Occupational exposure by inhalation to vanadium-containing particles such a
s residual oil fly ash results in respiratory tract inflammation. This infl
ammation, characterized by abundant neutrophilia, appears to be initiated b
y alveolar macrophages (AMs) encountering particles and the subsequent rele
ase of proinflammatory cytokines. Intracellular signaling events in these c
ells in response to particles or their components are largely unknown. We i
nvestigated two immediate responses of AMs to vanadium exposure in vitro, t
he production of reactive oxygen intermediates (ROI) or respiratory burst (
RB), and the tyrosine phosphorylation of cellular proteins. Macrophages exp
osed in vitro to 100 mu M vanadyl chloride/1 mu Gi V-48 incorporated 8.3% o
f the metal after 30 min. Exposure of AMs to increasing concentrations of s
odium metavanadate resulted in a dose-dependent increase in production of R
OI as measured by dichlorofluorescin oxidation. The lowest dose yielding a
significant response was 50 mu M, whereas 1000 mu M increased RE activity b
y 173%. NADPH oxidase inhibitors deoxy-D-glucose (100 mM) and diphenylene i
odonium (25 mu M) reduced the metavanadate-induced RE by 62 and 71%, respec
tively, implicating NADPH oxidase as the primary cellular source of ROI. En
hanced cerium chloride oxidation in response to metavanadate localized to t
he plasma membrane consistent with increased NADPH oxidase activity. Pretre
atment of AMs with the epidermal growth factor receptor inhibitor, tryphost
in B50 (10 mu M), reduced the metavanadate-induced RE, but did not influenc
e overall tyrosine phosphorylation, Metavanadate and H2O2 exposure greatly
increased overall tyrosine phosphorylation, yielding a similar but distingu
ishable pattern of phosphorylation in these cells. These observations demon
strate that in vitro metavanadate exposure regulates two distinct, yet rela
ted intracellular signaling pathways important in initiating inflammatory r
esponses in these cells: (1) activation of the NADPH oxidase complex with s
ubsequent increased ROI synthesis, and (2) enhanced tyrosine phosphorylatio
n of cellular proteins. (C) 1999 Academic Press.