The lack of an appropriate animal model with multiorgan pathology characteristic of the human form of cystic fibrosis has hampered our understanding of the pathobiology of the disease. We evaluated multiple organs of congenic C57BL/6J cystic fibrosis transmembrane regulator (Cftr)(-/-) and Cftr(+/+) mice maintained from weaning on a liquid diet then sacrificed between 1 and 24 months of age. The lungs of the Cftr(-/-) animals showed patchy alveolar overdistention, interstitial thickening, and fibrosis, with progression up to 6 months of age. The proximal and distal airway surface was encased with mucus-like material but lacked overt evidence of chronic bacterial infections or inflammation. All Cftr(-/-) animals showed progressive liver disease, with hepatosteatosis, focal cholangitis, inspissated secretions, and bile duct proliferation; after 1 year of age there was progression to focal biliary cirrhosis. The intercalated, intralobular and interlobular ducts and acinar lumina of the exocrine pancreas, the parotid and submaxillary glands of the Cftr(-/-) animals were dilated and filled with inspissated material, as well as mild inflammation and acinar cell drop out. Quantitative measurements of the pancreas showed significant acinar atrophy and increased acinar volume in comparison with age-matched Cftr(+/+) littermates. The ileal lumen and crypts were filled with adherent fibrillar material. After 3 months of age the vas deferens of the Cftr(-/-) animals could not be identified. None of the aforementioned pathological changes were observed in the Cftr(+/+) littermates fed the same liquid diet. We show, for the first time, that long-lived C578L/6J Cftr(-/-) mice develop manifestations of cystic fibrosis-like disease in all pathologically affected organs in the human form of cystic fibrosis.

Fibrinogen-like protein 2 (FGL2) is a multifunctional protein, which has been implicated in the pathogenesis of allograft and xenograft rejection. Previously, FGL2 was shown to inhibit maturation of BM-derived DC and T-cell proliferation. The mechanism of the immunosuppressive activity of FGL2 remains poorly elucidated. Here, we focus on identification of FGL2-specific receptor(s) and their ability to modulate APC activity and allograft survival. Using flow cytometry and surface plasmon resonance analysis, we show that FGL2 binds specifically to Fc gamma receptor (FccR)IIB and FccRIII receptors, which are expressed on the surface of APC, including B lymphocytes, macrophages and DC. Antibody to FccRIIB and FccRIII, or deficiency of these receptors, abrogated FGL2 binding. IntroductionFibrinogen-like protein 2 (FGL2), also known as fibroleukin, was first cloned from cytotoxic T lymphocytes and was classified as a member of the fibrinogen superfamily due to its homology (36%) with fibrinogen b and g chains [1]. Predicted as a type II transmembrane glycoprotein [2], cell membraneassociated FGL2 was shown to exhibit novel prothrombinase activity when associated with cell membranes/phospholipid vesicles [3,4], which has been implicated in the pathogenesis of à These authors contributed equally to this study.Correspondence: Dr. Gary Levy e-mail: glfgl2@attglobal.net DOI 10.1002/eji.200838338 Eur. J. Immunol. 2008. 38: 3114- [7]. The procoagulant activity was shown to depend on the serine 89 at the linear N terminal domain of FGL2 [4].The C terminal globular portion of FGL2 lacks the prothrombinase activity and contains a classical fibrinogen-related domain (FRED), which has been suggested to possess immunomodulatory activity based on several lines of evidence. First, other fibrinogen superfamily members including fibrinogen [8] Results Recombinant FcFGL2 protein binds to APCRecombinant FcFGL2 was generated and purified as described in the Materials and methods. The molecular size of the recombinant proteins was examined by SDS-PAGE, silver staining and Western blotting. FcFGL2 had a molecular weight of approximately 440 kDa under non-reducing conditions, which was confirmed by gel filtration (data not shown), and 110 kDa under reducing conditions (Fig. 1A). These data suggested that FcFGL2 exists as a tetramer, consistent with previous reports [13,21]. The Fc tag had a molecular size of 64 kDa under non-reducing conditions and 33 kDa under reducing conditions, suggesting that Fc is dimeric. Biotinylated FcFGL2 bound to the RAW264.7 cells (mouse macrophage cell lines), BMDC, the B-cell line A20 (which expresses only one Fcg receptor, FcgRIIB), thioglycolate-elicited peritoneal exudates cells (495% macrophages) from C57BL/6J mice, but FcFGL2 did not bind to the B-cell line A20IIA1.6, which does not express FcgRIIB (Fig. 1B) [22]. EL4 cells (a mouse T-cell line) did not bind FcFGL2 (data not shown). As expected, the Fc tag protein alone failed to bind to any of these cells and thus, Fc provided an appropriate negativ...

Hypertonicity suppresses neutrophil functions by unknown mechanisms. We investigated whether osmotically induced cytoskeletal changes might be related to the hypertonic inhibition of exocytosis. Hyperosmolarity abrogated the mobilization of all four granule types induced by diverse stimuli, suggesting that it blocks the process of exocytosis itself rather than individual signaling pathways. Concomitantly, osmotic stress provoked a twofold increase in F-actin, induced the formation of a submembranous F-actin ring, and abolished depolymerization that normally follows agonist-induced actin assembly. Several observations suggest a causal relationship between actin polymerization and inhibition of exocytosis: 1) prestimulus actin levels were inversely proportional to the stimulus-induced degranulation, 2) latrunculin B (LB) prevented the osmotic actin response and restored exocytosis, and 3) actin polymerization induced by jasplakinolide inhibited exocytosis under isotonic conditions. The shrinkage-induced tyrosine phosphorylation and the activation of the Na(+)/H(+) exchanger were not affected by LB. Inhibition of osmosensitive kinases failed to prevent the F-actin change, suggesting that the osmotic tyrosine phosphorylation and actin polymerization are independent phenomena. Thus cytoskeletal remodeling appears to be a key component in the neutrophil-suppressive, anti-inflammatory effects of hypertonicity.