RESUMO
Idiopathic pulmonary fibrosis (IPF) is a progressive and usually lethal disease of unknown aetiology. A growing body of evidence supports that IPF represents an epithelial-driven process characterised by aberrant epithelial cell behaviour, fibroblast/myofibroblast activation and excessive accumulation of extracellular matrix with the subsequent destruction of the lung architecture. The mechanisms involved in the abnormal hyper-activation of the epithelium are unclear, but we propose that recapitulation of pathways and processes critical to embryological development associated with a tissue specific age-related stochastic epigenetic drift may be implicated. These pathways may also contribute to the distinctive behaviour of IPF fibroblasts. Genomic and epigenomic studies have revealed that wingless/Int, sonic hedgehog and other developmental signalling pathways are reactivated and deregulated in IPF. Moreover, some of these pathways cross-talk with transforming growth factor-ß activating a profibrotic feedback loop. The expression pattern of microRNAs is also dysregulated in IPF and exhibits a similar expression profile to embryonic lungs. In addition, senescence, a process usually associated with ageing, which occurs early in alveolar epithelial cells of IPF lungs, likely represents a conserved programmed developmental mechanism. Here, we review the major developmental pathways that get twisted in IPF, and discuss the connection with ageing and potential therapeutic approaches.
Assuntos
Envelhecimento , Fibroblastos/citologia , Fibrose Pulmonar Idiopática/fisiopatologia , Pulmão/patologia , Miofibroblastos/patologia , Transdução de Sinais , Idoso , Animais , Epigênese Genética , Deriva Genética , Humanos , Camundongos , MicroRNAs/metabolismo , Processos Estocásticos , Transcrição GênicaRESUMO
Autophagy is a critical cellular homeostatic process that controls the turnover of damaged organelles and proteins. Impaired autophagic activity is involved in a number of diseases, including idiopathic pulmonary fibrosis suggesting that altered autophagy may contribute to fibrogenesis. However, the specific role of autophagy in lung fibrosis is still undefined. In this study, we show for the first time, how autophagy disruption contributes to bleomycin-induced lung fibrosis in vivo using an Atg4b-deficient mouse as a model. Atg4b-deficient mice displayed a significantly higher inflammatory response at 7 d after bleomycin treatment associated with increased neutrophilic infiltration and significant alterations in proinflammatory cytokines. Likewise, we found that Atg4b disruption resulted in augmented apoptosis affecting predominantly alveolar and bronchiolar epithelial cells. At 28 d post-bleomycin instillation Atg4b-deficient mice exhibited more extensive and severe fibrosis with increased collagen accumulation and deregulated extracellular matrix-related gene expression. Together, our findings indicate that the ATG4B protease and autophagy play a crucial role protecting epithelial cells against bleomycin-induced stress and apoptosis, and in the regulation of the inflammatory and fibrotic responses.
Assuntos
Autofagia/efeitos dos fármacos , Bleomicina/farmacologia , Cisteína Endopeptidases/metabolismo , Homeostase/efeitos dos fármacos , Fibrose Pulmonar Idiopática/metabolismo , Animais , Apoptose/genética , Autofagia/fisiologia , Proteínas Relacionadas à Autofagia , Cisteína Endopeptidases/genética , Citocinas/genética , Citocinas/metabolismo , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Fibrose Pulmonar Idiopática/induzido quimicamente , Camundongos KnockoutRESUMO
RATIONALE: Idiopathic pulmonary fibrosis (IPF) is a devastating disease characterized by epithelial phenotypic changes and fibroblast activation. Based on the temporal heterogeneity of IPF, we hypothesized that hyperplastic alveolar epithelial cells regulate the fibrotic response. OBJECTIVES: To identify novel mediators of fibrosis comparing the transcriptional signature of hyperplastic epithelial cells and conserved epithelial cells in the same lung. METHODS: Laser capture microscope and microarrays analysis were used to identify differentially expressed genes in IPF lungs. Bleomycin-induced lung fibrosis was evaluated in Mmp19-deficient and wild-type (WT) mice. The role of matrix metalloproteinase (MMP)-19 was additionally studied by transfecting the human MMP19 in alveolar epithelial cells. MEASUREMENTS AND MAIN RESULTS: Laser capture microscope followed by microarray analysis revealed a novel mediator, MMP-19, in hyperplastic epithelial cells adjacent to fibrotic regions. Mmp19(-/-) mice showed a significantly increased lung fibrotic response to bleomycin compared with WT mice. A549 epithelial cells transfected with human MMP19 stimulated wound healing and cell migration, whereas silencing MMP19 had the opposite effect. Gene expression microarray of transfected A549 cells showed that PTGS2 (prostaglandin-endoperoxide synthase 2) was one of the highly induced genes. PTGS2 was overexpressed in IPF lungs and colocalized with MMP-19 in hyperplastic epithelial cells. In WT mice, PTGS2 was significantly increased in bronchoalveolar lavage and lung tissues after bleomycin-induced fibrosis, but not in Mmp19(-/-) mice. Inhibition of Mmp-19 by siRNA resulted in inhibition of Ptgs2 at mRNA and protein levels. CONCLUSIONS: Up-regulation of MMP19 induced by lung injury may play a protective role in the development of fibrosis through the induction of PTGS2.
Assuntos
Ciclo-Oxigenase 2/metabolismo , Fibrose Pulmonar Idiopática/enzimologia , Metaloproteinases da Matriz Secretadas/metabolismo , Animais , Bleomicina , Células Cultivadas , Células Epiteliais/metabolismo , Regulação Enzimológica da Expressão Gênica , Humanos , Fibrose Pulmonar Idiopática/genética , Fibrose Pulmonar Idiopática/patologia , Microdissecção e Captura a Laser , Metaloproteinases da Matriz Secretadas/genética , Camundongos , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Alvéolos Pulmonares/metabolismo , Regulação para CimaRESUMO
The acyl-CoA synthetase 4 (ACSL4) is increased in breast cancer, colon and hepatocellular carcinoma. ACSL4 mainly esterifies arachidonic acid (AA) into arachidonoyl-CoA, reducing free AA intracellular levels, which is in contradiction with the need for AA metabolites in tumorigenesis. Therefore, the causal role of ACSL4 is still not established. This study was undertaken to determine the role of ACSL4 in AA metabolic pathway in breast cancer cells. The first novel finding is that ACSL4 regulates the expression of cyclooxygenase-2 (COX-2) and the production of prostaglandin in MDA-MB-231 cells. We also found that ACSL4 is significantly up-regulated in the highly aggressive MDA-MB-231 breast cancer cells. In terms of its overexpression and inhibition, ACSL4 plays a causal role in the control of the aggressive phenotype. These results were confirmed by the increase in the aggressive behaviour of MCF-7 cells stably transfected with a Tet-off ACSL4 vector. Concomitantly, another significant finding was that intramitochondrial AA levels are significantly higher in the aggressive cells. Thus, the esterification of AA by ACSL4 compartmentalizes the release of AA in mitochondria, a mechanism that serves to drive the specific lipooxygenase metabolization of the fatty acid. To our knowledge, this is the first report that ACSL4 expression controls both lipooxygenase and cyclooxygenase metabolism of AA. Thus, this functional interaction represents an integrated system that regulates the proliferating and metastatic potential of cancer cells. Therefore, the development of combinatory therapies that profit from the ACSL4, lipooxygenase and COX-2 synergistic action may allow for lower medication doses and avoidance of side effects.
Assuntos
Coenzima A Ligases/genética , Ciclo-Oxigenase 2/genética , Perfilação da Expressão Gênica , Lipoxigenases/genética , Araquidonato 12-Lipoxigenase/genética , Araquidonato 12-Lipoxigenase/metabolismo , Ácido Araquidônico/metabolismo , Western Blotting , Neoplasias da Mama/enzimologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Coenzima A Ligases/metabolismo , Ciclo-Oxigenase 2/metabolismo , Ácidos Graxos/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Lipoxigenases/metabolismo , Mitocôndrias/metabolismo , Prostaglandinas/metabolismo , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND: Matrix metalloproteinases (MMPs) participate in extracellular matrix degradation in physiological and pathological conditions. The available evidence suggests that MMP-13 plays a significant role in both the initiation and progress of bone resorption. The aim of our study was to identify the presence of MMP-13 in adult patients with untreated chronic periodontitis. We also determined the activity of MMP-13 present in lesions undergoing episodic attachment loss in gingival crevicular fluid (GCF) samples. METHODS: After monitoring at 2 and 4 months, 21 patients showed destructive periodontitis (periodontally affected sites presenting at least two sites with > or =2 mm clinical attachment loss), and GCF samples were collected both from active and inactive sites (21 GCF samples, each). GCF was collected during a 30-second interval using a paper strip, and an immunofluorescence assay was performed to determine the basal activity of MMP-13 and the relationship between 4-aminophenylmercuric acetate (APMA)-activated total MMP-13 and basal MMP-13 activity. Gingival tissues from five patients were fixed in formalin and MMP-13 expression was demonstrated using immunohistochemistry and in situ hybridization. MMP-13 molecular forms were examined by Western immunoblotting with monoclonal antibodies. RESULTS: MMP-13 was found in 100% of GCF samples from patients with chronic periodontitis. Active sites, associated with tissue destruction, had significantly higher proportions of active MMP-13 and MMP-13 activity levels than their inactive counterparts (1.49 versus 1.17 ng fluorescent product, respectively; P <0.05). Western blot, immunohistochemical staining, and in situ hybridization confirmed the presence of MMP-13 in periodontal disease, with observable differences between periodontitis and healthy subjects. MMP-13 immunoreactivities were seen mainly as 55 and 48 kDa, corresponding to partially and fully activated forms, respectively, and a smaller proportion of 60-kDa proenzyme form. CONCLUSION: MMP-13 activity in GCF samples was significantly increased in active sites from progressive periodontal disease, supporting its role in the alveolar bone loss developed in this disease.