Phagocyte-derived reactive oxygen metabolites: stimulation by pathogenic mineral dusts

Abstract

The inhalation of dusts is accompanied by pulmonary infiltration of phagocytic cells, macrophages and polymorphonuclearcytes (PMN) (Jaurand et al., 1980). Cellular activation by particulates results in the production of reactive oxygen metabolites (ROM), namely superoxide and hydroxyl free radicals and hydrogen peroxide and hypochlorite metabolites, which are associated with a variety of tissue injurious effects (Slater, 1984). Among these, oxidative alterations in serum sulphydryl levels have been demonstrated in inflammatory coalworkers’ pneumoconiosis (Evans, 1977). Inhalation of pathogenic dusts, e.g. silica and asbestos, causes lung fibrosis and in the latter case, cancer. Similarly, environmental exposure to the fibrous zeolite mineral erionite has indicated its extremely potent carcinogenic activity both in epidemiological surveys in Karain, Turkey (Baris et al., 1981), and experimentally (Wagner et al., 1985). Comparable erionite deposits in western U.S.A. have also aroused health concerns (Rom et al., 1983). The involvement of ROM in collagen biosynthesis (Myllyla et al., 1979) and in tumour promotion by phorbol myristate acetate (Birnboim, 1982) indicates the potential causal importance of such toxic oxidants in dust-induced diseases. The present study has been undertaken to determine the capacity of sample dusts to stimulate the production of ROM by human PMN in vitro. PMN were purified from heparinized blood by sedimentation in dextran (2%) and subsequent centrifugation with Ficoll-Paque to remove erythrocytes, monocytes and lymphocytes. Residual erythrocytes were lysed with NHdCl/EDTA. The resultant purified (95%) PMN were washed twice with Ca2+-free buffer before being finally resuspended in Krebs-Ringer Hepes buffer, pH 7.35 ( 1 x lo6 PMN/ml). A specially constructed computerized luminometer was used to monitor the associated luminol-dependent chemiluminescence (CL). The relative weight-specific PMN CL activities of the dusts are shown (Fig. 1). As evidenced, there appears to be a general positive correlation between the known pathogenicity of the dusts and the PMN CL response, ‘inert’ titanium dioxide producing the least and the highly pathogenic erionite dusts the most, with silica and asbestos causing an intermediate response. Complementary investigations into the pertinent particulate parameters, e.g. size, shape, charge and composition, indicated the relationship to be a complex one. Whereas comminution of crocidolite fibres by milling decreased the CL response, erionite dusts, which contain comparatively few fibres, exhibited very high activity. The possible effect of contaminating metals, e.g. ‘catalytic’ iron (Gutteridge et al., 1982), in mediating free-radical generation remains to be clarified. Accompanying studies into the cellular mechanisms of stimulation (Hallett & Campbell, 1983) showed that the microfilament reactant and phagocytic inhibitor cytochalasin B (1OpM) (Axilene & Reaven, 1974) largely abolished the particle-induced PMN CL. However, addition of the membrane receptor-mediated adenosine analogue 2-chloroadenosine (1OpM) (Roberts et al., 1985) which inhibits chemotactic peptide (N-formyl-L-methionylL-phenylalanine) stimulated PMN, produced no comparable reduction in dust-activated PMN CL.