Mechanisms of innate immune surveillance by pulmonary surfactant proteins and their clinical application to respiratory infection.

点击翻译按钮获取中文摘要

The purposes of this project are to investigate the molecular mechanisms of ligand recognition by pulmonary surfactant proteins (SP-A and SP-D ; pulmonary collectins) and Toll-like receptors and of the collectin-mediated bacterial phagocytosis by macrophages, and to establish the molecular basis for the clinical application to respiratory infections.1.We have shown that Mycobacterium avium is a ligand for pulmonary collectins, and that SP-D binds to lipoarabinomannan derived from M.avium. In addition, we have found that SP-A and SP-D enhance the phagocytosis of M.avium by increased activity of mannose receptor on macrophages.2.SP-A but not SP-D has been shown to augment the phagocytosis of Streptococcus pneumoniae and Staphylococcus aureus by alveolar macrophages. SP-A activates casein kinase 2 and increases cell surface localization of scavenger receptor A (SR-A). Thus, it is suggested that SP-A augment SR-A-mediated phagocytosis by casein kinase 2-dependent machanism. Analysis of SP- … More A/SP-D chimeras has revealed that the C-terminal Cys204-Cys218 is important for the SP-A's stimulatory effect on phagocytosis.3.We constructed a soluble form of recombinant extracellular TLR4 domain consisting of the region Glu24-Leu631 (sTLR4) and analysed the interactions with LPS and MD-2. MD-2 binds to the N-termnal region (Glu24-Pro34) of sTLR4. sTLR4 alone by itself does not bind to lipid A, but sTLR4 increases the binding affinity of MD-2 to lipid A and the amount of lipid A bound to MD-2. The sTLR4-MD-2 complex inhibits the binding of LPS to the cells expressing TLR4/MD-2. In addition, intratracheal instillation of sTLR4 and MD-2 dampens pulmonary inflammation caused by LPS in mice.4.SP-A and SP-D have been shown to directly bind to the extracellular domains of TLR2 and TLR4. Their bindings are Ca2+-dependent but are thought to be via protein-protein interaction because both collectins bind to deglycosylated sTLR proteins. Epitope mapping for anti-SP-D monoclonal antibodies and their effects on the SP-D binding to sTLRs have revealed that SP-D binds to sTLR proteins through the carbohydrate recognition domain (CRD). Collagenase-resistant fragment of SP-A (CRF), which has been shown to form trimer consisting of the CRD and the neck, binds to sTLR proteins with significantly lower affinity than that of wild type octadecameric SP-A. In addition, the ability of CRF to inhibit smooth LPS-induced NF-κB activation in TLR4/MD-2-tranfected cells was significantly attenuated. The results demonstrate the importance of supratrimeic oligomer formation by the N-terminal collagenous domain in interaction with TLR and in modulation of innate immune function by pulmonary collectins. Less