Gallium arsenide (GaAs) is a semiconductor used in the electronics industry. GaAs exposure of animals induces profound immunosuppression. The major objective of this research proposal is to define the mechanism by which GaAs modulates antigen processing by macrophages for activation of antigen-specific helper CD4+ T cells. Previous toxicological studies with GaAs-exposed mice showed that splenic macrophages at a distant site exhibit defective antigen processing, whereas macrophages at the exposure site have an augmented processing ability. Both consequences of GaAs exposure correlate with altered cathepsin proteolysis and will be studied. The impact of intratracheal GaAs exposure will be defined by examining several aspects of antigen processing. The relationship between the structure of a particulate antigen and the processing defect of GaAs-exposed splenic macrophages will be determined by modifying sheep erythrocytes. One form will maintain the particulate nature of sheep cells, and another will disrupt cellular integrity. Sheep cells will be coated with antibody for Fc receptor-mediated entry and delivery to secondary lysosomes. Third, the contribution of GaAs components to the antigen processing defect will be determined. Splenic macrophages will be incubated in vitro with metal chelators to reverse the defect. Fourth, inhibition of cathepsins, the proteases mediating antigen processing, by GaAs components will be determined by kinetic analyses. Proteolytic reactivation by metal chelators will be assessed. GaAs' interference with the transport sorting signal of cathepsins will be examined. Fifth, a correlation between cytokine production and altered macrophage antigen processing will be investigated. A panel of cytokines with varying functions will be measured by ELISA. Mutant nude mice that lack functional T cells, an important cytokine source, will be exposed to GaAs, and antigen processing by alveolar and splenic macrophages will be determined. Finally, co-stimulation is another critical function of presenting cells for T cell activation. GaAs' effect on macrophage co-stimulation will be studied. Expression of co-stimulatory molecules on alveolar and splenic macrophages will be measured by two-color immunofluorescence staining and flow cytometry. The function of macrophage co-stimulatory molecules will be determined by cell titration and antibody blocking experiments. Knowledge concerning the mechanism by which GaAs influences macrophage functions may provide insights into preventing the chemical's impact on the immune system.
