title: Assessment of preferential T-helper 1 or T-helper 2 induction by low molecular weight compounds using the local lymph node assay in conjunction with RT-PCR and ELISA for interferon-gamma and interleukin-4.
abstract: The local lymph node assay (LLNA) is a new and promising test in mice used to identify contact allergens by means of dermal exposure. Experimentally this assay, which comprises a sensitizing phase only, is also used to identify respiratory allergens. Another, experimentally used test in mice to identify allergens is also based on dermal exposure, but comprises both a sensitizing and effector phase. In this latter test, it has been shown that contact allergens preferentially induce a T-helper 1 (TH1) response, whereas respiratory allergens preferentially induce a T-helper 2 (TH2) response. These responses can be discriminated on the basis of cytokine production, such as IFN-gamma, which is produced by TH1 cells, and IL-4, which is produced by TH2 cells. The aim of the study was to establish whether the LLNA was sufficient to not only identify allergens but also mark them as either a contact or a respiratory allergen. To this end, LLNA responses to the contact allergen dinitrochlorobenzene (DNCB) and the respiratory allergen trimellitic anhydride (TMA) were determined using IFN-gamma and IL-4 mRNA expression and production as parameters. Topical application of TMA resulted in a threefold higher lymphocyte proliferation compared to DNCB 3 and 5 days after the first application, while a similar proliferation was found from Day 7 and onward. RT-PCR showed a similar induction of IFN-gamma and IL-4 mRNA expression. While both DNCB and TMA induced IFN-gamma production, TMA but not DNCB induced IL-4 production. Thus, only IL-4 production seemed a suitable parameter to discriminate between the two compounds. In a second study, the respiratory allergens toluene-2,4-diisocyanate (TDI) and phthalic anhydride (PA) were also assayed 7 days after the first application. Topical application of DNCB and PA resulted in a similar lymphocyte proliferation, while application of TMA and TDI resulted in a 1.8-fold higher proliferation. IFN-gamma production was similar for DNCB, TMA, and TDI, and fourfold lower for PA, while IL-4 production was similar for TMA, TDI, and PA, and 24-fold lower for DNCB. In summary, both studies showed induction of IL-4 production by respiratory allergens, with little or no induction by the contact allergen, holding promise for the possibility of identifying respiratory allergens within the LLNA by measuring IL-4 production 7 days after the first application.
authors.affiliation: Laboratory for Pathology and Immunobiology, National Institute of Public Health and the Environment, Bilthoven, 3720 BA, The Netherlands. r.vandebriel@rivm.nl
authors.forename: R J
authors.initials: RJ
authors.lastname: Vandebriel
authors.forename: W H
authors.initials: WH
authors.lastname: De Jong
authors.forename: S W
authors.initials: SW
authors.lastname: Spiekstra
authors.forename: M
authors.initials: M
authors.lastname: Van Dijk
authors.forename: A
authors.initials: A
authors.lastname: Fluitman
authors.forename: J
authors.initials: J
authors.lastname: Garssen
authors.forename: H
authors.initials: H
authors.lastname: Van Loveren
mesh_terms: D000485:Allergens / Q000494:pharmacology*; D000818:Animals; D002478:Cells, Cultured; D004137:Dinitrochlorobenzene / Q000276:immunology / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007371:Interferon-gamma / Q000096:biosynthesis*; D015847:Interleukin-4 / Q000096:biosynthesis*; D008198:Lymph Nodes / Q000166:cytology / Q000187:drug effects* / Q000276:immunology* / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D010796:Phthalic Anhydrides / Q000276:immunology / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018417:Th1 Cells / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D018418:Th2 Cells / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D014051:Toluene 2,4-Diisocyanate / Q000276:immunology / Q000494:pharmacology
chemical_list: D000485:Allergens; D004137:Dinitrochlorobenzene; D010796:Phthalic Anhydrides; D012333:RNA, Messenger; D014051:Toluene 2,4-Diisocyanate; D015847:Interleukin-4; C015559:trimellitic anhydride; D007371:Interferon-gamma; C043103:phthalic anhydride