MFAP4 (microfibrillar-associated protein 4) is an extracellular glycoprotein found in elastic fibers without a clearly defined role in elastic fiber assembly. In the present study, we characterized molecular interactions between MFAP4 and elastic fiber components. We established that MFAP4 primarily assembles into trimeric and hexameric structures of homodimers. Binding analysis revealed that MFAP4 specifically binds tropoelastin and fibrillin-1 and -2, as well as the elastin cross-linking amino acid desmosine, and that it co-localizes with fibrillin-1-positive fibers in vivo. Site-directed mutagenesis disclosed residues Phe 241 and Ser 203 in MFAP4 as being crucial for type I collagen, elastin, and tropoelastin binding. Furthermore, we found that MFAP4 actively promotes tropoelastin self-assembly. In conclusion, our data identify MFAP4 as a new ligand of microfibrils and tropoelastin involved in proper elastic fiber organization.Elastic fibers are key extracellular matrix structural elements of connective tissues that undergo repeated stretch, such as large arteries and the lung (1). The fibers consist of two major components: an amorphous elastin core surrounded by a sheath of fibrillin-rich microfibrils (2). Elastin is a highly hydrophobic polymer of the soluble precursor tropoelastin (3). Tropoelastin is known to undergo a self-assembly process known as coacervation (4), often believed to be a first step in the process of elastic fiber maturation. Because of the high content of lysine residues within the tropoelastin sequence, its assembly into a polymeric form is stabilized by formation of desmosine crosslinks, catalyzed by the lysyl oxidase (LOX) 3 enzyme family (5).Microfibrils, the other major component of elastic fibers, provide the structural scaffold for the deposition of elastin globules. They consist primarily of fibrillin-1 and fibrillin-2, large glycoproteins with a high degree of homology (6). Apart from fibrillins, numerous accessory proteins have been shown to associate with microfibrils or elastin and promote formation of mature fibers, including fibulins and microfibril-associated glycoproteins (MAGPs) (7-10). The importance of proper elastogenesis has been underscored by gene deficiency studies: mice lacking elastin, LOX, or fibrillin-1 die shortly after birth because of vascular abnormalities (11-13).MFAP4 (microfibrillar-associated protein 4) is an extracellular matrix protein belonging to the fibrinogen-related domain (FReD) family. The family includes several proteins engaged in tissue homeostasis and innate immunity, such as FIBCD1 (fibrinogen C domain-containing 1), ficolins, and angiopoietins (14 -16). The crystal structure of the FReD of several family members has been solved (17-19). The ligand-binding site, designated S1, is described in all the proteins and is located in close proximity to the calcium-binding site. MFAP4 has been reported to form homodimeric structures that further oligomerize, but its definite oligomerization pattern has not been established (20).MFAP4 is conside...

V ascular smooth muscle cell (VSMC) activation and phenotypic switching are critical for remodeling processes in vascular proliferative disorders, including intimal hyperplasia. Both the migratory and proliferative activities of VSMCs, as well as the interplay between the extracellular matrix (ECM) and integrin receptors essentially, contribute to neointimal hyperplasia and restrictive remodeling processes in the vessels.1 Among integrins, the particular role of integrin α V β 3 in the induction of VSMC responses has been shown both in vivo and in vitro.

MFAP4 promoted the development of asthmatic airway disease in vivo and pro-asthmatic functions of bronchial smooth muscle cells in vitro. Collectively, our results identify MFAP4 as a novel contributor to experimental asthma, acting through modulation of airway smooth muscle cells.

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Surfactant protein D (SP-D) is a pulmonary collectin important in lung immunity. SP-D-deficient mice (Sftpd(-/-)) are reported to be susceptible to ovalbumin (OVA)- and fungal allergen-induced pulmonary inflammation, while treatment with exogenous SP-D has therapeutic effects in such disease models. β-Glucans are a diverse group of polysaccharides previously suggested to serve as fungal ligands for SP-D. We set out to investigate if SP-D could interact with 1,3-β-glucan and attenuate allergic pulmonary inflammation in the presence of 1,3-β-glucan. Allergic airway disease was induced in Sftpd(-/-) and Sftpd(+/+) mice by OVA sensitization and subsequent challenge with OVA, 1,3-β-glucan, or OVA/1,3-β-glucan together. Mice in the combined treatment group were further treated with a high dose of recombinant fragment of human SP-D (rfhSP-D). We demonstrated direct interaction between SP-D and 1,3-β-glucan. OVA-induced mucous cell metaplasia was increased in Sftpd(-/-) mice, supporting previously reported protective effects of endogenous SP-D in allergy. OVA-induced parenchymal CCL11 levels and eosinophilic infiltration in bronchoalveolar lavage were unaffected by 1,3-β-glucan, but were reversed with rfhSP-D treatment. 1,3-β-Glucan treatment did, however, induce pulmonary neutrophilic infiltration and increased TNF-α levels in bronchoalveolar lavage, independently of OVA-induced allergy. This infiltration was also reversed by treatment with rfhSP-D. 1,3-β-Glucan reduced OVA-induced mucous cell metaplasia, T helper 2 cytokines, and IFN-γ production. rfhSP-D treatment further reduced mucous metaplasia and T helper 2 cytokine secretion to background levels. In summary, rfhSP-D treatment resulted in attenuation of both allergic inflammation and 1,3-β-glucan-mediated neutrophilic inflammation. Our data suggest that treatment with high-dose SP-D protects from mold-induced exacerbations of allergic asthma.