Hypoxemia regulates effect of lipopolysaccharide on polymorphonuclear leukocyte CD11b/CD18 expression

Expression of the receptor CD11b/CD18 on the polymorphonuclear leukocyte (PMN) surface is important for several aspects of PMN function during endotoxemia. The mechanisms underlying regulation of CD11b/CD18 expression during hypoxemia and endotoxemia, however, are less clear. We investigated the effects of exposure of whole blood PMNs to lipopolysaccharide (LPS) during hypoxemia. During hypoxemia (10–30% O2 saturation), LPS reduced CD11b/CD18 expression. Both kinetic assays and experiments with microfilament stabilizers (phalloidin, cytochalasin B) demonstrated that this was most likely due to receptor shedding. Incubation of whole blood PMNs with an anti-CD14 monoclonal antibody (MEM18) largely prevented the LPS-induced reduction of CD11b/CD18 expression. Decreased CD11b/CD18 expression reduced PMN functional capability, as the binding of its ligand (erythrocytes opsonized with the 3rd component of complement Cbi) and intracellular H2O2 production were reduced. After exposure to LPS, N-formyl-methionyl-leucyl-phenylalanine could rapidly induce new CD11b/CD18 receptors to the cell surface, and this was not inhibited by actinomycin D or cycloheximide. After reoxygenation (> 90% O2 saturation), CD11b/CD18 expression was restored, and this was abrogated by exposure to cytochalasin B. Lipid A was able to reproduce the effects of LPS during hypoxemia and hypoxemia-reoxygenation but required a 10-fold greater concentration to do so. These results demonstrate that during hypoxemia an important pathophysiological property of LPS is to reduce CD11b/CD18 expression. This results in diminished PMN functional capability, which would contribute to the pathogenicity of LPS during acute infectious states.

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Effects of Anti-Lipid a Human Monoclonal Antibody on Lipopolysaccharide-Induced Toxicity to the Kidney

The Journal of Urology ◽

10.1016/s0022-5347(17)41347-4 ◽

1989 ◽

Vol 141 (6) ◽

pp. 1463-1466 ◽

Cited By ~ 2

Author(s):

Bruce M. Tune ◽

Chieh-Yin Hsu ◽

Marcia M. Bieber ◽

Nelson N.H. Teng

Keyword(s):

Monoclonal Antibody ◽

Lipid A ◽

Human Monoclonal Antibody

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Development of a new monoclonal antibody-based point-of-care testing assay for the quantification of procalcitonin in whole blood for a rapid sepsis diagnostic

Critical Care ◽

10.1186/cc12913 ◽

2013 ◽

Vol 17 (Suppl 4) ◽

pp. P13

Author(s):

Martin Rieger ◽

Daniela Rascher ◽

Anton Hartmann

Keyword(s):

Monoclonal Antibody ◽

Whole Blood ◽

Point Of Care ◽

Point Of Care Testing

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Monoclonal Fluorescence Polarization Immunoassay Evaluated for Monitoring Cyclosporine in Whole Blood After Kidney, Heart, and Liver Transplantation

Clinical Chemistry ◽

10.1093/clinchem/38.1.123 ◽

1992 ◽

Vol 38 (1) ◽

pp. 123-126 ◽

Cited By ~ 28

Author(s):

M Winkler ◽

G Schumann ◽

D Petersen ◽

M Oellerich ◽

K Wonigeit

Keyword(s):

Monoclonal Antibody ◽

Liver Transplantation ◽

Whole Blood ◽

Fluorescence Polarization ◽

Fluorescence Polarization Immunoassay ◽

Transplant Patients ◽

Specific Radioimmunoassay ◽

Metabolite Concentrations ◽

A Prospective Study

Abstract In a prospective study we evaluated a novel fluorescence polarization immunoassay (FPIA) for determining cyclosporine (CsA) in whole blood. FPIA uses a monoclonal antibody and is performed on the TDx (Abbott). The within-series (CV less than 2%) and between-days (CV less than 3.3%) precision of the assay was excellent. The results obtained by the monoclonal FPIA in samples from transplant patients (n = 100) averaged 31.9% and 20.2% higher than those by HPLC and a specific radioimmunoassay (INCStar), respectively. Results by all three methods correlated well. Follow-up studies during the early course after liver transplantation, however, suggested that high metabolite concentrations affect FPIA results. This is explained by previously described cross-reactions of the monoclonal antibody with some CsA metabolites. The FPIA results in samples of such patients should be interpreted cautiously.

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Radioreceptor Assay for Quantifying FK-506 Immunosuppressant in Whole Blood

Clinical Chemistry ◽

10.1093/clinchem/38.7.1307 ◽

1992 ◽

Vol 38 (7) ◽

pp. 1307-1310 ◽

Cited By ~ 10

Author(s):

J N Murthy ◽

Y Chen ◽

V S Warty ◽

R Venkataramanan ◽

J G Donnelly ◽

...

Keyword(s):

Monoclonal Antibody ◽

Enzyme Immunoassay ◽

Whole Blood ◽

Binding Protein ◽

Radioreceptor Assay ◽

Fk 506 ◽

Blood Concentrations ◽

Chromatographic Columns ◽

Analytical Recovery

Abstract We describe a quantitative radioreceptor assay (RRA) for quantifying FK-506 in whole blood. FK-506 extracted from whole blood with a cyclohexyl-sorbent column competes with [3H]dihydro-FK-506 for binding to a partially purified preparation of FK-506 binding protein (FK-BP). Free and protein-bound FK-506 are separated on LH 20 Sephadex chromatographic columns. We compared the results of this method with those of an enzyme immunoassay that uses a monoclonal antibody: r = 0.97, Sy/x = 0.039. Between-day precisions (CV) at FK-506 concentrations of 8 and 17 micrograms/L were 9.2% and 8.2%, respectively. Within-run precisions were 5.9%, 8.1%, and 9.4%, respectively, at 4, 8, and 15 micrograms/L. Analytical recovery, evaluated at 5, 10, 15, 20, and 25 micrograms/L for FK-506 added to whole blood, ranged from 98% to 103%. The assay can reliably quantify FK-506 blood concentrations between 1.0 and 25 micrograms/L.

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Effect of anti-CD20 monoclonal antibody, Rituxan, on cynomolgus monkey and human B cells in a whole blood matrix

Cytometry ◽

10.1002/cyto.a.10030 ◽

2003 ◽

Vol 52A (2) ◽

pp. 101-109 ◽

Cited By ~ 34

Author(s):

Yulia Vugmeyster ◽

Kathy Howell ◽

Anahid Bakshl ◽

Clarissa Flores ◽

Eleanor Canova-Davis

Keyword(s):

Monoclonal Antibody ◽

B Cells ◽

Whole Blood ◽

Cynomolgus Monkey ◽

Human B Cells ◽

Anti Cd20

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Rapid one-step whole blood C–reactive protein magnetic permeability immunoassay with monoclonal antibody conjugated nanoparticles as superparamagnetic labels and enhanced sedimentation

Analytical and Bioanalytical Chemistry ◽

10.1007/s00216-005-0094-6 ◽

2005 ◽

Vol 384 (3) ◽

pp. 651-657 ◽

Cited By ~ 29

Author(s):

Filiz Ibraimi ◽

Dario Kriz ◽

Min Lu ◽

Lars-Olof Hansson ◽

Kirstin Kriz

Keyword(s):

Monoclonal Antibody ◽

Magnetic Permeability ◽

Whole Blood ◽

C Reactive Protein ◽

Reactive Protein ◽

One Step

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Phenotypic Comparison of Platelet- and Plasma-Derived Factor Va: Tyrosine Phosphorylation Differentiates the Two Cofactor Pools.

Blood ◽

10.1182/blood.v108.11.1696.1696 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1696-1696

Author(s):

Jeremy P. Wood ◽

Paula B. Tracy

Keyword(s):

Monoclonal Antibody ◽

Tyrosine Phosphorylation ◽

Light Chain ◽

Whole Blood ◽

Membrane Surface ◽

Tyrosine Phosphatase ◽

Light Chains ◽

Factor V ◽

Phosphatase Inhibitors ◽

Factor Va

Abstract Factor Va, the essential cofactor for thrombin formation via the Prothrombinase complex, exists in two pools in whole blood: 75–80% is found in the plasma, while platelet alpha-granules contain the remaining 20–25%. Platelet-derived factor Va possesses several modifications not present in its plasma counterpart, such as an O-glycosylation on Thr402, and is the more procoagulant molecule, exhibiting resistance to inactivation by activated protein C (APC) and plasmin when bound to the activated platelet membrane surface. As the platelet-derived cofactor pool originates from endocytosis of plasma-derived factor V by megakaryocytes, it can be hypothesized that, subsequent to its endocytosis, factor V is post-translationally modified, proteolytically processed, and packaged into alpha-granules to form the unique platelet-derived molecule. Platelet- and plasma-derived factor Va were purified from three individual donors. Platelets were isolated from whole blood and lysed with Triton X-100 in the presence of the tyrosine phosphatase inhibitors sodium orthovanadate and okadaic acid to prevent dephosphorylation by any released tyrosine phosphatases. Factor V in both the platelet lysates and the platelet-poor plasma was activated with thrombin, adsorbed onto a resin-coupled monoclonal antibody against human factor V, washed extensively, and eluted using 10% SDS. Western blotting analyses, using a monoclonal antibody against phosphotyrosine residues, revealed the presence of tyrosine phosphorylation on the light chain of plasma-derived, but not platelet-derived, factor Va. No such modification was observed on the heavy chain of either cofactor. Monoclonal antibodies against the heavy and light chains of factor V were used to verify that similar levels of factor V were present in all samples. This differential modification was observed in all three donors. Subsequent MALDI-TOF analyses of trypsin-digested plasma- and platelet-derived factor Va light chains were performed to identify the site(s) of tyrosine phosphorylation. These data identified phosphorylation of the C-terminal tyrosine residue (Tyr2196) in plasma-derived, but not platelet-derived, factor Va. The phosphorylated fragment (residues 2188–2196, m/z = 1152.5) is consistently present in spectra of plasma-derived factor Va, while only the non-phosphorylated fragment (m/z = 1072.5) has been observed in the platelet-derived cofactor. To demonstrate that the phosphatase inhibitors used were effective, the platelets were lysed in the presence of added plasma-derived factor Va. Tyrosine phosphorylation, as determined by immunoblotting, was still present on the light chain, indicating that the dephosphorylation was occurring prior to platelet lysis. Tyrosine phosphorylation of other proteins has been implicated in regulation of endocytosis. Further experiments will need to be performed to determine if the modification has a similar role in megakaryocyte endocytosis of factor V.

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A polyreactive human anti-lipid A monoclonal antibody having cross reactivity to polysaccharide portions ofPseudomonas aeruginosalipopolysaccharides

FEMS Immunology & Medical Microbiology ◽

10.1111/j.1574-695x.1996.tb00264.x ◽

1996 ◽

Vol 14 (1) ◽

pp. 31-38 ◽

Cited By ~ 7

Author(s):

Shin-ichi Yokota ◽

Hiroshi Ohtsuka ◽

Tsuneo Kohzuki ◽

Hiroshi Noguchi

Keyword(s):

Monoclonal Antibody ◽

Lipid A ◽

Cross Reactivity

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Deactivation of the lipopolysaccharide antagonist eritoran (E5564) by high-density lipoprotein-associated apolipoproteins

Innate Immunity ◽

10.1177/1753425910394395 ◽

2011 ◽

Vol 18 (1) ◽

pp. 171-178 ◽

Cited By ~ 6

Author(s):

Jacklyn G Fleischer ◽

Dan Rossignol ◽

Gordon A Francis ◽

Teddy Chan ◽

Melvyn Lynn ◽

...

Keyword(s):

Whole Blood ◽

Lipid A ◽

Density Lipoprotein ◽

Antagonistic Activity ◽

High Density ◽

High Density Lipoproteins ◽

Amyloid A ◽

Tnf Α ◽

Signalling Cascade ◽

Pro Inflammatory Cytokines

Lipid A, the active moiety of LPS, exerts its effects through interaction with TLR4, triggering a signalling cascade that results in the release of pro-inflammatory cytokines. Eritoran is a lipid A analogue that competes with LPS for binding to TLR4; however, after intravenous administration, it undergoes a time-dependent deactivation as a consequence of binding to high-density lipoproteins (HDLs). The site of eritoran association with HDL remains unknown. Therefore the aim of this study was to determine if HDL-associated apolipoproteins A1, A2, serum amyloid A (SAA) and C1, inhibit the ability of eritoran to block LPS-induced TNF-α release from whole blood. Eritoran activity after LPS stimulation in human whole blood was assessed in the presence of reconstituted HDL (rHDL) containing different apos. In rHDL, the major apolipoproteins in both the healthy and septic state, A1 and SAA, caused a significant reduction in eritoran antagonistic activity and had a greater effect than minor apolipoproteins A2 and C1. Apolipoproteins associated with HDL are likely to facilitate eritoran deactivation. Apolipoproteins A1 and SAA should be of particular focus as they are the major apos found on HDL in both the healthy and septic state. Further evaluation of the physical association between apolipoproteins and eritoran should be explored.

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