Pneumocystis carinii stimulates tumor necrosis factor-α release from alveolar macrophages through a β-glucan-mediated mechanism

Recent studies suggest that TNF-α plays a central role in host defenses during Pneumocystis carinii pneumonia. To determine whether P. carinii directly stimulates TNF-α secretion, rat alveolar macrophages were cultured in the presence of purified P. carinii. Whereas unstimulated alveolar macrophages released only 13.0 ± 2.7 pg/ml of TNF-α into the medium, macrophages stimulated with P. carinii released 108.2 ± 20.4 pg/ml of TNF- α after overnight culture (p = 0.0001). Maximal TNF-α release was observed after 8 h of incubation and required a P. carinii: macrophage ratio of at least 2.5:1. Autoclaved P. carinii were also able to trigger TNF-α release from macrophages albeit at a reduced level. In view of recent evidence that P. carinii is phylogenetically related to the fungi and contains a β- glucan-rich cell wall, we hypothesized that TNF-α release might in part be mediated by this cell wall component. Preincubation of macrophages with particulate β-glucan derived from Baker's yeast resulted in complete inhibition of TNF-α release in response to P. carinii. In addition, digestion of P. carinii with zymolyase, a preparation containing predominantly β-glucanase activity, substantially reduced the ability of P. carinii to cause TNF-α release from macrophages. In a similar manner, macrophages incubated with P. carinii in the presence of laminariheptaose, an oligosaccharide that binds to macrophage β-glucan receptors, also displayed decreased TNF-α release. Interestingly, TNF-α release may not be completely linked to the adherence of the organism to macrophages. Particulate β- glucan significantly reduced P. carinii adherence to macrophages and also impaired TNF-α release. However, yeast mannan also significantly reduced P. carinii adherence but had no effect on TNF-α release. These data demonstrate that P. carinii can directly stimulate the secretion of TNF-α from alveolar macrophages and that this effect is largely mediated by β-glucan components of the P. carinii cell wall.