RESUMO
Non-alcoholic fatty liver disease (NAFLD) can progress to the more serious non-alcoholic steatohepatitis (NASH), characterized by inflammatory injury and fibrosis. The pathogenic basis of NAFLD progressing to NASH is currently unknown, but growing evidence suggests MD2 (myeloid differentiation factor 2), an accessory protein of TLR4, is an important signalling component contributing to this disease. We evaluated the effectiveness of the specific MD2 inhibitor, L6H21, in reducing inflammatory liver injury in a relevant high-fat diet (HFD) mouse model of NASH and in the palmitic acid (PA)-stimulated human liver cell line (HepG2). For study, genetic knockout (MD2-/- ) mice were fed a HFD or control diet for 24 weeks, or wild-type mice placed on a similar diet regimen and treated with L6H21 for the last 8 or 16 weeks. Results indicated that MD2 inhibition with L6H21 was as effective as MD2 knockout in preventing the HFD-induced hepatic lipid accumulation, pro-fibrotic changes and expression of pro-inflammatory molecules. Direct challenge of HepG2 with PA (200 µM) increased MD2-TLR4 complex formation and expression of pro-inflammatory and pro-fibrotic genes and L6H21 pre-treatment prevented these PA-induced responses. Interestingly, MD2 knockout or L6H21 increased expression of the anti-inflammatory molecule, PPARγ, in liver tissue and the liver cell line. Our results provide further evidence for the critical role of MD2 in the development of NASH and conclude that MD2 could be a potential therapeutic target for NAFLD/NASH treatment. Moreover, the small molecule MD2 inhibitor, L6H21, was an effective and selective investigative agent for future mechanistic studies of MD2.
Assuntos
Progressão da Doença , Inflamação/patologia , Antígeno 96 de Linfócito/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Chalconas/farmacologia , Dieta Hiperlipídica , Regulação da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/genética , PPAR gama/metabolismo , Ácido PalmíticoRESUMO
BACKGROUND: Cigarette smoke is well known to worsen asthma symptoms in asthmatic patients and to make them refractory to treatment, but the underling molecular mechanism is unclear. We hypothesized that cigarette smoke can reduce the expression of HDAC2 in asthma and the process was achieved by activating the PI3K-δ/Akt signaling pathway. We further hypothesized that roxithromycin (RXM) can alleviate the impacts by cigarette smoke. METHODS: A murine model of asthma induced by ovalbumin (OVA) and cigarette smoke has been established. The infiltration of inflammatory cells and inflammatory factors was examined in this model. Finally, we evaluated the expression of HDAC2, Akt phosphorylation levels, and the effects of RXM treatment on the model described earlier. RESULTS: Cigarette smoke exposure reduced HDAC2 protein expression by enhancing the phosphorylation of Akt in PI3K-δ/Akt signaling pathway. Furthermore, RMX reduced the airway inflammation and improved the level of expression of HDAC2 in the cigarette smoke-exposed asthma mice. CONCLUSIONS: This study provides a novel insight into the mechanism of cigarette smoke exposure in asthma and the effects of RXM treatment on this condition. These results may be helpful for treating refractory asthma and emphasizing the need for a smoke-free environment for asthmatic patients.
Assuntos
Antiasmáticos/uso terapêutico , Antibacterianos/uso terapêutico , Asma/tratamento farmacológico , Histona Desacetilase 2/metabolismo , Nicotiana , Roxitromicina/uso terapêutico , Fumaça/efeitos adversos , Alérgenos , Animais , Antiasmáticos/farmacologia , Antibacterianos/farmacologia , Asma/genética , Asma/metabolismo , Asma/patologia , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Modelos Animais de Doenças , Eosinófilos/imunologia , Feminino , Histona Desacetilase 2/genética , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Camundongos Endogâmicos BALB C , Neutrófilos/imunologia , Ovalbumina , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Roxitromicina/farmacologiaRESUMO
Acute lung injury (ALI) is the leading cause of mortality in the intensive care unit. Currently, there is no effective pharmacological treatment for ALI. In our previous study, we reported that Lg25 and Lg26, two indole-2-carboxamide derivatives, inhibited the lipopolysaccharide (LPS)-induced inflammatory cytokines in vitro and attenuated LPS-induced sepsis in vivo. In the present study, we confirmed data from previous studies that LPS significantly induced pulmonary edema and pathological changes in lung tissue, increased protein concentration and number of inflammatory cells in bronchoalveolar lavage fluids (BALF), and increased inflammatory cytokine TNF-α expression in serum and BALF, pro-inflammatory genes expression, and macrophages infiltration in lung tissue. However, pretreatment with Lg25 and Lg26 significantly attenuated the LPS-induced changes in mice. Taken together, these data indicate that the newly discovered indole-2-carboxamide derivatives could be particularly useful in the treatment of inflammatory diseases such as ALI.
Assuntos
Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Amidas/farmacologia , Indóis/farmacologia , Inflamação/tratamento farmacológico , Lipopolissacarídeos/farmacologia , Lesão Pulmonar Aguda/metabolismo , Animais , Líquido da Lavagem Broncoalveolar , Citocinas/metabolismo , Expressão Gênica/efeitos dos fármacos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Edema Pulmonar/induzido quimicamente , Edema Pulmonar/tratamento farmacológico , Edema Pulmonar/metabolismoRESUMO
BACKGROUND: Acute lung injury (ALI) and its most severe form acute respiratory distress syndrome (ARDS) have been the leading cause of morbidity and mortality in intensive care units (ICU). Currently, there is no effective pharmacological treatment for acute lung injury. Curcumin, extracted from turmeric, exhibits broad anti-inflammatory properties through down-regulating inflammatory cytokines. However, the instability of curcumin limits its clinical application. METHODS: A series of new curcumin analogs were synthesized and screened for their inhibitory effects on the production of TNF-α and IL-6 in mouse peritoneal macrophages by ELISA. The evaluation of stability and mechanism of active compounds was determined using UV-assay and Western Blot, respectively. In vivo, SD rats were pretreatment with c26 for seven days and then intratracheally injected with LPS to induce ALI. Pulmonary edema, protein concentration in BALF, injury of lung tissue, inflammatory cytokines in serum and BALF, inflammatory cell infiltration, inflammatory cytokines mRNA expression, and MAPKs phosphorylation were analyzed. We also measured the inflammatory gene expression in human pulmonary epithelial cells. RESULTS: In the study, we synthesized 30 curcumin analogs. The bioscreeening assay showed that most compounds inhibited LPS-induced production of TNF-α and IL-6. The active compounds, a17, a18, c9 and c26, exhibited their anti-inflammatory activity in a dose-dependent manner and exhibited greater stability than curcumin in vitro. Furthermore, the active compound c26 dose-dependently inhibited ERK phosphorylation. In vivo, LPS significantly increased protein concentration and number of inflammatory cells in BALF, pulmonary edema, pathological changes of lung tissue, inflammatory cytokines in serum and BALF, macrophage infiltration, inflammatory gene expression, and MAPKs phosphorylation . However, pretreatment with c26 attenuated the LPS induced increase through ERK pathway in vivo. Meanwhile, compound c26 reduced the LPS-induced inflammatory gene expression in human pulmonary epithelial cells. CONCLUSIONS: These results suggest that the novel curcumin analog c26 has remarkable protective effects on LPS-induced ALI in rat. These effects may be related to its ability to suppress production of inflammatory cytokines through ERK pathway. Compound c26, with improved chemical stability and bioactivity, may have the potential to be further developed into an anti-inflammatory candidate for the prevention and treatment of ALI.
Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Anti-Inflamatórios/farmacologia , Curcumina/farmacologia , Pulmão/efeitos dos fármacos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Células Cultivadas , Curcumina/análogos & derivados , Modelos Animais de Doenças , Ativação Enzimática , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-6/metabolismo , Lipopolissacarídeos , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos Endogâmicos ICR , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Edema Pulmonar/induzido quimicamente , Edema Pulmonar/metabolismo , Edema Pulmonar/patologia , Edema Pulmonar/prevenção & controle , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Acute lung injury (ALI) is a major cause of acute respiratory failure in critically-ill patients. It has been reported that both resveratrol and chalcone derivatives could ameliorate lung injury induced by inflammation. A series of paralleled Aza resveratrol-chalcone compounds (5a-5m, 6a-6i) were designed, synthesized and screened for anti-inflammatory activity. A majority showed potent inhibition on the IL-6 and TNF-α expression-stimulated by LPS in macrophages, of which compound 6b is the most potent analog by inhibition of LPS-induced IL-6 release in a dose-dependent manner. Moreover, 6b exhibited protection against LPS-induced acute lung injury in vivo. These results offer further insight into the use of Aza resveratrol-chalcone compounds for the treatment of inflammatory diseases, and the use of compound 6b as a lead compound for the development of anti-ALI agents.
Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Anti-Inflamatórios/uso terapêutico , Compostos Aza/uso terapêutico , Chalconas/uso terapêutico , Pulmão/efeitos dos fármacos , Estilbenos/uso terapêutico , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/patologia , Animais , Anti-Inflamatórios/química , Compostos Aza/química , Linhagem Celular , Chalconas/química , Humanos , Interleucina-6/antagonistas & inibidores , Interleucina-6/imunologia , Lipopolissacarídeos/imunologia , Pulmão/imunologia , Pulmão/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Resveratrol , Estilbenos/química , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Inflammation is an important pathological process of many acute and chronic diseases, such as sepsis, arthritis, and cancer. Many factors can lead to an inflammatory state of the body, among which bacterial infection plays an important role. Bacterial infection often leads to sepsis, acute lung injury (ALI), or its more serious form of acute respiratory distress syndrome, which are the main fatal diseases in intensive care units. Costunolide has been reported to possess excellent anti-inflammatory activity; however, whether it can affect inflammation induced by gram-negative bacterial is still unclear. Lipopolysaccharide (LPS) stimulated mouse peritoneal macrophages (MPMs) to release proinflammatory cytokines was used as the cell model. The mouse model of sepsis and ALI was built through injecting intravenously and intratracheally of LPS. In the present study, costunolide inhibited LPS-induced inflammatory response through IKK/NF-κB signaling pathway in macrophages. In vivo, costunolide attenuated LPS-induced septic death in mice. Meanwhile, costunolide treatment alleviated LPS-induced lung injury and inflammation via inhibiting the infiltration of inflammatory cells and the expression of inflammatory cytokines. Taken together, these results demonstrated that costunolide could attenuate gram-negative bacterial induced inflammation and diseases and might be a potential candidate for the treatment of inflammatory diseases.
Assuntos
Lesão Pulmonar Aguda , Infecções Bacterianas , Sepse , Sesquiterpenos , Animais , Camundongos , NF-kappa B/metabolismo , Lipopolissacarídeos/toxicidade , Transdução de Sinais , Inflamação/patologia , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Citocinas/metabolismo , Sepse/induzido quimicamente , Sepse/tratamento farmacológico , Sepse/patologia , Infecções Bacterianas/patologia , Pulmão/patologiaRESUMO
Asthma represents a significant global challenge that affects individuals across all age groups and imposes substantial social and economic burden. Due to heterogeneity of the disease, not all patients obtain benefit with current treatments. The objective of this study was to explore the impact of MD2 on the progression of asthma using L6H21, a novel MD2 inhibitor, to identify potential targets and drug candidates for asthma treatment. To establish an asthma-related murine model and evaluate the effects of L6H21, ovalbumin (OVA) was used to sensitize and challenge mice. Pathological changes were examined with various staining techniques, such as H&E staining, glycogen staining, and Masson staining. Inflammatory cell infiltration and excessive cytokine secretion were evaluated by analyzing BALF cell count, RT-PCR, and ELISA. The TLR4/MD2 complex formation, as well as the activation of the MAPK and NF-кB pathways, was examined using western blot and co-IP. Treatment with L6H21 demonstrated alleviation of increased airway resistance, lung tissue injury, inflammatory cell infiltration and excessive cytokine secretion triggered by OVA. In addition, it also ameliorated mucus production and collagen deposition. In the L6H21 treatment group, inhibition of MAPK and NF-кB activation was observed, along with the disruption of TLR4/MD2 complex formation, in contrast to the model group. Thus, L6H21 effectively reduced the formation of the MD2 and TLR4 complex induced by OVA in a dose-dependent manner. This reduction resulted in the attenuation of MAPKs/NF-κB activation, enhanced suppression of inflammatory factor secretion, reduced excessive recruitment of inflammatory cells, and ultimately mitigated airway damage. MD2 emerges as a crucial target for asthma treatment, and L6H21, as an MD2 inhibitor, shows promise as a potential drug candidate for the treatment of asthma.
Assuntos
Asma , Chalcona , Chalconas , Humanos , Camundongos , Animais , Chalcona/uso terapêutico , Ovalbumina/uso terapêutico , NF-kappa B/genética , NF-kappa B/metabolismo , Chalconas/farmacologia , Chalconas/uso terapêutico , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Receptor 4 Toll-Like/uso terapêutico , Asma/induzido quimicamente , Asma/tratamento farmacológico , Asma/patologia , Pulmão/patologia , Citocinas/metabolismo , Modelos Animais de Doenças , Camundongos Endogâmicos BALB CRESUMO
Asthma is a type of chronic lung inflammation with restrictions in effective therapy. NF-κB pathway activation has been suggested to play an important role in the pathogenesis of asthma. Baicalein, one of the major active flavonoids found in Scutellaria baicalensis, exhibits potent anti-inflammatory properties by inhibiting NF-κB activity. Herein, we report that Baicalein significantly reduces OVA-induced airway hyperresponsiveness (AHR), airway inflammation, serum IgE levels, mucus production, and collagen deposition around the airway. Additionally, western blot analysis and immunofluorescence assay showed that Baicalein attenuates the activation of NF-κB, which was mainly reflected by IκBα phosphorylation and degradation, p65 nuclear translocation and downstream iNOS expression. Furthermore, in human epithelial cells, Baicalein blocked TNF-α-induced NF-κB activation. Our study provides evidence that Baicalein administration alleviates the pathological changes in asthma through inactivating the NF-κB/iNOS pathway. Baicalein might be a promising potential therapy agent for patients with allergic asthma in the future.
Assuntos
Antioxidantes/uso terapêutico , Asma/prevenção & controle , Flavanonas/uso terapêutico , NF-kappa B/metabolismo , Animais , Antioxidantes/farmacologia , Asma/imunologia , Linhagem Celular , Colágeno/metabolismo , Avaliação Pré-Clínica de Medicamentos , Flavanonas/farmacologia , Imunoglobulina E/sangue , Camundongos , Muco/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/metabolismo , Ovalbumina , Fitoterapia , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Scutellaria baicalensis , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND AND PURPOSE: Acute lung injury (ALI) is a challenging clinical syndrome, which manifests as an acute inflammatory response. Myeloid differentiation protein 2 (MD2) has an important role in mediating LPS-induced inflammation. Currently, there are no effective molecular-based therapies for ALI or viable biomarkers for predicting the severity of disease. Recent preclinical studies have shown that shikonin, a natural naphthoquinone, prevents LPS-induced inflammation. However, little is known about the underlying mechanisms. EXPERIMENTAL APPROACH: The binding affinity of shikonin to MD2 was analysed using computer docking, surface plasmon resonance analysis and elisa. In vitro, the anti-inflammatory effect and mechanism of shikonin were investigated through elisa, real-time quantitative reverse transcription PCR, Western blotting and immunoprecipitation assay. In vivo, lung injury was induced by intratracheal administration of LPS and assessed by changes in the histopathological and inflammatory markers. The underlying mechanisms were investigated by immunoprecipitation in lung tissue. KEY RESULTS: Shikonin directly bound to MD2 and interfered with the activation of toll-like receptor 4 (TLR4) induced by LPS. In cultured macrophages, shikonin inhibited TLR4 signalling and pro-inflammatory cytokine production. These effects were produced through suppression of key signalling proteins including the NF-κB and the MAPK pathway. We also showed that shikonin inhibits MD2-TLR4 complex formation and reduces LPS-induced inflammatory responses in a mouse model of ALI. CONCLUSIONS AND IMPLICATIONS: Our studies have uncovered the mechanism underlying the biological activity of shikonin in ALI and suggest that the targeting of MD2 may prove to be beneficial as a treatment option for this condition.
Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Inflamação/prevenção & controle , Antígeno 96 de Linfócito/metabolismo , Naftoquinonas/farmacologia , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Animais , Citocinas/metabolismo , Humanos , Inflamação/induzido quimicamente , Lipopolissacarídeos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Simulação de Acoplamento Molecular , Receptores Toll-Like/antagonistas & inibidores , Receptores Toll-Like/metabolismoRESUMO
Sepsis remains a leading cause of death worldwide. Despite years of extensive research, effective drugs to treat sepsis in the clinic are lacking. In this study, we found a novel imidazopyridine derivative, X22, which has powerful anti-inflammatory activity. X22 dose-dependently inhibited lipopolysaccharide (LPS)-induced proinflammatory cytokine production in mouse primary peritoneal macrophages and RAW 264.7 macrophages. X22 also downregulated the LPS-induced proinflammatory gene expression in vitro. In vivo, X22 exhibited a significant protection against LPS-induced death. Pretreatment or treatment with X22 attenuated the sepsis-induced lung and liver injury by inhibiting the inflammatory response. In addition, X22 showed protection against LPS-induced acute lung injury. We additionally found that pretreatment with X22 reduced the inflammatory pain in the acetic acid and formalin models and reduced the dimethylbenzene-induced ear swelling and acetic acid-increased vascular permeability. Together, these data confirmed that X22 has multiple anti-inflammatory effects and may be a potential therapeutic option in the treatment of inflammatory diseases.