Efeito de terapias anti-TNF-\F061 sobre a resposta inflamatória pulmonar crônica induzida por sílica em camundongos

Made available in DSpace on 2014-07-22T16:59:29Z (GMT). No. of bitstreams: 4 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) 69443.pdf: 30039667 bytes, checksum: ea70e0586b05a80c835ac7c80af39f40 (MD5) 69443.pdf.txt: 241427 bytes, checksum: f07e61db23ac87c25bde0ad04a330533 (MD5) 69443.pdf.jpg: 1051 bytes, checksum: 770309c47ccfffe998bf7bbc0d635cc9 (MD5) Previous issue date: 2014-05-07Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, BrasilA silicose é uma doença de caráter ocupacional, desencadeada pela inalação de partículas de sílica e associada à participação de uma ampla gama de mediadores. O TNF-\F061 é uma citocina pleiotrópica, que apresenta forma transmembranar (mTNF-\F061) e solúvel (sTNF-\F061), envolvidas em diversas doenças de caráter inflamatório. No presente estudo avaliamos o efeito de terapias neutralizantes de TNF-\F061, incluindo a inibição de geração com o composto talidomida, e o bloqueio da ação com o anticorpo infliximabe (Remicade®), no modelo de silicose experimental murina. Foi também testado o composto XPro 1595, um inibidor seletivo do sTNF-\F061. Camundongos Swiss-Webster apresentaram, 28 dias após instilação intranasal de sílica (10 mg), comprometimento da função pulmonar (aumento de resistência e elastância), além de hiper-reatividade à estimulação com metacolina. A análise morfológica e morfométrica revelou intenso infiltrado inflamatório com a presença de fibrose, incluindo aumento na deposição de colágeno e formação de granulomas dispersos no parênquima pulmonar. Vimos, também, aumento na produção de citocinas (TNF-\F061, INF-\F067, IL-1\F062, IL-6, TGF-\F062) e quimiocinas (MCP-1, MIP-1\F061 e MIP-2) no tecido pulmonar de camundongos silicóticos comparados aos animais controles. Aumento de células positivas para F4/80 e \F061- actina de músculo liso (\F061-SMA) foi detectado, indicando a presença de macrófagos e miofibroblastos, respectivamente, no pulmão dos animais silicóticos O tratamento terapêutico com talidomida (25 e 50 mg/kg, v.o.), infliximabe (1,25 mg/kg; i.p.), e com o composto XPro 1595 (10 mg/kg; i.p.), mostrou-se efetivo em reduzir parâmetros importantes do quadro silicótico tais como: i) comprometimento da função pulmonar (aumento de resistência e elastância); ii) componente inflamatório e fibrogênico (deposição de colágeno e formação de granuloma); iii) geração de citocinas inflamatórias/prófibróticas; e iv) presença de células positivas para F4/80 e \F061-SMA. De forma complementar, verificamos que o tratamento com infliximabe e com XPro 1595 foi também capaz de reduzir a ativação/translocação para o núcleo do fator de transcrição NF-\F06BB no pulmão de camundongos silicóticos. Em sistemas biológicos in vitro, vimos que macrófagos alveolares murinos (linhagem AMJ2C11) quando incubados com talidomida, infliximabe e XPro 1595 apresentaram diminuição da geração de TNF-\F061 frente à estimulação com sílica (300 \F06Dg/mL), sem alteração da taxa de viabilidade celular. No sistema de fibroblastos pulmonares, provenientes de camundongos silicóticos, houve redução da resposta proliferativa frente à estimulação com IL-13 (10 - 40 ng/mL) e TNF-\F061 (1 ng/mL) Os três compostos testados não apresentam efeito supressor sobre a produção de IL-8 por células epiteliais pulmonares (linhagem A549) estimuladas com IL-1\F062 (1 ng/mL). Tomados em conjunto, nossos resultados mostram que o tratamento de camundongos silicóticos com compostos neutralizantes da geração ou da ação do TNF-\F061 xii foram eficazes em inibir o comprometimento da função pulmonar e a resposta fibrogênica associadas ao quadro da silicose experimental murina, o que é indicativo de que o TNF-\F061 constitui um alvo terapêutico importante no contexto desta doença. Mais ainda, mostramos de maneira original que a forma solúvel do TNF-\F061 se coloca como de potencial relevância no que tange a terapia de doenças inflamatórias de caráter fibrótico como a silicoseSilicosis is an occupational disease, triggered by silica particle inhalation and is associated with a wide range of mediators. TNF- is a pleiotropic cytokine, which is presented as transmembrane (mTNF-) and soluble (sTNF-) forms, appearing to be involved in several diseases of inflammatory nature. In the present study we evaluated the effect of TNF-neutralizing therapies, including both inhibition of its generation by thalidomide and blockade of its action by the monoclonal antibody infliximab (Remicade®) on the experimental model of silicosis in mice. The selective inhibitor of sTNF- XPro 1595, was also tested. Swiss-Webster mice showed lung function failure (increased resistance and elastance) and hyperreactivity to methacholine aerosolization. Morphological and morphometric analyses revealed an intense inflammatory infiltrate and fibrosis, including increased collagen deposition and granulomas present spread in the lung parenchyma. Elevation in the levels of cytokines (TNF-, IFN-, IL-1, IL-6, TGF-) and chemokines (MCP-1, MIP-1 and MIP-2) in the lung tissue of silicotic mice was also detected as compared to controls. The number of positive cells for F4/80 and -smooth muscle actin (SMA) was shown to elevated in the lungs of silicotic mice, indicating the presence of macrophages and myofibroblasts, respectively. Therapeutic treatment with thalidomide (25 and 50 mg/kg, po), infliximab (1.25 mg/kg, ip) and compound XPro 1595 (10 mg/kg, ip) inhibited important parameters of silicosis such as: i) lung function impairment (increased resistance and elastance), ii) and inflammatory and fibrogenic components (collagen deposition and granuloma formation); iii) cytokine/chemokine generation and iv) presence of F4/80 and -SMA positive cells. In addition, we noted that infliximab and XPro 1595 were also able to reduce nucleus transcription factor NF-B activation/translocation in lung tissue of silicotic mice. We showed in vitro that murine alveolar macrophages (AMJ2C11 cell lineage) when incubated with thalidomide, infliximab and XPro 1595 responded with lower levels of TNF- after silica (300 g/mL) stimulation. No alteration of cell viability was noted. In the case of lung fibroblasts from silicotic mice, stimulated with IL-13 (10 - 40 ng/ml) and TNF- (1ng/ml), we showed that reduced proliferative response occurred after incubation with thalidomide, infliximab and XPro 1595. The three compounds have no effect on IL-8 production by human lung epithelial cells (A549) stimulated with silica (300 g/ml). Taken together, our findings show that treatment of silicotic mice with anti-TNF- therapy, either inhibiting its generation or blocking its activity, was effective to suppress ling function failure and fibrogenesis associated with murine experimental silicosis, which is an indicative that TNF- is an important therapeutic target in this disease. In addition, we showed for the first xiv time that the soluble TNF seems to be a potential therapeutic target in the case of lung fibrotic diseases such as silicosis

Aliskiren Reduces the Adrenal Zona Glomerulosa Apoptosis and Autophagy in Wistar Rats with 2K1C Hypertension

Hypertension is a disease classified as primary or secondary, manifested not only by elevation of blood pressure but also involved in structural and functional changes of target organs. Renal artery stenosis is a leading factor of secondary hypertension, and its progress is associated with overactivation of the renin-angiotensin-aldosterone system (RAAS). Aliskiren is a renin inhibiting drug that suppresses RAAS and culminates in decreased renin release, plasma angiotensin II concentration, and inhibition of aldosterone secretion. In this sense, the aim of the present study was to analyze the structural and ultrastructural morphophysiology of the adrenal glomerular zone, after treatment with aliskiren in Wistar rats with 2K1C hypertension. Parameters as structure and ultrastructure of the adrenal glomerular zone, cellular apoptosis, nuclear cell proliferation, and AT1 receptor expression were analyzed by immunostaining and electron microscopy. Our results showed that the hypertensive animals treated with aliskiren presented a reestablishment of AT1 receptor expression and decrease in apoptosis and autophagy. In addition, treatment with aliskiren improves the cell aspects in the adrenal glomerular zone, evidenced by ultrastructural analysis through preserved nuclei and well-developed mitochondria. Therefore, our evidence suggests that aliskiren has a beneficial effect on the adrenal glomerular zone remodeling in animals with renovascular hypertension