Regulation Of Cytokine Gene Expression In Mast Cells

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Abstract

In immunity, reactive oxygen species (ROS) and nitric oxide (NO) are important antimicrobial agents and regulators of cell signaling and activation pathways. Our aim was to determine the regulatory roles of such chemical species on mast cell activation. To determine the capability of mast cells to endogenously generate ROS and NO following IgE/antigen stimulation, we employed flow cytometry using the respective cell-permeable fluorescent probes dichlorodihydrofluorescein and diaminofluorescein. In rat peritoneal mast cells, mouse bone marrow-derived mast cells and human blood-derived mast cells, degranulation stimulated by IgE/antigen was accompanied by production of intracellular ROS but not NO. Likewise, bacterial lipopolysaccharide and interferon-gamma over a wide range of conditions failed to generate intracellular NO in mast cells, whereas these agents readily induced intracellular NO in macrophages. NO synthase protein, as assessed by Western blotting, was readily induced in macrophages but not mast cells. We concluded that rodent and human mast cells generate intracellular ROS but not NO and that intracellular ROS but not intracellular NO are functionally linked to mast cell degranulation. In terms of the effect of exogenous NO derived from other cells types on mast cell function, it has been reported that NO inhibits mast cell degranulation and subsequent allergic inflammation, although whether its action is nonspecific or occurs via specific molecular mechanisms remains unknown. To examine this question, we set out to determine whether NO inhibits mast cell cytokine production, and, if so, whether it also alters Fc RI-dependent signal transduction. As hypothesized, the radical inhibited IgE/Ag-induced IL-4, IL-6, and TNF production. Although NO did not influence phosphorylation of JNK, p38 MAPK, or p44/42 MAPK, it did inhibit phosphorylation of phospholipase C 1 and the AP-1 transcription factor protein c-Jun, but not NF- B or CREB. NO further completely abrogated IgE/Ag-induced DNA-binding activity of the nuclear AP-1 proteins Fos and Jun. These results show that NO is capable of inhibiting Fc RI-dependent mast cell cytokine production at the level of gene regulation, and suggest too that NO may contribute to resolution of allergic inflammation.