PubMed Health⌕ Search
Home/Archive/PubMed

PubMed · 10233059

Quantifying aggregation of IgE-FcepsilonRI by multivalent antigen.

W S Hlavacek·A S Perelson·B Sulzer·J Bold·J Paar·W Gorman·R G Posner
View original source ↗Export metadata ↓

Abstract

Aggregation of cell surface receptors by multivalent ligand can trigger a variety of cellular responses. A well-studied receptor that responds to aggregation is the high affinity receptor for IgE (FcepsilonRI), which is responsible for initiating allergic reactions. To quantify antigen-induced aggregation of IgE-FcepsilonRI complexes, we have developed a method based on multiparameter flow cytometry to monitor both occupancy of surface IgE combining sites and association of antigen with the cell surface. The number of bound IgE combining sites in excess of the number of bound antigens, the number of bridges between receptors, provides a quantitative measure of IgE-FcepsilonRI aggregation. We demonstrate our method by using it to study the equilibrium binding of a haptenated fluorescent protein, 2,4-dinitrophenol-coupled B-phycoerythrin (DNP25-PE), to fluorescein isothiocyanate-labeled anti-DNP IgE on the surface of rat basophilic leukemia cells. The results, which we analyze with the aid of a mathematical model, indicate how IgE-FcepsilonRI aggregation depends on the total concentrations of DNP25-PE and surface IgE. As expected, we find that maximal aggregation occurs at an optimal antigen concentration. We also find that aggregation varies qualitatively with the total concentration of surface IgE as predicted by an earlier theoretical analysis.

Explore related subjects

2,4-DinitrophenolAnimalsAntigensBiophysical PhenomenaBiophysicsCell MembraneFlow CytometryFluorescein-5-isothiocyanateFluorescent DyesImmunoglobulin ELigandsMacromolecular SubstancesMiceModels, BiologicalPhycoerythrinRatsReceptor AggregationReceptors, IgETumor Cells, CulturedJournal ArticleResearch Support, U.S. Gov't, Non-P.H.S.Research Support, U.S. Gov't, P.H.S.

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

W S Hlavacek, A S Perelson, B Sulzer, J Bold, J Paar, W Gorman, R G Posner. 1999. Quantifying aggregation of IgE-FcepsilonRI by multivalent antigen.. https://doi.org/10.1016/s0006-3495(99)77397-2

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

PubMed2007

Ketone bodies alter dinitrophenol-induced glucose uptake through AMPK inhibition and oxidative stress generation in adult cardiomyocytes.

In aerobic conditions, the heart preferentially oxidizes fatty acids. However, during metabolic stress, glucose becomes the major energy source, and enhanced glucose uptake has a protective effect on heart function and cardiomyocyte survival. Thus abnormal regulation of glucose uptake may contribute to the development of cardiac disease in diabetics. Ketone bodies are often elevated in poorly controlled diabetics and are associated with increased cellular oxidative stress. Thus we sought to determine the effect of the ketone body beta-hydroxybutyrate (OHB) on cardiac glucose uptake during metabolic stress. We used 2,4-dinitrophenol (DNP), an uncoupler of the mitochondrial oxidative chain, to mimic hypoxia in cardiomyocytes. Our data demonstrated that chronic exposure to OHB provoked a concentration-dependent decrease of DNP action, resulting in 56% inhibition of DNP-mediated glucose uptake at 5 mM OHB. This was paralleled by a diminution of DNP-mediated AMP-activated protein kinase (AMPK) and p38 MAPK phosphorylation. Chronic exposure to OHB also increased reactive oxygen species (ROS) production by 1.9-fold compared with control cells. To further understand the role of ROS in OHB action, cardiomyocytes were incubated with H(2)O(2). Our results demonstrated that this treatment diminished DNP-induced glucose uptake without altering activation of the AMPK/p38 MAPK signaling pathway. Incubation with the antioxidant N-acetylcysteine partially restored DNP-mediated glucose but not AMPK/p38 MAPK activation. In conclusion, these results suggest that ketone bodies, through inhibition of the AMPK/p38 MAPK signaling pathway and ROS overproduction, regulate DNP action and thus cardiac glucose uptake. Altered glucose uptake in hyperketonemic states during metabolic stress may contribute to diabetic cardiomyopathy.

2,4-Dinitrophenol↗
PubMed2006

Effect of 2,4-dinitrophenol under non-oxygenated condition in pons-medulla-spinal cord preparations in newborn rats: comparison with medulla-spinal cord preparations.

We tested whether depression of respiratory frequency (fR) under non-oxygenated artificial cerebrospinal fluid (aCSF) in pons-medulla-spinal cord (PMS) and medulla-spinal cord (MS) preparations is significantly influenced by the mitochondrial uncoupler 2,4-dinitrophenol (2,4-DNP) in newborn rats. Preparations were obtained from 0- to 4-day-old rats, and fR was monitored at the C4 ventral root in environmental temperature (24 degrees C). 2,4-DNP was dissolved in aCSF (1, 10 or 30 microM; pH 7.4), and we measured fR in PMS (n=19) and MS (n=16), both of which were superfused with aCSF equilibrated with oxygenated (95% O2-5% CO2) or non-oxygenated (10% O2-5% CO2, balanced with pure N2) gas. Our results showed that: (1) fR was significantly lower in PMS than MS, (2) fR was significantly decreased under non-oxygenated aCSF in both PMS and MS and (3) fR under non-oxygenated aCSF was significantly increased by 2,4-DNP applications at 10 and 30 microM in PMS but not in MS. Our results suggest that depression in fR under non-oxygenated aCSF in PMS and MS may not be due simply to O2 limitation, and 2,4-DNP has a stimulant effect on the medullary respiratory rhythm generator (RRG) through pontine RRG regulatory mechanisms under non-oxygenated aCSF.

2,4-Dinitrophenol↗
PubMed2006

Comblike dendrimers containing Tn antigen modulate natural killing and induce the production of Tn specific antibodies.

Comblike glycodendrimers were prepared by the chemoselective ligation of cysteine-modified glycopeptides (1-7) with a 3-maleimidopropionate-modified linear synthetic carrier (8). Glycodendrimers bearing mono-, di-, or tri-Tn clusters (9-11) were tested as inhibitors using plant and mammalian lectins. In the former group, the Codium fragile lectin showed moderate discrimination among 9, 10, and 11. In the latter group, A and B isoforms of rat NKR-P1 lectin strongly discriminated between 9 and 10. 10 caused a 4-fold increase in killing of the NK resistant tumor cell lines at concentrations as low as 10(-8) M. Surprisingly, 11 interacted exclusively with the rat NKR-P1B isoform and inhibited efficiently natural killing in both rats and humans, even in the presence of the activating compounds 9 and 10. Dinitrophenol haptenization or influenza virus hemagglutinin T-cell epitope conjugation increased the immunogenicity of the parent compounds and resulted in the production of Tn specific antibodies.

2,4-Dinitrophenol↗