T-cell activation by organic dust in vitro

Abstract

Inhalation of swine dust causes intense airway inflammation with a multifold increase of inflammatory cells and lymphocyte activation as assessed by bronchoalveolar lavage. To further investigate the mechanism for lymphocyte activation the present in vitro study focuses on the lymphocyte response to swine dust in whole blood. Various concentrations of phytohaemagglutinin (PHA) (final concentrations: 3·16, 10·0, 3·16 and 100μg ml-1) and swine dust (final concentrations: 10·0, 31·6, 100 and 316 μg ml-1) were added to heparinized whole blood from healthy donors. The blood samples were incubated in duplicate, using the homologous unstimulated blood as control, for 4, 24, 48 and 72h in a water bath at 37°C. The cells were stained with fluorochrome conjugated monoclonal antibodies. For analysis of T-cell activation CD3 was doublestained together with the activation markers CD69, CD25 and HLADR. Cell count percentages were analysed by flow cytometry. Soluble IL-2sRα in plasma was analysed using commercial sandwich ELISA techniqe. At baseline CD69, CD25 and HLA-DR were expressed in < 1%, approx 5% and <1% of the T-cells respectively. We found a dose response relationship between swine dust exposure and the expression of all three T-cell activation markers which appeared at different time-points. Maximal expression of CD69 (8%, P<0·05) and CD25 (15%, P<0·001) was found after 24h of activation. HLA-DR was significantly expressed after 48h (8%) and maximally expressed after 72h of activation (13%, P<0·05). The soluble IL-2sRα in plasma was maximally expressed after 24-48h (1200 pg ml-1 and 1500 pg ml-1, respectively In conclusion, T-cells were activated by swine dust in vitro. Thus, our previous findings of T-cell activation following swine dust exposure, in vivo may be an effect of the dust either directly on T-cells or on other cells which in turn contribute to the T-cell activation.