No abstract

Lung surfactant protein-D (SP-D), a collectin mainly produced by alveolar type II cells, initiates the effector mechanisms of innate immunity on binding to microbial carbohydrates. A panel of mRNAs from human tissues was screened for SP-D mRNA by RT-PCR. The lung was the main site of synthesis, but transcripts were readily amplified from trachea, brain, testis, salivary gland, heart, prostate gland, kidney, and pancreas. Minor sites of synthesis were uterus, small intestine, placenta, mammary gland, and stomach. The sequence of SP-D derived from parotid gland mRNA was identical with that of pulmonary SP-D. mAbs were raised against SP-D, and one was used to locate SP-D in cells and tissues by immunohistochemistry. SP-D immunoreactivity was found in alveolar type II cells, Clara cells, on and within alveolar macrophages, in epithelial cells of large and small ducts of the parotid gland, sweat glands, and lachrymal glands, in epithelial cells of the gall bladder and intrahepatic bile ducts, and in exocrine pancreatic ducts. SP-D was also present in epithelial cells of the skin, esophagus, small intestine, and urinary tract, as well as in the collecting ducts of the kidney. SP-D is generally present on mucosal surfaces and not restricted to a subset of cells in the lung. The localization and functions of SP-D indicate that this collectin is the counterpart in the innate immune system of IgA in the adaptive immune system.

The serum opsonin mannose-binding lectin (MBL) has been shown to be involved in the handling of apoptotic cells. However, at what stage in the process this happens and whether this mediates activation of complement is unknown. Cells rendered apoptotic or necrotic were incubated with purified MBL/MBL-associated serine protease (MASP) complexes and assessed by flow cytometry and fluorescence microscopy. MBL bound specifically to late apoptotic cells, as well as to apoptotic blebs and to necrotic cells, but not to early apoptotic cells. Binding of MBL could be inhibited by EDTA as well as with an antibody against the CRD region. Addition of C1q, another serum opsonin involved in the handling of apoptotic cells, prior to MBL partly inhibited MBL binding to apoptotic cells and vice versa. MBL/ MASP could initiate deposition of purified complement C4 on the target cells. However, addition of MBL/MASP to whole serum deficient for both C1q and MBL did not enhance deposition of C4, but MBL enhanced phagocytosis of apoptotic cells by macrophages. These results demonstrate that MBL interacts with structures exposed on cells rendered late apoptotic or necrotic and facilitates uptake by macrophages. Thus, MBL may promote noninflammatory sequestration of dying host cells.

No abstract

Deficiency of human mannose-binding lectin (MBL)caused by mutations in the coding part of the MBL2 gene is associated with increased risk and severity of infections and autoimmunity. To study the biological consequences of MBL mutations, we expressed wild type MBL and mutated MBL in Chinese hamster ovary cells. The normal MBL cDNA (WT MBL-A) was cloned, and the three known natural and two artificial variants were expressed in Chinese hamster ovary cells. When analyzed, WT MBL-A formed covalently linked higher oligomers with a molecular mass of about 300 -450 kDa, corresponding to 12-18 single chains or 4 -6 structural units. By contrast, all MBL variants formed a dominant band of about 50 kDa, with increasingly weaker bands at 75, 100, and 125 kDa corresponding to two, three, four, and five chains, respectively. In contrast to WT MBL-A, variant MBL formed noncovalent oligomers containing up to six chains (two structural units). MBL variants bound ligands with a markedly reduced capacity compared with WT MBL-A. Mutations in the collagenous region of human MBL compromise assembly of higher order oligomers, resulting in reduced ligand binding capacity and thus reduced capability to activate complement. Mannose-binding lectin (MBL)1 has been shown to be an important component of innate immunity and is a central recognition molecule of the lectin pathway of complement (for a recent review, see Ref. 1). MBL binds to an array of carbohydrate structures on surfaces of bacteria (2-4), yeast, viruses (5, 6), and parasitic protozoa (7,8). MBL functions as an opsonin (9), and the biological effect is mediated by direct killing via complement (10) through the lytic membrane attack complex or by promoting phagocytosis either by the MBL lectin pathway of complement or by direct binding to one or more cell surface receptors (11). The lectin pathway comprises at least three MBL-associated serine proteases (MASPs), namely MASP-1 (12), MASP-2 (13), and . Furthermore, the functional MBL-MASP complex contains a small MBL-associated protein (sMAP), also named MAp19, with no serine protease activity (15, 16). MASP-2 is a homologue of C1s of the classical complement pathway because it activates C4 and C2 (13). When MBL associated with MASP-2 binds to sugar groups on the surface of microbes, the MBL-MASP2 proenzyme is activated and cleaves sequentially C4 and C2, thereby creating the C4b2a complex, a potent C3 convertase. The MBL-MASP-1 complex is suggested to activate C3 directly (12). Whether both MASP-1 and MASP-2 are bound on the same MBL molecule is still unclear. Moreover, the biological role of sMAP, as well as the substrate for the recently discovered MASP-3, remains unclear at this moment (for a recent review on MASPs, see Ref. 17).MBL is a complex of six sets of homotrimers of a single polypeptide chain containing 228 amino acids (18 -21). This polypeptide consists of four domains ( Fig. 1): 1) a 20-amino acid N-terminal cysteine-rich domain involved in formation of intraand intersubunit disulfide bonds, 2) a collagen-like dom...