Studies on identification of environmental substances resulting in human respiratory diseases.

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In this study, we concentrated identification of environmental substances resulting in human respiratory diseases on 3 matters : (a) incorporation of environmental substances into macrophages, (b) production of cytokines in the macrophages, and (c) production of superoxide induced by cytokines. Since it was difficult to obtain human alveolar macrophages routinely, we substituted human peripheral blood monocytes. Similarly, we used bacterial cell wall skeletones in stead of environmental substances. Results are as follows : (1) Both neutrophiles and monocytes were able to be separated from other cells in purity of more than 90%. (2) Monocytes phagocytized the fluorescence-labeled cell wall skeletones dose-dependently. (3) Production of cytokines in the monocytes activated by phagocytosis of the cell wall skeletones increased dose-dependently, whereas there was difference in its peak time and amount between cytokines. (4) There was difference also in potency of superoxide production among cytokines; GM-CSF stimulated superoxide production most prominently, G-CSF, TNF-alpha, IL-6, IL-2 and IL-1beta did prominently next to GM-CSF, and any of IL-1alpha, IL-1beta and IFN-alpha scarcely stimulated.The study on relationship between superoxide production and proliferation of fibroblast has not been carried out, we, however, have established in vitro methods to measure incorporation of environmental substances into macrophages, secretion of cytokines from macrophages and stimulation of superoxide production by cytokines.In the near future, we would like to establish new leukemic cell lines with higher potency of the cytokine and superoxide production after their differentiation. These cell lines may be expected to get our method more general.