Reactive oxygen species (ROS) and nitric oxide (NO) have an inhibitory effect on Ag mediated mast cell degranulation but there is controversy over whether mast cells themselves can produce these immunoregulatory molecules. We investigated the intracellular generation of ROS/NO in mast cells and examined the effect of such production on degranulation.

Mast cells were purified from OVA-sensitized BN rats and cultured overnight with inhibitors of ROS/NO. Following a 1 h incubation with the intracellular probes DCF (ROS nonspecific) or DAF-2 (NO specific), cells were stimulated with either OVA or LPS/IFN-γ and fluorescence monitored over the next 24-48 h by flow cytometry. Peritoneal macrophages were used as a positive control. Degranulation was monitored by release of incorporated serotonin and changes in light side scatter.

Mast cells generated intracellular ROS following OVA challenge and this was accompanied by degranulation. Both ROS production and degranulation were inhibited by the flavoenzyme inhibitor DPI but not by the intracellular superoxide dismutase scavenger MnTmPyP. Challenge of mast cells with OVA or LPS + IFN-γ over a wide range of time periods failed to generate intracellular NO whereas peritoneal macrophages generated NO readily in response to LPS + IFN-γ.o:p>

Rat peritoneal mast cells generate intracellular ROS but not NO and a non-superoxide intracellular ROS product facilitates Ag-induced mast cell degranulation.