Utilization of the Ex Vivo LLNA

Abstract

Dermal exposure to chemicals may result in allergic or irritant contact dermatitis. In this study, we performed ex vivo local lymph node assay: bromodeoxyuridine-enzyme-linked immunosorbent assay (LLNA: BrdU-ELISA) to compare the differences between irritation and sensitization potency of some chemicals in terms of the 3 end points: lymphocyte proliferation, cytokine profiles (interleukin 2 [IL-2], interferon-γ (IFN-γ), IL-4, IL-5, IL-1, and tumor necrosis factor α [TNF-α]), and ear swelling. Different concentrations of the following well-known sensitizers and irritant chemicals were applied to mice: dinitrochlorobenzene, eugenol, isoeugenol, sodium lauryl sulfate (SLS), and croton oil. According to the lymph node results; the auricular lymph node weights and lymph node cell counts increased after application of both sensitizers and irritants in high concentrations. On the other hand, according to lymph node cell proliferation results, there was a 3-fold increase in proliferation of lymph node cells (stimulation index) for sensitizer chemicals and SLS in the applied concentrations; however, there was not a 3-fold increase for croton oil and negative control. The SLS gave a false-positive response. Cytokine analysis demonstrated that 4 cytokines including IL-2, IFN-γ, IL-4, and IL-5 were released in lymph node cell cultures, with a clear dose trend for sensitizers whereas only TNF-α was released in response to irritants. Taken together, our results suggest that the ex vivo LLNA: BrdU-ELISA method can be useful for discriminating irritants and allergens.