Intranasal Flunisolide Suppresses Pathological Alterations Caused by Silica Particles in the Lungs of Mice.

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-ß 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-ß, 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.

Reduced expression of IL-3 mediates intestinal mast cell depletion in diabetic rats: role of insulin and glucocorticoid hormones.

Rats turned diabetic by alloxan treatment are refractory to systemic anaphylactic shock, in a direct association with reduced intestinal haemorrhage and tissue response to antigen challenge. As diabetic rats show reduction in mast cell numbers in different body compartments, this study was undertaken to investigate the influence of alloxan diabetes on mast cell population as well as the expression of the mast cell growth factor interleukin (IL)-3 in the small intestine of rats. We also analysed the putative involvement of endogenous insulin and glucocorticoid hormones in this phenomenon. There was a significant decrease in the number of mast cells present in the small intestine (ileum segment) of diabetic rats. Likewise, the immunohistochemical analysis revealed that IL-3 labelling was markedly attenuated in diabetic rats, as compared with normal animals, a phenomenon which paralleled with a decreased mRNA expression as attested by Reverse transcriptase-polymerase chain reaction technique. Treatment with insulin and with the steroid receptor antagonist RU 486 restored basal mast cell numbers, normal levels of IL-3 labelling and mRNA expression for IL-3 in the ileum of diabetic rats. In conclusion, our findings show that there is a causative relationship between down-regulation of mast cell numbers and the expression of IL-3 associated with diabetic state. In addition, as both parameters were suppressed by administration of insulin and RU 486, it indicates that an imbalance between the systemic levels of insulin and glucocorticoid hormones seems to be implicated in the reduction in intestinal mast cell population and refractoriness to antigen provocation in alloxan diabetes.

Estudo da participação de mastócitos no processo de fibrose pulmonar induzido por sílica em camundongos / Study of mast cell involvement in silica-induced pulmonary fibrosis process in mice

A silicose é uma doença ocupacional, caracterizada pela presença de fibrose crônica nos pulmões. Evidências apontam para a implicação de mastócitos em algumas doenças de caráter fibrótico. Assim, no presente estudo, buscamos investigar a participação de mastócitos durante o estabelecimento do quadro de silicose pulmonar em camundongos, utilizando análise tanto qualitativa como quantitativa. A ativação de mastócitos por partículas de sílica in vitro e a potencial interação entre mástócitos e fibroblastos foram igualmente avaliadas. Partículas de sílica (10 mg) foram administradas intranasalmente em camundongos Swiss-Webster e as análises realizadas nos tempos de 7, 14 e 28 dias após a indução da silicose. Parâmetros como morfologia pulmonar e número de mastócitos, assim como expressão de stem-cell factor (SCF), foram avaliados por meio de colorações específicas e imunohistoquímica, respectivamente. Para análise da ativação de mastócitos in vitro, foram utilizadas células derivadas a partir da medula óssea (BMMCs) e a liberação de Beta-hexosaminidase (Beta-hex) como parâmetro de ativação. A análise do tecido pulmonar de camundongos silicóticos revelou a presença de uma intensa resposta fibrótica e numerosos granulomas, majoritariamente localizados na região peribrônquica, com início detectado no dia 7 e máximo sendo observado no tempo de 28 dias pós-sílica. Verificamos um aumento acentuado no número de mastócitos pulmonares, tanto em 14 quanto em 28 após a sílica, com claro predomínio de células apresentando fenótipo do tipo conjuntivo. De forma interessante, vimos que este fenômeno foi precedido por aumento na expressão de SCF em comparação ao verificado no caso dos animais controles. Na condição da exposição de BMMC às partículas de sílica in vitro evidenciamos sinais claros de ativação celular, conforme atestado pelo aumento na percentagem de Beta-hex liberada. Esta resposta mostrou-se sensível ao tratamento com a toxina pertussis sugerindo, então...murino.

Neutrophil infiltration is implicated in the sustained thermal hyperalgesic response evoked by allergen provocation in actively sensitized rats.

It has been proposed that allergen provocation induces hyperalgesia but the involvement of immunoglobulin E and leukocytes remains poorly understood. Here, we have compared the profile of allergen-evoked thermal hyperalgesic response in both passively and actively sensitized rats, and investigated the role of leukocytes in allergen-evoked nociception. Wistar rats were passively sensitized with an intraplantar injection of immunoglobulin E anti-dinitrophenylated bovine serum albumin monoclonal antibody (0.5 microg/paw), and challenged with dinitrophenylated bovine serum albumin (0.5 microg/paw) 24 h later. Alternatively, the animals were actively sensitized with a mixture of Al(OH)3 and ovalbumin and challenged intraplantarly with ovalbumin (12 microg/paw) 14 days later. We found that the thermal hyperalgesic responses set in very rapidly and with comparable intensity in both passively and actively sensitized rats. However, while in the former group the response was shorter, peaking within 1 h and reducing thereafter, a marked plateau was observed from 1 to 6 h post-challenge in the latter group. Actively sensitized rats also had higher neutrophil influx in the plantar tissue, as attested by both myeloperoxidase activity and histological analysis. Treatment of actively sensitized rats with either fucoidin (10 mg/kg, i.v) or anti-rat neutrophil antiserum (i.p.) reduced neutrophil accumulation and the late hyperalgesic response noted from 3 to 6 h post-challenge. Thus, we conclude that though immunoglobulin E-mediated mechanisms can cause thermal hyperalgesia, components of the cellular immune reaction are crucial in order to amplify and sustain the immediate hyperalgesic response triggered by allergen, in a process dependent on neutrophil recruitment.