Surfactant Protein D-Mediated Aggregation of Pneumocystis cariniiImpairs Phagocytosis by Alveolar Macrophages

ABSTRACTPneumocystis cariniiremains an important and potentially fatal cause of opportunistic pneumonia. Animal studies reveal that substantial quantities of surfactant protein D (SP-D) accumulate in the airspaces during P. cariniipneumonia and are particularly abundant in aggregates of organisms. Due to the multimeric structure of SP-D, we hypothesized that SP-D mediates aggregation of the organism. From previous clinical studies it is known that aggregated organisms are conspicuous in sections of lung tissue and bronchoalveolar lavage (BAL) fluids of humans with active P. cariniipneumonia. Herein, we observe that SP-D levels increased at least fourfold in BAL fluids of patients with P. cariniipneumonia. Next, a spectrophotometric sedimentation assay was developed to assess the aggregation of P. cariniiin vitro by SP-D. P. cariniiorganisms were first stripped with glutathione to remove bound SP-D and subsequently incubated in the presence of SP-D and 2 mM calcium. P. cariniiincubated with natural SP-D (10 μg/ml) containing dodecamers and higher-order forms exhibited aggregation and enhanced sedimentation compared to that of glutathione-stripped P. carinii. Aggregation was also enhanced by the concentrated supernatant of rat BAL fluid, and this effect was abolished by the selective removal of SP-D from the lavage fluid. P. cariniiaggregation was reduced by maltose, mannose, and EDTA, consistent with the role of the SP-D C-type lectin domain (CRD) in the aggregation event. Comparisons of different molecular forms of SP-D showed that dodecamers—but not trimeric subunits—mediate optimal aggregation of P. carinii. Aggregation of P. cariniiby SP-D was shown to be responsible for the impaired phagocytosis of the organisms by alveolar macrophages. Thus, SP-D-mediated aggregation of P. cariniimay represent one means by which the organism avoids elimination by the host.