RESUMO
Airway remodeling consists of the structural changes of airway walls, which is often considered the result of longstanding airway inflammation, but it may be present to an equivalent degree in the airways of children with asthma, raising the need for early and specific therapeutic interventions. The arachidonic acid cytochrome P-450 (CYP) pathway has thus far received relatively little attention in its relation to asthma. In this study, we studied the inhibition of soluble epoxide hydrolase (sEH) on airway remodeling and hyperresponsiveness (AHR) in a chronic asthmatic model which long-term exposure to antigen over a period of 12 weeks. The expression of sEH and CYP2J2, the level of 14, 15-epoxyeicosatrienoic acids (EETs), airway remodeling, hyperresponsiveness and inflammation were analyzed to determine the inhibition of sEH. The intragastric administration of 3 or 10 mg/kg ZDHXB-101, which is a structural derivative of natural product honokiol and a novel soluble epoxide hydrolase (sEH) inhibitor, daily for 9 weeks significantly increased the level of 14, 15-EETs by inhibiting the expression of sEH and increasing the expression of CYP2J2 in lung tissues. ZDHXB-101 reduced the expression of remodeling-related markers such as interleukin (IL)-13, IL-17, MMP-9 N-cadherin, α-smooth muscle actin, S100A4, Twist, goblet cell metaplasia, and collagen deposition in the lung tissue or in bronchoalveolar lavage fluid. Moreover, ZDHXB-101 alleviated AHR, which is an indicator that is used to evaluate the airway remodeling function. The inhibitory effects of ZDHXB-101 were demonstrated to be related to its direct inhibition of the extracellular signal-regulated kinase (Erk1/2) phosphorylation, as well as inhibition of c-Jun N-terminal kinases (JNK) and the signal transducer and activator of transcription-3 (STAT3) signal transduction. These findings first revealed the anti-remodeling potential of ZDHXB-101 lead in chronic airway disease.
Assuntos
Remodelação das Vias Aéreas/efeitos dos fármacos , Antiasmáticos/uso terapêutico , Asma/tratamento farmacológico , Asma/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Fator de Transcrição STAT3/metabolismo , Remodelação das Vias Aéreas/fisiologia , Animais , Antiasmáticos/química , Antiasmáticos/farmacologia , Citocromo P-450 CYP2J2 , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Dose-Resposta a Droga , Epóxido Hidrolases/metabolismo , Feminino , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Camundongos Endogâmicos ICR , Fator de Transcrição STAT3/antagonistas & inibidoresRESUMO
Cigarette smoke (CS) is a major risk factor for the development of lung cancer and chronic obstructive pulmonary disease (COPD). Epithelial-mesenchymal transition (EMT) commonly coexists in lung cancer and COPD. CS triggers many factors including matrix metalloproteinases (MMPs) production, contributing to EMT progression in the lungs. Here, how Shp2 signaling regulates the CS-induced MMP-9 production and EMT progression were investigated in mouse lungs and in pulmonary epithelial cell cultures (NCI-H292) found CS induced MMP-9 production, EMT progression (increased vimentin and α-SMA; decreased E-cadherin) and collagen deposition in lung tissues; cigarette smoke extract (CSE) induced MMP-9 production and EMT-related phenotypes in NCI-H292 cells, which were partially prevented by Shp2 KO/KD or Shp2 inhibition. The CSE exposure induced EMT phenotypes were suppressed by MMP-9 inhibition. Recombinant MMP-9 induced EMT, which was prevented by MMP-9 inhibition or Shp2 KD/inhibition. Mechanistically, CS and CSE exposure resulted in ERK1/2, JNK and Smad2/3 phosphorylation, which were suppressed by Shp2 KO/KD/inhibition. Consequentially, the CSE exposure-induced MMP-9 production and EMT progression were suppressed by ERK1/2, JNK and Smad2/3 inhibitors. Thus, CS induced MMP-9 production and EMT resulted from activation of Shp2/ERK1/2/JNK/Smad2/3 signaling pathways. Our study contributes to the underlying mechanisms of pulmonary epithelial structural changes in response to CS, which may provide novel therapeutic solutions for treating associated diseases, such as COPD and lung cancer.
Assuntos
Fumar Cigarros/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Metaloproteinase 9 da Matriz/biossíntese , Proteína Tirosina Fosfatase não Receptora Tipo 11/biossíntese , Animais , Linhagem Celular Tumoral , Fumar Cigarros/efeitos adversos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Exposição por Inalação/efeitos adversos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologiaRESUMO
Cigarette smoke (CS) is a major risk factor for the development of chronic obstructive pulmonary disease (COPD). Our previous studies have indicated that Rac1 is involved in lipopolysaccharide-induced pulmonary injury and CS-mediated epithelial-mesenchymal transition. However, the contribution of Rac1 activity to CS-induced lung inflammation remains not fully clear. In this study, we investigated the regulation of Rac1 in CS-induced pulmonary inflammation. Mice or 16HBE cells were exposed to CS or cigarette smoke extract (CSE) to induce acute inflammation. The lungs of mice exposed to CS showed an increase in the release of interleukin-6 (IL-6) and keratinocyte-derived chemokine (KC), as well as an accumulation of inflammatory cells, indicating high Rac1 activity. The exposure of 16HBE cells to CSE resulted in elevated Rac1 levels, as well as increased release of IL-6 and interleukin-8 (IL-8). Selective inhibition of Rac1 ameliorated the release of IL-6 and KC as well as inflammation in the lungs of CS-exposed mice. Histological assessment showed that treatment with a Rac1 inhibitor, NSC23766, led to a decrease in CD68 and CD11b positive cells and the infiltration of neutrophils and macrophages into the alveolar spaces. Selective inhibition or knockdown of Rac1 decreased IL-6 and IL-8 release in 16HBE cells induced by CSE, which correlated with CSE-induced Rac1-regulated Erk1/2 mitogen-activated protein kinase (MAPK) and signal transducer and activator of transcription-3 (STAT3) signaling. Our data suggest an important role for Rac1 in the pathological alterations associated with CS-mediated inflammation. Rac1 may be a promising therapeutic target for the treatment of CS-induced pulmonary inflammation.
Assuntos
Fumar Cigarros/efeitos adversos , Pulmão/metabolismo , Sistema de Sinalização das MAP Quinases , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Neuropeptídeos/metabolismo , Pneumonia/metabolismo , Fator de Transcrição STAT3/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo , Animais , Fumar Cigarros/genética , Fumar Cigarros/metabolismo , Citocinas/genética , Citocinas/metabolismo , Inflamação/etiologia , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Pulmão/patologia , Camundongos , Proteína Quinase 3 Ativada por Mitógeno/genética , Neuropeptídeos/genética , Infiltração de Neutrófilos/genética , Neutrófilos/metabolismo , Neutrófilos/patologia , Pneumonia/etiologia , Pneumonia/genética , Pneumonia/patologia , Fator de Transcrição STAT3/genética , Proteínas rac1 de Ligação ao GTP/genéticaRESUMO
BACKGROUND: Epithelial-mesenchymal transition (EMT) is the major pathophysiological process in lung fibrosis observed in chronic obstructive pulmonary disease (COPD) and lung cancer. Smoking is a risk factor for developing EMT, yet the mechanism remains largely unknown. In this study, we investigated the role of Rac1 in cigarette smoke (CS) induced EMT. METHODS: EMT was induced in mice and pulmonary epithelial cells by exposure of CS and cigarette smoke extract (CSE) respectively. RESULTS: Treatment of pulmonary epithelial cells with CSE elevated Rac1 expression associated with increased TGF-ß1 release. Blocking TGF-ß pathway restrained CSE-induced changes in EMT-related markers. Pharmacological inhibition or knockdown of Rac1 decreased the CSE exposure induced TGF-ß1 release and ameliorated CSE-induced EMT. In CS-exposed mice, pharmacological inhibition of Rac1 reduced TGF-ß1 release and prevented aberrations in expression of EMT markers, suggesting that Rac1 is a critical signaling molecule for induction of CS-stimulated EMT. Furthermore, Rac1 inhibition or knockdown abrogated CSE-induced Smad2 and Akt (PKB, protein kinase B) activation in pulmonary epithelial cells. Inhibition of Smad2, PI3K (phosphatidylinositol 3-kinase) or Akt suppressed CSE-induced changes in epithelial and mesenchymal marker expression. CONCLUSIONS AND GENERAL SIGNIFICANCE: Altogether, these data suggest that CS initiates EMT through Rac1/Smad2 and Rac1/PI3K/Akt signaling pathway. Our data provide new insights into the fundamental basis of EMT and suggest a possible new course of therapy for COPD and lung cancer.
Assuntos
Transição Epitelial-Mesenquimal , Neuropeptídeos/fisiologia , Nicotiana/efeitos adversos , Alvéolos Pulmonares/patologia , Fumaça/efeitos adversos , Proteínas rac1 de Ligação ao GTP/fisiologia , Animais , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Proteína Smad2/fisiologia , Fator de Crescimento Transformador beta1/análise , Fator de Crescimento Transformador beta1/biossínteseRESUMO
Bencycloquidium bromide (BCQB), a novel M3 receptor antagonist, alleviates airway hyperresponsiveness, inflammation, and airway remodeling in a murine model of asthma. The aim of this study was to investigate the anti-inflammatory activity of inhaled BCQB in a cigarette smoke (CS)-induced model of acute lung inflammation. Mice exposed to CS developed chronic obstructive pulmonary disease (COPD). Inhalation of BCQB suppressed the accumulation of neutrophils and macrophages in airways and lung and also inhibited the CS-induced increases in mRNA levels of keratinocyte-derived chemokine, monocyte chemotactic protein-1, tumor necrosis factor-alpha, and interleukin-1ß in lung and protein expression levels in bronchoalveolar lavage fluid. Moreover, BCQB (300 µg/ml) inhibited the CS-induced changes in superoxide dismutase and myeloperoxidase activities in the lungs. Our study suggests that BCQB might be a potential therapy for inflammation in CS-induced pulmonary diseases, including COPD.
Assuntos
Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Modelos Animais de Doenças , Nicotiana/efeitos adversos , Pneumonia/tratamento farmacológico , Receptor Muscarínico M3/antagonistas & inibidores , Fumaça/efeitos adversos , Administração por Inalação , Animais , Relação Dose-Resposta a Droga , Feminino , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos ICR , Pneumonia/metabolismo , Pneumonia/patologia , Fumar/efeitos adversos , Fumar/metabolismo , Fumar/patologia , Resultado do TratamentoRESUMO
Cigarette smoke (CS), the major cause of chronic obstructive pulmonary disease, contains a variety of oxidative components that were implicated in the regulation of Src homology domain 2-containing protein tyrosine phosphatase 2 (Shp2) activity. However, the contribution of Shp2 enzyme to chronic obstructive pulmonary disease pathogenesis remains unclear. We investigated the role of Shp2 enzyme in blockading CS-induced pulmonary inflammation. Shp2 levels were assessed in vivo and in vitro. Mice (C57BL/6) or pulmonary epithelial cells (NCI-H292) were exposed to CS or cigarette smoke extract (CSE) to induce acute injury and inflammation. Lungs of smoking mice showed increased levels of Shp2, compared with those of controls. Treatment of lung epithelial cells with CSE showed elevated levels of Shp2 associated with the increased release of IL-8. Selective inhibition or knockdown of Shp2 resulted in decreased IL-8 release in response to CSE treatment in pulmonary epithelial cells. In comparison with CS-exposed wild-type mice, selective inhibition or conditional knockout of Shp2 in lung epithelia reduced IL-8 release and pulmonary inflammation in CS-exposed mice. In vitro biochemical data correlate CSE-mediated IL-8 release with Shp2-regulated epidermal growth factor receptor/Grb-2-associated binders/MAPK signaling. Our data suggest an important role for Shp2 in the pathological alteration associated with CS-mediated inflammation. Shp2 may be a potential target for therapeutic intervention for inflammation in CS-induced pulmonary diseases.
Assuntos
Pneumonia/imunologia , Pneumonia/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/fisiologia , Fumar/efeitos adversos , Fumar/patologia , Produtos do Tabaco/toxicidade , Doença Aguda , Animais , Linhagem Celular , Modelos Animais de Doenças , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/prevenção & controle , Interleucina-8/metabolismo , Interleucina-8/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Pneumonia/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 11/antagonistas & inibidores , Proteína Tirosina Fosfatase não Receptora Tipo 11/deficiência , Alvéolos Pulmonares/imunologia , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/patologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Fumar/metabolismoRESUMO
Cigarette smoke contains reactive oxygen (ROS) that can cause oxidative stress. It increases the number of apoptotic and necrotic lung cells and further induces the development of chronic airway disease. In this study, we investigated the effects of cigarette smoke extract (CSE) on apoptosis in human bronchial epithelial cells (BEAS-2B). CSE exposure induced ROS generation and p38 mitogen-activated protein kinase (MAPK) activation that are associated with the activation of apoptosis-regulating signal kinase 1 (ASK-1). N-acetylcysteine (a general antioxidant) attenuated the CSE-induced ASK-1 and p38 MAPK activation and cell apoptosis, suggesting a triggering role of ROS in ASK-1/p38 MAPK activation during apoptotic progression. In contrast, the inhibition and knockdown of p38 attenuated the expression of anti-oxidant master NF-E2-related factor 2 (Nrf-2) and CSE-induced apoptosis, suggesting that p38 MAPK modulates Nrf-2 expression and presumably prevents cell apoptosis. Taken together, the data presented in this manuscript demonstrate that the ROS-dependent ASK-1/p38 signaling cascade regulates CSE-induced BEAS-2B cell apoptosis. In addition, anti-oxidative Nrf-2 is also up-regulated by the ROS/p38 signaling cascade in this progression.
Assuntos
Apoptose/efeitos dos fármacos , Brônquios/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/agonistas , Mucosa Respiratória/efeitos dos fármacos , Fumar/efeitos adversos , Regulação para Cima/efeitos dos fármacos , Acetilcisteína/farmacologia , Elementos de Resposta Antioxidante/efeitos dos fármacos , Antioxidantes/farmacologia , Brônquios/enzimologia , Brônquios/metabolismo , Linhagem Celular , Misturas Complexas/antagonistas & inibidores , Misturas Complexas/toxicidade , Ativação Enzimática/efeitos dos fármacos , Inativação Gênica , Humanos , MAP Quinase Quinase Quinase 5/antagonistas & inibidores , MAP Quinase Quinase Quinase 5/química , MAP Quinase Quinase Quinase 5/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Espécies Reativas de Oxigênio/agonistas , Espécies Reativas de Oxigênio/metabolismo , Mucosa Respiratória/enzimologia , Mucosa Respiratória/metabolismo , Fumaça , Produtos do Tabaco , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/química , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: Despite intensive studies focused on the pathophysiology of asthmatic inflammation, little is known about how cross-talk between neuroendocrine and immune systems regulates the inflammatory response during an asthmatic attack. We recently showed corresponding changes of cytokines and leukotriene B4 (LTB4) in brain and lung tissues of antigen-challenged asthmatic rats. Here, we investigated how LTB4 interacts with the neuroendocrine-immune system in regulating antigen-induced asthmatic responses in sensitized guinea pigs. METHODS: Ovalbumin-sensitized guinea pigs were challenged by inhalation of antigen. Vehicle, LTB4 or U75302 (a selective LTB4 BLT1 receptor inhibitor) was given via intracerebroventricular injection (i.c.v.) 30 min before challenge. Airway contraction response was evaluated using Penh values before and after antigen challenge. The inflammatory response in lung tissue was evaluated 24 h after challenge. The LTB4 content of lung and brain homogenate preparations was detected by reversed phase high-performance liquid chromatography (RP-HPLC). Plasma levels of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were measured using ELISA kits. RESULTS: Antigen challenge impaired pulmonary function and increased inflammatory cell infiltration in lung tissue. These responses could be significantly suppressed by LTB4, 30 ng i.c.v., in ovalbumin-sensitized guinea pigs. LTB4 content of lung and brain homogenates from antigen-challenged guinea pigs was significantly increased. In addition, administration of LTB4 via i.c.v. markedly increased CORT and ACTH level in plasma before antigen challenge, and there were further increases in CORT and ACTH levels in plasma after antigen challenge. U75302, 100 ng i.c.v., completely blocked the effects of LTB4. In addition, U75302, 100 ng via i.c.v. injection, markedly decreased LTB4 content in lung homogenates, but not in brain homogenates. CONCLUSIONS: Increased LTB4 levels in brain during asthmatic attacks down-regulates airway contraction response and inflammation through the BLT1 receptor. Stimulation of the hypothalamic-pituitary-adrenal axis by LTB4 may result in an increase in systemic glucocorticoids which, in turn, would feed back to suppress the asthmatic response.
Assuntos
Asma/tratamento farmacológico , Leucotrieno B4/administração & dosagem , Hormônio Adrenocorticotrópico/sangue , Análise de Variância , Animais , Asma/sangue , Asma/induzido quimicamente , Líquido da Lavagem Broncoalveolar , Cromatografia de Fase Reversa/métodos , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática/métodos , Álcoois Graxos/farmacologia , Glicóis/farmacologia , Cobaias , Hidrocortisona/sangue , Injeções Intraventriculares/métodos , Hemorragias Intracranianas/induzido quimicamente , Leucotrieno B4/antagonistas & inibidores , Pulmão/patologia , Pulmão/fisiopatologia , Ovalbumina , Fatores de TempoRESUMO
BACKGROUND: Basic and clinical studies suggest that hypothalamic-pituitary-adrenal (HPA) axis is the neuroendocrine-immune pathway that functionally regulates the chronic inflammatory disease including asthma. Our previous studies showed corresponding changes of cytokines and leukotriene B4 (LTB4) between brain and lung tissues in antigen-challenged asthmatic rats. Here, we investigated how the increased LTB4 level in brain interacts with HPA axis in regulating antigen-induced asthmatic response in sensitized rats. METHODS: Ovalbumin-sensitized rats were challenged by inhalation of antigen. Rats received vehicle, LTB4 or U75302 (a selective LTB4 BLT1 receptor inhibitor) was given via intracerebroventricular injection (i.c.v) 30 min before challenge. Lung resistance (RL) and dynamic lung compliance (Cdyn) were measured before and after antigen challenge. Inflammatory response in lung tissue was assessed 24 h after challenge. Expression of CRH mRNA and protein in hypothalamus were evaluated by RT-PCR and Western Blot, and plasma levels of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were measured using the ELISA kits. RESULTS: Antigen challenge decreased pulmonary function and induced airway inflammation, evoked HPA axis response in sensitized rats. Administration of LTB4 via i.c.v markedly attenuated airway contraction and inflammation. Meanwhile, LTB4 via i.c.v markedly increased CORT and ACTH level in plasma before antigen challenge, and followed by further increases in CORT and ACTH levels in plasma after antigen challenge in sensitized rats. Expression of CRH mRNA and protein in hypothalamus were also significantly increased by LTB4 via i.c.v in sensitized rats after antigen challenge. These effect were completely blocked by pre-treatment with BLT1 receptor antagonist U75302 (10 ng), but not by BLT2 antagonist LY255283. CONCLUSIONS: LTB4 administered via i.c.v down-regulates the airway contraction response and inflammation through activation of the HPA axis via its BLT1 receptor. This study expands our concept of the regulatory role of intracranial inflammatory mediators in inflammatory diseases including asthma. The favourable effects of LTB4 on the HPA axis may help to explain the phenomenon of self-relief after an asthmatic attack.
Assuntos
Asma/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Leucotrieno B4/metabolismo , Pulmão/metabolismo , Sistema Hipófise-Suprarrenal/metabolismo , Receptores do Leucotrieno B4/metabolismo , Hormônio Adrenocorticotrópico/sangue , Resistência das Vias Respiratórias , Animais , Asma/imunologia , Asma/fisiopatologia , Western Blotting , Hormônio Liberador da Corticotropina/genética , Hormônio Liberador da Corticotropina/metabolismo , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Álcoois Graxos/administração & dosagem , Feminino , Glicóis/administração & dosagem , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/imunologia , Hipotálamo/imunologia , Hipotálamo/metabolismo , Mediadores da Inflamação/metabolismo , Injeções Intraventriculares , Leucotrieno B4/administração & dosagem , Pulmão/imunologia , Pulmão/fisiopatologia , Complacência Pulmonar , Masculino , Ovalbumina , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/imunologia , Eosinofilia Pulmonar/imunologia , Eosinofilia Pulmonar/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores do Leucotrieno B4/agonistas , Receptores do Leucotrieno B4/antagonistas & inibidores , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Airway remodeling in asthma is difficult to treat because of its complex pathophysiology that involves proinflammatory cytokines, as well as the arachidonic acid cytochrome P-450 (CYP) pathway; however, it has received little attention. In this study, we assessed the efficacy of a soluble epoxide hydrolase (sEH) on airway remodeling in a mouse model of chronic asthma. The expression of sEH and CYP2J2 and the level of 14,15-epoxyeicosatrienoic acid (14,15-EET), airway remodeling and hyperresponsiveness (AHR) were analyzed to determine the level of sEH inhibition. AUDA, a sEH inhibitor, was given daily for 9 weeks orally, which significantly increased the level of 14,15-EET by inhibiting the expression of sEH and increasing the expression of CYP2J2 in lung tissues. The inhibition of sEH reduced the expression of remodeling-related molecular markers, such as interleukin (IL)-13, IL-17, matrix metalloproteinase 9, N-cadherin, α-smooth muscle actin (α-SMA), S100A4, Twist, epithelial goblet cell metaplasia, and collagen deposition in bronchoalveolar lavage fluid (BAL fluid) and lung tissues. Moreover, remodeling-related eosinophil accumulation in the BAL fluid and infiltration into the lung tissue were improved by AUDA. Finally, AUDA alleviated AHR, which is a functional indicator of airway remodeling. The effect of AUDA on airway remodeling was related to the downregulation of extracellular-regulated protein kinases (Erk1/2), c-Jun N-terminal kinases (JNK) and signal transducer and activator of transcription 3 (STAT3). To our knowledge, this is the first report to demonstrate that inhibition of sEH exerts significant protective effects on airway remodeling in asthma.
Assuntos
Adamantano/análogos & derivados , Remodelação das Vias Aéreas/efeitos dos fármacos , Asma/tratamento farmacológico , Epóxido Hidrolases/antagonistas & inibidores , Ácidos Láuricos/farmacologia , Pulmão/efeitos dos fármacos , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido 8,11,14-Eicosatrienoico/sangue , Adamantano/farmacologia , Adamantano/uso terapêutico , Remodelação das Vias Aéreas/imunologia , Animais , Asma/imunologia , Asma/patologia , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Citocromo P-450 CYP2J2 , Sistema Enzimático do Citocromo P-450/metabolismo , Modelos Animais de Doenças , Epóxido Hidrolases/metabolismo , Feminino , Humanos , Ácidos Láuricos/uso terapêutico , Pulmão/imunologia , Pulmão/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/imunologia , Camundongos , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologiaRESUMO
OBJECTIVES: Cigarette smoke (CS) is a major risk factor for the development of lung cancer and chronic obstructive pulmonary disease (COPD). Epithelial-mesenchymal transition (EMT) is found in invasive or metastatic phenotypes in lung cancer and COPD. MK-2206, a pan Akt inhibitor, has failed in clinical trials for solid tumors when administered alone at tolerated doses, but it has been shown to have synergistic effects when applied with certain molecular targeted agents. In this study, we investigated the working mechanism of MK-2206 in CS-induced pulmonary EMT both in vivo and in vitro. MATERIALS AND METHODS: The expression of Akt, epithelial-mesenchymal transition (EMT) markers and signaling proteins were analyzed by immunohistochemistry, real-time PCR and Western blot in cigarette smoke extract (CSE)-treated pulmonary epithelia and CS-treated lung tissues in mice. RESULTS AND CONCLUSION: We demonstrated that exposure of the epithelium to CSE and exposure of the mice to CS can induce EMT by activating the Akt signaling pathway. Intragastric application of MK-2206 at a low dose (50â¯mg/kg) reversed the changes of the key indicators of EMT in the lungs of CS-exposed mice, including TGF-ß1, α-SMA, vimentin, MMP-9, MMP-2, S100A4, collagen deposition, and E-cadherin. MK-2206 at a non-cytotoxic concentration (0.5⯵M) or Akt knockdown consistently reversed the changes of the key indicators of EMT in the pulmonary epithelia. Moreover, we found that the effects of Akt inhibition or knockdown on the CS/CSE-induced EMT acted via the TGF-ß1/Akt/Smad/mTOR and Akt/P38 MAPK pathways. Taken together, our data offer a novel perspective on the molecular mechanism of Akt for CS-induced EMT. This finding may enhance the understanding of the mechanism behind the synergistic use of a low dose of MK-2206 to achieve antitumor efficacy with reduced adverse reactions in patients with lung cancer and COPD.
Assuntos
Neoplasias Pulmonares/metabolismo , Pulmão/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Mucosa Respiratória/metabolismo , Animais , Células Cultivadas , Fumar Cigarros/efeitos adversos , Transição Epitelial-Mesenquimal , Feminino , Compostos Heterocíclicos com 3 Anéis/administração & dosagem , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Doença Pulmonar Obstrutiva Crônica/patologia , Mucosa Respiratória/patologia , Transdução de SinaisRESUMO
Bleomycin (BLM) has potent tumor cell-killing properties that have given it an important place in cancer chemotherapy, but pulmonary toxicity is its major adverse effect. Soluble epoxide hydrolase (sEH) inhibitors have been reported to have protective effects in fibrosis models, but the effects of AUDA, an sEH inhibitor of BLM-induced pulmonary toxicity and fibrosis, remain to be researched. In this study, we assessed the effects of AUDA on the BLM-induced pulmonary fibrosis in a mouse model, and transforming growth factor (TGF)-ß1-induced epithelial proliferation and epithelial-mesenchymal transition (EMT) in vitro by monitoring changes in pulmonary function, inflammatory response, fibrotic remodeling, and signaling pathways. AUDA was administered by intragastric administration (i.g) daily for three weeks, starting at seven days after intratracheal instillation of BLM. All examinations were performed 24h after the last i.g. In vivo, AUDA significantly improved BLM-induced decline in lung function and body weight, and inhibited inflammatory cell accumulation and the mRNA and protein expression of interleukin (IL)-1ß, TGF-ß1, and matrix metalloproteinase 9 (MMP-9) in lung tissue. Moreover, AUDA attenuated BLM-induced deposition of collagen fibers, destruction of alveolar structures, and pulmonary parenchyma. Additionally, AUDA regulated the expression of α-smooth muscle actin (α-SMA) and E-cadherin by inhibiting the Smad3/p38 signaling pathway. In vitro, AUDA significantly inhibited TGF-ß1-induced epithelial cells and fibroblast proliferation, reduced sEH expression and α-SMA expression, and increased epoxyeicosatrienoic acid (EET) levels and E-cadherin expression in epithelial cells. These effects were blocked by AUDA by downregulating the Smad3 and p38 signaling pathways. Taken together, these data indicate that treatment with sEH inhibitors may improve BLM-induced pulmonary toxicity.
Assuntos
Adamantano/análogos & derivados , Bleomicina , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Ácidos Láuricos/farmacologia , Pulmão/efeitos dos fármacos , Fibrose Pulmonar/prevenção & controle , Proteína Smad3/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adamantano/farmacologia , Animais , Biomarcadores/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Colágeno/metabolismo , Citocinas/metabolismo , Citoproteção , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Epóxido Hidrolases/metabolismo , Feminino , Humanos , Mediadores da Inflamação/metabolismo , Pulmão/enzimologia , Pulmão/patologia , Pulmão/fisiopatologia , Camundongos Endogâmicos ICR , Pneumonia/induzido quimicamente , Pneumonia/enzimologia , Pneumonia/prevenção & controle , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/enzimologia , Fibrose Pulmonar/patologia , Transdução de Sinais/efeitos dos fármacos , Fatores de TempoRESUMO
Pulmonary fibrosis is common in a variety of inflammatory lung diseases, such as interstitial pneumonia, chronic obstructive pulmonary disease, and silicosis. There is currently no effective clinical drug treatment. It has been reported that grape seed extracts (GSE) has extensive pharmacological effects with minimal toxicity. Although it has been found that GSE can improve the lung collagen deposition and fibrosis pathology induced by bleomycin in rat, its effects on pulmonary function, inflammation, growth factors, matrix metalloproteinases and epithelial-mesenchymal transition remain to be researched. In the present study, we studied whether GSE provided protection against bleomycin (BLM)-induced mouse pulmonary fibrosis. ICR strain mice were treated with BLM in order to establish pulmonary fibrosis models. GSE was given daily via intragastric administration for three weeks starting at one day after intratracheal instillation. GSE at 50 or 100mg/kg significantly reduced BLM-induced inflammatory cells infiltration, proinflammatory factor protein expression, and hydroxyproline in lung tissues, and improved pulmonary function in mice. Additionally, treatment with GSE also significantly impaired BLM-induced increases in lung fibrotic marker expression (collagen type I alpha 1 and fibronectin 1) and decreases in an anti-fibrotic marker (E-cadherin). Further investigation indicated that the possible molecular targets of GSE are matrix metalloproteinases-9 (MMP-9) and TGF-ß1, given that treatment with GSE significantly prevented BLM-induced increases in MMP-9 and TGF-ß1 expression in the lungs. Together, these results suggest that supplementation with GSE may improve the quality of life of lung fibrosis patients by inhibiting MMP-9 and TGF-ß1 expression in the lungs.
Assuntos
Anti-Inflamatórios/uso terapêutico , Antibióticos Antineoplásicos/toxicidade , Bleomicina/toxicidade , Extrato de Sementes de Uva/uso terapêutico , Pulmão/efeitos dos fármacos , Fibrose Pulmonar/prevenção & controle , Animais , Anti-Inflamatórios/administração & dosagem , Líquido da Lavagem Broncoalveolar/química , Feminino , Extrato de Sementes de Uva/administração & dosagem , Pulmão/imunologia , Pulmão/patologia , Metaloproteinase 9 da Matriz/metabolismo , Camundongos Endogâmicos ICR , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/imunologia , Fibrose Pulmonar/patologia , Testes de Função Respiratória , Fator de Crescimento Transformador beta/metabolismoRESUMO
Inhaled corticosteroid (ICS) therapy in combination with long-acting ß-adrenergic agonists (LABA) is the most important treatment for allergic asthma, although the mechanism still remains unclear. However, mast cells play a central role in the pathogenesis of asthma. In this study, we explored the sole or synergetic effects of des-ciclesonide (ICS) and formoterol (LABA) on the cytokines IL-4 and IL-13 and on histamine release from mast cells (RBL-2H3 cells). We found that des-ciclesonide (0.1, 1 and 10nM) and formoterol (0.1, 1 and 10µM) alone attenuated DNP-BSA-induced IL-4 and IL-13 production, respectively, in a concentration-dependent manner in DNP-IgE-sensitized mast cells. Des-ciclesonide (0.2nM) and formoterol (1µM) alone also reduced histamine production. However, the combination of des-ciclesonide (0.2nM) and formoterol (1µM) had a synergistic inhibition effect on IL-4 mRNA expression and protein production but not IL-13 and histamine release. The JNK inhibitor SP600125 (10µM) inhibited antigen-induced mRNA expression and protein production of IL-4. Des-ciclesonide and formoterol alone inhibited the activation of JNK in a concentration-dependent manner, and the combination of des-ciclesonide (0.2nM) and formoterol (1µM) exhibited greater inhibition effect compared with des-ciclesonide (0.2nM) or formoterol (1µM) alone. Taken together, these synergistic effects on mast cells might provide the rationale for the development of the most recent ICS/LABA combination approved for asthma therapy.
Assuntos
Corticosteroides/farmacologia , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Dinitrofenóis/imunologia , Fumarato de Formoterol/farmacologia , Imunoglobulina E/imunologia , Interleucina-4/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Mastócitos/efeitos dos fármacos , Pregnenodionas/farmacologia , Soroalbumina Bovina/imunologia , Animais , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Quimioterapia Combinada , Ativação Enzimática , Regulação da Expressão Gênica , Liberação de Histamina/efeitos dos fármacos , Interleucina-13/metabolismo , Interleucina-4/genética , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Mastócitos/enzimologia , Mastócitos/imunologia , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacosRESUMO
AIM: This systematic review and meta-analysis was performed to assess the efficacy and side effects between single-agent and doublet chemotherapy in first-line treatment of advanced non-small cell lung cancer with performance status 2 (PS2). METHODS: We searched for randomized controlled trials in online electronic databases and extracted data from eligible studies for meta-analysis. Pooled hazard ratio (HR) for overall survival (OS), pooled risk difference (RD) for 1-year survival, pooled risk ratio (RR) for objective response rate (ORR) and adverse effects were calculated using a fixed-effect model. RESULTS: Six trials with 386 participants in the single-agent group and 389 participants in the doublet group were included in this review. Compared with single-agent chemotherapy, doublet significantly improved OS (HR 0.72; 95% CI 0.61-0.84; P < 0.0001) and significantly increased 1-year survival rate (RD -0.09; 95% CI -0.14 to -0.03; P = 0.004) and ORR (RR 0.4; 95% CI 0.30-0.69; P = 0.0002). Doublet chemotherapy also significantly increased the risk of grade 3/4 neutropenia (RR = 4.97; 95% CI 2.93-8.43; P < 0.00001), thrombocytopenia (RR = 10.29; 95% CI 3.80-27.85; P < 0.00001) and anemia (RR = 2.50; 95% CI 1.27-4.90; P = 0.008). CONCLUSIONS: Our study implies that carboplatin-containing doublet chemotherapy improved OS, 1-year survival rate and ORR, but increased the risk of grade 3/4 hematotoxicity. Carboplatin-containing doublet may well be superior to non-carboplatin-containing treatment.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Feminino , Humanos , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Masculino , Taxa de SobrevidaRESUMO
In response to endothelial cell activation, arachidonic acid can be converted by cytochrome P450 (CYP) epoxygenases to epoxyeicosatrienoic acids (EETs), which have potent vasodilator and anti-inflammatory properties. In this study, we investigated the effects of exogenous EETs on cigarette smoke extract (CSE)-induced inflammation in human bronchial epithelial cells (NCI-H292). We found that CSE inhibited the expression of CYP2C8 and mildly stimulated the expression of epoxide hydrolase 2 (EPHX2) but did not change the expression of CYP2J2. Treatment with 11,12-EET or 14,15-EET attenuated the CSE-induced release of interleukin (IL)-8 by inhibiting the phosphorylation of p38 mitogen-activated protein kinases (MAPKs). Our results demonstrated that CSE may reduce the anti-inflammatory ability of epithelial cells themselves by lowering the EET level. EETs from pulmonary epithelial cells may play a critical protective role on epithelial cell injury.
Assuntos
Ácido 8,11,14-Eicosatrienoico/farmacologia , Anti-Inflamatórios/farmacologia , Brônquios/citologia , Células Epiteliais/efeitos dos fármacos , Interleucina-8/genética , Fumar/efeitos adversos , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Brônquios/efeitos dos fármacos , Linhagem Celular , Citocromo P-450 CYP2J2 , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Células Epiteliais/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-8/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Vasodilatadores/farmacologiaRESUMO
Cytochrome P-450 epoxygenase (EPOX)-derived epoxyeicosatrienoic acids (EETs), 5-lipoxygenase (5-LO), and leukotriene B4 (LTB4), the product of 5-LO, all play a pivotal role in the vascular inflammatory process. We have previously shown that EETs can alleviate oxidized low-density lipoprotein (ox-LDL)-induced endothelial inflammation in primary rat pulmonary artery endothelial cells (RPAECs). Here, we investigated whether ox-LDL can promote LTB4 production through the 5-LO pathway. We further explored how exogenous EETs influence ox-LDL-induced LTB4 production and activity. We found that treatment with ox-LDL increased the production of LTB4 and further led to the expression and release of both monocyte chemoattractant protein-1 (MCP-1/CCL2) and intercellular adhesion molecule-1 (ICAM-1). All of the above ox-LDL-induced changes were attenuated by the presence of 11,12-EET and 14,15-EET, as these molecules inhibited the 5-LO pathway. Furthermore, the LTB4 receptor 1 (BLT1 receptor) antagonist U75302 attenuated ox-LDL-induced ICAM-1 and MCP-1/CCL2 expression and production, whereas LY255283, a LTB4 receptor 2 (BLT2 receptor) antagonist, produced no such effects. Moreover, in RPAECs, we demonstrated that the increased expression of 5-LO and BLT1 following ox-LDL treatment resulted from the activation of nuclear factor-κB (NF-κB) via the p38 mitogen-activated protein kinase (MAPK) pathway. Our results indicated that EETs suppress ox-LDL-induced LTB4 production and subsequent inflammatory responses by downregulating the 5-LO/BLT1 receptor pathway, in which p38 MAPK phosphorylation activates NF-κB. These results suggest that the metabolism of arachidonic acid via the 5-LO and EPOX pathways may present a mutual constraint on the physiological regulation of vascular endothelial cells.
Assuntos
Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Araquidonato 5-Lipoxigenase/química , Leucotrieno B4/metabolismo , Lipoproteínas LDL/farmacologia , Artéria Pulmonar/metabolismo , Receptores do Leucotrieno B4/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Ácido 8,11,14-Eicosatrienoico/farmacologia , Animais , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/metabolismo , Western Blotting , Células Cultivadas , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Masculino , Fosforilação/efeitos dos fármacos , Artéria Pulmonar/citologia , Artéria Pulmonar/efeitos dos fármacos , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Receptores do Leucotrieno B4/genética , Receptores do Leucotrieno B4/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Vasodilatadores/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Oxidized low-density lipoprotein (Ox-LDL) is associated with atherosclerotic events through the modulation of arachidonic acid (AA) metabolism and activation of inflammatory signaling. Cytochrome P450 (CYP) epoxygenase-derived epoxyeicosatrienoic acids (EETs) mitigate inflammation through nuclear factor-κB (NF-κB). In this study, we explored the effects and mechanisms of exogenous EETs on the ox-LDL-induced inflammation of pulmonary artery endothelial cells (PAECs), which were cultured from rat pulmonary arteries. We determined that pre-treatment with 11,12-EET or 14,15-EET attenuated the ox-LDL-induced expression and release of intercellular adhesion molecule-1 (ICAM-1), E-selectin, and monocyte chemoattractant protein-1 (MCP-1) in a concentration-dependent manner. In addition, the ox-LDL-induced expression of CYP2J4 was upregulated by 11,12-EET and 14,15-EET (1µM). Furthermore, the endothelial receptor of lectin-like oxidized low-density lipoprotein (LOX-1) was downregulated in PAECs treated with EETs. The inflammatory responses evoked by ox-LDL (100µg/mL) were blocked by pharmacological inhibitors of Erk1/2 mitogen-activated protein kinase (MAPK) (U0126), p38 MAPK (SB203580), and NF-κB (PDTC). In addition, we confirmed that 11,12-EET suppresses phosphorylation of p38, degradation of IκBα, and activation of NF-κB (p65), whereas 14,15-EET can significantly suppress the phosphorylation of p38 and Erk1/2. Our results indicate that EETs exert beneficial effects on ox-LDL-induced inflammation primarily through the inhibition of LOX-1 receptor upregulation, MAPK phosphorylation, and NF-κB activation and through the upregulation of CYP2J4 expression. This study helps focus the current understanding of the contribution of EETs to the regulation of the inflammation of pulmonary vascular endothelial cells. Furthermore, the therapeutic potential of targeting the EET pathway in pulmonary vascular disease will be highlighted.