TNF-α release required CD14 to much lower extent. Among the two forms of LPS, rLPS-induced TNF-α production was less CD14-dependent and could proceed in the absence of serum as an LBP source. On the other hand, the involvement of CD14 was crucial for the binding of 1000 ng/mL of sLPS judging from an inhibitory effect of the anti-CD14 antibody. The binding of sLPS was also strongly inhibited by dextran sulfate, a competitive ligand of scavenger receptors (SR). In the presence of dextran sulfate, sLPS-induced production of TNF-α was upregulated about 1.6-fold. The data indicate that CD14 together with SR participates in the binding of high doses of sLPS. However, CD14 contribution to TNF α production induced by high concentrations of sLPS and rLPS can be limited.

  4. NMR spectral mapping of Lipid A molecular patterns affected by interaction with the innate immune receptor CD14

    SciTech Connect

    Albright, Seth; Agrawal, Prashansa; Jain, Nitin U.

    2009-01-23

    Soluble CD14 (sCD14) is a serum glycoprotein that binds to the Lipid A moiety of lipopolysaccharides (LPS) with high affinity as part of the innate immune response to bacterial endotoxins. In order to investigate structural interactions of Lipid A with sCD14, we have prepared an isotopically labeled form of a fully active and chemically defined endotoxin, Kdo{sub 2}-Lipid A, which allowed us to carry out detailed NMR spectral mapping of this agonist ligand bound to sCD14 and identify for the first time structural regions that are strongly affected during complex formation with sCD14. These map to two adjacent areas comprising the lower portions of the sugar headgroup and upper half of the acyl chains I, III, and V, which are spatially proximal to the 1- and 4'-phosphate ends. Additionally, we have detected for the first time, presence of differential dynamic behavior for the affected resonances, suggesting a likely role for dynamics in the mechanism of Lipid A pattern recognition by sCD14.

  5. Cytokine regulation of the myeloid glycoprotein CD14.

    PubMed

    Landmann, R; Wesp, M; Obrecht, J P

    1991-01-01

    Monocyte membrane CD14 (mCD14) antigen expression was measured in normal human monocytes and blood monocyte-derived macrophages. Seven-day culture of monocytes in serum-containing medium lead to an increase in mCD14. Addition of interferon-gamma (IFN gamma) or interleukin-4 (IL-4) to monocytes caused a dose-dependent reduction in mCD14 within 3 or 45 h respectively. These effects were strong in monocytes, weak in macrophages, and they were blocked by anti-IFN gamma and anti-IL-4 antibodies, respectively. Interleukin-2 and interferon-alpha produced a decrease in mCD14 in mononuclear leukocyte cultures but not in purified monocytes. Their effect was abolished in the presence of anti-IFN gamma. Other cytokines (TFN alpha, Ill beta, IL-6, IL-3, IL-5, GM-CSF, TGF beta 1) did not change mCD14. In conclusion, IFN gamma and IL-4 are revealed to be the only cytokines which directly affect monocyte CD14 expression.

  6. Lack of effect of the CD14 promoter gene C-159T polymorphism on nutritional status parameters in hemodialysis patients.

    PubMed

    Serwacka, Agnieszka; Protaziuk, Tomasz; Zagozda, Małgorzata; Popow, Anna-Maria; Kierzkiewicz, Maciej; Manitius, Jacek; Myśliwiec, Michał; Daniewska, Dorota; Gołebiewski, Sewer; Rydzewska-Rosołowska, Alicja; Flisiński, Mariusz; Stępień, Krystyna; Rydzewska, Grażyna; Olszewski, Waldemar L; Rydzewski, Andrzej

    2011-02-01

    CD14 is a membrane glycoprotein that acts as a co-receptor for the detection of bacterial lipopolysaccharide (LPS). Mutual interaction between CD14 and LPS plays an important role in the innate immune system. Increased serum soluble CD14 levels have been described in hemodialysis (HD) patients, and linked to increased mortality risk, inflammation and protein-energy wasting. The expression of CD14 may be influenced by CD14 promoter gene C-159T polymorphism. This study aimed to clarify the possible association between CD14 promoter gene C-159T polymorphism and nutritional status in hemodialysis patients. The study population consisted of 185 (104 males; 81 females) long-term HD patients treated in 5 dialysis centers. The control group consisted of 112 apparently healthy volunteers (32 males and 80 females). Nutritional status was assessed using a modified SGA scale, and anthropometric methods (BMI, WHR, waist, hip and mid-arm circumferences, biceps, triceps, subocular and subscapular skinfolds). Biochemical parameters evaluated included: CRP, albumin, creatinine, urea, cholesterol, triglycerides and TIBC. CD14 promoter gene C-159T polymorphism was determined by restriction fragment length polymorphism, after digestion of the PCR product with Hae III restriction endonuclease. Genotype and allele frequencies were similar to controls and compliant with Hardy-Weinberg equilibrium. No between-group differences were detected in measured variables with the exception of lower triglyceride levels in carriers of C allele in comparison to TT genotype. CD14 promoter gene C-159T polymorphism does not seem to be associated with nutritional status parameters in HD patients. It does seem, however, to influence triglyceride blood levels.

  7. Interaction between ambient pollutant exposure, CD14 (-159) polymorphism and respiratory outcomes among children in Kwazulu-Natal, Durban.

    PubMed

    Makamure, M T; Reddy, P; Chuturgoon, A; Naidoo, R N; Mentz, G; Batterman, S; Robins, T G

    2017-03-01

    The objective of this study was to determine if the association between exposure to ambient air pollutants such as sulfur dioxide, nitrogen dioxde (NO2), nitrous oxide (NO), and PM10, and variation in lung function measures was modified by genotype. A validated questionnaire was administered to 71 African children to evaluate prevalence of respiratory symptoms. Atopy was evaluated by skin-prick testing and bihourly measures of lung function (spirometry) were collected. Gaseous air pollutant concentrations were monitored continuously. CD14 polymorphism was genotyped and plasma CD14 levels were measured. There was no statistically significant association between the CD14 (159) CT+TT polymorphism with any asthma-related phenotype. There was a significant association between lung function (forced expiratory volume in 1 second intraday variability) and NO2 and NO among participants carrying the CD14 CT/TT genotype for lags 1, 2, and the 5-day average. Similarly, statistically significant gene-pollutant interactions ( p < 0.05) were found with NO and CD14 CT/TT at lag 2 and for the 5-day average. While there was no association with any respiratory phenotype (as determined by symptoms), the CD14 CT/TT genotype appeared to be protective to increased exposure to NO2 and NO.

  8. CD14{sup +} monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    SciTech Connect

    Wang, Ding; Chen, Ke; Du, Wei Ting; Han, Zhi-Bo; Ren, He; Chi, Ying; and others

    2010-09-10

    Here, the effect of CD14{sup +} monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-{gamma} (IFN-{gamma}) secretion capacities of CD4{sup +} and CD8{sup +} T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E{sub 2} (PGE{sub 2}) as an important soluble mediator. CD14{sup +} monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1{beta}, either exogenously added or produced by CD14{sup +} monocytes in culture, could trigger expression of high levels of PGE{sub 2} by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE{sub 2} expression, but also reversed the promotional effect of CD14{sup +} monocytes and partially restored CD4{sup +} and CD8{sup +} T cell proliferation and IFN-{gamma} secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  9. Circulating CD14+ monocytes in patients with aortic stenosis

    PubMed Central

    Shimoni, Sara; Meledin, Valery; Bar, Iris; Fabricant, Jacob; Gandelman, Gera; George, Jacob

    2016-01-01

    Background Calcific aortic stenosis (AS) is an active process sharing similarities with atherosclerosis and chronic inflammation. The pathophysiology of AS is notable for three cardinal components: inflammation, fibrosis and calcification. Monocytes play a role in each of these processes. The role of circulating monocytes in AS is not clear. The aim of the present study was to study an association between circulating apoptotic and non apoptotic CD14+ monocytes and AS features. Methods We assessed the number of CD14+ monocytes and apoptotic monocytes in 54 patients with significant AS (aortic valve area 0.74 ± 0.27 cm2) and compared them to 33 patients with similar risk factors and no valvular disease. The level of CD14+ monocytes and apoptotic monocytes was assessed by flow cytometry. Results There was no difference in the risk factor profile and known coronary or peripheral vascular diseases between patients with AS and controls. Patients with AS exhibited increased numbers of CD14+ monocytes as compared to controls (9.9% ± 4.9% vs. 7.7% ± 3.9%, P = 0.03). CD14+ monocyte number was related to age and the presence and severity of AS. In patients with AS, both CD14+ monocytes and apoptotic monocytes were inversely related to aortic valve area. Conclusions Patients with significant AS have increased number of circulating CD14+ monocytes and there is an inverse correlation between monocyte count and aortic valve area. These findings may suggest that inflammation is operative not only in early valve injury phase, but also at later developed stages such as calcification when AS is severe. PMID:26918018

  10. Close localization of mouse CD14 and CD32/16 in the cell surface of monocytic cell lines.

    PubMed

    Hisaka, H; Kataoka, M; Higuchi, Y; Matsuura, K; Yamamoto, S

    1999-01-01

    The binding of a rat anti-mouse CD14 monoclonal antibody (mAb) (rmC5-3) was inhibited by pretreatment of a mouse monocytic cell line WEHI-3 cells with anti-mouse CD32/16 mAb (2.4G2), whereas that of 2.4G2 was not inhibited by pretreatment of WEHI-3 cells with rmC5-3. An enzyme-linked immunosorbent assay showed that rmC5-3 detected peptide 9 corresponding to amino acid position 308-322 of CD14 but 2.4G2 did not. A Western blot analysis of sera revealed that rmC5-3 and 2.4G2 detected the bands thought to be soluble CD14 and CD32/16, respectively. rmC5-3 reacted with mouse CD14-transfected CHO cells, CD14-CHO-K1 cells, but 2.4G2 did not. Lipopolysaccharide-induced tumor necrosis factor (TNF)-alpha release was enhanced when a monocyte cell line (J774) was pretreated with rmC5-3. The enhancement was abolished by pretreatment with 2.4G2. The release of TNF-alpha was observed following treatment of J774 cells with 2.4G2 followed by anti-rat IgG F(ab')2.

  11. Decreased plasma levels of soluble CD18 link leukocyte infiltration with disease activity in spondyloarthritis

    PubMed Central

    2014-01-01

    Introduction Spondyloarthritis (SpA) comprises a group of diseases often associated with HLA-B27 and characterized by inflammation of the entheses and joints of the axial skeleton. The inflammatory process in SpA is presumably driven by innate immune cells but is still poorly understood. Thus, new tools for monitoring and treating inflammation are needed. The family of CD18 integrins is pivotal in guiding leukocytes to sites of inflammation, and CD18 hypomorphic mice develop a disease resembling SpA. Previously, we demonstrated that altered soluble CD18 (sCD18) complexes in the blood and synovial fluid of patients with arthritis have anti-inflammatory functions. Here, we study the mechanisms for these alterations and their association with SpA disease activity. Methods Plasma levels of sCD18 in a study population with 84 patients with SpA and matched healthy controls were analyzed with a time-resolved immunoflourometric assay (TRIFMA). Binding of sCD18 to endothelial cells and fibroblast-like synoviocytes (FLSs) was studied with confocal microscopy. Shedding of CD18 from peripheral blood mononuclear cells (PBMCs) was studied with flow cytometry and TRIFMA. Results Plasma levels of sCD18 were decreased in patients with SpA compared with healthy volunteers (P <0.001), and the lowest levels were in the HLA-B27-positive subgroup (P <0.05). In a multiple regression model, the sCD18 levels exhibited an inverse correlation with the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) (P <0.05), the level of morning stiffness (P <0.05), the Bath Ankylosing Spondilitis Metrology Index (P <0.05), the physician global assessment score (P <0.01), and the sacroiliac magnetic resonance imaging activity score (P <0.05). The mechanisms for these changes could be simulated in vitro. First, sCD18 in plasma adhered to inflammation-induced intercellular adhesion molecule 1 (ICAM-1) on endothelial cells and FLS, indicating increased consumption. Second, CD18 shedding from Sp

  12. Critical Role of CD14 for Production of Proinflammatory Cytokines and Cytokine Inhibitors during Sepsis with Failure To Alter Morbidity or Mortality

    PubMed Central

    Ebong, Samuel J.; Goyert, Sanna M.; Nemzek, Jean A.; Kim, Jiyoun; Bolgos, Gerald L.; Remick, Daniel G.

    2001-01-01

    We investigated the immunopathophysiologic responses during sepsis induced by cecal ligation and puncture (CLP) in CD4-deficient (CD14 knockout [CD14KO]) mice. Our studies were designed to specifically test the role of CD14 in the inflammatory response to sepsis and to ascertain if alterations would improve morbidity or mortality. Sepsis was induced using the CLP model with appropriate antibiotic treatment. The severity of sepsis increased in the CD14KO mice with increasing puncture size (18 gauge [18G], 21G, and 25G). Following CLP, body temperature (at 12 h) and gross motor activity levels of the sham and 25G CLP groups recovered to normal, while the 21G and 18G CLP groups exhibited severe hypothermia coupled with decreased gross motor activity and body weight. There were no significant differences in survival, temperature, body weight, or activity levels between CD14KO and control mice after 21G CLP. However, CD14KO mice expressed two- to fourfold less pro-inflammatory (interleukin-1β [IL-1β], tumor necrosis factor [TNF], and IL-6) and anti-inflammatory (IL-10, IL-1 receptor antagonist, and TNF receptors I and II) cytokines in the blood after 21G CLP. Plasma levels of the chemokines macrophage inflammatory protein 2α and KC were similarly reduced in CD14KO mice. A similar trend of decreased cytokine and cytokine inhibitor levels was observed in the peritoneal cavity of CD14KO mice. Our results indicate that the CD14 pathway of activation plays a critical role in the production of both pro-inflammatory cytokines and cytokine inhibitors but has minimal impact on the morbidity or mortality induced by the CLP model of sepsis. PMID:11254563

  13. Genetic polymorphisms in the CD14 gene are associated with monocyte activation and carotid intima-media thickness in HIV-infected patients on antiretroviral therapy.

    PubMed

    Yong, Yean K; Shankar, Esaki M; Westhorpe, Clare L V; Maisa, Anna; Spelman, Tim; Kamarulzaman, Adeeba; Crowe, Suzanne M; Lewin, Sharon R

    2016-08-01

    HIV-infected individuals on antiretroviral therapy (ART) are at increased risk of cardiovascular disease (CVD). Given the relationship between innate immune activation and CVD, we investigated the association of single-nucleotide polymorphisms (SNPs) in TLR4 and CD14 and carotid intima-media thickness (cIMT), a surrogate measurement for CVD, in HIV-infected individuals on ART and HIV-uninfected controls as a cross-sectional, case-control study. We quantified the frequency of monocyte subsets (CD14, CD16), markers of monocyte activation (CD38, HLA-DR), and endothelial adhesion (CCR2, CX3CR1, CD11b) by flow cytometry. Plasma levels of lipopolysaccharide, sCD163, sCD14, sCX3CL1, and sCCL2, were measured by ELISA. Genotyping of TLR4 and CD14 SNPs was also performed. The TT genotype for CD14/-260SNP but not the CC/CT genotype was associated with elevated plasma sCD14, and increased frequency of CD11b+CD14+ monocytes in HIV-infected individuals. The TT genotype was associated with lower cIMT in HIV-infected patients (n = 47) but not in HIV-uninfected controls (n = 37). The AG genotype for TLR4/+896 was associated with increased CX3CR1 expression on total monocytes among HIV-infected individuals and increased sCCL2 and fibrinogen levels in HIV-uninfected controls. SNPs in CD14/-260 and TLR4/+896 were significantly associated with different markers of systemic and monocyte activation and cIMT that differed between HIV-infected participants on ART and HIV-uninfected controls. Further investigation on the relationship of these SNPs with a clinical endpoint of CVD is warranted in HIV-infected patients on ART.

  14. Genetic polymorphisms in the CD14 gene are associated with monocyte activation and carotid intima-media thickness in HIV-infected patients on antiretroviral therapy

    PubMed Central

    Yong, Yean K.; Shankar, Esaki M.; Westhorpe, Clare L.V.; Maisa, Anna; Spelman, Tim; Kamarulzaman, Adeeba; Crowe, Suzanne M.; Lewin, Sharon R.

    2016-01-01

    Abstract HIV-infected individuals on antiretroviral therapy (ART) are at increased risk of cardiovascular disease (CVD). Given the relationship between innate immune activation and CVD, we investigated the association of single-nucleotide polymorphisms (SNPs) in TLR4 and CD14 and carotid intima-media thickness (cIMT), a surrogate measurement for CVD, in HIV-infected individuals on ART and HIV-uninfected controls as a cross-sectional, case-control study. We quantified the frequency of monocyte subsets (CD14, CD16), markers of monocyte activation (CD38, HLA-DR), and endothelial adhesion (CCR2, CX3CR1, CD11b) by flow cytometry. Plasma levels of lipopolysaccharide, sCD163, sCD14, sCX3CL1, and sCCL2, were measured by ELISA. Genotyping of TLR4 and CD14 SNPs was also performed. The TT genotype for CD14/−260SNP but not the CC/CT genotype was associated with elevated plasma sCD14, and increased frequency of CD11b+CD14+ monocytes in HIV-infected individuals. The TT genotype was associated with lower cIMT in HIV-infected patients (n = 47) but not in HIV-uninfected controls (n = 37). The AG genotype for TLR4/+896 was associated with increased CX3CR1 expression on total monocytes among HIV-infected individuals and increased sCCL2 and fibrinogen levels in HIV-uninfected controls. SNPs in CD14/−260 and TLR4/+896 were significantly associated with different markers of systemic and monocyte activation and cIMT that differed between HIV-infected participants on ART and HIV-uninfected controls. Further investigation on the relationship of these SNPs with a clinical endpoint of CVD is warranted in HIV-infected patients on ART. PMID:27495090

  15. Presepsin (sCD14-ST) Is a Novel Marker for Risk Stratification in Cardiac Surgery Patients.

    PubMed

    Bomberg, Hagen; Klingele, Matthias; Wagenpfeil, Stefan; Spanuth, Eberhard; Volk, Thomas; Sessler, Daniel I; Schäfers, Hans-Joachim; Groesdonk, Heinrich Volker

    2017-04-01

    Presepsin (soluble cluster-of-differentiation 14 subtype [sCD14-ST]) is a humoral risk stratification marker for systemic inflammatory response syndrome and sepsis. It remains unknown whether presepsin can be used to stratify risk in elective cardiac surgery. The authors therefore determined the usefulness of presepsin for risk stratification in patients having elective cardiac surgery. Eight hundred fifty-six cardiac surgical patients were prospectively studied. Preoperative plasma concentrations of presepsin, procalcitonin, N-terminal pro-hormone natriuretic peptide, cystatin C, and the additive European System of Cardiac Operative Risk Evaluation 2 were compared to mortality at 30 days (primary outcome), 6 months, and 2 yr. Discrimination was assessed with C statistic. Logistic regression analysis was used to calculate univariable and multivariable odds ratios. Thirty-day mortality was 3.2%, 6-month mortality was 6.1%, and 2-yr mortality was 10.4% across the population. Median preoperative presepsin concentrations were significantly greater in 30-day nonsurvivors than in survivors: 842 pg/ml (interquartile range, 306 to 1,246) versus 160 pg/ml (interquartile range, 122 to 234); difference, 167 pg/ml (interquartile range, 92 to 301; P < 0.001). The results were similar for 6-month and 2-yr mortality. Compared to the European System of Cardiac Operative Risk Evaluation 2, presepsin concentration provided better discrimination for postoperative mortality at all follow-up periods, including 30 days (C statistic 0.88 vs. 0.74), 6 months (0.87 vs. 0.76), and 2 yr (0.81 vs. 0.74). Presepsin also provided better discrimination than cystatin C, N-terminal pro-hormone natriuretic peptide, or procalcitonin. Elevated presepsin remained an independent risk predictor after adjustment for potential confounding factors. Elevated preoperative plasma presepsin concentration is an independent predictor of postoperative mortality in elective cardiac surgery patients and is a

  16. CD14 mediated endogenous TNF-alpha release in HL60 AML cells: a potential model for CD14 mediated endogenous cytokine release in the treatment of AML.

    PubMed

    Treon, S P; Anand, B; Ulevitch, R; Broitman, S A

    1994-01-01

    In previous studies, HL60 AML cells treated with tumor necrosis factor-alpha (TNF), interferon-gamma (IFN), and lipopolysaccharides (LPS) displayed decreased growth and viability, enhanced monocytic pathway differentiation and endogenous TNF release. Endogenous TNF release by LPS/TNF/IFN treated HL60 cells was postulated to play a role with the above findings. In these studies, HL60 cells expressed CD14 when treated with TNF, IFN, and LPS. CD14 mediates TNF release in monocytes/macrophages in response to binding of LPS with LPS binding protein (LBP). CD14 was not expressed in either untreated or LPS only treated HL60 cells. CD14 expression was present and greater with HL60 cells cultured with LPS/TNF/IFN vs TNF/IFN (47.47% vs 9.07% positive, respectively) suggesting synergism for LPS in CD14 induction. CD14 expression was associated with endogenous TNF release, and with significantly higher levels by HL60 cells treated with LPS/TNF/IFN vs TNF/IFN (p < 0.001). Addition of anti-CD14 antibody significantly reduced release of TNF in TNF/IFN (p < 0.001) and LPS/TNF/IFN (p = 0.0013) treated cells. KG1 and U937 AML cells treated with LPS, TNF, and IFN did not express CD14, nor release TNF. A model for inducing release of endogenous growth inhibitory cytokines by CD14 bearing AML cells is proposed as an approach to AML therapy.

  17. [Establishment of murine cell line transfected with human CD14 gene].

    PubMed

    Ning, Bo-Tao; Tang, Yong-Min; Xu, Yan; Chen, Yan-Fei; Cao, Jiang

    2006-04-01

    This study was aimed to construct the CD14 eukaryotic expression vector, establish the transgeneic CD14 positive cell line in order to facilitate the establishment of a mouse model of antibody targeting therapy for human acute monocytic leukemia (AML-M(5)). Total RNA extracted from peripheral blood mononuclear cells was treated with RNAase-free DNAase, the human CD14 gene was cloned and sequenced through the RT-PCR and T-A clone techniques. Eukaryotic expressional vector pcDNA3.1(+)/CD14 was constructed by cleaving with double restriction endonuleases and ligating with T4 ligase. A murine melanoma cell line B16 was transfected with the pcDNA3.1(+)/CD14 recombinant with Superfect transfection reagent. Positive clones were selected by G418 and the expression of human CD14 on the transfectant was confirmed by flow cytometry (FCM). The results indicated that the sequence of the human CD14 cDNA cloned was exact to be same as the one from GenBank database. The recombinant pcDNA3.1(+)/CD14 was identified with double-enzyme cleaving. The expression of the human CD14 on the transfectant (B16/CD14) was confirmed by FCM. In conclusion, the murine cell line B16/CD14 fransfected with human CD14 gene has been established which can be used for the study of human AML-M(5) antibody targeting therapy with mouse model.

  18. CD14 Is a Co-Receptor for TLR4 in the S100A9-Induced Pro-Inflammatory Response in Monocytes

    PubMed Central

    He, Zhifei; Riva, Matteo; Björk, Per; Swärd, Karl; Mörgelin, Matthias; Leanderson, Tomas; Ivars, Fredrik

    2016-01-01

    The cytosolic Ca2+-binding S100A9 and S100A8 proteins form heterodimers that are primarily expressed in human neutrophils and monocytes. We have recently shown that S100A9 binds to TLR4 in vitro and induces TLR4-dependent NF-κB activation and a pro-inflammatory cytokine response in monocytes. In the present report we have further investigated the S100A9-mediated stimulation of TLR4 in monocytes. Using transmission immunoelectron microscopy, we detected focal binding of S100A9 to monocyte membrane subdomains containing the caveolin-1 protein and TLR4. Furthermore, the S100A9 protein was detected in early endosomes of the stimulated cells, indicating that the protein could be internalized by endocytosis. Although stimulation of monocytes with S100A9 was strictly TLR4-dependent, binding of S100A9 to the plasma membrane and endocytosis of S100A9 was still detectable and coincided with CD14 expression in TLR4-deficient cells. We therefore investigated whether CD14 would be involved in the TLR4-dependent stimulation and could show that the S100A9-induced cytokine response was inhibited both in CD14-deficient cells and in cells exposed to CD14 blocking antibodies. Further, S100A9 was not internalized into CD14-deficient cells suggesting a direct role of CD14 in endocytosis of S100A9. Finally, we could detect satiable binding of S100A9 to CD14 in surface plasmon resonance experiments. Taken together, these results indicate that CD14 is a co-receptor of TLR4 in the S100A9-induced cytokine response. PMID:27228163

  19. Phagocytosis of gram-negative bacteria by a unique CD14-dependent mechanism.

    PubMed

    Schiff, D E; Kline, L; Soldau, K; Lee, J D; Pugin, J; Tobias, P S; Ulevitch, R J

    1997-12-01

    THP-1-derived cell lines were stably transfected with constructs encoding glycophosphatidylinositol (GPI)-anchored or transmembrane forms of human CD14. CD14 expression was associated with enhanced phagocytosis of serum (heat-inactivated)-opsonized Escherichia coli (opEc). Both the GPI-anchored and transmembrane forms of CD14 supported phagocytosis of opEc equally well. Lipopolysaccharide-binding protein (LBP) played a role in CD14-dependent phagocytosis as evidenced by inhibition of CD14-dependent phagocytosis of opEc with anti-LBP monoclonal antibody (mAb) and by enhanced phagocytosis of E. coli opsonized with purified LBP. CD14-dependent phagocytosis was inhibited by a phosphatidylinositol (PI) 3-kinase inhibitor (wortmannin) and a protein tyrosine kinase inhibitor (tyrphostin 23) but not a protein kinase C inhibitor (bisindolyl-maleimide) or a divalent cation chelator (ethylenediaminetetraacetate). Anti-LBP mAb 18G4 and anti-CD14 mAb 18E12 were used to differentiate between the pathways involved in CD14-dependent phagocytosis and CD14-dependent cell activation. F(ab')2 fragments of 18G4, a mAb to LBP that does not block cell activation, inhibited ingestion of opEc by THP1-wtCD14 cells. 18E12 (an anti-CD14 mAb that does not block LPS binding to CD14 but does inhibit CD14-dependent cell activation) did not inhibit phagocytosis of LBP-opEc by THP1-wtCD14 cells. Furthermore, CD14-dependent phagocytosis was not inhibited by anti-CD18 (CR3 and CR4 beta-chain) or anti-Fcgamma receptor mAb.

  20. Soluble Proteins Form Film by the Treatment of Low Temperature Plasma

    NASA Astrophysics Data System (ADS)

    Ikehara, Sanae; Sakakita, Hajime; Ishikawa, Kenji; Akimoto, Yoshihiro; Nakanishi, Hayao; Shimizu, Nobuyuki; Hori, Masaru; Ikehara, Yuzuru

    2015-09-01

    It has been pointed out that low temperature plasma in atmosphere was feasible to use for hemostasis without heat injury. Indeed, earlier studies demonstrated that low temperature plasma played an important role to stimulate platelets to aggregate and turned on the proteolytic activities of coagulation factors, resulting in the acceleration of the natural blood coagulation process. On the other hands, our developed equips could immediately form clots upon the contact with plasma flair, while the histological appearance was different from natural coagulation. Based on these findings in formed clots, we sought to determine if plasma flair supplied by our devices was capable of forming film using a series of soluble proteins Following plasma treatment, films were formed from bovine serum albumin, and the other plasma proteins at physiological concentration. Analysis of trans-electron microscope demonstrated that plasma treatment generated small protein particles and made them fuse to be larger aggregations The combined results demonstrated that plasma are capable of aggregating soluble proteins and that platelets and coagulation factors are not necessary for plasma induced blood coagulation. Supported in part by Grants-in-Aid for Scientific Research on Priority Area (21590454, 24590498, and 24108006 to Y. I.).

  1. A New Simulated Plasma for Assessing the Solubility of Mineral Trioxide Aggregate

    PubMed Central

    Samiei, Mohammad; Shahi, Shahriar; Aslaminabadi, Naser; Valizadeh, Hadi; Aghazadeh, Zahra; Pakdel, Seyyed Mahdi Vahid

    2015-01-01

    Introduction: Solubility of mineral trioxide aggregate (MTA) is an important characteristic that affects other properties such as microleakage and biocompatibility. Distilled water (DW) has previously been used for solubility tests. This experimental study compared the solubility of MTA in DW, synthetic tissue fluid (STF) and new simulated plasma (SP). Methods and Materials: In this study, 36 samples of tooth-colored ProRoot MTA were prepared and divided into three groups (n=12) to be immersed in three different solutions (DW, STF, and SP). Solubility tests were conducted at 2, 5, 9, 14, 21, 30, 50, and 78-day intervals. The unequal variance F-test (Welch test) was utilized to determine the effect of solubility media and Games-Howell analysis was used for pairwise comparisons. The repeated-measures ANOVA was used to assess the importance of immersion duration. Results: Welch test showed significant differences in solubility rates of samples between all the different solubility media at all the study intervals (P<0.05) except for the 14-day interval (P=0.094). The mixed repeated-measures ANOVA revealed a significant difference in solubility rate of MTA in three different solutions at all time-intervals (P=0.000). Games-Howell post-hoc test revealed that all pairwise comparisons were statistically significant at all time-intervals (P=0.000). Conclusion: Based on the findings of this study, the long-term solubility of MTA in simulated plasma was less than that in synthetic tissue fluid and distilled water. PMID:25598806

  2. Plasma Soluble Prion Protein, a Potential Biomarker for Sport-Related Concussions: A Pilot Study

    PubMed Central

    Pham, Nam; Akonasu, Hungbo; Shishkin, Rhonda; Taghibiglou, Changiz

    2015-01-01

    Sport-related mild traumatic brain injury (mTBI) or concussion is a significant health concern to athletes with potential long-term consequences. The diagnosis of sport concussion and return to sport decision making is one of the greatest challenges facing health care clinicians working in sports. Blood biomarkers have recently demonstrated their potential in assisting the detection of brain injury particularly, in those cases with no obvious physical injury. We have recently discovered plasma soluble cellular prion protein (PrPC) as a potential reliable biomarker for blast induced TBI (bTBI) in a rodent animal model. In order to explore the application of this novel TBI biomarker to sport-related concussion, we conducted a pilot study at the University of Saskatchewan (U of S) by recruiting athlete and non-athlete 18 to 30 year-old students. Using a modified quantitative ELISA method, we first established normal values for the plasma soluble PrPC in male and female students. The measured plasma soluble PrPC in confirmed concussion cases demonstrated a significant elevation of this analyte in post-concussion samples. Data collected from our pilot study indicates that the plasma soluble PrPC is a potential biomarker for sport-related concussion, which may be further developed into a clinical diagnostic tool to assist clinicians in the assessment of sport concussion and return-to-play decision making. PMID:25643046

  3. Plasma soluble prion protein, a potential biomarker for sport-related concussions: a pilot study.

    PubMed

    Pham, Nam; Akonasu, Hungbo; Shishkin, Rhonda; Taghibiglou, Changiz

    2015-01-01

    Sport-related mild traumatic brain injury (mTBI) or concussion is a significant health concern to athletes with potential long-term consequences. The diagnosis of sport concussion and return to sport decision making is one of the greatest challenges facing health care clinicians working in sports. Blood biomarkers have recently demonstrated their potential in assisting the detection of brain injury particularly, in those cases with no obvious physical injury. We have recently discovered plasma soluble cellular prion protein (PrP(C)) as a potential reliable biomarker for blast induced TBI (bTBI) in a rodent animal model. In order to explore the application of this novel TBI biomarker to sport-related concussion, we conducted a pilot study at the University of Saskatchewan (U of S) by recruiting athlete and non-athlete 18 to 30 year-old students. Using a modified quantitative ELISA method, we first established normal values for the plasma soluble PrP(C) in male and female students. The measured plasma soluble PrP(C) in confirmed concussion cases demonstrated a significant elevation of this analyte in post-concussion samples. Data collected from our pilot study indicates that the plasma soluble PrP(C) is a potential biomarker for sport-related concussion, which may be further developed into a clinical diagnostic tool to assist clinicians in the assessment of sport concussion and return-to-play decision making.

  4. Characterization of the CD14++CD16+ monocyte population in human bone marrow.

    PubMed

    Mandl, Manuela; Schmitz, Susanne; Weber, Christian; Hristov, Michael

    2014-01-01

    Numerous studies have divided blood monocytes according to their expression of the surface markers CD14 and CD16 into following subsets: classical CD14(++)CD16(-), intermediate CD14(++)CD16(+) and nonclassical CD14(+)CD16(++) monocytes. These subsets differ in phenotype and function and are further correlated to cardiovascular disease, inflammation and cancer. However, the CD14/CD16 nature of resident monocytes in human bone marrow remains largely unknown. In the present study, we identified a major population of CD14(++)CD16(+) monocytes by using cryopreserved bone marrow mononuclear cells from healthy donors. These cells express essential monocyte-related antigens and chemokine receptors such as CD11a, CD18, CD44, HLA-DR, Ccr2, Ccr5, Cx3cr1, Cxcr2 and Cxcr4. Notably, the expression of Ccr2 was inducible during culture. Furthermore, sorted CD14(++)CD16(+) bone marrow cells show typical macrophage morphology, phagocytic activity, angiogenic features and generation of intracellular oxygen species. Side-by-side comparison of the chemokine receptor profile with unpaired blood samples also demonstrated that these rather premature medullar monocytes mainly match the phenotype of intermediate and partially of (non)classical monocytes. Together, human monocytes obviously acquire their definitive CD14/CD16 signature in the bloodstream and the medullar monocytes probably transform into CD14(++)CD16- and CD14(+)CD16(++) subsets which appear enriched in the periphery.

  5. SuperSAGE evidence for CD14++CD16+ monocytes as a third monocyte subset.

    PubMed

    Zawada, Adam M; Rogacev, Kyrill S; Rotter, Björn; Winter, Peter; Marell, Rolf-R; Fliser, Danilo; Heine, Gunnar H

    2011-09-22

    Monocytes are a heterogeneous cell population with subset-specific functions and phenotypes. The differential expression of CD14 and CD16 distinguishes classical CD14(++)CD16(-), intermediate CD14(++)CD16(+), and nonclassical CD14(+)CD16(++) monocytes. Current knowledge on human monocyte heterogeneity is still incomplete: while it is increasingly acknowledged that CD14(++)CD16(+) monocytes are of outstanding significance in 2 global health issues, namely HIV-1 infection and atherosclerosis, CD14(++)CD16(+) monocytes remain the most poorly characterized subset so far. We therefore developed a method to purify the 3 monocyte subsets from human blood and analyzed their transcriptomes using SuperSAGE in combination with high-throughput sequencing. Analysis of 5 487 603 tags revealed unique identifiers of CD14(++)CD16(+) monocytes, delineating these cells from the 2 other monocyte subsets. Gene Ontology (GO) enrichment analysis suggests diverse immunologic functions, linking CD14(++)CD16(+) monocytes to Ag processing and presentation (eg, CD74, HLA-DR, IFI30, CTSB), to inflammation and monocyte activation (eg, TGFB1, AIF1, PTPN6), and to angiogenesis (eg, TIE2, CD105). In conclusion, we provide genetic evidence for a distinct role of CD14(++)CD16(+) monocytes in human immunity. After CD14(++)CD16(+) monocytes have earlier been discussed as a potential therapeutic target in inflammatory diseases, we are hopeful that our data will spur further research in the field of monocyte heterogeneity.

  6. Lipopolysaccharide modulation of a CD14-like molecule on porcine alveolar macrophages

    NASA Technical Reports Server (NTRS)

    Kielian, T. L.; Ross, C. R.; McVey, D. S.; Chapes, S. K.; Blecha, F.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    Cluster of differentiation antigen 14 (CD14) functions as a receptor for lipopolysaccharide (LPS) LPS-binding protein (LBP) complexes. Because LPS has varying effects on CD14 expression in vitro, we evaluated CD14 expression in response to LPS with a fully differentiated macrophage phenotype, the alveolar macrophage. By using flow microfluorometric analysis and a radioimmunoassay with an anti-human CD14 monoclonal antibody (My4) that cross-reacts with porcine CD14, we found that macrophages stimulated with LPS for 24 h exhibited a two- to fivefold increase in CD14-like antigen compared with unstimulated cells. At low concentrations of LPS, up-regulation of the CD14-like antigen was dependent on serum; at higher concentrations of LPS, serum was not required. In the absence of serum a 10-fold higher dose of LPS (10 ng/ml) was required to increase CD14-like expression. In addition, LPS-induced CD14-like up-regulation correlated with secretion of tumor necrosis factor-alpha, regardless of serum concentration. Blockade with My4 antibody significantly inhibited LPS-induced tumor necrosis factor-alpha secretion at 1 ng/ml of LPS. However, inhibition decreased as we increased the LPS concentration, suggesting the existence of CD14-independent pathways of macrophage activation in response to LPS. Alternatively, My4 may have a lower affinity for the porcine CD14 antigen than LPS, which may have only partially blocked the LPS-LBP binding site at high concentrations of LPS. Therefore, these data suggest that LPS activation of porcine alveolar macrophages for 24 h increased CD14-like receptor expression. The degree of CD14-like up-regulation was related to LPS concentration, however, activation did not require the presence of serum at high concentrations of LPS.

  7. Lipopolysaccharide modulation of a CD14-like molecule on porcine alveolar macrophages

    NASA Technical Reports Server (NTRS)

    Kielian, T. L.; Ross, C. R.; McVey, D. S.; Chapes, S. K.; Blecha, F.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    Cluster of differentiation antigen 14 (CD14) functions as a receptor for lipopolysaccharide (LPS) LPS-binding protein (LBP) complexes. Because LPS has varying effects on CD14 expression in vitro, we evaluated CD14 expression in response to LPS with a fully differentiated macrophage phenotype, the alveolar macrophage. By using flow microfluorometric analysis and a radioimmunoassay with an anti-human CD14 monoclonal antibody (My4) that cross-reacts with porcine CD14, we found that macrophages stimulated with LPS for 24 h exhibited a two- to fivefold increase in CD14-like antigen compared with unstimulated cells. At low concentrations of LPS, up-regulation of the CD14-like antigen was dependent on serum; at higher concentrations of LPS, serum was not required. In the absence of serum a 10-fold higher dose of LPS (10 ng/ml) was required to increase CD14-like expression. In addition, LPS-induced CD14-like up-regulation correlated with secretion of tumor necrosis factor-alpha, regardless of serum concentration. Blockade with My4 antibody significantly inhibited LPS-induced tumor necrosis factor-alpha secretion at 1 ng/ml of LPS. However, inhibition decreased as we increased the LPS concentration, suggesting the existence of CD14-independent pathways of macrophage activation in response to LPS. Alternatively, My4 may have a lower affinity for the porcine CD14 antigen than LPS, which may have only partially blocked the LPS-LBP binding site at high concentrations of LPS. Therefore, these data suggest that LPS activation of porcine alveolar macrophages for 24 h increased CD14-like receptor expression. The degree of CD14-like up-regulation was related to LPS concentration, however, activation did not require the presence of serum at high concentrations of LPS.

  8. Analysis of CD14 Expression Levels in Putative Mesenchymal Progenitor Cells Isolated from Equine Bone Marrow

    PubMed Central

    Hackett, Catherine H.; Flaminio, Maria Julia B.F.

    2011-01-01

    A long-term goal of mesenchymal progenitor cell (MPC) research is to identify cell-surface markers to facilitate MPC isolation. One reported MPC feature in humans and other species is lack of CD14 (lipopolysaccharide receptor) expression. The aim of this study was to evaluate CD14 as an MPC sorting marker. Our hypothesis was that cells negatively selected by CD14 expression would enrich MPC colony formation compared with unsorted and CD14-positive fractions. After validation of reagents, bone marrow aspirate was obtained from 12 horses. Fresh and cultured cells were analyzed by flow cytometry and reverse transcription and quantitative polymerase chain reaction to assess dynamic changes in phenotype. In fresh samples, cells did not consistently express protein markers used for lineage classification. Short-term (2-day) culture allowed distinction between hematopoietic and nonhematopoietic populations. Magnetic activated cell sorting was performed on cells from 6 horses to separate adherent CD14+ from CD14− cells. MPC colony formation was assessed at 7 days. Cells positively selected for CD14 expression were significantly more likely to form MPC colonies than both unsorted and negatively selected cells (P ≤ 0.005). MPCs from all fractions maintained low levels of CD14 expression long term, and upregulated CD14 gene and protein expression when stimulated with lipopolysaccharide. The equine CD14 molecule was trypsin-labile, offering a plausible explanation for the discrepancy with MPC phenotypes reported in other species. By definition, MPCs are considered nonhematopoietic because they lack expression of molecules such as CD14. Our results challenge this assumption, as equine MPCs appear to represent a descendant of a CD14-positive cell. PMID:20722500

  9. Developmental variations in plasma leptin, leptin soluble receptor and their molar ratio in healthy infants.

    PubMed

    Koo, Winston W K; Hammami, Mouhanad; Hockman, Elaine M

    2007-06-04

    Leptin and its soluble receptor (sOB-R) are important to regulation of body composition but there are no data on the developmental variations in these plasma variables and their relationship with body composition measurements, Weight, length, and body composition (bone, fat and lean mass) by dual energy absorptiometry, and plasma variables were measured in healthy infants at 2, 4, 8 and 12 months. 15 whites and 29 African Americans (21 males and 23 females) with mean birth weight 3357 +/- 45 (SEM) g and gestation of 39.3 +/- 0.17 weeks were studied. The overall Z score for weight, length and weight for length during the study were 0.00 +/- 0.15, -0.08 +/- 0.11 and 0.12 +/- 0.14 respectively. With increasing age, plasma leptin (1.0 to 18.2, median 5.5 ng/mL) and sOB-R:leptin molar ratio (10.1 to 247.4, median 59.9) were lowered (r = -0.47, p < 0.01; and r = -0.37, p < 0.05 respectively), best predicted by weight Z score and percentage of fat mass, and higher in African American and female. Presence of body composition measurements eliminated the race and gender effect on the plasma variables. Plasma sOB-R (49.5 to 173.9, median 81.3 ng/mL) did not change significantly with age and was correlated and predicted only by body composition measurements. In healthy growing infants, plasma leptin but not sOB-R decreases with age. Gender, race and anthropometric measurements are additional physiological determinants predictive of plasma leptin and the receptor:ligand ratio. However, body composition is the only variable that can predict plasma leptin and its soluble receptor and the receptor: ligand ratio; and body composition measurements eliminated the race and gender effect on these plasma variables.

  10. Luminescent, water-soluble silicon quantum dots via micro-plasma surface treatment

    NASA Astrophysics Data System (ADS)

    Wu, Jeslin J.; Kondeti, Vighneswara Siva Santosh Kumar; Bruggeman, Peter J.; Kortshagen, Uwe R.

    2016-03-01

    Silicon quantum dots (SiQDs), with their broad absorption, narrow and size-tunable emission, and potential biocompatibility are highly attractive materials in biological imaging applications. The inherent hydrophobicity and instability of hydrogen-terminated SiQDs are obstacles to their widespread implementation. In this work, we successfully produced highly luminescent, hydrophilic SiQDs with long-term stability in water using non-thermal plasma techniques. Hydrogen-terminated SiQDs were produced in a low-pressure plasma and subsequently treated in water using an atmospheric-pressure plasma jet for surface modification. Preliminary assessments of the chemical mechanism(s) involved in the creation of water-soluble SiQDs were performed using Fenton’s reaction and various plasma chemistries, suggesting both OH and O species play a key role in the oxidation of the SiQDs.

  11. Contribution of CD14 and TLR4 to changes of the PI(4,5)P2 level in LPS-stimulated cells.

    PubMed

    Płóciennikowska, Agnieszka; Hromada-Judycka, Aneta; Dembińska, Justyna; Roszczenko, Paula; Ciesielska, Anna; Kwiatkowska, Katarzyna

    2016-12-01

    LPS binds sequentially to CD14 and TLR4/MD2 receptor triggering production of proinflammatory mediators. The LPS-induced signaling is controlled by a plasma membrane lipid PI(4,5)P2 and its derivatives. Here, we show that stimulation of murine peritoneal macrophages with LPS induces biphasic accumulation of PI(4,5)P2 with peaks at 10 and 60-90 min that were still seen after silencing of TLR4 expression. In contrast, the PI(4,5)P2 elevation was abrogated when CD14 was removed from the cell surface. To assess the contribution of CD14 and TLR4 to the LPS-induced PI(4,5)P2 changes, we used HEK293 transfectants expressing various amounts of CD14 and TLR4. In cells with a low content of CD14 and high of TLR4, no accumulation of PI(4,5)P2 occurred. With an increasing amount of CD14 and concomitant decrease of TLR4, 2 peaks of PI(4,5)P2 accumulation appeared, eventually approaching those found in LPS-stimulated cells expressing CD14 alone. Mutation of the signaling domain of TLR4 let us conclude that the receptor activity can modulate PI(4,5)P2 accumulation in cells when expressed in high amounts compared with CD14. Among the factors limiting PI(4,5)P2 accumulation are its hydrolysis, phosphorylation, and availability of its precursor, PI(4)P. Inhibition of PLC and PI3K or overexpression of PI4K IIα that produces PI(4)P promoted PI(4,5)P2 elevation in LPS-stimulated cells. The elevation of PI(4,5)P2 was dispensable for TLR4 signaling yet enhanced its magnitude. Taken together, these data suggest that LPS-induced accumulation of PI(4,5)P2 that maximizes TLR4 signaling is controlled by CD14, whereas TLR4 can fine tune the process by affecting the PI(4,5)P2 turnover.

  12. Soluble ectodomain CD163 and extracellular vesicle-associated CD163 are two differently regulated forms of ‘soluble CD163’ in plasma

    PubMed Central

    Etzerodt, Anders; Berg, Ronan M. G.; Plovsing, Ronni R.; Andersen, Morten N.; Bebien, Magali; Habbeddine, Mohamed; Lawrence, Toby; Møller, Holger J.; Moestrup, Søren K.

    2017-01-01

    CD163 is the macrophage receptor for uptake of hemoglobin-haptoglobin complexes. The human receptor can be shed from the macrophage surface owing to a cleavage site for the inflammation-inducible TACE/ADAM17 enzyme. Accordingly, plasmasoluble CD163’ (sCD163) has become a biomarker for macrophage activity and inflammation. The present study disclosed that 10% of sCD163 in healthy persons is actually extracellular vesicle (EV)-associated CD163 not being cleaved and shed. Endotoxin injection of human volunteers caused a selective increase in the ectodomain CD163, while septic patients exhibited high levels of both soluble ectodomain CD163 and extracellular vesicle (EV) CD163, the latter representing up 60% of total plasma CD163. A poor prognosis of septic patients measured as the sequential organ failure assessment (SOFA) score correlated with the increase in membrane-associated CD163. Our results show that soluble ectodomain CD163 and EV CD163 in plasma are part of separate macrophage response in the context of systemic inflammation. While that soluble ectodomain CD163 is released during the acute systemic inflammatory response, this is not the case for EV CD163 that instead may be released during a later phase of the inflammatory response. A separate measurement of the two forms of CD163 constituting ‘soluble CD163’ in plasma may therefore add to the diagnostic and prognostic value. PMID:28084321

  13. Soluble ectodomain CD163 and extracellular vesicle-associated CD163 are two differently regulated forms of 'soluble CD163' in plasma.

    PubMed

    Etzerodt, Anders; Berg, Ronan M G; Plovsing, Ronni R; Andersen, Morten N; Bebien, Magali; Habbeddine, Mohamed; Lawrence, Toby; Møller, Holger J; Moestrup, Søren K

    2017-01-13

    CD163 is the macrophage receptor for uptake of hemoglobin-haptoglobin complexes. The human receptor can be shed from the macrophage surface owing to a cleavage site for the inflammation-inducible TACE/ADAM17 enzyme. Accordingly, plasma 'soluble CD163' (sCD163) has become a biomarker for macrophage activity and inflammation. The present study disclosed that 10% of sCD163 in healthy persons is actually extracellular vesicle (EV)-associated CD163 not being cleaved and shed. Endotoxin injection of human volunteers caused a selective increase in the ectodomain CD163, while septic patients exhibited high levels of both soluble ectodomain CD163 and extracellular vesicle (EV) CD163, the latter representing up 60% of total plasma CD163. A poor prognosis of septic patients measured as the sequential organ failure assessment (SOFA) score correlated with the increase in membrane-associated CD163. Our results show that soluble ectodomain CD163 and EV CD163 in plasma are part of separate macrophage response in the context of systemic inflammation. While that soluble ectodomain CD163 is released during the acute systemic inflammatory response, this is not the case for EV CD163 that instead may be released during a later phase of the inflammatory response. A separate measurement of the two forms of CD163 constituting 'soluble CD163' in plasma may therefore add to the diagnostic and prognostic value.

  14. CD14 is not involved in the uptake of synthetic CpG oligonucleotides.

    PubMed

    Li, Jessica; Ahmet, Fatma; O'Keeffe, Meredith; Lahoud, Mireille H; Heath, William R; Caminschi, Irina

    2017-01-01

    We have previously shown that DEC205, a surface receptor expressed at high levels on CD8(+)DC, is able to capture synthetic CpG oligonucleotides (ODN) and is required for optimal responsiveness. However, even in the absence of DEC205, CD8(+)DC are able to respond to CpG ODN, albeit suboptimally. This suggested that additional receptors might contribute to the uptake of CpG ODN. CD14 represented an ideal candidate as it is expressed by DC and has been shown to bind and facilitate the uptake of CpG ODN. However, when CD14-deficient (CD14(-/-)) mice and normal B6 mice were injected with CpG ODN, CD8(+)DC were equivalently activated as assessed by the upregulation of the co-stimulatory molecules CD40 and CD80. Furthermore, the level of serum IL-6 and IL-12 produced in response to CpG ODN was comparable in CD14(-/-) and B6 mice. Importantly, mice deficient in both DEC205 and CD14 had comparable responses to mice lacking DEC205 alone, both in terms of cytokine production and DC activation, arguing that CD14 did not contribute to responses to CpG ODN. For CD14 to act as an uptake receptor for CpG ODN, it must first capture CpG ODN. To this end we assessed the capacity of cell surface CD14 to bind CpG ODN. Although we unequivocally confirmed that CD14 is required for the binding of its known ligand LPS, CD14 was not required for binding or responses to A-, B-, and C- Class CpG ODN. Our studies dispute the claim that CD14 is involved in CpG ODN capture.

  15. Identification and immunolocalization of the innate immune receptor CD14 in hypertrophic adenoids and tonsils.

    PubMed

    Ben-Yaakov, Avraham; Maly, Bella; Abu-Ita, Rani; Elidan, Josef; Gross, Menachem

    2011-01-01

    The purpose of this study is to determine the expression of CD14 as a marker of the innate immunity in hypertrophic adenoids and tonsils. Twenty-four pediatric patients (age <12 years) with obstructive adenotonsillar hypertrophy, confirmed by sleep study were included in this study. Intensity and expression of positive CD14 infiltrating cells was assessed by immunohistochemistry in specific histologic areas. In tonsils, CD14 immunoreactivity was demonstrated in intraepithelial lymphocytes located in the basal layer of the stratified squamous mucoepithelium. CD14 expression was significantly higher in mucosal layers and inter-follicular areas of tonsils than adenoid tissues [(p < 0.001), (p = 0.021), respectively]. CD14 expression was significantly higher in the submucosal layers of adenoids than tonsil tissues (p = 0.002). Hypertrophic adenoids and tonsils from children with OSA are prominent sites of innate defense, with over expression of CD14. The enhanced expressions of CD14 cells in adenoids and tonsils may be an important factor for the development and persistence of adenoids and tonsils enlargement causing OSA in children. CD14 expression in adenoids and tonsils illustrates an important immunological sentinel function of the innate immunity of the upper airway.

  16. Activation of human and mouse Kupffer cells by lipopolysaccharide is mediated by CD14.

    PubMed

    Su, Grace L; Goyert, Sanna M; Fan, Ming-Hui; Aminlari, Alireza; Gong, Ke Qin; Klein, Richard D; Myc, Andrzej; Alarcon, William H; Steinstraesser, Lars; Remick, Daniel G; Wang, Stewart C

    2002-09-01

    Upregulation of CD14 in Kupffer cells has been implicated in the pathogenesis of several forms of liver injury, including alcoholic liver disease. However, it remains unclear whether CD14 mediates lipopolysaccharide (LPS) signaling in this specialized liver macrophage population. In this series of experiments, we determined the role of CD14 in LPS activation of Kupffer cells by using several complementary approaches. First, we isolated Kupffer cells from human livers and studied the effects of anti-CD14 antibodies on LPS activation of these cells. Kupffer cells were incubated with increasing concentrations of LPS in the presence and absence of recombinant human LPS binding protein (LBP). With increasing concentrations of LPS, human Kupffer cell tumor necrosis factor-alpha (TNF-alpha) production (a marker for Kupffer cell activation) increased in a dose-dependent manner in the presence and absence of LBP. In the presence of anti-human CD14 antibodies, the production of TNF-alpha was significantly diminished. Second, we compared LPS activation of Kupffer cells isolated from wild-type and CD14 knockout mice. Kupffer cells from CD14 knockout mice produced significantly less TNF-alpha in response to the same amount of LPS. Together, these data strongly support a critical role for CD14 in Kupffer cell responses to LPS.

  17. The CD14 rs2569190 TT Genotype is Associated with Chronic Periodontitis.

    PubMed

    Ishaan, Dilshi; Parthasarathy, Harinath; Ponnaiyan, Deepa; Tadepalli, Anupama

    2017-09-01

    The genetic variation in the CD14 rs2569190 promoter region contributes to the susceptibility of developing periodontitis. Since, CD14 gene polymorphism studies among various ethnic populations have produced conflicting results, the present study was designed to unearth the association between the CD14 rs2569190 gene polymorphism and generalized chronic periodontitis (GCP) in a South Indian population of Tamil ethnicity. Polymorphisms in the CD14 rs2569190 promoter region were analyzed in 96 subjects who were periodontally healthy and 96 patients with GCP. DNA extracted from peripheral venous blood was genotyped by polymerase chain reaction-restriction fragment length polymorphism analysis. One-way analysis of variance, Chi-square testing, and logistic regression analysis were used to test association between GCP and the CD14 rs2569190 single nucleotide polymorphism (SNP). The distribution of the CD14 rs2569190 genotypes and allele frequencies showed significant differences between the test and control groups. Prevalence of the polymorphic TT genotype and T allele of the CD14 rs2569190 were significantly increased in GCP patients when compared with healthy controls. The results of the present study suggest that the T allele and the TT genotype of the rs2569190 SNP in the promoter region of the CD14 gene are associated with GCP in a South Indian population of Tamil ethnicity.

  18. Double blockade of CD14 and complement C5 abolishes the cytokine storm and improves morbidity and survival in polymicrobial sepsis in mice.

    PubMed

    Huber-Lang, Markus; Barratt-Due, Andreas; Pischke, Søren E; Sandanger, Øystein; Nilsson, Per H; Nunn, Miles A; Denk, Stephanie; Gaus, Wilhelm; Espevik, Terje; Mollnes, Tom E

    2014-06-01

    Sepsis and septic shock, caused by an excessive systemic host-inflammatory response, are associated with high morbidity and mortality. The complement system and TLRs provide important pattern recognition receptors initiating the cytokine storm by extensive cross-talk. We hypothesized that double blockade of complement C5 and the TLR coreceptor CD14 could improve survival of experimental polymicrobial sepsis. Mice undergoing cecal ligation and puncture (CLP)-induced sepsis were treated with neutralizing anti-CD14 Ab biG 53, complement C5 inhibitor coversin (Ornithodoros moubata C inhibitor), or a combination thereof. The inflammatory study (24-h observation) revealed statistically significant increases in 22 of 24 measured plasma biomarkers in the untreated CLP group, comprising 14 pro- and anti-inflammatory cytokines and 8 chemokines, growth factors, and granulocyte activation markers. Single CD14 or C5 blockade significantly inhibited 20 and 19 of the 22 biomarkers, respectively. Combined CD14 and C5 inhibition significantly reduced all 22 biomarkers (mean reduction 85%; range 54-95%) compared with the untreated CLP group. Double blockade was more potent than single treatment and was required to significantly inhibit IL-6 and CXCL1. Combined inhibition significantly reduced morbidity (motility and eyelid movement) and mortality measured over 10 d. In the positive control CLP group, median survival was 36 h (range 24-48 h). Combined treatment increased median survival to 96 h (range 24-240 h) (p = 0.001), whereas survival in the single-treatment groups was not significantly increased (median and range for anti-CD14 and anti-C5 treatment were 36 h [24-48 h] and 48 h [24-96 h]). Combined with standard intervention therapy, specific blockade of CD14 and C5 might represent a promising new therapeutic strategy for treatment of polymicrobial sepsis.

  19. Double Blockade of CD14 and Complement C5 Abolishes the Cytokine Storm and Improves Morbidity and Survival in Polymicrobial Sepsis in Mice

    PubMed Central

    Huber-Lang, Markus; Barratt-Due, Andreas; Pischke, Søren E.; Sandanger, Øystein; Nilsson, Per H.; Nunn, Miles A.; Denk, Stephanie; Gaus, Wilhelm; Espevik, Terje

    2014-01-01

    Sepsis and septic shock, caused by an excessive systemic host-inflammatory response, are associated with high morbidity and mortality. The complement system and TLRs provide important pattern recognition receptors initiating the cytokine storm by extensive cross-talk. We hypothesized that double blockade of complement C5 and the TLR coreceptor CD14 could improve survival of experimental polymicrobial sepsis. Mice undergoing cecal ligation and puncture (CLP)–induced sepsis were treated with neutralizing anti-CD14 Ab biG 53, complement C5 inhibitor coversin (Ornithodoros moubata C inhibitor), or a combination thereof. The inflammatory study (24-h observation) revealed statistically significant increases in 22 of 24 measured plasma biomarkers in the untreated CLP group, comprising 14 pro- and anti-inflammatory cytokines and 8 chemokines, growth factors, and granulocyte activation markers. Single CD14 or C5 blockade significantly inhibited 20 and 19 of the 22 biomarkers, respectively. Combined CD14 and C5 inhibition significantly reduced all 22 biomarkers (mean reduction 85%; range 54–95%) compared with the untreated CLP group. Double blockade was more potent than single treatment and was required to significantly inhibit IL-6 and CXCL1. Combined inhibition significantly reduced morbidity (motility and eyelid movement) and mortality measured over 10 d. In the positive control CLP group, median survival was 36 h (range 24–48 h). Combined treatment increased median survival to 96 h (range 24–240 h) (p = 0.001), whereas survival in the single-treatment groups was not significantly increased (median and range for anti-CD14 and anti-C5 treatment were 36 h [24–48 h] and 48 h [24–96 h]). Combined with standard intervention therapy, specific blockade of CD14 and C5 might represent a promising new therapeutic strategy for treatment of polymicrobial sepsis. PMID:24790148

  20. Lipoteichoic acid (LTA) of Streptococcus pneumoniae and Staphylococcus aureus activates immune cells via Toll-like receptor (TLR)-2, lipopolysaccharide-binding protein (LBP), and CD14, whereas TLR-4 and MD-2 are not involved.

    PubMed

    Schröder, Nicolas W J; Morath, Siegfried; Alexander, Christian; Hamann, Lutz; Hartung, Thomas; Zähringer, Ulrich; Göbel, Ulf B; Weber, Joerg R; Schumann, Ralf R

    2003-05-02

    Lipoteichoic acid (LTA) derived from Streptococcus pneumoniae, purified employing a chloroform/methanol protocol, and from Staphylococcus aureus, prepared by the recently described butanol extraction procedure, was investigated regarding its interaction with lipopolysaccharide (LPS)-binding protein (LBP), CD14, Toll-like receptors (TLRs)-2 and -4, and MD-2. LTA from both organisms induced cytokine synthesis in human mononuclear phagocytes. Activation was LBP- and CD14-dependent, and formation of complexes of LTA with LBP and soluble CD14 as well as catalytic transfer of LTA to CD14 by LBP was verified by PhastGel(TM) native gel electrophoresis. Human embryonic kidney (HEK) 293/CD14 cells and Chinese hamster ovary (CHO) cells were responsive to LTA only after transfection with TLR-2. Additional transfection with MD-2 did not affect stimulation of these cells by LTA. Our data suggest that innate immune recognition of LTA via LBP, CD14, and TLR-2 represents an important mechanism in the pathogenesis of systemic complications in the course of infectious diseases brought about by the clinically most important Gram-positive pathogens. However, the involvement of TLR-4 and MD-2 in this process was ruled out.

  1. Increased plasma soluble CD40 ligand concentration in pelvic inflammatory disease.

    PubMed

    Ho, Tsung-chin; Yang, Shun-Fa; Wang, Po-Hui; Lin, Long-Yau; Tee, Yi-Torng; Liao, Wen-Chun; Chang, Hsiu-Ju; Tsai, Hsiu-Ting

    2015-01-01

    The role of soluble CD40 ligand (sCD40L) in pelvic inflammatory disease (PID) remains unclear. We sought to determine whether sCD40L was an efficient serum marker as with WBC and CRP in PID patients. Enzyme-linked immunosorbent assay was used to measure the plasma levels of sCD40L before and after routine protocol treatments in sixty-four PID patients and seventy healthy controls. The level of plasma sCD40L (pg/ml) was significantly elevated in PID patients (1632.83±270.91) compared to that in normal controls (700.33±58.77; p=0.001) and decreased significantly as compared to that in the same patients (928.77±177.25; p=0.0001) after they received treatment. The concentration of sCD40L was significantly correlated with the level of plasma C-reactive protein (CRP) in the blood (r=0.202, p=0.01, n=134). When the cutoff level of plasma sCD40L levels was determined to be 1612.26pg/ml based on ROC, the sensitivity, specificity, and the area under the curve of plasma sCD40L level for predicting PID were 0.26, 0.97, and 0.58 (95% confidence interval: 0.48-0.68), respectively, while the adjusted odds ratio (AOR) with their 95% CI of plasma sCD40L for PID risk was 7.09 (95% CI=1.14-43.87, p=0.03). The expression of plasma sCD40L was increased in patients with PID and detection of plasma sCD40L could be useful for the diagnosis of PID. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Bilirubin adsorption properties of water-soluble adsorbents with different cyclodextrin cavities in plasma dialysis system.

    PubMed

    Wang, Zhi; Cao, Yaming; Wei, Houliang; Jia, Lingyun; Xu, Li; Xie, Jian

    2012-02-01

    In this study, we explored the use of α-, β- or γ-cyclodextrin (CD)-grafted polyethyleneimine (PEI) as water-soluble adsorbent for removing excess plasma bilirubin. To evaluate the bilirubin-binding capacity of these adsorbents, bovine serum albumin (BSA) solution or plasma with high level of bilirubin were dialyzed against CD-PEI-spiked dialysate. In BSA solution with an initial biliurbin concentration of 171.5mg/L, α-CD-PEI, β-CD-PEI and γ-CD-PEI achieved adsorption capacities of 2.5, 5.8 and 3.8 mg/g, respectively. In a plasma dialysis system, 45.6% of bilirubin (260 mg/L) was removed from 200 mL plasma by 1L dialysate spiked with 10mg/mL β-CD-PEI, which was significantly higher than that removed by the same volume of BSA-spiked dialysate (P<0.05), demonstrating the strong bilirubin-binding ability of β-CD-PEI. The key feature of bilirubin adsorption was related to the CD functional group, not the PEI matrix. Subsequent molecular docking study indicated that the size of CD cavity could affect the affinity energy of CD-bilirubin complex. The cavity of β-CD was most suitable for accommodating the pyrrole rings of bilirubin. The inclusion complex of bilirubin and β-CD in the molar ratio of 1:2 was more logical in terms of affinity energy. All the results demonstrated the potential of β-CD-PEI (water-soluble adsorbent) as an effective agent for removing of bilirubin from plasma in dialysis system. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. Generation of Large Numbers of Antigen-Expressing Human Dendritic Cells Using CD14-ML Technology

    PubMed Central

    Imamura, Yuya; Haruta, Miwa; Tomita, Yusuke; Matsumura, Keiko; Ikeda, Tokunori; Yuno, Akira; Hirayama, Masatoshi; Nakayama, Hideki; Mizuta, Hiroshi; Nishimura, Yasuharu; Senju, Satoru

    2016-01-01

    We previously reported a method to expand human monocytes through lentivirus-mediated introduction of cMYC and BMI1, and we named the monocyte-derived proliferating cells, CD14-ML. CD14-ML differentiated into functional DC (CD14-ML-DC) upon addition of IL-4, resulting in the generation of a large number of DC. One drawback of this method was the extensive donor-dependent variation in proliferation efficiency. In the current study, we found that introduction of BCL2 or LYL1 along with cMYC and BMI1 was beneficial. Using the improved method, we obtained CD14-ML from all samples, regardless of whether the donors were healthy individuals or cancer patients. In vitro stimulation of peripheral blood T cells with CD14-ML-DC that were loaded with cancer antigen-derived peptides led to the establishment of CD4+ and CD8+ T cell lines that recognized the peptides. Since CD14-ML was propagated for more than 1 month, we could readily conduct genetic modification experiments. To generate CD14-ML-DC that expressed antigenic proteins, we introduced lentiviral antigen-expression vectors and subjected the cells to 2 weeks of culture for drug-selection and expansion. The resulting antigen-expressing CD14-ML-DC successfully induced CD8+ T cell lines that were reactive to CMVpp65 or MART1/MelanA, suggesting an application in vaccination therapy. Thus, this improved method enables the generation of a sufficient number of DC for vaccination therapy from a small amount of peripheral blood from cancer patients. Information on T cell epitopes is not necessary in vaccination with cancer antigen-expressing CD14-ML-DC; therefore, all patients, irrespective of HLA type, will benefit from anti-cancer therapy based on this technology. PMID:27050553

  4. Generation of Large Numbers of Antigen-Expressing Human Dendritic Cells Using CD14-ML Technology.

    PubMed

    Imamura, Yuya; Haruta, Miwa; Tomita, Yusuke; Matsumura, Keiko; Ikeda, Tokunori; Yuno, Akira; Hirayama, Masatoshi; Nakayama, Hideki; Mizuta, Hiroshi; Nishimura, Yasuharu; Senju, Satoru

    2016-01-01

    We previously reported a method to expand human monocytes through lentivirus-mediated introduction of cMYC and BMI1, and we named the monocyte-derived proliferating cells, CD14-ML. CD14-ML differentiated into functional DC (CD14-ML-DC) upon addition of IL-4, resulting in the generation of a large number of DC. One drawback of this method was the extensive donor-dependent variation in proliferation efficiency. In the current study, we found that introduction of BCL2 or LYL1 along with cMYC and BMI1 was beneficial. Using the improved method, we obtained CD14-ML from all samples, regardless of whether the donors were healthy individuals or cancer patients. In vitro stimulation of peripheral blood T cells with CD14-ML-DC that were loaded with cancer antigen-derived peptides led to the establishment of CD4+ and CD8+ T cell lines that recognized the peptides. Since CD14-ML was propagated for more than 1 month, we could readily conduct genetic modification experiments. To generate CD14-ML-DC that expressed antigenic proteins, we introduced lentiviral antigen-expression vectors and subjected the cells to 2 weeks of culture for drug-selection and expansion. The resulting antigen-expressing CD14-ML-DC successfully induced CD8+ T cell lines that were reactive to CMVpp65 or MART1/MelanA, suggesting an application in vaccination therapy. Thus, this improved method enables the generation of a sufficient number of DC for vaccination therapy from a small amount of peripheral blood from cancer patients. Information on T cell epitopes is not necessary in vaccination with cancer antigen-expressing CD14-ML-DC; therefore, all patients, irrespective of HLA type, will benefit from anti-cancer therapy based on this technology.

  5. Plasma soluble L-selectin in medicated patients with schizophrenia and healthy controls

    PubMed Central

    Yang, Fang; Amin, Pooja A.; Zunta-Soares, Giovana; Colpo, Gabriela D.; Stertz, Laura; Sharma, Ajaykumar N.; Fries, Gabriel R.; Walss-Bass, Consuelo; Soares, Jair C.; Okusaga, Olaoluwa O.

    2017-01-01

    Immune dysfunction has been implicated in the pathophysiology of schizophrenia. Leukocyte migration to the site of inflammation is a fundamental step of immune response which involves P-, E-, and L-selectins. Elevated selectin levels have been reported in un-medicated first-episode patients with schizophrenia but not in medicated patients with multi-episode schizophrenia. We measured fasting plasma soluble P-, E-, and L-selectin in 39 medicated patients with multi-episode schizophrenia and 19 healthy controls. In patients, psychotic symptom severity and cognitive function were assessed with the Positive and Negative Syndrome Scale (PANSS) and the NIH Toolbox Cognitive Test Battery respectively. C-reactive protein (CRP) and Body Mass Index (BMI) were measured in patients and controls. Comparison of selectin levels between patients and controls was done with t-tests and linear regression. Pearson correlation coefficients between plasma selectins and PANSS and cognitive measures were calculated. Geometric mean plasma soluble L-selectin level was lower in patients compared to controls from unadjusted (606.7 ± 1.2 ng/ml vs. 937.7 ± 1.15 ng/ml, p < 0.001) and adjusted analyses (β = 0.59; CI 0.41 to 0.88, p = 0.011). There was a trend towards higher plasma soluble P-selectin in patients compared to controls (90.4 ± 1.2ng/ml vs. 71.8 ± 1.2ng/ml, p = 0.059) in the unadjusted analysis. There was no association between the selectins and psychotic symptoms or cognitive function in the patients. In addition, the selectins were not significantly associated with CRP or BMI. The limitations of this study include small sample size and unavailability of information on medications and blood cell counts. The potential utility of soluble L-selectin as a biomarker of antipsychotic exposure in patients with schizophrenia and the concomitant change in immune response with the use of antipsychotics should be further evaluated. PMID:28334045

  6. The Th17/Treg Ratio, IL-1RA and sCD14 Levels in Primary HIV Infection Predict the T-cell Activation Set Point in the Absence of Systemic Microbial Translocation

    PubMed Central

    Chevalier, Mathieu F.; Petitjean, Gaël; Dunyach-Rémy, Catherine; Didier, Céline; Girard, Pierre-Marie; Manea, Maria Elena; Campa, Pauline; Meyer, Laurence; Rouzioux, Christine; Lavigne, Jean-Philippe; Barré-Sinoussi, Françoise; Scott-Algara, Daniel; Weiss, Laurence

    2013-01-01

    Impairment of the intestinal barrier and subsequent microbial translocation (MT) may be involved in chronic immune activation, which plays a central role in HIV pathogenesis. Th17 cells are critical to prevent MT. The aim of the study was to investigate, in patients with primary HIV infection (PHI), the early relationship between the Th17/Treg ratio, monocyte activation and MT and their impact on the T-cell activation set point, which is known to predict disease progression. 27 patients with early PHI were included in a prospective longitudinal study and followed-up for 6 months. At baseline, the Th17/Treg ratio strongly negatively correlated with the proportion of activated CD8 T cells expressing CD38/HLA-DR or Ki-67. Also, the Th17/Treg ratio was negatively related to viral load and plasma levels of sCD14 and IL-1RA, two markers of monocyte activation. In untreated patients, the Th17/Treg ratio at baseline negatively correlated with CD8 T-cell activation at month 6 defining the T-cell activation set point (% HLA-DR+CD38+ and %Ki-67+). Soluble CD14 and IL-1RA plasma levels also predicted the T-cell activation set point. Levels of I-FABP, a marker of mucosal damages, were similar to healthy controls at baseline but increased at month 6. No decrease in anti-endotoxin core antibody (EndoCAb) and no peptidoglycan were detected during PHI. In addition, 16S rDNA was only detected at low levels in 2 out 27 patients at baseline and in one additional patient at M6. Altogether, data support the hypothesis that T-cell and monocyte activation in PHI are not primarily driven by systemic MT but rather by viral replication. Moreover, the “innate immune set point” defined by the early levels of sCD14 and IL-1RA might be powerful early surrogate markers for disease progression and should be considered for use in clinical practice. PMID:23818854

  7. Prolonged interferon-gamma application by subcutaneous infusion in cancer patients: differential response of serum CD14, neopterin, and monocyte HLA class I and II antigens.

    PubMed

    Landmann, R; Ludwig, C; Wesp, M; Fisscher, A; Obrist, R; Knüsli, C; Denz, H; Obrecht, J P

    1992-04-01

    This study reports on biological response modification induced by prolonged continuous subcutaneous (s.c.) infusion of recombinant interferon-gamma (rIFN-gamma) with particular attention to changes of soluble CD14. This glycoprotein with an unknown function is derived from myeloid cells carrying membrane CD14, which is the receptor for lipopolysaccharide (LPS)-LPS-binding protein (LBP) complexes. Fifteen metastatic cancer patients received weekly escalating doses of rIFN-gamma starting at either 50 or 100 micrograms/24 h and increasing up to 400 micrograms/24 h for a median duration of 6 weeks. The maximum tolerated dose was higher (200 micrograms/24 h) with the lower (50 micrograms/24 h) starting dose. Biological activity of rIFN-gamma was evaluated by weekly measurements of CD14, neopterin, and beta 2-microglobulin concentrations in serum as well as monocyte HLA class I and II antigen expression and tumor cytotoxicity. Serum IFN-gamma concentrations increased 20-fold within 4 weeks of therapy. The levels were correlated to the mean dose (r = 0.95, p less than 0.05). Among the biological markers, two patterns were observed. First, serum CD14 concentration and expression of monocyte HLA class II antigens increased significantly during the first week, and marker expression correlated with serum IFN-gamma levels (p less than 0.05); CD14 and HLA class II antigens thereafter returned to pretreatment levels within 4 weeks of therapy despite persistently elevated serum IFN-gamma concentrations. Second, serum neopterin and beta 2-microglobulin concentrations as well as monocyte HLA class I expression also increased significantly within the first week, but remained elevated thereafter without any further dose relationship.(ABSTRACT TRUNCATED AT 250 WORDS)

  8. Human Lipopolysaccharide-binding Protein (LBP) and CD14 Independently Deliver Triacylated Lipoproteins to Toll-like Receptor 1 (TLR1) and TLR2 and Enhance Formation of the Ternary Signaling Complex*

    PubMed Central

    Ranoa, Diana Rose E.; Kelley, Stacy L.; Tapping, Richard I.

    2013-01-01

    Bacterial lipoproteins are the most potent microbial agonists for the Toll-like receptor 2 (TLR2) subfamily, and this pattern recognition event induces cellular activation, leading to host immune responses. Triacylated bacterial lipoproteins coordinately bind TLR1 and TLR2, resulting in a stable ternary complex that drives intracellular signaling. The sensitivity of TLR-expressing cells to lipoproteins is greatly enhanced by two lipid-binding serum proteins known as lipopolysaccharide-binding protein (LBP) and soluble CD14 (sCD14); however, the physical mechanism that underlies this increased sensitivity is not known. To address this, we measured the ability of LBP and sCD14 to drive ternary complex formation between soluble extracellular domains of TLR1 and TLR2 and a synthetic triacylated lipopeptide agonist. Importantly, addition of substoichiometric amounts of either LBP or sCD14 significantly enhanced formation of a TLR1·TLR2 lipopeptide ternary complex as measured by size exclusion chromatography. However, neither LBP nor sCD14 was physically associated with the final ternary complex. Similar results were obtained using outer surface protein A (OspA), a naturally occurring triacylated lipoprotein agonist from Borrelia burgdorferi. Activation studies revealed that either LBP or sCD14 sensitized TLR-expressing cells to nanogram levels of either the synthetic lipopeptide or OspA lipoprotein agonist. Together, our results show that either LBP or sCD14 can drive ternary complex formation and TLR activation by acting as mobile carriers of triacylated lipopeptides or lipoproteins. PMID:23430250

  9. Plasma Soluble CD163 Level Independently Predicts All-Cause Mortality in HIV-1-Infected Individuals.

    PubMed

    Knudsen, Troels Bygum; Ertner, Gideon; Petersen, Janne; Møller, Holger Jon; Moestrup, Søren K; Eugen-Olsen, Jesper; Kronborg, Gitte; Benfield, Thomas

    2016-10-15

    CD163, a monocyte- and macrophage-specific scavenger receptor, is shed as soluble CD163 (sCD163) during the proinflammatory response. Here, we assessed the association between plasma sCD163 levels and progression to AIDS and all-cause mortality among individuals infected with human immunodeficiency virus type 1 (HIV). Plasma sCD163 levels were measured in 933 HIV-infected individuals. Hazard ratios (HRs) with 95% confidence intervals (CIs) associated with mortality were computed by Cox proportional hazards regression. At baseline, 86% were receiving antiretroviral treatment, 73% had plasma a HIV RNA level of <50 copies/mL, and the median CD4(+) T-cell count was 503 cells/µL. During 10.5 years of follow-up, 167 (17.9%) died. Plasma sCD163 levels were higher in nonsurvivors than in survivors (4.92 mg/L [interquartile range {IQR}, 3.29-8.65 mg/L] vs 3.16 mg/L [IQR, 2.16-4.64 mg/L]; P = .0001). The cumulative incidence of death increased with increasing plasma sCD163 levels, corresponding to a 6% or 35% increased risk of death for each milligram per liter or quartile increase, respectively, in baseline plasma sCD163 level (adjusted HR, 1.06 [95% CI, 1.03-1.09] and 1.35 [95% CI, 1.13-1.63], respectively). Plasma sCD163 was an independent marker of all-cause mortality in a cohort of HIV-infected individuals, suggesting that monocyte/macrophage activation may play a role in HIV pathogenesis and be a target of intervention. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  10. Temporal plasma vitamin concentrations are altered by fat-soluble vitamin administration in suckling pigs.

    PubMed

    Jang, Y D; Ma, J Y; Monegue, J S; Monegue, H J; Stuart, R L; Lindemann, M D

    2015-11-01

    Piglets are born with purportedly low plasma vitamin D levels. The objective of this study was to investigate the effect of fat-soluble vitamin administration, primarily vitamin D, by different administration routes on plasma vitamin concentrations in suckling pigs. A total of 45 pigs from 5 litters were allotted at birth to 3 treatments within each litter. Pigs were administered 400 IU of α-tocopherol, 40,000 IU of retinyl palmitate, and 40,000 IU of vitamin D at d 1 of age either orally or by i.m. injection and compared with control pigs with no supplemental vitamin administration. Blood samples were collected at d 0 (initial), 1, 2, 3, 4, 6, 9, 14, and 20 after administration. Plasma 25-hydroxycholecalciferol (25OHD), α-tocopherol, retinyl palmitate, and retinol concentrations were analyzed. Except for retinol, the effects of treatment, day, and day × treatment interaction ( < 0.01) were observed on plasma vitamin concentrations. Plasma concentrations of 25OHD and α-tocopherol increased immediately regardless of administration routes to peak at d 2 and 1 after administration, respectively. Plasma retinyl palmitate concentrations increased only with the injection treatment, with the peak at d 1 after administration. Plasma concentrations of 25OHD in both administration treatments and α-tocopherol in the injection treatment were maintained at greater levels than those in the control treatment until d 20 after administration. With regard to the pharmacokinetic parameters for plasma 25OHD concentrations, the injection treatment had greater elimination half-life ( < 0.01), maximum plasma concentrations ( < 0.05), and all area under the curve parameters ( < 0.01) but a lower elimination rate constant ( < 0.01) than the oral treatment. Relative bioavailability of oral administration compared with injection administration was 55.26%. These results indicate that plasma status of 25OHD,α-tocopherol, and retinyl palmitate are differentially changed between types of

  11. Levels of plasma soluble complement receptor 1 (sCR1) in normal Indian adult population.

    PubMed

    Sivasankar, B; Raju, K R; Anand, V; Malu, S; Padmanabhan, S; Tiwari, S C; Das, N; Srivastava, L M

    1999-07-01

    A decrease in the membrane anchored erythrocyte complement receptor 1 (CR1) is reported as an acquired phenomenon in a number of inflammatory and autoimmune diseases with concomitant rise in soluble CR1 (sCR1) levels in plasma. There is a need to establish the normal range of sCR1 in Indian adults to assess the function and disease association of this protein. The plasma sCR1 levels of 50 healthy individuals have been estimated by an indigenously developed sandwich ELISA. sCR1 levels from 26 patients suffering from nephropathies had also been assayed which was much higher than the normal controls. This observation suggests sCR1 as a potential market for the assessment of disease activity in nephropathies.

  12. Synthesis, solubility, plasma stability, and pharmacological evaluation of novel sulfonylhydrazones designed as anti-diabetic agents

    PubMed Central

    Zapata-Sudo, Gisele; da Costa Nunes, Isabelle Karine; Araujo, Josenildo Segundo Chaves; da Silva, Jaqueline Soares; Trachez, Margarete Manhães; da Silva, Tiago Fernandes; da Costa, Filipe P; Sudo, Roberto Takashi; Barreiro, Eliezer J; Lima, Lídia Moreira

    2016-01-01

    Neuropathy is a serious complication of diabetes that has a significant socioeconomic impact, since it frequently demands high levels of health care consumption and compromises labor productivity. Recently, LASSBio-1471 (3) was demonstrated to improve oral glucose tolerance, reduce blood glucose levels, and display an anti-neuropathy effect in a murine streptozotocin-induced diabetes model. In the present work, we describe the design, synthesis, solubility, plasma stability, and pharmacological evaluation of novel sulfonylhydrazone derivatives (referred to herein as compounds 4–9), which were designed by molecular modification based on the structure of the prototype LASSBio-1471 (3). Among the compounds tested, better plasma stability was observed with 4, 5, and 9 in comparison to compounds 6, 7, and 8. LASSBio-1773 (7), promoted not only hypoglycemic activity but also the reduction of thermal hyperalgesia and mechanical allodynia in a murine model of streptozotocin-induced diabetic neuropathic pain. PMID:27672310

  13. Changes in plasma cytokines and their soluble receptors in complex regional pain syndrome.

    PubMed

    Alexander, Guillermo M; Peterlin, B Lee; Perreault, Marielle J; Grothusen, John R; Schwartzman, Robert J

    2012-01-01

    Complex Regional Pain Syndrome (CRPS) is a chronic and often disabling pain disorder. There is evidence demonstrating that neurogenic inflammation and activation of the immune system play a significant role in the pathophysiology of CRPS. This study evaluated the plasma levels of cytokines, chemokines, and their soluble receptors in 148 subjects afflicted with CRPS and in 60 gender- and age-matched healthy controls. Significant changes in plasma cytokines, chemokines, and their soluble receptors were found in subjects with CRPS as compared with healthy controls. For most analytes, these changes resulted from a distinct subset of the CRPS subjects. When the plasma data from the CRPS subjects was subjected to cluster analysis, it revealed 2 clusters within the CRPS population. The category identified as most important for cluster separation by the clustering algorithm was TNFα. Cluster 1 consisted of 64% of CRPS subjects and demonstrated analyte values similar to the healthy control individuals. Cluster 2 consisted of 36% of the CRPS subjects and demonstrated significantly elevated levels of most analytes and in addition, it showed that the increased plasma analyte levels in this cluster were correlated with disease duration and severity. The identification of biomarkers that define disease subgroups can be of great value in the design of specific therapies and of great benefit to the design of clinical trials. It may also aid in advancing our understanding of the mechanisms involved in the pathophysiology of CRPS, which may lead to novel treatments for this very severe condition. Copyright © 2012 American Pain Society. Published by Elsevier Inc. All rights reserved.

  14. Plasma soluble (pro)renin receptor is independent of plasma renin, prorenin, and aldosterone concentrations but is affected by ethnicity.

    PubMed

    Nguyen, Geneviève; Blanchard, Anne; Curis, Emmanuel; Bergerot, Damien; Chambon, Yann; Hirose, Takuo; Caumont-Prim, Aurore; Tabard, Sylvie Brailly; Baron, Stéphanie; Frank, Michael; Totsune, Kazuhito; Azizi, Michel

    2014-02-01

    A soluble (pro)renin receptor (sPRR) circulates in plasma and is able to bind renin and prorenin. It is not known whether plasma sPRR concentrations vary with the activity of the renin-angiotensin system. We measured plasma sPRR, renin, prorenin, and aldosterone concentrations in 121 white and 9 black healthy subjects, 40 patients with diabetes mellitus, 41 hypertensive patients with or without renin-angiotensin system blockers, 9 patients with primary aldosteronism, and 10 patients with Gitelman syndrome. Median physiological plasma sPRR concentration was 23.5 ng/mL (interquartile range, 20.9-26.5) under usual uncontrolled sodium diet. sPRR concentration in healthy subjects, unlike renin and prorenin, did not display circadian variation or dependence on age, sex, posture, or hormonal status. sPRR concentrations were ≈25% lower in black than in white subjects, whereas renin concentrations were ≈40% lower. Patients with diabetes mellitus (average renin-high prorenin levels) and with hypertension only (average renin-average prorenin levels) had sPRR concentrations similar to healthy subjects. Renin-angiotensin system blockade was associated with increase of sPRR concentration by ≈12%. sPRR in patients with primary aldosteronism (low renin-low prorenin) and Gitelman syndrome (high renin-high prorenin) were similar and ≈10% higher than in healthy subjects. There was no correlation between sPRR and renin or prorenin. In conclusion, our results show that plasma sPRR concentrations are dependent on ethnicity and independent of renin, prorenin, and aldosterone concentrations in healthy subjects and in patients with contrasted degrees of renin-angiotensin system activity.

  15. EV71-infected CD14(+) cells modulate the immune activity of T lymphocytes in rhesus monkeys.

    PubMed

    Wang, Jingjing; Pu, Jing; Huang, Hongtai; Zhang, Ying; Liu, Longding; Yang, Erxia; Zhou, Xiaofang; Ma, Na; Zhao, Hongling; Wang, Lichun; Xie, Zhenfeng; Tang, Donghong; Li, Qihan

    2013-07-01

    Preliminary studies of the major pathogen enterovirus 71 (EV71), a member of the Picornaviridae family, have suggested that EV71 may be a major cause of fatal hand, foot and mouth disease cases. Currently, the role of the pathological changes induced by EV71 infection in the immunopathogenic response remains unclear. Our study focused on the interaction between this virus and immunocytes and indicated that this virus has the ability to replicate in CD14(+) cells. Furthermore, these EV71-infected CD14(+) cells have the capacity to stimulate the proliferation of T cells and to enhance the release of certain functional cytokines. An adaptive immune response induced by the back-transfusion of EV71-infected CD14(+) cells was observed in donor neonatal rhesus monkeys. Based on these observations, the proposed hypothesis is that CD14(+) cells infected by the EV71 virus might modulate the anti-EV71 adaptive immune response by inducing simultaneous T-cell activation.

  16. CD14-DEPENDENT AIRWAY NEUTROPHIL RESPONSE TO INHALED LPS: ROLE OF ATOPY

    EPA Science Inventory

    Background: Inhaled endotoxin (LPS) is associated with airway neutrophilic (PMN) inflammation in both asthmatic and control subjects, with asthmatic subjects demonstrating possibly higher sensitivity. CD14 is the principal receptor mediating LPS responses in vivo. It is unkown ...

  17. Plasma levels of soluble tumour necrosis factor receptors are increased in coal miners with pneumoconiosis.

    PubMed

    Schins, R P; Borm, P J

    1995-10-01

    Among other cytokines, tumour necrosis factor (TNF)-alpha is considered to play a key role in the development of mineral dust related fibrosis. Previously, we showed that ex-vivo release of TNF by peripheral blood monocytes is a marker for progression of coal workers' pneumoconiosis (CWP). Since soluble TNF receptors (sTNF-Rs) are believed to play an important regulatory role in systemic effects of TNF, we measured plasma levels of sTNF-R55 and sTNF-R75 in coal miners with (n = 28) or without (n = 76) CWP and in nonexposed controls (n = 29). sTNF-R75 levels were significantly increased in miners with CWP (2.09 +/- 0.44 ng.mL-1) versus the nonexposed controls (1.86 +/- 0.23 ng.mL-1). Neither sTNF-R55 nor sTNF-R75 were related to exposure, stage of pneumoconiosis, smoking, or (spontaneous or ex-vivo induced) monocyte TNF-release. sTNF-R55 was increased in subjects with medication (especially those using cardiovascular drugs); upon exclusion of these subjects, sTNF-R55 was found also to be significantly increased in CWP. In conclusion, bearing in mind a confounding effect of medication, soluble TNF receptors are elevated in plasma of retired miners with coal workers' pneumoconiosis. These observations further support the important role of TNF-mediated pathways in the pathogenesis of mineral dust related fibrosis.

  18. Biochemical and mass spectrometric characterization of soluble ecto-5'-nucleotidase from bull seminal plasma.

    PubMed

    Fini, Carlo; Talamo, Fabio; Cherri, Silvia; Coli, Marcello; Floridi, Ardesio; Ferrara, Lino; Scaloni, Andrea

    2003-06-01

    Ecto-5'-nucleotidase (ecto-5'-NT) is a glycosylphosphatidylinositol-anchored membrane-bound protein that is ubiquitous in mammalian tissues. It is a target for a number of therapeutic drugs since increased levels of the enzyme correlate with various disease states. In this investigation, we describe the properties of a soluble ecto-5'-NT derived from bull seminal plasma. The protein was highly heterogeneous as demonstrated by chromatofocusing and two-dimensional PAGE. Sequencing analyses revealed a truncated polypeptide lacking the glycosylphospatidylinositol attachment site, suggesting that it is produced post-translationally by cleavage at Gln(547) and/or Phe(548). Heterogeneity was largely due to differential glycosylation, especially in the oligosaccharides linked to Asn(403). Significant differences in substrate specificity were observed between isoforms and, on the basis of molecular-modelling studies, were interpreted in terms of variable glycosylation causing steric hindrance of the substrate-binding site. Thus the soluble forms of ecto-5'-NT found in bull seminal plasma are unique both biochemically and structurally, and have a putative role in signalling interactions with spermatozoa following ejaculation and capacitation in the female reproductive tract.

  19. Biochemical and mass spectrometric characterization of soluble ecto-5'-nucleotidase from bull seminal plasma.

    PubMed Central

    Fini, Carlo; Talamo, Fabio; Cherri, Silvia; Coli, Marcello; Floridi, Ardesio; Ferrara, Lino; Scaloni, Andrea

    2003-01-01

    Ecto-5'-nucleotidase (ecto-5'-NT) is a glycosylphosphatidylinositol-anchored membrane-bound protein that is ubiquitous in mammalian tissues. It is a target for a number of therapeutic drugs since increased levels of the enzyme correlate with various disease states. In this investigation, we describe the properties of a soluble ecto-5'-NT derived from bull seminal plasma. The protein was highly heterogeneous as demonstrated by chromatofocusing and two-dimensional PAGE. Sequencing analyses revealed a truncated polypeptide lacking the glycosylphospatidylinositol attachment site, suggesting that it is produced post-translationally by cleavage at Gln(547) and/or Phe(548). Heterogeneity was largely due to differential glycosylation, especially in the oligosaccharides linked to Asn(403). Significant differences in substrate specificity were observed between isoforms and, on the basis of molecular-modelling studies, were interpreted in terms of variable glycosylation causing steric hindrance of the substrate-binding site. Thus the soluble forms of ecto-5'-NT found in bull seminal plasma are unique both biochemically and structurally, and have a putative role in signalling interactions with spermatozoa following ejaculation and capacitation in the female reproductive tract. PMID:12608891

  20. Senescent CD14+CD16+ monocytes exhibit proinflammatory and proatherosclerotic activity.

    PubMed

    Merino, Ana; Buendia, Paula; Martin-Malo, Alejandro; Aljama, Pedro; Ramirez, Rafael; Carracedo, Julia

    2011-02-01

    In elderly subjects and in patients with chronic inflammatory diseases, there is an increased subset of monocytes with a CD14(+)CD16(+) phenotype, whose origin and functional relevance has not been well characterized. In this study, we determined whether prolonged survival of human CD14(++)CD16(-) monocytes promotes the emergence of senescent cells, and we analyzed their molecular phenotypic and functional characteristics. We used an in vitro model to prolong the life span of healthy monocytes. We determined cell senescence, intracellular cytokine expression, ability to interact with endothelial cells, and APC activity. CD14(+)CD16(+) monocytes were senescent cells with shortened telomeres (215 ± 37 relative telomere length) versus CD14(++)CD16(-) cells (339 ± 44 relative telomere length; p < 0.05) and increased expression of β-galactosidase (86.4 ± 16.4% versus 10.3 ± 7.5%, respectively; p = 0.002). CD14(+)CD16(+) monocytes exhibited features of activated cells that included expression of CD209, release of cytokines in response to low-intensity stimulus, and increased capacity to sustain lymphocyte proliferation. Finally, compared with CD14(++)CD16(-) cells, CD14(+)CD16(+) monocytes showed elevated expression of chemokine receptors and increased adhesion to endothelial cells (19.6 ± 8.1% versus 5.3 ± 4.1%; p = 0.033). In summary, our data indicated that the senescent CD14(+)CD16(+) monocytes are activated cells, with increased inflammatory activity and ability to interact with endothelial cells. Therefore, accumulation of senescent monocytes may explain, in part, the development of chronic inflammation and atherosclerosis in elderly subjects and in patients with chronic inflammatory diseases.

  1. CD14++CD16+ monocytes and cardiovascular outcome in patients with chronic kidney disease.

    PubMed

    Rogacev, Kyrill S; Seiler, Sarah; Zawada, Adam M; Reichart, Birgit; Herath, Esther; Roth, Daniel; Ulrich, Christof; Fliser, Danilo; Heine, Gunnar H

    2011-01-01

    Patients with chronic kidney disease (CKD) pose a worldwide growing burden to health care systems due to accelerated atherosclerosis and subsequent high cardiovascular (CV) morbidity. Atherogenesis is prominently driven by monocytes and monocyte-derived macrophages. The expression of CD14 and CD16 characterizes three monocyte subsets: CD14(++)CD16(-), CD14(++)CD16(+), and CD14((+))CD16(+) cells; the latter two are often denoted as 'proinflammatory' CD16(+) monocytes. Despite an association between CD16(+) monocyte counts and higher CV risk in cross-sectional cohorts, the prognostic impact of elevated CD16(+) monocyte counts is poorly understood. We assessed monocyte heterogeneity using flow cytometry in 119 patients with non-dialysis CKD, who were prospectively followed for a median of 4.9 (inter-quartile range 4.8-5.0) years for the occurrence of CV events. In addition, we assessed expression of chemokine receptors on monocyte subsets. CD14(++)CD16(+) monocyte were independently associated with CV events [hazard ratio (for an increase of 10 cells/μL) 1.26 (confidence interval: 1.04-1.52; P = 0.018)] after adjustment for variables that significantly affected CD14(++)CD16(+) cell counts at baseline. Across the spectrum of CKD, CD14(++)CD16(+) monocytes selectively expressed CCR5. We found that CD14(++)CD16(+) monocytes were independently associated with CV events in non-dialysis CKD patients. Our results support the notion that CD16(+) monocytes rather than CD16(-) monocytes are involved in human atherosclerosis.

  2. Development of a Quantitative Bead Capture Assay for Soluble IL-7 Receptor Alpha in Human Plasma

    PubMed Central

    Faucher, Sylvie; Crawley, Angela M.; Decker, Wendy; Sherring, Alice; Bogdanovic, Dragica; Ding, Tao; Bergeron, Michele; Angel, Jonathan B.; Sandstrom, Paul

    2009-01-01

    Background IL-7 is an essential cytokine in T-cell development and homeostasis. It binds to the IL-7R receptor, a complex of the IL-7Rα (CD127) and common γ (CD132) chains. There is significant interest in evaluating the expression of CD127 on human T-cells as it often decreased in medical conditions leading to lymphopenia. Previous reports showed the usefulness of CD127 as a prognostic marker in viral infections such as HIV, CMV, EBV and HCV. A soluble CD127 (sCD127) is released in plasma and may contribute to disease pathogenesis through its control on IL-7 activities. Measuring sCD127 is important to define its role and may complement existing markers used in lymphopenic disease management. We describe a new quantitative assay for the measurement of sCD127 in plasma and report sCD127 concentrations in healthy adults. Methodology/Principal Findings We developed a quantitative bead-based sCD127 capture assay. Polyclonal CD127-specific antibodies were chosen for capture and a biotinylated monoclonal anti-CD127 antibody was selected for detection. The assay can detect native sCD127 and recombinant sCD127 which served as the calibrator. The analytical performance of the assay was characterized and the concentration and stability of plasma sCD127 in healthy adults was determined. The assay's range was 3.2–1000 ng/mL. The concentration of plasma sCD127 was 164±104 ng/mL with over a log variation between subjects. Individual sCD127 concentrations remained stable when measured serially during a period of up to one year. Conclusions/Significance This is the first report on the quantification of plasma sCD127 in a population of healthy adults. Soluble CD127 plasma concentrations remained stable over time in a given individual and sCD127 immunoreactivity was resistant to repeated freeze-thaw cycles. This quantitative sCD127 assay is a valuable tool for defining the potential role of sCD127 in lymphopenic diseases. PMID:19690616

  3. Molecular Characterization and SNP Detection of CD14 Gene of Crossbred Cattle

    PubMed Central

    Pal, Aruna; Sharma, Arjava; Bhattacharya, T. K.; Chatterjee, P. N.; Chakravarty, A. K.

    2011-01-01

    CD14 is an important molecule for innate immunity that can act against a wide range of pathogens. The present paper has characterized CD14 gene of crossbred (CB) cattle (Bos indicus×Bos taurus). Cloning and sequence analysis of CD14 cDNA revealed 1119 nucleotide long open reading frame encoding 373 amino acids protein and 20 amino acids signal peptide. CB cattle CD14 gene exhibited a high percentage of nucleotide identity (59.3–98.1%) with the corresponding mammalian homologs. Cattle and buffalo appear to have diverged from a common ancestor in phylogenetic analysis. 25 SNPs with 17 amino acid changes were newly reported and the site for mutational hot-spot was detected in CB cattle CD14 gene. Non-synonymous substitutions exceeding synonymous substitutions indicate the evolution of this protein through positive selection among domestic animals. Predicted protein structures obtained from deduced amino acid sequence indicated CB cattle CD14 molecule to be a receptor with horse shoe-shaped structure. The sites for LPS binding, LPS signalling, leucine-rich repeats, putative N-linked glycosylation, O-linked glycosylation, glycosyl phosphatidyl inositol anchor, disulphide bridges, alpha helix, beta strand, leucine rich nuclear export signal, leucine zipper and domain linker were predicted. Most of leucine and cysteine residues remain conserved across the species. PMID:22132326

  4. Design of pervaporation membrane for organic-liquid separation based on solubility control by plasma-graft filling polymerization technique

    SciTech Connect

    Yamaguchi, Takeo; Nakao, Shinichi; Kimura, Shoji )

    1993-05-01

    Pervaporation performance through the membranes showed the same tendency as solubility results. The authors have prepared the filling-polymerized membrane for pervaporation of organic-liquid mixtures by the plasma-graft polymerization technique. The membrane is composed of two different polymers: a porous substrate which can suppress membrane swelling and a grafted polymer which forms in the pores of the substrate and exhibits selectivity due to its solubility. The objectives of the present study are to design a suitable membrane for an organic-mixture system by the control of the filling-polymer solubility. Specifically, a porous high-density polyethylene membrane and poly(methylacrylate/acrylamide) copolymer were employed as the porous substrate and grafted polymer, respectively, and grafted copolymer solubility was predicted by Hansen solubility parameters (HSP). The grafted polymer composition and its solubility behavior could be controlled by varying the monomer composition, and the solubility change was in accordance with the prediction by HSP. Pervaporation performance through the membranes showed the same tendency as solubility results. The authors concluded that an optimum pervaporation membrane can be designed on the basis of solubility control through use of these techniques for polymerization and prediction.

  5. Plasma Levels of Soluble Interleukin 1 Receptor Accessory Protein Are Reduced in Obesity

    PubMed Central

    Attard, Chantal; Kulkarni, Hemant; Cummings, Nik; Diego, Vincent P.; Carless, Melanie A.; Shields, Katherine A.; Johnson, Matthew P.; Kowlessur, Sudhir; Dyer, Thomas D.; Comuzzie, Anthony G.; Almasy, Laura; Zimmet, Paul; Moses, Eric K.; Göring, Harald H. H.; Curran, Joanne E.; Blangero, John; Jowett, Jeremy B. M.

    2014-01-01

    Context: Adipokines actuate chronic, low-grade inflammation through a complex network of immune markers, but the current understanding of these networks is incomplete. The soluble isoform of the IL-1 receptor accessory protein (sIL1RAP) occupies an important position in the inflammatory pathways involved in obesity. The pathogenetic and clinical influences of sIL1RAP are unknown. Objective: The objective of the study was to elucidate whether plasma levels of sIL1RAP are reduced in obesity, using affluent clinical, biochemical, and genetic data from two diverse cohorts. Design, Setting, and Participants: The study was conducted in two cohorts: the San Antonio Family Heart Study (n = 1397 individuals from 42 families) and South Asians living in Mauritius, n = 230). Main Outcome Measures: Plasma sIL1RAP levels were measured using an ELISA. The genetic basis of sIL1RAP levels were investigated using both a large-scale gene expression profiling study and a genome-wide association study. Results: A significant decrease in plasma sIL1RAP levels were observed in obese subjects, even after adjustment for age and sex. The sIL1RAP levels demonstrated a strong inverse association with obesity measures in both populations. All associations were more significant in females. Plasma sIL1RAP levels were significantly heritable, correlated with IL1RAP transcript levels (NM_134470), showed evidence for shared genetic influences with obesity measures and were significantly associated with the rs2885373 single-nucleotide polymorphism (P = 6.7 × 10−23) within the IL1RAP gene. Conclusions: Plasma sIL1RAP levels are reduced in obesity and can potentially act as biomarkers of obesity. Mechanistic studies are required to understand the exact contribution of sIL1RAP to the pathogenesis of obesity. PMID:24915116

  6. Plasma levels of soluble interleukin 1 receptor accessory protein are reduced in obesity.

    PubMed

    Bozaoglu, Kiymet; Attard, Chantal; Kulkarni, Hemant; Cummings, Nik; Diego, Vincent P; Carless, Melanie A; Shields, Katherine A; Johnson, Matthew P; Kowlessur, Sudhir; Dyer, Thomas D; Comuzzie, Anthony G; Almasy, Laura; Zimmet, Paul; Moses, Eric K; Göring, Harald H H; Curran, Joanne E; Blangero, John; Jowett, Jeremy B M

    2014-09-01

    Adipokines actuate chronic, low-grade inflammation through a complex network of immune markers, but the current understanding of these networks is incomplete. The soluble isoform of the IL-1 receptor accessory protein (sIL1RAP) occupies an important position in the inflammatory pathways involved in obesity. The pathogenetic and clinical influences of sIL1RAP are unknown. The objective of the study was to elucidate whether plasma levels of sIL1RAP are reduced in obesity, using affluent clinical, biochemical, and genetic data from two diverse cohorts. The study was conducted in two cohorts: the San Antonio Family Heart Study (n = 1397 individuals from 42 families) and South Asians living in Mauritius, n = 230). Plasma sIL1RAP levels were measured using an ELISA. The genetic basis of sIL1RAP levels were investigated using both a large-scale gene expression profiling study and a genome-wide association study. A significant decrease in plasma sIL1RAP levels were observed in obese subjects, even after adjustment for age and sex. The sIL1RAP levels demonstrated a strong inverse association with obesity measures in both populations. All associations were more significant in females. Plasma sIL1RAP levels were significantly heritable, correlated with IL1RAP transcript levels (NM_134470), showed evidence for shared genetic influences with obesity measures and were significantly associated with the rs2885373 single-nucleotide polymorphism (P = 6.7 × 10(-23)) within the IL1RAP gene. Plasma sIL1RAP levels are reduced in obesity and can potentially act as biomarkers of obesity. Mechanistic studies are required to understand the exact contribution of sIL1RAP to the pathogenesis of obesity.

  7. CD14, CD16 and HLA-DR reliably identifies human monocytes and their subsets in the context of pathologically reduced HLA-DR expression by CD14(hi) /CD16(neg) monocytes: Expansion of CD14(hi) /CD16(pos) and contraction of CD14(lo) /CD16(pos) monocytes in acute liver failure.

    PubMed

    Abeles, Robin Daniel; McPhail, Mark J; Sowter, David; Antoniades, Charalambos G; Vergis, Nikhil; Vijay, Godhev K Manakkat; Xystrakis, Emmanuel; Khamri, Wafa; Shawcross, Debbie L; Ma, Yun; Wendon, Julia A; Vergani, Diego

    2012-10-01

    Changes in monocytes and their subsets (CD14(hi)/CD16(neg), CD14(hi)/CD16(pos) and CD14(lo)/CD16(pos)) have been described in several diseases. The combination of CD14, CD16 and HLA-DR has been suggested to discriminate monocytes from the CD16(pos) /HLA-DR(neg) NK-cells and neutrophils but no data exist whether this strategy can be used in situations when monocyte HLA-DR expression is pathologically reduced. Monocytes and their subsets were concurrently identified through negative (exclusion of CD66b(pos) neutrophils, CD56(pos) NKcells, CD19(pos) B-cells, and CD3(pos) T-cells) and positive gating (inclusion of monocytes by expression of CD14, CD16, and HLA-DR) strategies on 30 occasions [9 healthy controls (HC) and 21 patients with conditions associated with low monocyte HLA-DR expression]. Bland-Altman and Passing and Bablok regression statistics did not demonstrate any significant measurement bias between the two strategies of monocyte identification. Monocyte subset phenotype was then compared in 18 HC and 41 patients with acute liver failure (ALF). Compared with HC, in ALF, the percentage of CD14(hi)/CD16(pos) monocytes was higher (7% vs 4%) whilst the percentage of CD14(lo)/CD16(pos) was lower (1.9% vs. 7%) (P ≤ 0.001); HLA-DR and CD86 MFIs on all monocyte subsets were lower, whilst CCR5, CD64, and CD11b MFIs were higher (P < 0.05). The relative expression by monocyte subsets of HLA-DR, CCR2, CCR5, CX3CR1, and CD11a was similar in ALF patients and HCs. Repeat analysis of an identical antibody-fluorochrome "backbone" targeting HLA-DR, CD14, and CD16 was assessed in 189 samples across 5 different experiments. There was excellent agreement in the results obtained using the positive gating strategy (interclass correlation coefficients > 0.8). Monocytes and their subsets can be reliably identified using an antibody-fluorochrome "backbone" of HLA-DR, CD14, and CD16. CD16(pos) monocytes continue to constitutively express HLA-DR even in conditions where HLA-DR is

  8. Soluble CD44: quantification and molecular repartition in plasma of patients with colorectal cancer

    PubMed Central

    Masson, D; Denis, M G; Denis, M; Blanchard, D; Loirat, M J; Cassagnau, E; Lustenberger, P

    1999-01-01

    Based on the important role of CD44 in tumour progression and metastasis, we evaluated, in a prospective study, plasma-soluble CD44 (sCD44) as a serum marker in colorectal cancer. Blood plasma specimens from 89 patients with colorectal neoplasm, 22 patients with a gastrointestinal disease and 23 healthy donors were analysed for quantitation (ELISA assay) and purification of sCD44. The concentration of sCD44, indicating the concentration of all isoforms, was significantly higher in patients with colorectal cancer and intestinal disease than in normal individuals, but no significant differences were found between the two groups. We found no association between plasma levels and staging of the colorectal cancer patients according to Astler and Coller. A two-step batch purification combining ion exchange and immunoaffinity chromatography, followed by Western blot analysis, revealed a complex pattern with a major band corresponding to the standard form of CD44 and minor bands that may correspond to larger variant forms. No particular sCD44 isoform was clearly associated with anatomopathological or biological information. © 1999 Cancer Research Campaign PMID:10471052

  9. CD163+CD14+ macrophages, a potential immune biomarker for malignant pleural effusion.

    PubMed

    Wang, Fei; Yang, Li; Gao, Qun; Huang, Lan; Wang, Liping; Wang, Jing; Wang, Shengdian; Zhang, Bin; Zhang, Yi

    2015-08-01

    Malignant pleural effusion (MPE) is a common complication caused by malignant diseases. However, subjectivity, poor sensitivity, and substantial false-negative rates of cytology assay hamper accurate MPE diagnosis. The aim of this study was to assess whether CD163+CD14+ tumor-associated macrophages (TAMs) could be used as a biomarker for enabling sensitive and specific MPE diagnosis. Pleural effusion samples and peripheral blood samples were collected from 50 MPE patients and 50 non-malignant pleural effusion (NMPE) patients, respectively. Flow cytometry was performed to analyze cell phenotypes, and RT-qPCR was used to detect cytokine expression in these monocytes and macrophages. A blinded validation study (n = 40) was subsequently performed to confirm the significance of CD163+CD14+ TAMs in MPE diagnosis. Student's t test, rank sum test, and receiver operating characteristic curve analysis were used for statistical analysis. Notably, CD163+CD14+ cell frequency in MPE was remarkably higher than that in NMPE (P < 0.001). In a blinded validation study, a sensitivity of 78.9 % and a specificity of 100 % were obtained with CD163+CD14+ TAMs as a MPE biomarker. In total (n = 140), by using a cutoff level of 3.65 %, CD163+CD14+ cells had a sensitivity of 81.2 % and a specificity of 100 % for MPE diagnosis. Notably, MPE diagnosis by estimating CD163+CD14+ cells in pleural effusion could be obtained one week earlier than that obtained by cytological examination. CD163+CD14+ macrophages could be potentially used as an immune diagnostic marker for MPE and has better assay sensitivity than that of cytological analysis.

  10. Impact of Nonthermal Atmospheric Plasma on the Structure of Cellulose: Access to Soluble Branched Glucans.

    PubMed

    Delaux, Joakim; Ortiz Mellet, Carmen; Canaff, Christine; Fourré, Elodie; Gaillard, Cédric; Barakat, Abdellatif; García Fernández, José M; Tatibouët, Jean-Michel; Jérôme, François

    2016-11-07

    We have investigated the effect of non-thermal atmospheric plasma (NTAP) on the structure of microcrystalline cellulose. In particular, by means of different characterization methods, we demonstrate that NTAP promotes the partial cleavage of the β-1,4 glycosidic bond of cellulose leading to the release of short-chain cellodextrins that are reassembled in situ, preferentially at the C6 position, to form branched glucans with either a glucosyl or anhydroglucosyl terminal residue. The ramification of cellulosic chain induced by NTAP yields branched glucans that are soluble in DMSO or in water, thus opening a straightforward access to processable glucans from cellulose. Importantly, the absence of solvent and catalyst considerably facilitates downstream processing as compared to (bio)catalytic processes which typically occur in diluted conditions.

  11. Contaminating fibrin in CPD-blood: solubility in plasma and distribution in blood components following separation

    SciTech Connect

    Skjonsberg, O.H.; Kierulf, P.; Gravem, K.; Fagerhol, M.K.; Godal, H.C.

    1986-01-01

    In order to estimate the solubility of contaminating fibrin in CPD-blood, thrombin induced fibrin polymerzation in CPD-plasma was examined by light scattering and fibrinopeptide A (FPA) determinations. In addition, I-125 fibrin monomer enriched CPD-blood was used to investigate fibrin monomer retention in blood bags and transfusion filters (170 microns) and fibrin distribution in blood components derived from CPD-blood. Initial fibrin polymerization in CPD-blood occurred after conversion of 15 per cent of the fibrinogen to fibrin, implying that substantial amounts of fibrin may be kept solubilized in CPD-blood bags. Only minor amounts of I-125 fibrin monomers were retained in blood bags (2.4 per cent) and in transfusion filters (2.9 per cent) after sham transfusions. After separating I-125-fibrin monomer enriched CPD-blood into its constituent components, the major part of fibrin (75.0 per cent) could be traced in the cryoprecipitate.

  12. Interaction of Globular Plasma Proteins with Water-Soluble CdSe Quantum Dots.

    PubMed

    Pathak, Jyotsana; Rawat, Kamla; Sanwlani, Shilpa; Bohidar, H B

    2015-06-08

    The interactions between water-soluble semiconductor quantum dots [hydrophilic 3-mercaptopropionic acid (MPA)-coated CdSe] and three globular plasma proteins, namely, bovine serum albumin (BSA), β-lactoglobulin (β-Lg) and human serum albumin (HSA), are investigated. Acidic residues of protein molecules form electrostatic interactions with these quantum dots (QDs). To determine the stoichiometry of proteins bound to QDs, we used dynamic light scattering (DLS) and zeta potential techniques. Fluorescence resonance energy transfer (FRET) experiments revealed energy transfer from tryptophan residues in the proteins to the QD particles. Quenching of the intrinsic fluorescence of protein molecules was noticed during this binding process (hierarchy HSA<β-Lg

  13. The classical CD14⁺⁺ CD16⁻ monocytes, but not the patrolling CD14⁺ CD16⁺ monocytes, promote Th17 responses to Candida albicans.

    PubMed

    Smeekens, Sanne P; van de Veerdonk, Frank L; Joosten, Leo A B; Jacobs, Liesbeth; Jansen, Trees; Williams, David L; van der Meer, Jos W M; Kullberg, Bart Jan; Netea, Mihai G

    2011-10-01

    In the present study, we investigated the functional differences between cluster of differentiation (CD)14(++) CD16(-) and CD14(+) CD16(+) monocytes during anti-Candida host defense. CD14(++) CD16(-) are the "classical" monocytes and represent the majority of circulating monocytes in humans, while CD14(+) CD16(+) monocytes patrol the vasculature for maintenance of tissue integrity and repair. Both monocyte subsets inhibited the germination of live Candida albicans, and there was no difference in their capacity to phagocytose and kill Candida. Although production of IL-6 and IL-10 induced by C. albicans was found to be similar between monocyte subsets, IL-1β and prostaglandin E2 (PGE2) production was higher in CD14(++) CD16(-) compared with CD14(+) CD16(+) monocytes. In line with the increased production of IL-1β and PGE2, central mediators for inducing Th17 responses, CD14(++) CD16(-) monocytes induced greater Th17 responses upon stimulation with heat-killed C. albicans yeast. The percentage of cells that expressed mannose receptor (MR) was higher in the CD14(++) CD16(-) monocyte subset, and MR-specific stimulation induced higher Th17 responses only in co-cultures of CD14(++) CD16(-) monocytes and CD4 lymphocytes. In conclusion, both monocyte subsets have potent innate antifungal properties, but only CD14(++) CD16(-) monocytes are capable of inducing a potent Th17 response to C. albicans, an important component of antifungal host defense.

  14. Cardiomyogenic potential of mesenchymal progenitors derived from human circulating CD14+ monocytes.

    PubMed

    Kodama, Hiroaki; Inoue, Takafumi; Watanabe, Ryuichi; Yasuoka, Hidekata; Kawakami, Yutaka; Ogawa, Satoshi; Ikeda, Yasuo; Mikoshiba, Katsuhiko; Kuwana, Masataka

    2005-12-01

    Previously, we reported a unique CD14+CD45+CD34+type I collagen+ cell fraction derived from human circulating CD14+ monocytes, named monocyte-derived mesenchymal progenitors (MOMPs). These primitive cells differentiate along mesenchymal lineages, including bone, cartilage, fat, and skeletal muscle. Here, we demonstrate that CD14+ monocytes generate MOMPs that differentiate into cardiomyocytes. MOMPs labeled with a fluorescent marker and co-cultivated with rat cardiomyocytes for 4 weeks expressed the cardiomyocyte-specific transcription factors Nkx2.5, GATA-4, eHAND, and MEF2 and the hematopoietic/monocytic markers CD45 and CD14 within 10 days and retained their proliferative capacity for up to 16 days. A subpopulation of MOMPs subsequently expressed the cardiomyocyte-specific markers micro-sarcomeric actinin, troponin I, and atrial natriuretic peptide on day 21. Furthermore, fluorescence-labeled, spontaneously beating cells that formed gap junctions with adjacent rat cardiomyocytes appeared in these cultures and these cells exhibited electrophysiological properties typical of ventricular myocytes. The co-cultivation of human MOMPs with rat GFP-tagged cardiomyocytes resulted in the generation of human cardiomyocytes lacking green fluorescent protein (GFP) staining, suggesting that our observations could not solely be explained by cell fusion. Our results demonstrate for the first time that human circulating CD14+ monocytes include progenitors capable of proliferating and differentiating along the cardiomyogenic lineage via their differentiation into MOMPs.

  15. The "Intermediate" CD14(++)CD16(+) monocyte subset increases in severe peripheral artery disease in humans.

    PubMed

    Wildgruber, Moritz; Aschenbrenner, Teresa; Wendorff, Heiko; Czubba, Maria; Glinzer, Almut; Haller, Bernhard; Schiemann, Matthias; Zimmermann, Alexander; Berger, Hermann; Eckstein, Hans-Henning; Meier, Reinhard; Wohlgemuth, Walter A; Libby, Peter; Zernecke, Alma

    2016-12-19

    Monocytes are key players in atherosclerotic. Human monocytes display a considerable heterogeneity and at least three subsets can be distinguished. While the role of monocyte subset heterogeneity has already been well investigated in coronary artery disease (CAD), the knowledge about monocytes and their heterogeneity in peripheral artery occlusive disease (PAOD) still is limited. Therefore, we aimed to investigate monocyte subset heterogeneity in patients with PAOD. Peripheral blood was obtained from 143 patients suffering from PAOD (Rutherford stage I to VI) and three monocyte subsets were identified by flow cytometry: CD14(++)CD16(-) classical monocytes, CD14(+)CD16(++) non-classical monocytes and CD14(++)CD16(+) intermediate monocytes. Additionally the expression of distinct surface markers (CD106, CD162 and myeloperoxidase MPO) was analyzed. Proportions of CD14(++)CD16(+) intermediate monocyte levels were significantly increased in advanced stages of PAOD, while classical and non-classical monocytes displayed no such trend. Moreover, CD162 and MPO expression increased significantly in intermediate monocyte subsets in advanced disease stages. Likewise, increased CD162 and MPO expression was noted in CD14(++)CD16(-) classical monocytes. These data suggest substantial dynamics in monocyte subset distributions and phenotypes in different stages of PAOD, which can either serve as biomarkers or as potential therapeutic targets to decrease the inflammatory burden in advanced stages of atherosclerosis.

  16. CD14 Signaling Restrains Chronic Inflammation through Induction of p38-MAPK/SOCS-Dependent Tolerance

    PubMed Central

    Sahay, Bikash; Patsey, Rebeca L.; Eggers, Christian H.; Salazar, Juan C.; Radolf, Justin D.; Sellati, Timothy J.

    2009-01-01

    Current thinking emphasizes the primacy of CD14 in facilitating recognition of microbes by certain TLRs to initiate pro-inflammatory signaling events and the importance of p38-MAPK in augmenting such responses. Herein, this paradigm is challenged by demonstrating that recognition of live Borrelia burgdorferi not only triggers an inflammatory response in the absence of CD14, but one that is, in part, a consequence of altered PI3K/AKT/p38-MAPK signaling and impaired negative regulation of TLR2. CD14 deficiency results in increased localization of PI3K to lipid rafts, hyperphosphorylation of AKT, and reduced activation of p38. Such aberrant signaling leads to decreased negative regulation by SOCS1, SOCS3, and CIS, thereby compromising the induction of tolerance in macrophages and engendering more severe and persistent inflammatory responses to B. burgdorferi. Importantly, these altered signaling events and the higher cytokine production observed can be mimicked through shRNA and pharmacological inhibition of p38 activity in CD14-expressing macrophages. Perturbation of this CD14/p38-MAPK-dependent immune regulation may underlie development of infectious chronic inflammatory syndromes. PMID:20011115

  17. Expansion of CD14+CD16+ monocytes is related to acute leukemia

    PubMed Central

    Jiang, Xin-Quan; Zhang, Lei; Liu, Hong-Ai; Yuan, Ning; Hou, Pei-Qiang; Zhang, Rong-Qiang; Wu, Tuo

    2015-01-01

    Background: Objective: Aim to investigate the proportion of CD14+CD16+ monocytes and understand the pathogenesis of this monocyte subset in acute leukemia. Methods: Flow cytometry was utilized to study the phenotype expression of CD14+CD16+ monocytes and CD3+ T lymphocytes in peripheral blood derived from patients with acute leukemia. All the data were analyzed by SPSS 13.0 software. Results: The proportion of CD14+CD16+ monocytes including both intermediate and non-classical monocytes, increased significantly in patients with acute leukemia and changed negatively or positively according to the disease process. Meanwhile, the proportion of CD14+CD16+ monocytes was inversely correlated with absolute number of CD4+ T lymphocytes, ratio of CD4+/CD8+ T cells, and positively correlated with the proportion of neutrophil granulocytes. Conclusions: The proportion of CD14+CD16+ monocytes (especially the intermediate subpopulation) is related to the progression of acute leukemia, and the expansion of this monocyte subset could indicate the severity of the disease. PMID:26550139

  18. Plasma soluble thrombomodulin levels are associated with mortality in the acute respiratory distress syndrome

    PubMed Central

    Calfee, Carolyn S.; Liu, Kathleen D.; Kangelaris, Kirsten; Hansen, Helen; Pawlikowska, Ludmila; Ware, Lorraine B.; Alkhouli, Mustafa F.; Abbot, Jason; Matthay, Michael A.

    2015-01-01

    Objective Thombomodulin (TM) is an activator of protein C and a biomarker for endothelial injury. We hypothesized that (1) elevated plasma levels would be associated with clinical outcomes and (2) polymorphisms in the TM gene would be associated with plasma levels. Patients We studied 449 patients enrolled in the Fluid and Catheter Treatment Trial (FACTT) for whom both plasma and DNA were available. We used logistic regression and receiver operator curves (ROC) to test for associations between soluble TM (sTM) and mortality at 60 days. Measurements and results Plasma sTM levels were higher in non-survivors than survivors at baseline [median 147 (IQR, 95–218) vs. 89 (56–129) ng/mL, p < 0.0001] and on day 3 after study enrollment [205 (146–302) vs. 127 (85–189), p < 0.0001]. The odds of death increased by 2.4 (95 %CI 1.5–3.8, p < 0.001), and by 2.8 (1.7–4.7, P < 0.001) for every log increase in baseline and day 3 sTM levels, respectively, after adjustment for age, race, gender, severity of illness, fluid management strategy, baseline creatinine, and non-pulmonary sepsis as the primary cause of ARDS. By ROC analysis, plasma sTM levels discriminated between non-survivors and survivors [AUC = 72 % (66–78 %) vs. AUC = 54 % for severity based on Berlin criteria). Addition of sTM improved discrimination based on APACHE III from 77 to 80 % (P < 0.03). sTM levels at baseline were not statistically different among subjects stratified by genotypes of tag SNPs in the TM gene. Conclusions Higher plasma sTM levels are associated with increased mortality in ARDS. The lack of association between the sTM levels and genetic variants suggests that the increased levels of sTM may reflect severity of endothelial damage rather than genetic heterogeneity. These findings underscore the importance of endothelial injury in ARDS pathogenesis and suggest that, in combination with clinical markers, sTM could contribute to risk stratification. PMID:25643902

  19. α1-Acid Glycoprotein Up-regulates CD163 via TLR4/CD14 Protein Pathway

    PubMed Central

    Komori, Hisakazu; Watanabe, Hiroshi; Shuto, Tsuyoshi; Kodama, Azusa; Maeda, Hitoshi; Watanabe, Kenji; Kai, Hirofumi; Otagiri, Masaki; Maruyama, Toru

    2012-01-01

    CD163, a scavenger receptor that is expressed at high levels in the monocyte-macrophage system, is a critical factor for the efficient extracellular hemoglobin (Hb) clearance during hemolysis. Because of the enormous detrimental effect of liberated Hb on our body by its ability to induce pro-inflammatory signals and tissue damage, an understanding of the molecular mechanisms associated with CD163 expression during the acute phase response is a central issue. We report here that α1-acid glycoprotein (AGP), an acute phase protein, the serum concentration of which is elevated under various inflammatory conditions, including hemolysis, up-regulates CD163 expression in both macrophage-like differentiated THP-1 (dTHP-1) cells and peripheral blood mononuclear cells in a time- and concentration-dependent manner. Moreover, the subsequent induction of Hb uptake was also observed in AGP-treated dTHP-1 cells. Among representative acute phase proteins such as AGP, α1-antitrypsin, C-reactive protein, and haptoglobin, only AGP increased CD163 expression, suggesting that AGP plays a specific role in the regulation of CD163. Consistently, the physiological concentrations of AGP induced CD163, and the subsequent induction of Hb uptake as well as the reduction of oxidative stress in plasma were observed in phenylhydrazine-induced hemolytic model mice, confirming the in vivo role of AGP. Finally, AGP signaling through the toll-like receptor-4 (TLR4) and CD14, the common innate immune receptor complex that normally recognizes bacterial components, was identified as a crucial stimulus that induces the autocrine regulatory loops of IL-6 and/or IL-10 via NF-κB, p38, and JNK pathways, which leads to an enhancement in CD163 expression. These findings provide possible insights into how AGP exerts anti-inflammatory properties against hemolysis-induced oxidative stress. PMID:22807450

  20. Simultaneous chromatographic analysis of eight fat-soluble vitamins in plasma.

    PubMed

    Po, E S; Ho, J W; Gong, B Y

    1997-03-27

    A high-performance liquid chromatographic (HPLC) method for the simultaneous analysis of eight fat-soluble vitamins including A acid, retinol and retinal, vitamins D2 and D3 together with menadione (vitamin K3) is described. The eight vitamins extracted from plasma were analyzed by reversed-phase HPLC with UV detection at 245 nm. The mobile phase was composed of methanol and ethanol (85:15, v/v) with 0.1% triethylamine. The extraction efficiency of different solvents, namely, dichloromethane, benzene, methanol, ethanol, chloroform and hexane, with or without the addition of detergents was studied. There is no single solvent system that could extract all eight vitamins in one step. A recovery study of the vitamins was performed using rabbit plasma. An average recovery for individual vitamins was about 40% or more with the described extraction methods. The absorbance response was linear at the nanogram level. The present method may be broadly applied with alternative extraction methods for the eight vitamins from different sources.

  1. Association between arsenic exposure from drinking water and plasma levels of soluble cell adhesion molecules.

    PubMed

    Chen, Yu; Santella, Regina M; Kibriya, Muhammad G; Wang, Qiao; Kappil, Maya; Verret, Wendy J; Graziano, Joseph H; Ahsan, Habibul

    2007-10-01

    Epidemiologic studies of cardiovascular disease risk factors and appropriate biomarkers in populations exposed to a wide range of arsenic levels are a public health research priority. We investigated the relationship between inorganic arsenic exposure from drinking water and plasma levels of soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular adhesion molecule-1 (sVCAM-1), both markers of endothelial dysfunction and vascular inflammation, in an arsenic-exposed population in Araihazar, Bangladesh. The study participants included 115 individuals with arsenic-related skin lesions participating in a 2 x 2 randomized, placebo-controlled, double-blind trial of vitamin E and selenium supplementation. Arsenic exposure status and plasma levels of sICAM-1 and sVCAM-1 were assessed at baseline and after 6 months of follow-up. Baseline well arsenic, a long-term measure of arsenic exposure, was positively associated with baseline levels of both sICAM-1 and sVCAM-1 and with changes in the two markers over time. At baseline, for every 1-mug/L increase in well arsenic there was an increase of 0.10 ng/mL [95% confidence interval (CI), 0.00-0.20] and 0.33 ng/mL (95% CI, 0.15-0.51) in plasma sICAM-1 and sVCAM-1, respectively. Every 1-microg/L increase in well arsenic was associated with a rise of 0.11 ng/mL (95% CI, 0.01-0.22) and 0.17 ng/mL (95% CI, 0.00-0.35) in sICAM-1 and sVCAM-1 from baseline to follow-up, respectively, in spite of recent changes in urinary arsenic as well as vitamin E and selenium supplementation during the study period. The findings indicate an effect of chronic arsenic exposure from drinking water on vascular inflammation that persists over time and also suggest a potential mechanism underlying the association between arsenic exposure and cardiovascular disease.

  2. Sex differences in soluble markers vary before and after the initiation of antiretroviral therapy in chronically HIV infected individuals

    PubMed Central

    KREBS, Shelly J.; SLIKE, Bonnie M.; SITHINAMSUWAN, Pasiri; ALLEN, Isabel E.; CHALERMCHAI, Thep; TIPSUK, Somporn; PHANUPHAK, Nittaya; JAGODZINSKI, Linda; KIM, Jerome H.; ANANWORANICH, Jintanat; MAROVICH, Mary A.; VALCOUR, Victor G.

    2016-01-01

    Objective To evaluate differences in soluble inflammatory markers between chronically HIV-infected men and women, with or without cognitive impairment, and in response to treatment. Design Soluble biomarkers were measured in cryopreserved plasma and cerebrospinal fluid (CSF) of 60 treatment-naïve individuals (25 males and 35 females) with chronic HIV infection and 18 HIV-uninfected controls (9 males and 9 females) from Thailand. Following enrollment, participants began combination antiretroviral therapy (cART) and were evaluated for expression of these markers after 48 weeks. Methods Plasma and CSF levels of 19 soluble biomarkers (IFN-γ, TNFα, TNF-RII, IL-1α, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-15, MCP-1, t-Tau, IP-10, neopterin, IFNα, I-FABP, and sCD14) were measured using either a multi-parameter or standard ELISA assay. Results Prior to cART, females with impaired cognition had elevated levels of neopterin and TNF-RII compared to females with normal cognition in both the plasma and CSF, however levels did not differ between cognitively impaired or normal males. In a secondary outcome-hypothesis generating analysis, sex differences were also pronounced in plasma levels of MCP-1, IL-10, I-FABP, and sCD14 in response to treatment. Neopterin, IP-10, TNFα, TNF-RII, IFNα, MCP-1, IL-8, I-FABP, and sCD14 plasma levels remained elevated following 48 weeks of therapy in both sexes compared to uninfected controls. Conclusions We provide evidence of sustained immune activation after 48 weeks of treatment and identify possible sex differences in biomarkers previously linked to cognitive impairment, chronic inflammation, and gut integrity that may contribute to immunological differences between sexes in relationship to disease progression and response to therapy. PMID:26990631

  3. Interaction between CD14 and LXRbeta genes modulates Alzheimer's disease risk.

    PubMed

    Rodríguez-Rodríguez, Eloy; Sánchez-Juan, Pascual; Mateo, Ignacio; Infante, Jon; Sánchez-Quintana, Coro; García-Gorostiaga, Inés; Berciano, José; Combarros, Onofre

    2008-01-15

    A chronic inflammatory process with activation of microglial cells contribute to the neurodegeneration associated with Alzheimer's disease (AD). CD14 and LXRbeta are receptors involved in the regulation of inflammatory responses of microglia in response to bacterial infection or lipopolysaccharide stimulation. In a case-control study in 266 AD patients and 273 healthy controls, we examined whether the combined gene effects between CD14 (-260) polymorphism and LXRbeta (intron 5) polymorphism might be responsible for susceptibility to AD. Subjects carrying both the CD14 (-260) C/C and the LXRbeta (intron 5) G/G genotypes had a six times lower risk of developing AD than subjects without these risk genotypes (OR 0.16, 95% CI 0.04-0.67, p=0.01). These data support a role for innate immune response genes in risk for AD.

  4. c-Maf-dependent growth of Mycobacterium tuberculosis in a CD14(hi) subpopulation of monocyte-derived macrophages.

    PubMed

    Dhiman, Rohan; Bandaru, Anuradha; Barnes, Peter F; Saha, Sudipto; Tvinnereim, Amy; Nayak, Ramesh C; Paidipally, Padmaja; Valluri, Vijaya Lakshmi; Rao, L Vijaya Mohan; Vankayalapati, Ramakrishna

    2011-02-01

    Macrophages are a major component of the innate immune response, comprising the first line of defense against various intracellular pathogens, including Mycobacterium tuberculosis. In this report, we studied the factors that regulate growth of M. tuberculosis H37Rv in subpopulations of human monocyte-derived macrophages (MDMs). In healthy donors, M. tuberculosis H37Rv grew 5.6-fold more rapidly in CD14(hi) MDMs compared with that in CD14(lo)CD16(+) MDMs. Compared with CD14(lo)CD16(+) cells, M. tuberculosis H37Rv-stimulated CD14(hi) monocytes produced more IL-10 and had increased mRNA expression for c-Maf, a transcription factor that upregulates IL-10 gene expression. c-Maf small interfering RNA (siRNA) inhibited IL-10 production and growth of M. tuberculosis in CD14(hi) cells. Compared with CD14(lo)CD16(+) monocytes, M. tuberculosis H37Rv-stimulated CD14(hi) cells had increased expression of 22 genes whose promoters contained a c-Maf binding site, including hyaluronan synthase 1 (HAS1). c-Maf siRNA inhibited HAS1 expression in M. tuberculosis-stimulated CD14(hi) monocytes, and HAS1 siRNA inhibited growth of M. tuberculosis in CD14(hi) MDMs. M. tuberculosis H37Rv upregulated expression of HAS1 protein and its product, hyaluronan, in CD14(hi) MDMs. We conclude that M. tuberculosis grows more rapidly in CD14(hi) than in CD14(lo)CD16(+) MDMs because CD14(hi) cells have increased expression of c-Maf, which increases production of two key factors (hyaluronan and IL-10) that promote growth of M. tuberculosis.

  5. Angiogenesis related genes NOS3, CD14, MMP3 and IL4R are associated to VEGF gene expression and circulating levels in healthy adults.

    PubMed

    Saleh, Abdelsalam; Stathopoulou, Maria G; Dadé, Sébastien; Ndiaye, Ndeye Coumba; Azimi-Nezhad, Mohsen; Murray, Helena; Masson, Christine; Lamont, John; Fitzgerald, Peter; Visvikis-Siest, Sophie

    2015-10-05

    Vascular endothelial growth factor (VEGF) plays a key role in angiogenesis. The aim was to assess the genetic connections between the angiogenesis-related NOS3, CD14, MMP3, IL4R, IL4 genes and VEGF expression and plasma levels. The associations between VEGF plasma levels with the polymorphisms of NOS3, CD14, MMP3, IL4R, and IL4 were assessed in 403 healthy unrelated adults. The epistatic and environmental interactions were explored, including four VEGF-related polymorphisms previously identified. The VEGF expression in peripheral blood mononuclear cells was quantified (n = 65) for the VEGF121, VEGF145, VEGF165, and VEGF189 isoforms. The polymorphism rs1799983 of NOS3 was associated with the sum of all VEGF isoforms mRNA levels (P = 0.032) and VEGF145 (P = 0.033). Rs1800779 of NOS3 interacted with rs3918226 of the same gene and with the rs2569190 of CD14 (P = 0.022, P = 0.042, respectively) for VEGF plasma levels. Other epistatic interactions included the rs1801275 of IL4R with the rs6921438 (VEGF-related variant) and rs3025058 of MMP3 (P = 0.042, P = 0.010 respectively) and the rs2569190 of CD14 with the rs3025058 of MMP3 (P = 0.0119). We also identified an interaction of rs1800779 with obesity, high-density lipoprotein cholesterol and triglycerides (P = 0.018, P = 0.005, P = 0.043, respectively) as well as the interaction of rs6921438 with hypertension (P = 0.028). Our findings indicated that genetic variants of NOS3, CD14, MMP3 and IL4R are implicated in the determination of VEGF expression and plasma levels. Thus, they support the hypothesis that in physiological conditions there are complex biological relationships between pathways (such as angiogenesis and inflammation), which are involved in the development of chronic diseases.

  6. Alteration in plasma testosterone levels in male mice lacking soluble epoxide hydrolase.

    PubMed

    Luria, Ayala; Morisseau, Christophe; Tsai, Hsing-Ju; Yang, Jun; Inceoglu, Bora; De Taeye, Bart; Watkins, Steven M; Wiest, Michelle M; German, J Bruce; Hammock, Bruce D

    2009-08-01

    Soluble epoxide hydrolase (Ephx2, sEH) is a bifunctional enzyme with COOH-terminal hydrolase and NH(2)-terminal phosphatase activities. sEH converts epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids (DHETs), and the phosphatase activity is suggested to be involved in cholesterol metabolism. EETs participate in a wide range of biological functions, including regulation of vascular tone, renal tubular transport, cardiac contractility, and inflammation. Inhibition of sEH is a potential approach for enhancing the biological activity of EETs. Therefore, disruption of sEH activity is becoming an attractive therapeutic target for both cardiovascular and inflammatory diseases. To define the physiological role of sEH, we characterized a knockout mouse colony lacking expression of the Ephx2 gene. Lack of sEH enzyme is characterized by elevation of EET to DHET ratios in both the linoleate and arachidonate series in plasma and tissues of both female and male mice. In male mice, this lack of expression was also associated with decreased plasma testosterone levels, sperm count, and testicular size. However, this genotype was still able to sire litters. Plasma cholesterol levels also declined in this genotype. Behavior tests such as anxiety-like behavior and hedonic response were also examined in Ephx2-null and WT mice, as all can be related to hormonal changes. Null mice showed a level of anxiety with a decreased hedonic response. In conclusion, this study provides a broad biochemical, physiological, and behavioral characterization of the Ephx2-null mouse colony and suggests a mechanism by which sEH and its substrates may regulate circulating levels of testosterone through cholesterol biosynthesis and metabolism.

  7. Immunosuppressive CD14+HLA-DRlow/− Monocytes in Prostate Cancer

    PubMed Central

    Vuk-Pavlović, Stanimir; Bulur, Peggy A.; Lin, Yi; Qin, Rui; Szumlanski, Carol L.; Zhao, Xinghua; Dietz, Allan B.

    2010-01-01

    Objective To determine if the levels of circulating myeloid-derived suppressor cells increase with progression of prostate cancer (PCa); to determine if such cells could contribute to the relative inefficiency of PCa immunotherapy. Materials and Methods We analyzed peripheral blood mononuclear cells isolated from untreated PCa patients (uPCa; N = 18; mean age ± SD: 72.1 ± 6.9 years), tPCa (N = 22; 72.8 ± 9.8 years) and age matched controls (AMC; N = 12; 68.8 ± 7.5 years). We quantified surface marker phenotype, differentiation potential, effects on T cell proliferation and intracellular cytokines. Results We observed an unexpectedly high percentage of a type of myeloid-derived suppressor cells, CD14+HLA-DRlow/− monocytes, in tPCa (30.7 ± 15.0% of CD14+ cells) relative to AMC (4.1 ± 6.5%, P < 0.0001) and uPCa (10.6 ± 14.3%, P = 0.0001). The levels of CD14+ HLA-DRlow/− cells were significantly correlated with circulating PSA levels and treatment with LHRH-agonist leuprolide in combination with either an antiandrogen or dexamethasone. Monocytes from tPCa inhibited autologous T cell proliferation statistically significantly more effectively than AMC monocytes and were defective in their ability to differentiate into phenotypically mature dendritic cells. Isolated CD14+HLA-DRlow/− cells expressed higher levels of intracellular interleukin-10 and suppressed T cell proliferation more effectively than isolated CD14+HLA-DR+ cells. Conclusions This is the first report of CD14+ cells exhibiting reduced expression of HLA-DR molecules in PCa patients. These cells suppress immune cell function in vitro and, plausibly, in vivo, a finding that must be factored into the design of immunotherapy protocols for PCa patients. PMID:19902470

  8. Equations for O2 and CO2 solubilities in saline and plasma: combining temperature and density dependences.

    PubMed

    Christmas, Kevin M; Bassingthwaighte, James B

    2017-05-01

    Solubilities of respiratory gasses in water, saline, and plasma decrease with rising temperatures and solute concentrations. Henry's Law, C = α·P, states that the equilibrium concentration of a dissolved gas is solubility times partial pressure. Solubilities in the water of a solution depend on temperature and the content of other solutes. Blood temperatures may differ more than 20°C between skin and heart, and an erythrocyte will undergo that range as blood circulates. The concentrations of O2 and CO2 are the driving forces for diffusion, exchanges, and for reactions. We provide an equation for O2 and CO2 solubilities, α, that allows for continuous changes in temperature, T, and solution density, ρ, in dynamically changing states:[Formula: see text]This two-exponential expression with a density scalar γ, and a density exponent β, accounts for solubility changes due to density changes of an aqueous solution. It fits experimental data on solubilities in water, saline, and plasma over temperatures from 20 to 40°C, and for plasma densities, ρsol up to 1.020 g/ml with ~0.3% error. The amounts of additional bound O2 (to Hb) and CO2 (bicarbonate and carbamino) depend on the concentrations in the local water space and the reaction parameters. During exercise, solubility changes are large; both ρsol and T change rapidly with spatial position and with time. In exercise hemoconcentration plasma, ρsol exceeds 1.02, whereas T may range over 20°C. The six parameters for O2 and the six for CO2 are constants, so solubilities are calculable continuously as T and ρsol change.NEW & NOTEWORTHY Solubilities for oxygen and carbon dioxide are dependent on the density of the solution, on temperature, and on the partial pressure. We provide a brief equation suitable for hand calculators or mathematical modeling, accounting for these factors over a wide range of temperatures and solution densities for use in rapidly changing conditions, such as extreme exercise or osmotic

  9. CD14(+)CD16(++) "nonclassical" monocytes are associated with endothelial dysfunction in patients with coronary artery disease.

    PubMed

    Urbanski, Karol; Ludew, Dominik; Filip, Grzegorz; Filip, Magdalena; Sagan, Agnieszka; Szczepaniak, Piotr; Grudzien, Grzegorz; Sadowski, Jerzy; Jasiewicz-Honkisz, Barbara; Sliwa, Tomasz; Kapelak, Boguslaw; McGinnigle, Eilidh; Mikolajczyk, Tomasz; Guzik, Tomasz J

    2017-02-23

    Endothelial dysfunction and inflammation are key mechanisms of vascular disease. We hypothesised that heterogeneity of monocyte subpopulations may be related to the development of vascular dysfunction in coronary artery disease (CAD). Therefore, we examined the relationships between monocyte subsets (CD14(++)CD16(-) "classical - Mon1", CD14(++)CD16(+) "intermediate - Mon2" and CD14(+)CD16(++) "nonclassical - Mon3"), endothelial function and risk factor profiles in 130 patients with CAD undergoing coronary artery bypass grafting. This allowed for direct nitric oxide (NO) bioavailability assessment using isometric tension studies ex vivo (acetylcholine; ACh- and sodium-nitropruside; SNP-dependent) in segments of internal mammary arteries. The expression of CD14 and CD16 antigens and activation markers were determined in peripheral blood mononuclear cells using flow cytometry. Patients with high CD14(+)CD16(++) "nonclassical" and low CD14(++)CD16(-) "classical" monocytes presented impaired endothelial function. High frequency of CD14(+)CD16(++) "nonclassical" monocytes was associated with increased vascular superoxide production. Furthermore, endothelial dysfunction was associated with higher expression of activation marker CD11c selectively on CD14(+)CD16(++) monocytes. Nonclassical and classical monocyte frequencies remained independent predictors of endothelial dysfunction when major risk factors for atherosclerosis were taken into account (β=0.18 p=0.04 and β=-0.19 p=0.03, respectively). In summary, our data indicate that CD14(+)CD16(++) "nonclassical" monocytes are associated with more advanced vascular dysfunction measured as NO- bioavailability and vascular reactive oxygen species production.

  10. Highly Active Antiretroviral Therapy (HAART)-Related Hypertriglyceridemia Is Associated With Failure of Recovery of CD14lowCD16+ Monocyte Subsets in AIDS Patients.

    PubMed

    Han, Junyan; Zhao, Hongxin; Ma, Yaluan; Zhou, Haiwei; Hao, Yu; Li, Yanmei; Song, Chuan; Han, Ning; Liu, Xiangyi; Zeng, Hui; Qin, Mingzhao

    2015-07-01

    As cellular reservoirs, CD16 monocyte subsets play important roles in the progression of HIV infection. Previous studies have shown that highly active antiretroviral therapy (HAART) reduced the percentages of CD14CD16 monocyte subsets, but did not recover the percentages of CD14CD16 subsets. Eighty-four chronic HIV-infected, HAART-naïve individuals and 55 HIV-negative subjects (31 without hyperlipidemia and 24 with hypertriglyceridemia) were enrolled. Plasma HIV-1 RNA levels, CD4 T-cell counts, triglycerides, total cholesterol, high-density lipoprotein, and low-density lipoprotein were followed up for 48 weeks during HAART treatment in the longitudinal study. We found that mild hypertriglyceridemia in HIV-negative subjects and HIV-infected patients, naïve to HAART, did not affect the percentage of monocyte subsets. However, a failure of CD14CD16 subset recovery was observed in patients with HAART-related hypertriglyceridemia at 48 weeks. Thus, HAART-related hypertriglyceridemia altered homeostasis of monocyte subsets to antiviral therapy, which might further affect immune reconstitution.

  11. Highly Active Antiretroviral Therapy (HAART)-Related Hypertriglyceridemia Is Associated With Failure of Recovery of CD14lowCD16+ Monocyte Subsets in AIDS Patients

    PubMed Central

    Han, Junyan; Zhao, Hongxin; Ma, Yaluan; Zhou, Haiwei; Hao, Yu; Li, Yanmei; Song, Chuan; Han, Ning; Liu, Xiangyi; Zeng, Hui; Qin, Mingzhao

    2015-01-01

    Abstract As cellular reservoirs, CD16+ monocyte subsets play important roles in the progression of HIV infection. Previous studies have shown that highly active antiretroviral therapy (HAART) reduced the percentages of CD14highCD16+ monocyte subsets, but did not recover the percentages of CD14lowCD16+ subsets. Eighty-four chronic HIV-infected, HAART-naïve individuals and 55 HIV-negative subjects (31 without hyperlipidemia and 24 with hypertriglyceridemia) were enrolled. Plasma HIV-1 RNA levels, CD4+ T-cell counts, triglycerides, total cholesterol, high-density lipoprotein, and low-density lipoprotein were followed up for 48 weeks during HAART treatment in the longitudinal study. We found that mild hypertriglyceridemia in HIV-negative subjects and HIV-infected patients, naïve to HAART, did not affect the percentage of monocyte subsets. However, a failure of CD14lowCD16+ subset recovery was observed in patients with HAART-related hypertriglyceridemia at 48 weeks. Thus, HAART-related hypertriglyceridemia altered homeostasis of monocyte subsets to antiviral therapy, which might further affect immune reconstitution. PMID:26166108

  12. Polymorphism of the CD14 and TLR4 genes and post-treatment apical periodontitis.

    PubMed

    Rôças, Isabela N; Siqueira, José F; Del Aguila, Camila A; Provenzano, José C; Guilherme, Bianca P S; Gonçalves, Lucio S

    2014-02-01

    This study investigated the association of CD14 -260C>T and TLR4 +896A>G gene polymorphisms with post-treatment apical periodontitis in Brazilian individuals. The study population consisted of 41 patients with post-treatment apical periodontitis and 42 individuals with root canal-treated teeth exhibiting healed/healing periradicular tissues (controls). All teeth had apical periodontitis lesions at the time of treatment, which was completed at least 1 year previously. Saliva was collected from the participants; DNA was extracted and used for CD14 and TLR4 genotyping using the polymerase chain reaction-restriction fragment length polymorphism approach and a real-time polymerase chain reaction TaqMan assay (Applied Biosystems, Foster City, CA), respectively. No specific genotype or allele of the CD14 and TLR4 genes or any combination thereof was positively associated with post-treatment apical periodontitis (P > .05). Data from the present study suggest that polymorphisms in the CD14 and TLR4 genes do not influence the response to endodontic treatment of teeth with apical periodontitis. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  13. Enhanced host immune recognition of E.coli causing mastitis in CD-14 transgenic mice.

    USDA-ARS?s Scientific Manuscript database

    Escherchia coli causes mastitis, an economically significant disease in dairy animals. E. coli endotoxin (lipopolysaccharide, LPS) when bound by host membrane proteins such as CD-14, causes release of pro-inflammatory cytokines recruiting neutrophils as a early innate immune response. Excessive pr...

  14. Elevation of plasma-soluble HLA-G in childhood nephrotic syndrome is associated with IgE.

    PubMed

    Liu, Yanqing; Lai, Meimei; Lou, Yunyan; Han, Qiuyue; Yang, Qing; Chen, Minguang; Li, Jingbo; Wang, Huiyan; Yan, Weihua; Zheng, Xiaoqun

    2017-01-01

    Background Nephrotic syndrome is related to immune system dysfunction. Soluble human leukocyte antigen-G has been suggested to have an immunomodulatory role. Additionally, human leukocyte antigen-G expression may be influenced by the 14-base pair insertion/deletion polymorphism. However, this molecule has not been investigated in nephrotic syndrome. Methods Fifty-five children with nephrotic syndrome were enrolled: 24 primary nephrotic syndrome patients and 31 recurrent nephrotic syndrome patients. A group of 120 healthy subjects were included as reference controls. Additionally, 22 patients in nephrotic syndrome remission after treatments were also included. Both nephrotic syndrome patients and healthy subjects were genotyped for the 14-base pair insertion/deletion polymorphism. Plasma soluble human leukocyte antigen-G concentrations and serum immunoglobulin concentrations were determined. Results Nephrotic syndrome patients showed significantly higher levels of both soluble human leukocyte antigen-G and immunoglobulin E compared to normal controls. Nephrotic syndrome patients presented a higher frequency of the -14-base pair allele than did normal controls. Soluble human leukocyte antigen-G concentrations in remission patients were dramatically lower compared to in nephrotic syndrome patients. Moreover, soluble human leukocyte antigen-G and immunoglobulin E were moderately correlated in nephrotic syndrome patients. Conclusions The present study demonstrated that plasma soluble human leukocyte antigen-G concentrations were significantly elevated and that a relationship between serum total immunoglobulin E in nephrotic syndrome patients and the human leukocyte antigen-G -14-base pair allele may be a risk factor for nephrotic syndrome. These findings suggest that soluble human leukocyte antigen-G may be used as a monitoring marker for nephrotic syndrome patients' condition.

  15. Increased plasma soluble human leukocyte antigen-G in persistent wheezy infants.

    PubMed

    Tahan, Fulya; Eke Gungor, Hatice; Akar, Himmet Haluk; Saraymen, Berkay

    2017-05-01

    Human leukocyte antigen (HLA)-G is a non-classical major histocompatibility complex class I antigen characterized by limited polymorphism in its coding region, unique tissue expression pattern in physiologic conditions and immunomodulatory properties. Recently, the level of soluble (s)HLA-G was found to be higher in atopic asthma and allergic rhinitis, but this remains to be clarified in wheezy infants. The aim of the present study was therefore to investigate sHLA-G in wheezy infants. The subjects consisted of infants with persistent wheezing and positive modified asthma predictive index (mAPI; n = 30; persistent group) and those with transient wheezing and negative mAPI (n = 17; transient group). sHLA-G was measured in plasma using enzyme-linked immunosorbent assay. Total immunoglobulin E (IgE) and eosinophil count were measured, and skin testing was performed with a battery of 13 antigens with appropriate positive and negative controls. sHLA-G was significantly higher in the persistent wheezing (positive mAPI) group compared with the transient wheezing (negative mAPI) group (P = 0.008). There was no significant difference in peripheral blood eosinophil count and total IgE between the groups. The increased sHLA-G in infants with persistent wheeze suggests that sHLA-G may be able to be used to distinguish persistent from transient wheeze. Further comprehensive studies are needed on this topic. © 2016 Japan Pediatric Society.

  16. Soluble glycoprotein VI is raised in the plasma of patients with acute ischemic stroke.

    PubMed

    Al-Tamimi, Mohammad; Gardiner, Elizabeth E; Thom, Jim Y; Shen, Yang; Cooper, Matthew N; Hankey, Graeme J; Berndt, Michael C; Baker, Ross I; Andrews, Robert K

    2011-02-01

    Ischemic stroke induced by thrombosis may be triggered by atherosclerotic plaque rupture and collagen-induced platelet activation. Collagen induces glycoprotein VI shedding. We measured plasma-soluble glycoprotein VI (sGPVI) by enzyme-linked immunosorbent assay in 159 patients with acute (<7-day) ischemic stroke and age/sex-matched community-based control subjects. sGPVI was elevated in stroke compared with controls (P=0.0168). ORs were higher in Quartile 4 for stroke cases (P=0.0121), and sGPVI was significantly elevated in stroke associated with large artery disease across Quartiles 2 to 4 and small artery disease in Quartile 4. sGPVI decreased 3 to 6 months after antiplatelet treatment, consistent with elevated sGPVI due to platelet activation during the thrombotic event. sGPVI correlated with P-selectin (P=0.0007) and was higher in individuals with the GPVIa haplotype (P=0.024). Glycoprotein VI shedding is implicated in the pathology of acute ischemic stroke.

  17. No simple answers for the Finnish and Russian Karelia allergy contrast: Methylation of CD14 gene.

    PubMed

    Khoo, Siew-Kim; Mäkelä, Mika; Chandler, David; Schultz, En Nee; Jamieson, Sarra E; Goldblatt, Jack; Haahtela, Tari; LeSouëf, Peter; Zhang, Guicheng

    2016-11-01

    Finnish and Russian Karelian children have a highly contrasting occurrence of asthma and allergy. In these two environments, we studied associations between total serum immunoglobulin E (IgE) with methylation levels in cluster of differentiation 14 (CD14). Five hundred Finnish and Russian Karelian children were included in four groups: Finnish children with high IgE (n = 126) and low IgE (n = 124) as well as Russian children with high IgE (n = 125) and low IgE (n = 125). DNA was extracted from whole blood cells and pyrosequenced. Three CpG sites were selected in the promoter region of CD14. Methylation levels in two of the three CpG sites were higher in the Finnish compared to Russian Karelian children. In the promoter area of CD14, the Finnish compared to Russian children with low IgE had a significant (p < 0.0001) increase in methylation levels at the Amp5Site 2. Likewise, the Finnish compared to Russian children with high IgE had a significant (p = 0.003) increase in methylation levels at the Amp5Site 3. In Russian children with low vs. high IgE, there were significant differences in methylation levels, but this was not the case on the Finnish side. In the regression analysis, adding the methylation variation of CD14 to the model did not explain the higher asthma and allergy risk in the Finnish children. The methylation levels in the promoter region of CD14 gene were higher in the Finnish compared to Russian Karelian children. However, the methylation variation of this candidate gene did not explain the asthma and allergy contrast between these two areas. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. CD14, a Key Candidate Gene Associated with Specific Immune Response to Cockroach

    PubMed Central

    Gao, Peisong; Grigoryev, Dmitry N.; Rafaels, Nicholas M.; Mu, Deguang; Wright, Jerry M.; Cheadle, Christopher; Togias, Alkis; Beaty, Terri H.; Mathias, Rasika A.; Schroeder, John T.; Barnes, Kathleen C.

    2010-01-01

    Summary Background Sensitization to cockroach allergen is one of the strongest predictors of asthma morbidity, especially among African Americans. Objective Our aims were to determine the genomic basis of cockroach sensitization and the specific response to cockroach antigen. Methods We investigated the Th1/Th2 cytokine profile of co-cultured plasmacytoid DCs (pDCs) and CD4+ T cells and the “transcript signature” of the immune response to cockroach antigen using high-throughput expression profiling of co-cultured cells. Results We observed significantly elevated levels of IL-13, IL-10 and TNF-α, but undetectable levels of IL-12p70 and IFN-α, when cultures were exposed to crude cockroach antigen. A significant difference was observed for IL-13 between cockroach allergic and non-allergic individuals (p = 0.039). Microarray analyses demonstrated a greater response at 48 hours compared to 4 hours, with 50 genes being uniquely expressed in cockroach antigen-treated cells, including CD14, S100A8, CCL8, and IFI44L. The increased CD14 expression was further observed in purified pDCs, human monocytic THP-1 cells, and supernatant of co-cultured pDCs and CD4+ T cells in exposure to cockroach extract. Furthermore, the most differential expression of CD14 between cockroach allergy and non-cockroach allergy was only observed among individuals with the CC “high-risk” genotype of the CD14 -260C/T. Ingenuity Pathways Analysis (IPA) analyses suggested the interferon-signaling as the most significant canonical pathway. Conclusion Our results suggest these differentially expressed genes, particularly CD14, and genes in the interferon-signaling pathway may be important candidates for further investigation of their role in the immune response to cockroach allergen. PMID:20618347

  19. Elevated levels of peripheral blood CD14(bright) CD16+ and CD14(dim) CD16+ monocytes may contribute to the development of retinopathy in patients with juvenile onset type 1 diabetes.

    PubMed

    Ryba-Stanisławowska, Monika; Myśliwska, Jolanta; Juhas, Ulana; Myśliwiec, Małgorzata

    2015-09-01

    The study aimed to analyze the CD14(bright) CD16(+) and CD14(dim) CD16(+) monocyte subsets in juvenile-onset complication-free diabetes mellitus type 1 in the context of their association with microvascular complications. 61 children with type 1 diabetes and 30 healthy individuals were enrolled in a study. CD14(bright) CD16(+) and CD14(dim) CD16(+) monocytes were quantified in peripheral blood by means of flow cytometry. At the time of sampling blood glucose concentration was taken along with biochemical measurement of renal function, CRP and glycosylated hemoglobin. The Spearman's correlations were used to compare the relationship between CD16(+) monocyte subsets and the clinical parameters that can predict the development of microangiopathies. The flow cytometric analysis of monocyte subsets in peripheral blood of analyzed subjects revealed that the numbers of CD14(bright) CD16(+) and CD14(dim) CD16(+) monocytes were significantly higher in patients with type 1 diabetes than in the healthy individuals. As to the relationship between CD16(+) monocyte subsets and the clinical parameters that can predict development of microangiopathies, it was shown that both CD16(+) subsets were associated with increased risk of retinopathy development, defined as retinopathy development value. Elevated levels of intermediate CD14(bright) CD16(+) and non-classical CD14(dim) CD16(+) monocytes predict development of diabetic retinopathy in patients with type 1 diabetes.

  20. Directed Evolution of an LBP/CD14 Inhibitory Peptide and Its Anti-Endotoxin Activity

    PubMed Central

    Fang, Li; Xu, Zhi; Wang, Guan-song; Ji, Fu-yun; Mei, Chun-xia; Liu, Juan; Wu, Guo-ming

    2014-01-01

    Background LPS-binding protein (LBP) and its ligand CD14 are located upstream of the signaling pathway for LPS-induced inflammation. Blocking LBP and CD14 binding might prevent LPS-induced inflammation. In previous studies, we obtained a peptide analog (MP12) for the LBP/CD14 binding site and showed that this peptide analog had anti-endotoxin activity. In this study, we used in vitro directed evolution for this peptide analog to improve its in vivo and in vitro anti-endotoxin activity. Methods We used error-prone PCR (ep-PCR) and induced mutations in the C-terminus of LBP and attached the PCR products to T7 phages to establish a mutant phage display library. The positive clones that competed with LBP for CD14 binding was obtained by screening. We used both in vivo and in vitro experiments to compare the anti-endotoxin activities of a polypeptide designated P1 contained in a positive clone and MP12. Results 11 positive clones were obtained from among target phages. Sequencing showed that 9 positive clones had a threonine (T) to methionine (M) mutation in amino acid 287 of LBP. Compared to polypeptide MP12, polypeptide P1 significantly inhibited LPS-induced TNF-α expression and NF-κB activity in U937 cells (P<0.05). Compared to MP12, P1 significantly improved arterial oxygen pressure, an oxygenation index, and lung pathology scores in LPS-induced ARDS rats (P<0.05). Conclusion By in vitro directed evolution of peptide analogs for the LBP/CD14 binding site, we established a new polypeptide (P1) with a threonine (T)-to-methionine (M) mutation in amino acid 287 of LBP. This polypeptide had high anti-endotoxin activity in vitro and in vivo, which suggested that amino acid 287 in the C-terminus of LBP may play an important role in LBP binding with CD14. PMID:25025695

  1. The CD14+CD16+ inflammatory monocyte subset displays increased mitochondrial activity and effector function during acute Plasmodium vivax malaria.

    PubMed

    Antonelli, Lis R V; Leoratti, Fabiana M S; Costa, Pedro A C; Rocha, Bruno C; Diniz, Suelen Q; Tada, Mauro S; Pereira, Dhelio B; Teixeira-Carvalho, Andrea; Golenbock, Douglas T; Gonçalves, Ricardo; Gazzinelli, Ricardo T

    2014-09-01

    Infection with Plasmodium vivax results in strong activation of monocytes, which are important components of both the systemic inflammatory response and parasite control. The overall goal of this study was to define the role of monocytes during P. vivax malaria. Here, we demonstrate that P. vivax-infected patients display significant increase in circulating monocytes, which were defined as CD14(+)CD16- (classical), CD14(+)CD16(+) (inflammatory), and CD14loCD16(+) (patrolling) cells. While the classical and inflammatory monocytes were found to be the primary source of pro-inflammatory cytokines, the CD16(+) cells, in particular the CD14(+)CD16(+) monocytes, expressed the highest levels of activation markers, which included chemokine receptors and adhesion molecules. Morphologically, CD14(+) were distinguished from CD14lo monocytes by displaying larger and more active mitochondria. CD14(+)CD16(+) monocytes were more efficient in phagocytizing P. vivax-infected reticulocytes, which induced them to produce high levels of intracellular TNF-α and reactive oxygen species. Importantly, antibodies specific for ICAM-1, PECAM-1 or LFA-1 efficiently blocked the phagocytosis of infected reticulocytes by monocytes. Hence, our results provide key information on the mechanism by which CD14(+)CD16(+) cells control parasite burden, supporting the hypothesis that they play a role in resistance to P. vivax infection.

  2. CD14++CD16+ Monocytes Are Enriched by Glucocorticoid Treatment and Are Functionally Attenuated in Driving Effector T Cell Responses.

    PubMed

    Liu, Baoying; Dhanda, Ashwin; Hirani, Sima; Williams, Emily L; Sen, H Nida; Martinez Estrada, Fernando; Ling, Diamond; Thompson, Ian; Casady, Megan; Li, Zhiyu; Si, Han; Tucker, William; Wei, Lai; Jawad, Shayma; Sura, Amol; Dailey, Jennifer; Hannes, Susan; Chen, Ping; Chien, Jason L; Gordon, Siamon; Lee, Richard W J; Nussenblatt, Robert B

    2015-06-01

    Human peripheral monocytes have been categorized into three subsets based on differential expression levels of CD14 and CD16. However, the factors that influence the distribution of monocyte subsets and the roles that each subset plays in autoimmunity are not well studied. In this study, we show that circulating monocytes from patients with autoimmune uveitis exhibit a skewed phenotype toward intermediate CD14(++)CD16(+) cells, and that this is associated with glucocorticoid therapy. We further demonstrate that CD14(++)CD16(+) monocytes from patients and healthy control donors share a similar cell-surface marker and gene expression profile. Comparison of the effects of intermediate CD14(++)CD16(+) monocytes with classical CD14(++)CD16(-) and nonclassical CD14(+)CD16(++) monocytes revealed that the intermediate CD14(++)CD16(+) subset had an attenuated capacity to promote both naive CD4(+) T cell proliferation and polarization into a Th1 phenotype, and memory CD4(+) T cell proliferation and IL-17 expression. Furthermore, CD14(++)CD16(+) cells inhibit CD4(+) T cell proliferation induced by other monocyte subsets and enhance CD4(+) T regulatory cell IL-10 expression. These data demonstrate the impact of glucocorticoids on monocyte phenotype in the context of autoimmune disease and the differential effects of monocyte subsets on effector T cell responses.

  3. Prognostic and diagnostic value of plasma soluble ST2 concentrations in Acute Respiratory Distress Syndrome

    PubMed Central

    Bajwa, Ednan K.; Volk, Jessica A.; Christiani, David C.; Harris, R. Scott; Matthay, Michael A.; Thompson, B. Taylor; Januzzi, James L.

    2013-01-01

    Objective Soluble ST2 (sST2) is a biomarker of myocardial strain and inflammation. The characteristics of acute respiratory distress syndrome (ARDS) include inflammation and cardiovascular dysfunction. We sought to determine whether plasma sST2 concentration is associated with outcome and response to conservative fluid management, and whether sST2 concentration discriminates ARDS from decompensated heart failure (HF). Design, Setting, and Patients We assayed plasma sST2 concentrations in 826 patients in the Fluid and Catheter Treatment Trial (FACTT), a multi-center randomized controlled trial of conservative fluid management in ARDS, as well as a cohort of patients with decompensated HF. We tested whether sST2 was associated with outcome, response to therapy, and diagnostic utility for ARDS vs. HF. Measurements and Main Results Non-survivors had higher day 0 (P<.0001) and day 3 (P<.0001) sST2 concentrations. After adjustment for severity of illness, higher sST2 concentration was associated with mortality, with odds ratio (ORadj) 1.47 (95% confidence interval [CI] 0.99 – 2.20, P=.06) at day 0, 2.94 (95% CI 2.00 – 4.33, P<.0001) at day 3, and 3.63 (95% CI 2.38 – 5.53, P<.0001) if sST2 increased between days. Cumulative fluid balance was more positive among patients with higher day 0 (median 5212 mL, interquartile range [IQR] 200 – 12284 vs. 2020 mL, −2034 – 7091; P<0.0001), and day 3 sST2 (median 7678 mL, IQR 2217 – 14278 vs. 1492 mL, −2384 – 6239; P<0.0001). sST2 showed excellent discriminative ability between the FACTT and HF populations (Area under ROC curve=0.98, P<0.0001). Conclusions Higher sST2 concentrations are associated with worse outcome in ARDS and may have value for discriminating ARDS from heart failure. PMID:23939353

  4. Clinical Proteomics Identifies Urinary CD14 as a Potential Biomarker for Diagnosis of Stable Coronary Artery Disease

    PubMed Central

    Lee, Min-Yi; Huang, Chun-Hao; Kuo, Chao-Jen; Lin, Chen-Lung Steve; Lai, Wen-Ter; Chiou, Shyh-Horng

    2015-01-01

    Inflammation plays a key role in coronary artery disease (CAD) and other manifestations of atherosclerosis. Recently, urinary proteins were found to be useful markers for reflecting inflammation status of different organs. To identify potential biomarker for diagnosis of CAD, we performed one-dimensional SDS-gel electrophoresis followed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Among the proteins differentially expressed in urine samples, monocyte antigen CD14 was found to be consistently expressed in higher amounts in the CAD patients as compared to normal controls. Using enzyme-linked immunosorbent assays to analyze the concentrations of CD14 in urine and serum, we confirmed that urinary CD14 levels were significantly higher in patients (n = 73) with multi-vessel and single vessel CAD than in normal control (n = 35) (P < 0.001). Logistic regression analysis further showed that urinary CD14 concentration level is associated with severity or number of diseased vessels and SYNTAX score after adjustment for potential confounders. Concomitantly, the proportion of CD14+ monocytes was significantly increased in CAD patients (59.7 ± 3.6%) as compared with healthy controls (14.9 ± 2.1%) (P < 0.001), implicating that a high level of urinary CD14 may be potentially involved in mechanism(s) leading to CAD pathogenesis. By performing shotgun proteomics, we further revealed that CD14-associated inflammatory response networks may play an essential role in CAD. In conclusion, the current study has demonstrated that release of CD14 in urine coupled with more CD14+ monocytes in CAD patients is significantly correlated with severity of CAD, pointing to the potential application of urinary CD14 as a novel noninvasive biomarker for large-scale diagnostic screening of susceptible CAD patients. PMID:25668619

  5. Neurogenic potential of progenitors derived from human circulating CD14+ monocytes.

    PubMed

    Kodama, Hiroaki; Inoue, Takafumi; Watanabe, Ryuichi; Yasutomi, Daisuke; Kawakami, Yutaka; Ogawa, Satoshi; Mikoshiba, Katsuhiko; Ikeda, Yasuo; Kuwana, Masataka

    2006-04-01

    We previously reported a primitive cell fraction derived from human circulating CD14+ monocytes, named monocyte-derived multipotential cells (MOMC), that can differentiate along mesenchymal lineages, including bone, cartilage, fat, skeletal muscle and cardiac muscle. In this study, we investigated whether MOMC can differentiate into the neuronal lineage. MOMC were fluorescently labelled and cocultivated with a primary culture of rat neurons for up to 4 weeks. The protein and gene expressions of neuron-specific markers in the human MOMC were evaluated over time using immunohistochemistry, in situ hybridization and reverse transcription followed by PCR. Shortly after cocultivation with rat neurons, nearly all the MOMC expressed early neuroectodermal markers, Mash1, Neurogenin2 and NeuroD, together with nestin, an intermediate filament expressed in neurogenesis. After 14 days of coculture, a subpopulation of MOMC displayed a multipolar morphology with elongated neurites and expressed mature neuron-specific markers, including neurofilament, microtubule-associated protein type 2, beta3-tubulin, NeuN and Hu. Transdifferentiation of monocytes into the neuroectodermal lineage was shown by the simultaneous expression of proneural markers and CD45/CD14 early in the differentiation process. The cocultivated MOMC retained their proliferative capacity for at least 16 days. Finally, the neuronal differentiation of MOMC was observed when they were cultured with neurons without cell-to-cell contact. The capacity of MOMC to differentiate into both mesodermal and neuroectodermal lineages suggests that circulating CD14+ monocytes are more multipotential than previously thought.

  6. Association between plasma soluble RAGE and renal function is unaffected by medication usage and enzymatic antioxidants in chronic kidney disease with type 2 diabetes.

    PubMed

    Wong, Foo Nian; Tan, Jin Ai Mary Anne; Keng, Tee Chau; Ng, Kok Peng; Chua, Kek Heng; Kuppusamy, Umah Rani

    2016-01-30

    This study aimed to investigate the relationship between soluble RAGE and estimated glomerular filtration rate (eGFR) in patients with chronic kidney disease (CKD) after controlling for the potential confounding factors such as medication usage and enzymatic antioxidants. A total of 222 CKD patients whose eGFR is less than 60ml/min/1.73m(2) and 111 non-CKD individuals were recruited. The study subjects were classified based on their diabetes status. The plasma glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities as well as plasma soluble RAGE level were measured. The plasma GPx and SOD activities were significantly lower and the plasma soluble RAGE level was significantly higher in the CKD patients than in the non-CKD individuals, regardless of the diabetes status. Soluble RAGE was significantly correlated with eGFR in both diabetic CKD (D-CKD) and non-diabetic CKD (ND-CKD) patients. The association between soluble RAGE and eGFR remained largely unaffected by the confounding factors in D-CKD patients. However, the confounding effect of enzymatic antioxidants in the relationship between eGFR and soluble RAGE was observed in ND-CKD patients. The increased plasma level of soluble RAGE is a better indicator of renal function decline in diabetic CKD patients instead of non-diabetic CKD patients. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. HLA-E polymorphism and soluble HLA-E plasma levels in chronic hepatitis B patients.

    PubMed

    Zidi, I; Laaribi, A B; Bortolotti, D; Belhadj, M; Mehri, A; Yahia, H B; Babay, W; Chaouch, H; Zidi, N; Letaief, A; Yacoub, S; Boukadida, J; Di Luca, D; Hannachi, N; Rizzo, R

    2016-03-01

    Chronic hepatitis B virus (HBV) infection occurs in association to a deregulation of immune system. Human leukocyte antigen E (HLA-E) is an immune-tolerant nonclassical HLA class I molecule that could be involved in HBV progression. To measure soluble (s) HLA-E in patients with chronic HBV hepatitis (CHB). We tested the potential association of HLA-E*01:01/01:03 A > G gene polymorphism to CHB. Our cohort consisted of 93 Tunisian CHB patients (stratified in CHB with high HBV DNA levels and CHB with low HBV DNA levels) and 245 healthy donors. Plasma sHLA-E was determined using enzyme-linked immunosorbent assay (ELISA). Genotyping was performed using polymerase chain reaction sequence-specific primer. No association between HLA-E*01:01/01:03 A > G polymorphism and HBV DNA levels in CHB patients was found. G/G genotype is less frequent in CHB patients without significance. sHLA-E is significantly enhanced in CHB patients compared with healthy controls (P = 0.0017). Stratification according to HBV DNA levels showed that CHB patients with low HBV DNA levels have higher sHLA-E levels compared with CHB patients with high HBV DNA levels. CHB patients with G/G genotype have enhanced sHLA-E levels compared with other genotypes (P = 0.037). This significant difference is maintained only for CHB women concerning G/G genotypes (P = 0.042). Finally, we reported enhanced sHLA-E in CHB patients with advanced stages of fibrosis (P = 0.032). We demonstrate, for the first time, the association of sHLA-E to CHB. Owing to the positive correlation of HLA-E*01:01/01:03 A > G polymorphism and the association of sHLA-E to advanced fibrosis stages, HLA-E could be a powerful predictor for CHB progression. Further investigations will be required to substantiate HLA-E role as a putative clinical biomarker of CHB.

  8. IL21R expressing CD14(+)CD16(+) monocytes expand in multiple myeloma patients leading to increased osteoclasts.

    PubMed

    Bolzoni, Marina; Ronchetti, Domenica; Storti, Paola; Donofrio, Gaetano; Marchica, Valentina; Costa, Federica; Agnelli, Luca; Toscani, Denise; Vescovini, Rosanna; Todoerti, Katia; Bonomini, Sabrina; Sammarelli, Gabriella; Vecchi, Andrea; Guasco, Daniela; Accardi, Fabrizio; Palma, Benedetta Dalla; Gamberi, Barbara; Ferrari, Carlo; Neri, Antonino; Aversa, Franco; Giuliani, Nicola

    2017-04-01

    Bone marrow monocytes are primarily committed to osteoclast formation. It is, however, unknown whether potential primary alterations are specifically present in bone marrow monocytes from patients with multiple myeloma, smoldering myeloma or monoclonal gammopathy of undetermined significance. We analyzed the immunophenotypic and transcriptional profiles of bone marrow CD14(+) monocytes in a cohort of patients with different types of monoclonal gammopathies to identify alterations involved in myeloma-enhanced osteoclastogenesis. The number of bone marrow CD14(+)CD16(+) cells was higher in patients with active myeloma than in those with smoldering myeloma or monoclonal gammopathy of undetermined significance. Interestingly, sorted bone marrow CD14(+)CD16(+) cells from myeloma patients were more pro-osteoclastogenic than CD14(+)CD16-cells in cultures ex vivo Moreover, transcriptional analysis demonstrated that bone marrow CD14(+) cells from patients with multiple myeloma (but neither monoclonal gammopathy of undetermined significance nor smoldering myeloma) significantly upregulated genes involved in osteoclast formation, including IL21RIL21R mRNA over-expression by bone marrow CD14(+) cells was independent of the presence of interleukin-21. Consistently, interleukin-21 production by T cells as well as levels of interleukin-21 in the bone marrow were not significantly different among monoclonal gammopathies. Thereafter, we showed that IL21R over-expression in CD14(+) cells increased osteoclast formation. Consistently, interleukin-21 receptor signaling inhibition by Janex 1 suppressed osteoclast differentiation from bone marrow CD14(+) cells of myeloma patients. Our results indicate that bone marrow monocytes from multiple myeloma patients show distinct features compared to those from patients with indolent monoclonal gammopathies, supporting the role of IL21R over-expression by bone marrow CD14(+) cells in enhanced osteoclast formation. Copyright© Ferrata Storti

  9. IL21R expressing CD14+CD16+ monocytes expand in multiple myeloma patients leading to increased osteoclasts

    PubMed Central

    Bolzoni, Marina; Ronchetti, Domenica; Storti, Paola; Donofrio, Gaetano; Marchica, Valentina; Costa, Federica; Agnelli, Luca; Toscani, Denise; Vescovini, Rosanna; Todoerti, Katia; Bonomini, Sabrina; Sammarelli, Gabriella; Vecchi, Andrea; Guasco, Daniela; Accardi, Fabrizio; Palma, Benedetta Dalla; Gamberi, Barbara; Ferrari, Carlo; Neri, Antonino; Aversa, Franco; Giuliani, Nicola

    2017-01-01

    Bone marrow monocytes are primarily committed to osteoclast formation. It is, however, unknown whether potential primary alterations are specifically present in bone marrow monocytes from patients with multiple myeloma, smoldering myeloma or monoclonal gammopathy of undetermined significance. We analyzed the immunophenotypic and transcriptional profiles of bone marrow CD14+ monocytes in a cohort of patients with different types of monoclonal gammopathies to identify alterations involved in myeloma-enhanced osteoclastogenesis. The number of bone marrow CD14+CD16+ cells was higher in patients with active myeloma than in those with smoldering myeloma or monoclonal gammopathy of undetermined significance. Interestingly, sorted bone marrow CD14+CD16+ cells from myeloma patients were more pro-osteoclastogenic than CD14+CD16-cells in cultures ex vivo. Moreover, transcriptional analysis demonstrated that bone marrow CD14+ cells from patients with multiple myeloma (but neither monoclonal gammopathy of undetermined significance nor smoldering myeloma) significantly upregulated genes involved in osteoclast formation, including IL21R. IL21R mRNA over-expression by bone marrow CD14+ cells was independent of the presence of interleukin-21. Consistently, interleukin-21 production by T cells as well as levels of interleukin-21 in the bone marrow were not significantly different among monoclonal gammopathies. Thereafter, we showed that IL21R over-expression in CD14+ cells increased osteoclast formation. Consistently, interleukin-21 receptor signaling inhibition by Janex 1 suppressed osteoclast differentiation from bone marrow CD14+ cells of myeloma patients. Our results indicate that bone marrow monocytes from multiple myeloma patients show distinct features compared to those from patients with indolent monoclonal gammopathies, supporting the role of IL21R over-expression by bone marrow CD14+ cells in enhanced osteoclast formation. PMID:28057743

  10. Dopamine increases CD14+CD16+ monocyte migration and adhesion in the context of substance abuse and HIV neuropathogenesis.

    PubMed

    Coley, Jacqueline S; Calderon, Tina M; Gaskill, Peter J; Eugenin, Eliseo A; Berman, Joan W

    2015-01-01

    Drug abuse is a major comorbidity of HIV infection and cognitive disorders are often more severe in the drug abusing HIV infected population. CD14+CD16+ monocytes, a mature subpopulation of peripheral blood monocytes, are key mediators of HIV neuropathogenesis. Infected CD14+CD16+ monocyte transmigration across the blood brain barrier mediates HIV entry into the brain and establishes a viral reservoir within the CNS. Despite successful antiretroviral therapy, continued influx of CD14+CD16+ monocytes, both infected and uninfected, contributes to chronic neuroinflammation and the development of HIV associated neurocognitive disorders (HAND). Drug abuse increases extracellular dopamine in the CNS. Once in the brain, CD14+CD16+ monocytes can be exposed to extracellular dopamine due to drug abuse. The direct effects of dopamine on CD14+CD16+ monocytes and their contribution to HIV neuropathogenesis are not known. In this study, we showed that CD14+CD16+ monocytes express mRNA for all five dopamine receptors by qRT-PCR and D1R, D5R and D4R surface protein by flow cytometry. Dopamine and the D1-like dopamine receptor agonist, SKF38393, increased CD14+CD16+ monocyte migration that was characterized as chemokinesis. To determine whether dopamine affected cell motility and adhesion, live cell imaging was used to monitor the accumulation of CD14+CD16+ monocytes on the surface of a tissue culture dish. Dopamine increased the number and the rate at which CD14+CD16+ monocytes in suspension settled to the dish surface. In a spreading assay, dopamine increased the area of CD14+CD16+ monocytes during the early stages of cell adhesion. In addition, adhesion assays showed that the overall total number of adherent CD14+CD16+ monocytes increased in the presence of dopamine. These data suggest that elevated extracellular dopamine in the CNS of HIV infected drug abusers contributes to HIV neuropathogenesis by increasing the accumulation of CD14+CD16+ monocytes in dopamine rich brain

  11. Dopamine Increases CD14+CD16+ Monocyte Migration and Adhesion in the Context of Substance Abuse and HIV Neuropathogenesis

    PubMed Central

    Coley, Jacqueline S.; Calderon, Tina M.; Gaskill, Peter J.; Eugenin, Eliseo A.; Berman, Joan W.

    2015-01-01

    Drug abuse is a major comorbidity of HIV infection and cognitive disorders are often more severe in the drug abusing HIV infected population. CD14+CD16+ monocytes, a mature subpopulation of peripheral blood monocytes, are key mediators of HIV neuropathogenesis. Infected CD14+CD16+ monocyte transmigration across the blood brain barrier mediates HIV entry into the brain and establishes a viral reservoir within the CNS. Despite successful antiretroviral therapy, continued influx of CD14+CD16+ monocytes, both infected and uninfected, contributes to chronic neuroinflammation and the development of HIV associated neurocognitive disorders (HAND). Drug abuse increases extracellular dopamine in the CNS. Once in the brain, CD14+CD16+ monocytes can be exposed to extracellular dopamine due to drug abuse. The direct effects of dopamine on CD14+CD16+ monocytes and their contribution to HIV neuropathogenesis are not known. In this study, we showed that CD14+CD16+ monocytes express mRNA for all five dopamine receptors by qRT-PCR and D1R, D5R and D4R surface protein by flow cytometry. Dopamine and the D1-like dopamine receptor agonist, SKF38393, increased CD14+CD16+ monocyte migration that was characterized as chemokinesis. To determine whether dopamine affected cell motility and adhesion, live cell imaging was used to monitor the accumulation of CD14+CD16+ monocytes on the surface of a tissue culture dish. Dopamine increased the number and the rate at which CD14+CD16+ monocytes in suspension settled to the dish surface. In a spreading assay, dopamine increased the area of CD14+CD16+ monocytes during the early stages of cell adhesion. In addition, adhesion assays showed that the overall total number of adherent CD14+CD16+ monocytes increased in the presence of dopamine. These data suggest that elevated extracellular dopamine in the CNS of HIV infected drug abusers contributes to HIV neuropathogenesis by increasing the accumulation of CD14+CD16+ monocytes in dopamine rich brain

  12. Vitamin D, d-dimer, Interferon γ, and sCD14 Levels are Independently Associated with Immune Reconstitution Inflammatory Syndrome: A Prospective, International Study☆

    PubMed Central

    Musselwhite, Laura W.; Andrade, Bruno B.; Ellenberg, Susan S.; Tierney, Ann; Belaunzaran-Zamudio, Pablo F.; Rupert, Adam; Lederman, Michael M.; Sanne, Ian; Sierra Madero, Juan G.; Sereti, Irini

    2016-01-01

    To determine the immunological profile most important for IRIS prediction, we evaluated 20 baseline plasma biomarkers in Acquired Immunodeficiency Syndrome (AIDS) patients initiating antiretroviral therapy (ART). Patients were enrolled in a randomized, placebo-controlled ART initiation trial in South Africa and Mexico to test whether maraviroc could prevent IRIS. Participants were classified prospectively as having IRIS within 6 months of ART initiation. Twenty plasma biomarkers were measured at study enrollment for 267 participants. Biomarkers were tested for predicting IRIS with adjustment for covariates chosen through forward stepwise selection. Sixty-two participants developed IRIS and of these 19 were tuberculosis (TB)-IRIS. Baseline levels of vitamin D and higher d-dimer, interferon gamma (IFNγ), and sCD14 were independently associated with risk of IRIS in multivariate analyses. TB-IRIS cases exhibited a distinct biosignature from IRIS related to other pathogens, with increased levels of C-reactive protein (CRP), sCD14, IFNγ, and lower levels of Hb that could be captured by a composite risk score. Elevated markers of Type 1 T helper (Th1) response, monocyte activation, coagulation and low vitamin D were independently associated with IRIS risk. Interventions that decrease immune activation and increase vitamin D levels warrant further study. PMID:26981576

  13. Monocyte activation is a feature of common variable immunodeficiency irrespective of plasma lipopolysaccharide levels

    PubMed Central

    Barbosa, R R; Silva, S P; Silva, S L; Tendeiro, R; Melo, A C; Pedro, E; Barbosa, M P; Santos, M C P; Victorino, R M M; Sousa, A E

    2012-01-01

    Common variable immunodeficiency disorders (CVID), the most frequent cause of symptomatic primary immunodeficiency, are defined by impaired antibody production. Notwithstanding, T cell activation and granulomatous manifestations represent the main causes of CVID morbidity even in patients receiving immunoglobulin (Ig) G replacement therapy. Additionally, gut pathology is a frequent feature of CVID. In this study, we investigated monocyte imbalances and their possible relationship with increased microbial translocation in CVID patients. Monocyte subsets were defined according to CD14 and CD16 expression levels and evaluated in terms of human leucocyte antigen D-related (HLA-DR), CD86 and programmed death-1 molecule ligand 1 (PD-L1) expression by flow cytometry, in parallel with the quantification of plasma lipopolysaccharide (LPS) and serum levels of soluble CD14 (sCD14), LPS-binding protein (LBP) and anti-LPS antibodies. CVID patients (n = 31) featured significantly increased levels of serum sCD14 and an expansion of CD14brightCD16+ monocytes in direct correlation with T cell and B cell activation, the latter illustrated by the frequency of the CD21lowCD38low subset. Such alterations were not observed in patients lacking B cells due to congenital agammaglobulinaemia (n = 4). Moreover, we found no significant increase in circulating LPS or LBP levels in CVID patients, together with a relative preservation of serum anti-LPS antibodies, in agreement with their presence in commercial IgG preparations. In conclusion, CVID was associated with monocyte imbalances that correlated directly with T cell activation markers and with B cell imbalances, without an association with plasma LPS levels. The heightened monocyte activated state observed in CVID may represent an important target for complementary therapeutic strategies. PMID:22861366

  14. BLUNTING AIRWAYS EOSINOPHILIC INFLAMMATION RESULTS IN A DECREASED AIRWAY NEUTROPHIL RESPONSE TO INHALED LPS IN ATOPIC ASTHMATICS A ROLE FOR CD-14

    EPA Science Inventory

    Recent data demonstrate that atopic inflammation might enhance airway responses to inhaled LPS in individuals with atopic asthma by increasing CD14 expression on airway macrophages. We sought to determine whether blunting airway eosinophilic inflammation decreases CD14 expressio...

  15. BLUNTING AIRWAYS EOSINOPHILIC INFLAMMATION RESULTS IN A DECREASED AIRWAY NEUTROPHIL RESPONSE TO INHALED LPS IN ATOPIC ASTHMATICS A ROLE FOR CD-14

    EPA Science Inventory

    Recent data demonstrate that atopic inflammation might enhance airway responses to inhaled LPS in individuals with atopic asthma by increasing CD14 expression on airway macrophages. We sought to determine whether blunting airway eosinophilic inflammation decreases CD14 expressio...

  16. Expansion and preferential activation of the CD14(+)CD16(+) monocyte subset during multiple sclerosis.

    PubMed

    Chuluundorj, Delgertsetseg; Harding, Scott A; Abernethy, David; La Flamme, Anne Camille

    2014-07-01

    Multiple sclerosis (MS) is an immune-driven, demyelinating disease of the central nervous system (CNS). Although many types of immune cells are involved in disease progression, activated monocytes are believed to be one of the first to arrive to the brain and initiate inflammation. However, little is known about how the two main monocyte subsets, CD14(++)CD16(-) and CD14(+)CD16(+), are involved in MS. To understand how the phenotype and responses of these monocyte subsets are altered during MS, total monocytes and the purified monocyte subsets from healthy subjects (n=29) and MS patients (n=20) were characterized ex vivo and stimulated in vitro with lipopolysaccharide (LPS). The ex vivo analyses showed that total monocytes from MS patients had significantly elevated levels of CD40, CD86, HLA-DR, CD64 and C-C motif chemokine receptor 2 (CCR2), and this elevation was most marked on CD16(+) monocytes. In vitro stimulation with LPS led to an increase in CD86, HLA-DR, CD64 and IL-6 production by monocytes from MS patients. Furthermore, in purified cultures, CD14(+) monocytes were found to be the main producers of IL-10 while CD16(+) monocytes produced more IL-12. In monocytes from MS patients, both subsets produced substantially more IL-6, and the production of IL-10 by the CD16(+) subset was also significantly elevated compared with healthy monocytes. Together these findings highlight the important contribution of the CD16(+) monocyte subset in driving inflammatory responses during MS.

  17. YKL-40 expression in CD14+ liver cells in acute and chronic injury

    PubMed Central

    Pizano-Martínez, Oscar; Yañez-Sánchez, Irinea; Alatorre-Carranza, Pilar; Miranda-Díaz, Alejandra; Ortiz-Lazareno, Pablo C; García-Iglesias, Trinidad; Daneri-Navarro, Adrian; Mercado, Mónica Vázquez-Del; Fafutis-Morris, Mary; Delgado-Rizo, Vidal

    2011-01-01

    AIM: To demonstrate that CD14+ cells are an important source of the growth factor YKL-40 in acute and chronic liver damage. METHODS: Rats were inoculated with one dose of CCl4 to induce acute damage. Liver biopsies were obtained at 0, 6, 12, 24, 48 and 72 h. For chronic damage, CCl4 was administered three days per week for 6 or 8 wk. Tissue samples were collected, and cellular populations were isolated by liver digestion and purified by cell sorting. YKL-40 mRNA and protein expression were evaluated by real-time polymerase chain reaction and western blot. RESULTS: Acute liver damage induced a rapid increase of YKL-40 mRNA beginning at 12 h. Expression peaked at 24 h, with a 26-fold increase over basal levels. By 72 h however, YKL-40 expression levels had nearly returned to control levels. On the other hand, chronic damage induced a sustained increase in YKL-40 expression, with 7- and 9-fold higher levels at 6 and 8 wk, respectively. The pattern of YKL-40 expression in different subpopulations showed that CD14+ cells, which include Kupffer cells, are a source of YKL-40 after acute damage at 72 h [0.09 relative expression units (REU)] as well as after chronic injury at 6 wk (0.11 REU). Hepatocytes, in turn, accounted for 0.06 and 0.01 REU after 72 h (acute) or 6 wk (chronic), respectively. The rest of the CD14- cells (including T lymphocytes, B lymphocytes, natural killer and natural killer T cells) yielded 0.07 and 0.15 REU at 72 h and 6 wk, respectively. YKL-40 protein expression in liver was detected at 72 h as well as 6 and 8 wk, with the highest expression relative to controls (11-fold; P ≤ 0.05) seen at 6 wk. Macrophages were stimulated by lipopolysaccharide. We demonstrate that under these conditions, these cells showed maximum expression of YKL-40 at 12 h, with P < 0.05 compared with controls. CONCLUSION: Hepatic CD14+ cells are an YKL-40 mRNA and protein source in acute and chronic liver injury, with expression patterns similar to growth factors implicated

  18. A differential protein solubility approach for the depletion of highly abundant proteins in plasma using ammonium sulfate.

    PubMed

    Bollineni, Ravi Chand; Guldvik, Ingrid J; Grönberg, Henrik; Wiklund, Fredrik; Mills, Ian G; Thiede, Bernd

    2015-12-21

    Depletion of highly abundant proteins is an approved step in blood plasma analysis by mass spectrometry (MS). In this study, we explored a precipitation and differential protein solubility approach as a fractionation strategy for abundant protein removal from plasma. Total proteins from plasma were precipitated with 90% saturated ammonium sulfate, followed by differential solubilization in 55% and 35% saturated ammonium sulfate solutions. Using a four hour liquid chromatography (LC) gradient and an LTQ-Orbitrap XL mass spectrometer, a total of 167 and 224 proteins were identified from the 55% and 35% ammonium sulfate fractions, whereas 235 proteins were found in the remaining protein fractions with at least two unique peptides. SDS-PAGE and exclusive total spectrum counts from LC-MS/MS analyses clearly showed that majority of the abundant plasma proteins were solubilized in 55% and 35% ammonium sulfate solutions, indicating that the remaining protein fraction is of potential interest for identification of less abundant plasma proteins. Serum albumin, serotransferrin, alpha-1-antitrypsin and transthyretin were the abundant proteins that were highly enriched in 55% ammonium sulfate fractions. Immunoglobulins, complement system proteins, and apolipoproteins were among other abundant plasma proteins that were enriched in 35% ammonium sulfate fractions. In the remaining protein fractions a total of 40 unique proteins were identified of which, 32 proteins were identified with at least 10 exclusive spectrum counts. According to PeptideAtlas, 9 of these 32 proteins were estimated to be present at low μg ml(-1) (0.12-1.9 μg ml(-1)) concentrations in the plasma, and 17 at low ng ml(-1) (0.1-55 ng ml(-1)) range.

  19. Plasma Levels of Pentosidine, Carboxymethyl-Lysine, Soluble Receptor for Advanced Glycation End Products, and Metabolic Syndrome: The Metformin Effect

    PubMed Central

    Haddad, Mohamed; Knani, Ines; Bouzidi, Hsan; Berriche, Olfa; Hammami, Mohamed

    2016-01-01

    Metabolic syndrome (MetS) is considered one of the most important public health problems. Several and controversial studies showed that the role of advanced glycation end products (AGEs) and their receptor in the development of metabolic syndrome and therapeutic pathways is still unsolved. We have investigated whether plasma pentosidine, carboxymethyl-lysine (CML), and soluble receptor for advanced glycation end products (sRAGE) levels were increased in patients with MetS and the effect of metformin in plasma levels of pentosidine, CML, and sRAGE. 80 control subjects and 86 patients were included in this study. Pentosidine, CML, and sRAGE were measured in plasma by enzyme-linked immunosorbent assay (ELISA). Plasma pentosidine, CML, and sRAGE levels were significantly increased in patients compared to control subjects (P < 0.001, P < 0.001, and P = 0.014, resp.). Plasma levels of pentosidine were significantly decreased in patients who received metformin compared to untreated patients (P = 0.01). However, there was no significant difference between patients treated with metformin and untreated patients in plasma CML levels. Plasma levels of sRAGE were significantly increased in patients who received metformin and ACE inhibitors (P < 0.001 and P = 0.002, resp.). However, in a multiple stepwise regression analysis, pentosidine, sRAGE, and drugs treatments were not independently associated. Patients with metabolic syndrome showed increased levels of AGEs such as pentosidine and CML. Metformin treatment showed a decreased level of pentosidine but not of CML. Therapeutic pathways of AGEs development should be taken into account and further experimental and in vitro studies merit for advanced research. PMID:27829696

  20. CD14+ cells from peripheral blood positively regulate hematopoietic stem and progenitor cell survival resulting in increased erythroid yield.

    PubMed

    Heideveld, Esther; Masiello, Francesca; Marra, Manuela; Esteghamat, Fatemehsadat; Yağcı, Nurcan; von Lindern, Marieke; Migliaccio, Anna Rita F; van den Akker, Emile

    2015-11-01

    Expansion of erythroblasts from human peripheral blood mononuclear cells is 4- to 15-fold more efficient than that of CD34(+) cells purified from peripheral blood mononuclear cells. In addition, purified CD34(+) and CD34(-) populations from blood do not reconstitute this erythroid yield, suggesting a role for feeder cells present in blood mononuclear cells that increase hematopoietic output. Immunodepleting peripheral blood mononuclear cells for CD14(+) cells reduced hematopoietic stem and progenitor cell expansion. Conversely, the yield was increased upon co-culture of CD34(+) cells with CD14(+) cells (full contact or transwell assays) or CD34(+) cells re-constituted in conditioned medium from CD14(+) cells. In particular, CD14(++)CD16(+) intermediate monocytes/macrophages enhanced erythroblast outgrowth from CD34(+) cells. No effect of CD14(+) cells on erythroblasts themselves was observed. However, 2 days of co-culturing CD34(+) and CD14(+) cells increased CD34(+) cell numbers and colony-forming units 5-fold. Proliferation assays suggested that CD14(+) cells sustain CD34(+) cell survival but not proliferation. These data identify previously unrecognized erythroid and non-erythroid CD34(-) and CD34(+) populations in blood that contribute to the erythroid yield. A flow cytometry panel containing CD34/CD36 can be used to follow specific stages during CD34(+) differentiation to erythroblasts. We have shown modulation of hematopoietic stem and progenitor cell survival by CD14(+) cells present in peripheral blood mononuclear cells which can also be found near specific hematopoietic niches in the bone marrow.

  1. PLASMA SOLUBLE SGP130 LEVELS ARE INCREASED IN OLDER SUBJECTS WITH METABOLIC SYNDROME. THE ROLE OF INSULIN RESISTANCE

    PubMed Central

    Zuliani, Giovanni; Galvani, Matteo; Maggio, Marcello; Volpato, Stefano; Bandinelli, Stefania; Corsi, Anna Maria; Lauretani, Fulvio; Cherubini, Antonio; Guralnik, Jack M.; Fellin, Renato; Ferrucci, Luigi

    2010-01-01

    Objective Increased interleukin-6 plasma levels have been reported in metabolic syndrome (MS); nevertheless, it is unclear whether interleukin-6 activity is exerted through direct signalling only or also through the “trans-signalling”. This issue is important to clarify since signalling and “trans-signalling” affect different tissues. We investigated the relationship between MS and the interleukin-6 system in an older population. Methods Data from 997 older community dwelling individuals (age ≥ 65 years; females: 56.2%) enrolled the InChianti study were analyzed. Interleukin-6, soluble interleukin-6 receptor (sIL-6r), and soluble glycoprotein 130 (sgp130) were measured on plasma by ELISA. MS was defined by the NCEP-ATPIII criteria; 309 individuals (31%) resulted affected by MS. Results Subjects with MS had higher interleukin-6 and sgp130 levels compared to controls; a trend toward higher levels of sIL-6R was also observed. The risk of having MS was increased in individuals with high sIL-6r or/and sgp130 levels, independent of age, gender, and interleukin-6 levels. Elevated sgp130 levels were associated with higher plasma glucose, HOMA, triglycerides, and with diabetes both in subjects with and without MS. Although the risk of high sgp130 levels was generally associated with MS (O.R.:1.77, 95%C.I.: 1.39-2.25), this excess of risk was not present in MS phenotypes excluding the criteria “elevated glucose” or “elevated triglycerides”. Furthermore, the association between sgp130 and MS disappeared after adjustment for HOMA. Conclusions We found that older individuals with MS have increased sgp130 plasma levels compared with controls; nevertheless, our data suggest that this association might be mediated by insulin resistance. PMID:20869059

  2. Plasma soluble gp130 levels are increased in older subjects with metabolic syndrome. The role of insulin resistance.

    PubMed

    Zuliani, Giovanni; Galvani, Matteo; Maggio, Marcello; Volpato, Stefano; Bandinelli, Stefania; Corsi, Anna Maria; Lauretani, Fulvio; Cherubini, Antonio; Guralnik, Jack M; Fellin, Renato; Ferrucci, Luigi

    2010-11-01

    Increased interleukin-6 plasma levels have been reported in metabolic syndrome (MS); nevertheless, it is unclear whether interleukin-6 activity is exerted through direct signalling only or also through the "trans-signalling". This issue is important to clarify since signalling and "trans-signalling" affect different tissues. We investigated the relationship between MS and the interleukin-6 system in an older population. Data from 997 older community dwelling individuals (age ≥ 65 years; females: 56.2%) enrolled the InChianti study were analysed. Interleukin-6, soluble interleukin-6 receptor (sIL-6r), and soluble glycoprotein 130 (sgp130) were measured on plasma by ELISA. MS was defined by the NCEP ATP III criteria; 309 individuals (31%) resulted affected by MS. Subjects with MS had higher interleukin-6 and sgp130 levels compared to controls; a trend toward higher levels of sIL-6R was also observed. The risk of having MS was increased in individuals with high sIL-6r or/and sgp130 levels, independent of age, gender, and interleukin-6 levels. Elevated sgp130 levels were associated with higher plasma glucose, HOMA, triglycerides, and with diabetes both in subjects with and without MS. Although the risk of high sgp130 levels was generally associated with MS (O.R.: 1.77, 95%C.I.: 1.39-2.25), this excess of risk was not present in MS phenotypes excluding the criteria "elevated glucose" or "elevated triglycerides". Furthermore, the association between sgp130 and MS disappeared after adjustment for HOMA. We found that older individuals with MS have increased sgp130 plasma levels compared with controls; nevertheless, our data suggest that this association might be mediated by insulin resistance. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  3. Increased Soluble Urokinase-Type Plasminogen Activator Receptor (suPAR) Levels in Plasma of Suicide Attempters

    PubMed Central

    Ventorp, Filip; Gustafsson, Anna; Träskman-Bendz, Lil; Westrin, Åsa; Ljunggren, Lennart

    2015-01-01

    The soluble form of the urokinase receptor, suPAR, has been suggested as a novel biomarker of low-grade inflammation. Activation of the immune system has been proposed to contribute to the development of depression and suicidal behavior. In order to identify depressed and suicidal individuals who could benefit from an anti-inflammatory treatment, a reliable biomarker of low-grade inflammation is vital. This study evaluates plasma suPAR levels as a biomarker of low-grade inflammation in patients with major depressive disorder and in patients who recently attempted suicide. The plasma suPAR and an established biomarker, C reactive protein (CRP) of suicide attempters (n = 54), depressed patients (n = 19) and healthy controls (n = 19) was analyzed with enzyme-linked immunosorbent assays. The biomarker attributes of sensitivity and sensibility were evaluated using ROC curve analysis. Both the depressed patients and suicide attempters had increased plasma suPAR. The levels of suPAR discriminated better between controls and suicide attempters than did CRP. In the future, plasma suPAR might be a superior prognosticator regarding outcome of treatment applying conventional antidepressants in conjunction with anti-inflammatory drugs. PMID:26451727

  4. Ethanol alters cellular activation and CD14 partitioning in lipid rafts

    SciTech Connect

    Dai Qun; Zhang Jun; Pruett, Stephen B. . E-mail: spruet@lsuhsc.edu

    2005-06-24

    Alcohol consumption interferes with innate immunity. In vivo EtOH administration suppresses cytokine responses induced through Toll-like receptor 4 (TLR4) and inhibits TLR4 signaling. Actually, EtOH exhibits a generalized suppressive effect on signaling and cytokine responses induced by through most TLRs. However, the underlying mechanism remains unknown. RAW264.7 cells were treated with LPS or co-treated with EtOH or with lipid raft-disrupting drugs. TNF-{alpha} production, IRAK-1 activation, and CD14 partition were evaluated. EtOH or nystatin, a lipid raft-disrupting drug, suppressed LPS-induced production of TNF-{alpha}. The suppressive effect of EtOH on LPS-induced TNF-{alpha} production was additive with that of methyl-{beta}-cyclodextrin (MCD), another lipid raft-disrupting drug. EtOH interfered with IRAK-1 activation, an early TLR4 intracellular signaling event. Cell fractionation analyses show that acute EtOH altered LPS-related partition of CD14, a critical component of the LPS receptor complex. These results suggest a novel mechanism of EtOH action that involves interference with lipid raft clustering induced by LPS. This membrane action of EtOH might be one of the mechanisms by which EtOH acts as a generalized suppressor for TLR signaling.

  5. Membrane bound IL-15 is increased on CD14 monocytes in early stages of MS.

    PubMed

    Vaknin-Dembinsky, Adi; Brass, Steven D; Brass, Steven; Gandhi, Roopali; Weiner, Howard L

    2008-03-01

    IL-15 is a pro-inflammatory cytokine whose three-dimensional structure is similar to that of IL-2. IL-2 and IL-15 have similar as well as distinct biological functions. An active form of IL-15 that is membrane bound has also been described. Furthermore, IL-15 is known to play a role in autoimmune diseases. We thus investigated the expression of membrane bound IL-15 on monocytes (CD14+ cells) and studied its effect on T cell activation in MS patients. We found that unstimulated CD14+ cells from relapsing remitting MS patients had increased membrane bound IL-15. Those with high surface levels of IL-15 on monocytes were in the early stages of the disease. In addition, we found that T cells of MS patients had enhanced responsiveness to IL-15 and there was increased expression of IL-15 receptor on CD4+ T cells. Thus, IL-15 may be an important cytokine that drives Th1 responses early in the course of the disease and could serve as a target for immunotherapy and as an early marker in the immunologic staging of MS.

  6. The accuracy of presepsin (sCD14-ST) for the diagnosis of sepsis in adults: a meta-analysis.

    PubMed

    Zhang, Xin; Liu, Dan; Liu, You-Ning; Wang, Rui; Xie, Li-Xin

    2015-09-11

    The early diagnosis of sepsis remains a challenge. Recently, soluble cluster of differentiation 14 subtype (sCD14-ST), also known as presepsin, has been identified as a potential biomarker of sepsis. We performed a meta-analysis to assess the diagnostic accuracy of presepsin for sepsis in patients with systemic inflammation. We systematically searched the PubMed, Embase, Web of Knowledge and Cochrane databases. Studies were included if they assessed the diagnostic accuracy of presepsin for sepsis in adult patients with systemic inflammatory response syndrome (SIRS). Furthermore, a 2 × 2 contingency table was constructed based on these results. Two authors independently judged the studies and extracted the data. The diagnostic accuracy of presepsin in sepsis was calculated using a bivariate meta-analysis model. The Q-test and I (2) index were used to test the heterogeneity. Eight studies involving a total of 1,815 patients were included in the present study. The pooled sensitivity, specificity, diagnostic odds ratio, positive likelihood ratio and negative likelihood ratio were 0.86 (95% CI: 0.79-0.91), 0.78 (95% CI: 0.68-0.85), 22 (95% CI: 10-48), 3.8 (95% CI: 2.6-5.7), and 0.18 (95% CI: 0.11-0.28), respectively. The area under the summary receiver operator characteristic curve was 0.89 (95% CI: 0.86-0.92). Meta-regression analysis revealed that consecutive patient selection, sample size and setting significantly accounted for the heterogeneity of sensitivity. Our findings suggest that presepsin exhibits very good diagnostic accuracy (AUC=0.89) for the diagnosis for sepsis. Nevertheless, an overall assessment of all the clinical indexes for sepsis diagnosis and continual re-evaluation of presepsin during the course of the disease are needed.

  7. Soluble forms of CD46 are detected in Bos taurus plasma and neutralize BVDV, the bovine pestivirus.

    PubMed

    Alzamel, Nidal; Bayrou, Calixte; Decreux, Annabelle; Desmecht, Daniel

    2016-12-01

    The pestivirus bovine viral diarrhea virus (BVDV) is known to bind to the CD46 molecule, which subsequently promotes entry of the virus. Mapping of the BVD-virion-binding site has shown that two peptides, 66EQIV69 and 82GQVLAL87, located on antiparallel beta sheets in the most distal complement control protein module (CCP1), provide the attachment platform. In the present study, we reveal new CD46-encoding transcripts that are predicted to encode CCP1-containing soluble forms. Further, we show that the serum of most adult cattle contains soluble CD46 (sCD46) and that a recombinant soluble isoform neutralizes BVDV infectivity in an in vitro assay. We have then established an ELISA for determination of plasma sCD46 in a large cohort of animals. Overall, serum sCD46 amounts to 8±18ng/mL (mean±SD, n=440), with a IC [95-105] ranging from 6,4 to 9,8ng/mL and extreme values ​​between 0 and 178ng/mL. We found that sCD46 is not detectable in fetal and neonatal sera and that its plasma concentration increases progressively up to adulthood. We also detected high- and low-sCD46 performers and show that this phenotype does not depend of environment. As modern rearing techniques make it possible to disseminate genetically-determined phenotypes very quickly in a population, a large-scale study examining whether high-sCD46 animals provide epidemiological protection against BVDV infection and transmission should be undertaken. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Vasoactive intestinal peptide induces CD14+HLA-DR‑/low myeloid-derived suppressor cells in gastric cancer.

    PubMed

    Li, Gang; Wu, Ke; Tao, Kaixiong; Lu, Xiaoming; Ma, Jianhua; Mao, Zhengqiang; Li, Hang; Shi, Liang; Li, Jing; Niu, Yanfeng; Xiang, Fan; Wang, Guobin

    2015-07-01

    Myeloid-derived suppressor cells (MDSCs) are a heterogeneous group of cells, which have been revealed to inhibit T-cell responses in tumor-bearing mice. In addition, a number of immune suppressive mechanisms have linked MDSCs and the development of human cancer. However, the role of MDSCs in human gastric cancer tissue remains to be elucidated as specific markers are lacking. Therefore, the aim of the present study was to investigate the frequency and immune suppressive function of MDSCs denoted in the present study as cluster of differentiation 14 (CD14)+human leukocyte antigen (HLA)-DR-/low in gastric cancer patients. In the present study, MDSCs were directly isolated and characterized from the tumor and adjacent normal tissue of gastric cancer patients. Functional analysis of the CD14+HLA-DR-/low MDSCs co-cultured with allogeneic CD4+ T cells were performed and compared with controls. In addition, the interferon-γ (IFN-γ) and interleukin (IL)-2 production was compared in order to investigate the capacity of vasoactive intestinal peptide (VIP) to induce CD14+HLA-DR(-/low) MDSC-mediated CD4+ T-cell dysfunction and whether IL-10 secretion is involved in this mechanism. As a result, the quantity of CD14+HLA-DR(-/low) cells in tumor tissue from gastric cancer patients was significantly higher than that in the adjacent normal tissue. In addition, CD14+HLA-DR-/low MDSCs isolated from tumor tissue were observed to inhibit the CD4+ T-cells' immune responses in comparison with those from the adjacent normal tissue. Furthermore, VIP was able to induce the differentiation of CD14+ mononuclear cells isolated from healthy donor peripheral blood mononuclear cells into activated MDSC cells. Of note, the immunosuppressive effect of VIP-induced CD14+HLA-DR(-/low) MDSCs on CD4+ T cells was mediated by IL-10 secretion, which was demonstrated in the subsequent decrease of IFN-γ and IL-2 production. In conclusion, CD14+HLA-DR(-/low) cells were significantly increased in gastric

  9. Impaired Cd14 and Cd36 expression, bacterial clearance, and Toll-like receptor 4-Myd88 signaling in caveolin-1-deleted macrophages and mice.

    PubMed

    Tsai, Tsung-Huang; Chen, Shu-Fen; Huang, Tai-Yu; Tzeng, Chun-Fu; Chiang, Ann-Shyn; Kou, Yu Ru; Lee, Tzong-Shyuan; Shyue, Song-Kun

    2011-01-01

    An overwhelming immune response, particularly from macrophages, with gram-negative bacteria-induced sepsis plays a critical role in survival of and organ damage in infected patients. Caveolin-1 (Cav-1), a major structure protein of caveolae, regulates many cellular functions. We examined the vital role of Cav-1 in the response of macrophages and mice to bacteria or LPS exposure. Deletion of Cav-1 decreased the expression of CD14 and CD36 during macrophage differentiation and suppressed their phagocytotic ability. As well, the ability to kill bacteria was inhibited in Cav-1 macrophages and mice peritoneal cavity, tissue, and plasma, which was partly attributed to hindered expression of iNOS induced by bacteria or LPS. Furthermore, deletion of Cav-1 attenuated the expression of Toll-like receptor 4 and myeloid differentiation factor 88 and the activation of nuclear factor κB, all of which impeded the production of inflammatory cytokines in response to bacterial exposure in Cav-1 macrophages and mice. Thus, Cav-1 participates in the regulation of CD14, CD36, Toll-like receptor 4 and myeloid differentiation factor 88 protein expression and is crucial for the immune response of macrophages to bacterial infection. Cav-1 may be a therapeutic target in the treatment of sepsis.

  10. The plasma levels of soluble ST2 as a marker of gut mucosal damage in early HIV infection

    PubMed Central

    Mehraj, Vikram; Jenabian, Mohammad-Ali; Ponte, Rosalie; Lebouché, Bertrand; Costiniuk, Cecilia; Thomas, Réjean; Baril, Jean-Guy; LeBlanc, Roger; Cox, Joseph; Tremblay, Cécile; Routy, Jean-Pierre

    2016-01-01

    Objective: Following tissue barrier breaches, interleukin-33 (IL-33) is released as an ‘alarmin’ to induce inflammation. Soluble suppression of tumorigenicity 2 (sST2), as an IL-33 decoy receptor, contributes to limit inflammation. We assessed the relationship between the IL-33/ST2 axis and markers of gut mucosal damage in patients with early (EHI) and chronic HIV infection (CHI) and elite controllers. Design: Analyses on patients with EHI and CHI were conducted to determine IL-33/sST2 changes over time. Methods: IL-33 and sST2 levels were measured in plasma. Correlations between sST2 levels and plasma viral load, CD4+ and CD8+ T-cell counts, expression of T-cell activation/exhaustion markers, gut mucosal damage, microbial translocation and inflammation markers, as well as kynurenine/tryptophan ratio were assessed. Results: Plasma sST2 levels were elevated in EHI compared with untreated CHI and uninfected controls, whereas IL-33 levels were comparable in all groups. In EHI, sST2 levels were positively correlated with the CD8+ T-cell count and the percentage of T cells expressing activation and exhaustion markers, but not with viral load or CD4+ T-cell count. Plasma sST2 levels also correlated with plasma levels of gut mucosal damage, microbial translocation and kynurenine/tryptophan ratio and for some markers of inflammation. Prospective analyses showed that early antiretroviral therapy had no impact on sST2 levels, whereas longer treatment duration initiated during CHI normalized sST2. Conclusion: As sST2 levels were elevated in EHI and were correlated with CD8+ T-cell count, immune activation, and microbial translocation, sST2 may serve as a marker of disease progression, gut damage and may directly contribute to HIV pathogenesis. PMID:27045377

  11. Lutein supplementation leads to decreased soluble complement membrane attack complex sC5b-9 plasma levels.

    PubMed

    Tian, Yuan; Kijlstra, Aize; van der Veen, Rob L P; Makridaki, Maria; Murray, Ian J; Berendschot, Tos T J M

    2015-03-01

    The purpose of this study was to investigate the effect of lutein on systemic complement activation in elderly individuals. Seventy patients with signs of early age-related macular degeneration (AMD) were included in this study. All subjects were randomly assigned to receive a 10 mg daily dose of lutein or a placebo for a time period of 1 year. EDTA blood was collected before and at various time-points during the study (0, 4, 8 and 12 months). The plasma level of the soluble complement membrane attack complex sC5b-9 was measured by ELISA. We found a significant 1.1 ng/ml monthly decrease in the plasma sC5b-9 concentration in the lutein group (p<0.001), resulting in a decrease from 60.3 ng/ml at baseline to 46.3 ng/ml at 12 months. For the placebo group, we found a significant 0.6 ng/ml monthly increase in plasma sC5b-9 concentration (p=0.001), resulting in an increase from 51.6 ng/ml at baseline to 58.4 ng/ml at 12 months. Lutein supplementation inhibits the systemic activation of the complement system, which provides further functional evidence for the reported beneficial effects of this carotenoid in the management of AMD. © 2014 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  12. Preliminary study of the effects of condensed barley distillers soluble on rumen fermentation and plasma metabolites in Japanese Black cows.

    PubMed

    Tsuruoka, Katsuhiko; Kanamaru, Hidenobu; Takahashi, Hideyuki; Gotoh, Takafumi

    2017-04-01

    In Japan, condensed barley distillers soluble (CBDS) is a widely known liquor byproduct that contains a high level of protein and is used as a supplementary protein feed for cattle. The present study evaluated the effects of CBDS feed on rumen fermentation and plasma metabolites in Japanese Black cows. Applying a replicated 3 × 3 Latin square design, nine cows were offered CBDS and hay (CBDS-t), soy bean meal and hay (Soybean-t) and only hay (Hay-t) over 35 days. We collected ruminal fluid and plasma just before feeding and at 3 h after feeding. The concentrations of propionate and butyrate in the rumen before feeding were lower in the CBDS-t than in the Soybean-t group (P < 0.05). However, after 3 h, the concentrations were higher in the CBDS-t than in the Soybean-t and Hay-t groups (P < 0.05). Although, there were no differences in the compositions (% mol) of propionate and butyrate in the rumen and the concentration of plasma β-hydroxybutyric acid before feeding between treatments, after 3 h they were significantly higher in the CBDS-t than in the Soybean-t and Hay-t groups (P < 0.05). These results indicate that feeding CBDS promotes rumen fermentation and butyrate metabolism. © 2016 Japanese Society of Animal Science.

  13. Plasma interleukin-6 and soluble IL-6 receptors are associated with psychological well-being in aging women.

    PubMed

    Friedman, Elliot M; Hayney, Mary; Love, Gayle D; Singer, Burton H; Ryff, Carol D

    2007-05-01

    This study tested the hypothesis that psychological well-being would predict lower plasma levels of inflammatory factors in aging women. One hundred thirty-five women ages 61-91 years (M = 74.5 years) participated in this study. After completing self-administered questionnaires in their homes, participants stayed overnight at the General Clinical Research Center (GCRC) at the University of Wisconsin-Madison. Blood samples for cytokine analyses were obtained in participants' homes after the GCRC visit. Psychological well-being and ill-being, history of health problems, and health behaviors were assessed via self-administered questionnaires. Detailed medical history and concurrent health measures were obtained during the GCRC stay. Enzyme-linked immunosorbent assays were used to determine interleukin-6 (IL-6) and soluble IL-6 receptor (sIL-6R) concentrations in plasma. Regression analyses showed that plasma IL-6 levels were lower in women scoring higher on positive relationships, whereas sIL-6R levels were lower in women scoring higher on purpose in life, even after a variety of sociodemographic and health factors were controlled. These outcomes, combined with the absence of significant links with other measures of well-being and ill-being, suggest selective patterns of association between later life inflammatory processes and psychological factors, particularly those focused on positive ties with others and purposeful engagement. Copyright (c) 2007 APA, all rights reserved.

  14. Solution NMR studies provide structural basis for endotoxin pattern recognition by the innate immune receptor CD14

    SciTech Connect

    Albright, Seth; Chen Bin; Holbrook, Kristen; Jain, Nitin U.

    2008-04-04

    CD14 functions as a key pattern recognition receptor for a diverse array of Gram-negative and Gram-positive cell-wall components in the host innate immune response by binding to pathogen-associated molecular patterns (PAMPs) at partially overlapping binding site(s). To determine the potential contribution of CD14 residues in this pattern recognition, we have examined using solution NMR spectroscopy, the binding of three different endotoxin ligands, lipopolysaccharide, lipoteichoic acid, and a PGN-derived compound, muramyl dipeptide to a {sup 15}N isotopically labeled 152-residue N-terminal fragment of sCD14 expressed in Pichia pastoris. Mapping of NMR spectral changes upon addition of ligands revealed that the pattern of residues affected by binding of each ligand is partially similar and partially different. This first direct structural observation of the ability of specific residue combinations of CD14 to differentially affect endotoxin binding may help explain the broad specificity of CD14 in ligand recognition and provide a structural basis for pattern recognition. Another interesting finding from the observed spectral changes is that the mode of binding may be dynamically modulated and could provide a mechanism for binding endotoxins with structural diversity through a common binding site.

  15. Intermediate CD14(++)CD16(+) monocyte predicts severe coronary stenosis and extensive plaque involvement in asymptomatic individuals.

    PubMed

    Lo, Shyh-Chyi; Lee, Wen-Jeng; Chen, Ching-Yi; Lee, Bai-Chin

    2017-02-26

    Circulating leukocyte subtypes and monocyte subsets are independent predictors of cardiovascular events. We hypothesized that an increased leukocyte subtype would predict severe coronary stenosis and extensive plaque involvement. We retrospectively analyzed clinical, laboratory, and coronary CT data in a total of 588 asymptomatic adults (69% men; mean age, 57 ± 9 years) undergoing a general health check-up. Intermediate CD14(++)CD16(+) monocyte count had the strongest association with mixed and calcified plaque scores, whereas the numbers of neutrophils and classical CD14(++)CD16(-) monocytes were significantly associated with non-calcified plaque score. Only high CD14(++)CD16(+) monocyte count (>12 cells/μL) significantly predicted extensive plaque involvement [odds ratio 3.16 (95% confidence interval 1.84-5.43), P < 0.001; quartile 4 vs. 1-3] and severe coronary stenosis [3.67 (1.84-7.33), P < 0.001; quartile 4 vs. 1-3] after adjustments for Framingham Risk Score (FRS), metabolic syndrome, and C-reactive protein. The CD14(++)CD16(+) monocyte count, when added to FRS, significantly reclassified 30.4 and 26.7% of the overall and 50.2 and 36.2% of the intermediate-risk population (FRS 6-20%) for predicting extensive plaque involvement and severe coronary stenosis, respectively. Thus, in asymptomatic individuals, intermediate CD14(++)CD16(+) monocyte could independently predict severe CAD and improve risk stratification.

  16. CD14(++) CD16(-) monocytes are the main source of 11β-HSD type 1 after IL-4 stimulation.

    PubMed

    Kunnathully, Vidya; Gomez-Lira, Macarena; Bassi, Giulio; Poli, Fabio; Zoratti, Elisa; La Verde, Valentina; Idolazzi, Luca; Gatti, Davide; Viapiana, Ombretta; Adami, Silvano; Rossini, Maurizio

    2017-02-01

    The anti-inflammatory actions of IL-4 are well established through earlier findings. However, the exact mechanism it uses to downregulate the pro-inflammatory cytokine production through monocytes and macrophages is poorly understood. In this study, we examined the effect of IL-4 in the induction of 11β-HSD1 in the two main classes of monocytes, CD14(++) CD16(-) (CD14) and CD14(+) CD16(+) (CD16). Peripheral Blood Mononuclear Cells (PBMCs) were isolated from 17 healthy donors and were sorted into CD14 and CD16 subpopulations using cell sorting. Effect of IL-4 on 11β-HSD1-enzyme activity was measured in sorted and unsorted monocytes using Homogeneous Time-Resolved Fluorescence (HTRF) and M1/M2 polarization analysis was performed by flow cytometry. Our results indicate that CD14 cells are the major source of 11β-HSD1 enzyme after IL-4 stimulation and that M2 phenotype is not a pre-requisite for its synthesis.

  17. Signaling of high mobility group box 1 (HMGB1) through toll-like receptor 4 in macrophages requires CD14.

    PubMed

    Kim, Sodam; Kim, Sun Young; Pribis, John P; Lotze, Michael; Mollen, Kevin P; Shapiro, Richard; Loughran, Patricia; Scott, Melanie J; Billiar, Timothy R

    2013-05-20

    High mobility group box 1 (HMGB1) is a DNA-binding protein that possesses cytokinelike, proinflammatory properties when released extracellularly in the C23-C45 disulfide form. HMGB1 also plays a key role as a mediator of acute and chronic inflammation in models of sterile injury. Although HMGB1 interacts with multiple pattern recognition receptors (PRRs), many of its effects in injury models occur through an interaction with toll-like receptor 4 (TLR4). HMGB1 interacts directly with the TLR4/myeloid differentiation protein 2 (MD2) complex, although the nature of this interaction remains unclear. We demonstrate that optimal HMGB1-dependent TLR4 activation in vitro requires the coreceptor CD14. TLR4 and MD2 are recruited into CD14-containing lipid rafts of RAW264.7 macrophages after stimulation with HMGB1, and TLR4 interacts closely with the lipid raft protein GM1. Furthermore, we show that HMGB1 stimulates tumor necrosis factor (TNF)-α release in WT but not in TLR4(-/-), CD14(-/-), TIR domain-containing adapter-inducing interferon-β (TRIF)(-/-) or myeloid differentiation primary response protein 88 (MyD88)(-/-) macrophages. HMGB1 induces the release of monocyte chemotactic protein 1 (MCP-1), interferon gamma-induced protein 10 (IP-10) and macrophage inflammatory protein 1α (MIP-1α) in a TLR4- and CD14-dependent manner. Thus, efficient recognition of HMGB1 by the TLR4/MD2 complex requires CD14.

  18. The “Intermediate” CD14++CD16+ monocyte subset increases in severe peripheral artery disease in humans

    PubMed Central

    Wildgruber, Moritz; Aschenbrenner, Teresa; Wendorff, Heiko; Czubba, Maria; Glinzer, Almut; Haller, Bernhard; Schiemann, Matthias; Zimmermann, Alexander; Berger, Hermann; Eckstein, Hans-Henning; Meier, Reinhard; Wohlgemuth, Walter A.; Libby, Peter; Zernecke, Alma

    2016-01-01

    Monocytes are key players in atherosclerotic. Human monocytes display a considerable heterogeneity and at least three subsets can be distinguished. While the role of monocyte subset heterogeneity has already been well investigated in coronary artery disease (CAD), the knowledge about monocytes and their heterogeneity in peripheral artery occlusive disease (PAOD) still is limited. Therefore, we aimed to investigate monocyte subset heterogeneity in patients with PAOD. Peripheral blood was obtained from 143 patients suffering from PAOD (Rutherford stage I to VI) and three monocyte subsets were identified by flow cytometry: CD14++CD16− classical monocytes, CD14+CD16++ non-classical monocytes and CD14++CD16+ intermediate monocytes. Additionally the expression of distinct surface markers (CD106, CD162 and myeloperoxidase MPO) was analyzed. Proportions of CD14++CD16+ intermediate monocyte levels were significantly increased in advanced stages of PAOD, while classical and non-classical monocytes displayed no such trend. Moreover, CD162 and MPO expression increased significantly in intermediate monocyte subsets in advanced disease stages. Likewise, increased CD162 and MPO expression was noted in CD14++CD16− classical monocytes. These data suggest substantial dynamics in monocyte subset distributions and phenotypes in different stages of PAOD, which can either serve as biomarkers or as potential therapeutic targets to decrease the inflammatory burden in advanced stages of atherosclerosis. PMID:27991581

  19. Porcine CD14 gene silencing partially inhibited the bacterial immune response mediated by TLR4 signaling pathway.

    PubMed

    Dai, Chaohui; Wang, Haifei; Zhu, Guoqiang; Wu, Shenglong; Bao, Wenbin

    2017-09-10

    Cluster of differentiation antigen 14 (CD14) is the membrane receptor protein in Toll-like Receptor 4 (TLR4) signaling pathway, which plays an important regulation role in not only innate immune response but also adaptive immune response. In this study, the pig kidney epithelial cell (PK15) line with CD14 gene silencing mediated by lentivirus was established and cells of CD14-RNAi and NC group were exposed to three kinds of Escherichia coli (E. coli F18ab, E. coli F18ac and E. coli K88ac) and LPS. Then qPCR and western blot were used to detect expression levels of TLR4 signaling pathway-related genes. Finally, ELISA was used to detect the level of proinflammatory cytokines in the cell culture supernatant. The results showed that the expression level of TLR4 signaling pathway-related genes in the entire signal pathway had obvious increases when cells were exposed to the stimulation induced by E. coli and LPS. In addition, the expression levels of CD14-RNAi group were overall significantly lower than NC group (P<0.05 or P<0.01), which was the same with the release levels of proinflammatory cytokines. This study revealed that pig CD14 gene silencing partially inhibited immune response to E. coli F18 invasion mediated by TLR4 signaling pathway. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. The effects of electrical stunning voltage on meat quality, plasma parameters, and protein solubility of broiler breast meat.

    PubMed

    Huang, J C; Yang, J; Huang, M; Chen, K J; Xu, X L; Zhou, G H

    2017-03-01

    This study was designed to compare the effects of different stunning voltages of pulsed direct current on meat quality of broilers. For this purpose, plasma parameters, blood loss, carcass damage, and meat water holding capacity, color, shear force, pH, and protein solubility were analyzed. A total of 400 broilers were divided into 5 treatment groups and stunned with 5, 15, 25, 35, and 45 V at 750 Hz and 10 s, respectively. Blood samples were collected immediately after cutting the neck. Pectoralis major muscles were removed from the carcass after chilling and placed on ice. Breast muscle pH and meat color were determined at both 2 and 24 h postmortem. Dripping loss, cooking loss, pressing loss, and cooked breast meat shear values were determined after 24 h postmortem. The 5 V treatment significantly increased (P < 0.05) blood plasma corticosterone and lactate concentration compared with the other groups. The carcass damage of wings, Pectoralis major, and Pectoralis minor was significant in the 5, 35, and 45 V groups. The pH of 2 h postmortem in the 5 and 45 V groups was significantly lower (P < 0.05) than in the 15 and 25 V groups. In the 5 and 45 V groups, the protein solubility and shear force value were significantly lower (P < 0.05) and dripping loss was significantly higher (P < 0.05) than the other groups. © 2016 Poultry Science Association Inc.

  1. Plasma soluble vascular adhesion molecule-1 levels are persistently elevated during the first month after colorectal cancer resection.

    PubMed

    Shantha Kumara, H M C; Tohme, Samer T; Herath, Sonali A C; Yan, Xiaohong; Senagore, Anthony J; Nasar, Abu; Kalady, Matthew F; Baxter, Raymond; Whelan, Richard L

    2012-06-01

    Plasma from the second and third weeks after minimally invasive colorectal resection (MICR) has high levels of the proangiogenic proteins VEGF and angiopoietin 2 and also stimulates, in vitro, endothelial cell (EC) proliferation and migration, which are critical to wound and tumor angiogenesis. Soluble vascular cell adhesion molecule-1 (sVCAM-1) stimulates EC chemotaxis and angiogenesis. The impact of MICR on blood levels of sVCAM-1 is unknown. This study's purpose was to determine plasma sVCAM-1 levels after MICR in colorectal cancer (CRC) patients. Blood samples from 90 patients (26% rectal, 74% colon) were obtained preoperatively, on postoperative days (POD) 1 and 3, and at other points during the next 2 months. The late samples were bundled into 7-day time blocks. sVCAM-1 levels were determined in duplicate via ELISA and reported as ng/ml. Student's t test was used for data analysis (significance, P < 0.008 after Bonferroni correction). The mean incision length was 7.3 ± 3.1 cm, and the conversion rate was 3%. Compared with preoperative (PreOp) levels (811.3 ± 233.2), the mean plasma sVCAM-1 level was significantly higher on POD 1 (905.7 ± 292.4, P < 0.001) and POD 3 (977.7 ± 271.8, P < 0.001). Levels remained significantly elevated for the POD 7-13, POD 14-20, POD 21-27, and POD 28-67 time blocks. MICR for CRC is associated with a persistent increase in plasma sVCAM-1 levels during the first month. This sustained increase may promote angiogenesis and stimulate the growth of residual tumor cells early after surgery.

  2. Dopamine Increases CD14(+)CD16(+) Monocyte Transmigration across the Blood Brain Barrier: Implications for Substance Abuse and HIV Neuropathogenesis.

    PubMed

    Calderon, Tina M; Williams, Dionna W; Lopez, Lillie; Eugenin, Eliseo A; Cheney, Laura; Gaskill, Peter J; Veenstra, Mike; Anastos, Kathryn; Morgello, Susan; Berman, Joan W

    2017-01-29

    In human immunodeficiency virus-1 (HIV) infected individuals, substance abuse may accelerate the development and/or increase the severity of HIV associated neurocognitive disorders (HAND). It is proposed that CD14(+)CD16(+) monocytes mediate HIV entry into the central nervous system (CNS) and that uninfected and infected CD14(+)CD16(+) monocyte transmigration across the blood brain barrier (BBB) contributes to the establishment and propagation of CNS HIV viral reservoirs and chronic neuroinflammation, important factors in the development of HAND. The effects of substance abuse on the frequency of CD14(+)CD16(+) monocytes in the peripheral circulation and on the entry of these cells into the CNS during HIV neuropathogenesis are not known. PBMC from HIV infected individuals were analyzed by flow cytometry and we demonstrate that the frequency of peripheral blood CD14(+)CD16(+) monocytes in HIV infected substance abusers is increased when compared to those without active substance use. Since drug use elevates extracellular dopamine concentrations in the CNS, we examined the effects of dopamine on CD14(+)CD16(+) monocyte transmigration across our in vitro model of the human BBB. The transmigration of this monocyte subpopulation is increased by dopamine and the dopamine receptor agonist, SKF 38393, implicating D1-like dopamine receptors in the increase in transmigration elicited by this neurotransmitter. Thus, elevated extracellular CNS dopamine may be a novel common mechanism by which active substance use increases uninfected and HIV infected CD14(+)CD16(+) monocyte transmigration across the BBB. The influx of these cells into the CNS may increase viral seeding and neuroinflammation, contributing to the development of HIV associated neurocognitive impairments.

  3. CD14 dependence of TLR4 endocytosis and TRIF signaling displays ligand specificity and is dissociable in endotoxin tolerance.

    PubMed

    Rajaiah, Rajesh; Perkins, Darren J; Ireland, Derek D C; Vogel, Stefanie N

    2015-07-07

    Dimerization of Toll-like receptor 4 (TLR4)/myeloid differentiation factor 2 (MD2) heterodimers is critical for both MyD88- and TIR-domain-containing adapter-inducing IFN-β (TRIF)-mediated signaling pathways. Recently, Zanoni et al. [(2011) Cell 147(4):868-880] reported that cluster of differentiation 14 (CD14) is required for LPS-/Escherichia coli- induced TLR4 internalization into endosomes and activation of TRIF-mediated signaling in macrophages. We confirmed their findings with LPS but report here that CD14 is not required for receptor endocytosis and downstream signaling mediated by TLR4/MD2 agonistic antibody (UT12) and synthetic small-molecule TLR4 ligands (1Z105) in murine macrophages. CD14 deficiency completely ablated the LPS-induced TBK1/IRF3 signaling axis that mediates production of IFN-β in murine macrophages without affecting MyD88-mediated signaling, including NF-κB, MAPK activation, and TNF-α and IL-6 production. However, neither the MyD88- nor TRIF-signaling pathways and their associated cytokine profiles were altered in the absence of CD14 in UT12- or 1Z105-treated murine macrophages. Eritoran (E5564), a lipid A antagonist that binds the MD2 "pocket," completely blocked LPS- and 1Z105-driven, but not UT12-induced, TLR4 dimerization and endocytosis. Furthermore, TLR4 endocytosis is induced in macrophages tolerized by exposure to either LPS or UT12 and is independent of CD14. These data indicate that TLR4 receptor endocytosis and the TRIF-signaling pathway are dissociable and that TLR4 internalization in macrophages can be induced by UT12, 1Z105, and during endotoxin tolerance in the absence of CD14.

  4. CD14 dependence of TLR4 endocytosis and TRIF signaling displays ligand specificity and is dissociable in endotoxin tolerance

    PubMed Central

    Rajaiah, Rajesh; Perkins, Darren J.; Ireland, Derek D. C.; Vogel, Stefanie N.

    2015-01-01

    Dimerization of Toll-like receptor 4 (TLR4)/myeloid differentiation factor 2 (MD2) heterodimers is critical for both MyD88- and TIR-domain–containing adapter-inducing IFN-β (TRIF)-mediated signaling pathways. Recently, Zanoni et al. [(2011) Cell 147(4):868–880] reported that cluster of differentiation 14 (CD14) is required for LPS-/Escherichia coli- induced TLR4 internalization into endosomes and activation of TRIF-mediated signaling in macrophages. We confirmed their findings with LPS but report here that CD14 is not required for receptor endocytosis and downstream signaling mediated by TLR4/MD2 agonistic antibody (UT12) and synthetic small-molecule TLR4 ligands (1Z105) in murine macrophages. CD14 deficiency completely ablated the LPS-induced TBK1/IRF3 signaling axis that mediates production of IFN-β in murine macrophages without affecting MyD88-mediated signaling, including NF-κB, MAPK activation, and TNF-α and IL-6 production. However, neither the MyD88- nor TRIF-signaling pathways and their associated cytokine profiles were altered in the absence of CD14 in UT12- or 1Z105-treated murine macrophages. Eritoran (E5564), a lipid A antagonist that binds the MD2 “pocket,” completely blocked LPS- and 1Z105-driven, but not UT12-induced, TLR4 dimerization and endocytosis. Furthermore, TLR4 endocytosis is induced in macrophages tolerized by exposure to either LPS or UT12 and is independent of CD14. These data indicate that TLR4 receptor endocytosis and the TRIF-signaling pathway are dissociable and that TLR4 internalization in macrophages can be induced by UT12, 1Z105, and during endotoxin tolerance in the absence of CD14. PMID:26106158

  5. Plasma and cerebrospinal fluid biomarkers predict cerebral injury in HIV-infected individuals on stable combination antiretroviral therapy

    PubMed Central

    Anderson, Albert M.; Harezlak, Jaroslaw; Bharti, Ajay; Mi, Deming; Taylor, Michael J.; Daar, Eric S.; Schifitto, Giovanni; Zhong, Jianhui; Alger, Jeffry R.; Brown, Mark S.; Singer, Elyse J.; Campbell, Thomas B.; McMahon, Deborah D.; Buchthal, Steven; Cohen, Ronald; Yiannoutsos, Constantin; Letendre, Scott L.; Navia, Bradford A.

    2015-01-01

    Objectives HIV-associated brain injury persists despite antiretroviral therapy (cART), but contributing factors remain poorly understood. We postulated that inflammation-associated biomarkers will be associated with cerebral injury on proton magnetic resonance spectroscopy (MRS) in chronically HIV-infected subjects. Methods Five biomarkers were measured in 197 HIV-infected subjects: soluble CD14, MCP-1, IP-10, MIP-1β, and fractalkine. Levels of N-acetyl aspartate (NAA), Choline (Cho), Myoinositol (MI), Glutamate+Glutamine (Glx), and Creatine (Cr) were acquired in the midfrontal cortex (MFC), frontal white matter (FWM), and basal ganglia (BG). Predictive models were built via linear regression and the best models were chosen using the Akaike Information Criterion. Results Increases in plasma or CSF MCP-1 were associated with lower NAA/Cr in the MFC and BG while metabolite changes in the FWM for NAA/Cr, GlxCr and Cho/Cr were explained almost exclusively by a single factor, sCD14. Plasma and CSF levels of this factor were also significantly associated with Glx/Cr in MFC and BG. Higher CSF FKN was associated with higher NAA/Cr in BG. Best predictors for higher Cho/Cr in BG and MFC were CSF sCD14 and CSF MIP-1β. Plasma and CSF IP-10 were only associated with Cho/Cr in MFC. Of the three models that simultaneously accounted for both plasma and CSF, there were more associations between CSF biomarkers and MRS metabolites. Conclusions Markers of inflammation and immune activation, in particular MCP-1 and sCD14, predominantly reflecting CNS sources, contribute to the persistence of brain injury in a metabolite and region dependent manner in chronically HIV-infected patients on stable cART. PMID:25622053

  6. Vitamin D effects on monocytes' CCL-2, IL6 and CD14 transcription in Addison's disease and HLA susceptibility.

    PubMed

    Kraus, A U; Penna-Martinez, M; Meyer, G; Badenhoop, K

    2017-07-29

    Addison's disease is a rare autoimmune disorder leading to adrenal insufficiency and life-long glucocorticoid dependency. Vitamin D receptor (VDR) polymorphisms and vitamin D deficiency predispose to Addison's disease. Aim of the current study was, to investigate potential anti-inflammatory vitamin D effects on monocytes in Addison's disease, focusing on inflammatory CCL-2 and IL6, as well on monocyte CD14 markers. Addison's disease is genetically linked to distinct HLA susceptibility alleles. Therefore we analyzed, whether HLA genotypes differed for vitamin D effects on monocyte markers. CD14(+) monocytes were isolated from Addison's disease patients (AD, n=13) and healthy controls (HC, n=15) and stimulated with 1,25-dihydroxyvitamin D3 and IL1β as an inflammatory stimulant. Cells were processed for mRNA expression of CCL-2, IL6 and CD14 and DNA samples were genotyped for major histocompatibility class (MHC) class II-encoded HLA- DQA1-DQB1 haplotypes. We found a downregulation of CCL-2 after vitamin D treatment in IL1β-stimulated monocytes both from AD patients and HC (AD p<0.001; HC p<0.0001). CD14 expression however, was upregulated in both HC and AD patients after vitamin D treatment (p<0.001, respectively). HC showed higher CD14 transcription level than AD patients after vitamin D treatment (p=0.04). Compared to IL1β-induced inflammation, HC have increased CD14 levels after vitamin D treatment (p<0.001), whereas the IL1β-induced CD14 expression of AD patients' monocytes did not change after vitamin D treatment (p=0.8). AD patients carrying HLA high-risk haplotypes showed an increased CCL-2 expression after IL1β-induced inflammation compared to intermediate-risk HLA carriers (p=0.05). Also HC monocytes' CD14 transcription after IL1β and vitamin D co-stimulation differed according to HLA risk profile. We show that vitamin D can exert anti-inflammatory effects on AD patients' monocytes which may be modulated by HLA risk genotypes. Copyright © 2017 Elsevier

  7. Plasma levels of soluble HLA-E and HLA-F at diagnosis may predict overall survival of neuroblastoma patients.

    PubMed

    Morandi, Fabio; Cangemi, Giuliana; Barco, Sebastiano; Amoroso, Loredana; Giuliano, Maria; Gigliotti, Anna Rita; Pistoia, Vito; Corrias, Maria Valeria

    2013-01-01

    The purpose of this study was to identify the plasma/serum biomarkers that are able to predict overall survival (OS) of neuroblastoma (NB) patients. Concentration of soluble (s) biomarkers was evaluated in plasma (sHLA-E, sHLA-F, chromogranin, and B7H3) or serum (calprotectin) samples from NB patients or healthy children. The levels of biomarkers that were significantly higher in NB patients were then analyzed considering localized or metastatic subsets. Finally, biomarkers that were significantly different in these two subsets were correlated with patient's outcome. With the exception of B7H3, levels of all molecules were significantly higher in NB patients than those in controls. However, only chromogranin, sHLA-E, and sHLA-F levels were different between patients with metastatic and localized tumors. sHLA-E and -F levels correlated with each other but not chromogranin. Chromogranin levels correlated with different event-free survival (EFS), whereas sHLA-E and -F levels also correlated with different OS. Association with OS was also detected considering only patients with metastatic disease. In conclusion, low levels of sHLA-E and -F significantly associated with worse EFS/OS in the whole cohort of NB patients and in patients with metastatic NB. Thus, these molecules deserve to be tested in prospective studies to evaluate their predictive power for high-risk NB patients.

  8. Genome-wide association study identifies polymorphisms in LEPR as determinants of plasma soluble leptin receptor levels

    PubMed Central

    Sun, Qi; Cornelis, Marilyn C.; Kraft, Peter; Qi, Lu; van Dam, Rob M.; Girman, Cynthia J.; Laurie, Cathy C.; Mirel, Daniel B.; Gong, Huizi; Sheu, Chau-Chyun; Christiani, David C.; Hunter, David J.; Mantzoros, Christos S.; Hu, Frank B.

    2010-01-01

    Plasma soluble leptin receptor (sOB-R) levels were inversely associated with diabetes risk factors, including adiposity and insulin resistance, and highly correlated with the expression levels of leptin receptor, which is ubiquitously expressed in most tissues. We conducted a genome-wide association study of sOB-R in 1504 women of European ancestry from the Nurses' Health Study. The initial scan yielded 26 single nucleotide polymorphisms (SNPs) significantly associated with sOB-R levels (P < 5 × 10−8); all mapping to the leptin receptor gene (LEPR). Analysis of imputed genotypes on autosomal chromosomes revealed an additional 106 SNPs in and adjacent to this gene that reached genome-wide significance level. Of these 132 SNPs (including two non-synonymous SNPs, rs1137100 and rs1137101), rs2767485, rs1751492 and rs4655555 remained associated with sOB-R levels at the 0.05 level (P = 9.1 × 10−9, 0.0105 and 0.0267, respectively) after adjustment for other univariately associated SNPs in a forward selection procedure. Significant associations with these SNPs were replicated in an independent sample of young males (n = 875) residing in Cyprus (P < 1 × 10−4). These data provide novel evidence revealing the role of polymorphisms in LEPR in modulating plasma levels of sOB-R and may further our understanding of the complex relationships among leptin, leptin receptor and diabetes-related traits. PMID:20167575

  9. Plasma soluble erythropoietin receptor is decreased during sleep in Andean highlanders with Chronic Mountain Sickness.

    PubMed

    Villafuerte, Francisco C; Corante, Noemí; Anza-Ramírez, Cecilia; Figueroa-Mujíca, Rómulo; Vizcardo-Galindo, Gustavo; Mercado, Andy; Macarlupú, José Luis; León-Velarde, Fabiola

    2016-07-01

    Excessive erythrocytosis (EE) is the main sign of Chronic Mountain Sickness (CMS), a highly prevalent syndrome in Andean highlanders. Low pulse O2 saturation (SpO2) during sleep and serum androgens have been suggested to contribute to EE in CMS patients. However, whether these factors have a significant impact on the erythropoietin (Epo) system leading to EE is still unclear. We have recently shown that morning soluble Epo receptor (sEpoR), an endogenous Epo antagonist, is decreased in CMS patients suggesting increased Epo availability (increased Epo/sEpoR). The present study aimed to characterize the nocturnal concentration profile of sEpoR and Epo and their relationship with SpO2, Hct, and serum testosterone in healthy highlanders (HH) and CMS patients. Epo and sEpoR concentrations were evaluated every 4 h (6 PM to 6 AM) and nighttime SpO2 was continuously monitored (10 PM to 6 AM) in 39 male participants (CMS, n = 23; HH, n = 16) aged 21-65 yr from Cerro de Pasco, Peru (4,340 m). CMS patients showed higher serum Epo concentrations throughout the night and lower sEpoR from 10 PM to 6 AM. Consequently, Epo/sEpoR was significantly higher in the CMS group at every time point. Mean sleep-time SpO2 was lower in CMS patients compared with HH, while the percentage of sleep time spent with SpO2 < 80% was higher. Multiple-regression analysis showed mean sleep-time SpO2 and Epo/sEpoR as significant predictors of hematocrit corrected for potential confounders (age, body mass index, and testosterone). Testosterone levels were associated neither with Hct nor with erythropoietic factors. In conclusion, our results show sustained erythropoietic stimulus driven by the Epo system in CMS patients, further enhanced by a continuous exposure to accentuated nocturnal hypoxemia.

  10. Plasma soluble erythropoietin receptor is decreased during sleep in Andean highlanders with Chronic Mountain Sickness

    PubMed Central

    Corante, Noemí; Anza-Ramírez, Cecilia; Figueroa-Mujíca, Rómulo; Vizcardo-Galindo, Gustavo; Mercado, Andy; Macarlupú, José Luis; León-Velarde, Fabiola

    2016-01-01

    Excessive erythrocytosis (EE) is the main sign of Chronic Mountain Sickness (CMS), a highly prevalent syndrome in Andean highlanders. Low pulse O2 saturation (SpO2) during sleep and serum androgens have been suggested to contribute to EE in CMS patients. However, whether these factors have a significant impact on the erythropoietin (Epo) system leading to EE is still unclear. We have recently shown that morning soluble Epo receptor (sEpoR), an endogenous Epo antagonist, is decreased in CMS patients suggesting increased Epo availability (increased Epo/sEpoR). The present study aimed to characterize the nocturnal concentration profile of sEpoR and Epo and their relationship with SpO2, Hct, and serum testosterone in healthy highlanders (HH) and CMS patients. Epo and sEpoR concentrations were evaluated every 4 h (6 PM to 6 AM) and nighttime SpO2 was continuously monitored (10 PM to 6 AM) in 39 male participants (CMS, n = 23; HH, n = 16) aged 21-65 yr from Cerro de Pasco, Peru (4,340 m). CMS patients showed higher serum Epo concentrations throughout the night and lower sEpoR from 10 PM to 6 AM. Consequently, Epo/sEpoR was significantly higher in the CMS group at every time point. Mean sleep-time SpO2 was lower in CMS patients compared with HH, while the percentage of sleep time spent with SpO2 < 80% was higher. Multiple-regression analysis showed mean sleep-time SpO2 and Epo/sEpoR as significant predictors of hematocrit corrected for potential confounders (age, body mass index, and testosterone). Testosterone levels were associated neither with Hct nor with erythropoietic factors. In conclusion, our results show sustained erythropoietic stimulus driven by the Epo system in CMS patients, further enhanced by a continuous exposure to accentuated nocturnal hypoxemia. PMID:27125843

  11. Lung carcinomas decrease the number of monocytes/macrophages (CD14+ cells) that produce TNF-alpha.

    PubMed

    Lopez-Gonzalez, Jose Sullivan; Avila-Moreno, Federico; Prado-Garcia, Heriberto; Aguilar-Cazares, Dolores; Mandoki, Juan Jose; Meneses-Flores, Manuel

    2007-03-01

    The role that inflammation plays in cancer is puzzling. In peripheral blood, TNF-alpha-producing monocytes (CD14+ cells) were compared among patients with lung cancer, patients with tuberculosis and healthy donors; also, in pleural effusion TNF-alpha-producing CD14+ cells were compared between tuberculous patients and lung cancer patients. To analyze the level of the cellular alteration in TNF-alpha production, an experimental model was set up. TNF-alpha-producing CD14+ cells in peripheral blood from lung cancer patients were significantly lower than those from healthy donors. In pleural effusion, TNF-alpha-producing CD14+ cells were significantly lower in lung cancer patients than in tuberculous patients. Based on the results obtained from an experimental model, we suggest that this phenomenon was attributed to a reduced expression of TNF-alpha transcript. These findings provide evidence that lung carcinomas reduce TNF-alpha production by macrophages, possibly by inducing in these cells an M2 phenotype, which favor tumor progression.

  12. Inhibition of mitochondrial function in HL60 cells is associated with an increased apoptosis and expression of CD14.

    PubMed

    Mills, K I; Woodgate, L J; Gilkes, A F; Walsh, V; Sweeney, M C; Brown, G; Burnett, A K

    1999-09-24

    The myelomonocytic cell line HL60 can be induced by a variety of chemical agents to differentiation to either neutrophils or monocytes. Examination of gene expression, by differential display, in cells induced to monocytes with 1alpha,25-dihydroxyvitamin D(3) or neutrophils with all-trans retinoic acid (ATRA) identified a number of clones with altered patterns of expression over the period of differentiation. One of these clones was the mitochondrial gene NADH dehydrogenase subunit 4 (ND4) which showed a differential pattern of expression between the neutrophil and monocyte lineages. The potential of mitochondrial inhibitors to induce differentiation was investigated by treating the HL60 cells with either the NADH dehydrogenase inhibitor, Rotenone, the complex III inhibitor, Antimycin A, or the highly specific mitochondrial ATP-synthase inhibitor, Oligomycin. Although functional assays of differentiation did not produce any positive results, all the inhibitors resulted in a dramatic increase in CD14 expression at day 1, with CD38 markers not observed until day 3. The increased expression of CD14 was accompanied by a decrease in viability and all CD14 positive cells were also positive for Annexin V, a marker of apoptosis. These results suggest that inhibition of the components of the mitochondrial pathways may lead to the marking of some cells, via CD14, for cell death, whilst allowing commitment to differentiation to occur in the surviving population. Copyright 1999 Academic Press.

  13. Human endotoxin tolerance is associated with enrichment of the CD14+ CD16+ monocyte subset.

    PubMed

    Domínguez-Nieto, Aimée; Zentella, Alejandro; Moreno, José; Ventura, José L; Pedraza, Sigifredo; Velázquez, Juan R

    2015-01-01

    Prior exposure to lipopolysaccharides (LPS) induces a state of cell resistance to subsequent LPS restimulation, known as endotoxin tolerance, mainly by repressing the expression of pro-inflammatory cytokines. We established an endotoxin tolerance model in human monocytes Endotoxin-tolerant cells showed a decrease in IκBα degradation and diminished expression of Tumor necrosis factor (TNF) (both messenger RNA [mRNA] and protein content). The myeloid differentiation factor 88 (MyD88)/MyD88 splice variant (MyD88s) ratio, an indirect way to test the Toll-like receptor 4 (TLR4) MyD88-dependent signaling cascade, did not change in endotoxin-tolerant cells when compared to LPS-stimulated or -unstimulated ones. Remarkably, cell population analysis indicated a significant increase of the CD14+ CD16+ subset only under the endotoxin-tolerant condition. Furthermore, endotoxin-tolerant cells produced higher amounts of C-X-C motif chemokine 10 (CXCL10), a typical MyD88-independent cytokine.

  14. TLR4 and CD14 receptors expressed in rat pineal gland trigger NFKB pathway.

    PubMed

    da Silveira Cruz-Machado, Sanseray; Carvalho-Sousa, Claudia Emanuele; Tamura, Eduardo Koji; Pinato, Luciana; Cecon, Erika; Fernandes, Pedro Augusto Carlos Magno; de Avellar, Maria Christina Werneck; Ferreira, Zulma Silva; Markus, Regina Pekelmann

    2010-09-01

    Nuclear factor-kappa B (NFKB), a pivotal player in inflammatory responses, is constitutively expressed in the pineal gland. Corticosterone inhibits pineal NFKB leading to an enhancement of melatonin production, while tumor necrosis factor (TNF) leads to inhibition of Aa-nat transcription and the production of N-acetylserotonin in cultured glands. The reduction in nocturnal melatonin surge favors the mounting of the inflammatory response. Despite these data, there is no clear evidence of the ability of the pineal gland to recognize molecules that signal infection. This study investigated whether the rat pineal gland expresses receptors for lipopolysaccharide (LPS), the endotoxin from the membranes of Gram-negative bacteria, and to establish the mechanism of action of LPS. Here, we show that pineal glands possess both CD14 and toll-like receptor 4 (TLR4), membrane proteins that bind LPS and trigger the NFKB pathway. LPS induced the nuclear translocation of p50/p50 and p50/RELA dimers and the synthesis of TNF. The maximal expression of TNF in cultured glands coincides with an increase in the expression of TNF receptor 1 (TNFR1) in isolated pinealocytes. In addition, LPS inhibited the synthesis of N-acetylserotonin and melatonin. Therefore, the pineal gland transduces Gram-negative endotoxin stimulation by producing TNF and inhibiting melatonin synthesis. Here, we provide evidence to reinforce the idea of an immune-pineal axis, showing that the pineal gland is a constitutive player in the innate immune response.

  15. Combined inhibition of complement C5 and CD14 markedly attenuates inflammation, thrombogenicity, and hemodynamic changes in porcine sepsis.

    PubMed

    Barratt-Due, Andreas; Thorgersen, Ebbe B; Egge, Kjetil; Pischke, Søren; Sokolov, Andrey; Hellerud, Bernt C; Lindstad, Julie K; Pharo, Anne; Bongoni, Anjan K; Rieben, Robert; Nunn, Miles; Scott, Helge; Mollnes, Tom E

    2013-07-15

    Complement and the TLR family constitute two important branches of innate immunity. We previously showed attenuating effects on inflammation and thromogenicity by inhibiting the TLR coreceptor CD14 in porcine sepsis. In the present study, we explored the effect of the C5 and leukotriene B4 inhibitor Ornithodoros moubata complement inhibitor (OmCI; also known as coversin) alone and combined with anti-CD14 on the early inflammatory, hemostatic, and hemodynamic responses in porcine Escherichia coli-induced sepsis. Pigs were randomly allocated to negative controls (n = 6), positive controls (n = 8), intervention with OmCI (n = 8), or with OmCI and anti-CD14 (n = 8). OmCI ablated C5 activation and formation of the terminal complement complex and significantly decreased leukotriene B4 levels in septic pigs. Granulocyte tissue factor expression, formation of thrombin-antithrombin complexes (p < 0.001), and formation of TNF-α and IL-6 (p < 0.05) were efficiently inhibited by OmCI alone and abolished or strongly attenuated by the combination of OmCI and anti-CD14 (p < 0.001 for all). Additionally, the combined therapy attenuated the formation of plasminogen activator inhibitor-1 (p < 0.05), IL-1β, and IL-8, increased the formation of IL-10, and abolished the expression of wCD11R3 (CD11b) and the fall in neutrophil cell count (p < 0.001 for all). Finally, OmCI combined with anti-CD14 delayed increases in heart rate by 60 min (p < 0.05) and mean pulmonary artery pressure by 30 min (p < 0.01). Ex vivo studies confirmed the additional effect of combining anti-CD14 with OmCI. In conclusion, upstream inhibition of the key innate immunity molecules, C5 and CD14, is a potential broad-acting treatment regimen in sepsis as it efficiently attenuated inflammation and thrombogenicity and delayed hemodynamic changes.

  16. The role of TLR4 and CD14 polymorphisms in the pathogenesis of respiratory syncytial virus bronchiolitis in greek infants.

    PubMed

    Goutaki, M; Haidopoulou, K; Pappa, S; Tsakiridis, P; Frydas, E; Eboriadou, M; Hatzistylianou, M

    2014-01-01

    Clinical manifestations of respiratory syncytial virus (RSV) infection vary from minimal disease to severe acute bronchiolitis. The structural complex of TLR4/CD14 participates in the virus recognition as a component of natural immune response. Genetic variations of TLR4/CD14 may explain great variations in disease severity. The aim of this study was to investigate the possible role of polymorphisms of TLR4, Asp299Gly and Thr399Ile and CD14, C-159T and C-550T in the development of RSV bronchiolitis. Our study included two groups of Greek infants and young children (A and B). Group A consisted of 50 infants ≤2 years of age hospitalised with bronchiolitis and group B of 99 previously healthy children aged 4-14 years (control group) with a free past medical history. RSV was identified by PCR of genetic material that was extracted from nasopharyngeal samples collected from all patients. Blood samples were used to extract DNA and by using the PCR-RFLP method we performed TLR4 and CD14 genotyping. We found no association between TLR4 polymorphisms (Asp299Gly and Thr399Ile) and the development of acute bronchiolitis. For CD14 polymorphisms, a positive association was found between the C-159T and the development of bronchiolitis (p=0.05) but not for the other loci. There were no differences detected in the frequencies of the four polymorphisms studied among infants with RSV and non-RSV bronchiolitis. It is concluded that protein CD14 may have a functional role in the viral conjunction to the structural complex TLR4/CD14. The association between the polymorphism C-159T and the manifestation of disease found in our study points out that the severity in the development of acute bronchiolitis is not specified exclusively by the pathogen, but the immune response of the host also plays a significant role. More extensive multicentric studies need to take place, in order to lead to safer conclusions.

  17. Soluble P-selectin in human plasma: effect of anticoagulant matrix and its levels in patients with cardiovascular disorders.

    PubMed

    Amin, H M; Ahmad, S; Walenga, J M; Hoppensteadt, D A; Leitz, H; Fareed, J

    2000-04-01

    P-Selectin represents a cell surface glycoprotein that is constitutively present in the Weibel-Palade bodies of endothelial cells and in the alpha-granules of platelets. In inflammation and thrombogenic conditions, plasmatic P-selectin levels are markedly elevated, indicating the leakage of this marker from these sites. In this study, a quantitative sandwich enzyme-linked immunosorbent assay (ELISA) utilizing a monoclonal soluble P-selectin (sP-selectin) antibody was employed to assess this marker in blood samples collected in various anticoagulants such as heparin, hirudin, sodium citrate (3.2% and 3.8%), and ethylenediaminetetraacetic acid (EDTA). The soluble P-selectin levels ranged from 26 ng/mL to 44 ng/mL. Sodium citrate (3.8%) was used to collect platelet-poor plasma (PPP) from patients with heparin-induced thrombocytopenia (HIT), coronary angioplasty (CA), or coronary atherectomy (CAT). In comparison with the control group (approximately 30 ng/mL), all of these patient groups showed a marked elevation of sP-selectin levels (HIT = 96 ng/mL [n = 18], CA = 46 ng/mL [n = 6] and CAT = 60 ng/mL [n = 10]). In platelet-rich plasma (PRP) preparations using various anticoagulants, the sP-selectin levels were markedly higher, ranging from 87 ng/ mL to 117 ng/mL (n = 10). In patients recruited into a clinical trial (the argatroban [ARG] 911 Study), in which argatroban was used as an alternate anticoagulant in patients with HIT, a 25% to 35% decrease in sP-selectin levels was observed after 72 hours of argatroban treatment. In addition, the relative ratio between levels in PRP and PPP in these patients differed, suggesting that the anticoagulant matrix influences the sP-selectin levels. These data clearly suggest that the anticoagulant matrix and blood collection procedures may significantly influence the plasmatic P-selectin levels. Furthermore, in different clinical conditions, elevation of this marker may reflect endogenous platelet activation; however, optimal

  18. Soluble Urokinase-Type Plasminogen Activator Receptor Plasma Concentration May Predict Susceptibility to High Altitude Pulmonary Edema

    PubMed Central

    Zügel, Stefanie; Schoeb, Michele; Auinger, Katja; Dehnert, Christoph; Maggiorini, Marco

    2016-01-01

    Introduction. Acute exposure to high altitude induces inflammation. However, the relationship between inflammation and high altitude related illness such as high altitude pulmonary edema (HAPE) and acute mountain sickness (AMS) is poorly understood. We tested if soluble urokinase-type plasminogen activator receptor (suPAR) plasma concentration, a prognostic factor for cardiovascular disease and marker for low grade activation of leukocytes, will predict susceptibility to HAPE and AMS. Methods. 41 healthy mountaineers were examined at sea level (SL, 446 m) and 24 h after rapid ascent to 4559 m (HA). 24/41 subjects had a history of HAPE and were thus considered HAPE-susceptible (HAPE-s). Out of the latter, 10/24 HAPE-s subjects were randomly chosen to suppress the inflammatory cascade with dexamethasone 8 mg bid 24 h prior to ascent. Results. Acute hypoxic exposure led to an acute inflammatory reaction represented by an increase in suPAR (1.9 ± 0.4 at SL versus 2.3 ± 0.5 at HA, p < 0.01), CRP (0.7 ± 0.5 at SL versus 3.6 ± 4.6 at HA, p < 0.01), and IL-6 (0.8 ± 0.4 at SL versus 3.3 ± 4.9 at HA, p < 0.01) in all subjects except those receiving dexamethasone. The ascent associated decrease in PaO2 correlated with the increase in IL-6 (r = 0.46, p < 0.001), but not suPAR (r = 0.27, p = 0.08); the increase in IL-6 was not correlated with suPAR (r = 0.16, p = 0.24). Baseline suPAR plasma concentration was higher in the HAPE-s group (2.0 ± 0.4 versus 1.8 ± 0.4, p = 0.04); no difference was found for CRP and IL-6 and for subjects developing AMS. Conclusion. High altitude exposure leads to an increase in suPAR plasma concentration, with the missing correlation between suPAR and IL-6 suggesting a cytokine independent, leukocyte mediated mechanism of low grade inflammation. The correlation between IL-6 and PaO2 suggests a direct effect of hypoxia, which is not the case for suPAR. However, suPAR plasma concentration measured before hypoxic exposure may predict

  19. Signaling of High Mobility Group Box 1 (HMGB1) through Toll-like Receptor 4 in Macrophages Requires CD14

    PubMed Central

    Kim, Sodam; Kim, Sun Young; Pribis, John P; Lotze, Michael; Mollen, Kevin P; Shapiro, Richard; Loughran, Patricia; Scott, Melanie J; Billiar, Timothy R

    2013-01-01

    High mobility group box 1 (HMGB1) is a DNA-binding protein that possesses cytokinelike, proinflammatory properties when released extracellularly in the C23–C45 disulfide form. HMGB1 also plays a key role as a mediator of acute and chronic inflammation in models of sterile injury. Although HMGB1 interacts with multiple pattern recognition receptors (PRRs), many of its effects in injury models occur through an interaction with toll-like receptor 4 (TLR4). HMGB1 interacts directly with the TLR4/myeloid differentiation protein 2 (MD2) complex, although the nature of this interaction remains unclear. We demonstrate that optimal HMGB1-dependent TLR4 activation in vitro requires the coreceptor CD14. TLR4 and MD2 are recruited into CD14-containing lipid rafts of RAW264.7 macrophages after stimulation with HMGB1, and TLR4 interacts closely with the lipid raft protein GM1. Furthermore, we show that HMGB1 stimulates tumor necrosis factor (TNF)-α release in WT but not in TLR4−/−, CD14−/−, TIR domain-containing adapter-inducing interferon-β (TRIF)−/− or myeloid differentiation primary response protein 88 (MyD88)−/− macrophages. HMGB1 induces the release of monocyte chemotactic protein 1 (MCP-1), interferon gamma–induced protein 10 (IP-10) and macrophage inflammatory protein 1α (MIP-1α) in a TLR4- and CD14-dependent manner. Thus, efficient recognition of HMGB1 by the TLR4/MD2 complex requires CD14. PMID:23508573

  20. Systemic immune suppression in glioblastoma: the interplay between CD14+HLA-DRlo/neg monocytes, tumor factors, and dexamethasone

    PubMed Central

    Gustafson, Michael P.; Lin, Yi; New, Kent C.; Bulur, Peggy A.; O'Neill, Brian Patrick; Gastineau, Dennis A.; Dietz, Allan B.

    2010-01-01

    Patients with glioblastoma (GBM) exhibit profound systemic immune defects that affect the success of conventional and immune-based treatments. A better understanding of the contribution of the tumor and/or therapy on systemic immune suppression is necessary for improved therapies, to monitor negative effects of novel treatments, to improve patient outcomes, and to increase understanding of this complex system. To characterize the immune profile of GBM patients, we phenotyped peripheral blood and compared these to normal donors. In doing so, we identified changes in systemic immunity associated with both the tumor and dexamethasone treated tumor bearing patients. In particular, dexamethasone exacerbated tumor associated lymphopenia primarily in the T cell compartment. We have also identified unique tumor and dexamethasone dependent altered monocyte phenotypes. The major population of altered monocytes (CD14+HLA-DRlo/neg) had a phenotype distinct from classical myeloid suppressor cells. These cells inhibited T cell proliferation, were unable to fully differentiate into mature dendritic cells, were associated with dexamethasone-mediated changes in CCL2 levels, and could be re-created in vitro using tumor supernatants. We provide evidence that tumors express high levels of CCL2, can contain high numbers of CD14+ cells, that tumor supernatants can transform CD14+HLA-DR+ cells into CD14+HLA-DRlo/neg immune suppressors, and that dexamethasone reduces CCL2 in vitro and is correlated with reduction of CCL2 in vivo. Consequently, we have developed a model for tumor mediated systemic immune suppression via recruitment and transformation of CD14+ cells. PMID:20179016

  1. Large-scale immunomagnetic selection of CD14+ monocytes to generate dendritic cells for cancer immunotherapy: a phase I study.

    PubMed

    Babatz, J; Röllig, C; Oelschlägel, U; Zhao, S; Ehninger, G; Schmitz, M; Bornhäuser, M

    2003-10-01

    Dendritic cells (DC) are professional antigen-presenting cells that are widely used in the experimental immunotherapy of cancer. For clinical use GMP-like protocols for the preparation of functionally active dendritic cells (DC) in large numbers and at high purity are needed. However, the currently available protocols have certain disadvantages. In this study we tested the generation and clinical applicability of DC from monocyte preparations produced by immunomagnetic CD14(+) selection using a semiautomated clinical scale immunomagnetic column. Peripheral blood mononuclear cells (PBMC) of 10 patients with metastatic solid tumors were used. With the immunomagnetic separation, we obtained a cell suspension of high CD14(+) purity (median 97.4%, range 94.9-99.0) with a high monocyte yield (median 82.3%, range 63.9-100.0). Differentiation of CD14(+) cells into mature monocyte-derived DC was induced by incubation with IL-4, GM-CSF, TNF-alpha, PGE(2), IL-1 beta, and IL-6. Mature DC showed a high expression of CD83, HLA-DR, and the co-stimulatory molecules CD80 and CD86. Overall CD83(+) yield was 12.1% (range 4.0-29.4). Allogeneic T stimulatory capacity could be demonstrated for all DC preparations in proliferation assays. No significant differences in marker expression or T cell stimulation was detected between fresh DC and those derived from cryopreserved immature DC. Clinical administration of autologous DC by three different parenteral routes was tolerated by all 10 patients without systemic signs of toxicity. Our results indicate that immunomagnetic isolation of CD14(+) monocytes using the CliniMACS device is a suitable method for clinical-scale generation of functional DC under GMP-grade conditions. The selection can be performed in a closed system. Therefore, immunomagnetic CD14(+) selection can be seen as an alternative way to generate DC for clinical tumor vaccination protocols.

  2. Soluble Lutheran/basal cell adhesion molecule is detectable in plasma of hepatocellular carcinoma patients and modulates cellular interaction with laminin-511 in vitro.

    PubMed

    Kikkawa, Yamato; Miwa, Takahiro; Tanimizu, Naoki; Kadoya, Yuichi; Ogawa, Takaho; Katagiri, Fumihiko; Hozumi, Kentaro; Nomizu, Motoyoshi; Mizuguchi, Toru; Hirata, Koichi; Mitaka, Toshihiro

    2014-10-15

    Lutheran (Lu), an immunoglobulin superfamily transmembrane receptor, is also known as basal cell adhesion molecule (B-CAM). Lu/B-CAM is a specific receptor for laminin α5, a subunit of laminin-511 (LM-511) that is a major component of basement membranes in various tissues. Our previous study showed that Lu/B-CAM was cleaved by MT1-MMP and released from cell surfaces. In this study we examined the soluble Lu/B-CAM in culture media and in plasma of mice bearing HuH-7 hepatocellular carcinoma (HCC) cells and patients with HCC. Two HCC cell lines, HepG2 and HuH-7, released Lu/B-CAM into the culture media. Although Lu/B-CAM was cleaved by MT1-MMP in HuH-7 cells, HepG2 cells released Lu/B-CAM in a MMP-independent manner. The concentration of Lu/B-CAM released into mouse plasma correlated with tumor size. Moreover the soluble Lu/B-CAM in plasma of HCC patients was significantly decreased after resection of the tumor. Immunohistochemical studies showed that although the expression of Lu/B-CAM was observed in most HCCs, MT1-MMP was not always expressed in tumor tissues, suggesting that a part of Lu/B-CAM in plasma of HCC patients was also released in a MMP-independent manner. In vitro studies showed that the soluble Lu/B-CAM released from HCC cells bound to LM-511. Moreover the soluble Lu/B-CAM influenced cell migration on LM-511. These results suggest that soluble Lu/B-CAM serves as not only a novel marker for HCC but also a modulator in tumor progression.

  3. Functional characterization of a STAT3-dependent dendritic cell-derived CD14+ cell population arising upon IL-10-driven maturation

    PubMed Central

    Lindenberg, Jelle J.; van de Ven, Rieneke; Lougheed, Sinéad M.; Zomer, Anoek; Santegoets, Saskia J.A.M.; Griffioen, Arjan W.; Hooijberg, Erik; van den Eertwegh, Alfons J.M.; Thijssen, Victor L.; Scheper, Rik J.; Oosterhoff, Dinja; de Gruijl, Tanja D.

    2013-01-01

    Interleukin (IL)-10 is a major cancer-related immunosuppressive factor, exhibiting a unique ability to hamper the maturation of dendritic cells (DCs). We have previously reported that IL-10 induces the conversion of activated, migratory CD1a+ DCs found in the human skin to CD14+CD141+ macrophage-like cells. Here, as a model of tumor-conditioned DC maturation, we functionally assessed CD14- and CD14+ DCs that matured in vitro upon exposure to IL-10. IL-10-induced CD14+ DCs were phenotypically characterized by a low maturation state as well as by high levels of BDCA3 and DC-SIGN, and as such they closely resembled CD14+ cells infiltrating melanoma metastases. Compared with DC matured under standard conditions, CD14+ DCs were found to express high levels of B7-H1 on the cell surface, to secrete low levels of IL-12p70, to preferentially induce TH2 cells, to have a lower allogeneic TH cell and tumor antigen-specific CD8+ T-cell priming capacity and to induce proliferative T-cell anergy. In contrast to their CD14+ counterparts, CD14- monocyte-derived DCs retained allogeneic TH priming capacity but induced a functionally anergic state as they completely abolished the release of effector cytokines. Transcriptional and cytokine release profiling studies indicated a more profound angiogenic and pro-invasive signature of CD14+ DCs as compared with DCs matured in standard conditions or CD14− DCs matured in the presence of IL-10. Importantly, signal transducer and activator of transcription 3 (STAT3) depletion by RNA interference prevented the development of the IL-10-associated CD14+ phenotype, allowing for normal DC maturation and providing a potential means of therapeutic intervention. PMID:23734330

  4. Elevated circulating CD14(low)CD16(+) monocyte subset in primary biliary cirrhosis correlates with liver injury and promotes Th1 polarization.

    PubMed

    Peng, Anping; Ke, Peifeng; Zhao, Rong; Lu, Xinyi; Zhang, Cheng; Huang, Xianzhang; Tian, Guangjun; Huang, Jun; Wang, Jinli; Invernizzi, Pietro; Chen, Qubo; Zhuang, Junhua

    2016-11-01

    Primary biliary cirrhosis (PBC) is a progressive autoimmune liver disease in which monocytes/macrophages infiltration and skewed T helper type (Th) 1 and Th17 cell responses participate in the development of the disease. Human peripheral blood monocytes are heterogeneous and can be divided into classical CD14(high)CD16(-), intermediate CD14(high)CD16(+), and nonclassical CD14(low)CD16(+) monocyte subsets. Compared to classical monocytes, CD16(+) monocytes are generally termed pro-inflammatory monocytes and play an important pathogenic role in autoimmune diseases. However, little is known about the immunophenotype and immunopathogenic role of peripheral blood CD16(+) monocytes in PBC. Thus, we investigated the phenotype and function of these circulating monocyte subsets from PBC patients. The frequencies of circulating CD14(high)CD16(+) and CD14(low)CD16(+) subpopulation were increased in disease compared with healthy controls. Among them, CD14(low)CD16(+) monocyte subset positively correlated with disease progress, liver damage indicators and serum C-reactive protein, respectively. Furthermore, the frequencies of Th1 and Th17 cells were upregulated and CD14(low)CD16(+) monocyte subset was also positively associated with Th1 cell frequency in PBC. Using a vitro coculture model, we further found that CD14(low)CD16(+) monocytes promoted Th1 cell polarization compared to classical monocytes. Interleukin-12 (IL-12) and direct contact of patient CD4(+)T cell and CD14(low)CD16(+) monocytes, were responsible for CD14(low)CD16(+) monocytes promotion of Th1 cells polarization in PBC. Our study demonstrated that the enhanced CD14(low)CD16(+) monocyte subset participated in fostering liver damage and inflammatory responses, and promoted Th1 cells skewing in PBC.

  5. Plasma soluble Tim-3 emerges as an inhibitor in sepsis: sepsis contrary to membrane Tim-3 on monocytes.

    PubMed

    Ren, F; Li, J; Jiang, X; Xiao, K; Zhang, D; Zhao, Z; Ai, J; Hou, C; Jia, Y; Han, G; Xie, L

    2015-11-01

    Immune dysfunction is the main characteristic of sepsis. T cell Ig and mucin domain protein 3 (Tim-3) on the monocytes has been reported to promote immune homeostasis during sepsis, but the influences of plasm soluble Tim-3 (sTim-3) on the immune system during sepsis remain unknown. Here, 100 patients with different severities of sepsis (40 sepsis, 42 severe sepsis, and 18 septic shock) were enrolled in this study. The Tim-3 and human leukocyte antigen-DR (HLA-DR) on the circulating monocytes were detected using flow cytometry. Plasma sTim-3 was detected by enzyme-linked immunosorbent assay. Inflammatory factors and two kinds of A disintegrin and metalloprotease (ADAM) - ADAM10 and ADAM17 were assessed. The Tim-3 and HLA-DR on the monocytes decreased with increasing sepsis severity. The sTim-3 was reduced in the sepsis and severe sepsis patients but was elevated in the septic shock patients who exhibited significant immunosuppression as predicted by HLA-DR. sTim-3 levels were negatively correlated with IL-12 and TNF-α. ADAM10 and ADAM17, sheddases of Tim-3, exhibited trends toward elevations in the septic shock group. In conclusion, sTim-3 was involved in the development of sepsis. The homeostasis-promoting role of the Tim-3 on the monocytes was disrupted, while the inhibitory role of sTim-3 emerged during sepsis-induced immunosuppression.

  6. CD40L expression in plasma of volunteers following LPS administration: A comparison between assay of CD40L on platelet microvesicles and soluble CD40L.

    PubMed

    Mobarrez, Fariborz; Sjövik, Carolina; Soop, Anne; Hållström, Lars; Frostell, Claes; Pisetsky, David S; Wallén, Håkan

    2015-01-01

    CD40 ligand (CD40L) is a transmembrane protein that is mainly expressed on activated T cells and platelets. This protein, however, may also be shed from cells and circulate in the blood in a soluble form. "Soluble CD40L" has attracted interest as a biomarker as it can interact with CD40 and elicit cellular responses involved in the pathophysiology of various thrombotic and inflammatory conditions. As platelets can release microvesicles following activation, we investigated the expression of CD40L on circulating microvesicles as well as CD40L in plasma, in an experimental model of inflammation in healthy volunteers (i.e., intravenous lipopolysaccharide administration). We studied CD40L quantified as CD40L-positive platelet microvesicles by flow cytometry, and as CD40L in plasma ("soluble CD40L") by an ELISA. Results of these studies showed that levels of CD40L exposed on platelet microvesicles were significantly increased after lipopolysaccharide administration. ELISA measurements of CD40L in plasma ("soluble CD40L") did not show any significant increase in plasma levels over time. Separation of soluble and vesicle-bound CD40L by high-speed centrifugation indicated that the ELISA can also detect CD40L on microvesicles, as a trend toward increased concentrations were observed in the pellet of high-speed centrifuged samples (i.e., in samples in which microvesicles are enriched). Together, these findings suggest that platelet microvesicles are a source of CD40L in the circulation and that CD40L exposure on platelet microvesicles increases following experimentally induced inflammation. Our data also suggest that determining levels of CD40L on microvesicles in plasma samples may provide a more sensitive detection of changes in CD40L expression than measurement of "soluble CD40L" in plasma with an ELISA. In addition, information regarding the cellular source of CD40L can be obtained with a flow cytometry-based microvesicle assay in a way not possible with an ordinary

  7. The Effect of Tacrolimus and Mycophenolic Acid on CD14+ Monocyte Activation and Function

    PubMed Central

    Kannegieter, Nynke M.; Hesselink, Dennis A.; Dieterich, Marjolein; Kraaijeveld, Rens; Rowshani, Ajda T.; Leenen, Pieter J. M.; Baan, Carla C.

    2017-01-01

    Monocytes and macrophages play key roles in many disease states, including cellular and humoral rejection after solid organ transplantation (SOT). To suppress alloimmunity after SOT, immunosuppressive drug therapy is necessary. However, little is known about the effects of the immunosuppressive drugs tacrolimus and mycophenolic acid (MPA) on monocyte activation and function. Here, the effect of these immunosuppressants on monocytes was investigated by measuring phosphorylation of three intracellular signaling proteins which all have a major role in monocyte function: p38MAPK, ERK and Akt. In addition, biological functions downstream of these signaling pathways were studied, including cytokine production, phagocytosis and differentiation into macrophages. To this end, blood samples from healthy volunteers were spiked with diverse concentrations of tacrolimus and MPA. Tacrolimus (200 ng/ml) inhibited phosphorylation of p38MAPK by 30% (mean) in CD14+ monocytes which was significantly less than in activated CD3+ T cells (max 60%; p < 0.05). This immunosuppressive agent also partly inhibited p-AKT (14%). MPA, at a therapeutic concentration showed the strongest effect on p-AKT (27% inhibition). p-ERK was inhibited with a maximum of 15% after spiking with either tacrolimus or MPA. The production of IL-1β and phagocytosis by monocytes were not affected by tacrolimus concentrations, whereas MPA did inhibit IL-1β production by 50%. Monocyte/macrophage polarization was shifted to an M2-like phenotype in the presence of tacrolimus, while MPA increased the expression of M2 surface markers, including CD163 and CD200R, on M1 macrophages. These results show that tacrolimus and MPA do not strongly affect monocyte function, apart from a change in macrophage polarization, to a clinically relevant degree. PMID:28122021

  8. Plasmatic presepsin (sCD14-ST) concentrations in acute pyelonephritis in adult patients.

    PubMed

    Claessens, Yann-Erick; Trabattoni, Eloise; Grabar, Sophie; Quinquis, Laurent; Der Sahakian, Guillaume; Anselmo, Marine; Schmidt, Jeannot; de la Coussaye, Jean-Emmanuel; Plaisance, Patrick; Casalino, Enrique; Potel, Gilles; Lecomte, François; Borderie, Didier; Chenevier-Gobeaux, Camille

    2017-01-01

    Presepsin (sCD14-ST) is an emerging biomarker for infection. We hypothesized that presepsin could specifically increase during acute pyelonephritis and correlate with severity. We compared presepsin values in patients with acute pyelonephritis and controls, and we assessed its capacity to predict bacteraemia and admission in patients. In 312 patients with acute pyelonephritis (median age 33years), presepsin concentrations were higher than in controls (476 vs 200ng/L, p<0.001). ROC curve indicated an AUC at 0.90 [for presepsin (vs. 0.99 and 0.98 for CRP and PCT, respectively; p<0.05) and an optimal threshold at 340ng/L (74% sensitivity, 94% specificity). Presepsin concentrations increased in acute pyelonephritis patients with bacteraemia (614 vs. 461ng/L, p,=0.001) and in those requiring admission (614ng/L vs. 320ng/L, p<0.001). Performance of presepsin to predict bacteraemia [AUC=0.63, 95%CI: 0.55-0.72] was similar to CRP (AUC=0.64, p=0.87) and less accurate than PCT (AUC=0.78, p<0.001). AUC for presepsin to detect the need for admission was 0.67, and comparable to CRP (p=0.26) and PCT (p=0.18). Presepsin is a valuable biomarker to detect patients with acute pyelonephritis. However, it presents mild performance to predict bacteraemia and the need for admission, and offers no advantage as compared to CRP and PCT. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Beryllium increases the CD14(dim)CD16+ subset in the lung of chronic beryllium disease.

    PubMed

    Li, Li; Hamzeh, Nabeel; Gillespie, May; Elliott, Jill; Wang, Jieru; Gottschall, Eva Brigitte; Mroz, Peggy M; Maier, Lisa A

    2015-01-01

    CD14dimCD16+ and CD14brightCD16+ cells, which compose a minor population of monocytes in human peripheral blood mononuclear cells (PBMC), have been implicated in several inflammatory diseases. The aim of this study was to investigate whether this phenotype was present as a subset of lung infiltrative alveolar macrophages (AMs) in the granulomatous lung disease, chronic beryllium disease (CBD). The monocytes subsets was determined from PBMC cells and bronchoalveolar lavage (BAL) cells from CBD, beryllium sensitized Non-smoker (BeS-NS) and healthy subjects (HS) using flow cytometry. The impact of smoking on the AMs cell phenotype was determined by using BAL cells from BeS smokers (BeS-S). In comparison with the other monocyte subpopulations, CD14dimCD16+ cells were at decreased frequency in PBMCs of both BeS-NS and CBD and showed higher HLA-DR expression, compared to HS. The AMs from CBD and BeS-NS demonstrated a CD14dimCD16+phenotype, while CD14brightCD16+ cells were found at increased frequency in AMs of BeS, compared to HS. Fresh AMs from BeS-NS and CBD demonstrated significantly greater CD16, CD40, CD86 and HLA-DR than HS and BeS-S. The expression of CD16 on AMs from both CBD and BeS-NS was downregulated significantly after 10μM BeSO4 stimulation. The phagocytic activity of AMs decreased after 10μM BeSO4 treatment in both BeS-NS and CBD, although was altered or reduced in HS and BeS-S. These results suggest that Be increases the CD14dimCD16+ subsets in the lung of CBD subjects. We speculate that Be-stimulates the compartmentalization of a more mature CD16+ macrophage phenotype and that in turn these macrophages are a source of Th1 cytokines and chemokines that perpetuate the Be immune response in CBD. The protective effect of cigarette smoking in BeS-S may be due to the low expression of co-stimulatory markers on AMs from smokers as well as the decreased phagocytic function.

  10. Melanoma-educated CD14+ cells acquire a myeloid-derived suppressor cell phenotype through COX-2-dependent mechanisms.

    PubMed

    Mao, Yumeng; Poschke, Isabel; Wennerberg, Erik; Pico de Coaña, Yago; Egyhazi Brage, Suzanne; Schultz, Inkeri; Hansson, Johan; Masucci, Giuseppe; Lundqvist, Andreas; Kiessling, Rolf

    2013-07-01

    Tumors can suppress the host immune system by employing a variety of cellular immune modulators, such as regulatory T cells, tumor-associated macrophages, and myeloid-derived suppressor cells (MDSC). In the peripheral blood of patients with advanced stage melanoma, there is an accumulation of CD14(+)HLA-DR(lo/-) MDSC that suppress autologous T cells ex vivo in a STAT-3-dependent manner. However, a precise mechanistic basis underlying this effect is unclear, particularly with regard to whether the MDSC induction mechanism relies on cell-cell contact of melanoma cells with CD14(+) cells. Here, we show that early-passage human melanoma cells induce phenotypic changes in CD14(+) monocytes, leading them to resemble MDSCs characterized in patients with advanced stage melanoma. These MDSC-like cells potently suppress autologous T-cell proliferation and IFN-γ production. Notably, induction of myeloid-suppressive functions requires contact or close proximity between monocytes and tumor cells. Further, this induction is largely dependent on production of cyclooxygenase-2 (COX-2) because its inhibition in these MDSC-like cells limits their ability to suppress T-cell function. We confirmed our findings with CD14(+) cells isolated from patients with advanced stage melanoma, which inhibited autologous T cells in a manner relying up prostaglandin E2 (PGE2), STAT-3, and superoxide. Indeed, PGE2 was sufficient to confer to monocytes the ability to suppress proliferation and IFN-γ production by autologous T cells ex vivo. In summary, our results reveal how immune suppression by MDSC can be initiated in the tumor microenvironment of human melanoma.

  11. CD14 and Toll-like receptors 2 and 4 are required for fibrillar Aβ-stimulated microglial activation

    PubMed Central

    Reed-Geaghan, Erin G.; Savage, Julie C.; Hise, Amy G.; Landreth, Gary E.

    2009-01-01

    Microglia are the brain's tissue macrophages and are found in an activated state surrounding β-amyloid plaques in the Alzheimer's disease brain. Microglia interact with fibrillar β-amyloid (fAβ) through an ensemble of surface receptors composed of the α6β1 integrin, CD36, CD47, and the class A scavenger receptor. These receptors act in concert to initiate intracellular signaling cascades and phenotypic activation of these cells. However, it is unclear how engagement of this receptor complex is linked to the induction of an activated microglial phenotype. We report that the response of microglial cells to fibrillar forms of Aβ requires the participation of Toll like receptors (TLRs) and the co-receptor CD14. The response of microglia to fAβ is reliant upon CD14, which act together with TLR4 and TLR2 to bind fAβ and to activate intracellular signaling. We find that cells lacking these receptors could not initiate a Src-Vav-Rac signaling cascade leading to reactive oxygen species production and phagocytosis. The fAβ-mediated activation of p38 MAPK also required CD14, TLR4, and TLR2. Inhibition of p38 abrogated fAβ-induced reactive oxygen species production and attenuated the induction of phagocytosis. Microglia lacking CD14, TLR4, and TLR2 showed no induction of phosphorylated IκBα following fAβ. These data indicate these innate immune receptors function as members of the microglial fAβ receptor complex and identify the signaling mechanisms whereby they contribute to microglial activation. PMID:19776284

  12. Dengue virus infection induces expansion of a CD14(+)CD16(+) monocyte population that stimulates plasmablast differentiation.

    PubMed

    Kwissa, Marcin; Nakaya, Helder I; Onlamoon, Nattawat; Wrammert, Jens; Villinger, Francois; Perng, Guey Chuen; Yoksan, Sutee; Pattanapanyasat, Kovit; Chokephaibulkit, Kulkanya; Ahmed, Rafi; Pulendran, Bali

    2014-07-09

    Dengue virus (DENV) infection induces the expansion of plasmablasts, which produce antibodies that can neutralize DENV but also enhance disease upon secondary infection with another DENV serotype. To understand how these immune responses are generated, we used a systems biological approach to analyze immune responses to dengue in humans. Transcriptomic analysis of whole blood revealed that genes encoding proinflammatory mediators and type I interferon-related proteins were associated with high DENV levels during initial symptomatic disease. Additionally, CD14(+)CD16(+) monocytes increased in the blood. Similarly, in a nonhuman primate model, DENV infection boosted CD14(+)CD16(+) monocyte numbers in the blood and lymph nodes. Upon DENV infection in vitro, monocytes upregulated CD16 and mediated differentiation of resting B cells to plasmablasts as well as immunoglobulin G (IgG) and IgM secretion. These findings provide a detailed picture of innate responses to dengue and highlight a role for CD14(+)CD16(+) monocytes in promoting plasmablast differentiation and anti-DENV antibody responses.

  13. Trappin-2 promotes early clearance of Pseudomonas aeruginosa through CD14-dependent macrophage activation and neutrophil recruitment.

    PubMed

    Wilkinson, Thomas S; Dhaliwal, Kevin; Hamilton, Thomas W; Lipka, Alexander F; Farrell, Lesley; Davidson, Donald J; Duffin, Rodger; Morris, Andrew Conway; Haslett, Chris; Govan, John R W; Gregory, Christopher D; Sallenave, Jean-Michel; Simpson, A John

    2009-04-01

    Microaspiration of Pseudomonas aeruginosa contributes to the pathogenesis of nosocomial pneumonia. Trappin-2 is a host defense peptide that assists with the clearance of P. aeruginosa through undefined mechanisms. A model of macrophage interactions with replicating P. aeruginosa (strain PA01) in serum-free conditions was developed, and the influence of subantimicrobial concentrations of trappin-2 was subsequently studied. PA01 that was pre-incubated with trappin-2 (at concentrations that have no direct antimicrobial effects), but not control PA01, was cleared by alveolar and bone marrow-derived macrophages. However, trappin-2-enhanced clearance of PA01 was completely abrogated by CD14- null macrophages. Fluorescence microscopy demonstrated the presence of trappin-2 on the bacterial cell surface of trappin-2-treated PA01. In a murine model of early lung infection, trappin-2-treated PA01 was cleared more efficiently than control PA01 2 hours of intratracheal instillation. Furthermore, trappin-2-treated PA01 up-regulated the murine chemokine CXCL1/KC after 2 hours with a corresponding increase in neutrophil recruitment 1 hour later. These in vivo trappin-2-treated PA01 effects were absent in CD14-deficient mice. Trappin-2 appears to opsonize P. aeruginosa for more efficient, CD14-dependent clearance by macrophages and contributes to the induction of chemokines that promote neutrophil recruitment. Trappin-2 may therefore play an important role in innate recognition and clearance of pathogens during the very earliest stages of pulmonary infection.

  14. Importance of the plasma soluble HLA-G levels for prognostic stratification with traditional prognosticators in colorectal cancer.

    PubMed

    Li, Jing-Bo; Ruan, Yan-Yun; Hu, Bin; Dong, Shan-Shan; Bi, Tie-Nan; Lin, Aifen; Yan, Wei-Hua

    2017-07-25

    An increased peripheral soluble HLA-G (sHLA-G) expression has been observed in various malignancies while its prognostic significance was rather limited. In this study, the prognostic value of plasma sHLA-G in 178 colorectal cancer (CRC) patients was investigated. sHLA-G levels were analyzed by specific enzyme-linked immunosorbent assay. Data showed sHLA-G levels were significantly increased in CRC patients compared with normal controls (36.8 U/ml vs 25.4 U/ml, p = 0.009). sHLA-G in the died were obviously higher than that of alive CRC patients (46.8 U/ml vs 27.4 U/ml, p = 0.012). Patients with sHLA-G above median levels (≥ 36.8 U/ml, sHLA-Ghigh) had a significantly shorter survival time than those with sHLA-Glow (< 36.8 U/ml, p < 0.001), and sHLA-G could be an independent prognostic factor for CRC patients. With stratification of clinical parameters in survival by sHLA-Glow and sHLA-Ghigh, sHLA-G exhibited a significant predictive value for CRC patients of the female (p = 0.036), the elder (p = 0.009), advanced tumor burden (T3 + 4, p = 0.038), regional lymph node status (N0, p = 0.041), both metastasis status (M0, p = 0.014) and (M1, p=0.018), and clinical stage (I + II, p = 0.018), respectively. Summary, our data demonstrated for the first time that sHLA-G levels is an independent prognosis factor and improves the prognostic stratification offered by traditional prognosticators in CRC patients.

  15. Effect of intensive insulin therapy on macular biometrics, plasma VEGF and its soluble receptor in newly diagnosed diabetic patients.

    PubMed

    Hernández, Cristina; Zapata, Miguel A; Losada, Eladio; Villarroel, Marta; García-Ramírez, Marta; García-Arumí, José; Simó, Rafael

    2010-07-01

    To evaluate whether intensive insulin therapy leads to changes in macular biometrics (volume and thickness) in newly diagnosed diabetic patients with acute hyperglycaemia and its relationship with serum levels of vascular endothelial growth factor (VEGF) and its soluble receptor (sFlt-1). Twenty-six newly diagnosed diabetic patients admitted to our hospital to initiate intensive insulin treatment were prospectively recruited. Examinations were performed on admission (day 1) and during follow-up (days 3, 10 and 21) and included a questionnaire regarding the presence of blurred vision, standardized refraction measurements and optical coherence tomography. Plasma VEGF and sFlt-1 were assessed by ELISA at baseline and during follow-up. At study entry seven patients (26.9%) complained of blurred vision and five (19.2%) developed burred vision during follow-up. Macular volume and thickness increased significantly (p = 0.008 and p = 0.04, respectively) in the group with blurred vision at day 3 and returned to the baseline value at 10 days. This pattern was present in 18 out of the 24 eyes from patients with blurred vision. By contrast, macular biometrics remained unchanged in the group without blurred vision. We did not detect any significant changes in VEGF levels during follow-up. By contrast, a significant reduction of sFlt-1 was observed in those patients with blurred vision at day 3 (p = 0.03) with normalization by day 10. Diabetic patients with blurred vision after starting insulin therapy present a significant transient increase in macular biometrics which is associated with a decrease in circulating sFlt-1. Copyright (c) 2010 John Wiley & Sons, Ltd.

  16. Evaluation and analysis of plasma soluble adhesion molecules in patients with coronary ectasia and atherosclerotic coronary artery disease.

    PubMed

    Soto, Maria Elena; Reyes-Villatoro, Martha Abigail; Márquez, Ricardo; Cardoso, Guillermo; Posadas-Sánchez, Rosalinda; Juárez-Orozco, Luis Eduardo

    2014-08-01

    The pathogenesis of coronary artery ectasia (CE) is poorly understood. An underlying inflammatory process is suspected; current therapy aims to prevent thrombotic events. Presently, there is no evidence to support an anti-inflammatory approach for CE patients. We undertook this study to determine the presence and levels of adhesion molecules as well as other inflammation-related markers and evaluate their invasive angiography findings to compare between patients with CE or atherosclerotic coronary artery disease (CAD). We included 23 patients with CE alone and 23 patients with CAD alone reported by diagnostic angiography. The two groups were paired according to gender and age. Clinical variables, angiographic findings (STEMI, NSTEMI, previous coronary stenting and previous CABG), serum VCAM, ICAM, C-reactive protein (CRP), IL-6 and adiponectin were compared between groups. In patients with CE, median for serum VCAM was 1700 ng/mL with a range from 900-4050 and for serum ICAM was 550 ng/mL (370-1530); in contrast to the obstructive lesions group where we found a median for VCAM of 1150 ng/mL (650-2000) and for ICAM of 370 ng/mL (150-480). Both measurements achieved statistical significance with a p value = 0.0001. There was no important variation between patients measured at different times from diagnosis. Plasma soluble adhesion molecules in CE are elevated in comparison to CAD. The presence of high levels of these molecules, along with not uncommon multivessel and extensive coronary affection suggest the participation of subclinical inflammation, which may have an important role in the development of CE. Copyright © 2014 IMSS. Published by Elsevier Inc. All rights reserved.

  17. The relationship between asthma and bronchiolitis is modified by TLR4, CD14, and IL-13 polymorphisms.

    PubMed

    Jung, Young-Ho; Seo, Ju-Hee; Kim, Hyung Young; Kwon, Ji-Won; Kim, Byoung-Ju; Kim, Hyo-Bin; Lee, So-Yeon; Jang, Gwang Cheon; Song, Dae Jin; Kim, Woo Kyung; Shim, Jung Yeon; Hong, Soo-Jong

    2015-01-01

    Asthma is a complex genetic disorder that is associated with both genetic and environmental factors. The aim of study was to investigate the combined effect of toll-like receptor 4 (TLR4), cluster of differentiation 14 (CD14), and interleukin-13 (IL-13) polymorphisms and bronchiolitis in the development of childhood asthma. A modified International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire was used to survey 1,341 elementary school children and 919 nursery children in Seoul, Korea. TLR4 (rs1927911), CD14 (rs2569190), and IL-13 (rs20541) polymorphisms were genotyped by the TaqMan assay. In elementary school and nursery children, parental history of asthma (adjusted odds ratio [aOR] 2.56 [95% CI 1.16-5.63], aOR 3.60 [95% CI 1.66-7.76], respectively), and past history of bronchiolitis (aOR 3.11 [95% CI 1.84-5.24], aOR 3.94 [95% CI 2.27-6.84], respectively) were independent risk factors for asthma diagnosis. When compared to children with each CC of TLR4 polymorphism or TT of CD14 polymorphism or GG of IL13 polymorphism and no past history of bronchiolitis, children with CT or TT of TLR4 polymorphism and past history of bronchiolitis had 4.23 and 5.34 times higher risk to develop asthma, respectively; children with TT of CD14 polymorphism and past history of bronchiolitis had 3.57 and 7.22 times higher risk for asthma, respectively; children with GA or AA of IL-13 polymorphism and past history of bronchiolitis had 3.21 and 4.13 times higher risk for asthma, respectively. Family history of asthma or allergic rhinitis and past history of bronchiolitis could be independent risk factors for the development of childhood asthma. The relationship between asthma and bronchiolitis is modified by the TLR4, CD14, and IL-13 polymorphisms in Korean children. © 2013 Wiley Periodicals, Inc.

  18. Angiotensin II type 1 receptor antagonist decreases plasma levels of tumor necrosis factor alpha, interleukin-6 and soluble adhesion molecules in patients with chronic heart failure.

    PubMed

    Tsutamoto, T; Wada, A; Maeda, K; Mabuchi, N; Hayashi, M; Tsutsui, T; Ohnishi, M; Sawaki, M; Fujii, M; Matsumoto, T; Kinoshita, M

    2000-03-01

    To evaluate the effects of an angiotensin (Ang II) type 1 receptor antagonist on immune markers in patients with congestive heart failure (CHF). Ang II stimulates production of immune factors via the Ang II type 1 receptor in vitro, and the long-term effects of Ang II type 1 receptor antagonists on plasma markers of immune activation are unknown in patients with CHF. Twenty-three patients with mild to moderate CHF with left ventricular dysfunction were randomly divided into two groups: treatment with Ang II type 1 receptor (candesartan cilexetil) (n = 14) or placebo (n = 9). We measured plasma levels of immune factors such as tumor necrosis factor alpha (TNFalpha), interleukin-6 (IL-6), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1). We also measured plasma levels of the neurohumoral factors such as atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) and cyclic guanosine monophosphate (cGMP), a biological marker of ANP and BNP. Plasma levels of TNFalpha, IL-6, sICAM-1 and sVCAM-1 were increased in the 23 CHF patients compared with normal subjects and significantly decreased after 14 weeks of candesartan cilexetil treatment, but did not change in the placebo group. Plasma levels of BNP, which is a marker of ventricular injury, significantly decreased, and the molar ratio of plasma cGMP to cardiac natriuretic peptides (ANP + BNP) was significantly increased after candesartan cilexetil treatment, but did not change in the placebo group. These findings suggest that 14 weeks of treatment with an Ang II type 1 receptor antagonist (candesartan cilexetil) decreased plasma levels of the immune markers such as TNFalpha, IL-6, sICAM-1 and sVCAM-1 and that it improved the biological compensatory action of endogenous cardiac natriuretic peptides in patients with mild to moderate CHF.

  19. Effect of biliary drainage on inducible nitric oxide synthase, CD14 and TGR5 expression in obstructive jaundice rats

    PubMed Central

    Wang, Zi-Kai; Xiao, Jian-Guo; Huang, Xue-Fei; Gong, Yi-Chun; Li, Wen

    2013-01-01

    AIM: To investigate the effect of biliary drainage on inducible nitric oxide synthase (iNOS), CD14 and TGR5 expression in rats with obstructive jaundice (OJ). METHODS: Male adult Sprague-Dawley rats were randomly assigned to four groups: OJ, sham operation (SH), internal biliary drainage (ID) and external biliary drainage (ED). Rat models were successfully established by two operations and succumbed for extraction of Kupffer cells (KCs) and liver tissue collection on the 8th and 15th day. KCs were isolated by in situ hepatic perfusion and digested with collagen IV, density gradient centrifuged by percoll reagent and purified by cell culture attachment. The isolated KCs were cultured with the endotoxin lipopolysaccharide (LPS) with and without the addition of ursodeoxycholic acid (UDCA). The expression of iNOS, CD14 and bile acid receptor-TGR5 protein in rat liver tissues was determined by immunohistochemistry. The expression of iNOS and CD14 messenger RNA (mRNA) on the isolated KCs was detected by reverse transcription polymerase chain reaction (PCR) and the TGR5 mRNA level in KCs was measured by real-time quantitative PCR. RESULTS: The iNOS protein was markedly expressed in the liver of OJ rats, but rare expressed in SH rats. After relief of OJ, the iNOS expression was decidedly suppressed in the ID group (ID vs OJ, P < 0.01), but obviously increased in rats of ED (ED vs OJ, P = 0.004). When interfered only with LPS, the expression of iNOS mRNA by KCs was increased in the OJ group compared with the SH group (P = 0.004). After relief of biliary obstruction, the iNOS mRNA expression showed slight changes in the ED group (ED vs OJ, P = 0.71), but dropped in the ID group (ID vs OJ, P = 0.001). Compared with the simple intervention with LPS, the expressions of iNOS mRNA were significantly inhibited in all four groups after interfered with both LPS and UDCA (P < 0.01, respectively). After bile duct ligation, the CD14 protein expression in rat liver was significantly

  20. An unbalanced PD-L1/CD86 ratio in CD14(++)CD16(+) monocytes is correlated with HCV viremia during chronic HCV infection.

    PubMed

    Zheng, Jiajia; Liang, Hua; Xu, Chunhui; Xu, Qiang; Zhang, Ting; Shen, Tao; Lu, Fengmin

    2014-05-01

    Circulating monocyte subsets with distinct functions play important roles in hepatitis C virus (HCV) infection. However, the mechanisms have not been well studied. In this study, we analyzed the distributions and phenotypic characteristics of three circulating monocyte subsets-CD14(++)CD16(-), CD14(++)CD16(+) and CD14(+/dim)CD16(+)-in chronic HCV-infected patients, HCV spontaneous resolvers and healthy controls, and we evaluated the possible link between HCV viremia and disease progression. Our results indicated that the frequency of the CD14(++)CD16(+) monocyte subset was decreased, and negatively correlated with HCV RNA and core antigen levels during chronic HCV infection. PD-L1 expression and the PD-L1/CD86 ratio in CD14(++)CD16(+) monocytes were higher during chronic HCV infection than in spontaneous HCV resolvers and healthy controls. The PD-L1/CD86 ratio positively correlated with HCV viral load and core antigen levels. Finally, PD-L1 was significantly increased, while cytokine secretions were dramatically decreased upon Toll-like receptor (TLR) ligand binding and HCV JFH-1stimulation. These findings indicates the compromised immune status of the CD14(++)CD16(+) monocytes during chronic HCV infection and provides new insights into the specific role of the CD14(++)CD16(+) monocytes and their significance in chronic HCV infection.

  1. Increased Serum CD14 Level Is Associated with Depletion of TNF-α in Monocytes in Migraine Patients during Interictal Period.

    PubMed

    Michalak, Slawomir; Kalinowska-Lyszczarz, Alicja; Wegrzyn, Danuta; Niezgoda, Adam; Losy, Jacek; Osztynowicz, Krystyna; Kozubski, Wojciech

    2017-02-13

    The aim of the present study was to investigate the levels of circulating CD14 in relation to the expression of tumor necrosis factor alpha (TNF-α) in monocytes, and serum levels of TNF-α and macrophage inflammatory protein-1 (MIP-1) in migraine patients. Numerous studies revealed controversial changes in the components of the immune system during attacks and the interictal period in migraine patients. Our study included 40 migraineurs and 39 controls. The levels of TNF-α, MIP-1 and CD14 were measured in peripheral monocytes and in sera with the Enzyme-Linked Immunosorbent Assay (ELISA) method, and the monocyte expression of TNF-α was also analysed by immunostaining. Serum CD14 concentrations were higher and the expression of TNF-α in monocytes was decreased in migraineurs. The serum MIP-1 level correlated with Verbal Rating Scale (VRS); the MIP-1:CD14 ratio in monocytes correlated with Visual Analogue Scale (VAS); the MIP-1:CD14 ratio correlated with Migraine Severity (MIGSEV)-Pain scores; and serum CD14 concentration correlated with migraine duration in years. Increased serum CD14 and depletion of TNF-α in monocytes can orchestrate other components of the immune system during the interictal period.

  2. Increased Serum CD14 Level Is Associated with Depletion of TNF-α in Monocytes in Migraine Patients during Interictal Period

    PubMed Central

    Michalak, Slawomir; Kalinowska-Lyszczarz, Alicja; Wegrzyn, Danuta; Niezgoda, Adam; Losy, Jacek; Osztynowicz, Krystyna; Kozubski, Wojciech

    2017-01-01

    The aim of the present study was to investigate the levels of circulating CD14 in relation to the expression of tumor necrosis factor alpha (TNF-α) in monocytes, and serum levels of TNF-α and macrophage inflammatory protein-1 (MIP-1) in migraine patients. Numerous studies revealed controversial changes in the components of the immune system during attacks and the interictal period in migraine patients. Our study included 40 migraineurs and 39 controls. The levels of TNF-α, MIP-1 and CD14 were measured in peripheral monocytes and in sera with the Enzyme-Linked Immunosorbent Assay (ELISA) method, and the monocyte expression of TNF-α was also analysed by immunostaining. Serum CD14 concentrations were higher and the expression of TNF-α in monocytes was decreased in migraineurs. The serum MIP-1 level correlated with Verbal Rating Scale (VRS); the MIP-1:CD14 ratio in monocytes correlated with Visual Analogue Scale (VAS); the MIP-1:CD14 ratio correlated with Migraine Severity (MIGSEV)-Pain scores; and serum CD14 concentration correlated with migraine duration in years. Increased serum CD14 and depletion of TNF-α in monocytes can orchestrate other components of the immune system during the interictal period. PMID:28208835

  3. Plasma kinetics and biodistribution of water-soluble CdTe quantum dots in mice: a comparison between Cd and Te

    NASA Astrophysics Data System (ADS)

    Han, Ying; Xie, Guangyun; Sun, Zhiwei; Mu, Ying; Han, Sihai; Xiao, Yang; Liu, Na; Wang, Hui; Guo, Caixia; Shi, Zhixiong; Li, Yanbo; Huang, Peili

    2011-10-01

    Water-soluble quantum dots (QDs) have shown potential as tumor diagnostic agents. However, little is known about their biological behaviors in vivo. Male ICR mice were intravenously given a single dose (2.5 μmol kg-1 body weight) of water-soluble cadmium-telluride (CdTe) QDs (the QDs are approximately 4 nm in diameter and have maximal emission at 630 nm). Inductively coupled plasma mass spectrometry (ICP-MS) was used for measuring the kinetic action of 111Cd and 125Te for 7 days. The plasma kinetics of Cd and Te followed a two-compartment model, in which Cd exhibited greater apparent volume of distribution, greater clearance, faster distribution half-life, and significantly slower elimination half-life compared to Te. Contrary to its relatively transient fate in the plasma, high levels of Cd persisted in the liver and kidneys. Te accumulated primarily in the spleen. The different plasma kinetics and distribution patterns of Cd and Te imply that CdTe QDs have been part of the degradation or aggregation in vivo.

  4. Interleukin-13 Inhibits Expression of cyp27b1 in Peripheral CD14+ Cells That Is Correlated With Vertebral Bone Mineral Density of Patients With Ulcerative Colitis.

    PubMed

    Ding, Qingfeng; Zhou, Hao; Yun, Bo; Zhou, Lingjie; Zhang, Ning; Yin, Guoyong; Fan, Jin

    2017-02-01

    Osteoporosis is a common problem in aged people and those with related diseases, such as inflammatory bowel diseases. Deregulation of vitamin D metabolism plays a role in the pathogenesis of osteoporosis. Micro RNA (miR) can regulate cytokine expression in cells. This study test a hypothesis that inflammatory cytokine interleukin (IL)-13 increases miR-19a to compromise cyp27b1 (a vitamin D hydroxylase) in peripheral CD14(+) cells. Bone mineral density of L2-L4 was measured in 20 patients with ulcerative colitis (UC) and 20 healthy subjects. Peripheral CD14(+) cells were isolated from healthy people and patients with UC. Expression of cyp27b1 by CD14(+) cells was analyzed in the presence or absence of IL-13 in the culture. We observed that bone mineral density (BMD) in UC patients was significantly lower than healthy subjects. The BMD is negatively correlated with miR-19a in peripheral CD14(+) cells. MiR-19a in peripheral CD14(+) cell was correlated with serum IL-13 in UC patients. Expression of cyp27b1 in peripheral CD14(+) cells was correlated with miR-19a and serum IL-13 in UC patients. IL-13 suppressed cyp27b1 expression in CD14(+) cells. IL-13 increased expression of miR-19a in CD14(+) cells. IL-13 suppresses cyp27b1 expression in peripheral CD14(+) cells via up regulating miR-19a expression. J. Cell. Biochem. 118: 376-381, 2017. © 2016 Wiley Periodicals, Inc.

  5. Combined Measurement of 6 Fat-Soluble Vitamins and 26 Water-Soluble Functional Vitamin Markers and Amino Acids in 50 μL of Serum or Plasma by High-Throughput Mass Spectrometry.

    PubMed

    Midttun, Øivind; McCann, Adrian; Aarseth, Ove; Krokeide, Marit; Kvalheim, Gry; Meyer, Klaus; Ueland, Per M

    2016-11-01

    Targeted metabolic profiling characterized by complementary platforms, multiplexing and low volume consumption are increasingly used for studies using biobank material. Using liquid-liquid extraction, we developed a sample workup suitable for quantification of 6 fat- and 26 water-soluble biomarkers. 50 μL of serum/plasma was mixed with dithioerythritol, ethanol, and isooctane/chloroform. The organic layer was used for analysis of the fat-soluble vitamins all-trans retinol (A), 25-hydroxyvitamin D2, 25-hydroxyvitamin D3, α-tocopherol (E), γ-tocopherol (E), and phylloquinone (K1) by LC-MS/MS. The remaining aqueous fraction was mixed with ethanol, water, pyridine, and methylchloroformate (in toluene) to derivatize the water-soluble biomarkers. The resulting toluene layer was used for GC-MS/MS analysis of alanine, α-ketoglutarate, asparagine, aspartic acid, cystathionine, total cysteine, glutamic acid, glutamine, glycine, histidine, total homocysteine, isoleucine, kynurenine, leucine, lysine, methionine, methylmalonic acid, ornithine, phenylalanine, proline, sarcosine, serine, threonine, tryptophan, tyrosine, and valine. Isotope-labeled internal standards were used for all analytes. Chromatographic run times for the LC-MS/MS and GC-MS/MS were 4.5 and 11 min, respectively. The limits of detection (LOD) for the low-concentration analytes (25-hydroxyvitamin D2, 25-hydroxyvitamin D3, and phylloquinone) were 25, 17, and 0.33 nM, respectively, while all other analytes demonstrated sensitivity significantly lower than endogenous concentrations. Recoveries ranged from 85.5-109.9% and within- and between-day coefficients of variance (CVs) were 0.7-9.4% and 1.1-17.5%, respectively. This low-volume, high-throughput multianalyte assay is currently in use in our laboratory for quantification of 32 serum/plasma biomarkers in epidemiological studies.

  6. During Hepatitis C Virus (HCV) Infection and HCV-HIV Coinfection, an Elevated Plasma Level of Autotaxin Is Associated With Lysophosphatidic Acid and Markers of Immune Activation That Normalize During Interferon-Free HCV Therapy.

    PubMed

    Kostadinova, Lenche; Shive, Carey L; Judge, Chelsey; Zebrowski, Elizabeth; Compan, Anita; Rife, Kelsey; Hirsch, Amy; Falck-Ytter, Yngve; Schlatzer, Daniela M; Li, Xiaolin; Chance, Mark R; Rodriguez, Benigno; Popkin, Daniel L; Anthony, Donald D

    2016-11-01

     Immune activation predicts morbidity during hepatitis C virus (HCV) infection and human immunodeficiency virus (HIV) infection, although mechanisms underlying immune activation are unclear. Plasma levels of autotaxin and its enzymatic product, lysophosphatidic acid (LPA), are elevated during HCV infection, and LPA activates immunocytes, but whether this contributes to immune activation is unknown.  We evaluated plasma levels of autotaxin, interleukin 6 (IL-6), soluble CD14 (sCD14), soluble CD163 (sCD163), and Mac2 binding protein (Mac2BP) during HCV infection, HIV infection, and HCV-HIV coinfection, as well as in uninfected controls, before and after HIV antiretroviral therapy (ART) initiation and during interferon-free HCV therapy.  We observed greater plasma autotaxin levels in HCV-infected and HCV-HIV-coinfected participants, compared with uninfected participants, primarily those with a higher ratio of aspartate aminotransferase level to platelet count. Autotaxin levels correlated with IL-6, sCD14, sCD163, Mac2BP, and LPA levels in HCV-infected participants and with Mac2BP levels in HCV-HIV-coinfected participants, while in HIV-infected individuals, sCD14 levels correlated with Mac2BP levels. Autotaxin, LPA, and sCD14 levels normalized, while sCD163 and Mac2BP levels partially normalized within 6 months of starting interferon-free HCV therapy. sCD163 and IL-6 levels normalized within 6 months of starting ART for HIV infection. In vitro, LPA activated monocytes.  These data indicate that elevated levels of autotaxin and soluble markers of immune activation during HCV infection are partially reversible within 6 months of initiating interferon-free HCV treatment and that autotaxin may be causally linked to immune activation during HCV infection and HCV-HIV coinfection. Published by Oxford University Press for the Infectious Diseases Society of America 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  7. Availability of zinc from infant formula by in vitro methods (solubility and dialyzability) and size-exclusion chromatography coupled to inductively coupled plasma-mass spectrometry.

    PubMed

    Gomez, Beatriz Gomez; Perez-Corona, M Teresa; Madrid, Yolanda

    2016-12-01

    Zinc bioaccessibility from infant formula was estimated by in vitro methods (solubility and dialyzability) and size-exclusion chromatography (SEC) coupled to inductively coupled plasma-mass spectrometry (ICP-MS). Infant formula samples were first characterized in terms of Zn bound to lipids and proteins and Zn distribution in the aqueous soluble protein fraction. We found that Zn is not incorporated into the lipid fraction of the samples, being mainly associated with the protein fraction (around 100%). Fractionation of Zn-containing proteins in the soluble protein fraction was achieved by SEC-ICP-MS after performing protein extraction with a solution of 100mM (pH 6.8) Tris-HCl. The percentages of zinc in the soluble protein fraction in the soy-based and lactose-free infant formula were very low, around 7 and 24%, respectively, whereas the content of Zn in the soluble protein fraction of milk-based formula was around 90%. By SEC-ICP-MS, we found that Zn is associated with low-molecular weight compounds (around 10kDa) in all the infant formulas tested. The percentages of Zn estimated in the in vitro gastrointestinal digests of the infant formula ranged from 30 to 70% and from 1 to 10% for solubility and dialyzability assays, respectively. The dialyzability test resulted in lower than expected scores, as SEC-ICP-MS analysis of the gastrointestinal extracts revealed that Zn is bound to biomolecules with a molecular weight ranging from 1 to 7kDa, which suggests that dialysis data should be interpreted with caution. Speciation studies are a valuable tool for establishing availability of nutrients and for validating data from dialyzable in vitro methods. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  8. Soluble TNF-alpha receptor 1 and IL-6 plasma levels in humans subjected to the sleep deprivation model of spaceflight

    NASA Technical Reports Server (NTRS)

    Shearer, W. T.; Reuben, J. M.; Mullington, J. M.; Price, N. J.; Lee, B. N.; Smith, E. O.; Szuba, M. P.; Van Dongen, H. P.; Dinges, D. F.

    2001-01-01

    BACKGROUND: The extent to which sleep loss may predispose astronauts to a state of altered immunity during extended space travel prompts evaluation with ground-based models. OBJECTIVE: We sought to measure plasma levels of selected cytokines and their receptors, including the putative sleep-regulation proteins soluble TNF-alpha receptor (sTNF-alpha R) I and IL-6, in human subjects undergoing 2 types of sleep deprivation during environmental confinement with performance demands. METHODS: Healthy adult men (n = 42) were randomized to schedules that varied in severity of sleep loss: 4 days (88 hours) of partial sleep deprivation (PSD) involving two 2-hour naps per day or 4 days of total sleep deprivation (TSD). Plasma samples were obtained every 6 hours across 5 days and analyzed by using enzyme-linked immunoassays for sTNF-alpha RI, sTNF-alpha RII, IL-6, soluble IL-2 receptor, IL-10, and TNF-alpha. RESULTS: Interactions between the effects of time and sleep deprivation level were detected for sTNF-alpha RI and IL-6 but not for sTNF-alpha RII, soluble IL-2 receptor, IL-10, and TNF-alpha. Relative to the PSD condition, subjects in the TSD condition had elevated plasma levels of sTNF-alpha RI on day 2 (P =.04), day 3 (P =.01), and across days 2 to 4 of sleep loss (P =.01) and elevated levels of IL-6 on day 4 (P =.04). CONCLUSIONS: Total sleep loss produced significant increases in plasma levels of sTNF-alpha RI and IL-6, messengers that connect the nervous, endocrine, and immune systems. These changes appeared to reflect elevations of the homeostatic drive for sleep because they occurred in TSD but not PSD, suggesting that naps may serve as the basis for a countermeasures approach to prolonged spaceflight.

  9. Effects of a carbohydrate-restricted diet with and without supplemental soluble fiber on plasma low-density lipoprotein cholesterol and other clinical markers of cardiovascular risk.

    PubMed

    Wood, Richard J; Fernandez, Maria Luz; Sharman, Matthew J; Silvestre, Ricardo; Greene, Christine M; Zern, Tosca L; Shrestha, Sudeep; Judelson, Daniel A; Gomez, Ana L; Kraemer, William J; Volek, Jeff S

    2007-01-01

    Carbohydrate-restricted diets (CRDs) promote weight loss, reductions in plasma triacylglycerol (TAG) levels, and increases in high-density lipoprotein cholesterol (HDL-C) levels but may cause undesirable low-density lipoprotein cholesterol (LDL-C) responses in some people. The objective of the present study was to determine the effect of adding soluble fiber to a CRD on plasma LDL-C and other traditionally measured markers of cardiovascular disease. Using a parallel-arm, double-blind, placebo-controlled design, 30 overweight and obese men (body mass index, 25-35 kg/m(2)) were randomly assigned to supplement a CRD with soluble fiber (Konjac-mannan, 3g/d) (n = 15) or placebo (n = 15). Plasma lipids, anthropometrics, body composition, blood pressure, and nutrient intake were evaluated at baseline and at 6 and 12 weeks. Compliance was excellent as assessed by 7-day weighed dietary records and ketonuria. Both groups experienced decreases in (P < .01) body weight, percent body fat, systolic blood pressure, waist circumference, and plasma glucose levels. After 12 weeks, HDL-C and TAG improved significantly in the fiber (10% and -34%) and placebo (14%, -43%) groups. LDL-C decreased by 17.6% (P < .01) at week 6 and 14.1% (P < .01) at week 12 in the fiber group. Conversely, LDL-C reductions were significant in the placebo group only after 12 weeks (-6.0%, P < .05). We conclude that although clearly effective at lowering LDL-C, adding soluble fiber to a CRD during active and significant weight loss provides no additional benefits to the diet alone. Furthermore, a CRD led to clinically important positive alterations in cardiovascular disease risk factors.

  10. Soluble TNF-alpha receptor 1 and IL-6 plasma levels in humans subjected to the sleep deprivation model of spaceflight

    NASA Technical Reports Server (NTRS)

    Shearer, W. T.; Reuben, J. M.; Mullington, J. M.; Price, N. J.; Lee, B. N.; Smith, E. O.; Szuba, M. P.; Van Dongen, H. P.; Dinges, D. F.

    2001-01-01

    BACKGROUND: The extent to which sleep loss may predispose astronauts to a state of altered immunity during extended space travel prompts evaluation with ground-based models. OBJECTIVE: We sought to measure plasma levels of selected cytokines and their receptors, including the putative sleep-regulation proteins soluble TNF-alpha receptor (sTNF-alpha R) I and IL-6, in human subjects undergoing 2 types of sleep deprivation during environmental confinement with performance demands. METHODS: Healthy adult men (n = 42) were randomized to schedules that varied in severity of sleep loss: 4 days (88 hours) of partial sleep deprivation (PSD) involving two 2-hour naps per day or 4 days of total sleep deprivation (TSD). Plasma samples were obtained every 6 hours across 5 days and analyzed by using enzyme-linked immunoassays for sTNF-alpha RI, sTNF-alpha RII, IL-6, soluble IL-2 receptor, IL-10, and TNF-alpha. RESULTS: Interactions between the effects of time and sleep deprivation level were detected for sTNF-alpha RI and IL-6 but not for sTNF-alpha RII, soluble IL-2 receptor, IL-10, and TNF-alpha. Relative to the PSD condition, subjects in the TSD condition had elevated plasma levels of sTNF-alpha RI on day 2 (P =.04), day 3 (P =.01), and across days 2 to 4 of sleep loss (P =.01) and elevated levels of IL-6 on day 4 (P =.04). CONCLUSIONS: Total sleep loss produced significant increases in plasma levels of sTNF-alpha RI and IL-6, messengers that connect the nervous, endocrine, and immune systems. These changes appeared to reflect elevations of the homeostatic drive for sleep because they occurred in TSD but not PSD, suggesting that naps may serve as the basis for a countermeasures approach to prolonged spaceflight.

  11. Plasma interferon-gamma, interleukin-10 and soluble markers of immune activation in infants with primary adenovirus (ADV) and respiratory syncytial virus (RSV) infection.

    PubMed

    Fernández, J Alonso; Tapia, Lorena; Palomino, M Angélica; Larrañaga, Carmen; Peña, Mónica; Jaramillo, Héctor

    2005-01-01

    Adenovirus (ADV) and respiratory syncytial virus (RSV) are etiological agents of acute respiratory tract infection in infants. Long-term prognosis of ADV infection includes severe lung damage, bronchiectasis and hyperlucent lung, while RSV infection is associated with development of recurrent wheezing and subsequent asthma. These differences may be related to differences in the primary immune responses elicited by these viruses. In this paper, we investigated the type of cytokine responses and the magnitude of immune activation in ADV and RSV infections in infants. We examined plasma concentrations of interferon-gamma (IFN-gamma), interleukin-10 (IL-10), soluble interleukin-2 receptor (sCD25) and soluble tumor necrosis factor receptor II (sTNFR-II) in previously healthy infants during the acute phase of primary ADV infection (n = 21) and RSV infection (n = 68), and in uninfected controls (n = 44). In ADV-infected infants, IFN-gamma plasma levels were significantly higher than those observed in RSV cases and the control group (p < 0.05). RSV cases did not show any differences in IFN-gamma plasma levels compared to the other groups. sCD25 levels were significantly higher in ADV- and RSV-infected infants than in controls (p < 0.0001), and higher in ADV than in RSV cases (p < 0.05). sTNFR-II levels were significantly higher in RSV- and ADV-infected infants than in controls (p < 0.0001, p < 0.05, respectively), and higher in RSV than in ADV infection (p < 0.05). No significant differences were observed in IL-10 plasma concentrations between the three groups. These results indicate that ADV and RSV infections in infants differ significantly with regard to the magnitude of production of interferon-gamma and soluble immune activation markers sCD25 and sTNFR-II. These immunological differences may be involved in the different clinical outcomes associated with these viral infections.

  12. Asian Zika virus strains target CD14(+) blood monocytes and induce M2-skewed immunosuppression during pregnancy.

    PubMed

    Foo, Suan-Sin; Chen, Weiqiang; Chan, Yen; Bowman, James W; Chang, Lin-Chun; Choi, Younho; Yoo, Ji Seung; Ge, Jianning; Cheng, Genhong; Bonnin, Alexandre; Nielsen-Saines, Karin; Brasil, Patrícia; Jung, Jae U

    2017-08-21

    Blood CD14(+) monocytes are frontline immunomodulators categorized into classical, intermediate or non-classical subsets, and subsequently differentiated into M1 pro- or M2 anti-inflammatory macrophages on stimulation. Although the Zika virus (ZIKV) rapidly establishes viraemia, the target cells and immune responses, particularly during pregnancy, remain elusive. Furthermore, it is unknown whether African- and Asian-lineage ZIKV have different phenotypic impacts on host immune responses. Using human blood infection, we identified CD14(+) monocytes as the primary target for African- or Asian-lineage ZIKV infection. When immunoprofiles of human blood infected with ZIKV were compared, a classical/intermediate monocyte-mediated M1-skewed inflammation by the African-lineage ZIKV infection was observed, in contrast to a non-classical monocyte-mediated M2-skewed immunosuppression by the Asian-lineage ZIKV infection. Importantly, infection of the blood of pregnant women revealed an enhanced susceptibility to ZIKV infection. Specifically, Asian-lineage ZIKV infection of pregnant women's blood led to an exacerbated M2-skewed immunosuppression of non-classical monocytes in conjunction with a global suppression of type I interferon-signalling pathway and an aberrant expression of host genes associated with pregnancy complications. Also, 30 ZIKV(+) sera from symptomatic pregnant patients showed elevated levels of M2-skewed immunosuppressive cytokines and pregnancy-complication-associated fibronectin-1. This study demonstrates the differential immunomodulatory responses of blood monocytes, particularly during pregnancy, on infection with different lineages of ZIKV.Both African and epidemic strains of Zika virus are shown to target CD14(+) monocytes, which are more susceptible in pregnant women, but African strains are associated with inflammatory responses, and epidemic strains with immunotolerance.

  13. CD14-dependent monocyte isolation enhances phagocytosis of listeria monocytogenes by proinflammatory, GM-CSF-derived macrophages.

    PubMed

    Neu, Caroline; Sedlag, Anne; Bayer, Carina; Förster, Sabine; Crauwels, Peter; Niess, Jan-Hendrik; van Zandbergen, Ger; Frascaroli, Giada; Riedel, Christian U

    2013-01-01

    Macrophages are an important line of defence against invading pathogens. Human macrophages derived by different methods were tested for their suitability as models to investigate Listeria monocytogenes (Lm) infection and compared to macrophage-like THP-1 cells. Human primary monocytes were isolated by either positive or negative immunomagnetic selection and differentiated in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF) into pro- or anti-inflammatory macrophages, respectively. Regardless of the isolation method, GM-CSF-derived macrophages (GM-Mφ) stained positive for CD206 and M-CSF-derived macrophages (M-Mφ) for CD163. THP-1 cells did not express CD206 or CD163 following incubation with PMA, M- or GM-CSF alone or in combination. Upon infection with Lm, all primary macrophages showed good survival at high multiplicities of infection whereas viability of THP-1 was severely reduced even at lower bacterial numbers. M-Mφ generally showed high phagocytosis of Lm. Strikingly, phagocytosis of Lm by GM-Mφ was markedly influenced by the method used for isolation of monocytes. GM-Mφ derived from negatively isolated monocytes showed low phagocytosis of Lm whereas GM-Mφ generated from positively selected monocytes displayed high phagocytosis of Lm. Moreover, incubation with CD14 antibody was sufficient to enhance phagocytosis of Lm by GM-Mφ generated from negatively isolated monocytes. By contrast, non-specific phagocytosis of latex beads by GM-Mφ was not influenced by treatment with CD14 antibody. Furthermore, phagocytosis of Lactococcus lactis, Escherichia coli, human cytomegalovirus and the protozoan parasite Leishmania major by GM-Mφ was not enhanced upon treatment with CD14 antibody indicating that this effect is specific for Lm. Based on these observations, we propose macrophages derived by ex vivo differentiation of negatively selected human primary monocytes as the most suitable model

  14. Use of pressurized hot water extraction and high performance liquid chromatography-inductively coupled plasma-mass spectrometry for water soluble halides speciation in atmospheric particulate matter.

    PubMed

    Moreda-Piñeiro, Jorge; Alonso-Rodríguez, Elia; Moreda-Piñeiro, Antonio; Moscoso-Pérez, Carmen; López-Mahía, Purificación; Muniategui-Lorenzo, Soledad; Prada-Rodríguez, Darío

    2012-11-15

    The feasibility of pressurized hot water extraction (PHWE) has been novelty investigated to speed up water soluble halide species (bromide, Br(-); bromate, BrO(3)(-); iodide, I(-) and iodate, IO(3)(-)) leaching from atmospheric particulate matter (PM(10) and PM(2.5)). Total bromine and iodine and total water soluble bromine and iodine have been assessed by inductively coupled plasma-mass spectrometry (ICP-MS). Water-soluble bromine and iodine species were also measured by ICP-MS after anion exchange high performance liquid chromatography (HPLC). Variables inherent to the pressurized hot water extraction process (temperature, modifier concentration, static time, pressure, number of cycles and dispersing agent mass) were fully studied. Results showed that the pressurized leaching procedure can be performed in 9 min (5 min for pre-heating, 2 min of static time, 1 min of purge time, and 1 min of end relief time). The use of diluted acetic acid as a modifier did not improve the target recoveries. Dispersing agent (diatomaceous earth) was not needed, which reduces the time for filling the cells. Water-soluble halides were reached under the following extraction conditions: extraction temperature of 100 °C, pressure of 1500 psi, static time of 2 min and 1 extraction cycle. Optimized HPLC conditions consisted of an isocratic elution with 175 mM ammonium nitrate plus 15% (v/v) methanol as mobile phase (optimum flow rate of at 1.5 mL min(-1)). Analytical performances, such as limits of detection and quantification, repeatability and analytical recoveries of the over-all procedure have been established. Results obtained show water soluble halides accounted for approximately 20.9±1.3 and 11.8±0.6% of the total bromine and total iodine, respectively. A 79 and 89% of bromine and iodine was non-water soluble, which may be organic non-water soluble species. Br(-) and IO(3)(-) were found to be the major species, and they accounted for 100% of the total water-soluble bromine and

  15. Helicobacter pylori lipopolysaccharide alters ECL cell DNA synthesis via a CD14 receptor and polyamine pathway in mastomys.

    PubMed

    Kidd, M; Tang, L H; Schmid, S; Lauffer, J; Louw, J A; Modlin, I M

    2000-01-01

    Chronic Helicobacter pylori infection is associated with alterations in gastric mucosal cell proliferation. Despite the recognition that bacterial lipopolysaccharide (LPS) is present in biologically active quantities in the gastric mucosa, the mechanisms by which it stimulates cells are largely unknown. We have previously established a gastric enterochromaffin-like (ECL) cell neoplasia model in the African rodent species Mastomys and identified that tumor ECL cell proliferation is associated with polyamine biosynthesis and ornithine decarboxylase (ODC) activity. In addition, we have shown that H. pylori LPS exhibits a specific mitogenic effect on naive ECL cells in vitro. The aim of this study was to evaluate whether H. pylori has a direct effect on tumor ECL cell proliferation in vitro and further to evaluate the possible molecular mechanisms for this effect. ECL cell neoplasia was generated in Mastomys by endogenous hypergastrinemia induced by H(2) blockade (loxtidine 1 g/kg/day) and tumor ECL cells prepared. The DNA synthesis in 24-hour cultured tumor cells was measured by bromodeoxyuridine uptake and ODC activity by (14)CO(2) formation from (14)C-ornithine. The putative LPS receptor, CD14, was evaluated by reverse-transcription polymerase chain reaction. Our results demonstrated: (1) H. pylori LPS (10(-12) to 10(-7) M) stimulated basal DNA synthesis (2.2-fold) with an estimated EC(50) of 10(-10) M; (2) this proliferative response correlated with an increase in ODC activity (1.4-fold, EC(50) approximately 10(-10) M) which could be inhibited by a specific ODC inhibitor, difluoromethyl ornithine, at 10(-9) M; (3) the CD14 receptor was identified in both naive and transformed ECL cells by reverse-transcription polymerase chain reaction, and (4) the effects of LPS were inhibited by blocking the CD14 receptor with its specific monoclonal antibody (1:100). Thus, H. pylori LPS appears to influence tumor ECL cell proliferation by activation of the intracellular

  16. Differential distribution of proteins and lipids in detergent-resistant and detergent-soluble domains in rod outer segment plasma membranes and disks

    PubMed Central

    Elliott, Michael H.; Nash, Zack A.; Takemori, Nobuaki; Fliesler, Steven J.; McClellan, Mark E.; Naash, Muna I.

    2009-01-01

    Membrane heterogeneity plays a significant role in regulating signal transduction and other cellular activities. We examined the protein and lipid components associated with the detergent-resistant membrane (DRM) fractions from retinal rod outer segment (ROS) disk and plasma membrane-enriched preparations. Proteomics and correlative western blot analysis revealed the presence of α and β subunits of the rod cGMP-gated ion channel and glucose transporter type 1, among other proteins. The glucose transporter was present exclusively in ROS plasma membrane (not disks) and was highly enriched in DRMs, as was the cGMP-gated channel β-subunit. In contrast, the majority of rod opsin and ATP-binding cassette transporter A4 was localized to detergent-soluble domains in disks. As expected, the cholesterol: fatty acid mole ratio was higher in DRMs than in the corresponding parent membranes (disk and plasma membranes, respectively) and was also higher in disks compared to plasma membranes. Furthermore, the ratio of saturated: polyunsaturated fatty acids was also higher in DRMs compared to their respective parent membranes (disk and plasma membranes). These results confirm that DRMs prepared from both disks and plasma membranes are enriched in cholesterol and in saturated fatty acids compared to their parent membranes. The dominant fatty acids in DRMs were 16: 0 and 18: 0; 22: 6n3 and 18: 1 levels were threefold higher and twofold lower, respectively, in disk-derived DRMs compared to plasma membrane-derived DRMs. We estimate, based on fatty acid recovery that DRMs account for only ~ 8% of disks and ~ 12% of ROS plasma membrane. PMID:17944869

  17. Plasma levels of soluble CD27: a simple marker to monitor immune activation during potent antiretroviral therapy in HIV-1-infected subjects

    PubMed Central

    DE MILITO, A; ALEMAN, S; MARENZI, R; SÖNNERBORG, A; FUCHS, D; ZAZZI, M; CHIODI, F

    2002-01-01

    Plasma levels of soluble CD27 (sCD27) are elevated in diseases characterized by T cell activation and are used as a marker of immune activation. We assessed the usefulness of determining plasma sCD27 as a marker for monitoring immune activation in HIV-1-infected patients treated with highly active antiretroviral therapy (HAART). A first cross-sectional examination of 68 HIV-1-infected and 18 normal subjects showed high levels of sCD27 in HIV-1 infection; plasma sCD27 was correlated to HIV-1 viraemia and inversely correlated to CD4+ T cell count. Twenty-six HIV-1-infected patients undergoing HAART were studied at baseline and after 6, 12, 18 and 24 months of therapy. Seven additional patients under HAART were analysed at baseline, during and after interruption of therapy. In the total population, HAART induced a significant and progressive reduction, but not a normalization, of plasma levels of sCD27 after 24 months. A full normalization of plasma sCD27 was observed in the virological responders (undetectable HIV-1 RNA at months 18 and 24) and also in patients with moderate immunodeficiency at baseline (CD4+ T cell count >200 cells/mm3). Changes in plasma neopterin paralleled the changes in sCD27 but only baseline sCD27 levels were predictive of a greater increase in CD4+ T cell count during the follow-up. Discontinuation of therapy resulted in a rapid increase of sCD27 plasma levels associated with viraemia rebound and drop in CD4+ T cell count. Our findings suggest that plasma sCD27 may represent an alternative and simple marker to monitor immune activation during potent antiretroviral therapy. HIV-1-induced immune activation can be normalized by HAART in successfully treated patients where the disease is not advanced. PMID:11966765

  18. Detection of cell surface determinants for anti-Leu M3 (CD14), MY9 (CD33) and MY4 (CD14) and phagocytic function of cord blood monocytes in the course of gestational age.

    PubMed

    Gengenbacher, D; Salm, H; Vogt, A; Schneider, H

    1998-07-01

    To investigate immunological functions of cord blood cells, cord blood monocytes of 84 term and preterm newborn infants were examined in presenting cell surface determinants for the monoclonal antibodies anti-Leu M3 (CD14), My9 (CD33) and My4 (CD14) using an alkaline-phosphatase-anti-alkaline-phosphatase staining method. We also tested the phagocytic function of these cells by incorporation of Candida albicans in a 60 min incubation assay. Mean values for antigen expression ranged from 60 to 70%, whereby My4 showed a statistically significant higher level compared to Anti-Leu M3 and My9 (P = 0.05). Changes of antigen expression were statistically significant by the time of maturation. Statistically significant differences between male and female newborns were observed in 26-33 and 36-37 weeks of gestation. Considering the modus of delivery, no significantly higher amount of antigen-presenting monocytes were found in newborns of spontaneous birth compared to cesarean sections. However, no statistically significant differences in phagocytosis were observed between term and preterm newborn infants, mature newborns showed higher levels of both antigen expressing and phagocytic monocytes in contrast to a higher number of phagocytic cord blood monocytes only in premature infants.

  19. [The frequency of peripheral blood CD14(+)HLA-DR(-/low) MDSCs is negatively correlated with the inflammation in patients with chronic hepatitis B].

    PubMed

    Zhang, Hao; Guan, Shihe; Yang, Kai; Ye, Jun; Yan, Kaili; Pan, Ying; Wu, Yuanyuan; Wang, Aihua; Sun, Beibei

    2015-10-01

    To study the frequency of CD14⁺HLA-DR(-/low) myeloid-derived suppressor cells (MDSCs) in the peripheral blood of chronic hepatitis B (CHB) patients and the relationship with biochemical characteristics, viral load and liver pathology. The frequency of CD14⁺HLA-DR(-/low) MDSCs in the peripheral blood of 96 patients with CHB and 20 healthy control cases were detected by flow cytometry. Ultrasound-guided liver biopsies as well as HBV-related serological tests were performed in HBV-infected individuals to analyze the biochemical characteristics, viral load and pathology. The data were assessed using Spearman correlation analysis. The frequency of the peripheral blood CD14⁺HLA-DR(-/low) MDSCs in the 96 CHB cases was (6.03 ± 0.09)%, which was significantly higher than that of the 20 healthy control cases (1.87 ± 0.05)%. The group of HBeAg positive cases had a significantly higher frequency of the peripheral blood CD14⁺HLA-DR(-/low) MDSCs compared with the group of HBeAg negative cases and the healthy control group. The frequency of CD14⁺HLA-DR(-/low) MDSCs in the peripheral blood was negatively correlated with serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. There was no correlation between the frequency of peripheral blood CD14⁺HLA-DR(-/low) MDSCs and HBV load. The frequency of CD14⁺HLA-DR(-/low) MDSCs in the peripheral blood was negatively correlated with the liver inflammation grade, but not related with the fibrosis stage in patients with CHB. The frequency of CD14⁺HLA-DR(-/low) MDSCs is negatively correlated with the inflammation of CHB.

  20. Probiotic Treatment Decreases the Number of CD14-Expressing Cells in Porcine Milk Which Correlates with Several Intestinal Immune Parameters in the Piglets

    PubMed Central

    Scharek-Tedin, Lydia; Kreuzer-Redmer, Susanne; Twardziok, Sven Olaf; Siepert, Bianca; Klopfleisch, Robert; Tedin, Karsten; Zentek, Jürgen; Pieper, Robert

    2015-01-01

    Modulating the mucosal immune system of neonates by probiotic treatment of their mothers is a promising approach which can only be investigated through the use of animal models. Here, we used sows and their piglets to investigate the impact of a bacterial treatment on the sow’s milk and on the neonate piglet intestinal immune system. In previous experiments, feed supplementation of sows with the probiotic Enterococcus faecium NCIMB 10415 during pregnancy and lactation had been shown to affect intestinal microbiota and cytokine expression of the offspring during the suckling and weaning periods. We therefore investigated the composition of the milk from treated sows in comparison to samples from a control group. In treated sows, the amount of lactose increased, and the somatic cell numbers were reduced. In all milk samples, the percentage of cells expressing membranous CD14 (mCD14) was greater than the fractions of immune cells, indicating expression of mCD14 on mammary epithelial cells. However, in the milk of E. faecium-treated sows, mCD14+ cells were reduced. Furthermore, the number of CD14+ milk cells was positively correlated with the percentages of B cells and activated T cells in the ileal MLN of the piglets. This study provides evidence for the expression of mCD14 by the porcine mammary epithelium, and suggests an immunological effect of mCD14+ milk cells on the piglets’ intestinal immune system. Our study further suggests that mCD14+ mammary epithelial cell populations can be modulated by probiotic feed supplementation of the sow. PMID:25806034

  1. Microglial Heparan Sulfate Proteoglycans Facilitate the Cluster-of-Differentiation 14 (CD14)/Toll-like Receptor 4 (TLR4)-Dependent Inflammatory Response*

    PubMed Central

    O'Callaghan, Paul; Li, Jin-Ping; Lannfelt, Lars; Lindahl, Ulf; Zhang, Xiao

    2015-01-01

    Microglia rapidly mount an inflammatory response to pathogens in the central nervous system (CNS). Heparan sulfate proteoglycans (HSPGs) have been attributed various roles in inflammation. To elucidate the relevance of microglial HSPGs in a pro-inflammatory response we isolated microglia from mice overexpressing heparanase (Hpa-tg), the HS-degrading endoglucuronidase, and challenged them with lipopolysaccharide (LPS), a bacterial endotoxin. Prior to LPS-stimulation, the LPS-receptor cluster-of-differentiation 14 (CD14) and Toll-like receptor 4 (TLR4; essential for the LPS response) were similarly expressed in Ctrl and Hpa-tg microglia. However, compared with Ctrl microglia, Hpa-tg cells released significantly less tumor necrosis factor-α (TNFα), essentially failed to up-regulate interleukin-1β (IL1β) and did not initiate synthesis of proCD14. Isolated primary astroyctes expressed TLR4, but notably lacked CD14 and in contrast to microglia, LPS challenge induced a similar TNFα response in Ctrl and Hpa-tg astrocytes, while neither released IL1β. The astrocyte TNFα-induction was thus attributed to CD14-independent TLR4 activation and was unaffected by the cells HS status. Equally, the suppressed LPS-response in Hpa-tg microglia indicated a loss of CD14-dependent TLR4 activation, suggesting that microglial HSPGs facilitate this process. Indeed, confocal microscopy confirmed interactions between microglial HS and CD14 in LPS-stimulated microglia and a potential HS-binding motif in CD14 was identified. We conclude that microglial HSPGs facilitate CD14-dependent TLR4 activation and that heparanase can modulate this mechanism. PMID:25869127

  2. In situ Delivery of Antigen to DC-SIGN(+)CD14(+) Dermal Dendritic Cells Results in Enhanced CD8(+) T-Cell Responses.

    PubMed

    Fehres, Cynthia M; van Beelen, Astrid J; Bruijns, Sven C M; Ambrosini, Martino; Kalay, Hakan; van Bloois, Louis; Unger, Wendy W J; Garcia-Vallejo, Juan J; Storm, Gert; de Gruijl, Tanja D; van Kooyk, Yvette

    2015-09-01

    CD14(+) dendritic cells (DCs) present in the dermis of human skin represent a large subset of dermal DCs (dDCs) that are considered macrophage-like cells with poor antigen (cross)-presenting capacity and limited migratory potential to the lymph nodes. CD14(+) dDC highly express DC-specific ICAM-3-grabbing non-integrin (DC-SIGN), a receptor containing potent endocytic capacity, facilitating intracellular routing of antigens to major histocompatibility complex I and II (MHC-I andII) loading compartments for the presentation to antigen-specific CD8(+) and CD4(+) T cells. Here we show using a human skin explant model that the in situ targeting of antigens to DC-SIGN using glycan-modified liposomes enhances the antigen-presenting capacity of CD14(+) dDCs. Intradermal vaccination of liposomes modified with the DC-SIGN-targeting glycan Lewis(X), containing melanoma antigens (MART-1 or Gp100), accumulated in CD14(+) dDCs and resulted in enhanced Gp100- or MART-1-specific CD8(+) T-cell responses. Simultaneous intradermal injection of the cytokines GM-CSF and IL-4 as adjuvant enhanced the migration of the skin DCs and increased the expression of DC-SIGN on the CD14(+) and CD1a(+) dDCs. These data demonstrate that human CD14(+) dDCs exhibit potent cross-presenting capacity when targeted in situ through DC-SIGN.

  3. Circulating CD14(+)HLA-DR(-/low) myeloid-derived suppressor cell is an indicator of poor prognosis in patients with ESCC.

    PubMed

    Huang, Haitao; Zhang, Guangbo; Li, Guangbin; Ma, Haitao; Zhang, Xueguang

    2015-09-01

    Accumulating evidences demonstrate that a population of suppressive cells known as myeloid-derived suppressor cells (MDSCs) is key immune modulators which suppress antitumor immunity. In this study, we found that the level of circulating CD14(+)HLA-DR(-/low) cells in patients was significantly higher than that of healthy donors and was correlated with tumor burden, lymph node metastasis, and tumor, node, and metastasis (TNM) clinical stage. More importantly, we for the first time find the level of CD14(+)HLA-DR(-/low) is a biological indicator of poor prognosis through the analysis of 3-year overall survival. Furthermore, we evidenced that the proportion of CD14(+)HLA-DR(-/low) cells in the tumor metastatic tumor-draining lymph nodes (TDLNs) was notably higher compared to tumor-free TDLNs. Additionally, CD14(+)HLA-DR(-/low) cells from esophageal squamous cell carcinoma (ESCC) patients expressed dramatically increased programmed death ligand 1 (PD-L1) comparing to that from healthy control. Subsequently, blocking PD-L1 pathway by antibody could effectively reverse the suppressive effect on autologous T cell proliferation mediated by CD14(+)HLA-DR(-/low) cells in vitro. In conclusion, our data revealed CD14(+)HLA-DR(-/low) MDSCs which increase in ESCC patients is a novel poor prognostic indicator and may exert immunosuppressive properties through PD-L1/PD-1 pathway.

  4. CD14 (C-260T) polymorphism is not associated with sudden infant death syndrome (SIDS) in a large South Australian cohort.

    PubMed

    Highet, Amanda R; Gibson, Catherine S; Goldwater, Paul N

    2011-02-01

    Similarities have been drawn between models of endotoxic shock and gross and microscopic pathology observed in sudden infant death syndrome (SIDS) cases. Polymorphisms in genes that influence the expression of endotoxin receptors could affect the outcome of toxaemia, and could, therefore, play a role in SIDS. The CD14 gene promoter contains a single nucleotide polymorphism that affects the level of CD14 gene expression. The TT genotype of the CD14 (C-260T) polymorphism causes a significantly higher density of CD14 receptor expression on monocytes which makes the individual more sensitive to endotoxin than those with the wild-type (CC). This investigation was designed to determine whether SIDS infants have a higher frequency of the CD14 (C-260T) polymorphism compared with non-SIDS controls. One hundred and sixteen SIDS and 228 control infants were genotyped using PCR followed by restriction fragment length analysis of amplified product. Carriage of the TT or CT genotypes did not significantly differ between SIDS and control infants (P = 0.218 and 0.081, respectively). The frequencies observed in the control group were consistent with Hardy-Weinberg equilibrium and did not differ significantly from the published frequencies in Caucasian Australians. These results suggest that CD14 (C-260T) polymorphism is unlikely to be implicated in SIDS.

  5. The soluble form of Alzheimer's amyloid beta protein is complexed to high density lipoprotein 3 and very high density lipoprotein in normal human plasma.

    PubMed

    Koudinov, A; Matsubara, E; Frangione, B; Ghiso, J

    1994-12-15

    The amyloid fibrils of Alzheimer's neuritic plaques and cerebral blood vessels are mainly composed of aggregated forms of a 39 to 44 amino acids peptide, named amyloid beta (A beta). A similar although soluble form of A beta (sA beta) has been identified in plasma, cerebrospinal fluid and cell culture supernatants, indicating that it is produced under physiologic conditions. We report here that sA beta in normal human plasma is associated with lipoprotein particles, in particular to the HDL3 and VHDL fractions where it is complexed to ApoJ and, to a lesser extent, to ApoAI. This was assessed by immunoprecipitation experiments of purified plasma lipoproteins and lipoprotein-depleted plasma and confirmed by means of amino acid sequence analysis. Moreover, biotinylated synthetic peptide A beta 1-40 was traced in normal human plasma in in vitro experiments. As in the case of sA beta, biotinylated A beta 1-40 was specifically recovered in the HDL3 and VHDL fractions. This data together with the previous demonstration that A beta 1-40 is taken up into the brain via a specific mechanism and possibly as an A beta 1-40-ApoJ complex indicate a role for HDL3- and VHDL-containing ApoJ in the transport of the peptide in circulation and suggest their involvement in the delivery of sA beta across the blood-brain barrier.

  6. Optimization of protein solubilization for the analysis of the CD14 human monocyte membrane proteome using LC-MS/MS.

    PubMed

    Ye, Xiaoying; Johann, Donald J; Hakami, Ramin M; Xiao, Zhen; Meng, Zhaojing; Ulrich, Robert G; Issaq, Haleem J; Veenstra, Timothy D; Blonder, Josip

    2009-11-02

    Proteomic profiling of membrane proteins is of vital importance in the search for disease biomarkers and drug development. However, the slow pace in this field has resulted mainly from the difficulty to analyze membrane proteins by mass spectrometry (MS). The objective of this investigation was to explore and optimize solubilization of membrane proteins for shotgun membrane proteomics of the CD14 human monocytes by examining different systems that rely on: i) an organic solvent (methanol) ii) an acid-labile detergent 3-[3-(1,1-bisalkyloxyethyl)pyridin-1-yl]propane-1-sulfonate (PPS), iii) a combination of both agents (methanol+PPS). Solubilization efficiency of different buffers was first compared using bacteriorhodopsin as a model membrane protein. Selected approaches were then applied on a membrane subproteome isolated from a highly enriched human monocyte population that was approximately 98% positive for CD14 expression as determined by FACS analysis. A methanol-based buffer yielded 194 proteins of which 93 (48%) were mapped as integral membrane proteins. The combination of methanol and acid-cleavable detergent gave similar results; 203 identified proteins of which 93 (46%) were mapped integral membrane proteins. However, employing PPS 216 proteins were identified of which 75 (35%) were mapped as integral membrane proteins. These results indicate that methanol alone or in combination with PPS yielded significantly higher membrane protein identification/enrichment than the PPS alone.

  7. Transient Migration of Large Numbers of CD14++ CD16+ Monocytes to the Draining Lymph Node after Onset of Inflammation

    PubMed Central

    Lund, Hege; Boysen, Preben; Åkesson, Caroline Piercey; Lewandowska-Sabat, Anna Monika; Storset, Anne K.

    2016-01-01

    The dynamics of skin-draining cells following infection or vaccination provide important insight into the initiation of immune responses. In this study, the local recruitment and activation of immune cells in draining lymph nodes (LNs) was studied in calves in an adjuvant-induced inflammation. A transient but remarkably strong recruitment of monocytes was demonstrated after onset of inflammation, constituting up to 41% of live cells in the draining LNs after 24 h. Numerous CD14+ cells were visualized in subcutaneous tissues and draining LNs, and the majority of these cells did not express dendritic cell-associated markers CD205 and CD11c. In the LNs, recruited cells were predominately of a CD14++ and CD16+ phenotype, consistent with an intermediate monocyte subset characterized to possess a high inflammatory potential. Moreover, monocytes from the draining LN showed a high expression of genes coding for pro-inflammatory cytokines, including IL-1β, IL-6, TNFa, and TGFβ. Shortly after their appearance in the LN cortical areas, the monocytes had moved into the medulla followed by an increase in peripheral blood. In conclusion, this study provides novel information on in vivo monocyte recruitment and migration after onset of inflammation. PMID:27621730

  8. A phased strategy to differentiate human CD14+monocytes into classically and alternatively activated macrophages and dendritic cells.

    PubMed

    Zarif, Jelani C; Hernandez, James R; Verdone, James E; Campbell, Scott P; Drake, Charles G; Pienta, Kenneth J

    2016-01-01

    There are currently several in vitro strategies to differentiate human CD14(+) monocytes isolated from peripheral blood mononuclear cells (PBMCs) into the M1 or M2 macrophage cell types. Each cell type is then verified using flow cytometric analysis of cell-surface markers. Human CD14(+) monocytes have the potential to differentiate into M1 and M2 macrophages, both of which demonstrate varying degrees of cell-surface antigen overlap. Using multiple surface markers with current macrophage polarization protocols, our data reveal several limitations of currently used methods, such as highly ambiguous cell types that possess cell-surface marker overlap and functional similarities. Utilizing interleukin-6 (IL-6) and two phases of cytokine exposure, we have developed a protocol to differentiate human monocytes into M1, M2, or dendritic cells (DCs) with greater efficiency and fidelity relative to macrophages and DCs that are produced by commonly used methods. This is achieved via alterations in cytokine composition, dosing, and incubation times, as well as improvements in verification methodology. Our method reliably reproduces human in vitro monocyte-derived DCs and macrophage models that will aid in better defining and understanding innate and adaptive immunity, as well as pathologic states.

  9. The association of C (-260)-->T polymorphism in CD14 promoter and Chlamydia pneumoniae infection in ischemic stroke patients.

    PubMed

    Lin, Tsun-Mei; Chen, Chih-Hung; Wu, Hua-Lin; Wang, Chiou-Huey; Chen, Yi-Ling; Eng, Hock-Liew

    2008-10-01

    The C (-260) --> T polymorphism has been reported to regulate CD14 gene expression. It has also been implicated in atherosclerotic diseases, and in addition, it could be a genetic factor responsible for interindividual differences in the susceptibility to Chlamydia pneumoniae infection. This case-control study is aimed at evaluating the association between CD14 promoter polymorphisms, frequency of persistent C pneumoniae infection, and anti-chlamydial heat shock protein 60 (cHsp60) induction in stroke patients. Persistent C pneumoniae infection was observed in 43.3% of control subjects and 53.3% of patients (P = .005). The odds ratio of persistent infection was 3.25 for the CC genotype in stroke patients (P = .018). However, the serologic positive responses to cHsp60 in people with the TT genotype were significantly lower in stroke patients (9.3% vs 38.6%; P < .001). Our findings suggest that persistent C pneumoniae infection is independently associated with stroke in CC genotypes and the lowering of cHsp60 antibody levels in TT genotypes.

  10. Rosiglitazone influences the expression of leukocyte adhesion molecules and CD14 receptor in type 2 diabetes mellitus patients.

    PubMed

    Štulc, T; Svobodová, H; Krupičková, Z; Doležalová, R; Marinov, I; Češka, R

    2014-01-01

    Diabetes mellitus is associated with increased inflammatory response, which may contribute to atherosclerosis progression. Experimental results demonstrated anti-inflammatory activity of glitazones; their effect on leukocyte adhesion molecules has not been studied to date. We therefore studied the effect of rosiglitazone treatment on leukocyte surface expression of adhesion molecules in patients with type 2 diabetes mellitus and compared our results with findings in healthy subjects. 33 subjects with type 2 diabetes and 32 healthy controls were included; patients were examined at baseline and after 5 months of rosiglitazone treatment (4 mg/d). Leukocyte expression of adhesion molecules LFA-1, CD18 and ICAM-1 was quantified using flow cytometry; in addition, CD14 (lipopolysaccharide receptor) expression was analyzed as a marker of nonspecific immunity. The expression of examined molecules at baseline was higher in patients compared to controls. Despite only mild decrease in blood glucose, rosiglitazone treatment induced substantial decrease of CD18 and CD14 expression and borderline decrease of LFA-1 and ICAM-1 expression (on monocytes only). We thus observed improvement in the expression of leukocyte inflammatory markers after rosiglitazone treatment. This effect is supposed to be mediated by direct effect of rosiglitazone on PPAR-gamma receptors on leukocytes.

  11. Death ligand TRAIL, secreted by CD1a+ and CD14+ cells in blister fluids, is involved in killing keratinocytes in toxic epidermal necrolysis.

    PubMed

    de Araujo, Elisabeth; Dessirier, Valérie; Laprée, Geneviève; Valeyrie-Allanore, Laurence; Ortonne, Nicolas; Stathopoulos, Efstathios N; Bagot, Martine; Bensussan, Armand; Mockenhaupt, Maja; Roujeau, Jean-Claude; Tsapis, Andreas

    2011-02-01

    Toxic epidermal necrolysis (TEN) is characterized by an acute detachment and destruction of keratinocytes, affecting large areas of the skin. It is often related to adverse drug reactions. Previous studies have shown that effector CD8+ T cells, which accumulate in the blister fluid, are functionally cytotoxic and act through a classical perforin/granzyme B pathway. It has recently been shown that these cytotoxic T cells also secrete granulysin peptide, which is lethal to keratinocytes. These cytotoxic T cells exert their killer activity against autologous keratinocytes in the presence of the drug. However, they are unlikely to be the only effectors of TEN. We therefore searched for soluble death factors in the blister fluids that might kill keratinocytes. We found that the amounts of interferon-γ, TRAIL and TNF-α proteins were significantly greater in TEN blister fluids than in all controls (normal sera, TEN sera, burns and Eosinophilic pustular folliculitis blister fluids) and TNF-like weak inducer of apoptosis (TWEAK) amounts are also greater in all controls except burns. We showed that these proteins acted in synergy to induce the death of keratinocytes in vitro. We also found that TRAIL and TWEAK were secreted by CD1a+ and CD14+ cells present in the blister fluids. Thus, in addition to MHC class I-restricted cytotoxic T lymphocytes (CTLs), which lyse keratinocytes, ligands secreted by non-lymphoid cells capable of inducing keratinocyte death in an MHC class I-independent manner, also seem to be present in the blister fluids of patients with TEN.

  12. Association between intraoperative ventilator settings and plasma levels of soluble receptor for advanced glycation end-products in patients without pre-existing lung injury.

    PubMed

    Jabaudon, Matthieu; Futier, Emmanuel; Roszyk, Laurence; Sapin, Vincent; Pereira, Bruno; Constantin, Jean-Michel

    2015-10-01

    The soluble form of the receptor for advanced glycation end-products (sRAGE) is elevated and correlated with severity in patients with acute respiratory distress syndrome (ARDS). The impact of ventilator settings on plasma levels of sRAGE, in patients with or without pre-existing lung injury, remains under-investigated to date. Our objective was to assess the effects of a lung-protective ventilation strategy (combining low tidal volume, positive end-expiratory pressure and recruitment maneuvers), as compared with a non-protective approach (with high tidal volume and zero end-expiratory pressure), on plasma levels of sRAGE in patients without lung injury undergoing major abdominal surgery. Plasma samples were obtained from 95 patients enrolled in a large randomized controlled trial of lung-protective ventilation for major abdominal surgery. Plasma levels of sRAGE were measured in duplicate with an enzyme-linked immunoassay on day 1, immediately after surgery, and on postoperative days 1, 3 and 7. Early postoperative plasma levels of sRAGE were significantly lower in the lung-protective ventilation group (n = 47) than in the non-protective ventilation group (n = 48) (mean (standard deviation), 1782 (836) vs 2171 (1678) pg/mL, respectively, P = 0.03). Intraoperative changes in plasma sRAGE were associated with postoperative hypoxemia and ARDS. A lung-protective ventilation strategy decreased plasma sRAGE in patients without lung injury undergoing major abdominal surgery compared with the patients with non-protective ventilation. This intraoperative decrease could reflect a lesser degree of epithelial injury. © 2015 Asian Pacific Society of Respirology.

  13. A Novel HPLC Method for the Concurrent Analysis and Quantitation of Seven Water-Soluble Vitamins in Biological Fluids (Plasma and Urine): A Validation Study and Application

    PubMed Central

    Grotzkyj Giorgi, Margherita; Howland, Kevin; Martin, Colin; Bonner, Adrian B.

    2012-01-01

    An HPLC method was developed and validated for the concurrent detection and quantitation of seven water-soluble vitamins (C, B1, B2, B5, B6, B9, B12) in biological matrices (plasma and urine). Separation was achieved at 30°C on a reversed-phase C18-A column using combined isocratic and linear gradient elution with a mobile phase consisting of 0.01% TFA aqueous and 100% methanol. Total run time was 35 minutes. Detection was performed with diode array set at 280 nm. Each vitamin was quantitatively determined at its maximum wavelength. Spectral comparison was used for peak identification in real samples (24 plasma and urine samples from abstinent alcohol-dependent males). Interday and intraday precision were <4% and <7%, respectively, for all vitamins. Recovery percentages ranged from 93% to 100%. PMID:22536136

  14. Lipopolysaccharide Causes an Increase in Intestinal Tight Junction Permeability in Vitro and in Vivo by Inducing Enterocyte Membrane Expression and Localization of TLR-4 and CD14

    PubMed Central

    Guo, Shuhong; Al-Sadi, Rana; Said, Hamid M.; Ma, Thomas Y.

    2014-01-01

    Bacterial-derived lipopolysaccharides (LPS) play an essential role in the inflammatory process of inflammatory bowel disease. A defective intestinal tight junction (TJ) barrier is an important pathogenic factor of inflammatory bowel disease and other inflammatory conditions of the gut. Despite its importance in mediating intestinal inflammation, the physiological effects of LPS on the intestinal epithelial barrier remain unclear. The major aims of this study were to determine the effects of physiologically relevant concentrations of LPS (0 to 1 ng/mL) on intestinal barrier function using an in vitro (filter-grown Caco-2 monolayers) and an in vivo (mouse intestinal perfusion) intestinal epithelial model system. LPS, at physiologically relevant concentrations (0 to 1 ng/mL), in the basolateral compartment produced a time-dependent increase in Caco-2 TJ permeability without inducing cell death. Intraperitoneal injection of LPS (0.1 mg/kg), leading to clinically relevant plasma concentrations, also caused a time-dependent increase in intestinal permeability in vivo. The LPS-induced increase in intestinal TJ permeability was mediated by an increase in enterocyte membrane TLR-4 expression and a TLR-4–dependent increase in membrane colocalization of membrane-associated protein CD14. In conclusion, these studies show for the first time that LPS causes an increase in intestinal permeability via an intracellular mechanism involving TLR-4–dependent up-regulation of CD14 membrane expression. PMID:23201091

  15. The Effects of Fat-soluble Vitamin Administration on Plasma Vitamin Status of Nursing Pigs Differ When Provided by Oral Administration or Injection

    PubMed Central

    Jang, Y. D.; Lindemann, M. D.; Monegue, H. J.; Stuart, R. L.

    2014-01-01

    Four experiments were conducted to investigate the effect of fat-soluble vitamin administration to sows or newborn pigs on plasma vitamin status. In Exp. 1 and 2, a total of 24 and 43 newborn pigs were allotted to control and vitamin treatments (vitamin D3 with variable addition of vitamins A and E) orally or by i.m. injection. In Exp. 3, pigs from Exp. 2 were allotted to 2 treatments (±vitamins D3 and E in drinking water) for 14 d postweaning. In Exp. 4, twenty-four gestating sows were used for 2 treatments (±injection of a vitamin D3/A/E product 2 wk prepartum). In Exp. 1 and 2, when vitamin D3 was administrated orally or by i.m. injection on d 1 of age, pigs had increased plasma 25-hydroxycholecalciferol (25-OH D3) concentration 10 d after administration compared with control pigs (p<0.05). The injectable administration with vitamin D3 and E was able to achieve higher plasma 25-OH D3 (p<0.05) and α-tocopherol (p<0.05) concentrations than oral administration. At weaning, the pigs in the injection group had higher plasma 25-OH D3 concentration than those in the other groups in both studies (p<0.05). In Exp. 3, water supplementation of vitamin D3 and E postweaning increased plasma 25-OH D3 and α-tocopherol concentrations at d 14 postweaning (p<0.01). In Exp. 4, when sows were injected with the vitamin D3 product prepartum, serum 25-OH D3 concentrations of sows at farrowing (p<0.01), and in their progeny at birth (p<0.01) and weaning (p<0.05) were increased. These results demonstrated that fat-soluble vitamin administration to newborn pigs increased plasma 25-OH D3 concentration regardless of administration routes and α-tocopherol concentration by the injectable route, and that water supplementation of vitamin D3 and E to nursery pigs increased plasma 25-OH D3 and α-tocopherol concentrations. Additionally, injecting sows with vitamin D3 prepartum increased 25-OH D3 in sows and their offspring. If continued research demonstrates that the serum levels of 25-OH D

  16. Soluble arsenic and selenium species in fly ash/organic waste-amended soils using ion chromatography-inductively coupled plasma mass spectrometry

    SciTech Connect

    Jackson, B.P.; Miller, W.P.

    1999-01-15

    Mixing coal fly ash with an organic waste increases macronutrient content and may make land application of fly ash a viable disposal alternative. However, trace element chemistry of these mixed waste products warrants investigation. Speciation of As and Se in soil solutions of fly ash-, poultry litter- and sewage sludge-amended soils was determined over a 10-day period by ion chromatography coupled to inductively coupled plasma mass spectrometry (IC-ICP-MS). Detection limits were 0.031, 0.028, 0.051, 0.161, 0.497, and 0.660 {micro}g L{sup {minus}1} for dimethylarsinate (DMA), As(III), monomethylarsonate (MMA), As(V), Se(IV), and Se(VI), respectively. Arsenic was highly water-soluble from poultry litter and appeared to be predominantly As(V). Arsenic(V) was the predominant species in soil amended with two fly ashes. Application of fly ash/poultry litter mixtures increased As solubility and led to the prevalence of DMA as the major As species. DMA concentrations of these soil solutions decreased rapidly over the sampling period relative to As(V), suggesting that DMA readily underwent mineralization in the soil solution. Se(VI) was the predominant soluble Se species in all treatments indicating rapid oxidation of Se(IV) initially solubilized from the fly ashes.

  17. Development of an Advanced HPLC–MS/MS Method for the Determination of Carotenoids and Fat-Soluble Vitamins in Human Plasma

    PubMed Central

    Hrvolová, Barbora; Martínez-Huélamo, Miriam; Colmán-Martínez, Mariel; Hurtado-Barroso, Sara; Lamuela-Raventós, Rosa Maria; Kalina, Jiří

    2016-01-01

    The concentration of carotenoids and fat-soluble vitamins in human plasma may play a significant role in numerous chronic diseases such as age-related macular degeneration and some types of cancer. Although these compounds are of utmost interest for human health, methods for their simultaneous determination are scarce. A new high pressure liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS) method for the quantification of selected carotenoids and fat-soluble vitamins in human plasma was developed, validated, and then applied in a pilot dietary intervention study with healthy volunteers. In 50 min, 16 analytes were separated with an excellent resolution and suitable MS signal intensity. The proposed HPLC–MS/MS method led to improvements in the limits of detection (LOD) and quantification (LOQ) for all analyzed compounds compared to the most often used HPLC–DAD methods, in some cases being more than 100-fold lower. LOD values were between 0.001 and 0.422 µg/mL and LOQ values ranged from 0.003 to 1.406 µg/mL, according to the analyte. The accuracy, precision, and stability met with the acceptance criteria of the AOAC (Association of Official Analytical Chemists) International. According to these results, the described HPLC-MS/MS method is adequately sensitive, repeatable and suitable for the large-scale analysis of compounds in biological fluids. PMID:27754400

  18. Development of an Advanced HPLC-MS/MS Method for the Determination of Carotenoids and Fat-Soluble Vitamins in Human Plasma.

    PubMed

    Hrvolová, Barbora; Martínez-Huélamo, Miriam; Colmán-Martínez, Mariel; Hurtado-Barroso, Sara; Lamuela-Raventós, Rosa Maria; Kalina, Jiří

    2016-10-14

    The concentration of carotenoids and fat-soluble vitamins in human plasma may play a significant role in numerous chronic diseases such as age-related macular degeneration and some types of cancer. Although these compounds are of utmost interest for human health, methods for their simultaneous determination are scarce. A new high pressure liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS) method for the quantification of selected carotenoids and fat-soluble vitamins in human plasma was developed, validated, and then applied in a pilot dietary intervention study with healthy volunteers. In 50 min, 16 analytes were separated with an excellent resolution and suitable MS signal intensity. The proposed HPLC-MS/MS method led to improvements in the limits of detection (LOD) and quantification (LOQ) for all analyzed compounds compared to the most often used HPLC-DAD methods, in some cases being more than 100-fold lower. LOD values were between 0.001 and 0.422 µg/mL and LOQ values ranged from 0.003 to 1.406 µg/mL, according to the analyte. The accuracy, precision, and stability met with the acceptance criteria of the AOAC (Association of Official Analytical Chemists) International. According to these results, the described HPLC-MS/MS method is adequately sensitive, repeatable and suitable for the large-scale analysis of compounds in biological fluids.

  19. Variation in the Lectin-like Oxidized LDL Receptor 1 (LOX-1) Gene Is Associated With Plasma Soluble LOX-1 Levels

    PubMed Central

    Brinkley, Tina E.; Kume, Noriaki; Mitsuoka, Hirokazu; Brown, Michael D.; Phares, Dana A.; Ferrell, Robert. E.; Kita, Toru; Hagberg, James M.

    2009-01-01

    The lectin-like ox-LDL receptor 1 (LOX-1) expressed on vascular cells plays a major role in atherogenesis by internalizing and degrading oxidized LDL. LOX-1 can be cleaved from the cell surface and released as soluble LOX-1 (sLOX-1), and elevated sLOX-1 levels may be indicative of atherosclerotic plaque instability. We examined associations between the LOX-1 3′UTR-C/T and G501C polymorphisms and plasma sLOX-1 levels in 97 healthy older men and women. The frequencies for the 3′UTR-T and 501C alleles were 46% and 10%, respectively. Plasma sLOX-1 levels were significantly higher in the 3′UTR CC genotype group compared to both the CT (p=0.02) and TT (p=0.002) genotype groups. Plasma sLOX-1 were also significantly higher in the 501GC genotype group compared to the GG genotype group (p=0.004). In univariate analyses, sLOX-1 levels were significantly associated with both the 3′UTR-C/T and G501 C polymorphisms. These associations remained significant after adjusting for age, gender, race, and BMI. In conclusion, variation in the LOX-1 gene is associated with plasma sLOX-1 levels in older men and women. PMID:18469066

  20. Variation in the human lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) gene is associated with plasma soluble LOX-1 levels.

    PubMed

    Brinkley, Tina E; Kume, Noriaki; Mitsuoka, Hirokazu; Brown, Michael D; Phares, Dana A; Ferrell, Robert E; Kita, Toru; Hagberg, James M

    2008-09-01

    The lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) expressed on vascular cells plays a major role in atherogenesis by internalizing and degrading oxidized low-density lipoprotein. LOX-1 can be cleaved from the cell surface and released as soluble LOX-1 (sLOX-1), and elevated sLOX-1 levels may be indicative of atherosclerotic plaque instability. We examined associations between the LOX-1 gene 3'UTR-C/T and G501C polymorphisms and plasma sLOX-1 levels in 97 healthy older men and women. The frequencies for the 3'UTR-T and 501C alleles were 46 and 10%, respectively. Plasma sLOX-1 levels were significantly higher in the 3'UTR CC genotype group compared with both the CT (P=0.02) and TT genotype groups (P=0.002). Plasma sLOX-1 levels were also significantly higher in the 501GC genotype group compared with the GG genotype group (P=0.004). In univariate analyses, sLOX-1 levels were significantly associated with both the 3'UTR-C/T and G501C polymorphisms. These associations remained significant after adjusting for age, sex, race and body mass index. In conclusion, variation in the LOX-1 gene is associated with plasma sLOX-1 levels in older men and women.

  1. Soluble HLA-G and HLA-E Levels in Bone Marrow Plasma Samples Are Related to Disease Stage in Neuroblastoma Patients

    PubMed Central

    Pozzi, Sarah; Carlini, Barbara; Amoroso, Loredana; Corrias, Maria Valeria

    2016-01-01

    The role of nonclassical HLA-class Ib molecules HLA-G and HLA-E in the progression of Neuroblastoma (NB), the most common pediatric extracranial solid tumor, has been characterized in the last years. Since BM infiltration by NB cells is an adverse prognostic factor, we have here analyzed for the first time the concentration of soluble (s)HLA-G and HLA-E in bone marrow (BM) plasma samples from NB patients at diagnosis and healthy donors. sHLA-G and sHLA-E are present in BM plasma samples, and their levels were similar between NB patients and controls, thus suggesting that these molecules are physiologically released by resident or stromal BM cell populations. This hypothesis was supported by the finding that sHLA-G and sHLA-E levels did not correlate with BM infiltration and other adverse prognostic factors (MYCN amplification and age at diagnosis). In contrast, BM plasma levels of both molecules were higher in patients with metastatic disease than in patients with localized NB, thus suggesting that concentration of these molecules might be correlated with disease progression. The prognostic role of sHLA-G and sHLA-E concentration in the BM plasma for NB patients will be evaluated in future studies, by analyzing the clinical outcome of the same NB patients at follow-up. PMID:27610393

  2. Milk fat globule epidermal growth factor-factor VIII is down-regulated in sepsis via the lipopolysaccharide-CD14 pathway.

    PubMed

    Komura, Hidefumi; Miksa, Michael; Wu, Rongqian; Goyert, Sanna M; Wang, Ping

    2009-01-01

    Phagocytosis prevents the release of potentially harmful or immunogenic materials from dying cells. Milk fat globule epidermal growth factor (EGF)-factor VIII (MFG-E8) mediates the clearance of apoptotic cells. We have previously shown that the administration of MFG-E8-rich exosomes from immature dendritic cells promotes the phagocytosis of apoptotic cells and improves survival in sepsis. Because endotoxin is elevated in polymicrobial sepsis, we hypothesized that down-regulation of MFG-E8 is mediated via the LPS-CD14 pathway, eventually leading to the accruement of apoptotic cells. Polymicrobial sepsis was induced by cecal ligation and puncture (CLP) in CD14-deficient (CD14(-/-)), TLR4-mutated and wild-type (WT) mice. In addition, endotoxemia was elicited by i.p. injection of LPS. LPS was also neutralized by pretreating CLP-induced WT mice with polymyxin B. Splenic MFG-E8 expression, phagocytic activity, and apoptosis were assessed 5 and 20 h after CLP or 5 h after LPS administration. In septic WT mice, MFG-E8 mRNA and protein levels were suppressed by 49 and 33%, respectively. Endotoxemia reduced MFG-E8 mRNA expression in a dose dependent manner and the down-regulation of MFG-E8 mRNA expression in CLP-induced sepsis was attenuated by polymyxin B. This CLP-induced suppression was not observed in both CD14(-/-) and TLR4-mutated mice. CLP significantly decreased phagocytic activity of peritoneal macrophages in WT (by 30%), but not in CD14(-/-) mice. CLP also induced significant apoptosis in the spleen of WT (by 61%), but less in CD14(-/-) mice. Thus, MFG-E8 production is down-regulated in sepsis by LPS-CD14 dependent fashion, leading to a reduction of phagocytosis of apoptotic cells.

  3. Adsorptive depletion of CD14⁺CD16⁺ proinflammatory monocyte phenotype in patients with generalized pustular psoriasis: clinical efficacy and effects on cytokines.

    PubMed

    Fujisawa, Tomomi; Murase, Kana; Kanoh, Hiroyuki; Takemura, Masao; Ohnishi, Hidenori; Seishima, Mariko

    2012-10-01

    Generalized pustular psoriasis (GPP) is a subtype of psoriasis with strong association with activated neutrophils. Adsorptive granulocyte and monocyte apheresis (GMA) is an extracorporeal intervention for selective depletion of activated granulocytes and monocytes. However, the immunological mechanism(s) for the effect of GMA on patients is not fully defined yet. We investigated the effects of GMA on the ratio of CD14(+) CD16(+) proinflammatory monocytes/CD14(+) monocytes and cytokine/chemokine production by these leukocytes including CXCL8, CCL2, CCL3, CCL4, CCL5 and tumor necrosis factor (TNF)-α in five patients with active GPP. CD14(+) CD16(+) monocytes were significantly elevated in patients with active GPP, and GMA markedly reduced the CD14(+) CD16(+) /CD14(+) ratio together with improvement of patients' clinical symptoms. The serum levels of CXCL8, CCL3 and CCL4 were increased in active GPP patients. Likewise, CCL2 production from monocytes was increased in active GPP patients. Further, CCL3 and CCL4 production from monocytes in active GPP patients were reduced after a course of GMA. Serum CCL5 level and the release of CCL5 from monocytes in active GPP were significantly reduced, but TNF-α level in active GPP was similar to controls. Based on these results, we believe that in addition to neutrophils, elevated CD14(+) CD16(+) proinflammatory monocytes are part of the immune pathology in GPP. Accordingly, selective depletion of CD14(+) CD16(+) monocytes by GMA should be therapeutic in this condition. © 2012 The Authors. Therapeutic Apheresis and Dialysis © 2012 International Society for Apheresis.

  4. HCMV protein LUNA is required for viral reactivation from latently infected primary CD14⁺ cells.

    PubMed

    Keyes, Lisa R; Hargett, Danna; Soland, Melisa; Bego, Mariana G; Rossetto, Cyprian C; Almeida-Porada, Graca; St Jeor, Stephen

    2012-01-01

    Human cytomegalovirus (HCMV) is a member of the Herpesviridae family that infects individuals throughout the world. Following an initial lytic stage, HCMV can persist in the individual for life in a non-active (or latent) form. During latency, the virus resides within cells of the myeloid lineage. The mechanisms controlling HCMV latency are not completely understood. A latency associated transcript, UL81-82ast, encoding the protein LUNA (Latency Unique Natural Antigen) was identified from latently infected donors in vivo. To address the role of the UL81-82ast protein product LUNA, in the context of the viral genome, we developed a recombinant HCMV bacterial artificial chromosome (BAC) that does not express LUNA. This construct, LUNA knockout FIX virus (FIX-ΔLUNA), was used to evaluate LUNA's role in HCMV latency. The FIX-ΔLUNA virus was able to lytically infect Human Fibroblast (HF) cells, showing that LUNA is not required to establish a lytic infection. Interestingly, we observed significantly higher viral copy numbers in HF cells infected with FIX-ΔLUNA when compared to FIX-WT virus. Furthermore, FIX-WT and FIX-ΔLUNA genomic DNA and transcription of UL81-82ast persisted over time in primary monocytes. In contrast, the levels of UL138 transcript expression in FIX-ΔLUNA infected HF and CD14⁺ cells was 100 and 1000 fold lower (respectively) when compared to the levels observed for FIX-WT infection. Moreover, FIX-ΔLUNA virus failed to reactivate from infected CD14⁺ cells following differentiation. This lack of viral reactivation was accompanied by a lack of lytic gene expression, increase in viral copy numbers, and lack of the production of infectious units following differentiation of the cells. Our study suggests that the LUNA protein is involved in regulating HCMV reactivation, and that in the absence of LUNA, HCMV may not be able to enter a proper latent state and therefore cannot be rescued from the established persistent infection in CD14⁺ cells.

  5. Potential utility of soluble p3-alcadein α plasma levels as a biomarker for sporadic Alzheimer's disease.

    PubMed

    Kamogawa, Kenji; Kohara, Katsuhiko; Tabara, Yasuharu; Takita, Rie; Miki, Tetsuro; Konno, Tomoko; Hata, Saori; Suzuki, Toshiharu

    2012-01-01

    Alcadeins (Alcs) constitute a family of neuronal type I membrane proteins (α, β, γ) that share identical localization and function to the amyloid-β protein precursor (AβPP) in the brain. Alcs are proteolyzed in neurons through successive cleavages via secretases, resulting in non-aggregative p3-Alc, where p3 corresponds to the AβPP-fragment. We found p3-Alcα detected in human plasma reflected the pathological process of amyloid-β accumulation in Alzheimer's disease (AD) patients and therefore investigated the utility of p3-Alcα as a plasma biomarker in AD. We measured p3-Alcα plasma levels in 83 sporadic-AD, 18 mild cognitive impaired (MCI), and 24 control subjects using the sandwich-ELISA system. Pooled samples with previously published data (171 AD and 45 controls) were also analyzed. The plasma p3-Alcα concentrations in patients with AD and MCI were significantly higher compared with control subjects (224.7 ± 40.4, 223.3 ± 53.9, and 189.1 ± 32.9 pg/ml, respectively; p = 0.0012). In AD patients, the plasma p3-Alcα concentration significantly correlated with age (r = 0.23, p = 0.037) and serum creatinine levels (r = 0.23, p = 0.0012). Even after adjusting for confounding factors of age, gender, renal function, and ApoE-ε4, high plasma p3-Alcα levels were correlated with significant AD risk, with an odds ratio 1.47 (95% confidence interval: 1.18-1.93, p = 0.0019) for every 10 pg/ml increase. Pooled analysis further confirmed these findings. Increased plasma p3-Alcα, evident in the early stages of cognitive impairment, suggests that Alc metabolites are useful plasma biomarkers of AD.

  6. Recombinant interferon gamma up-regulates in vivo and down-regulates in vitro monocyte CD14 antigen expression in cancer patients.

    PubMed

    Landmann, R; Wesp, M; Ludwig, C; Obrist, R; Knüsli, C; Obrecht, J P

    1990-01-01

    The expression of the monocyte membrane glycoprotein CD14 was measured and related to the serum interferon gamma (IFN gamma) concentration in thirteen patients with disseminated cancer during treatment with human recombinant interferon gamma (rIFN gamma). The drug was administered by continuous subcutaneous infusion using an escalating dose schedule, starting at 50 micrograms/day or 100 micrograms/day and increasing weekly up to 600 micrograms/day, if tolerated. Treatment was continued at a mean maximal tolerated dose of 200 micrograms/day for a median duration of 43 days. Serum IFN gamma concentration and monocyte CD14 antigen expression (immunofluorescence with the monoclonal antibody LeuM3 and fluorescence-activated cell sorting analysis) were determined weekly. The serum IFN gamma concentration was positively correlated with the rIFN gamma dose (P less than 0.05). Therapy induced a dose-dependent enhancement of CD14 antigen expression. The increase in mean CD14 fluorescence intensity was on average 60% after 3 weeks of treatment at a mean dose of 220 micrograms rIFN gamma/day and was reversed after withdrawal of therapy. Patients with a rapidly rising serum IFN gamma concentration (starting dose 100 micrograms/day) showed a smaller increment in CD14 fluorescence intensity than those with slowly rising serum IFN gamma levels (starting dose 50 micrograms/day). Since rIFN gamma is known to down-regulate CD14 antigen expression in vitro, monocytes from patients off therapy and from healthy volunteers were cultured with this cytokine. A similar decrease of CD14 fluorescence was observed in both groups. In patients several factors, such as IFN gamma concentration, duration of drug effect and type of serum, were evaluated and could not explain the discrepant in vivo and in vitro findings. In conclusion, the monocyte marker CD14 was found to be differentially regulated by rIFN gamma in vivo and in vitro. In vivo, secondary mediators, induced by rIFN gamma and acting on

  7. Negative Regulation of Toll-like Receptor-4 Signaling through the Binding of Glycosylphosphatidylinositol-anchored Glycoprotein, CD14, with the Sialic Acid-binding Lectin, CD33*

    PubMed Central

    Ishida, Akiko; Akita, Kaoru; Mori, Yugo; Tanida, Shuhei; Toda, Munetoyo; Inoue, Mizue; Nakada, Hiroshi

    2014-01-01

    When monocyte-derived immature dendritic cells (imDCs) were stimulated with LPS in the presence of anti-CD33/Siglec-3 mAb, the production of IL-12 and phosphorylation of NF-κB decreased significantly. The cell surface proteins of imDCs were chemically cross-linked, and CD33-linked proteins were analyzed by SDS-PAGE and immunoblotting. It was CD14 that was found to be cross-linked with CD33. A proximity ligation assay also indicated that CD33 was colocalized with CD14 on the cell surface of imDCs. Sialic acid-dependent binding of CD33 to CD14 was confirmed by a plate assay using recombinant CD33 and CD14. Three types of cells (HEK293T cells expressing the LPS receptor complex (Toll-like receptor (TLR) cells), and the LPS receptor complex plus either wild-type CD33 (TLR/CD33WT cells) or mutated CD33 without sialic acid-binding activity (TLR/CD33RA cells)) were prepared, and then the binding and uptake of LPS were investigated. Although the level of LPS bound on the cell surface was similar among these cells, the uptake of LPS was reduced in TLR/CD33WT cells. A higher level of CD14-bound LPS and a lower level of TLR4-bound LPS were detected in TLR/CD33WT cells compared with the other two cell types, probably due to reduced presentation of LPS from CD14 to TLR4. Phosphorylation of NF-κB after stimulation with LPS was also compared. Wild-type CD33 but not mutated CD33 significantly reduced the phosphorylation of NF-κB. These results suggest that CD14 is an endogenous ligand for CD33 and that ligation of CD33 with CD14 modulates with the presentation of LPS from CD14 to TLR4, leading to down-regulation of TLR4-mediated signaling. PMID:25059667

  8. Negative regulation of Toll-like receptor-4 signaling through the binding of glycosylphosphatidylinositol-anchored glycoprotein, CD14, with the sialic acid-binding lectin, CD33.

    PubMed

    Ishida, Akiko; Akita, Kaoru; Mori, Yugo; Tanida, Shuhei; Toda, Munetoyo; Inoue, Mizue; Nakada, Hiroshi

    2014-09-05

    When monocyte-derived immature dendritic cells (imDCs) were stimulated with LPS in the presence of anti-CD33/Siglec-3 mAb, the production of IL-12 and phosphorylation of NF-κB decreased significantly. The cell surface proteins of imDCs were chemically cross-linked, and CD33-linked proteins were analyzed by SDS-PAGE and immunoblotting. It was CD14 that was found to be cross-linked with CD33. A proximity ligation assay also indicated that CD33 was colocalized with CD14 on the cell surface of imDCs. Sialic acid-dependent binding of CD33 to CD14 was confirmed by a plate assay using recombinant CD33 and CD14. Three types of cells (HEK293T cells expressing the LPS receptor complex (Toll-like receptor (TLR) cells), and the LPS receptor complex plus either wild-type CD33 (TLR/CD33WT cells) or mutated CD33 without sialic acid-binding activity (TLR/CD33RA cells)) were prepared, and then the binding and uptake of LPS were investigated. Although the level of LPS bound on the cell surface was similar among these cells, the uptake of LPS was reduced in TLR/CD33WT cells. A higher level of CD14-bound LPS and a lower level of TLR4-bound LPS were detected in TLR/CD33WT cells compared with the other two cell types, probably due to reduced presentation of LPS from CD14 to TLR4. Phosphorylation of NF-κB after stimulation with LPS was also compared. Wild-type CD33 but not mutated CD33 significantly reduced the phosphorylation of NF-κB. These results suggest that CD14 is an endogenous ligand for CD33 and that ligation of CD33 with CD14 modulates with the presentation of LPS from CD14 to TLR4, leading to down-regulation of TLR4-mediated signaling. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. JAM-A and ALCAM are therapeutic targets to inhibit diapedesis across the BBB of CD14+CD16+ monocytes in HIV-infected individuals.

    PubMed

    Williams, Dionna W; Anastos, Kathryn; Morgello, Susan; Berman, Joan W

    2015-02-01

    Monocyte transmigration across the BBB is a critical step in the development of cognitive deficits termed HAND that affect 40-70% of HIV-infected individuals, even with successful antiretroviral therapy. The monocyte subsets that enter the CNS during HIV infection are not fully characterized. We examined PBMC from HIV-positive individuals from 2 distinct cohorts and enumerated monocyte populations, characterized their transmigration properties across an in vitro human BBB model, and identified surface proteins critical for the entry of these cells into the CNS. We demonstrated that the frequency of peripheral blood CD14(+)CD16(+) and CD14(low)CD16(+) monocytes was increased in HIV-seropositive compared with -seronegative individuals, despite virologic control. We showed that CD14(+)CD16(+) monocytes selectively transmigrated across our BBB model as a result of their increased JAM-A and ALCAM expression. Antibody blocking of these proteins inhibited diapedesis of CD14(+)CD16(+) monocytes but not of T cells from the same HIV-infected people across the BBB. Our data indicate that JAM-A and ALCAM are therapeutic targets to decrease the entry of CD14(+)CD16(+) monocytes into the CNS of HIV-seropositive individuals, contributing to the eradication of neuroinflammation, HAND, and CNS viral reservoirs.

  10. CD14 and Complement Crosstalk and Largely Mediate the Transcriptional Response to Escherichia coli in Human Whole Blood as Revealed by DNA Microarray

    PubMed Central

    Lau, Corinna; Nygård, Ståle; Fure, Hilde; Olstad, Ole Kristoffer; Holden, Marit; Lappegård, Knut Tore; Brekke, Ole-Lars; Espevik, Terje; Hovig, Eivind; Mollnes, Tom Eirik

    2015-01-01

    Systemic inflammation like in sepsis is still lacking specific diagnostic markers and effective therapeutics. The first line of defense against intruding pathogens and endogenous damage signals is pattern recognition by e.g., complement and Toll-like receptors (TLR). Combined inhibition of a key complement component (C3 and C5) and TLR-co-receptor CD14 has been shown to attenuate certain systemic inflammatory responses. Using DNA microarray and gene annotation analyses, we aimed to decipher the effect of combined inhibition of C3 and CD14 on the transcriptional response to bacterial challenge in human whole blood. Importantly, combined inhibition reversed the transcriptional changes of 70% of the 2335 genes which significantly responded to heat-inactivated Escherichia coli by on average 80%. Single inhibition was less efficient (p<0.001) but revealed a suppressive effect of C3 on 21% of the responding genes which was partially counteracted by CD14. Furthermore, CD14 dependency of the Escherichia coli-induced response was increased in C5-deficient compared to C5-sufficient blood. The observed crucial distinct and synergistic roles for complement and CD14 on the transcriptional level correspond to their broad impact on the inflammatory response in human blood, and their combined inhibition may become inevitable in the early treatment of acute systemic inflammation. PMID:25706641

  11. Reconstruction of LPS Transfer Cascade Reveals Structural Determinants within LBP, CD14, and TLR4-MD2 for Efficient LPS Recognition and Transfer.

    PubMed

    Ryu, Je-Kyung; Kim, Soo Jin; Rah, Sang-Hyun; Kang, Ji In; Jung, Hi Eun; Lee, Dongsun; Lee, Heung Kyu; Lee, Jie-Oh; Park, Beom Seok; Yoon, Tae-Young; Kim, Ho Min

    2017-01-17

    Lipopolysaccharide (LPS), the major component of the outer membrane of Gram-negative bacteria, binds Toll-like receptor 4 (TLR4)-MD2 complex and activates innate immune responses. LPS transfer to TLR4-MD2 is catalyzed by both LPS binding protein (LBP) and CD14. To define the sequential molecular interactions underlying this transfer, we reconstituted in vitro the entire LPS transfer process from LPS micelles to TLR4-MD2. Using electron microscopy and single-molecule approaches, we characterized the dynamic intermediate complexes for LPS transfer: LBP-LPS micelles, CD14-LBP-LPS micelle, and CD14-LPS-TLR4-MD2 complex. A single LBP molecule bound longitudinally to LPS micelles catalyzed multi-rounds of LPS transfer to CD14s that rapidly dissociated from LPB-LPS complex upon LPS transfer via electrostatic interactions. Subsequently, the single LPS molecule bound to CD14 was transferred to TLR4-MD2 in a TLR4-dependent manner. The definition of the structural determinants of the LPS transfer cascade to TLR4 may enable the development of targeted therapeutics for intervention in LPS-induced sepsis.

  12. Chimeric Anti-CD14 IGG2/4 Hybrid Antibodies for Therapeutic Intervention in Pig and Human Models of Inflammation

    PubMed Central

    Lau, Corinna; Gunnarsen, Kristin S.; Høydahl, Lene S.; Andersen, Jan Terje; Berntzen, Gøril; Pharo, Anne; Lindstad, Julie K.; Ludviksen, Judith K.; Brekke, Ole-Lars; Barratt-Due, Andreas; Nielsen, Erik Waage; Stokes, Christopher R.; Espevik, Terje; Sandlie, Inger

    2013-01-01

    CD14 is a key recognition molecule of innate immune responses, interacting with several TLRs. TLR signaling cross-talks extensively with the complement system, and combined CD14 and complement inhibition has been proved effective in attenuating inflammatory responses. Pig models of human diseases have emerged as valuable tools to study therapeutic intervention, but suitable neutralizing Abs are rare. Undesired Fc-mediated functions, such as platelet activation and IL-8 release induced by the porcine CD14-specific clone Mil2, limit further studies. Therefore, an inert human IgG2/IgG4 hybrid C region was chosen for an rMil2. As revealed in ex vivo and in vivo pig experiments, rMil2 inhibited the CD14-mediated proinflammatory cytokine response similar to the original clone, but lacked the undesired Fc-effects, and inflammation was attenuated further by simultaneous complement inhibition. Moreover, rMil2 bound porcine FcRn, a regulator of t1/2 and biodistribution. Thus, rMil2, particularly combined with complement inhibitors, should be well suited for in vivo studies using porcine models of diseases, such as sepsis and ischemia-reperfusion injury. Similarly, the recombinant anti-human CD14 IgG2/4 Ab, r18D11, was generated with greatly reduced Fc-mediated effects and preserved inhibitory function ex vivo. Such Abs might be drug candidates for the treatment of innate immunity-mediated human diseases. PMID:24062486

  13. Differential expression of CD14, CD36 and the LDL receptor on human monocyte-derived macrophages. A novel cell culture system to study macrophage differentiation and heterogeneity.

    PubMed

    Wintergerst, E S; Jelk, J; Asmis, R

    1998-09-01

    Macrophages are key players in many aspects of human physiology and disease. It has been hypothesized that in a given microenvironment monocytes differentiate into specific subpopulations with distinct functions. In order to study the role of macrophage heterogeneity in atherogenesis, we established a novel isolation and culture technique for human monocyte-derived macrophages. The present technique does not select for monocyte subpopulations prior to the onset of differentiation. Monocytes were cultured for 2 weeks in the presence of autologous lymphocytes before being plated quantitatively. They differentiated into mature macrophages in terms of morphology, lipid composition, and biological activity. Based on phagocytic activity as well as on the expression of CD14, CD36, and the low-density lipoprotein (LDL) receptor, we have identified macrophage subpopulations that may play distinct roles in atherogenesis. While virtually all adherence-purified monocytes expressed CD14, CD36, and the LDL-R, we characterized three subpopulations of macrophages based on the expression of these antigens: CD36+CD14-LDL-R-(58+/-12%), CD36+CD14+LDL-R+(18+/-5%), the remaining cells being CD36-CD14- LDL-R-. The first two subsets decreased in size during further differentiation (51+/-12% and 8+/-3%, respectively). Our culture technique may also serve as a good model for studying the implications of macrophage heterogeneity in diseases other than atherosclerosis.

  14. Inhibition of transglutaminase 2 reduces efferocytosis in human macrophages: Role of CD14 and SR-AI receptors.

    PubMed

    Eligini, S; Fiorelli, S; Tremoli, E; Colli, S

    2016-10-01

    Transglutaminase 2 (TGM2), a member of the transglutaminase family of enzymes, is a multifunctional protein involved in numerous events spanning from cell differentiation, to signal transduction, apoptosis, and wound healing. It is expressed in a variety of cells, macrophages included. Macrophage TGM2 promotes the clearance of apoptotic cells (efferocytosis) and emerging evidence suggests that defective efferocytosis contributes to the consequences of inflammation-associated diseases, including atherosclerotic lesion progression and its sequelae. Of interest, active TGM2 identified in human atherosclerotic lesions plays critical roles in plaque stability through effects on matrix cross-linking and TGFβ activity. This study explores the mechanisms by which TGM2 controls efferocytosis in human macrophages. Herein we show that TGM2 increases progressively during monocyte differentiation towards macrophages and controls their efferocytic potential as well as morphology and viability. Two experimental approaches that took advantage of the inhibition of TGM2 activity and protein silencing give proof that TGM2 reduction significantly impairs macrophage efferocytosis. Among the mechanisms involved we highlighted a role of the receptors CD14 and SR-AI whose levels were markedly reduced by TGM2 inhibition. Conversely, CD36 receptor and αvβ3 integrin levels were not influenced. Of note, lipid accumulation and IL-10 secretion were reduced in macrophages displaying defective efferocytosis. Overall, our data define a crucial role of TGM2 activity during macrophage differentiation via mechanisms involving CD14 and SR-AI receptors and show that TGM2 inhibition triggers a pro-inflammatory phenotype. Copyright © 2016 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

  15. Role of a novel soluble nucleotide phospho-hydrolase from sheep plasma in inhibition of platelet reactivity: hemostasis, thrombosis, and vascular biology.

    PubMed

    Birk, Alex V; Bubman, Darya; Broekman, M Johan; Robertson, Hugh D; Drosopoulos, Joan H F; Marcus, Aaron J; Szeto, Hazel H

    2002-02-01

    Ecto- and exoenzymes that metabolize extracellular adenosine diphosphate (ADP), the major promoter of platelet activation and recruitment, are of potential clinical importance because they can metabolically prevent excessive thrombus growth. An ecto-ADPase (CD39, NTPDase1) has been identified on endothelial cells. We demonstrate that ADP and adenosine triphosphate (ATP) are rapidly metabolized to adenosine monophosphate (AMP) in sheep plasma at pH 7.4. This hydrolysis is sensitive to P(1), P(5)-di-(adenosine-5') pentaphosphate (Ap(5)A), and ethylene glycol bis (beta-aminoethyl ether) - N, N, N(-), N(-) tetra-acetate (EGTA) but insensitive to tetramisole (an alkaline phosphatase inhibitor). A specific phosphodiesterase substrate, p -nitrophenol-5'-thymidine monophosphate (TMP) (p -Nph-5'-TMP), was readily hydrolyzed in sheep plasma at a rate of approximately 0.25 nmol/min/mg protein, and this hydrolysis was inhibited by ADP, ATP, and Ap(5)A. Furthermore, 200-fold purified p -Nph-5'-TMP-hydrolyzing activity also hydrolyzed ATP and ADP directly to AMP. When ADP was preincubated in plasma, its ability to induce platelet aggregation was inhibited in a time-dependent manner. This effect was abolished by Ap(5)A. The inhibitory effects on platelet aggregation correlated with hydrolysis of the ADP in plasma. These data suggest that the endogenous soluble plasma phosphohydrolase metabolizes ATP and ADP by means of cleavage of the alpha-beta-phosphodiester bond of nucleoside 5'-phosphate derivatives. This novel biochemical activity inhibits platelet reactivity through hydrolysis of extracellular nucleotides released by activated platelets during (patho)physiological processes, serving a homeostatic and antithrombotic function in vivo.

  16. Plasma levels of soluble CD30 are increased in children with chronic renal failure and with primary growth deficiency and decrease during treatment with recombination human growth hormone.

    PubMed

    Barbano, G; Cappa, F; Prigione, I; Pistoia, V; Cohen, A; Chiesa, S; Gusmano, R; Perfumo, F

    2001-09-01

    Previous studies have suggested that in vivo Th2 lymphocyte activation is related to increased soluble CD30 (sCD30) plasma levels. As various hormones (dehydroepiandrosterone, glucocorticoids, progesterone) can regulate the Th1/Th2 balance, and because growth hormone (GH) enhances lymphocyte function, we measured sCD30 plasma levels, before and after treatment with recombinant human GH (rhGH), in children with growth failure due to chronic renal failure (CRF) or to isolated GH deficiency in order to evaluate the potential effects of rhGH treatment on Th1/Th2 balance. sCD30 plasma levels were determined by ELISA assay in 30 children with CRF (mean age 10.7+/-3.7 years), in five children with isolated GH deficiency (mean age 11.4+/-2.6 years), and in 10 normal controls (mean age 10.1+/-3.5 years). sCD30 levels were higher in the 30 children with CRF than in the 10 controls (179.8+/-79.4 vs 11.3+/-10.9 U/ml, P<0.001) exhibiting an inverse correlation with glomerular filtration rate (GFR) (r=-0.7860, P<0.001). In 11 children with CRF, after 19.9+/-16.7 months of rhGH treatment, a decrease of sCD30 plasma level (170+/-50 vs 134+/-49 U/ml, P<0.01) was observed. The five children with primary GH deficiency had higher sCD30 plasma level than controls (mean 147+/-105 vs 11+/-10 U/ml, P<0.004) and sCD30 plasma levels decreased to 95.2+/-109.6 U/ml after rhGH treatment. The finding that rhGH treatment decreased sCD30 plasma levels in children with CRF, and that children with primary GH deficiency had higher sCD30 plasma levels than controls, suggest that GH may regulate CD30 expression and possibly the balance of Th1/Th2. Whether the uraemia-induced increase in sCD30 is due to decreased renal excretion, to overproduction or both, remains to be determined.

  17. Quantification of phenolic acids and their methylates, glucuronides, sulfates and lactones metabolites in human plasma by LC-MS/MS after oral ingestion of soluble coffee.

    PubMed

    Marmet, Cynthia; Actis-Goretta, Lucas; Renouf, Mathieu; Giuffrida, Francesca

    2014-01-01

    Chlorogenic acids and derivatives like phenolic acids are potentially bioactive phenolics, which are commonly found in many foods. Once absorbed, chlorogenic and phenolic acids are highly metabolized by the intestine and the liver, producing glucuronidated and/or sulphated compounds. These metabolites were analyzed in human plasma using a validated liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method. After protein precipitation, phenolic acids and their metabolites were extracted by using ethanol and chromatographic separation was achieved by reversed-phase using an Acquity UPLC BEH C18 column combined with a gradient elution system using 1% acetic acid aqueous solution and 1% acetic acid with 100% acetonitrile. The method was able to quantify 56 different compounds including 24 phenolic acids, 4 lactones, 15 sulfates and 13 glucuronides metabolites between 5 and 1000nM in plasma for most of them, except for m-dihydrocoumaric acid, 5-ferulloylquinic-glucuronide, 4-methoxycinnamic acid, 3-phenylpropionic acid, 3-(4-methoxyphenyl)propionic acid (25 to 1000nM) and p-dihydrocoumaric acid (50-1000nM). Values of repeatability and intermediate reproducibility were below 15% of deviation in general, and maximum 20% for the lowest concentrations. The validated method was successfully applied to quantify phenolic acids and their metabolites in plasma obtained after oral ingestion of soluble coffee. In conclusion, the developed and validated method is proved to be very sensitive, accurate and precise for the quantification of these possible dietary phenols.

  18. Posttransfusion purpura: conversion of PLA1-negative platelets to the PLA1-positive phenotype by stored plasma is not due to the presence of soluble PLA1 antigen.

    PubMed

    Ehmann, W C; Dancis, A; Ferziger, R; Karpatkin, S

    1990-11-01

    Three previous articles have reported a PLA1 antigen in the plasma of stored blood which is capable of binding to PLA1-negative platelets in the presence of divalent cations, rendering them PLA1 positive. Such a mechanism could explain the enigma of posttransfusion purpura (PTP), i.e., severe thrombocytopenia in a healthy subject with PLA1-negative platelets secondary to the infusion of blood containing PLA1-positive platelets. We find that the PLA1 antigen of stored blood is due to the presence of platelet fragments which can be removed by centrifugation and that divalent cation-chelating agents play no role in the apparent binding of these fragments to platelets. The apparent conversion of PLA1-negative platelets to the PLA1-positive phenotype by incubation with stored plasma from a PLA1-positive subject is due to the cosedimentation of platelet fragments with the platelets. No soluble PLA1 antigen was found in the plasma of five patients with acute posttransfusion purpura.

  19. Heterotrimeric G proteins physically associated with the lipopolysaccharide receptor CD14 modulate both in vivo and in vitro responses to lipopolysaccharide.

    PubMed Central

    Solomon, K R; Kurt-Jones, E A; Saladino, R A; Stack, A M; Dunn, I F; Ferretti, M; Golenbock, D; Fleisher, G R; Finberg, R W

    1998-01-01

    Septic shock induced by lipopolysaccharide (LPS) triggering of cytokine production from monocytes/macrophages is a major cause of morbidity and mortality. The major monocyte/macrophage LPS receptor is the glycosylphosphatidylinositol (GPI)-anchored glycoprotein CD14. Here we demonstrate that CD14 coimmunoprecipitates with Gi/Go heterotrimeric G proteins. Furthermore, we demonstrate that heterotrimeric G proteins specifically regulate CD14-mediated, LPS-induced mitogen-activated protein kinase (MAPK) activation and cytokine production in normal human monocytes and cultured cells. We report here that a G protein binding peptide protects rats from LPS-induced mortality, suggesting a functional linkage between a GPI-anchored receptor and the intracellular signaling molecules with which it is physically associated. PMID:9835628

  20. Xuemaitong granules attenuate carotid atherosclerosis by decreasing the expression of CD14+CD16+ monocytes, IL-6, TNF-α, and hsCRP.

    PubMed

    Zhang, Q; Qian, G; Ding, Z

    2014-09-12

    Carotid atherosclerosis (CAS) has been extensively studied because its position can be easily observed. Our objective was to investigate the effects of Xuemaitong granules on the generation and activation of CD14+CD16+ monocytes on the inflammatory reaction in CAS patients. In this study, 22 male apolipoprotein E (apoE)-deficient mice were fed a high-fat diet for 13 weeks. After induction of an atherosclerotic plaque, the animals were randomly divided into the Xuemaitong granule group (450.5 mg/kg via intragastric administration, N=11) and the control group (equal volume saline via intragastric administration, N=11). Venous blood was obtained to analyze monocyte and CD14+CD16+ inflammatory monocyte levels, as well as interleukin (IL)-6, tumor necrosis factor (TNF)-a, and high-sensitivity C-reactive protein (hsCRP). For clinical studies, 100 CAS patients received oral administration of Xuemaitong granules for 6 months. Monocytes, CD14+CD16+ inflammatory monocytes, and the inflammatory cytokines IL-6, TNF-a, and hsCRP were analyzed. Compared with the control group, a remarkable decrease in the number of monocytes and CD14+CD16+ inflammatory monocytes as well as TNF-a, hsCRP, and IL-6 was noted in the Xuemaitong group. Compared with before treatment levels, the proportions of monocytes and their subsets of CD14+CD16+ inflammatory monocytes and the concentration of the inflammatory cytokines IL-6, TNF-a, and hsCRP significantly decreased. Xuemaitong granules played a significant role in the anti-inflammatory reactions. In addition, the granules attenuated the expression of the CD14+CD16+ inflammatory monocytes, resulting in the downregulation of the cytokines IL-6, TNF-a, and hsCRP.

  1. A novel CD14(high) CD16(high) subset of peritoneal macrophages from cirrhotic patients is associated to an increased response to LPS.

    PubMed

    Ruiz-Alcaraz, Antonio José; Tapia-Abellán, Ana; Fernández-Fernández, María Dolores; Tristán-Manzano, María; Hernández-Caselles, Trinidad; Sánchez-Velasco, Eduardo; Miras-López, Manuel; Martínez-Esparza, María; García-Peñarrubia, Pilar

    2016-04-01

    The aim of this study was to characterize monocyte-derived macrophages (M-DM) from blood and ascites of cirrhotic patients comparatively with those obtained from blood of healthy controls. The phenotypic profile based on CD14/CD16 expression was analyzed by flow cytometry. Cells were isolated and stimulated in vitro with LPS and heat killed Candida albicans. Phosphorylation of ERK, c-Jun, p38 MAPK, and PKB/Akt was analyzed by Western blotting. A novel CD14(high)CD16(high) M-DM subpopulation is present in ascites (∼33%). The CD14(++)CD16(+) intermediate subset is increased in the blood of cirrhotic patients (∼from 4% to 11%) and is predominant in ascites (49%), while the classical CD14(++)CD16(-) subpopulation is notably reduced in ascites (18%). Basal hyperactivation of ERK and JNK/c-Jun pathways observed in ascites M-DM correlates with CD14/CD16 high expressing subsets, while PI3K/PKB does it with the CD16 low expressing cells. In vitro LPS treatment highly increases ERK1/2, PKB/Akt and c-Jun phosphorylation, while that of p38 MAPK is decreased in M-DM from ascites compared to control blood M-DM. Stimulation of healthy blood M-DM with LPS and C. albicans induced higher phosphorylation levels of p38 than those from ascites. Regarding cytokines secretion, in vitro activated M-DM from ascites of cirrhotic patients produced significantly higher amounts of IL-6, IL-10 and TNF-α, and lower levels of IL-1β and IL-12 than control blood M-DM. In conclusion, a new subpopulation of CD14(high)CD16(high) peritoneal M-DM has been identified in ascites of cirrhotic patients, which is very sensitive to LPS stimulation.

  2. CD14(+)CD16(+) monocytes are the main target of Zika virus infection in peripheral blood mononuclear cells in a paediatric study in Nicaragua.

    PubMed

    Michlmayr, Daniela; Andrade, Paulina; Gonzalez, Karla; Balmaseda, Angel; Harris, Eva

    2017-10-02

    The recent Zika pandemic in the Americas is linked to congenital birth defects and Guillain-Barré syndrome. White blood cells (WBCs) play an important role in host immune responses early in arboviral infection. Infected WBCs can also function as 'Trojan horses' and carry viruses into immune-sheltered spaces, including the placenta, testes and brain. Therefore, defining which WBCs are permissive to Zika virus (ZIKV) is critical. Here, we analyse ZIKV infectivity of peripheral blood mononuclear cells (PBMCs) in vitro and from Nicaraguan Zika patients and show CD14(+)CD16(+) monocytes are the main target of infection, with ZIKV replication detected in some dendritic cells. The frequency of CD14(+) monocytes was significantly decreased, while the CD14(+)CD16(+) monocyte population was significantly expanded during ZIKV infection compared to uninfected controls. Viral RNA was detected in PBMCs from all patients, but in serum from only a subset, suggesting PBMCs may be a reservoir for ZIKV. In Zika patients, the frequency of infected cells was lower but the percentage of infected CD14(+)CD16(+) monocytes was significantly higher compared to dengue cases. The gene expression profile in monocytes isolated from ZIKV- and dengue virus-infected patients was comparable, except for significant differences in interferon-γ, CXCL12, XCL1, interleukin-6 and interleukin-10 levels. Thus, our study provides a detailed picture of the innate immune profile of ZIKV infection and highlights the important role of monocytes, and CD14(+)CD16(+) monocytes in particular.The main targets of Zika virus infection in human peripheral blood mononuclear cells are monocytes, particularly CD14(+)CD16(+) monocytes, which are expanded in Zika patients and in in vitro-infected samples.

  3. Role of CD14 and TLR4 in type I, type III collagen expression, synthesis and secretion in LPS-induced normal human skin fibroblasts

    PubMed Central

    Yang, Hongming; Li, Juncong; Wang, Yihe; Hu, Quan

    2015-01-01

    Objectives: The primary aim of this study was to investigate the role of CD14 and TLR4 in type I, type III collagen expression, synthesis and secretion in LPS-induced normal human skin fibroblasts. The secondary aim was to provide theoretical basis for the molecular mechanisms of scar formation induced by LPS. Methods: The normal skin fibroblasts cultured in vitro were randomly divided into four groups: 0.1 μg/mL LPS reference group, CD14 pretreatment + LPS, TLR4 pretreatment + LPS, CD14 and TLR4 pretreatment + LPS. The collagen DNA synthesis was assessed by 3H-proline incorporation method. Real-time Quantitative PCR was used to detect type I, type III collagen mRNA expression. Results: Similar results were revealed for mRNA expression levels. The immunofluorescence staining suggested that type I and type III collagen were expressed in all investigated groups and that the expression was differentially downregulated in groups B, C, D. ELISA demonstrated markedly decreased levels in secreting type I, type III collagens and hydroxyproline in groups B, C, D (P<0.05), and the lowest level was detected in group D (P<0.01). Conclusion: Pretreatment with CD14 or TLR4 alone or their combination can significantly reduce the levels of type I and type III collagen expression, synthesis and secretion, with the most notable reduction detected in case of CD14 and TLR4 combined. We could thus conclude that both CD14 and TLR4 are involved in type I and type III collagen expression, synthesis and secretion in LPS-induced skin fibroblasts. PMID:25932184

  4. HIV-1 Tat Protein Induces Production of Proinflammatory Cytokines by Human Dendritic Cells and Monocytes/Macrophages through Engagement of TLR4-MD2-CD14 Complex and Activation of NF-κB Pathway

    PubMed Central

    Leghmari, Kaoutar; Serrero, Manutea; Delobel, Pierre; Izopet, Jacques; BenMohamed, Lbachir; Bahraoui, Elmostafa

    2015-01-01

    We recently reported that the human immunodeficiency virus type-1 (HIV-1) Tat protein induced the expression of programmed death ligand-1 (PD-L1) on dendritic cells (DCs) through a TLR4 pathway. However, the underlying mechanisms by which HIV-1 Tat protein induces the abnormal hyper-activation of the immune system seen in HIV-1 infected patients remain to be fully elucidated. In the present study, we report that HIV-1 Tat protein induced the production of significant amounts of the pro-inflammatory IL-6 and IL-8 cytokines by DCs and monocytes from both healthy and HIV-1 infected patients. Such production was abrogated in the presence of anti-TLR4 blocking antibodies or soluble recombinant TLR4-MD2 as a decoy receptor, suggesting TLR4 was recruited by Tat protein. Tat-induced murine IL-6 and CXCL1/KC a functional homologue of human IL-8 was abolished in peritoneal macrophages derived from TLR4 KO but not from Wt mice, confirming the involvement of the TLR4 pathway. Furthermore, the recruitment of TLR4-MD2-CD14 complex by Tat protein was demonstrated by the activation of TLR4 downstream pathways including NF-κB and SOCS-1 and by down-modulation of cell surface TLR4 by endocytosis in dynamin and lipid-raft-dependent manners. Collectively, these findings demonstrate, for the first time, that HIV-1 Tat interacts with TLR4-MD2-CD14 complex and activates the NF-κB pathway, leading to overproduction of IL-6 and IL-8 pro-inflammatory cytokines by myeloid cells from both healthy and HIV-1 infected patients. This study reveals a novel mechanism by which HIV-1, via its early expressed Tat protein, hijacks the TLR4 pathway, hence establishing abnormal hyper-activation of the immune system. PMID:26090662

  5. Lower Plasma Soluble Transferrin Receptor Range in Healthy Indian Pediatric Cohort as Compared to Asian and Western Data.

    PubMed

    Bhatia, P; Siyaram, D; Deepshikha; Marathe, R; Dayal, D

    2017-09-01

    Soluble serum transferrin receptor is derived from erythroid transferrin receptor expressed on surface of developing erythroid cells. It can be detected in blood using sensitive ELISA methodology and blood levels reflect physiological iron dependent erythropoiesis state in bone marrow. Normal adult levels vary from 2 to 5 mg/l. However, pediatric studies are few and describe normal ranges to the tune of 1.0-3.0 mg/l, which are relatively lower than that of adults. In present study 40 healthy children (2-12 years) were evaluated to establish normal soluble transferrin receptor range. The mean transferrin receptor levels were 0.39 mg/l with a range of 0.17-2.1 mg/l. The levels were low as compared to mean levels described in other studies from West and our country (4.39 and 2.0 mg/l respectively). Since, no internationally standard method for reporting and testing for transferrin receptor levels are yet available, hence it is imperative to establish normal control ranges in different population cohorts, especially in pediatric age group, to better interpret their levels in diagnostic context.

  6. Bezafibrate, a peroxisome proliferator-activated receptor α agonist, decreases circulating CD14(+)CD16(+) monocytes in patients with type 2 diabetes.

    PubMed

    Terasawa, Tomoko; Aso, Yoshimasa; Omori, Kyoko; Fukushima, Maiko; Momobayashi, Atsushi; Inukai, Toshihiko

    2015-02-01

    CD14(+)CD16(+) monocytes are proinflammatory cells that produce tumor necrosis factor and interleukin (IL)-1β. The number of circulating CD14(+)CD16(+) monocytes is increased in patients with chronic renal failure or coronary artery disease. We investigated the effect of bezafibrate, a peroxisome proliferator-activated receptor α agonist, on circulating CD14(+)CD16(+) monocytes in patients with type 2 diabetes. Using cells isolated from type 2 diabetic subjects, we also examined the in vitro expression of CD16 messenger RNA (mRNA) by mononuclear cells (MNCs) exposed to bezafibrate. The percentage of CD14(+)CD16(+) monocytes among all CD14(+) monocytes was significantly higher in subjects with impaired glucose tolerance (P < 0.01) or type 2 diabetes (P < 0.05) than in those with normal glucose tolerance. The percentage of CD14(+)CD16(+) monocytes was significantly lower in patients with type 2 diabetes who were taking bezafibrate (400 mg/d) than in patients not taking it (P < 0.01). Treatment with bezafibrate for 12 weeks significantly reduced the percentage of circulating CD14(+)CD16(+) monocytes from 45.4 ± 25.2% to 38.3 ± 21.8% (P = 0.0144). In an in vitro study, the expression of CD16 mRNA by MNCs from 6 diabetic subjects was decreased after 24 hours of treatment with 10 μg/mL of bezafibrate (P < 0.05). Expression of IL-1β mRNA by MNCs was also decreased after 24 hours of treatment with 10 μg/mL of bezafibrate, whereas the IL-1β level in the culture supernatant was significantly decreased after treatment of MNCs with either 1 or 10 μg/mL of bezafibrate. In conclusion, bezafibrate decreased circulating CD14(+)CD16(+) monocytes in patients with type 2 diabetes, probably by inhibiting the expression of CD16 mRNA.

  7. Matched molecular pairs as a guide in the optimization of pharmaceutical properties; a study of aqueous solubility, plasma protein binding and oral exposure.

    PubMed

    Leach, Andrew G; Jones, Huw D; Cosgrove, David A; Kenny, Peter W; Ruston, Linette; MacFaul, Philip; Wood, J Matthew; Colclough, Nicola; Law, Brian

    2006-11-16

    By identifying every pair of molecules that differ only by a particular, well-defined, structural transformation in a database of measured properties and computing the corresponding change in property, we obtain an overview of the effect that structural change has upon the property and set an expectation for what will happen when that transformation is applied elsewhere. The mean change indicates the expected magnitude of the change in the property and the number of cases in which the property increases give the probability that the structural transformation will cause the property to increase. Outliers indicate potential ways of avoiding the general trend. Comparing to changes in lipophilicity highlights structural transformations that have unusual effects, some of which can be explained by conformational changes. In this paper, we focus upon the effects on aqueous solubility, plasma protein binding and oral exposure of adding substituents to aromatic rings and methylating heteroatoms.

  8. Fat-soluble vitamins and plasma and erythrocyte membrane fatty acids in chylothorax pediatric patients receiving a medium-chain triglyceride-rich diet.

    PubMed

    Densupsoontorn, Narumon; Jirapinyo, Pipop; Tirapongporn, Hathaichanok; Wongarn, Renu; Chotipanang, Kwanjai; Phuangphan, Phakkanan; Chongviriyaphan, Nalinee

    2014-11-01

    Post-operative chylothorax can be cured by a medium-chain triglyceride (MCT)-rich diet. However, there is concern that an MCT-rich diet results in clinical and biochemical deficiencies in fat-soluble vitamins and fatty acids. We compared fat-soluble vitamins status and fatty acids status before and after administration of an MCT-rich diet. Nine children with congenital heart disease developed chylothorax after cardiac surgery. Blood samples were drawn from each subject twice, first prior to administration of an MCT-rich diet and secondly when the chylothorax was clinically cured and the MCT diet discontinued. Both blood samples were analyzed for retinol and 25-hydroxy vitamin D concentrations, the ratio of α-tocopherol to total lipids (α-TE/TL), coagulogram, and the fatty acid composition in plasma and erythrocyte membrane phospholipids. In spite of a decrease in the α-TE/TL ratio (3.78 ± 0.89 vs 2.36 ± 0.44 mg/g, p<0.05), this decrease did not reach the deficiency cut-off level. Linoleic acid in both plasma and erythrocyte membrane lipids decreased significantly (25.25 ± 8.06 vs 14.25 ± 2.88%, and 11.19 ± 2.15 vs 6.89 ± 2.45%, respectively). Administration of an MCT-rich diet for treatment of postoperative chylothorax caused a reduction in vitamin E status and linoleic acid, but without any symptoms of deficiency.

  9. Combination of plasma-soluble fms-like tyrosine kinase 1 and uterine artery Doppler for the prediction of preeclampsia in cases of elderly gravida.

    PubMed

    Kulmala, Lalita; Phupong, Vorapong

    2014-06-01

    The aim of this study was to determine the predictive value of the combination of plasma-soluble fms-like tyrosine kinase 1 (sFlt-1) and uterine artery Doppler for the detection of preeclampsia in women of advanced age at 16-18 weeks of gestation and to identify associations between other pregnancy complications and abnormalities of these combined tests. The maternal plasma sFlt-1 level was measured, and uterine artery Doppler was performed at 16-18 weeks of gestation in 314 cases of elderly gravida. The main outcome was preeclampsia. Fourteen women (4.46%) developed preeclampsia. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of uterine artery Doppler combined with plasma sFlt-1 for preeclampsia prediction were 28.6, 95.7, 23.5 and 96.6%, respectively. For the prediction of early-onset preeclampsia, the sensitivity, specificity, PPV and NPV were 80, 95.8, 23.5 and 99.7%, respectively. Patients with abnormal uterine artery Doppler findings and an abnormal plasma s Flt-1 level (greater than 1724.5 pg ml(-1)) had a higher risk of preterm delivery (relative risk (RR)=3.38, 95% confidence interval (CI) 1.47-7.59), neonatal respiratory distress syndrome (RR=52.06, 95% CI 5.71-474.45) and perinatal death (RR=17.35, 95% CI 1.13-265.64). Our findings indicate that the combination of uterine artery Doppler and sFlt-1 level at 16-18 weeks of gestation in cases of elderly gravida has a high predictive value for early-onset preeclampsia, but not for overall preeclampsia. This combination test may be a useful early second trimester screening test for the prediction of early-onset preeclampsia in cases of elderly gravida.

  10. Comparative effects of pioglitazone and rosiglitazone on plasma levels of soluble receptor for advanced glycation end products in type 2 diabetes mellitus patients.

    PubMed

    Oz Gul, Ozen; Tuncel, Ercan; Yilmaz, Yusuf; Ulukaya, Engin; Gul, Cuma Bulent; Kiyici, Sinem; Oral, Arzu Yilmaztepe; Guclu, Metin; Ersoy, Canan; Imamoglu, Sazi

    2010-01-01

    Low levels of soluble receptor for advanced glycation end products (sRAGE) have been associated with the occurrence of vascular complications in patients with type 2 diabetes mellitus. Preliminary evidence has suggested that thiazolidinediones have the ability to modulate circulating levels of this molecule in the hyperglycemic milieu. The aim of this pilot study was to assess the differential effect of 2 different thiazolidinediones-pioglitazone and rosiglitazone-on plasma levels of sRAGE in type 2 diabetes mellitus patients. Sixty type 2 diabetes mellitus subjects were randomly assigned to receive pioglitazone (30 mg/d, n = 19), rosiglitazone (4 mg/d, n = 20), or placebo (medical nutrition therapy, n = 21) for 12 weeks. Changes in plasma glucose, glycosylated hemoglobin, insulin resistance (homeostasis model assessment), total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, triglycerides, and sRAGE were evaluated at baseline and after 12 weeks. At 12 weeks, the pioglitazone (P < .001) group had a significant increase from baseline in sRAGE values that was not seen in the medical nutrition therapy and rosiglitazone groups. We conclude that, in type 2 diabetes mellitus patients, pioglitazone-but not rosiglitazone-significantly raised sRAGE, which may contribute to its antiatherogenic effects.

  11. Clinical Value of Plasma Soluble Urokinase-Type Plasminogen Activator Receptor Levels in Term Neonates with Infection or Sepsis: A Prospective Study

    PubMed Central

    Siahanidou, Tania; Margeli, Alexandra; Charoni, Stavroula; Giannaki, Maria; Vavourakis, Eustathios; Charisiadou, Athina; Papassotiriou, Ioannis

    2014-01-01

    Background. suPAR, the soluble form of the urokinase-type plasminogen activator receptor, has been identified as a biomarker of infection in adults but its properties in neonatal infection are not known. Methods. Plasma suPAR levels were determined by ELISA in 47 term neonates with infection (19 bacterial and 28 viral) and in 18 healthy neonates as controls. Thirteen out of 47 infected neonates were septic. In all infected neonates, suPAR levels were repeated at 24 hours, 48 hours, 3–5 days, and 7–10 days following admission. Results. Plasma suPAR levels were significantly increased in infected neonates upon admission, whereas they were highest in septic neonates, in comparison with controls (P < 0.001) and correlated positively with serum CRP levels (P = 0.001). At infection subsidence, suPAR concentrations decreased significantly in comparison with baseline (P < 0.001) but remained higher than in controls (P = 0.01). Receiver operating characteristic analysis resulted in significant areas under the curve for detecting either infected or septic neonates, but not for discriminating between bacterial and viral cause of infection. Conclusions. suPAR is a diagnostic biomarker of infection or sepsis in term neonates; however, it cannot discriminate bacterial from viral infections and also its utility for monitoring the response to treatment is questioned. PMID:24882949

  12. Infiltrating CD16+ Are Associated with a Reduction in Peripheral CD14+CD16++ Monocytes and Severe Forms of Lupus Nephritis

    PubMed Central

    Barrera García, Anabel; Arias, Luis F.; Burbano, Catalina; Restrepo, Mauricio; Vanegas, Adriana L.; Muñoz, Carlos H.; Rojas, Mauricio; González, Luis A.

    2016-01-01

    Our aim was to characterize glomerular monocytes (Mo) infiltration and to correlate them with peripheral circulating Mo subsets and severity of lupus nephritis (LN). Methods. We evaluated 48 LN biopsy samples from a referral hospital. Recognition of Mo cells was done using microscopic view and immunohistochemistry stain with CD14 and CD16. Based on the number of cells, we classified LN samples as low degree of diffuse infiltration (<5 cells) and high degree of diffuse infiltration (≥5 cells). Immunophenotyping of peripheral Mo subsets was done using flow cytometry. Results. Mean age was 34.0 ± 11.7 years and the mean SLEDAI was 17.5 ± 6.9. The most common SLE manifestations were proteinuria (91%) and hypocomplementemia (75%). Severe LN was found in 70% of patients (Class III, 27%; Class IV, 43%). Severe LN patients and patients with higher grade of CD16+ infiltration had lower levels of nonclassical (CD14+CD16++) Mo in peripheral blood. Conclusions. Our results might suggest that those patients with more severe forms of LN had a higher grade of CD14+CD16+ infiltration and lower peripheral levels of nonclassical (CD14+CD16++) Mo and might reflect a recruitment process in renal tissues. However, given the small sample, our results must be interpreted carefully. PMID:28070418

  13. CD14 in the TLRs signaling pathway is associated with the resistance to E. coli F18 in Chinese domestic weaned piglets

    PubMed Central

    Wu, Zhengchang; Liu, Ying; Dong, Wenhua; Zhu, Guo-qiang; Wu, Shenglong; Bao, Wenbin

    2016-01-01

    Escherichia coli F18 (E. coli F18) is mainly responsible for post-weaning diarrhea (PWD) in piglets. The genetic basis and regulatory mechanism of E. coli F18 resistance in Chinese domestic weaned piglets remain unclear. Meishan piglets were used as model animals to test their susceptibility to E. coli F18. By performing a comparative transcriptome study on duodenum tissues of sensitive and resistant pigs, we identified 198 differentially expressed genes (DEGs; 125 upregulated and 73 downregulated) in the resistant pigs. DEGs were predominately involved in immune system pathways, including the Toll-like receptor (TLR) signaling pathway. qPCR and western blot showed CD14, IFN-α, TLR4 and IL-1β, etc. in the TLR signaling pathway had significantly higher expression levels in lipopolysaccharide (LPS)-induced small intestinal epithelial cell lines (IPEC-J2) than those in normal IPEC-J2 cells. Immunohistochemical analysis showed the increased expression of CD14 gene in the E. coli F18-resistant individuals. After CD14 knockdown, the levels of cytokines IL-6 and IL-12 were significantly reduced in IPEC-J2 cell supernatants. The adhesion ability of F18ab strain with IPEC-J2 cells was significantly increased (p < 0.01). This study revealed the TLR signaling pathway, and especially CD14, probably plays an important role in resistance to E. coli F18 infection in Chinese domestic piglets. PMID:27098998

  14. Modulation of interferon-gamma-induced major histocompatibility (MHC) and CD14 antigen changes by lipophilic muramyltripeptide MTP-PE in human monocytes.

    PubMed

    Landmann, R; Wesp, M; Dukor, P

    1988-11-01

    The lipophilic muramylpeptide derivative muramyltripeptide-phosphatidylethanolamine (MTP-PE, 0.05 to 5 micrograms/ml) and human recombinant interferon-gamma (rIFN-gamma, 1 to 100 U/ml) were applied singly or in combination to fresh human mononuclear blood leucocytes in vitro. After 15 to 72 hr incubation, culture- and drug-induced changes in beta 2-microglobulin (MHC class I associated), HLA-DR (MHC class II), and Leu-M3 (CD14) antigen expression were investigated by flow cytometry; changes in monocyte morphology (forward light scatter and side scatter) were assessed by scatter analysis. It was found that (1) rIFN-gamma caused a simultaneous down-regulation of the CD14 antigen and an up-regulation of MHC class I and class II molecules on the surface of cultured monocytes; (2) MTP-PE, which by itself failed to influence the expression of these antigens, synergized with rIFN-gamma in increasing MHC antigens and reducing CD14; (3) at high concentrations rIFN-gamma reduced monocyte viability to a small but significant extent and this effect was further potentiated by MTP-PE; and (4) untreated monocytes in culture showed an apparently MTP-PE-insensitive increase in size, density, and beta 2-microglobulin, HLA-DR, and CD14 antigen expression. The influence of MTP-PE on rIFN-gamma-induced surface marker changes may contribute to its immunoadjuvant activity in vivo.

  15. Infiltrating CD16(+) Are Associated with a Reduction in Peripheral CD14(+)CD16(++) Monocytes and Severe Forms of Lupus Nephritis.

    PubMed

    Barrera García, Anabel; Gómez-Puerta, José A; Arias, Luis F; Burbano, Catalina; Restrepo, Mauricio; Vanegas, Adriana L; Muñoz, Carlos H; Rojas, Mauricio; González, Luis A; Vásquez, Gloria

    2016-01-01

    Our aim was to characterize glomerular monocytes (Mo) infiltration and to correlate them with peripheral circulating Mo subsets and severity of lupus nephritis (LN). Methods. We evaluated 48 LN biopsy samples from a referral hospital. Recognition of Mo cells was done using microscopic view and immunohistochemistry stain with CD14 and CD16. Based on the number of cells, we classified LN samples as low degree of diffuse infiltration (<5 cells) and high degree of diffuse infiltration (≥5 cells). Immunophenotyping of peripheral Mo subsets was done using flow cytometry. Results. Mean age was 34.0 ± 11.7 years and the mean SLEDAI was 17.5 ± 6.9. The most common SLE manifestations were proteinuria (91%) and hypocomplementemia (75%). Severe LN was found in 70% of patients (Class III, 27%; Class IV, 43%). Severe LN patients and patients with higher grade of CD16(+) infiltration had lower levels of nonclassical (CD14(+)CD16(++)) Mo in peripheral blood. Conclusions. Our results might suggest that those patients with more severe forms of LN had a higher grade of CD14(+)CD16(+) infiltration and lower peripheral levels of nonclassical (CD14(+)CD16(++)) Mo and might reflect a recruitment process in renal tissues. However, given the small sample, our results must be interpreted carefully.

  16. Aberrant overexpression of CD14 on granulocytes sensitizes the innate immune response in mDia1 heterozygous del(5q) MDS.

    PubMed

    Keerthivasan, Ganesan; Mei, Yang; Zhao, Baobing; Zhang, Ling; Harris, Chad E; Gao, Juehua; Basiorka, Ashley A; Schipma, Matthew J; McElherne, James; Yang, Jing; Verma, Amit K; Pellagatti, Andrea; Boultwood, Jacqueline; List, Alan F; Williams, David A; Ji, Peng

    2014-07-31

    The myelodysplastic syndromes (MDSs) include a spectrum of stem cell malignancies characterized by an increased risk of developing acute myeloid leukemia. Heterozygous loss of chromosome 5q (del[5q]) is the most common cytogenetic abnormality in MDS. DIAPH1 is localized to 5q31 and encodes one of the formin proteins, mDia1, which is involved in linear actin polymerization. Mice with mDia1 deficiency develop hematologic features with age mimicking human myeloid neoplasm, but its role in the pathogenesis of MDS is unclear. Here we report that mDia1 heterozygous and knockout mice develop MDS phenotypes with age. In these mice, CD14 was aberrantly overexpressed on granulocytes in a cell-autonomous manner, leading to a hypersensitive innate immune response to lipopolysaccharide (LPS) stimuli through CD14/Toll-like receptor 4 signaling. Chronic stimulation with LPS accelerated the development of MDS in mDia1 heterozygous and knockout mice that can be rescued by lenalidomide. Similar findings of CD14 overexpression were observed on the bone marrow granulocytes of del(5q) MDS patients. Mechanistically, mDia1 deficiency led to a downregulation of membrane-associated genes and a specific upregulation of CD14 messenger RNA in granulocytes, but not in other lineages. These results underscore the significance of mDia1 heterozygosity in deregulated innate immune responses in del(5q) MDS.

  17. Preparations of intravenous immunoglobulins diminish the number and proinflammatory response of CD14+CD16++ monocytes in common variable immunodeficiency (CVID) patients.

    PubMed

    Siedlar, Maciej; Strach, Magdalena; Bukowska-Strakova, Karolina; Lenart, Marzena; Szaflarska, Anna; Węglarczyk, Kazimierz; Rutkowska, Magdalena; Baj-Krzyworzeka, Monika; Pituch-Noworolska, Anna; Kowalczyk, Danuta; Grodzicki, Tomasz; Ziegler-Heitbrock, Loems; Zembala, Marek

    2011-05-01

    We have studied the effect of intravenous immunoglobulins (IVIG) on monocyte subpopulations and cytokine production in patients with CVID. The absolute number of CD14(+)CD16(++) monocytes decreased on average 2.5-fold 4h after IVIG and after 20h returned to the baseline. The cytokine level in the supernatants of peripheral blood mononuclear cells (PBMC) after ex vivo LPS stimulation demonstrated the >2-fold decrease in TNF production 4h after IVIG. The TNF expression, which is higher in the CD14(+)CD16(++) monocytes, was decreased in these cells by IVIG in 4/7 CVID cases. In vitro exposure of the healthy individuals' monocytes to the IVIG preparation resulted in reduced TNF production, which was overcome by blockade of the FcγRIIB in the CD14(+)CD16(++) CD32B(high) monocytes. Our data suggest that reduction in the number of CD14(+)CD16(++) monocytes and the blockade of their cytokine production via triggering CD32B can contribute to the anti-inflammatory action of IVIG. Copyright © 2011 Elsevier Inc. All rights reserved.

  18. Genetic polymorphism in CD14 gene, a co-receptor of TLR4 associated with non-alcoholic fatty liver disease

    PubMed Central

    Kapil, Shweta; Duseja, Ajay; Sharma, Bal Krishan; Singla, Bhupesh; Chakraborti, Anuradha; Das, Ashim; Ray, Pallab; Dhiman, Radha K; Chawla, Yogesh

    2016-01-01

    AIM To evaluate the pathogenic role of toll-like receptor (TLR) gene polymorphisms in patients with non-alcoholic fatty liver disease (NAFLD). METHODS Two hundred and fifty subjects (NAFLD = 200, healthy volunteers = 50) underwent polymerase chain reaction and restriction fragment length polymorphism to assess one polymorphism in the toll-like receptor 2 (TLR2) gene (A753G), two polymorphisms in the TLR4 gene (TLR4 Asp299Gly and Thr399Ile allele), and two polymorphisms in the cluster of differentiation 14 (CD14) (C-159T and C-550T) gene, a co-receptor of TLR4. Association of TLR gene polymorphisms with NAFLD and its severity was evaluated by genetic models of association. RESULTS On both multiplicative and recessive models of gene polymorphism association, there was significant association of CD14 C (-159) T polymorphism with NAFLD; patients with TT genotype had a 2.6 fold increased risk of developing NAFLD in comparison to CC genotype. There was no association of TLR2 Arg753Gln, TLR4 Asp299Gly, Thr399Ile, and CD14 C (-550) T polymorphisms with NAFLD. None of the TLR gene polymorphisms had an association with histological severity of NAFLD. CONCLUSION Patients with CD14 C (-159) T gene polymorphism, a co-receptor of TLR4, have an increased risk of NAFLD development. PMID:27895422

  19. Characterization and use of new monoclonal antibodies to CD11c, CD14, and CD163 to analyze the phenotypic complexity of ruminant monocyte subsets.

    PubMed

    Elnaggar, Mahmoud M; Abdellrazeq, Gaber S; Mack, Victoria; Fry, Lindsay M; Davis, William C; Park, Kun Taek

    2016-10-01

    The sequencing of the bovine genome and development of mass spectrometry, in conjunction with flow cytometry (FC), have afforded an opportunity to complete the characterization of the specificity of monoclonal antibodies (mAbs), only partially characterized during previous international workshops focused on antibody development for livestock (1991, Leukocyte Antigens in Cattle, Sheep, and Goats; 1993, Leukocyte Antigens of Cattle and Sheep; 1996, Third Workshop on Ruminant Leukocyte Antigens). The objective of this study was to complete the characterization of twelve mAbs incompletely characterized during the workshops that reacted with molecules predominantly expressed on bovine monocytes and use them to provide further information on the phenotypic complexity of monocyte subsets in ruminants. Analysis revealed that the mAbs could be grouped into three clusters that recognize three different molecules: CD11c, CD14, and CD163. Following characterization, comparison of the patterns of expression of CD14 and CD163 with expression of CD16, CD172a, and CD209 revealed the mononuclear cell population is comprised of multiple subsets with differential expression of these molecules. Further analysis revealed the epitopes recognized by mAbs to CD14 and CD163 are conserved on orthologues in sheep and goats. In contrast to CD14 that is also expressed on sheep and goat granulocytes, CD163 is a definitive marker for their monocytes. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. T cell immunoglobulin and mucin domain-containing molecule 3 on CD14(+) monocytes serves as a novel biological marker for diabetes duration in type 2 diabetes mellitus.

    PubMed

    Yan, Wen-Jiang; Sun, Peng; Wei, Dan-Dan; Wang, Shuang-Xi; Yang, Jing-Jing; Li, Yi-Hui; Zhang, Cheng

    2016-11-01

    Type 2 diabetes is a worldwide disease that is associated with increased rates of obesity and reduced physical activity. Obesity-associated insulin resistance in type 2 diabetes is a disorder in the balance between pro-inflammatory and anti-inflammatory signals. T cell immunoglobulin and mucin domain-containing molecule 3 (Tim-3) has been reported as an important regulatory inflammation molecule, and plays a pivotal role in several inflammation-related diseases. Peripheral blood mononuclear cells were obtained from type 2 diabetes patients (n = 31) and healthy donors (n = 18), and Tim-3 expression on peripheral blood mononuclear cells was evaluated by flow cytometry. We showed the downregulated expression of Tim-3 on CD14(+) monocytes from type 2 diabetes patients. In addition, the upregulated expression of Tim-3 on peripheral CD4(+) T cells and CD8(+) T cells was observed in the present study. The correlation analysis between Tim-3 expression on CD14(+) monocytes and diabetes duration showed the longer diabetes duration time, the lower Tim-3 expression on CD14 monocytes. The present results suggest that Tim-3 might participate in the progression of type 2 diabetes by its negative regulation on these immune cells, and Tim-3 on CD14(+) monocytes serves as a novel biological marker for diabetes duration in type 2 diabetes patients. © 2016 The Authors. Journal of Diabetes Investigation published by Asian Association for the Study of Diabetes (AASD) and John Wiley & Sons Australia, Ltd.

  1. Bromelain inhibits lipopolysaccharide-induced cytokine production in human THP-1 monocytes via the removal of CD14.

    PubMed

    Huang, Jing-Rong; Wu, Chia-Chuan; Hou, Rolis Chien-Wei; Jeng, Kee-Ching

    2008-01-01

    Bromelain has been reported to have anti-inflammatory and immunomodulatory effects. However, the anti-inflammatory mechanism of bromelain is unclear. Therefore, we investigated the effect of bromelain on cytokine production from lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells (PBMC) and monocytic leukemia THP-1 cells. The result showed that bromelain (50-100 microg/ml) significantly and reversibly reduced tumor necrosis factor (TNF)-alpha interleukin- (IL)-1beta and IL-6 from LPS-induced PBMC and THP-1 cells. This effect was correlated with reduced LPS-induced TNF-alpha mRNA and NF-kappaB activity in THP-1 cells. In addition, bromelain dose-dependently inhibited LPS-induced prostaglandin E(2), thromboxane B(2) and COX-2 mRNA but not COX-1 mRNA. Importantly, bromelain degraded TNF-alpha and IL-1beta molecules, reduced the expression of surface marker CD14 but not Toll-like receptor 4 from THP-1 cells. Taken together, the results suggest that the suppression of signaling pathways by bromelain's proteolytic activity may contribute to the anti-inflammatory activity of bromelain.

  2. Molecular Basis of the Functional differences between soluble human vs. murine MD-2: Role of Val-135 in transfer of LPS from CD14 to MD-21

    PubMed Central

    Vašl, Jožica; Oblak, Alja; Peternelj, Tina T.; Klett, Javier; Martín-Santamaría, Sonsoles; Gioannini, Theresa L.; Weiss, Jerrold P.; Jerala, Roman

    2017-01-01

    Myeloid differentiation factor (MD-2) is an extracellular protein, associated with the ectodomain of Toll-like receptor 4 (TLR4), that plays a critical role in the recognition of bacterial lipopolysaccharide (LPS). Despite high overall structural and functional similarity, human (h) and murine (m) MD-2 exhibit several species-related differences. hMD-2 is capable of binding LPS in the absence of TLR4, whereas mMD-2 supports LPS responsiveness only when mMD-2 and mTLR4 are coexpressed in the same cell. Previously, charged residues at the edge of LPS binding pocket have been attributed to this difference. In this study, site-directed mutagenesis was used to explore the hydrophobic residues within MD-2 binding pocket, as the source of functional differences between hMD-2 and mMD-2. While decreased hydrophobicity of residues 61 and 63 in hMD-2 binding pocket retained the characteristics of wild type hMD-2, a relatively minor change of valine to alanine at position 135 completely abolished the binding of LPS to the hMD-2 mutant. The mutant, however, retained the LPS binding in complex with TLR4 and also cell activation, resulting in a murine-like phenotype. These results were supported by the molecular dynamics simulation. We propose that the residue at position 135 of MD-2 governs the dynamics of the binding pocket and its ability to accommodate lipid A, which is allosterically affected by bound TLR4. PMID:26826249

  3. Molecular Basis of the Functional Differences between Soluble Human Versus Murine MD-2: Role of Val135 in Transfer of Lipopolysaccharide from CD14 to MD-2.

    PubMed

    Vašl, Jožica; Oblak, Alja; Peternelj, Tina T; Klett, Javier; Martín-Santamaría, Sonsoles; Gioannini, Theresa L; Weiss, Jerrold P; Jerala, Roman

    2016-03-01

    Myeloid differentiation factor 2 (MD-2) is an extracellular protein, associated with the ectodomain of TLR4, that plays a critical role in the recognition of bacterial LPS. Despite high overall structural and functional similarity, human (h) and murine (m) MD-2 exhibit several species-related differences. hMD-2 is capable of binding LPS in the absence of TLR4, whereas mMD-2 supports LPS responsiveness only when mMD-2 and mTLR4 are coexpressed in the same cell. Previously, charged residues at the edge of the LPS binding pocket have been attributed to this difference. In this study, site-directed mutagenesis was used to explore the hydrophobic residues within the MD-2 binding pocket as the source of functional differences between hMD-2 and mMD-2. Whereas decreased hydrophobicity of residues 61 and 63 in the hMD-2 binding pocket retained the characteristics of wild-type hMD-2, a relatively minor change of valine to alanine at position 135 completely abolished the binding of LPS to the hMD-2 mutant. The mutant, however, retained the LPS binding in complex with TLR4 and also cell activation, resulting in a murine-like phenotype. These results were supported by the molecular dynamics simulation. We propose that the residue at position 135 of MD-2 governs the dynamics of the binding pocket and its ability to accommodate lipid A, which is allosterically affected by bound TLR4. Copyright © 2016 by The American Association of Immunologists, Inc.

  4. STEC:O111-HUS complicated by acute encephalopathy in a young girl was successfully treated with a set of hemodiafiltration, steroid pulse, and soluble thrombomodulin under plasma exchange.

    PubMed

    Yada, Noritaka; Fujioka, Masayuki; Bennett, Charles L; Inoki, Kazuya; Miki, Toyokazu; Watanabe, Akihiko; Yoshida, Toshiko; Hayakawa, Masaki; Matsumoto, Masanori; Fujimura, Yoshihiro

    2015-04-01

    We report a 14-year-old girl, who developed shigatoxin-producing E. coli (STEC)-HUS complicated by encephalopathy. She was successfully treated with hemodiafiltration, high-dose methylprednisolone pulse therapy, and soluble recombinant thrombomodulin under plasma exchange. von Willebrand factor multimers analysis provides potential insights into how the administered therapies might facilitate successful treatment of STEC-HUS.

  5. Chromium in exhaled breath condensate (EBC), erythrocytes, plasma and urine in the biomonitoring of chrome-plating workers exposed to soluble Cr(VI).

    PubMed

    Goldoni, Matteo; Caglieri, Andrea; De Palma, Giuseppe; Acampa, Olga; Gergelova, Petra; Corradi, Massimo; Apostoli, Pietro; Mutti, Antonio

    2010-02-01

    Chromium (Cr) levels measured in exhaled breath condensate (EBC-Cr) and urine (Cr-U) at the beginning and end of working shifts were related to those measured in erythrocytes (Cr-RBC) and plasma in 14 non-smoking male chrome-plating workers exposed to Cr(VI) in soluble aerosol form who did not report any significant current or past respiratory disease. Cr-U mainly correlated with Cr-P (Cr in plasma) at the end of the working shift (r(2) = 0.59, p < 0.01), whereas Cr-RBC correlated with EBC-Cr (r(2) = 0.32, p < 0.05); at the beginning of the shift, the only significant correlation was between Cr-U and Cr-RBC (r(2) = 0.74, p < 0.01). The clearance of Cr(iii) arising from Cr(VI) reduction was rapid, thus making Cr-U and Cr-P ideal biomarkers of the most recent exposure, whereas Cr-RBC may represent the fraction of Cr(VI) that reaches the bloodstream in non-reduced form and therefore depends on the airway inhaled dose represented by EBC-Cr. Cr-RBC clearance is slower and not only involves the free diffusion of Cr(iii) from RBC to plasma, but probably also involves more complicated kinetic phenomena involving other tissues and organs, which may explain the correlation between Cr-RBC and Cr-U and the lack of correlation Cr-RBC and Cr-P at least 36 h after the last exposure. In conclusion, our findings reinforce the idea that measuring Cr in EBC can significantly contribute to traditional biomonitoring by providing specific information at the target organ level and integrating our knowledge of Cr toxicokinetics.

  6. Protein/ionic liquid/glassy carbon sensors following analyte focusing by ionic liquid micelle collapse for simultaneous determination of water soluble vitamins in plasma matrices.

    PubMed

    Abd El-Hady, D; Albishri, H M

    2015-07-01

    Two novel sensors based on human serum albumin (HSA)-ionic liquid (IL) and bovine serum albumin (BSA)-ionic liquid (IL) composites modified glassy carbon electrode (GCE) were produced for simultaneous determination of water soluble vitamins B2, B6 and C in human plasma following analytes focusing by IL micelles collapse (AFILMC). For selective and efficient extraction, vitamins were dissolved in 3.0molL(-1) micellar solution of 1-octyl-3-methyl imidazolium bromide IL. The extracted vitamins were hydrodynamically injected by 25mbar for 20s into a running buffer of 12.5mmolL(-1) phosphate at pH 6.0 followed by electrochemical detection (ECD) on protein/1-octyl-3-methyl imidazolium hexafluorophosphate IL/GC sensors. The chemical stability of proposed sensors was achieved up to 7 days without any decomposition of PF6-based IL/protein and adsorption of interfering ions. In the current work, the sensitivity enhancement factor (SEF) up to 5000-fold was achieved using the AFILMC/ECD setup compared to conventional CE/UV. Under optimal conditions, linear calibration graphs were obtained from 0.5, 0.5 and 1.0 to 1500.0µgmL(-1) of vitamins B2, B6 and C, respectively. Detection limits of analytes were ranged from 180.0 to 520.0ngmL(-1). The proposed AFILMC/ECD setup was successfully applied to the assay of trace level quantification of vitamins in human plasma samples and also their binding constants with HSA and BSA were determined. The concurrent use of IL micelles for the proposed separation and detection processes exhibited some advantages, such as, a reduction of use toxic solvents, an efficient extraction and a direct injection of samples with a short-single run. Furthermore, IL micelles, having variable possibility of interactions, facilitated the successful achievements of AFILMC/ECD setup for the quantification of vitamins in plasma matrices.

  7. Plasma levels of soluble intercellular adhesion molecule-1 as a biomarker for disease severity of patients with community-acquired pneumonia.

    PubMed

    Chang, Pin-Yu; Tsao, Shih-Ming; Chang, Jer-Hwa; Chien, Ming-Hsien; Hung, Wen-Yueh; Huang, Yi-Wen; Yang, Shun-Fa

    2016-12-01

    Community-acquired pneumonia (CAP) is characterized as an acute inflammation of the lung associated with the activation of macrophages and neutrophils. Intercellular adhesion molecule-1 (ICAM-1) is an essential adhesion molecule involved in immune cell recruitment in lung inflammation. We investigated whether ICAM-1 is a useful biomarker for assessing the disease severity of hospitalized adult patients with CAP. Plasma soluble ICAM-1 (sICAM-1) levels were measured in 78 patients with CAP and 69 healthy controls by using a commercial enzyme-linked immunosorbent assay. The pneumonia severity index scores were used to determine CAP severity in patients upon initial hospitalization. The sICAM-1 and C-reactive protein (CRP) levels decreased significantly in patients with CAP after antibiotic treatment. The plasma concentration of sICAM-1 alone, but not CRP, was correlated with CAP severity according to the pneumonia severity index scores (r=0.431, p<0.001). The sICAM-1 levels in patients with CAP with high mortality risk were significantly higher than those in patients with CAP with medium or low mortality risk. Moreover, the sICAM-1 level showed a significant correlation with the length of hospital stay (r=0.488, p<0.001). Mechanistic investigations found that bacterial lipopolysaccharide induced upregulation of ICAM-1 expression through the c-Jun N-terminal kinase pathway in RAW264.7 macrophages. Plasma sICAM-1 levels may play a role in the diagnosis and clinical assessment of CAP severity. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Association of CD14 −159 (−260C/T) polymorphism and asthma risk: an updated genetic meta-analysis study

    PubMed Central

    Wang, Duan; Yang, Yang; Xu, Jin; Zhou, Zong-Ke; Yu, Hai-Yang

    2016-01-01

    Abstract Background: It has been reported that the cluster of differentiation 14 (CD14) gene −159C/T variant may be associated with asthma risk. However, some studies yielded conflicting results. Therefore, a comprehensive meta-analysis was designed to assess the precise association. Methods: A systematic search in PubMed, Embase (Ovid), China National Knowledge Internet (CNKI), and Wan fang databases was conducted up to August 15, 2015. Odds ratio (OR) and 95% confidence interval (CI) were used to pool the effect size. We used I2 to assess heterogeneity, and a funnel plot and Egger test to assess publication bias. Results: In total, 34 studies involving 15,641 subjects were included in this meta-analysis. There was a statistically significant association between CD14 −159C/T polymorphism and asthma risk observed in dominant model (TT+TC vs CC: OR = 0.86, 95% CI = 0.77–0.97, P = 0.012) and codominant model (TC vs CC: OR = 0.88, 95% CI = 0.78–0.99, P = 0.035) in adults. However, there may be no significant association between CD14 159C/T and atopic and nonatopic asthma risk. Conclusion: In summary, the overall results suggested that the CD14 −159C/T variant may decrease the risk of asthma susceptibility in adults. However, no significant association between CD14 159C/T and atopic and nonatopic asthma susceptibility was identified. More studies with larger sample size are needed to validate the findings from this study. PMID:27684840

  9. Micrococcus luteus Teichuronic Acids Activate Human and Murine Monocytic Cells in a CD14- and Toll-Like Receptor 4-Dependent Manner

    PubMed Central

    Yang, Shuhua; Sugawara, Shunji; Monodane, Toshihiko; Nishijima, Masahiro; Adachi, Yoshiyuki; Akashi, Sachiko; Miyake, Kensuke; Hase, Sumihiro; Takada, Haruhiko

    2001-01-01

    Teichuronic acid (TUA), a component of the cell walls of the gram-positive organism Micrococcus luteus (formerly Micrococcus lysodeikticus), induced inflammatory cytokines in C3H/HeN mice but not in lipopolysaccharide (LPS)-resistant C3H/HeJ mice that have a defect in the Toll-like receptor 4 (TLR4) gene, both in vivo and in vitro, similarly to LPS (T. Monodane, Y. Kawabata, S. Yang, S. Hase, and H. Takada, J. Med. Microbiol. 50:4–12, 2001). In this study, we found that purified TUA (p-TUA) induced tumor necrosis factor alpha (TNF-α) in murine monocytic J774.1 cells but not in mutant LR-9 cells expressing membrane CD14 at a lower level than the parent J774.1 cells. The TNF-α-inducing activity of p-TUA in J774.1 cells was completely inhibited by anti-mouse CD14 monoclonal antibody (MAb). p-TUA also induced interleukin-8 (IL-8) in human monocytic THP-1 cells differentiated to macrophage-like cells expressing CD14. Anti-human CD14 MAb, anti-human TLR4 MAb, and synthetic lipid A precursor IVA, an LPS antagonist, almost completely inhibited the IL-8-inducing ability of p-TUA, as well as LPS, in the differentiated THP-1 cells. Reduced p-TUA did not exhibit any activities in J774.1 or THP-1 cells. These findings strongly suggested that M. luteus TUA activates murine and human monocytic cells in a CD14- and TLR4-dependent manner, similar to LPS. PMID:11254554

  10. Micrococcus luteus teichuronic acids activate human and murine monocytic cells in a CD14- and toll-like receptor 4-dependent manner.

    PubMed

    Yang, S; Sugawara, S; Monodane, T; Nishijima, M; Adachi, Y; Akashi, S; Miyake, K; Hase, S; Takada, H

    2001-04-01

    Teichuronic acid (TUA), a component of the cell walls of the gram-positive organism Micrococcus luteus (formerly Micrococcus lysodeikticus), induced inflammatory cytokines in C3H/HeN mice but not in lipopolysaccharide (LPS)-resistant C3H/HeJ mice that have a defect in the Toll-like receptor 4 (TLR4) gene, both in vivo and in vitro, similarly to LPS (T. Monodane, Y. Kawabata, S. Yang, S. Hase, and H. Takada, J. Med. Microbiol. 50:4-12, 2001). In this study, we found that purified TUA (p-TUA) induced tumor necrosis factor alpha (TNF-alpha) in murine monocytic J774.1 cells but not in mutant LR-9 cells expressing membrane CD14 at a lower level than the parent J774.1 cells. The TNF-alpha-inducing activity of p-TUA in J774.1 cells was completely inhibited by anti-mouse CD14 monoclonal antibody (MAb). p-TUA also induced interleukin-8 (IL-8) in human monocytic THP-1 cells differentiated to macrophage-like cells expressing CD14. Anti-human CD14 MAb, anti-human TLR4 MAb, and synthetic lipid A precursor IV(A), an LPS antagonist, almost completely inhibited the IL-8-inducing ability of p-TUA, as well as LPS, in the differentiated THP-1 cells. Reduced p-TUA did not exhibit any activities in J774.1 or THP-1 cells. These findings strongly suggested that M. luteus TUA activates murine and human monocytic cells in a CD14- and TLR4-dependent manner, similar to LPS.

  11. Toll-like receptor 4 and CD14 mRNA expression are lower in resistive exercise-trained elderly women.

    PubMed

    Flynn, Michael G; McFarlin, Brian K; Phillips, Melody D; Stewart, Laura K; Timmerman, Kyle L

    2003-11-01

    The purpose of this study was to examine the influence of resistive exercise training and hormone status on mRNA expression of toll-like receptor 4 (TLR4), CD14, IL-1beta, IL-6, and TNF-alpha. Resistive exercise-trained women on "traditional" hormone replacements [hormone replacement therapy (HRT), n = 9], not taking hormones (NHR, n = 6), or taking medications known to influence bone (MIB, n = 7) were compared with untrained subjects not taking supplemental hormones (Con, n = 6). Blood was taken from trained subjects before, immediately after, and 2 h after resistive exercise (same time points for resting Con). TLR4 mRNA expression (RT-PCR) was not different among groups or across time but was significantly (P = 0.044) lower (1.9-fold) when trained groups were collapsed and compared with Con. There was also a significant group effect (P < 0.0001) for TLR4 mRNA when expressed per monocyte. CD14 expression was significantly (P = 0.006) lower (2.3-fold) for training groups collapsed and compared with Con. CD14 mRNA, expressed per monocyte, was significantly lower immediately after resistive exercise for NHR, HRT, and MIB compared with Con. There were few significant effects detected for IL-6, IL-1beta, and TNF-alpha mRNA, but there was a significant group effect (P < 0.0001) for TNF-alpha mRNA expressed per monocyte (Con > HRT, NHR, MIB). These findings suggest that there may be a resistive exercise training-induced reduction in TLR4/CD14 expression in older women. Further research is needed to determine whether lower TLR4/CD14 could explain the lower LPS-stimulated inflammatory cytokines observed in these women.

  12. Rapid Accumulation of CD14+CD11c+ Dendritic Cells in Gut Mucosa of Celiac Disease after in vivo Gluten Challenge

    PubMed Central

    Beitnes, Ann-Christin Røberg; Ráki, Melinda; Brottveit, Margit; Lundin, Knut Erik Aslaksen

    2012-01-01

    Background Of antigen-presenting cells (APCs) expressing HLA-DQ molecules in the celiac disease (CD) lesion, CD11c+ dendritic cells (DCs) co-expressing the monocyte marker CD14 are increased, whereas other DC subsets (CD1c+ or CD103+) and CD163+CD11c− macrophages are all decreased. It is unclear whether these changes result from chronic inflammation or whether they represent early events in the gluten response. We have addressed this in a model of in vivo gluten challenge. Methods Treated HLA-DQ2+ CD patients (n = 12) and HLA-DQ2+ gluten-sensitive control subjects (n = 12) on a gluten-free diet (GFD) were orally challenged with gluten for three days. Duodenal biopsies obtained before and after gluten challenge were subjected to immunohistochemistry. Single cell digests of duodenal biopsies from healthy controls (n = 4), treated CD (n = 3) and untreated CD (n = 3) patients were analyzed by flow cytometry. Results In treated CD patients, the gluten challenge increased the density of CD14+CD11c+ DCs, whereas the density of CD103+CD11c+ DCs and CD163+CD11c− macrophages decreased, and the density of CD1c+CD11c+ DCs remained unchanged. Most CD14+CD11c+ DCs co-expressed CCR2. The density of neutrophils also increased in the challenged mucosa, but in most patients no architectural changes or increase of CD3+ intraepithelial lymphocytes (IELs) were found. In control tissue no significant changes were observed. Conclusions Rapid accumulation of CD14+CD11c+ DCs is specific to CD and precedes changes in mucosal architecture, indicating that this DC subset may be directly involved in the immunopathology of the disease. The expression of CCR2 and CD14 on the accumulating CD11c+ DCs indicates that these cells are newly recruited monocytes. PMID:22438948

  13. Rapid accumulation of CD14+CD11c+ dendritic cells in gut mucosa of celiac disease after in vivo gluten challenge.

    PubMed

    Beitnes, Ann-Christin Røberg; Ráki, Melinda; Brottveit, Margit; Lundin, Knut Erik Aslaksen; Jahnsen, Frode Lars; Sollid, Ludvig Magne

    2012-01-01

    Of antigen-presenting cells (APCs) expressing HLA-DQ molecules in the celiac disease (CD) lesion, CD11c(+) dendritic cells (DCs) co-expressing the monocyte marker CD14 are increased, whereas other DC subsets (CD1c(+) or CD103(+)) and CD163(+)CD11c(-) macrophages are all decreased. It is unclear whether these changes result from chronic inflammation or whether they represent early events in the gluten response. We have addressed this in a model of in vivo gluten challenge. Treated HLA-DQ2(+) CD patients (n = 12) and HLA-DQ2(+) gluten-sensitive control subjects (n = 12) on a gluten-free diet (GFD) were orally challenged with gluten for three days. Duodenal biopsies obtained before and after gluten challenge were subjected to immunohistochemistry. Single cell digests of duodenal biopsies from healthy controls (n = 4), treated CD (n = 3) and untreated CD (n = 3) patients were analyzed by flow cytometry. In treated CD patients, the gluten challenge increased the density of CD14(+)CD11c(+) DCs, whereas the density of CD103(+)CD11c(+) DCs and CD163(+)CD11c(-) macrophages decreased, and the density of CD1c(+)CD11c(+) DCs remained unchanged. Most CD14(+)CD11c(+) DCs co-expressed CCR2. The density of neutrophils also increased in the challenged mucosa, but in most patients no architectural changes or increase of CD3(+) intraepithelial lymphocytes (IELs) were found. In control tissue no significant changes were observed. Rapid accumulation of CD14(+)CD11c(+) DCs is specific to CD and precedes changes in mucosal architecture, indicating that this DC subset may be directly involved in the immunopathology of the disease. The expression of CCR2 and CD14 on the accumulating CD11c(+) DCs indicates that these cells are newly recruited monocytes.

  14. Neonatal Plasma Polarizes TLR4-Mediated Cytokine Responses towards Low IL-12p70 and High IL-10 Production via Distinct Factors

    PubMed Central

    Belderbos, Mirjam E.; Levy, Ofer; Stalpers, Femke; Kimpen, Jan L.; Meyaard, Linde; Bont, Louis

    2012-01-01

    Human neonates are highly susceptible to infection, which may be due in part to impaired innate immune function. Neonatal Toll-like receptor (TLR) responses are biased against the generation of pro-inflammatory/Th1-polarizing cytokines, yet the underlying mechanisms are incompletely defined. Here, we demonstrate that neonatal plasma polarizes TLR4-mediated cytokine production. When exposed to cord blood plasma, mononuclear cells (MCs) produced significantly lower TLR4-mediated IL-12p70 and higher IL-10 compared to MC exposed to adult plasma. Suppression by neonatal plasma of TLR4-mediated IL-12p70 production, but not induction of TLR4-mediated IL-10 production, was maintained up to the age of 1 month. Cord blood plasma conferred a similar pattern of MC cytokine responses to TLR3 and TLR8 agonists, demonstrating activity towards both MyD88-dependent and MyD88-independent agonists. The factor causing increased TLR4-mediated IL-10 production by cord blood plasma was heat-labile, lost after protein depletion and independent of lipoprotein binding protein (LBP) or soluble CD14 (sCD14). The factor causing inhibition of TLR4-mediated IL-12p70 production by cord blood plasma was resistant to heat inactivation or protein depletion and was independent of IL-10, vitamin D and prostaglandin E2. In conclusion, human neonatal plasma contains at least two distinct factors that suppress TLR4-mediated IL-12p70 production or induce IL-10 or production. Further identification of these factors will provide insight into the ontogeny of innate immune development and might identify novel targets for the prevention and treatment of neonatal infection. PMID:22442690

  15. Validation of an HPLC method for analysis of DB-67 and its water soluble prodrug in mouse plasma.

    PubMed

    Horn, Jamie; Jordan, Sherri L; Song, Lin; Roberts, Michael J; Anderson, Bradley D; Leggas, Markos

    2006-11-21

    A method for the quantitation of DB-67 ((20S)-10-hydroxy-7-tert-butyldimethylsilylcamptothecin) lactone and carboxylate in mouse plasma has been developed, validated, and applied in pharmacokinetic studies. The analytes were separated by reversed-phase chromatography with fluorescence detection. Validation demonstrated the selectivity and specificity for the carboxylate and lactone, with linearity between 1-300ng/mL and 2.5-300ng/mL for the carboxylate and lactone, respectively (accuracy 90-110% of theory and coefficient of variation < or =5.7%). Carboxylate to lactone conversion was <4% using this method. The assay was found to be suitable for the analysis of DB-67 lactone and carboxylate in pharmacokinetic studies following intravenous administration of DB-67 or its delta-aminobutyric acid ester derivative.

  16. Fusion of the Fc part of human IgG1 to CD14 enhances its binding to gram-negative bacteria and mediates phagocytosis by Fc receptors of neutrophils.

    PubMed

    Vida, András; Bardoel, Bart; Milder, Fin; Majoros, László; Sümegi, Andrea; Bácsi, Attila; Vereb, György; van Kessel, Kok P M; van Strijp, Jos A G; Antal-Szalmás, Péter

    2012-08-30

    Microbial resistance to antimicrobial drugs is promoting a search for new antimicrobial agents that target highly conservative structures of pathogens. Human CD14 - a known pattern recognition receptor (PRR) which recognizes multiple ligands from different microbes might be a worthy candidate. The aim of our work was to create a CD14/Fc dimer protein and evaluate its whole bacteria binding and opsonizing capabilities. Fusion of CD14 with the fragment crystallisable (Fc) part of human IgG1 could not only lead to an artificial opsonin but the dimerization through the Fc part might also increase its affinity to different ligands. Human CD14 and the Fc part of human IgG1 was fused and expressed in HEK293 cells. A histidine tagged CD14 (CD14/His) was also expressed as control. Using flow cytometry we could prove that CD14/Fc bound to whole Gram-negative bacteria, especially to short lipopolysaccharide (Ra and Re) mutants, and weak interaction was observed between the fusion protein and Listeria monocytogenes. Other Gram-positive bacteria and fungi did not show any association with CD14/Fc. CD14/His showed about 50-times less potent binding to Gram-negative bacteria. CD14/Fc acted as an opsonin and enhanced phagocytosis of these bacteria by neutrophil granulocytes, monocyte-derived macrophages and dendritic cells. Internalization of bacteria was confirmed by trypan blue quenching and confocal microscopy. On neutrophils the Fc part of the fusion protein was recognized by Fc receptors (CD16, CD32), as determined by blocking experiments. CD14/Fc enhanced the killing of bacteria in an ex vivo whole blood assay. Our experiments confirm that PRR/Fc fusion proteins can give a boost to FcR dependent phagocytosis and killing provided the antimicrobial part binds efficiently to microbes.

  17. Hansen solubility parameters for assay method optimization of simvastatin, ramipril, atenolol, hydrochlorothiazide and aspirin in human plasma using liquid chromatography with tandem mass spectrometry.

    PubMed

    Devalapalli, Murali Mohan Reddy; Cheruvu, Hanumanth Srikanth; Yertha, Thejaswi; Veeravalli, Vijaya Bhaskar; Sampathi, Sunitha; Shivakumar, Savithri

    2017-09-01

    A simple, specific, sensitive, validated method was developed using liquid chromatography with tandem mass spectrometry with electrospray ionization of human plasma for the simultaneous estimation of drugs (simvastatin, ramipril, atenolol, hydrochlorothiazide, and aspirin) of Polycap(TM) capsule used in cardiovascular therapy. The interaction of these actives including internal standards between the stationary and mobile phase were investigated using Hansen solubility parameters. Chromatographic separation was performed on Phenomenex Synergi Polar-RP (30 × 2 mm, 4 μm) column with a gradient mobile phase composition of acetonitrile and 5 mM ammonium formate for positive mode and 0.1% formic acid in both water and acetonitrile for negative mode. The flow rate and runtime were 1.0 mL/min and 3.5 min, respectively. Sample extraction was done by protein precipitation using acetonitrile, enabling a fast analysis. The calibration ranges from 0.1 to 100, 0.1 to 100, and 1 to 1000 ng/mL for simvastatin, ramipril, and atenolol using internal standard carbamazepine in positive mode, respectively, whereas it was 0.3-300 and 2-2000 ng/mL for hydrochlorothiazide and aspirin using internal standard 7-hydroxy coumarin in negative mode, respectively. Hansen solubility parameters can be used as a high-throughput optimizing tool for column and mobile phase selection in bioanalysis. This validated bioanalytical method has the potential for future fixed dose combination based preclinical and clinical studies that can save analysis time. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. [Can mannose-binding lectin and plasma level of soluble urokinase receptor be used in diagnosis and treatment monitorization of brucellosis patients?].

    PubMed

    Karsen, Hasan; Cesur, Salih; Karaağaç, Leman; Binici, Irfan; Fidan, Yasemin; Oğüş, Elmas; Demiröz, Ali Pekcan

    2012-07-01

    The aim of this study was to evaluate the diagnostic value of serum mannose-binding lectin (MBL) and plasma soluble urokinase plasminogen activator receptor (SuPAR) levels in monitoring the treatment in patients with brucellosis, by comparing their levels before and after treatment with the values obtained from healthy control group. Thirty brucellosis patients (mean age: 25.8 ± 12.2 years; 15 were male) and 28 healthy controls (mean age: 29.3 ± 12.3 years; 15 were male) were included in the study. Patients were diagnosed with brucellosis according to the characteristic clinical findings and by brucella standard tube agglutination test (SAT) titer ≥ 1/160 and/or blood culture positivity. Serum MBL (Antibodyshop, Denmark) and plasma SuPAR (Virogates, Denmark) levels were investigated with commercial ELISA kits. In our study, no statistical significance was observed between the pre-treatment (13.8 ± 13.4 ng/ml) and post-treatment (12.4 ± 13.1 ng/ml) MBL levels of the patient group and MBL levels of the control group (16.5 ± 14.8 ng/ml) (p> 0.05). Moreover, the mean SuPAR levels measured in pre-treatment and post-treatment plasma samples of the brucellosis patients was 5.1 ± 1.9 ng/ml and 2.9 ± 1.3 ng/ml, respectively, while the mean SuPAR level was 1.8 ± 0.5 ng/ml in the control group. The difference between mean SuPAR levels of patients in pre- and post-treatment samples was found statistically significant (p< 0.001). In addition SuPAR levels were significantly higher in patients before and after treatment than the control group (p> 0.001). In conclusion, plasma SuPAR level would be a useful marker for the diagnosis and treatment follow up of the patients with brucellosis.

  19. High habitual dietary α-linolenic acid intake is associated with decreased plasma soluble interleukin-6 receptor concentrations in male twins123

    PubMed Central

    Ziegler, Thomas R; Bostick, Roberd M; Manatunga, Amita K; Jones, Dean P; Goldberg, Jack; Miller, Andrew; Vogt, Gerald; Wilson, Peter W; Jones, Linda; Shallenberger, Lucy; Vaccarino, Viola

    2010-01-01

    Background: α-Linolenic acid (ALA) is associated with a low risk of cardiovascular disease; however, the underlying mechanism is not completely known. Objective: The objective was to examine whether habitual dietary ALA intake is associated with plasma concentrations of inflammatory biomarkers after control for shared genetic and common environmental factors. Design: We cross-sectionally studied 353 middle-aged male twins. Habitual diet was assessed with the Willett food-frequency questionnaire. Fasting plasma concentrations of interleukin-6 (IL-6) and its soluble receptor (sIL-6R), high-sensitivity C-reactive protein (hsCRP), and tumor necrosis factor-α (TNF-α) were measured. Linear mixed-effect regression analysis was used to partition the overall association into within- and between-pair associations. Results: A 1-g increment in habitual dietary ALA intake was associated with 11.0% lower concentrations of sIL-6R (P = 0.004) but not of IL-6 (P = 0.31), TNF-α (P = 0.16), or hsCRP (P = 0.36) after adjustment for energy intake, nutritional factors, known cardiovascular disease risk factors, and medications. After further control for shared genetic and common environmental factors by comparison of brothers within a twin pair, a twin with a 1-g higher ALA intake was likely to have 10.9% (95% CI: 3.7%, 17.6%; P = 0.004) lower sIL-6R concentrations than his co-twin with a low intake, whereas ALA intake was not significantly associated with plasma concentrations of IL-6, TNF-α, or hsCRP. These results were validated by using 1000 bootstrap samples. Conclusions: Habitual dietary ALA intake is inversely associated with plasma sIL-6R concentrations independent of shared genetic and common environmental influences. Lowering sIL-6R may be a mechanism underlying the cardioprotective properties of habitual dietary ALA. This study was registered at clinicaltrials.gov as NCT00017836. PMID:20463041

  20. Systemic increase in human maternal circulating CD14+CD16- MCP-1+ monocytes as a marker of labor.

    PubMed

    Bardou, Marc; Hadi, Tarik; Mace, Guillaume; Pesant, Matthieu; Debermont, Julie; Barrichon, Marina; Wendremaire, Maeva; Laurent, Nicole; Sagot, Paul; Lirussi, Frédéric

    2014-01-01

    To study the influence of pregnancy and labor on the proportion and level of activation of monocyte subpopulations in human pregnancy. Peripheral blood samples were obtained from healthy nonpregnant women (n = 6); women in the third-trimester of healthy pregnancies (n = 18) and women with preterm premature rupture of membranes (n = 46), just before delivery for the last 2 groups. Monocyte subpopulations were characterized by flow cytometry using CD14, CD16, and activation level using macrophage chemoattractant protein-1 (MCP-1) and CCR2 antibodies. The relative proportion of each monocyte subset in nonpregnant women was similar to that in women with healthy or complicated pregnancies. However, pregnancy was associated with a significant decrease in MCP-1 expressing monocytes (79.5% ± 19.8% vs 9.3% ± 6.8% and 11.9% ± 8.3% for nonpregnant, healthy pregnancy, and preterm premature rupture of membranes (respectively, P < .05). Spontaneous labor was associated with a return to nonpregnant values for the proportion of MCP-1 expressing monocytes in both normal (74.4% ± 16.9) and preterm premature rupture of membranes pregnancy (68.4% ± 35.6), irrespective of the mode of delivery (vaginal or cesarean section). This was not observed in women who delivered without spontaneous labor onset. CCR-2 (MCP-1 receptor) expression was not modified in monocytes at the time of labor, but was significantly increased in granulocytes (3646 ± 1080 vs 7338 ± 2718 for nonlaboring and laboring preterm premature rupture of membranes, respectively, P < .05) CONCLUSION: In light of previous reports of a role for MCP-1 in labor, our results suggest the downregulation of activation levels of monocytes, via MCP-1 expression might be involved in maternofetal immune tolerance. Monocyte reactivation might be associated with labor. Copyright © 2014 Mosby, Inc. All rights reserved.

  1. Increased Th17-inducing activity of CD14+ CD163 low myeloid cells in intestinal lamina propria of patients with Crohn's disease.

    PubMed

    Ogino, Takayuki; Nishimura, Junichi; Barman, Soumik; Kayama, Hisako; Uematsu, Satoshi; Okuzaki, Daisuke; Osawa, Hideki; Haraguchi, Naotsugu; Uemura, Mamoru; Hata, Taishi; Takemasa, Ichiro; Mizushima, Tsunekazu; Yamamoto, Hirofumi; Takeda, Kiyoshi; Doki, Yuichiro; Mori, Masaki

    2013-12-01

    Abnormal activity of innate immune cells and T-helper (Th) 17 cells has been implicated in the pathogenesis of autoimmune and inflammatory diseases, including Crohn's disease (CD). Intestinal innate immune (myeloid) cells have been found to induce development of Th17 cells in mice, but it is not clear if this occurs in humans or in patients with CD. We investigated whether human intestinal lamina propria cells (LPCs) induce development of Th17 cells and whether these have a role in the pathogenesis of CD. Normal intestinal mucosa samples were collected from patients with colorectal cancer and noninflamed and inflamed regions of mucosa were collected from patients with CD. LPCs were isolated by enzymatic digestion and analyzed for expression of HLA-DR, lineage markers CD14 and CD163 using flow cytometry. Among HLA-DR(high) Lin(-) cells, we identified a subset of CD14(+) CD163(low) cells in intestinal LPCs; this subset expressed Toll-like receptor (TLR) 2, TLR4, and TLR5 mRNAs and produced interleukin (IL)-6, IL-1β, and tumor necrosis factor in response to lipopolysaccharide. In vitro co-culture with naïve T cells revealed that CD14(+) CD163(low) cells induced development of Th17 cells. CD14(+) CD163(low) cells from inflamed regions of mucosa of patients with CD expressed high levels of IL-6, IL-23p19, and tumor necrosis factor mRNAs, and strongly induced Th17 cells. CD14(+) CD163(low) cells from the noninflamed mucosa of patients with CD also had increased abilities to induce Th17 cells compared with those from normal intestinal mucosa. CD14(+) CD163(low) cells in intestinal LPCs from normal intestinal mucosa induce differentiation of naive T cells into Th17 cells; this activity is increased in mucosal samples from patients with CD. These findings show how intestinal myeloid cell types could contribute to pathogenesis of CD and possibly other Th17-associated diseases. Copyright © 2013 AGA Institute. Published by Elsevier Inc. All rights reserved.

  2. Unchanged plasma levels of the soluble urokinase plasminogen activator receptor in elective coronary artery bypass graft surgery patients and cardiopulmonary bypass use.

    PubMed

    Gozdzik, Waldemar; Adamik, Barbara; Gozdzik, Anna; Rachwalik, Maciej; Kustrzycki, Wojciech; Kübler, Andrzej

    2014-01-01

    The soluble urokinase plasminogen activator receptor (suPAR) has been recently recognized as a potential biological marker of various disease states, but the impact of a major surgical intervention on the suPAR level has not yet been established. The aim of our study was to investigate if the induction of a systemic inflammatory reaction in response to cardiopulmonary bypass would be accompanied by an increase in the plasma suPAR level. Patients undergoing coronary artery bypass grafting under cardiopulmonary bypass (CPB) were added. Based on the baseline suPAR level, patients were divided into group 1 (suPAR within normal range) or group 2 (suPAR above range). Blood was collected before the induction of anesthesia and 6 and 24 hours after surgery. Plasma suPAR, IL-6, IL-8, TNF-α, troponin I, NT-proBNP, and NGAL were quantified to assess the impact of surgical trauma on these markers. The baseline suPAR level was within the normal range in 31 patients (3.3 ng/mL), and elevated in 29 (5.1 ng/mL) (p<0.001). Baseline mediators of systemic inflammatory reaction concentrations (IL-6, TNF-α, and IL-8) and organ injury indices (troponin I, NT-proBNP, and NGAL) were low and increased after surgery in all patients (p<0.05). The surgery did not cause significant changes in the suPAR level either at 6 or 24 hours after, however the difference between groups observed at baseline remained substantial during the postoperative period. There was no change in the suPAR level observed in patients subjected to elective cardiac coronary artery bypass surgery and CPB, despite activation of a systemic inflammatory reaction.

  3. Solubility Database

    National Institute of Standards and Technology Data Gateway

    SRD 106 IUPAC-NIST Solubility Database (Web, free access)   These solubilities are compiled from 18 volumes (Click here for List) of the International Union for Pure and Applied Chemistry(IUPAC)-NIST Solubility Data Series. The database includes liquid-liquid, solid-liquid, and gas-liquid systems. Typical solvents and solutes include water, seawater, heavy water, inorganic compounds, and a variety of organic compounds such as hydrocarbons, halogenated hydrocarbons, alcohols, acids, esters and nitrogen compounds. There are over 67,500 solubility measurements and over 1800 references.

  4. Plasma fat-soluble vitamin and carotenoid concentrations after plant sterol and plant stanol consumption: a meta-analysis of randomized controlled trials.

    PubMed

    Baumgartner, Sabine; Ras, Rouyanne T; Trautwein, Elke A; Mensink, Ronald P; Plat, Jogchum

    2017-04-01

    Plant sterols and stanols interfere with intestinal cholesterol absorption, and it has been questioned whether absorption and plasma concentrations of fat-soluble vitamins and carotenoids are also affected. We conducted a meta-analysis to assess the effects of plant sterol and stanol consumption on plasma fat-soluble vitamin and carotenoid concentrations. Forty-one randomized controlled trials involving 3306 subjects were included. Weighted absolute and relative changes of non-standardized and total cholesterol (TC)-standardized values (expressed as summary estimates and 95 % CIs) were calculated for three fat-soluble vitamins (α- and γ-tocopherol, retinol and vitamin D) and six carotenoids (β-carotene, α-carotene, lycopene, lutein, zeaxanthin and β-cryptoxanthin) using a random effects model. Heterogeneity was assessed using predefined subject and treatment characteristics. Average plant sterol or stanol intake was 2.5 g/d. Relative non-standardized and TC-standardized concentrations of β-carotene decreased by, respectively, -16.3 % (95 % CI -18.3; -14.3) and -10.1 % (-12.3; -8.0), α-carotene by -14.4 % (-17.5; 11.3) and -7.8 % (-11.3; -4.3), and lycopene by -12.3 % (-14.6; -10.1) and -6.3 % (-8.6; -4.0). Lutein concentrations decreased by -7.4 % (-10.1; -4.8), while TC-standardized concentrations were not changed. For zeaxanthin, these values were -12.9 % (-18.9; -6.8) and -7.7 % (-13.8; -1.7) and for β-cryptoxanthin -10.6 % (-14.3; -6.9) and -4.8 % (-8.7; -0.9). Non-standardized α-tocopherol concentrations decreased by -7.1 % (-8.0; -6.2) and γ-tocopherol by -6.9 % (-9.8; -3.9), while TC-standardized tocopherol concentrations were not changed. Non-standardized retinol and vitamin D concentrations were not affected. Results were not affected by baseline concentrations, dose, duration and type of plant sterols/stanols, except for significant effects of duration (≤4 vs. >4 weeks) on TC-standardized lutein concentrations (1.0 vs. -5.6

  5. Ion-pair cloud-point extraction: a new method for the determination of water-soluble vitamins in plasma and urine.

    PubMed

    Heydari, Rouhollah; Elyasi, Najmeh S

    2014-10-01

    A novel, simple, and effective ion-pair cloud-point extraction coupled with a gradient high-performance liquid chromatography method was developed for determination of thiamine (vitamin B1 ), niacinamide (vitamin B3 ), pyridoxine (vitamin B6 ), and riboflavin (vitamin B2 ) in plasma and urine samples. The extraction and separation of vitamins were achieved based on an ion-pair formation approach between these ionizable analytes and 1-heptanesulfonic acid sodium salt as an ion-pairing agent. Influential variables on the ion-pair cloud-point extraction efficiency, such as the ion-pairing agent concentration, ionic strength, pH, volume of Triton X-100, extraction temperature, and incubation time have been fully evaluated and optimized. Water-soluble vitamins were successfully extracted by 1-heptanesulfonic acid sodium salt (0.2% w/v) as ion-pairing agent with Triton X-100 (4% w/v) as surfactant phase at 50°C for 10 min. The calibration curves showed good linearity (r(2) > 0.9916) and precision in the concentration ranges of 1-50 μg/mL for thiamine and niacinamide, 5-100 μg/mL for pyridoxine, and 0.5-20 μg/mL for riboflavin. The recoveries were in the range of 78.0-88.0% with relative standard deviations ranging from 6.2 to 8.2%. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Identification of soluble N-ethylmaleimide-sensitive factor attachment protein receptor exocytotic machinery in human plasma cells: SNAP-23 is essential for antibody secretion.

    PubMed

    Reales, Elena; Mora-López, Francisco; Rivas, Verónica; García-Poley, Antonio; Brieva, José A; Campos-Caro, Antonio

    2005-11-15

    Plasma cells (PC) are B-lymphocytes terminally differentiated in a postmitotic state, with the unique purpose of manufacturing and exporting Igs. Despite the importance of this process in the survival of vertebrates, no studies have been made to understand the molecular events that regulate Ig exocytosis by PC. The present study explores the possible presence of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) system in human PC, and examines its functional role in Ig secretion. Syntaxin-2, Syntaxin-3, Syntaxin-4, vesicle-associated membrane protein (VAMP)-2, VAMP-3, and synaptosome-associated protein (SNAP)-23 could be readily detected in normal human PC obtained from intestinal lamina propria and blood, as well as in human PC lines. Because SNAP-23 plays a central role in SNAREs complex formation, it was chosen to examine possible functional implications of the SNARE system in PC Ig secretion. When recombinant SNAP-23 fusion protein was introduced into the cells, a complete abolishment of Ig production was observed in the culture supernatants of PC lines, as well as in those of normal PC. These results provide insights, for the first time, into the molecular machinery of constitutive vesicular trafficking in human PC Ig secretion and present evidence indicating that at least SNAP-23 is essential for Ab production.

  7. Etomidate Mitigates Lipopolysaccharide-Induced CD14 and TREM-1 Expression, NF-κB Activation, and Pro-inflammatory Cytokine Production in Rat Macrophages.

    PubMed

    Liu, Ming; Zhang, Yu; Xiong, Jun-Yu; Wang, Yan; Lv, Shen

    2016-02-01

    This study was aimed at investigating the effect of etomidate on the viability of rat macrophages and the function of lipopolysaccharide (LPS)-stimulated macrophages as well as the potential mechanisms. Rat macrophages were isolated and treated with different doses of etomidate for 24 h, and their viability was determined by the CCK-8 assay. Furthermore, macrophages were treated with, or without, 1 μg/ml of LPS, and/or 2.5 or 5 μM etomidate in the presence or absence of a TREM-1 inhibitor (LP17, 100 ng/ml), and the levels of TNF-α, IL-6, CD14, and TREM-1 in the different groups of cells were determined by quantitative RT-PCR, ELISA, and Western blot assays. The levels of NF-κB activation in the different groups of cells were analyzed by an electrophoretic mobility shift assay (EMSA). Etomidate at 31.25 μM or a low dose did not affect the viability of rat macrophages, while etomidate at higher doses reduced the viability of macrophages in vitro. Treatment with 2.5 or 5 μM etomidate or with LP17 alone did not affect the levels of TNF-α, IL-6, CD-14, and TREM-1 in macrophages. Treatment with etomidate significantly mitigated LPS-stimulated TNF-α, IL-6, CD-14, and TREM-1 expression (p < 0.05 for all) and inhibited LPS-induced NF-κB activation in macrophages in vitro. However, treatment with both etomidate and LP17 did not enhance the inhibitory effects in macrophages. Hence, etomidate mitigates LPS-up-regulated pro-inflammatory cytokine production and inhibits LPS-enhanced CD14 and TREM-1 expression and NF-κB activation in macrophages.

  8. Cis and Trans Acting Factors Involved in Human Cytomegalovirus Experimental and Natural Latent Infection of CD14 (+) Monocytes and CD34 (+) Cells

    PubMed Central

    Pari, Gregory S.

    2013-01-01

    The parameters involved in human cytomegalovirus (HCMV) latent infection in CD14 (+) and CD34 (+) cells remain poorly identified. Using next generation sequencing we deduced the transcriptome of HCMV latently infected CD14 (+) and CD34 (+) cells in experimental as well as natural latency settings. The gene expression profile from natural infection in HCMV seropositive donors closely matched experimental latency models, and included two long non-coding RNAs (lncRNAs), RNA4.9 and RNA2.7 as well as the mRNAs encoding replication factors UL84 and UL44. Chromatin immunoprecipitation assays on experimentally infected CD14 (+) monocytes followed by next generation sequencing (ChIP-Seq) were employed to demonstrate both UL84 and UL44 proteins interacted with the latent viral genome and overlapped at 5 of the 8 loci identified. RNA4.9 interacts with components of the polycomb repression complex (PRC) as well as with the MIE promoter region where the enrichment of the repressive H3K27me3 mark suggests that this lncRNA represses transcription. Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE), which identifies nucleosome-depleted viral DNA, was used to confirm that latent mRNAs were associated with actively transcribed, FAIRE analysis also showed that the terminal repeat (TR) region of the latent viral genome is depleted of nucleosomes suggesting that this region may contain an element mediating viral genome maintenance. ChIP assays show that the viral TR region interacts with factors associated with the pre replication complex and a plasmid subclone containing the HCMV TR element persisted in latently infected CD14 (+) monocytes, strongly suggesting that the TR region mediates viral chromosome maintenance. PMID:23717203

  9. Helicobacter pylori Lipopolysaccharide Binds to CD14 and Stimulates Release of Interleukin-8, Epithelial Neutrophil-Activating Peptide 78, and Monocyte Chemotactic Protein 1 by Human Monocytes

    PubMed Central

    Bliss, Charles M.; Golenbock, Douglas T.; Keates, Sarah; Linevsky, Joanne K.; Kelly, Ciarán P.

    1998-01-01

    Helicobacter pylori gastritis is characterized by leukocyte infiltration of the gastric mucosa. The aims of this study were to determine whether H. pylori-derived factors stimulate chemokine release from human monocytes and to ascertain whether H. pylori lipopolysaccharide (LPS) may be responsible for this effect. Human peripheral blood monocytes were exposed to an H. pylori water extract (HPE) or to purified H. pylori LPS. Levels of the chemokines interleukin-8 (IL-8), epithelial neutrophil-activating peptide 78 (ENA-78), and monocyte chemotactic protein 1 (MCP-1) were measured by enzyme-linked immunosorbent assay. The contribution of H. pylori LPS to monocyte activation was determined by using the LPS antagonist Rhodobacter sphaeroides lipid A (RSLA) and a blocking monoclonal antibody to CD14 (60bca). HPE increased monocyte secretion of IL-8, ENA-78, and MCP-1. Heat treatment of HPE did not reduce its ability to activate monocytes. Purified H. pylori LPS also stimulated monocyte chemokine production but was 1,000-fold less potent than Salmonella minnesota lipid A. RSLA blocked H. pylori LPS-induced monocyte IL-8 release in a dose-dependent fashion (maximal inhibition 82%, P < 0.001). RSLA also inhibited HPE-induced IL-8 release (by 93%, P < 0.001). The anti-CD14 monoclonal antibody 60bca substantially inhibited IL-8 release from HPE-stimulated monocytes (by 88%, P < 0.01), whereas the nonblocking anti-CD14 monoclonal antibody did not. These experiments with potent and specific LPS inhibitors indicate that the main monocyte-stimulating factor in HPE is LPS. H. pylori LPS, acting through CD14, stimulates human monocytes to release the neutrophil-activating chemokines IL-8 and ENA-78 and the monocyte-activating chemokine MCP-1. Despite its low relative potency, H. pylori LPS may play an important role in the pathogenesis of H. pylori gastritis. PMID:9784544

  10. CD14-159C/T and TLR9-1237T/C polymorphisms are not associated with gastric cancer risk in Caucasian populations

    PubMed Central

    Hold, Georgina L.; Rabkin, Charles S.; Gammon, Marilie D.; Berry, Susan H.; Smith, Malcolm G.; Lissowska, Jolanta; Risch, Harvey A.; Chow, Wong-Ho; Mowat, N. Ashley G.; Vaughan, Thomas L.; El-Omar, Emad M.

    2009-01-01

    Host genetic factors play an important role in modifying the risk of human disease, including cancers of the upper gastrointestinal tract, with increasing interest in Toll-like receptor (TLR) signaling and the impact of genetic polymorphisms in these systems. The CD14-159C/T and the TLR9-1237T/C promoter polymorphisms have previously been shown to be associated with various inflammatory conditions including Helicobacter pylori-induced gastritis in Caucasian populations. In this study, we assessed the association of these two functional single nucleotide polymorphisms with gastric cancer in two independent Caucasian population-based case–control studies of upper gastrointestinal tract cancer, initially in 312 noncardia gastric carcinoma cases and 419 controls and then in 184 noncardia gastric carcinomas, 123 cardia carcinomas, 159 esophageal cancers, and 211 frequency-matched controls. Odds ratios were computed from logistic models and adjusted for potential confounding factors. No significant association was found between the CD14-159C/T and the TLR9-1237T/C promoter polymorphisms and increased risk of gastric cancer. Neither single nucleotide polymorphism has been assessed in a Caucasian gastric cancer case–control study before; although the CD14-159C/T polymorphism has been reported to show no apparent association with H. pylori-related gastric malignancy in a Taiwanese Chinese population. In conclusion, although our earlier preliminary studies suggested that the CD14-159C/T and the TLR9-1237T/C promoter polymorphisms increase the risk of precancerous outcomes, they do not seem to increase the risk of gastric cancer itself. This discrepancy merits further examination. PMID:19337058

  11. Induction of Experimental Arthritis by Borrelial Lipoprotein and CpG Motifs: Are Toll-Like Receptors 2, 4, 9 or CD-14 Involved?

    SciTech Connect

    Batsford, S.; Dunn, J.; Mihatsch, M.

    2011-06-01

    Bacterial lipoproteins and CpG-DNA are ligands for Toll-Like-Receptors (TLR) 2 and 9 respectively. Both classes of molecules were reported to induce experimental arthritis in rodents following direct intra-articular injection. Here we studied: (1) whether arthritis induction by Outer surface (Lipo)protein A (OspA) (B.burgdorferi) involved the TLR-2 as well as the TLR-4 or the CD-14 receptors in addition, and (2) re-examined the arthritogenic potential of CpG-DNA motifs in mice. Following intra-articular injection of the test substances [20 {micro}g recombinant, lipidated OspA; 1nM(6 {micro}g) to 10nM(60 {micro}g) synthetic CpG-DNA], inflammation was monitored by {sup 99}Tc scintigraphy (ratio left/right knee joint uptake > 1.1 indicates inflammation) and by histology. Lipoprotein OspA induced severe, acute arthritis in TLR-2{sup +/+} w.t. but not in TLR-2{sup -/-} mice (p<0.01). There were no significant differences in the severity of arthritis induced in TLR-4{sup +/+} w.t. and TLR-4{sup -/-} mutant mice, or between CD14{sup +/+} w.t. and CD14{sup -/-} mice. CpG-DNA (1or 10 nM) did not cause notable inflammation in C57BL/6 mice; {sup 99}Tc ratios were < 1.0 and histology showed only minimal changes. Induction of arthritis by the OspA lipoprotein of B.burgdorferi involves the TLR-2 receptor, no evidence for additional participation of TLR-4 or CD14 receptors was found. Intra-articular injection of CpG-DNA did not produce manifest joint injury in mice, at variance with previous reports.

  12. NOD2/CARD15, TLR4 and CD14 mutations in Scottish and Irish Crohn's disease patients: evidence for genetic heterogeneity within Europe?

    PubMed

    Arnott, I D R; Nimmo, E R; Drummond, H E; Fennell, J; Smith, B R K; MacKinlay, E; Morecroft, J; Anderson, N; Kelleher, D; O'Sullivan, M; McManus, R; Satsangi, J

    2004-08-01

    NOD2/caspase recruitment domain (CARD)15 variants are identified in up to 50% of Crohn's disease (CD) patients. Functional variants of toll-like receptor-4 (TLR4) and CD14 genes may also be relevant to disease pathophysiology. We aimed to assess the contribution of NOD2/CARD15, TLR4 and CD14 variants in Scottish and Irish CD patients. In all, 612 patients with well-characterised inflammatory bowel disease (252 Scottish CD, 247 Scottish UC, 113 Irish CD) and 304 controls were genotyped for variants of NOD2/CARD15 (1007fsinsC, G908R, R702W, P268S), TLR4 (A299G) and CD14 (T-159C). Genotype-phenotype analyses were performed. Variant 1007fsinsC (P=0.003) and G908R (P=0.008) but not R702W (P=0.269) alleles were more prevalent in Scottish CD (4.7, 1.8 and 7.1%, respectively) than Scottish control (2.3, 0.3 and 5.4%). CD allelic frequencies were lower than the series from Europe (P<0.00001) and North America (P<0.00001) but not Scandinavia (P<0.7). Associations were identified with age at diagnosis (P=0.002), ileal disease (P<0.02), penetrating disease (P=0.04) and inflammatory joint disease (P<0.02). TLR4 and CD14 variants did not differ between CD and controls. In conclusion, we present compelling evidence for genetic heterogeneity within Europe. These NOD2/CARD15 variants have a minor contribution in Scottish and Irish CD patients, consistent with an emerging pattern from Northern Europe.

  13. A novel subset of B7-H3(+)CD14(+)HLA-DR(-/low) myeloid-derived suppressor cells are associated with progression of human NSCLC.

    PubMed

    Zhang, Guangbo; Huang, Haitao; Zhu, Yibei; Yu, Gehua; Gao, Xin; Xu, Yunyun; Liu, Cuiping; Hou, Jianquan; Zhang, Xueguang

    2015-02-01

    Myeloid-derived suppressor cells (MDSC) potently inhibit antitumor immune responses, and thereby promoti tumor progression and metastasis. However, the nature of human tumor-infiltrating MDSC remains poorly characterized. Here, we find B7-H3 is exclusively expressed on a subset of intratumoral CD14(+)HLA-DR(-/low) MDSC but absent from adjacent normal lung tissues of patients with non-small cell lung carcinoma (NSCLC). Cytokine analysis revealed that B7-H3(+)CD14(+)HLA-DR(-/low) MDSC (B7-H3(+)MDSC) produced higher levels of IL-10 and TNFα but lower levels of IL-1β and IL-6 when compared with B7-H3(-)CD14(+)HLA-DR(-/low) myeloid-derived suppressor cells (B7-H3(-)MDSC). In a murine lung cancer model, B7-H3(+)MDSCs were found only in the tumor microenvironment and their frequencies increased during tumor progression. Clinical data analysis indicated that a higher frequency of B7-H3(+)MDSCs was associated with reduced recurrence-free survival in patients with NSCLC. Taken together, we identify a novel subset of MDSCs within the tumor microenvironment that fosters tumor progression.

  14. Duration and severity of symptoms and levels of plasma interleukin-1 receptor antagonist, soluble tumor necrosis factor receptor, and adhesion molecules in patients with common cold treated with zinc acetate.

    PubMed

    Prasad, Ananda S; Beck, Frances W J; Bao, Bin; Snell, Diane; Fitzgerald, James T

    2008-03-15

    Zinc lozenges have been used for treatment of the common cold; however, the results remain controversial. Fifty ambulatory volunteers were recruited within 24 h of developing symptoms of the common cold for a randomized, double-blind, placebo-controlled trial of zinc. Participants took 1 lozenge containing 13.3 mg of zinc (as zinc acetate) or placebo every 2-3 h while awake. The subjective scores for common cold symptoms were recorded daily. Plasma zinc, soluble interleukin (IL)-1 receptor antagonist (sIL-1ra), soluble tumor necrosis factor receptor 1, soluble vascular endothelial cell adhesion molecule, and soluble intercellular adhesion molecule (sICAM)-1 were assayed on days 1 and 5. Compared with the placebo group, the zinc group had a shorter mean overall duration of cold (4.0 vs. 7.1 days; P < .0001) and shorter durations of cough (2.1 vs. 5.0 days; P < .0001) and nasal discharge (3.0 vs. 4.5 days, P = .02) Blinding of subjects was adequate, and adverse effects were comparable in the 2 groups. Symptom severity scores were decreased significantly in the zinc group. Mean changes in plasma levels of zinc, sIL-1ra, and ICAM-1 differed significantly between groups. Administration of zinc lozenges was associated with reduced duration and severity of cold symptoms. We related the improvement in cold symptoms to the antioxidant and anti-inflammatory properties of zinc.

  15. GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia

    PubMed Central

    2012-01-01

    Background Microglia are resident macrophage-like cells in the central nervous system (CNS) and cause innate immune responses via the LPS receptors, Toll-like receptor (TLR) 4 and CD14, in a variety of neuroinflammatory disorders including bacterial infection, Alzheimer’s disease, and amyotrophic lateral sclerosis. Granulocyte macrophage-colony stimulating factor (GM-CSF) activates microglia and induces inflammatory responses via binding to GM-CSF receptor complex composed of two different subunit GM-CSF receptor α (GM-CSFRα) and common β chain (βc). GM-CSF has been shown to be associated with neuroinflammatory responses in multiple sclerosis and Alzheimer’s disease. However, the mechanisms how GM-CSF promotes neuroinflammation still remain unclear. Methods Microglia were stimulated with 20 ng/ml GM-CSF and the levels of TLR4 and CD14 expression were evaluated by RT-PCR and flowcytometry. LPS binding was analyzed by flowcytometry. GM-CSF receptor complex was analyzed by immunocytechemistry. The levels of IL-1β, IL-6 and TNF-α in culture supernatant of GM-CSF-stimulated microglia and NF-κB nuclear translocation were determined by ELISA. Production of nitric oxide (NO) was measured by the Griess method. The levels of p-ERK1/2, ERK1/2, p-p38 and p38 were assessed by Western blotting. Statistically significant differences between experimental groups were determined by one-way ANOVA followed by Tukey test for multiple comparisons. Results GM-CSF receptor complex was expressed in microglia. GM-CSF enhanced TLR4 and CD14 expressions in microglia and subsequent LPS-binding to the cell surface. In addition, GM-CSF priming increased LPS-induced NF-κB nuclear translocation and production of IL-1β, IL-6, TNF-α and NO by microglia. GM-CSF upregulated the levels of p-ERK1/2 and p-p38, suggesting that induction of TLR4 and CD14 expression by GM-CSF was mediated through ERK1/2 and p38, respectively. Conclusions These results suggest that GM-CSF upregulates TLR4 and

  16. A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

    PubMed

    Tundup, Smanla; Srivastava, Leena; Norberg, Thomas; Watford, Wendy; Harn, Donald

    2015-01-01

    The milk pentasaccharide LNFPIII has therapeutic action for metabolic and autoimmune diseases and prolongs transplant survival in mice when presented as a neoglycoconjugate. Within LNFPIII is the Lewisx trisaccharide, expressed by many helminth parasites. In humans, LNFPIII is found in human milk and also known as stage-specific embryonic antigen-1. LNFPIII-NGC drives alternative activation of macrophages and dendritic cells via NFκB activation in a TLR4 dependent mechanism. However, the connection between LNFPIII-NGC activation of APCs, TLR4 signaling and subsequent MAP kinase signaling leading to anti-inflammatory activation of APCs remains unknown. In this study we determined that the innate receptor CD14 was essential for LNFPIII-NGC induction of both ERK and NFkB activation in APCs. Induction of ERK activation by LNFPIII-NGC was completely dependent on CD14/TLR4-Ras-Raf1/TPL2-MEK axis in bone marrow derived dendritic cells (BMDCs). In addition, LNFPIII-NGC preferentially induced the production of Th2 "favoring" chemokines CCL22 and matrix metalloprotease protein-9 in a CD14 dependent manner in BMDCs. In contrast, LNFPIII-NGC induces significantly lower levels of Th1 "favoring" chemokines, MIP1α, MIP1β and MIP-2 compared to levels in LPS stimulated cells. Interestingly, NGC of the identical human milk sugar LNnT, minus the alpha 1-3 linked fucose, failed to activate APCs via TLR4/MD2/CD14 receptor complex, suggesting that the alpha 1-3 linked fucose in LNFPIII and not on LNnT, is required for this process. Using specific chemical inhibitors of the MAPK pathway, we found that LNFPIII-NGC induction of CCL22, MMP9 and IL-10 production was dependent on ERK activation. Over all, this study suggests that LNFPIII-NGC utilizes CD14/TLR4-MAPK (ERK) axis in modulating APC activation to produce anti-inflammatory chemokines and cytokines in a manner distinct from that seen for the pro-inflammatory PAMP LPS. These pathways may explain the in vivo therapeutic effect of

  17. The Role of TLR2, TLR4 and CD14 Genetic Polymorphisms in Gastric Carcinogenesis: A Case-Control Study and Meta-Analysis

    PubMed Central

    Castaño-Rodríguez, Natalia; Kaakoush, Nadeem O.; Goh, Khean-Lee; Fock, Kwong Ming; Mitchell, Hazel M.

    2013-01-01

    Background In addition to Helicobacter pylori infection, host genetic factors contribute to gastric cancer (GC). Recognition of H. pylori is known to involve Toll-like receptors (TLR), which subsequently leads to activation of NF-κB. Thus, the overall aim of this study was to estimate for the first time the pooled effect size of polymorphisms in TLR2, TLR4 and CD14 on GC development through a meta-analysis. Methods A case-control study comprising 284 ethnic Chinese individuals (70 non-cardia GC cases and 214 functional dyspepsia controls) was conducted for the genotyping of TLR2 -196 to -174del, CD14 -260 C/T and TLR4 rs11536889 using PCR, RT-PCR and mass spectrometry. Case-control studies of TLR2, TLR4 and CD14 polymorphisms and GC were searched up to June 2012. Pooled odds ratios and 95% confidence intervals were obtained by means of the random effects model. Results In our ethnic Chinese case-control study, the TLR4 rs11536889 C allele increased the risk of GC (OR: 1.89, 95%CI: 1.23–2.92) while the CD14 -260 T allele was protective (OR: 0.62, 95%CI: 0.42–0.91). TLR2 -196 to -174 increased the risk of GC only in H. pylori-infected individuals (OR: 3.10, 95%CI: 1.27–7.60). In the meta-analysis, TLR4 Asp299Gly showed borderline results in the general analysis (pooled OR: 1.58, 95%CI: 0.98–2.60), nevertheless, stratified analysis by ethnicity showed that the mutant allele was a definitive risk factor for GC in Western populations (pooled OR: 1.87, 95%CI: 1.31–2.65). There was a potential association between the TLR2 -196 to -174 deletion allele and GC in Japanese (pooled OR: 1.18, 95%CI: 0.96–1.45). TLR4 Thr399Ile did not provide significant results. Conclusions TLR4 rs11536889 and CD14 -260 C/T are associated with non-cardia GC in Chinese. Based on our meta-analysis, the TLR signalling pathway is involved in gastric carcinogenesis, TLR4 Asp299Gly and TLR2 -196 to -174del showing associations with GC in an ethnic-specific manner. PMID:23565226

  18. Soluble T Cell Immunoglobulin Mucin Domain 3 Is Shed from CD8+ T Cells by the Sheddase ADAM10, Is Increased in Plasma during Untreated HIV Infection, and Correlates with HIV Disease Progression

    PubMed Central

    Douglas-Vail, Matthew B.; Rahman, A. K. M. Nur-ur; Medcalf, Karyn E.; Xie, Irene Y.; Chew, Glen M.; Tandon, Ravi; Lanteri, Marion C.; Norris, Philip J.; Deeks, Steven G.; Ndhlovu, Lishomwa C.

    2015-01-01

    ABSTRACT Chronic HIV infection results in a loss of HIV-specific CD8+ T cell effector function, termed “exhaustion,” which is mediated, in part, by the membrane coinhibitory receptor T cell immunoglobulin mucin domain-3 (Tim-3). Like many other receptors, a soluble form of this protein has been described in human blood plasma. However, soluble Tim-3 (sTim-3) is poorly characterized, and its role in HIV disease is unknown. Here, we show that Tim-3 is shed from the surface of responding CD8+ T cells by the matrix metalloproteinase ADAM10, producing a soluble form of the coinhibitory receptor. Despite previous reports in the mouse model, no alternatively spliced, soluble form of Tim-3 was observed in humans. Shed sTim-3 was found in human plasma and was significantly elevated during early and chronic untreated HIV infection, but it was not found differentially modulated in highly active antiretroviral therapy (HAART)-treated HIV-infected subjects or in elite controllers compared to HIV-uninfected subjects. Plasma sTim-3 levels were positively correlated with HIV load and negatively correlated with CD4 counts. Thus, plasma sTim-3 shedding correlated with HIV disease progression. Despite these correlations, we found that shedding Tim-3 did not improve the function of CD8+ T cells in terms of gamma interferon production or prevent their apoptosis through galectin-9. Further characterization studies of sTim-3 function are needed to understand the contribution of sTim-3 in HIV disease pathogenesis, with implications for novel therapeutic interventions. IMPORTANCE Despite the overall success of HAART in slowing the progression to AIDS in HIV-infected subjects, chronic immune activation and T cell exhaustion contribute to the eventual deterioration of the immune system. Understanding these processes will aid in the development of interventions and therapeutics to be used in combination with HAART to slow or reverse this deterioration. Here, we show that a soluble form of

  19. The relationship between levels of plasma-soluble urokinase plasminogen activator receptor (suPAR) and presence of migraine attack and aura.

    PubMed

    Yılmaz, Nigar; Yılmaz, Mustafa; Sirin, Burcu; Yılmaztekin, Sureyya; Kutlu, Gülnihal

    2017-10-01

    Migraine is one of the most common types of pain associated with sterile inflammatory conditions. Soluble urokinase plasminogen activator receptor (suPAR) is a potential novel inflammatory marker. We aim to determine the association between serum values of suPAR, procalcitonin, fibrinogen, and high-sensitivity C-reactive protein (hs-CRP) and migraine disease characteristics. The study involved a total of 60 migraine patients (33 patients in the interictal period, 27 patients in the attack period) and 30 healthy individuals. The serum values of suPAR were found to be significantly higher in migraine patients in the attack period than in migraine patients in the interictal period, and in healthy individuals (p < .01 for both). In addition, levels of suPAR were determined to be higher in migraine with aura patients than in migraine without aura patients. When we subdivided migraine patients according to frequency of attack (attacks/month), significant differences were found between the suPAR and procalcitonin levels (measured during the attack period) of those in the frequent-attack group (4-5 or more) versus those in the less frequent attack group (less than 4). Serum levels of procalcitonin were shown to be significantly higher in migraine patients during the attack period compared with migraine patients in the interictal period and in control subjects (p = .001 for both). Significant differences were found between plasma levels of fibrinogen in migraine patients versus control subjects (p < .01). No statistically significant difference was found between levels of hs-CRP in migraine patients versus the control group. These findings may show that presenting a high level of suPAR in migraine patients with attack and aura results to predisposition to occurring on the symptoms and that high levels of suPAR, procalcitonin and fibrinogen in patients with migraine result in neurogenic inflammation during migraine headaches.

  20. Plasma levels of soluble programmed death ligand-1 may be associated with overall survival in nonsmall cell lung cancer patients receiving thoracic radiotherapy

    PubMed Central

    Zhao, Jing; Zhang, Peng; Wang, Jianhua; Xi, Qingsong; Zhao, Xueqi; Ji, Minghua; Hu, Guangyuan

    2017-01-01

    Abstract Immune-checkpoint signaling plays an important role in immunosuppression of tumors. We aimed to investigate the association of soluble programmed death-ligand 1 (sPD-L1) level in plasma with overall survival (OS) in locally advanced or inoperable nonsmall-cell lung cancer (NSCLC) patients treated with thoracic radiotherapy (TRT). We used ELISA to evaluate the sPD-L1 levels at diagnosis and during TRT in 126 clinically inoperable NSCLC patients. OS rates were followed up and recorded. SPSS software and GraphPad Prism 5 were used for statistics. In this study, the average sPD-L1 levels at baseline, week 2, and week 4 during TRT and post-TRT were 107.2, 51.3, 65.4, and 111.1 pg/mL, respectively. Levels of sPD-L1 at week 2 and week 4 were significantly less than at baseline, with both P values < 0.001. Using 96.5 pg/mL as the cutoff, patients with lower baseline sPD-L1 level had longer OS than those with higher sPD-L1 level (27.8 months vs 15.5 months, P = 0.005). Using multivariate analysis, the following factors were significantly associated with longer OS: female, adenocarcinoma, higher TRT dose, and lower baseline sPD-L1 level. Patients with both characteristics of lower baseline sPD-L1 level and higher TRT dose (BED10 ≥84 Gy) had the longest OS. To conclude, the lower baseline sPD-L1 level was significantly associated with longer OS in NSCLC patients treated with TRT, which may serve as an independent biomarker and needs further clinical study. PMID:28207525

  1. Serum Soluble Urokinase-Type Plasminogen Activator Receptor Is Associated with Low Left Ventricular Ejection Fraction and Elevated Plasma Brain-Type Natriuretic Peptide Level

    PubMed Central

    Fujita, Shu-ichi; Tanaka, Suguru; Maeda, Daichi; Morita, Hideaki; Fujisaka, Tomohiro; Takeda, Yoshihiro; Ito, Takahide; Ishizaka, Nobukazu

    2017-01-01

    Background Recent studies have suggested that soluble urokinase plasminogen activator receptor (suPAR), a biomarker of subclinical levels of inflammation, is significantly correlated with cardiovascular events. Purpose We investigated the association between suPAR and left ventricular ejection fraction (LVEF), left ventricular mass index (LVMI), and plasma B-type natriuretic peptide (BNP) among cardiac inpatients. Methods and Results In total, 242 patients (mean age 71.3 ± 9.8 years; 70 women) admitted to the cardiology department were enrolled in the study. suPAR was significantly correlated with LVEF (R = -0.24, P<0.001), LVMI (R = 0.16, P = 0.014) and BNP (R = 0.46, P<0.001). In logistic regression analysis, the highest suPAR tertile (> 3236 pg/mL) was associated with low LVEF (< 50%) and elevated BNP (> 300 pg/mL) with an odds ratio of 3.84 (95% confidence interval [CI], 1.22–12.1) and 5.36 (95% CI, 1.32–21.8), respectively, after adjusting for age, sex, log-transformed estimated glomerular filtration rate (log(eGFR)), C-reactive protein, and diuretic use. The association between suPAR and LVMI was not statistically significant. In multivariate receiver operating characteristic analysis, addition of log(suPAR) to the combination of age, sex, log(eGFR) and CRP incrementally improved the prediction of low LVEF (area under the curve [AUC], 0.827 to 0.852, P = 0.046) and BNP ≥ 300 pg/mL (AUC, 0.869 to 0.906; P = 0.029). Conclusions suPAR was associated with low LVEF and elevated BNP, but not with left ventricular hypertrophy, independent of CRP, renal function, and diuretic use among cardiac inpatients who were not undergoing chronic hemodialysis. PMID:28135310

  2. Combined Inhibition of Complement and CD14 Attenuates Bacteria-Induced Inflammation in Human Whole Blood More Efficiently Than Antagonizing the Toll-like Receptor 4–MD2 Complex

    PubMed Central

    Gustavsen, Alice; Nymo, Stig; Landsem, Anne; Christiansen, Dorte; Ryan, Liv; Husebye, Harald; Lau, Corinna; Pischke, Søren E.; Lambris, John D.; Espevik, Terje; Mollnes, Tom E.

    2016-01-01

    Background. Single inhibition of the Toll-like receptor 4 (TLR4)–MD2 complex failed in treatment of sepsis. CD14 is a coreceptor for several TLRs, including TLR4 and TLR2. The aim of this study was to investigate the effect of single TLR4-MD2 inhibition by using eritoran, compared with the effect of CD14 inhibition alone and combined with the C3 complement inhibitor compstatin (Cp40), on the bacteria-induced inflammatory response in human whole blood. Methods. Cytokines were measured by multiplex technology, and leukocyte activation markers CD11b and CD35 were measured by flow cytometry. Results. Lipopolysaccharide (LPS)–induced inflammatory markers were efficiently abolished by both anti-CD14 and eritoran. Anti-CD14 was significantly more effective than eritoran in inhibiting LPS-binding to HEK-293E cells transfected with CD14 and Escherichia coli–induced upregulation of monocyte activation markers (P < .01). Combining Cp40 with anti-CD14 was significantly more effective than combining Cp40 with eritoran in reducing E. coli–induced interleukin 6 (P < .05) and monocyte activation markers induced by both E. coli (P < .001) and Staphylococcus aureus (P < .01). Combining CP40 with anti-CD14 was more efficient than eritoran alone for 18 of 20 bacteria-induced inflammatory responses (mean P < .0001). Conclusions. Whole bacteria–induced inflammation was inhibited more efficiently by anti-CD14 than by eritoran, particularly when combined with complement inhibition. Combined CD14 and complement inhibition may prove a promising treatment strategy for bacterial sepsis. PMID:26977050

  3. Resistin Gene Expression is Downregulated in CD4(+) T Helper Lymphocytes and CD14(+) Monocytes in Rheumatoid Arthritis Responding to TNF-α Inhibition.

    PubMed

    Nagaev, I; Andersen, M; Olesen, M K; Nagaeva, O; Wikberg, J; Mincheva-Nilsson, L; Andersen, G N

    2016-10-01

    Rheumatoid arthritis (RA) is caused by complex interactions between immune cells and sustained by Th1 response cytokines. Resistin [resistance to insulin; (RETN)] is an inflammatory cytokine, first discovered in murine adipocytes. In man, RETN is mainly secreted by monocytes. The distinct role of RETN in the immune reaction is uncertain; however, RETN has pro-inflammatory, pro-fibrotic and possibly tolerogenic properties. The aim was to assess the reaction of RETN gene expression to TNF-α inhibition (I) in pathogenetic immune cell subsets in RA, in the context of Th1, inflammatory and regulatory cytokine gene expressions. Accordingly, we measured RETN, IFN-γ, TNF-β, IL-1β, TNF-α, TGF-β and IL-10 gene expressions in CD14(+) monocytes, CD4(+) T helper (Th) lymphocytes (ly), CD8(+) T cytotoxic (Tc) ly and CD19(+) B ly in active RA before and 3 months after start of TNF-αI. Leucocyte subsets were separated by specific monoclonal antibody-covered beads, RNA extracted and levels of RETN, Th1 response, inflammatory and regulatory cytokine mRNAs measured by quantitative reverse transcription-polymerase chain reaction technique. We found that TNF-αI caused a significant downregulation of RETN gene expression in CD14(+) monocytes and CD4(+) Th ly and was unchanged in CD8(+) Tc ly and CD19(+) B ly. Both in active RA and during TNF-αI, RETN mRNA levels were significantly higher in CD14(+) monocytes than in all other examined cell types. In monocytes, fold change in RETN and TGF-β gene expressions upon TNF-αI correlated significantly. Our findings indicate that RETN has pro-inflammatory as well as proresolving roles in active RA.

  4. The interaction between farming/rural environment and TLR2, TLR4, TLR6 and CD14 genetic polymorphisms in relation to early- and late-onset asthma

    PubMed Central

    Lau, Melisa Y. Z.; Dharmage, Shyamali C.; Burgess, John A.; Win, Aung K.; Lowe, Adrian J.; Lodge, Caroline; Perret, Jennifer; Hui, Jennie; Thomas, Paul S.; Morrison, Stephen; Giles, Graham G.; Hopper, John; Abramson, Michael J.; Walters, E. Haydn; Matheson, Melanie C.

    2017-01-01

    Asthma phenotypes based on age-of-onset may be differently influenced by the interaction between variation in toll-like receptor (TLR)/CD14 genes and environmental microbes. We examined the associations between single-nucleotide polymorphisms (SNP) in the TLR/CD14 genes and asthma, and their interaction with proxies of microbial exposure (childhood farm exposure and childhood rural environment). Ten SNPs in four genes (TLR2, TLR4, TLR6, CD14) were genotyped for 1,116 participants from the Tasmanian Longitudinal Health Study (TAHS). Using prospectively collected information, asthma was classified as never, early- (before 13 years) or late-onset (after 13 years). Information on childhood farm exposure/childhood rural environment was collected at baseline. Those with early-onset asthma were more likely to be males, had a family history of allergy and a personal history of childhood atopy. We found significant interaction between TLR6 SNPs and childhood farm exposure. For those with childhood farm exposure, carriers of the TLR6-rs1039559 T-allele (p-interaction = 0.009) and TLR6-rs5743810 C-allele (p-interaction = 0.02) were associated with lower risk of early-onset asthma. We suggest the findings to be interpreted as hypothesis-generating as the interaction effect did not withstand correction for multiple testing. In this large, population-based longitudinal study, we found that the risk of early- and late-onset asthma is differently influenced by the interaction between childhood farming exposure and genetic variations. PMID:28262750

  5. Impaired Functions of Macrophage from Cystic Fibrosis Patients: CD11b, TLR-5 Decrease and sCD14, Inflammatory Cytokines Increase

    PubMed Central

    Simonin-Le Jeune, Karin; Le Jeune, André; Jouneau, Stéphane; Belleguic, Chantal; Roux, Pierre-François; Jaguin, Marie; Dimanche-Boitre, Marie-Thérèse; Lecureur, Valérie; Leclercq, Caroline; Desrues, Benoît; Brinchault, Graziella; Gangneux, Jean-Pierre; Martin-Chouly, Corinne

    2013-01-01

    Background Early in life, cystic fibrosis (CF) patients are infected with microorganisms. The role of macrophages has largely been underestimated in literature, whereas the focus being mostly on neutrophils and epithelial cells. Macrophages may however play a significant role in the initiating stages of this disease, via an inability to act as a suppressor cell. Yet macrophage dysfunction may be the first step in cascade of events leading to chronic inflammation/infection in CF. Moreover, reports have suggested that CFTR contribute to altered inflammatory response in CF by modification of normal macrophage functions. Objectives In order to highlight possible intrinsic macrophage defects due to impaired CFTR, we have studied inflammatory cytokines secretions, recognition of pathogens and phagocytosis in peripheral blood monocyte-derived macrophages from stable adult CF patients and healthy subjects (non-CF). Results In CF macrophage supernatants, concentrations of sCD14, IL-1β, IL-6, TNF-α and IL-10 were strongly raised. Furthermore expression of CD11b and TLR-5 were sorely decreased on CF macrophages. Beside, no difference was observed for mCD14, CD16, CD64, TLR-4 and TLR1/TLR-2 expressions. Moreover, a strong inhibition of phagocytosis was observed for CF macrophages. Elsewhere CFTR inhibition in non-CF macrophages also led to alterations of phagocytosis function as well as CD11b expression. Conclusions Altogether, these findings demonstrate excessive inflammation in CF macrophages, characterized by overproduction of sCD14 and inflammatory cytokines, with decreased expression of CD11b and TLR-5, and impaired phagocytosis. This leads to altered clearance of pathogens and non-resolution of infection by CF macrophages, thereby inducing an exaggerated pro-inflammatory response. PMID:24098711

  6. Tumor-infiltrating CD14-positive myeloid cells and CD8-positive T-cells prolong survival in patients with cervical carcinoma.

    PubMed

    de Vos van Steenwijk, P J; Ramwadhdoebe, T H; Goedemans, R; Doorduijn, E M; van Ham, J J; Gorter, A; van Hall, T; Kuijjer, M L; van Poelgeest, M I E; van der Burg, S H; Jordanova, E S

    2013-12-15

    One of the hallmarks of cancer is the influx of myeloid cells. In our study, we investigated the constitution of tumor-infiltrating myeloid cells and their relationship to other tumor-infiltrating immune cells, tumor characteristics and the disease-specific survival of patients with cervical cancer (CxCa). Triple-color immunofluorescence confocal microscopy was used to locate, identify and quantify macrophages (CD14), their maturation status (CD33) and their polarization (CD163) in a cohort of 86 patients with cervical carcinoma. Quantification of the numbers of myeloid cells revealed that a strong intraepithelial infiltration of CD14+ cells, and more specifically the population of CD14+CD33-CD163- matured M1 macrophages, is associated with a large influx of intraepithelial T lymphocytes (p = 0.008), improved disease-specific survival (p = 0.007) and forms an independent prognostic factor for survival (p = 0.033). The intraepithelial CD8+ T-cell and regulatory T-cell (Treg) ratio also forms an independent prognostic factor (p = 0.010) and combination of these two factors reveals a further increased benefit in survival for patients whose tumor displays a dense infiltration with intraepithelial matured M1 macrophages and a high CD8 T-cell/Treg ratio, indicating that both populations of immune cells simultaneously improve survival. Subsequently, we made a heatmap including all known immune parameters for these patients, whereby we were able to identify different immune signatures in CxCa. These results indicate that reinforcement and activation of the intratumoral M1 macrophages may form an attractive immunotherapeutic option in CxCa.

  7. Association between CD14 SNP -159 C/T and gastric cancer: an independent case–control study and an updated meta-analysis

    PubMed Central

    Gong, Ai-Min; Li, Xin-Yuan; Xie, Yi-Qiang; Jia, Zhan-Dong; Li, Yuan-Xin; Zou, Yong-Yan; Xu, Chang-Qing; Wang, Zhen-Yu

    2016-01-01

    Purpose The association between CD14 -159C/T polymorphism and the susceptibility to gastric cancer (GC) has been reported. However, the results were inconclusive. In the present study, a case–control study and a meta-analysis were performed to assess the possible association between -159C/T in the CD14 gene and GC risk. Patients and methods Relevant studies were searched in several databases including PubMed, Web of Science, EMBASE, Chinese National Knowledge Infrastructure database, and Wanfang database (last search was performed on December 30, 2015). In addition, a case–control study involving 164 GC cases and 169 controls was also performed in the analysis. Statistical analysis was performed by the software Revman5.3. Results A total of ten published studies and the present case–control study involving 2,844 GC and 3,983 controls were included for the meta-analysis. The analysis result indicated that the T allele of CD14 -159C/T polymorphism did not confer risk for GC (in our study: [P=0.93]; in the meta-analysis: T vs 2N odds ratio =1.28 and 95% confidence interval (CI) =0.95–1.24, [P=0.24]). However, we found a significant association in the recessive model (in our study: TT vs TC+CC [P=0.04]; in the meta-analysis: TT vs TC+CC odds ratio =1.12 and 95% CI =1.01–1.26, [P=0.04]). Furthermore, a subgroup analysis by ethnicity showed that TT genotype was significantly associated with GC in Asian (odds ratio =1.17 and 95% CI =1.02–1.34, [P=0.02]) but not in Caucasian. Conclusion Our results highlight the TT genotype of CD14 -159C/T as a genetic susceptibility factor for gastric cancer, particularly, in Asians and population-based controls. PMID:27486336

  8. VB-201, an oxidized phospholipid small molecule, inhibits CD14- and Toll-like receptor-2-dependent innate cell activation and constrains atherosclerosis

    PubMed Central

    Mendel, I; Feige, E; Yacov, N; Salem, Y; Levi, I; Propheta-Meiran, O; Shoham, A; Ishai, E; George, J; Harats, D; Breitbart, E

    2014-01-01

    Atherosclerosis is an inflammatory disease of the vascular wall. Activated monocytes and dendritic cells (DC) in the intima layer of the vasculature promote atherogenesis. Toll-like receptor (TLR)-2 and TLR-4, which are predominantly expressed on these cells and mediate their activation, are essential for atherosclerosis development. In this study we demonstrate that VB-201, an oxidized phospholipid (Ox-PL) small molecule, inhibits TLR signalling restricted to TLR-2 and TLR-4 in human and mouse monocytes and DC. Mechanistically, we show that VB-201 binds directly to TLR-2 and CD14, the TLR-4 co-receptor, to impair downstream cues and cytokine production. In a rabbit model, oral administration of VB-201 constrained atherosclerosis progression. This effect was not due to reduced cholesterol abundance, as hyperlipidaemia was sustained. We suggest that VB-201 may counter inflammation where TLR-2 and/or CD14 complicity is essential, and is therefore beneficial for the treatment of atherosclerosis. PMID:24116867

  9. Early Dynamics of Cerebrospinal CD14+ Monocytes and CD15+ Granulocytes in Patients after Severe Traumatic Brain Injury: A Cohort Study

    PubMed Central

    Postl, Lukas Kurt; Bogner, Viktoria; Beirer, Marc; Kanz, Karl Georg; Egginger, Christoph; Schmitt-Sody, Markus; Biberthaler, Peter; Kirchhoff, Chlodwig

    2015-01-01

    In traumatic brain injury (TBI) the analysis of neuroinflammatory mechanisms gained increasing interest. In this context certain immunocompetent cells might play an important role. Interestingly, in the actual literature there exist only a few studies focusing on the role of monocytes and granulocytes in TBI patients. In this regard it has recently reported that the choroid plexus represents an early, selective barrier for leukocytes after brain injury. Therefore the aim of this study was to evaluate the very early dynamics of CD14+ monocytes and CD15+ granulocyte in CSF of patients following severe TBI with regard to the integrity of the BBB. Cytometric flow analysis was performed to analyze the CD14+ monocyte and CD15+ granulocyte population in CSF of TBI patients. The ratio of CSF and serum albumin as a measure for the BBB's integrity was assessed in parallel. CSF samples of patients receiving lumbar puncture for elective surgery were obtained as controls. Overall 15 patients following severe TBI were enrolled. 10 patients were examined as controls. In patients, the monocyte population as well as the granulocyte population was significantly increased within 72 hours after TBI. The BBB's integrity did not have a significant influence on the cell count in the CSF. PMID:26568661

  10. Immature immunosuppressive CD14+HLA-DR-/low cells in melanoma patients are Stat3hi and overexpress CD80, CD83, and DC-sign.

    PubMed

    Poschke, Isabel; Mougiakakos, Dimitrios; Hansson, Johan; Masucci, Giuseppe V; Kiessling, Rolf

    2010-06-01

    Myeloid-derived suppressor cells (MDSC) have emerged as key immune modulators in various tumor models and human malignancies, but their characteristics in humans remain to be unequivocally defined. In this study, we have examined circulating CD14(+)HLA-DR(-/low) MDSC in 34 advanced malignant melanoma (MM) patients. Their frequency is significantly increased and associated with disease activity. Contrary to the common notion that MDSC are a heterogeneous population of exclusively immature cells, we find the coexpression of markers associated with mature phenotype. We show for the first time the overexpression of CD80, CD83, and DC-Sign in human MDSC. Further, increased levels of signal transducer and activator of transcription 3 (Stat3), an important regulator in MDSC development and function, were noted in MM-MDSC. Stat3 was altered toward an active, phosphorylated state in the HLA-DR(-) population of CD14(+) cells and was more reactive to activating stimuli in patients. Importantly, inhibition of Stat3 abolished their suppressive activity almost completely. The described MM-MDSC use arginase in conjunction with other yet undefined mechanisms to suppress CD4(+) and CD8(+) T cells. Several observations suggest a redox imbalance in MDSC and indicate an important role of Stat3-dependent oxidative stress in MDSC-mediated T-cell suppression. These results emphasize the diversity of MDSC in human cancer and provide potential targets for therapeutic interventions.

  11. The antiviral activity of poly-γ-glutamic acid, a polypeptide secreted by Bacillus sp., through induction of CD14-dependent type I interferon responses.

    PubMed

    Lee, Wooseong; Lee, Seung-Hoon; Ahn, Dae-Gyun; Cho, Hee; Sung, Moon-Hee; Han, Seung Hyun; Oh, Jong-Won

    2013-12-01

    Poly-γ-glutamic acid (γ-PGA) is an anionic polypeptide secreted by Bacillus sp. that has been shown to activate immune cells through interactions with toll-like receptor 4 (TLR4). However, its ability to induce the type I interferon (IFN) response has not yet been characterized. Here, we demonstrate that γ-PGA induces type I IFN signaling pathway via the TLR4 signaling pathway. The induction required both myeloid differentiation factor 2 (MD2) and the pattern-recognition receptor CD14, which are two TLR4-associated accessory proteins. The γ-PGA with high molecular weights (2000 and 5000 kDa) was able to activate the subsequent signals through TLR4/MD2 to result in dimerization of IRF-3, a transcription factor required for IFN gene expression, leading to increases in mRNA levels of the type I IFN-response genes, 2'-5' OAS and ISG56. Moreover, γ-PGA (2000 kDa) displayed an antiviral activity against SARS coronavirus and hepatitis C virus. Our results identify high-molecular weight γ-PGA as a TLR4 ligand and demonstrate that γ-PGA requires both CD14 and MD2 for the activation of type I IFN responses. Our results suggest that the microbial biopolymer γ-PGA may have therapeutic potential against a broad range of viruses sensitive to type I IFNs.

  12. DNA-Containing Immunocomplexes Promote Inflammasome Assembly and Release of Pyrogenic Cytokines by CD14+ CD16+ CD64high CD32low Inflammatory Monocytes from Malaria Patients

    PubMed Central

    Hirako, Isabella C.; Gallego-Marin, Carolina; Ataide, Marco A.; Andrade, Warrison A.; Gravina, Humberto; Rocha, Bruno C.; de Oliveira, Rosane B.; Pereira, Dhelio B.; Vinetz, Joseph; Diamond, Betty; Ram, Sanjay; Golenbock, Douglas T.

    2015-01-01

    ABSTRACT High levels of circulating immunocomplexes (ICs) are found in patients with either infectious or sterile inflammation. We report that patients with either Plasmodium falciparum or Plasmodium vivax malaria have increased levels of circulating anti-DNA antibodies and ICs containing parasite DNA. Upon stimulation with malaria-induced ICs, monocytes express an NF-κB transcriptional signature. The main source of IC-induced proinflammatory cytokines (i.e., tumor necrosis factor alpha [TNF-α] and interleukin-1β [IL-1β])in peripheral blood mononuclear cells from acute malaria patients was found to be a CD14+ CD16 (FcγRIIIA)+ CD64 (FcγRI)high CD32 (FcγRIIB)low monocyte subset. Monocytes from convalescent patients were predominantly of the classical phenotype (CD14+ CD16−) that produces high levels of IL-10 and lower levels of TNF-α and IL-1β in response to ICs. Finally, we report a novel role for the proinflammatory activity of ICs by demonstrating their ability to induce inflammasome assembly and caspase-1 activation in human monocytes. These findings illuminate our understanding of the pathogenic role of ICs and monocyte subsets and may be relevant for future development of immunity-based interventions with broad applications to systemic inflammatory diseases. PMID:26578679

  13. Lipopolysaccharide induces alveolar macrophage necrosis via CD14 and the P2x7 receptor leading to Interleukin-1α release

    PubMed Central

    Dagvadorj, Jargalsaikhan; Shimada, Kenichi; Chen, Shuang; Jones, Heather D.; Tumurkhuu, Gantsetseg; Zhang, Wenxuan; Wawrowsky, Kolja A.; Crother, Timothy R.; Arditi, Moshe

    2015-01-01

    SUMMARY Acute lung injury (ALI) remains a serious health issue with little improvement in our understanding of the pathophysiology and therapeutic approaches. We investigated the mechanism that lipopolysaccharide (LPS) induces early neutrophil recruitment to lungs and increases pulmonary vascular permeability during ALI. Intratracheal LPS induced release of pro-interleukin-1α (IL-1α) from necrotic alveolar macrophages (AM), which activated endothelial cells (EC) to induce vascular leakage via loss of vascular endothelial (VE)-cadherin. LPS triggered the AM purinergic receptor P2X7(R) to induce Ca2+ influx and ATP depletion, which led to necrosis. P2X7R deficiency significantly reduced necrotic death of AM and release of pro-IL-1α into the lung. CD14 was required for LPS binding to P2X7R, as CD14 neutralization significantly diminished LPS induced necrotic death of AM and pro-IL-1α release. These results demonstrate a key role for pro-IL-1α from necrotic alveolar macrophages in LPS-mediated ALI, as a critical initiator of increased vascular permeability and early neutrophil infiltration. PMID:25862090

  14. Lipopolysaccharide Induces Alveolar Macrophage Necrosis via CD14 and the P2X7 Receptor Leading to Interleukin-1α Release.

    PubMed

    Dagvadorj, Jargalsaikhan; Shimada, Kenichi; Chen, Shuang; Jones, Heather D; Tumurkhuu, Gantsetseg; Zhang, Wenxuan; Wawrowsky, Kolja A; Crother, Timothy R; Arditi, Moshe

    2015-04-21

    Acute lung injury (ALI) remains a serious health issue with little improvement in our understanding of the pathophysiology and therapeutic approaches. We investigated the mechanism that lipopolysaccharide (LPS) induces early neutrophil recruitment to lungs and increases pulmonary vascular permeability during ALI. Intratracheal LPS induced release of pro-interleukin-1α (IL-1α) from necrotic alveolar macrophages (AM), which activated endothelial cells (EC) to induce vascular leakage via loss of vascular endothelial (VE)-cadherin. LPS triggered the AM purinergic receptor P2X7(R) to induce Ca(2+) influx and ATP depletion, which led to necrosis. P2X7R deficiency significantly reduced necrotic death of AM and release of pro-IL-1α into the lung. CD14 was required for LPS binding to P2X7R, as CD14 neutralization significantly diminished LPS induced necrotic death of AM and pro-IL-1α release. These results demonstrate a key role for pro-IL-1α from necrotic alveolar macrophages in LPS-mediated ALI, as a critical initiator of increased vascular permeability and early neutrophil infiltration.

  15. Mechanisms of HIV entry into the CNS: increased sensitivity of HIV infected CD14+CD16+ monocytes to CCL2 and key roles of CCR2, JAM-A, and ALCAM in diapedesis.

    PubMed

    Williams, Dionna W; Calderon, Tina M; Lopez, Lillie; Carvallo-Torres, Loreto; Gaskill, Peter J; Eugenin, Eliseo A; Morgello, Susan; Berman, Joan W

    2013-01-01

    As HIV infected individuals live longer, the prevalence of HIV associated neurocognitive disorders is increasing, despite successful antiretroviral therapy. CD14(+)CD16(+) monocytes are critical to the neuropathogenesis of HIV as they promote viral seeding of the brain and establish neuroinflammation. The mechanisms by which HIV infected and uninfected monocytes cross the blood brain barrier and enter the central nervous system are not fully understood. We determined that HIV infection of CD14(+)CD16(+) monocytes resulted in their highly increased transmigration across the blood brain barrier in response to CCL2 as compared to uninfected cells, which did not occur in the absence of the chemokine. This exuberant transmigration of HIV infected monocytes was due, at least in part, to increased CCR2 and significantly heightened sensitivity to CCL2. The entry of HIV infected and uninfected CD14(+)CD16(+) monocytes into the brain was facilitated by significantly increased surface JAM-A, ALCAM, CD99, and PECAM-1, as compared to CD14(+) cells that are CD16 negative. Upon HIV infection, there was an additional increase in surface JAM-A and ALCAM on CD14(+)CD16(+) monocytes isolated from some individuals. Antibodies to ALCAM and JAM-A inhibited the transmigration of both HIV infected and uninfected CD14(+)CD16(+) monocytes across the BBB, demonstrating their importance in facilitating monocyte transmigration and entry into the brain parenchyma. Targeting CCR2, JAM-A, and ALCAM present on CD14(+)CD16(+) monocytes that preferentially infiltrate the CNS represents a therapeutic strategy to reduce viral seeding of the brain as well as the ongoing neuroinflammation that occurs during HIV pathogenesis.

  16. Gene expression profiling of Gram-negative bacteria-induced inflammation in human whole blood: The role of complement and CD14-mediated innate immune response.

    PubMed

    Lau, Corinna; Olstad, Ole Kristoffer; Holden, Marit; Nygård, Ståle; Fure, Hilde; Lappegård, Knut Tore; Brekke, Ole-Lars; Espevik, Terje; Hovig, Eivind; Mollnes, Tom Eirik

    2015-09-01

    Non-sterile pathogen-induced sepsis and sterile inflammation like in trauma or ischemia-reperfusion injury may both coincide with the life threatening systemic inflammatory response syndrome and multi-organ failure. Consequently, there is an urgent need for specific biomarkers in order to distinguish sepsis from sterile conditions. The overall aim of this study was to uncover putative sepsis biomarkers and biomarker pathways, as well as to test the efficacy of combined inhibition of innate immunity key players complement and Toll-like receptor co-receptor CD14 as a possible therapeutic regimen for sepsis. We performed whole blood gene expression analyses using microarray in order to profile Gram-negative bacteria-induced inflammatory responses in an ex vivo human whole blood model. The experiments were performed in the presence or absence of inhibitors of complement proteins (C3 and CD88 (C5a receptor 1)) and CD14, alone or in combination. In addition, we used blood from a C5-deficient donor. Anti-coagulated whole blood was challenged with heat-inactivated Escherichia coli for 2 h, total RNA was isolated and microarray analyses were performed on the Affymetrix GeneChip Gene 1.0 ST Array platform. The initial experiments were performed in duplicates using blood from two healthy donors. C5-deficiency is very rare, and only one donor could be recruited. In order to increase statistical power, a technical replicate of the C5-deficient samples was run. Subsequently, log2-transformed intensities were processed by robust multichip analysis and filtered using a threshold of four. In total, 73 microarray chips were run and analyzed. The normalized and filtered raw data have been deposited in NCBI's Gene Expression Omnibus (GEO) and are accessible with GEO Series accession number GSE55537. Linear models for microarray data were applied to estimate fold changes between data sets and the respective multiple testing adjusted p-values (FDR q-values). The interpretation of the

  17. Immune profiling of BALB/C and C57BL/6 mice reveals a correlation between Ureaplasma parvum-Induced fetal inflammatory response syndrome-like pathology and increased placental expression of TLR2 and CD14.

    PubMed

    Allam, Ayman B; von Chamier, Maria; Brown, Mary B; Reyes, Leticia

    2014-03-01

    Both BALB/c and C57BL/6 mice are susceptible to intrauterine infection with Ureaplasma parvum, but only protypical TH2/M2 BALB/c mice develop severe chorioamnionitis, fetal infection, and fetal inflammatory response syndrome-like (FIRS) pathology. Microscopy, gene expression analysis, and ELISA were used to identify placental innate immune responses relevant to macrophage polarity, severe chorioamnionitis, and fetal infection. Both mouse strains exhibited a pro-M2 cytokine profile at the maternal/fetal interface. In BALB/c mice, expression of CD14 and TLRs 1, 2, 6 was increased in infected placentas; TLR2 and CD14 were localized to neutrophils. Increased TLR2/CD14 was also observed in BALB/c syncytiotrophoblasts in tissues with pathological evidence of FIRS. In contrast, expression in C57BL/6 placentas was either unchanged or down-regulated. Our findings show a link between increased syncytiotrophoblast expression of CD14/TLR2 and FIRS-like pathology in BALB/c mice. Functional studies are required to determine if CD14 is contributing to fetal morbidity during chorioamnionitis. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Involvement of lipopolysaccharide binding protein, CD14, and Toll-like receptors in the initiation of innate immune responses by Treponema glycolipids.

    PubMed

    Schröder, N W; Opitz, B; Lamping, N; Michelsen, K S; Zähringer, U; Göbel, U B; Schumann, R R

    2000-09-01

    Culture supernatants from Treponema maltophilum associated with periodontitis in humans and Treponema brennaborense found in a bovine cattle disease accompanied with cachexia caused a dose-dependent TNF-alpha synthesis in human monocytes increasing with culture time. This activity could be reduced significantly by blocking the CD14-part of the LPS receptor using the My 4 mAb and by polymyxin B. In the murine macrophage cell line RAW 264.7, Treponema culture supernatants induced TNF-alpha secretion in a LPS binding protein (LBP)-dependent fashion. To enrich for active compounds, supernatants were extracted with butanol, while whole cells were extracted using a phenol/water method resulting in recovery of material exhibiting a similar activity profile. An LPS-LBP binding competition assay revealed an interaction of the treponeme phenol/water extracts with LBP, while precipitation studies implied an affinity to polymyxin B and endotoxin neutralizing protein. Macrophages obtained from C3H/HeJ mice carrying a Toll-like receptor (TLR)-4 mutation were stimulated with treponeme extracts for NO release to assess the role of TLRs in cell activation. Furthermore, NF-kappaB translocation in TLR-2-negative Chinese hamster ovary (CHO) cells was studied. We found that phenol/water-extracts of the two strains use TLRs differently with T. brennaborense-stimulating cells in a TLR-4-dependent fashion, while T. maltophilum-mediated activation apparently involved TLR-2. These results indicate the presence of a novel class of glycolipids in Treponema initiating inflammatory responses involving LBP, CD14, and TLRs.

  19. Phenotypic and Functional Properties of Human Steady State CD14+ and CD1a+ Antigen Presenting Cells and Epidermal Langerhans Cells

    PubMed Central

    Fehres, Cynthia. M.; Bruijns, Sven C. M.; Sotthewes, Brigit N.; Kalay, Hakan; Schaffer, Lana; Head, Steven R.; de Gruijl, Tanja D.; Garcia-Vallejo, Juan J.; van Kooyk, Yvette

    2015-01-01

    Cutaneous antigen presenting cells (APCs) are critical for the induction and regulation of skin immune responses. The human skin contains phenotypically and functionally distinct APCs subsets that are present at two separated locations. While CD1ahigh LCs form a dense network in the epidermis, the CD14+ and CD1a+ APCs reside in the dermal compartment. A better understanding of the biology of human skin APC subsets is necessary for the improvement of vaccine strategies that use the skin as administration route. In particular, progress in the characterization of uptake and activatory receptors will certainly improve APC-targeting strategies in vaccination. Here we performed a detailed analysis of the expression and function of glycan-binding and pattern-recognition receptors in skin APC subsets. The results demonstrate that under steady state conditions human CD1a+ dermal dendritic cells (DCs) were phenotypically most mature as measured by the expression of CD83 and CD86, whereas the CD14+ cells showed a higher expression of the CLRs DC-SIGN, mannose receptor and DCIR and had potent antigen uptake capacity. Furthermore, steady state LCs showed superior antigen cross-presentation as compared to the dermal APC subsets. Our results also demonstrate that the TLR3 ligand polyribosinic-polyribocytidylic acid (pI:C) was the most potent stimulator of cytokine production by both LCs and dDCs. These studies warrant further exploration of human CD1a+ dDCs and LCs as target cells for cancer vaccination to induce anti-tumor immune responses. PMID:26605924

  20. Actinobacillus actinomycetemcomitans lipopolysaccharide stimulates the phosphorylation of p44 and p42 MAP kinases through CD14 and TLR-4 receptor activation in human gingival fibroblasts.

    PubMed

    Gutiérrez-Venegas, Gloria; Kawasaki-Cárdenas, Perla; Cruz-Arroyo, Santa Rita; Pérez-Garzón, Miguel; Maldonado-Frías, Silvia

    2006-04-25

    Tyrosine phosphorylation is an early step in lipopolysaccharide (LPS) stimulated monocytes and macrophages that appears to play a key role in signal transduction. We have demonstrated that LPS purified from Actinobacillus actinomycetemcomitans also increases protein tyrosine phosphorylation in human gingival fibroblasts (HGF). This effect was elicited rapidly after LPS stimulation at concentrations that stimulate anti-bacterial responses in human gingival fibroblasts. Two main proteins, with an apparent molecular weight of 44 and 42 kDa, were phosphorylated after LPS stimulation of the human gingival fibroblasts. The phosphorylation was detected after 5 to 15 min and reached the maximum at 30 min of treatment. The increase in tyrosine phosphorylation was apparent following stimulation with LPS at 10 ng/ml and the response was dose dependent up to 10 microg/ml. Pretreatment with the tyrosine kinase inhibitors, herbimycin A and genistein inhibited the LPS-stimulated phosphorylation of p44 and p42 MAP kinases in a dose dependent manner. Pretreatment of human gingival fibroblasts with antibodies anti-CD14 or anti-TLR-4 but not anti-TLR-2 inhibited the LPS-induced tyrosine phosphorylation of p44 and p42. Additionally, LPS-induced p44 and p42 phosphorylation was inhibited by polymyxin treatment. These findings demonstrate that LPS from A. actinomycetemcomintans increases rapidly p44 and p42 phosphorylation (ERK 1 and ERK 2, respectively) in human gingival fibroblasts. Our data also suggest that CD14 and TLR-4 receptors are involved in the LPS effects in human gingival fibroblasts.

  1. Moraxella catarrhalis adhesin UspA1-derived recombinant fragment rD-7 induces monocyte differentiation to CD14+CD206+ phenotype.

    PubMed

    Xie, Qi; Brackenbury, Louise S; Hill, Darryl J; Williams, Neil A; Qu, Xun; Virji, Mumtaz

    2014-01-01

    Circulating monocytes in the bloodstream typically migrate to other tissues and differentiate into tissue resident macrophages, the process being determined by the constituents of the microenvironments encountered. These may include microbes and their products. In this study, we investigated whether Moraxella catarrhalis Ubiquitous Surface Protein A1 (UspA1), known to bind to a widely expressed human cell surface receptor CEACAM1, influences monocyte differentiation as receptor engagement has been shown to have profound effects on monocytes. We used the recombinant molecules corresponding to the regions of UspA1 which either bind (rD-7; UspA1527-665) or do not bind (r6-8; UspA1659-863) to CEACAM1 and investigated their effects on CD206, CD80 and CD86 expression on freshly isolated human CD14+ monocytes from peripheral blood mononuclear cells (PBMC). Exposure to rD-7, but not r6-8, biased monocyte differentiation towards a CD14+CD206+ phenotype, with reduced CD80 expression. Monocytes treated with rD-7 also secreted high levels of IL-1ra and chemokine IL-8 but not IL-10 or IL-12p70. The effects of rD-7 were independent of any residual endotoxin. Unexpectedly, these effects of rD-7 were also independent of its ability to bind to CEACAM1, as monocyte pre-treatment with the anti-CEACAM antibody A0115 known to inhibit rD-7 binding to the receptor, did not affect rD-7-driven differentiation. Further, another control protein rD-7/D (a mutant form of rD-7, known not to bind to CEACAMs), also behaved as the parent molecule. Our data suggest that specific regions of M. catarrhalis adhesin UspA1 may modulate inflammation during infection through a yet unknown receptor on monocytes.

  2. Soluble and membrane levels of molecules involved in the interaction between clonal plasma cells and the immunological microenvironment in multiple myeloma and their association with the characteristics of the disease.

    PubMed

    Perez-Andres, Martin; Almeida, Julia; Martin-Ayuso, Marta; De Las Heras, Natalia; Moro, Maria Jesus; Martin-Nuñez, Guillermo; Galende, Josefina; Cuello, Rebeca; Abuín, Isabel; Moreno, Inmaculada; Domínguez, Mercedes; Hernandez, Jose; Mateo, Gema; San Miguel, Jesus F; Orfao, Alberto

    2009-01-15

    Clonal plasma cells (PC) from different types of monoclonal gammopathies (MG) display distinct phenotypes consistent with an increased antigen-presentation and T-cell costimulation in MG of undetermined significance that deteriorates in malignant conditions. Expression of other cell surface and soluble molecules (e.g. adhesion/proliferation molecules) involved in the interaction between clonal PC and the bone marrow (BM) microenvironment has also been related to malignant PC, although the exact clinical significance of their expression remains largely unknown. Analysis of cell surface levels of several of these molecules in multiple myeloma (MM) patients shows an association between lower expression on BMPC of the HLA-I and beta2-microglobulin antigen-presenting molecules, the CD126 and CD130 IL6 receptor (IL6R) chains, and CD38, and adverse prognostic features of the disease. Likewise, patients showing higher soluble levels of antigen-presenting molecules (HLA-I and beta2-microglobulin), IL6R and CD95 tended to be associated with more aggressive disease behavior. In contrast, CD40, CD86, CD56, CD19, and CD45 were not associated with patients' outcome. Interestingly, upon considering the ratio between the soluble and PC membrane expression of each molecule, an increased adverse prognostic impact was observed for both HLA-I and beta2-microglobulin, but not for the other molecules. Multivariate analysis confirmed the independent prognostic value of cell surface expression of CD126 on BMPC together with serum beta2-microglobulin and LDH. In summary, our results show an abnormal distribution of the cellular and soluble compartments of the HLA-I, IL6R, and to a lower extent, CD95 molecules, in MM, associated with the clinical characteristics and behavior of the disease. Copyright (c) 2008 Wiley-Liss, Inc.

  3. Inhibitory FcγRIIb-Mediated Soluble Antigen Clearance from Plasma by a pH-Dependent Antigen-Binding Antibody and Its Enhancement by Fc Engineering

    PubMed Central

    Iwayanagi, Yuki; Maeda, Atsuhiko; Haraya, Kenta; Wada, Naoko A.; Shibahara, Norihito; Ohmine, Ken; Nambu, Takeru; Nakamura, Genki; Mimoto, Futa; Katada, Hitoshi; Ito, Shunsuke; Tachibana, Tatsuhiko; Jishage, Kou-ichi; Hattori, Kunihiro

    2015-01-01

    Fc engineering can modulate the Fc–FcγR interaction and thus enhance the potency of Abs that target membrane-bound Ags, but it has not been applied to Abs that target soluble Ags. In this study, we revealed a previously unknown function of inhibitory FcγRII in vivo and, using an Ab that binds to Ag pH dependently, demonstrated that the function can be exploited to target soluble Ag. Because pH-dependent Ab dissociates Ag in acidic endosome, its Ag clearance from circulation reflects the cellular uptake rate of Ag/Ab complexes. In vivo studies showed that FcγR but not neonatal FcR contributes to Ag clearance by the pH-dependent Ab, and when Fc binding to mouse FcγRII and III was increased, Ag clearance was markedly accelerated in wild-type mice and FcR γ-chain knockout mice, but the effect was diminished in FcγRII knockout mice. This demonstrates that mouse FcγRII efficiently promotes Ab uptake into the cell and its subsequent recycling back to the cell surface. Furthermore, when a human IgG1 Fc variant with selectively increased binding to human FcγRIIb was tested in human FcγRIIb transgenic mice, Ag clearance was accelerated without compromising the Ab half-life. Taken together, inhibitory FcγRIIb was found to play a prominent role in the cellular uptake of monomeric Ag/Ab immune complexes in vivo, and when the Fc of a pH-dependent Ab was engineered to selectively enhance human FcγRIIb binding, the Ab could accelerate soluble Ag clearance from circulation. We assume such a function would enhance the therapeutic potency of Abs that target soluble Ags. PMID:26320252

  4. The number of circulating CD14+ cells is related to infarct size and postinfarct volumes in ST segment elevation myocardial infarction but not non-ST segment elevation myocardial infarction

    PubMed Central

    Montange, Damien; Davani, Siamak; Deschaseaux, Frédéric; Séronde, Marie France; Chopard, Romain; Schiele, François; Jehl, Jérome; Bassand, Jean Pierre; Kantelip, Jean-Pierre; Meneveau, Nicolas

    2012-01-01

    OBJECTIVE: To determine the relationship between the number of CD14+ cells, myocardial infarct (MI) size and left ventricular (LV) volumes in ST segment elevation MI (STEMI) and non-ST segment elevation MI (NSTEMI) patients. METHODS: A total of 62 patients with STEMI (n=34) or NSTEMI (n=28) were enrolled. The number of CD14+ cells was assessed at admission. Infarct size, left ventricular ejection fraction (LVEF) and LV volumes were measured using magnetic resonance imaging five days after MI and six months after MI. Results: In STEMI patients, the number of CD14+ cells was positively and significantly correlated with infarct size at day 5 (r=0.40; P=0.016) and after six months (r=0.34; P=0.047), negatively correlated with LVEF at day 5 (r=−0.50; P=0.002) and after six months (r=−0.46; P=0.005) and positively correlated with end-diastolic (r=0.38; P=0.02) and end-systolic (r=0.49; P=0.002) volumes after six months. In NSTEMI patients, no significant correlation was found between the number of CD14+ cells and infarct size, LVEF or LV volumes at day 5 or after six months. CONCLUSIONS: The number of CD14+ cells at admission was associated with infarct size and LV remodelling in STEMI patients with large infarct size, whereas in NSTEMI patients, no relationship was observed between numbers of CD14+ cells and LV remodelling. PMID:23620701

  5. Glucocorticoid treatment at moderate doses of SIVmac251-infected rhesus macaques decreases the frequency of circulating CD14+CD16++ monocytes but does not alter the tissue virus reservoir.