BACKGROUND AND PURPOSEEndogenous glucocorticoids are pro-resolving mediators, an example of which is the endogenous glucocorticoid-regulated protein annexin A1 (ANXA1). Because silicosis is an occupational lung disease characterized by unabated inflammation and fibrosis, in this study we tested the therapeutic properties of the N-terminal ANXA1-derived peptide annexin 1-(2-26) (Ac2-26) on experimental silicosis. EXPERIMENTAL APPROACHSwiss-Webster mice were administered silica particles intranasally and were subsequently treated with intranasal peptide Ac2-26 (200 μg per mouse) or dexamethasone (25 μg per mouse) for 7 days, starting 6 h post-challenge. Ac2-26 abolished the leukocyte infiltration, collagen deposition, granuloma formation and generation of pro-inflammatory cytokines evoked by silica; these variables were only partially inhibited by dexamethasone. KEY RESULTSA clear exacerbation of the silica-induced pathological changes was observed in ANXA1 knockout mice as compared with their wild-type (WT) littermate controls. Incubation of lung fibroblasts from WT mice with Ac2-26 in vitro reduced IL-13 or TGF-β-induced production of CCL2 (MCP-1) and collagen, but this peptide did not affect the production of CCL2 (MCP-1) by stimulated fibroblasts from formyl peptide receptor type 1 (FPR1) knockout mice. Ac2-26 also inhibited the production of CCL2 (MCP-1) from fibroblasts of FPR2 knockout mice. CONCLUSIONS AND IMPLICATIONSCollectively, our findings reveal novel protective properties of the ANXA1 derived peptide Ac2-26 on the inflammatory and fibrotic responses induced by silica, and suggest that ANXA1 mimetic agents might be a promising strategy as innovative anti-fibrotic approaches for the treatment of silicosis. IntroductionSilicosis is a lung occupational disease caused by inhalation of free crystalline silica particles over a prolonged period of time. This disease mainly affects workers in the ceramic industry, glass manufacturing, sandblasting, mining, quarrying and civil construction (Leung et al., 2012;Singer et al., 2012). Silicosis is a problem worldwide, but it is prevalent in countries of low and middle income. China had more than 500 000 cases recorded between 1991 and 1995, and 6000 new cases and more than 24 000 deaths are reported annually (Leung et al., 2012). In Brazil more than 4500 workers were reported to have silicosis between 1978 and 1998, especially in the gold-mining area in Minas Gerais (Carneiro et al., 2006). The irritative action of silica in the lungs results in the formation of nodular lesions, which may coalesce resulting in massive areas of fibrous tissue and reduction of lung elasticity (Davis, 1986). Because of its surface properties, crystalline silica leads to activation of alveolar macrophages [acting on 'scavenger' receptors such as the MARCO (macrophage receptor with collagenous structure)], epithelial cells and fibroblasts (Mossman and Churg, 1998;Fubini and Hubbard, 2003;Huaux, 2007). The granulomatous inflammation induced by silica is characterized by the accumulati...

MSCs therapeutically administered exert anti-inflammatory effects in the airway of HDM-challenged mice, but do not ameliorate lung function or remodelling. Although MSC pre-treatment can increase Treg cell numbers, it is highly unlikely that the MSCs will induce Treg cell expansion when lymphocytes are allergenically primed in an established lung inflammation.

Inhaled lidocaine prevents eosinophilic inflammation, overproduction of mucus, and peribronchial fibrosis in a murine model of asthma, and impaired airway hyperreactivity, possibly by inhibiting allergen-evoked GATA-3 expression and the subsequent up-regulation of proinflammatory cytokines and chemokines.

Instillation of silica into the lungs of rodents results in pathological changes that strongly mimic human silicosis, an occupational lung disease marked by restrictive airway obstruction, inflammation, and fibrosis. Because IL-13 is a pivotal proinflammatory and fibrogenic cytokine, we examined whether a recombinant immunotoxin comprised of human IL-13 and a mutated form of Pseudomonas exotoxin (IL-13–PE) might affect pathological features of experimental silicosis. Mice received a single intranasal instillation of silica particles and were treated with intranasal IL-13–PE every other day from days 21 to 27 postsilica. The sensitivity of putative cell targets to IL-13–PE was also assessed in in vitro settings. Upregulation of IL-13, its receptor subunits IL-13Rα1 and IL-13Rα2, and shared receptor IL-4Rα were associated with development of granulomatous lung inflammation triggered by silica. IL-13–PE inhibited silica-induced granuloma and fibrotic responses noted at 24 h and 15 d after the last treatment. Upregulation of TNF-α, TGF-β, and chemokines, as well as increased collagen deposition and airway hyperreactivity to methacholine were all clearly sensitive to IL-13–PE. In addition, IL-13–PE inhibited both IL-13–induced proliferation of cultured lung fibroblasts from silicotic mice and silica-induced IL-8 generation from A549 cells. In conclusion, our findings show that therapeutic treatment with IL-13–PE can reverse important pathological features caused by inhalation of silica particles, suggesting that this recombinant immunotoxin is a promising molecular template in drug discovery for the treatment of silicosis.

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Previous studies described that allergic diseases, including asthma, occur less often than expected in patients with type 1 diabetes. Here, we investigated the influence of diabetes on allergic airway inflammation in a model of experimental asthma in mice. Diabetes was induced by intravenous injection of alloxan into 12 h-fasted A/J mice, followed by subcutaneous sensitization with ovalbumin (OVA) and aluminum hydroxide (Al(OH)3), on days 5 and 19 after diabetes induction. Animals were intranasally challenged with OVA (25 μg), from day 24 to day 26. Alloxan-induced diabetes significantly attenuated airway inflammation as attested by the lower number of total leukocytes in the bronchoalveolar lavage fluid, mainly neutrophils and eosinophils. Suppression of eosinophil infiltration in the peribronchiolar space and generation of eosinophilotactic mediators, such as CCL-11/eotaxin, CCL-3/MIP-1α, and IL-5, were noted in the lungs of diabetic sensitized mice. In parallel, reduction of airway hyperreactivity (AHR) to methacholine, mucus production, and serum IgE levels was also noted under diabetic conditions. Our findings show that alloxan diabetes caused attenuation of lung allergic inflammatory response in A/J mice, by a mechanism possibly associated with downregulation of IgE antibody production.

We previously reported that alloxan-induced diabetes results in reduction inIt has been demonstrated that diabetic patients present many functional abnormalities which can be partially responsible for their failure in mounting an appropriate inflammatory response (Garcia-Leme 1989). In line with this concept, some authors reported that the occurrence of allergic disorders concomitantly with type 1 diabetes is markedly reduced when compared with the incidence of each disease alone (Olesen et al. 2001, Rosenbauer et al. 2003. It is thus possible that the resistance to allergic provocation results from an imbalance in the T-helper 1/ T-helper 2 (Th1/Th2) response. Accordingly, since autoimmune type 1 diabetes is Th1-dependent and allergy is Th2-dependent, the susceptibility to one disease might lead a state of refractoriness to the other (Huang 1999).We previously described that alloxan-diabetic rats showed reduction of the allergen-induced acute plasma leakage and late eosinophil accumulation in the pleural cavity, phenomena which simultaneously occurred in parallel with decrease in the number of mast cells recovered from the pleural space (Diaz et al. 1996). Antigen-evoked plasma leakage in diabetic rats was restored by adoptive transfer of mast cells from sensitized non-diabetic rats (de Oliveira Barreto et al. 2003), supporting the idea that refractoriness of diabetic animals to allergen stimulation seems to be at least partially accounted for by depletion in the mast cell population. Mast cells act as central effector and regulatory cells in many inflammatory disorders, such as autoimmunity, allergy and parasite infections (Galli et al. 1999). They were shown to be widely distributed in many tissues including skin, airways, gastrointestinal tract (Wasserman 1990), and also in the thymus (Huntley et al. 1990). Based on the above observations, we investigated herein the interference of alloxan-diabetes on thymic mast cell population. MATERIALS AND METHODSMale Wistar rats from the Oswaldo Cruz Foundation breeding were used. All procedures involving care and use of laboratory animals in this study were examined and approved by the Animal Ethics Committee of the Oswaldo Cruz Foundation (License 0085-02). Diabetes was induced by a single injection of alloxan monohydrate (Sigma St. Louis, US) (40 mg/kg, i.v.) into 12 h fasted rats. Control animals were injected with the vehicle alone. Blood glycaemia was determined by means of a glucose monitor in samples obtained from the tail vein and only rats with glucose levels above 200 mg/dl were considered for further experiments. To evaluate thymus atrophy, the organ was removed from diabetic and control animals, dried for 24 h at 40ºC and weighed. For cell analyses, the thymus was passed through a mesh stainless steel grid and the supernatant centrifuged at 150 xg for 10 min. Cells were harvested, washed three times with phosphate buffer solution (PBS) and counted in a Neubauer chamber, after dilution of samples in Türk solution. For histological analysis, thymus fragments ...

Silicosis is an occupational disease triggered by the inhalation of fine particles of crystalline silica and characterized by inflammation and scarring in the form of nodular lesions in the lungs. In spite of the therapeutic arsenal currently available, there is no specific treatment for the disease. Flunisolide is a potent corticosteroid shown to be effective for controlling chronic lung inflammatory diseases. In this study, the effect of flunisolide on silica-induced lung pathological changes in mice was investigated. Swiss-Webster mice were injected intranasally with silica particles and further treated with flunisolide from day 21 to 27 post-silica challenge. Lung function was assessed by whole body invasive plethysmography. Granuloma formation was evaluated morphometrically, collagen deposition by Picrus sirius staining and quantitated by Sircol. Chemokines and cytokines were evaluated using enzyme-linked immunosorbent assay. The sensitivity of lung fibroblasts was also examined in in vitro assays. Silica challenge led to increased leukocyte numbers (mononuclear cells and neutrophils) as well as production of the chemokine KC/CXCL-1 and the cytokines TNF-α and TGF-β in the bronchoalveolar lavage. These alterations paralleled to progressive granuloma formation, collagen deposition and impairment of lung function. Therapeutic administration of intranasal flunisolide inhibited granuloma and fibrotic responses, noted 28 days after silica challenge. The upregulation of MIP-1α/CCL-3 and MIP-2/CXCL-2 and the cytokines TNF-α and TGF-β, as well as deposition of collagen and airway hyper-reactivity to methacholine were shown to be clearly sensitive to flunisolide, as compared to silica-challenge untreated mice. Additionally, flunisolide effectively suppressed the responses of proliferation and MCP-1/CCL-2 production from IL-13 stimulated lung fibroblasts from silica-or saline-challenged mice. In conclusion, we report that intranasal treatment with the corticosteroid flunisolide showed protective properties on pathological features triggered by silica particles in mice, suggesting that the compound may constitute a promising strategy for the treatment of silicosis.