Detection of and discrimination between total and free human interleukin-4 and free soluble interleukin-4 receptor by ELISA

Jung, Thomas; Bews, John P.A.; Enssle, Karl-Heinz; Wagner, Katrin; Neumann, Christine; Heusser, Christoph

doi:10.1016/s0022-1759(98)00084-2

Cited by 13 publications

(14 citation statements)

References 28 publications

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“…IL-10 and IL-5 are potent Th2 cytokines involved in autoantigen-induced tolerance in EAE (36) and in drug-induced immune suppression in EAE (37) and MS (38). Although IL-4 production was undetectable, probably due to a rapid and pronounced uptake of IL-4 by high affinity IL-4Rs on the cells in the culture (39,40), the glucosamine-induced increase in IL-5 and IL-10 production ex vivo and in vitro (see below) indicates that glucosamine abrogates EAE not simply as an immunosuppressant, but as an immunomodulator. Administration of glucosamine, thus, provides a novel immunoregulatory approach in autoimmune disorders.…”

Section: Discussionmentioning

confidence: 99%

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

Zhang

Gran

et al. 2005

The Journal of Immunology

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Glucosamine, a natural glucose derivative and an essential component of glycoproteins and proteoglycans, has been safely used to relieve osteoarthritis in humans. Recent studies have shown that glucosamine also possesses immunosuppressive properties and is effective in prolonging graft survival in mice. Whether this reagent is effective in human multiple sclerosis (MS), an inflammatory demyelination in the CNS, is not known. We thus investigated the therapeutic effect of glucosamine on experimental autoimmune encephalomyelitis (EAE), an animal model of MS. We demonstrated that oral, i.p., or i.v. administration of glucosamine significantly suppressed acute EAE, with reduced CNS inflammation and demyelination. A significant, albeit not strong, blockade of Th1 response and an up-regulation of Th2 cytokines (IL-5 and IL-10) are observed in the splenocytes of glucosamine-treated mice. Glucosamine also regulates IL-5 and IL-10 in vitro. As glucosamine is able to effectively suppress acute EAE, has low or absent toxicity, and has been safely used in humans orally, our study suggests a potential use for this drug alone or in combination with other disease-modifying immunotherapies to enhance their efficacy and reduce their doses in MS and possibly other autoimmune disorders. Furthermore, because glucosamine functions not simply as an immunosuppressant, but as a mild immunomodulator, administration of glucosamine provides a novel immunoregulatory approach for autoimmune disorders.

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Section: Discussionmentioning

confidence: 99%

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

Zhang

Gran

et al. 2005

The Journal of Immunology

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“…Samples were measured in triplicates on a Wallac Betacounter. The neutralizing anti-IL-4 mAb 8F12 and anti-IL-4 mAb 3H4 were a kind gift from C. Heusser (Novartis, Basel, Switzerland) [44]. The ELISA for determination of total IL-4 was done as described [44].…”

Section: Cells Cell Culture Antibodies Proliferationmentioning

confidence: 99%

“…The neutralizing anti-IL-4 mAb 8F12 and anti-IL-4 mAb 3H4 were a kind gift from C. Heusser (Novartis, Basel, Switzerland) [44]. The ELISA for determination of total IL-4 was done as described [44]. The following antibodies were used for intracellular staining: anti-STAT6, anti-STAT5, anti-SHP-2 (Transduction Laboratories), anti-GATA3, anti-JAK3 (Santa Cruz), anti-IRS-2 (Upstate Biotechnology).…”

Section: Cells Cell Culture Antibodies Proliferationmentioning

confidence: 99%

CD45 isoform expression is associated with different susceptibilities of human naive and effector CD4⁺ T cells to respond to IL‐4

et al. 2005

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Interleukin-4 (IL-4) is the major factor promoting the development of T helper type 2 (Th2) cells from naive precursor T cells. Minute amounts of IL-4 produced by naive T cells seem to be sufficient; however, the molecular mechanisms explaining this efficient utilization of are not yet known. Here, we show that human CD4 + CD45RA + naive T cells, in contrast to CD4 + CD45R0 + effector T cells, show responsiveness to endogenous as well as exogenous IL-4 to proliferate and differentiate towards Th2 cells in vitro. Despite production levels of IL-4 below conventional detection limits, CD45RA + T cell-derived IL-4 could clearly activate STAT6. Although the expression levels of IL-4R and STAT6 were not different between naive and effector T cells, only naive T cells responded to IL-4 in a STAT6-dependent reporter gene assay. Transfecting a trans-dominant negative form of STAT6 abrogated IL-4-induced proliferation in CD45RA + cells. A significantly higher protein tyrosine phosphatase (PTPase) activity was detected in CD45R0 + T cells as compared to CD45RA + T cells. Cross-linking CD45 potently reduced PTPase activity in CD45R0 + T cells and restored their ability to proliferate in response to IL-4. Thus, CD45 PTPase activity contributes to the susceptibility of naive and memory T cells to respond to IL-4.

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“…Cytokines are often bound to various molecules present in biological fluids (such as soluble receptors, a 2 -macroglobulin and autoantibodies), and these are usually present in excess [72][73][74][75][76][77][78]. A cytokine may not be recognised by the capture antibody of the assay if the relevant epitope is hidden when the cytokine is bound within a complex.…”

Section: Factors Specific To the Soluble Mediatormentioning

confidence: 99%

“…If the lines are not parallel, the presence of a substance that interferes with recognition of mediator epitopes by the immunoassay and which is dilutable is suggested. This may be due to masking of epitopes by autoantibodies [78,115], soluble receptors [72,73,116,117] or other binding proteins such as albumin and a 2 -macroglobulin [78,86]. Specific assays are available for the soluble receptors of only certain cytokines, e.g.…”

Section: What To Do If Validity Is Poormentioning

confidence: 99%

Analysis of fluid phase mediators

Keatings

Leigh²,

Peterson³

et al. 2002

European Respiratory Journal

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Sputum cellular indices are valid, reliable and responsive to change [1][2][3][4][5][6][7]. Increasingly, numerous inflammatory mediators are being measured in the fluid phase of sputum; these include cytokines, chemokines, granulocyte proteins, markers of vascular leakage, eicosanoids, proteases and others. Many of these mediators are included in table 1, which gives details of methods used by investigators to process sputum and measure mediator levels. The table also provides the median/mean levels measured in studied subject groups to give an indication of the expected levels of these mediators in sputum. However, the reproducibility, precision and validity of many of these measurements in sputum have not been investigated and, therefore, their utility as a research and clinical tool remains uncertain and requires confirmation.The following issues are important in the analysis of mediators: 1) choice of methods for measuring fluidphase mediators; 2) points to consider when planning an immunoassay; and 3) evaluation of the measurement of a soluble mediator in sputum, i.e. validation of the measurement method. Methods for measuring fluid-phase mediatorsThe three main types of method used for the measurement of sputum fluid-phase mediators are bioassays, enzyme assays and immunoassays. BioassaysBioassays rely on the retention of biological activity and the ability to exert a measurable effect, such as proliferation of cells, bone marrow colony formation or chemotaxis [3,11,58]. Sputum processing with mucolytics such as dithiothreitol (DTT) or dithioerythritol, which are strong reducing agents, may decrease the biological activity of cytokines, many of which rely on disulphide bonds to provide a stable structure for bioactivity. Bioassays also have the disadvantage of being inconvenient, time-consuming and lacking in specificity [59]. In addition, the presence of commonly occurring endogenous cytokine inhibitors, although allowing an estimate of net activity, may result in significant underestimation of total cytokine levels [58]. Enzyme assaysMany of the mediators of the inflammatory response are enzymes, released from cellular sites of synthesis into an environment replete with enzyme inhibitors. Again, an estimate of net activity is important when evaluating their potential contribution to tissue responses. The proteases neutrophil elastase [20,22,60,61], cathepsin G [20, 61] and cathepsin B [22] have been assayed in sputum using specific chromogenic substrates, from which proteases release a coloured product that can be quantified spectrophotometrically. Net protease activity is determined using purified enzyme standards. Active forms of matrix metalloproteinases in sputum are identified by means of substrate gel zymography, and net activity can be quantified using radiolabelled substrates [62]. Other proteases, such as chymase and tryptase, are less robust. Their activity rapidly diminishes on freezing/thawing sputum samples, and immunoassays are the best means of detecting them (see below).The activity of e...

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Detection of and discrimination between total and free human interleukin-4 and free soluble interleukin-4 receptor by ELISA

Cited by 13 publications

References 28 publications

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

CD45 isoform expression is associated with different susceptibilities of human naive and effector CD4⁺ T cells to respond to IL‐4

Analysis of fluid phase mediators

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Detection of and discrimination between total and free human interleukin-4 and free soluble interleukin-4 receptor by ELISA

Cited by 13 publications

References 28 publications

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

Glucosamine Abrogates the Acute Phase of Experimental Autoimmune Encephalomyelitis by Induction of Th2 Response

CD45 isoform expression is associated with different susceptibilities of human naive and effector CD4+ T cells to respond to IL‐4

Analysis of fluid phase mediators

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CD45 isoform expression is associated with different susceptibilities of human naive and effector CD4⁺ T cells to respond to IL‐4