SURFACTANT PROTEIN-D BINDING TO ALVEOLAR MACROPHAGES

Surfactant protein D (SP-D) is a lung-specific protein, synthesized an d secreted by lung epithelial cells. It belongs to group III of the fa mily of C-type lectins; each member of this group has an unusual overa ll structure consisting of multiple globular 'head' regions (which con tain the C-type lectin domains) linked by triplehlical, collagen-like, strands. This group includes the surfactant protein A (SP-A) and the serum proteins mannan-binding protein, conglutinin and collectin-43, a ll of which have been shown to bind to the Clq receptor found on a wid e variety of cells, including macrophages. Both SP-D and SP-A have bee n shown to enhance oxygen radical production by alveolar macrophages. Although this strongly suggests a direct interaction between SP-D and a specific receptor on alveolar macrophages, it is still unclear wheth er SP-D binds to the same receptor used by SP-A and/or Clq. Human SP-D was isolated from amniotic fluid and was radiolabelled using I-125. A lveolar macrophages were isolated from human bronchioalveolar lavage f luid, and also from bovine lung washings, by differential adhesion to 24-well tissue-culture plates. The study was carried out using EDTA-co ntaining buffers, to eliminate Ca2+-dependent C-type lectin binding, a nd was also carried out at 4 degrees C to eliminate possible internali zation by the cells. I-125-SP-D showed specific binding to alveolar ma crophages in both a time- and concentration-saturable manner. The bind ing was inhibited, by approx. 90 %, on addition of a 200-fold excess o f unlabelled SP-D. The apparent dissociation constant (K-d) was (3.6 /- 1.3) x 10(-11) M, based on the assumption that native SP-D is assem bled as a dodecamer of 12 identical polypeptides of 43 kDa to yield a protein of 516 kDa. Clq was also shown to bind alveolar macrophages (K -d 3 x 10(-6) M), but addition of Clq did not show inhibition of the b inding of I-125-Sp-D to the macrophages. We conclude that SPD binds sp ecifically to alveolar macrophages and the receptor involved is differ ent from that utilized by Clq.