Fibrosis involves increasing amounts of scar tissue appearing in a tissue, but what drives this is unclear. In fibrotic lesions in human and mouse lungs, we found extensive desialylation of glycoconjugates, and upregulation of sialidases. The fibrosis-associated cytokine TGF-β1 upregulates sialidases in human airway epithelium cells, lung fibroblasts, and immune system cells. Conversely, addition of sialidases to human peripheral blood mononuclear cells induces accumulation of extracellular TGF-β1, forming what appears to be a sialidase - TGF-β1 - sialidase positive feedback loop. Monocyte-derived cells called fibrocytes also activate fibroblasts, and we found that sialidases potentiate fibrocyte differentiation. A sialylated glycoprotein called serum amyloid P (SAP) inhibits fibrocyte differentiation, and sialidases attenuate SAP function. Injections of the sialidase inhibitors DANA and oseltamivir (Tamiflu) starting either 1 day or 10 days after bleomycin strongly attenuate pulmonary fibrosis in the mouse bleomycin model, and by breaking the feedback loop, cause a downregulation of sialidase and TGF-β1 accumulation. Together, these results suggest that a positive feedback loop involving sialidases potentiates fibrosis, and suggest that sialidase inhibitors could be useful for the treatment of fibrosis.

Fibrosis is caused by scar tissue formation in internal organs and is associated with 45% of deaths in the United States. Two closely related human serum proteins, serum amyloid P (SAP) and C-reactive protein (CRP), strongly affect fibrosis. In multiple animal models, and in Phase 1 and Phase 2 clinical trials, SAP affects several aspects of the innate immune system to reduce fibrosis, whereas CRP appears to potentiate fibrosis. However, SAP and CRP bind the same Fcγ receptors (FcγR) with similar affinities, and why SAP and CRP have opposing effects is unknown. Here, we report that SAP but not CRP binds the receptor DC-SIGN (SIGN-R1) to affect the innate immune system, and that FcγR are not necessary for SAP function. A polycyclic aminothiazole DC-SIGN ligand and anti-DC-SIGN antibodies mimic SAP effects in vitro. In mice, the aminothiazole reduces neutrophil accumulation in a model of acute lung inflammation and, at 0.001 mg/kg, alleviates pulmonary fibrosis by increasing levels of the immunosuppressant IL-10. DC-SIGN (SIGN-R1) is present on mouse lung epithelial cells, and SAP and the aminothiazole potentiate IL-10 production from these cells. Our data suggest that SAP activates DC-SIGN to regulate the innate immune system differently from CRP, and that DC-SIGN is a target for antifibrotics.F ibrosing diseases such scleroderma, pulmonary fibrosis, and renal fibrosis are caused by aberrant scar tissue formation in internal organs and are associated with 45% of deaths in the United States (1). At a fibrotic lesion, monocytes leave the blood, enter the tissue, and differentiate into cells such as macrophages and fibrocytes (2). Fibrocytes and macrophages then secrete extracellular matrix (ECM) proteins, ECM modifying enzymes, and/or cytokines such as IL-4 to promote scar tissue formation and fibrosis (3, 4).Pentraxins are a family of highly conserved secreted proteins that have a profound effect on the development of fibrosis and the regulation of the innate immune system (5-7). The pentraxin serum amyloid P (SAP) reduces neutrophil activation and recruitment (8, 9), inhibits the differentiation of monocytes into fibroblast-like cells called fibrocytes (8, 10), and promotes IL-10-secreting macrophages (11-13). In animal models and two human trials (6, 14, 15), injections of SAP decrease fibrosis, indicating that SAP has a dominant effect on a disease that is mediated in part by the innate immune system. Conversely, the closely related pentraxin C-reactive protein (CRP) is proinflammatory and promotes fibrosis (5, 16). However, under some conditions, CRP decreases inflammation, indicating that much remains to be understood about this molecule (5, 17). Despite the strong effects of pentraxins on the innate immune system and fibrosis (5, 6), little is known about their mechanism of action. For instance, pentraxins such as SAP and CRP appear to bind the same Fcγ receptors (FcγR) with similar affinities (7,8,18), but they generally have opposite effects. What causes this functional difference is not known.Dendritic ce...

RNA aptamers are synthetic oligonucleotide-based affinity molecules that utilize unique three-dimensional structures for their affinity and specificity to a target such as a protein. They hold the promise of numerous advantages over biologically produced antibodies; however, the binding affinity and specificity of RNA aptamers are often insufficient for successful implementation in diagnostic assays or as therapeutic agents. Strong binding affinity is important to improve the downstream applications. We report here the use of the phosphorodithioate (PS2) substitution on a single nucleotide of RNA aptamers to dramatically improve target binding affinity by ∼1000-fold (from nanomolar to picomolar). An X-ray co-crystal structure of the α-thrombin:PS2-aptamer complex reveals a localized induced-fit rearrangement of the PS2-containing nucleotide which leads to enhanced target interaction. High-level quantum mechanical calculations for model systems that mimic the PS2 moiety and phenylalanine demonstrate that an edge-on interaction between sulfur and the aromatic ring is quite favorable, and also confirm that the sulfur analogs are much more polarizable than the corresponding phosphates. This favorable interaction involving the sulfur atom is likely even more significant in the full aptamer-protein complexes than in the model systems.

In healing wounds and fibrotic lesions, fibroblasts and monocyte-derived fibroblast-like cells called fibrocytes help to form scar tissue. Although fibrocytes promote collagen production by fibroblasts, little is known about signaling from fibroblasts to fibrocytes. In this report, we show that fibroblasts stimulated with the fibrocyte-secreted inflammatory signal tumor necrosis factor-α secrete the small leucine-rich proteoglycan lumican, and that lumican, but not the related proteoglycan decorin, promotes human fibrocyte differentiation. Lumican competes with the serum fibrocyte differentiation inhibitor serum amyloid P, but dominates over the fibroblast-secreted fibrocyte inhibitor Slit2. Lumican acts directly on monocytes, and unlike other factors that affect fibrocyte differentiation, lumican has no detectable effect on macrophage differentiation or polarization. α2β1, αMβ2, and αXβ2 integrins are needed for lumican-induced fibrocyte differentiation. In lung tissue from pulmonary fibrosis patients with relatively normal lung function, lumican is present at low levels throughout the tissue, whereas patients with advanced disease have pronounced lumican expression in the fibrotic lesions. These data may explain why fibrocytes are increased in fibrotic tissues, suggest that the levels of lumican in tissues may have a significant effect on the decision of monocytes to differentiate into fibrocytes, and indicate that modulating lumican signaling may be useful as a therapeutic for fibrosis.

Tissue-resident macrophages are present in most tissues with developmental, self-renewal, or functional attributes that do not easily fit into a textbook picture of a plastic and multifunctional macrophage originating from hematopoietic stem cells; nor does it fit a pro- versus anti-inflammatory paradigm. This review presents and discusses current knowledge on the developmental biology of macrophages from an evolutionary perspective focused on the function of macrophages, which may aid in study of developmental, inflammatory, tumoral, and degenerative diseases. We also propose a framework to investigate the functions of macrophages in vivo and discuss how inherited germline and somatic mutations may contribute to the roles of macrophages in diseases.