An essential role of NAD(P)H oxidase 2 in UVA-induced calcium oscillations in mast cells

Abstract

Solar UVA radiation (320–400 nm) is known to have immunomodulatory effects, but the detailed mechanisms involved are not fully elucidated. UVA irradiation has been shown to induce calcium oscillations in rat peritoneal mast cells due to NAD(P)H oxidase (NOX) activation, but the specific NOX isoforms have not been identified. In the present work effects of UVA irradiation were investigated in isolated rat peritoneal mast cells, in cultured rat mast cell line RBL-2H3, and in mouse bone marrow-derived mast cells (BMMC). It was found that UVA irradiation by alternate 340/380 nm (3.2–5.6 μW cm⁻²) or by LED (380 nm, 80 μW cm⁻²) induced calcium oscillations in isolated rat peritoneal mast cells, in RBL-2H3, and in BMMC. Such UVA-induced calcium oscillations resembled closely those induced by surface IgE receptor (FcεRI) activation. It was found that RBL-2H3 expressed high levels of gp91^phox (NOX2), p22^phox, p67^phox, p47^phox, p40^phox, Rac1, Rac2, moderate levels of DUOX2, but did not express NOX1, NOX3, NOX4, or DUOX1. The specific cellular localizations of gp91^phox (NOX2), p22^phox, p47^phox, p67^phox, p40^phox and Rac1/2 were confirmed by immunocytochemistry. UVA-induced reactive oxygen species (ROS) production in RBL-2H3 was completely suppressed by the NOX inhibitor diphenyleneiodonium chloride (DPI) or by the antioxidant N-acetyl-L-cysteine (NAC). siRNA suppression of gp91^phox (NOX2), p22^phox and p47^phox expression inhibited markedly UVA-induced calcium oscillations, ROS and IL-6/LTC₄ production in RBL-2H3. Taken together these data indicate that NOX2 plays an essential role in UVA irradiation-induced calcium oscillations, ROS and mediator production in mast cells.