Search

Item does not contain fulltext Solid-phase multiplex-bead assays are widely used in transplantation to detect anti-human leukocyte antigen (HLA) antibodies. These assays enable high resolution detection of low levels of HLA antibodies. However, multiplex-bead assays are costly and yield variable measurements that limit the comparison of results between laboratories. In the context of a Dutch national Consortium study we aimed to determine the inter-assay and inter-machine variability of multiplex-bead assays, and we assessed how to reduce the assay reagents costs. Fifteen sera containing a variety of HLA antibodies were used yielding in total 7092 median fluorescence intensities (MFI) values. The inter-assay and inter-machine mean absolute relative differences (MARD) of the screening assay were 12% and 13%, respectively. The single antigen bead (SAB) inter-assay MARD was comparable, but showed a higher lot-to-lot variability. Reduction of screening assay reagents to 50% or 40% of manufacturers' recommendations resulted in MFI values comparable to 100% of the reagents, with an MARD of 12% or 14%, respectively. The MARD of the 50% and 40% SAB assay reagent reductions were 11% and 22%, respectively. From this study, we conclude that the reagents can be reliably reduced at least to 50% of manufacturers' recommendations with virtually no differences in HLA antibody assignments.

Contains fulltext : 13844.pdf (Publisher’s version ) (Closed access)

Activation of the complement system may contribute to the pathogenesis of many diseases. Hence, an effective inhibitor of complement might be useful to reduce tissue damage. Some glycosaminoglycans (GAG), such as heparin, are known to inhibit the interaction of C1q with activators and the assembly of the classical and the alternative pathway C3 convertases. Furthermore, they may potentiate C1 inhibitor-mediated inactivation of C1s. To search for potential complement inhibitors, we systematically investigated the complement inhibitory properties of various synthetic and naturally occurring GAG (dextran sulfates 500,000 and 5,000, heparin, N-acetylheparin, heparan sulfate, dermatan sulfate, and chondroitin sulfates A and C). First, we assessed the effect of GAG on the second-order rate constant of the inactivation of C1s by C1 inhibitor. This rate constant increased 6- to 130-fold in the presence of the GAG, dextran sulfate being the most effective. Second, all tested GAG were found to reduce deposition of C4 and C3 on immobilized aggregated human IgG (AHG) and to reduce fluid phase formation of C4b/c and C3b/c in recalcified plasma upon incubation with AHG. Dextran sulfate again was found to be most effective. We conclude that GAG modulate complement activation in vitro and that the low molecular weight dextran sulfate (m.w. 5000) may be a candidate for pharmacologic manipulation of complement activation via potentiation of C1 inhibitor.

Contains fulltext : 26146.PDF (Publisher’s version ) (Open Access)

Background: The massive pulmonary neutrophil influx in respiratory distress syndrome (RDS) in preterm infants has been ascribed to the effect of leukotriene B-4 (LTB4). Objectives: To investigate whether secretory phospholipase A(2) (sPLA(2)), the rate-limiting enzyme in LTB4 production, is present in lungs of RDS infants and stimulates neutrophil migration. Methods: sPLA(2) was measured in tracheal aspirates from 15 preterm infants with RDS. The effect of aspirates on cord blood neutrophil migration was first measured, and the contribution of sPLA(2) was assessed by addition of its endogenous inhibitor Clara cell protein (CC16) or absorption of sPLA(2) from the aspirates. The role of intracellular signal transduction activation and LTB4 formation in sPLA(2)-induced neutrophil migration was determined using purified sPLA(2), several inhibitors of signal transduction, a LTB4 synthesis inhibitor and a LTB4 receptor antagonist. Results: All aspirates contained sPLA(2), which significantly stimulated neutrophil migration. Addition of CC16 or absorption of sPLA(2) abolished the stimulatory effect. All inhibitors significantly reduced sPLA(2)-induced neutrophil migration. Conclusions: sPLA(2) is present in tracheal aspirates of preterm infants with RDS. Human recombinant sPLA(2) and pancreatic type sPLA(2) stimulate in vitro cord blood neutrophil migration via activation of intracellular signal transduction pathways, LTB4 production and receptor binding. We speculate that sPLA(2) contributes to pulmonary neutrophil influx in RDS. Further studies are needed to determine the potential of sPLA(2) inhibition as a treatment for RDS. Copyright (C) 2009 S. Karger AG, Basel

The serine protease inhibitor C1-Inhibitor (C1-Inh) inhibits several complement- and contact-system proteases, which play an important role in inflammation. C1-Inh has a short reactive site loop (RSL) compared to other serpins. RSL length determines the inhibitory activity of serpins. We investigated the effect of RSL elongation on inhibitory activity of C1-Inh by insertion of one or two alanine residues in the RSL. One of five mutants had an increased association rate with kallikrein, but was nevertheless a poor inhibitor because of a simultaneous high stoichiometry of inhibition (>10). The association rate of the other variants was lower than that of wild-type C1-Inh. These data suggest that the relatively weak inhibitory activity of C1-Inh is not the result of its short RSL. The short RSL of C1-Inh has, surprisingly, the optimal length for inhibition. (C) 2004 Elsevier B.V. All rights reserved.

Item does not contain fulltext OBJECTIVE: To investigate the patterns of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-6, interferon-gamma (IFN-gamma) and interleukin-1 receptor antagonist (IL-1Ra) during the course of dengue shock syndrome. DESIGN: Prospective clinical study. SETTING: Pediatric Intensive Care Unit, Dr. Kariadi Hospital, the university hospital of Diponegoro University, Semarang, Indonesia. PATIENTS: Fifty children with dengue shock syndrome. MEASUREMENTS: The plasma concentration and the ex vivo production, with and without lipopolysaccharide (LPS), of TNF-alpha, IL-1beta and IL-1Ra were measured in duplicate by nonequilibrium radioimmunoassay (RIA); IFN-gamma and IL-6 were measured by ELISA. RESULTS: During the acute phase, the plasma concentrations and the ex vivo production without LPS of IL-1Ra were considerably elevated and returned to normal on recovery. However, the ex vivo LPS-stimulated production of the proinflammatory cytokines TNF-alpha and IL-1beta were considerably depressed. Also, these concentrations returned towards normal on recovery. In non-survivors, the plasma concentrations of IL-6 and IL-1Ra were significantly higher than in survivors (p = 0.00001 and p = 0.0005, respectively). In addition, the ex vivo production of IL-1Ra in non-survivors was significantly higher than in survivors, both without LPS stimulation (p = 0.0008) and with LPS (p < 0.004). IL-1Ra was significantly associated with mortality (p = 0.007). CONCLUSION: Since IL-1Ra was significantly associated with mortality, this measurement may be used as an index of disease severity in dengue shock syndrome.

Nederlands Tijdschrift voor Klinische Chemie 27(2),69(2002) -- Nederlandse Vereniging voor Klinische Chemie en Laboratoriumgeneeskunde -- 1

Item does not contain fulltext In a prospective clinical study of 50 patients with Dengue Shock Syndrome (DSS), we investigated the association of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-1 receptor antagonist (IL-1Ra), and IL-6 with activation markers of coagulation (F1+2 and TATc) and fibrinolysis (t-PA, PAPc, and D-dimer). We found that TNF-alpha, IL-1beta and Il-1Ra, but not IL-6, concentrations were elevated in the circulation during the early stage of infection and at discharge from hospital. TNF-alpha was significantly associated with D-dimer, an activation marker of fibrinolysis (p < 0.003), but not with activation markers of coagulation. IL-1beta was significantly associated with t-PA (p < 0.03). IL-1Ra was significantly associated with F1+2, TATc (p < 0.04 and p < 0.02, respectively), whereas IL-6 was significantly associated with both, activation markers of coagulation (F1+2; p < 0.03) and fibrinolysis (PAPc; p = 0.002). Our data are in line with studies in bacterial sepsis. In severe dengue virus infection the same cytokines are involved in the onset and regulation of hemostasis.

Item does not contain fulltext 2 p.

Item does not contain fulltext

BACKGROUND: Rises in circulating C-reactive protein (CRP), the prototypical acute-phase protein in humans, correlate with clinical outcome in patients with myocardial ischemia and infarction. We hypothesized that these correlations might reflect active participation of CRP in the local inflammatory response ensuing in the jeopardized myocardium because on binding to a ligand, CRP is able to activate the classic pathway of complement, and in addition, complement activation has been shown to occur locally in infarcted myocardium.METHODS AND RESULTS: To verify our hypothesis, we investigated localization of CRP in relation to deposition of complement in tissue specimens of infarcted and healthy heart tissue obtained from 17 patients who had died after acute myocardial infarction. CRP was found to be deposited only in infarcted regions and not in normal-appearing areas of the myocardium, being colocalized with depositions of C4 and C3 activation fragments of the complement system. Deposition of CRP and complement in infarcted myocardium appeared to be time dependent, because it was found in all infarctions except for one of young age (< 12 hours old) and two of greater age (> 1 year old), whereas another tissue specimen of an infarct < 12 hours old showed only moderate but positive staining for both CRP and complement in comparison with older infarctions.CONCLUSIONS: We conclude that in humans, CRP may localize in infarcted heart tissue and suggest that this acute-phase protein promotes local complement activation, and hence tissue damage, in acute myocardial infarction.

To assess the role of interleukin-12 (IL-12) and gamma interferon (IFN-gamma) in children with bacterial meningitis, bioactive IL-12 (p70) and the inactive subunit p40 and IFN-gamma were measured in serum and cerebrospinal fluid (CSF) from 35 children with bacterial meningitis and 10 control subjects. The production of IFN-gamma is induced by IL-12 with tumor necrosis factor alpha (TNF-alpha) as a costimulator and inhibited by IL-10. CSF concentrations of IL-12 p40 as well as those of IFN-gamma were markedly elevated, whereas IL-12 p70 was hardly detectable. Detectable CSF levels of IFN-gamma correlated positively with IL-12 p40 (r = 0.40, P = 0.02) and TNF-alpha (r = 0.46, P = 0.04) but not with IL-6, IL-8, or IL-10. In contrast to CSF levels of TNF-alpha, IL-12, and IL-10, those of IFN-gamma were significantly higher in patients with pneumococcal meningitis than in children with meningitis caused by Haemophilus influenzae and Neisseria meningitidis, presumably because of a high CSF TNF-alpha/IL-10 ratio in the former. We suggest that IL-12- and TNF-alpha-induced IFN-gamma production may contribute to the natural immunity against microorganisms in the CSF compartment during the acute phase of bacterial meningitis. -------------------------------------------------------------------------------- Write to PMC | PMC Home | PubMed NCBI | U.S. National Library of Medicine NIH | Department of Health and Human Services Privacy Policy | Disclaimer | Freedom of Information Act

Contains fulltext : 25390___.PDF (Publisher’s version ) (Open Access)