The OX-81monoclonal antibody reacts with rat IL-4（interleukin-4）amultifunctional14kDa cytokine。IL-4is expressed primarily by activated Th2cells and NK cells，and at lower levels by mast cells，and basophils。IL-4signals through the IL-4Rα。Upon receptor binding IL-4stimulates activated B and tlymphocyte proliferation，and the differentiation of B cells into plasma cells。It also induces B cell class switching to IgE，and up-regulates MHC class II production while decreasing the production of Th1 cells，macrophages，IFNγ，and dendritic cell IL-12。Like other Th2associated cytokines，IL-4is involved in the airway inflammation observed in the lungs of patients with allergic asthma。The OX-81antibody has been shown to neutralize the bioactivity of rat IL-4。
Mitsuoka N，Iwagaki H，Ozaki M，Sheng SD，Sadamori H，Matsukawa H，Morimoto Y，Matsuoka J，Tanaka N，Yagi T.（2004）。“Theimpact of portal infusion with donor-derived bone marrow cells and intracelllar cytokine expression of graft-infiltrating lymphocytes on the graft survival in rat small bowel transplant model”Transpl Immunol13（3）：155-60。PubMed
Intraportal administration of alloantigen is reported to reduce antigen-specific immune responses，although the underlying mechanisms for the reduced immunological reactions，especially those of the graft，are poorly understood。We examined intracelllular cytokine production by graft-infiltrating lymphocytes（GILs）and peripheral blood lymphocytes（PBLs）to elucidate the underlying mechanism of beneficial effects on intraportal infusion of donor cells inrat small bowel transplantation（SBT）。
ELISA
Salomon I，Netzer N，Wildbaum G，Schif-Zuck S，Maor G，Karin N.（2002）。“Targating the function of IFN-gamma-inducible protein10 suppresses ongoing adjuvant arthritis”J Immunol169（5）：2685-93。PubMed
IFN-gamma-inducible protein10（IP-10）isa CXC chemokine that is thought to manifest a proinflammatory role because it stimulates the directional migration of activated T cells，particularly Th1cells。It is an open question whether this chemokine is also directly involved in T cell polarization。We show here that during the course of adjuvant-induced arthritis the immune system mounts a notable Ab titer against self-IP-10。Upon the administration of naked DNA encoding IP-10，this titer rapidly accelerates to provide protective immunity。Self-specific Ab to IP-10developed in protected animals，as well as neutralizing Ab to IP-10that we have generated in rabbits，could inhibit leukocyte migration，alter thein vivoand，andin vitroTh1/Th2balance toward low IFN-gamma，low TNF-alpha，high IL-4-producing T cells，and adoptively transfer disease suppression。This not only demonstrates the pivotal role of this chemokine in T cell polarization during experimentally induced arthritis but also suggests a practical way to interfere in the regulation of disease to provide protective immunity。自支撑式通风装置，this study challenges the paradigm ofin vivoredundancy。After all，we did not neutralize the activity of other chemokines that bind CXCR3（i.e.，macrophage-induced gene and IFN-inducible T cell alpha chemoattractant）and yet significantly blocked not only adjuvant-induced arthritis but also thein vivocompetence to mount delayed-type hypersensitivity。
in vivoIL-4neutralization
Tran GT，Carter N，He XY，Spicer TS，Plain KM，Nicolls M，Hall BM，Hodgkinson SJ.（2001）。“Reversal of experimental allergic encephalomyelitis with non-mitogenic，non-depleting anti-CD3mAb therapy with a preferential effect on T（h）1cells that is augmented by IL-4”Int Immunol13（9）：1109-20。PubMed
This study examined whether therapy with anon-mitogenic，non-activating anti-CD3mAb（G4.18）alone，or in combination with the T（h）2cytokines，could inhibit induction or facilitate recovery from experimental allergic encephalomyelitis（EAE）in Lewis rats。G4.18，but not rIL-4，rIL-5 or anti-IL-4mAb，reduced the severity and accelerated recovery from active EAE。A combination of rIL-4with G4.18was more effective than G4.18alone。The infiltrate of CD4（+）and CD8（+）T cells，B cells，dendritic cells，and macrophages in the brain stem was less with combined G4.18and IL-4than G4.18therapy or no treatment。Residual cells had preferential sparing of T（r）1cytokines IL-5 and transforming growth factor-beta with loss of T（h）1markers IL-2，IFN-gamma and IL-12Rbeta2，and the T（h）2cytokine IL-4as well as macrophage cytokines IL-10and tumor necrosis factor-alpha。Lymph nodes draining the site of immunization had less mRNA for T（h）1cytokines，but T（h）2and T（r）1cytokine expression was spared。Treatment with G4.18，rIL-4 or rIL-5 from the time of immunization had no effect on the course of active EAE。AC OX-81，amAb that blocks IL-4，delayed onset by2days，but had no effect on severity of active EAE。G4.18also inhibited the ability of activated T cells from rats with active EAE to transfer passive EAE。This study demonstrated that T cell-mediated inflammation was rapidly reversed byanon-activating anti-CD3mAb that blocked effector T（h）1cells，and spared cells expressing T（h）2andT（r）1cytokines。
ELISA
Uchikawa R，Matsuda S，Arizono N.（2000）。“Suppression of gamma interferon transcription and production by nematode excretory-secretory antigen during polyclonal stimulation of rat lymph node T cells”Infect Immun68（11）：6233-9。PubMed
Although certain helminth infections preferentially induce type2T-cell responses，the immunological mechanism responsible for type2T-cell polarization remain unclear。In the present study，we investigated the effects of excretory-secretory（ES）antigen from the nematode Nippostrongylus brasiliensis on cytokine production by mesenteric lymph node（MLN）cells isolated from naiverats。LN cells produced considerable levels of gamma interferon（IFN-gamma）during a72-h stimulation with concanavlin A（ConA）or with immobilized anti-CD3plus soluble anti-CD28antibodies（anti-CD3/CD28）。With either stimulation，10microg of ES antigen per ml significantly suppressed IFN-gamma and interleukin-2（IL-2）production without cytotoxic activity。The copresence of anti-IL-4，anti-IL-10，or transforming growth factor beta（TGF-beta）blocking antibodies did not alter the suppressive effect of ES antigen on IFN-gamma production。ES antigen did not affect IL-10 production。Kinetic studies of the effect of ES antigen indicated that the antigen suppressed even ongoing IFN-gamma production。Reverse transcription-PCR study showed that in the presence of ES antigen，IFN-gamma mRNA expression by MLN cells was suppressed6and12h after ConA or anti-CD3/CD28 stimulation。ES antigen also significantly suppressed IFN-gamma production by purified CD4（+）or CD8（+）T cells during anti-CD3/CD28 stimulation but did not affect IL-4 production by CD4（+）T cells。These findings suggested that the nematode antigen suppressed production of IFN-gamma and IL-2but not IL-4or IL-10 production。ES antigen-mediated suppression of IFN-gamma during the initiation of the immune response may provide a microenvironment that helps generation of type2T cells。
in vivoIL-4neutralization
Saoudi A，Simmonds S，Huitinga I，Mason D.（1995）。“Prevention of experimental allergic encephalomyelitis in rats by targeting autoantigen to B cells:evidence that the protective mechanism depends on changes in the cytokine response and migratory properties of the autoantigen-specific T cells”J Exp Med182（2）：335-44。PubMed
Previous experiments from experimental allergic encephalomyelitis（EAE）induced by the injection of myelin basic protein（MBP）in Freund's complete adjuvant if they were treated with the encephalitogenic peptide of MBP covalently to mulin g）D。It was suggested that this protection developed because the antibody-peptide conjugate targeted the peptide to B cells and that this mode of presentation induced a Th2-like T cell response that controlled the concomitant encephalitogenic Th1 reaction to the autoantigen。The current experiments were carried out to test this hypothesis and to examine the alternative explanation for the protective effect of the conjugate pretreatment，namely that it induced a state of nonresponsiveness in the autoantigenspecific T cells。It was shown that EAE induction was suppressed in Lewis rats when the antibody-peptide conjugate was injected intravenously14and7d before immunization with MBP in adjuvant，but that antibody titers were at least as high in these animals as in controls that were not pretreated the munith。Lymph node cells from these pretreated animals，while proliferatingin vitroto MBP as vigorously as those from controls，produced less interferon gamma and were very inferior in their ability to transfer disease after thisin vitroactivation。In contrast，these same lymph node cells from protected rats generated markedly increased levels of messenger RNA for interleukin（IL）-4and IL-13。When thesein vitroexperiments were repeated using the encephalitogenic peptide rather than MBP as the stimulus，the proliferative response of lymph node cells from pretreated donors was less than that from controls but was still readily detectable in the majority of experiments。Furthermore，the cytokine expression induced by the peptide was similar to that elicited by whole MBP。While these results support the original hypothesis that the anti-IgD-peptide conjugate pretreatment protected rats from EAE by inducing a Th2-type cytokine response，attally unexpected finding was that this pretreatment greatly reduced the level of leukocyte infiltration into the cento the centrastem。This result provides a direct explanation for the protective effect of the pretreatment，but it raises questions regarding migratory and homing patterns of leukocytes activated by different immunological stimuli。
