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Induction of Expression of Genes Encoding Components of the Respiratory Burst Oxidase during Differentiation of Human Myeloid Cell Lines Induced by Tumor Necrosis Factor and -Interferon¹

The Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104 [J. W. G., M. K., L. H., G. T.], and Istituto di Patologia Generale, University of Verona, 37134 Verona, Italy [M. C.]

In HL-60 and ML-3 human myeloid cell lines, -interferon (IFN-) and/or tumor necrosis factor (TNF) induce synergistic accumulation of transcripts of the genes encoding the heavy chain (gp91-phox) of cytochrome b₅₅₈ and the cytosolic factors p47-phox and p67-phox, components of the superoxide-generating NADPH oxidase system. The accumulation of transcripts for gp91-phox and p47-phox, as quantitated at the single-cell level by in situ hybridization, is extremely heterogeneous; however, when the cells are stimulated by IFN- and TNF together, most or all the cells in the induced cultures express higher accumulation of gp91-phox and p47-phox transcripts than cells from uninduced culture. In situ hybridization was performed on cellular subsets separated by fluorescence-activated cell sorting on the basis of surface expression of differentiation antigens or respiratory burst activity. The accumulation of gp91-phox and p47-phox transcripts correlated positively with the expression of the CD14 and CD11b antigens, two markers expressed on mature myelomonocytic cells. Similarly, accumulation of the two transcripts correlated with respiratory burst activity in cells separated by fluorescence-activated cell sorting after being loaded with dichlorofluorescein diacetate and stimulated with 12-O-tetradecanoylphorbol-13-acetate. These results suggest that all the cells in the culture are induced to differentiate by TNF and IFN- but that at the time of analysis there is heterogeneity in the level of differentiation and a proportion of cells is present that shows more mature characteristics with a coordinate expression of the various differentiation markers and functions.

¹ Supported in part by U.S.P.H.S. Grants CA 10815, CA 20833, CA 32898, and CA 40256, by Consiglio Nazionale delle Richerche Grants 89.012781.04 and 90.00053.70, and by a North Atlantic Treaty Organization travel grant.

² To whom requests for reprints should be addressed, at The Wistar Institute, 3601 Spruce Street, Philadelphia, PA 19104.

Received 12/12/91. Accepted 2/21/92.

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