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

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 t his assay, which comprises a sensitizing phase only, is also used to identi fy respiratory allergens. Another, experimentally used test in mice to iden tify allergens is also based on dermal exposure, but comprises both a sensi tizing and effector phase. In this latter test, it has been shown that cont act allergens preferentially induce a T-helper 1 (TH1) response, whereas re spiratory allergens preferentially induce a T-helper 2 (TH2) response. Thes e 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 b y TH2 cells. The aim of the study was to establish whether the UNA was suff icient to not only identify allergens but also mark them as either a contac t or a respiratory allergen. To this end, LLNA responses to the contact all ergen dinitrochlorobenzene (DNCB) and the respiratory allergen trimellitic anhydride (TMA) were determined using IFN-gamma and IL-4 mRNA expression an d production as parameters. Topical application of TMA resulted in a threef old 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 expres sion. While both DNCB and TMA induced IFN-gamma production, TMA but not DNC B induced IL-4 production. Thus, only IL-4 production seemed a suitable par ameter to discriminate between the two compounds. In a second study, the re spiratory allergens toluene-2,4-diisacyanate (TDI) and phthalic anhydride ( PA) were also assayed 7 days after the first application. Topical applicati on of DNCB and PA resulted in a similar lymphocyte proliferation, while app lication of TMA and TDI resulted in a 1.8-fold higher proliferation. IFN-ga mma production was similar for DNCB, TMA, and TDI, and fourfold lower for P A, while IL-4 production was similar for TMA, TDI, and PA, and 24-fold lowe r 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 applica tion. (C) 2000 Academic Press.