RESUMO
IFN-γ, a proinflammatory cytokine produced primarily by T cells and NK cells, activates macrophages and engages mechanisms to control pathogens. Although there is evidence of IFN-γ production by murine macrophages, IFN-γ production by normal human macrophages and their subsets remains unknown. Herein, we show that human M1 macrophages generated by IFN-γ and IL-12- and IL-18-stimulated monocyte-derived macrophages (M0) produce significant levels of IFN-γ. Further stimulation of IL-12/IL-18-primed macrophages or M1 macrophages with agonists for TLR-2, TLR-3, or TLR-4 significantly enhanced IFN-γ production in contrast to the similarly stimulated M0, M2a, M2b, and M2c macrophages. Similarly, M1 macrophages generated from COVID-19-infected patients' macrophages produced IFN-γ that was enhanced following LPS stimulation. The inhibition of M1 differentiation by Jak inhibitors reversed LPS-induced IFN-γ production, suggesting that differentiation with IFN-γ plays a key role in IFN-γ induction. We subsequently investigated the signaling pathway(s) responsible for TLR-4-induced IFN-γ production in M1 macrophages. Our results show that TLR-4-induced IFN-γ production is regulated by the ribosomal protein S6 kinase (p70S6K) through the activation of PI3K, the mammalian target of rapamycin complex 1/2 (mTORC1/2), and the JNK MAPK pathways. These results suggest that M1-derived IFN-γ may play a key role in inflammation that may be augmented following bacterial/viral infections. Moreover, blocking the mTORC1/2, PI3K, and JNK MAPKs in macrophages may be of potential translational significance in preventing macrophage-mediated inflammatory diseases.
Assuntos
Interferon gama/biossíntese , Macrófagos/efeitos dos fármacos , Poli I-C/farmacologia , COVID-19/imunologia , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/farmacologia , MAP Quinase Quinase Quinases/antagonistas & inibidores , MAP Quinase Quinase Quinases/imunologia , Macrófagos/imunologia , Fosfatidilinositol 3-Quinases/imunologia , Proteínas Quinases S6 Ribossômicas 70-kDa/antagonistas & inibidores , Proteínas Quinases S6 Ribossômicas 70-kDa/imunologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/imunologia , Receptor 4 Toll-Like/agonistasRESUMO
BACKGROUND/AIM: We investigated the effect of Kumaizasa leaf extract (KLE) on innate immunity using the HEK293 and RAW 264.7 cell lines. MATERIALS AND METHODS: KLE, lipopolysaccharides (LPS), or KLE with LPS were added to RAW 264.7 cells. The TNF-α and IL-1ß mRNA expression was then quantified. The expression of MAPKs, NFĸB, TNF-α and IL-1ß proteins was also quantified. In addition, KLE was added to HEK293 cells and the IL-8 concentration was measured. RESULTS: In RAW 264.7 cells, KLE increased the levels of TNF-α and IL-1ß mRNA. By contrast, when KLE and LPS were added to RAW 264.7 cells, the increase in TNF-α and IL-1ß mRNA was ameliorated. Similarly, the expression of JNK and ERK proteins was reduced. The addition of KLE to HEK293 cells induced IL-8 production. CONCLUSION: Based on these results, a KLE-mediated mechanism may regulate immunity by suppressing the expression of JNK and ERK, which are involved in inflammatory signal transduction.
Assuntos
Imunidade Inata/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Sasa , Animais , Citocinas/genética , Citocinas/imunologia , MAP Quinases Reguladas por Sinal Extracelular/imunologia , Células HEK293 , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Camundongos , Folhas de Planta , Células RAW 264.7RESUMO
The induction of major histocompatibility complex (MHC) class II proteins by interferon gamma (IFN-γ) in macrophages play an important role during immune responses. Here we explore the signaling pathways involved in the induction by IFN-γ of the MHC II transactivator (CIIta) required for MHC II transcriptional activation. Cyclophilin A (CypA) is required for IFN-γ-dependent induction of MHC II in macrophages, but not when it is mediated by GM-CSF. The effect of CypA appears to be specific because it does not affect the expression of other molecules or genes triggered by IFN-γ, such as FcγR, NOS2, Lmp2, and Tap1. We found that CypA inhibition blocked the IFN-γ-induced expression of CIIta at the transcriptional level in two phases. In an early phase, during the first 2 h of IFN-γ treatment, STAT1 is phosphorylated at Tyrosine 701 and Serine 727, residues required for the induction of the transcription factor IRF1. In a later phase, STAT1 phosphorylation and JNK activation are required to trigger CIIta expression. CypA is needed for STAT1 phosphorylation in this last phase and to bind the CIIta promoter. Our findings demonstrate that STAT1 is required in a two-step induction of CIIta, once again highlighting the significance of cross talk between signaling pathways in macrophages.
Assuntos
Interferon gama/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Janus Quinases/imunologia , Proteínas Nucleares/imunologia , Fator de Transcrição STAT1/imunologia , Transativadores/imunologia , Animais , Linhagem Celular , Ciclosporina/farmacologia , Lactonas/farmacologia , Camundongos Endogâmicos BALB C , Proteínas Nucleares/genética , Compostos de Espiro/farmacologia , Transativadores/genéticaRESUMO
Hypertension emerged from early reports as a potential risk factor for worse outcomes for persons with coronavirus disease 2019 (COVID-19). Among the putative links between hypertension and COVID-19 is a key counter-regulatory component of the renin-angiotensin system (RAS): angiotensin-converting enzyme 2 (ACE2). ACE2 facilitates entry of severe acute respiratory syndrome coronavirus 2, the virus responsible for COVID-19, into host cells. Because RAS inhibitors have been suggested to increase ACE2 expression, health-care providers and patients have grappled with the decision of whether to discontinue these medications during the COVID-19 pandemic. However, experimental models of analogous viral pneumonias suggest RAS inhibitors may exert protective effects against acute lung injury. We review how RAS and ACE2 biology may affect outcomes in COVID-19 through pulmonary and other systemic effects. In addition, we briefly detail the data for and against continuation of RAS inhibitors in persons with COVID-19 and summarize the current consensus recommendations from select specialty organizations.
Assuntos
Lesão Pulmonar Aguda/metabolismo , Antagonistas de Receptores de Angiotensina/uso terapêutico , Enzima de Conversão de Angiotensina 2/metabolismo , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , COVID-19/metabolismo , Hipertensão/tratamento farmacológico , Lesão Pulmonar Aguda/epidemiologia , Lesão Pulmonar Aguda/imunologia , Angiotensina I/imunologia , Angiotensina I/metabolismo , Angiotensina II/imunologia , Angiotensina II/metabolismo , Enzima de Conversão de Angiotensina 2/imunologia , COVID-19/epidemiologia , COVID-19/imunologia , Comorbidade , Humanos , Hipertensão/epidemiologia , Hipertensão/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Pulmão/imunologia , Pulmão/metabolismo , Sistema de Sinalização das MAP Quinases , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/metabolismo , Fatores de Proteção , Receptores de Coronavírus/imunologia , Receptores de Coronavírus/metabolismo , Sistema Renina-Angiotensina , Fatores de Risco , SARS-CoV-2 , Regulação para CimaRESUMO
Resveratrol was reported to inhibit inflammatory responses; however, the role of this polyphenol in obesity-induced skeletal muscle inflammation remains unknown. Mice fed a high fat diet (HFD) were treated with resveratrol for 16 weeks. Resveratrol treatment decreased macrophage infiltration into skeletal muscle of HFD-fed mice. Resveratrol also led to the polarization of macrophages to the M2 direction, as well as decreasing the expression of a number of M1 pro-inflammatory cytokines [tumor necrosis factor α (TNF-α), interleukin 1 ß (IL-1ß) and interleukin 6 (IL-6)]. In addition, increased infiltration of regulatory T cells (Treg cells) was found following resveratrol treatment in skeletal muscle of mice. Decreased intramyocellular lipid deposition was associated with reduced expression levels of toll-like receptors 2 (TLR2) and TLR4 in resveratrol treated mice. We also found that diminished inflammation in skeletal muscle following resveratrol treatment was accompanied by increasing phosphorylation of 5'-adenosine monophosphate-activated protein kinase (AMPK) and decreasing phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Taken together, these findings suggest that resveratrol ameliorates inflammation in skeletal muscle of HFD-induced model of obesity. Therefore, resveratrol might represent a potential treatment for attenuation of inflammation in skeletal muscle tissue.
Assuntos
Macrófagos/imunologia , Músculo Esquelético/imunologia , Obesidade/tratamento farmacológico , Resveratrol/administração & dosagem , Linfócitos T Reguladores/imunologia , Animais , Polaridade Celular/efeitos dos fármacos , Humanos , Interleucina-6/genética , Interleucina-6/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Músculo Esquelético/efeitos dos fármacos , Obesidade/imunologia , Obesidade/fisiopatologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Liang-Ge-San (LGS) is a traditional Chinese medicine formula that commonly used in acute inflammatory diseases. However, the anti-inflammatory effects and the underlying mechanisms of LGS are not fully studied. AIM OF THE STUDY: This study aims to investigate the anti-inflammatory activity and explore the underlying mechanisms of LGS in zebrafish and cell inflammation models. MATERIALS AND METHODS: LPS-induced zebrafish inflammation model was established by LPS-yolk microinjection. The protective effect of LGS on zebrafish injected with LPS was observed using survival analysis. Infiltration of inflammatory cells was determined by H&E staining assay. Expression levels of key inflammatory cytokines TNF-α and IL-6 were measured by q-PCR assay. Recruitment of neutrophils and macrophages were observed by fluorescence microscopy, SB staining and NR staining. In vitro anti-inflammatory effects of LGS were evaluated on LPS-stimulated RAW 264.7â¯cells. The generation of IL-6 and TNF-α was detected by ELISA. The protein expression levels of JNK, p-JNK (Thr183/Tyr185), Nur77 and p-Nur77 (Ser351) were determined by Western blotting. Finally, two additional inflammatory models in zebrafish, which were induced by CuSO4 or tail fin injury, were also established and the recruitment of neutrophils and macrophages were observed for the determination of the anti-inflammatory activity of LGS. RESULTS: LGS protected zebrafish against LPS-induced death and dose-dependently inhibited LPS-induced acute inflammatory response in zebrafish, as indicated by increased survival rate, reduced infiltration of inflammatory cells, decreased recruitment of macrophages and neutrophils, and downregulated expression levels of TNF-α and IL-6. Additionally, LGS inhibited the secretion of TNF-α and IL-6, increased the expression of Nur77, and reduced the expression of p-Nur77 (Ser351) and p-JNK (Thr183/Tyr185) in LPS-stimulated RAW 264.7â¯cells. The anti-inflammatory action of LGS was also observed in another two zebrafish inflammation models, which was supported by the inhibition on neutrophils and macrophages recruitment. CONCLUSION: The present study demonstrates that LGS possesses anti-inflammatory activity in zebrafish inflammation models and LPS-stimulated RAW 264.7â¯cells, which is related to the inhibition on p-JNK and p-Nur77. This finding provides a pharmacological basis for LGS in the control of inflammatory disorder.
Assuntos
Anti-Inflamatórios/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Inflamação/tratamento farmacológico , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Doença Aguda/terapia , Animais , Anti-Inflamatórios/uso terapêutico , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/uso terapêutico , Feminino , Humanos , Inflamação/imunologia , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Masculino , Camundongos , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/imunologia , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Fosforilação/efeitos dos fármacos , Células RAW 264.7 , Peixe-ZebraRESUMO
OBJECTIVE: Iguratimod, a novel disease-modifying anti-rheumatic drug for the treatment of rheumatoid arthritis, has been approved in China and Japan. Here, we aimed to find whether iguratimod can inhibit the aggressive behavior and promote apoptosis of rheumatoid fibroblast-like synoviocytes (RA-FLSs). METHODS: The proliferation of RA-FLSs was assessed by 5-ethynyl-2'-deoxyuridine test and Cell Counting Kit-8. Migration and invasion were determined by the wound test and a transwell assay. Apoptosis was tested by flow cytometry. The mRNA expression of matrix metalloproteinases (MMPs) and proinflammatory cytokines in RA-FLSs were measured by quantitative PCR and ELISA. To gain insight into the molecular signaling mechanisms, we determined the effect of iguratimod on the activation of mitogen-activated protein kinases (MAPK) signaling pathways by the cellular thermal shift assay (CETSA) and western blot. RESULTS: Iguratimod treatment significantly reduced the proliferation, migration, and invasive capacities of RA-FLSs in a dose-dependent manner in vitro. MMP-1, MMP-3, MMP-9, Interleukin-6 (IL-6), and monocyte chemoattractant protein-1 mRNA and protein levels were all decreased after treatment with iguratimod. Furthermore, tumor necrosis factor-alpha- (TNF-α-) induced expression of phosphorylated c-Jun N-terminal kinases (JNK) and P38 MAPK were inhibited by iguratimod. Additionally, iguratimod promoted the apoptosis of RA-FLSs. Most importantly, iguratimod was shown to directly interact with JNK and P38 protein by CETSA assay. Moreover, activating transcription factor 2 (ATF-2), a substrate of both JNK and P38, was suppressed by iguratimod. CONCLUSIONS: Our findings suggested that the therapeutic effects of iguratimod on RA might be, in part, due to targeting the aggressive behavior and apoptosis of RA-FLSs.
Assuntos
Antirreumáticos/farmacologia , Cromonas/farmacologia , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Imunossupressores/farmacologia , Sulfonamidas/farmacologia , Sinoviócitos/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Artrite Reumatoide/cirurgia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Quimiocina CCL2/genética , Quimiocina CCL2/imunologia , Feminino , Fibroblastos/imunologia , Fibroblastos/patologia , Regulação da Expressão Gênica/imunologia , Humanos , Interleucina-6/genética , Interleucina-6/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/imunologia , Metaloproteinase 3 da Matriz/genética , Metaloproteinase 3 da Matriz/imunologia , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/imunologia , Cultura Primária de Células , Transdução de Sinais , Sinovectomia , Membrana Sinovial/imunologia , Membrana Sinovial/patologia , Sinoviócitos/imunologia , Sinoviócitos/patologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Grass carp septicemia is a systemic inflammatory response that develops following a bacterial infection. The hyperinflammatory state develops could lead to septic shock and lethality. There is increasing evidence that microRNAs are involved in the regulation of the inflammatory response. In the present study, miR-21 was confirmed to be involved in the inflammatory response following infection with Aeromonas hydrophila and LPS stimulation. Both jnk and ccr7 were identified as target gene of miR-21 by overexpression, inhibition, and dual luciferase reporter assays experiments. Meanwhile, miR-21 targets the jnk and ccr7 to modulate downstream pro-inflammatory factors tnf-α, il-1ß, il-6, and il-12. Our results provide a theoretical basis for exploring the molecular mechanism of grass carp miR-21 regulating inflammation.
Assuntos
Carpas/genética , Carpas/imunologia , Doenças dos Peixes/microbiologia , Inflamação/veterinária , Proteínas Quinases JNK Ativadas por Mitógeno/genética , MicroRNAs/genética , Receptores CCR7/genética , Aeromonas hydrophila/fisiologia , Animais , Sequência de Bases , Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Infecções por Bactérias Gram-Negativas/genética , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Inflamação/genética , Inflamação/imunologia , Inflamação/microbiologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos/farmacologia , MicroRNAs/imunologia , Receptores CCR7/imunologiaRESUMO
OBJECTIVE: Tumour necrosis factor-α-induced protein 8-like 2 (TIPE2) has strong anti-inflammatory properties. However, it is unknown whether increased TIPE2 is protective against lipopolysaccharide (LPS)-induced ALI. In the current study, we aimed to investigate whether increased TIPE2 can exert protective effects in a mouse model of ALI induced by LPS. METHODS: We administered TIPE2 adeno-associated virus (AAV-TIPE2) intratracheally into the lungs of mice. Three weeks later, ALI was induced by intratracheal injection of LPS into BALB/c mice. Twenty-four hours later, lung bronchoalveolar lavage fluid (BALF) was acquired to analyse cells and protein, arterial blood was collected for arterial blood gas analysis and the determination of pro-inflammatory factor levels, and lung issues were collected for histologic examination, transmission electron microscopy (TEM), TUNEL staining, wet/dry (W/D) weight ratio analysis, myeloperoxidase (MPO) activity analysis and blot analysis of protein expression. RESULTS: We found that TIPE2 overexpression markedly mitigated LPS-induced lung injury, which was evaluated by the deterioration of histopathology, histologic scores, the W/D weight ratio, and total protein expression in the BALF. Moreover, TIPE2 overexpression markedly attenuated lung inflammation, as evidenced by the downregulation of polymorphonuclear neutrophils (PMNs) in the BALF, lung MPO activity, and pro-inflammatory cytokine levels in the serum. Moreover, TIPE2 overexpression not only dramatically prevented LPS-induced pulmonary cell apoptosis in mice but also blocked LPS-activated JNK phosphorylation and NF-κB p65 nuclear translocation. CONCLUSIONS: Our study shows that the increased expression of AAV-mediated TIPE2 in the lungs of mice inhibits acute inflammation and apoptosis and suppresses the activation of NF-κB and JNK in a murine model of ALI.
Assuntos
Lesão Pulmonar Aguda/terapia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/patologia , Animais , Apoptose , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Citocinas/sangue , Dependovirus/genética , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Inflamação/terapia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Contagem de Leucócitos , Lipopolissacarídeos , Pulmão/patologia , Pulmão/ultraestrutura , Masculino , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão , NF-kappa B/imunologia , Transdução GenéticaRESUMO
Acute lung injury (ALI) and its severe manifestation of acute respiratory distress syndrome (ARDS) in human lung are induced by inflammatory cytokines and endogenous factors such as miRNAs. However, the role of miR-150 in lipopolysaccharide (LPS)-induced ALI is not clear. Here, we found miR-150 expression was significantly reduced in the serum of patients with ARDS, and negatively associated with the disease severity and 28-day survival of ARDS. In vivo, miR-150 decreased total cell and neutrophil counts, and production of inflammatory cytokines interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) as well as levels of total protein, albumin and IgM in the bronchoalveolar lavage (BAL) fluid in LPS-induced ALI mice. Meanwhile, miR-150 improved the 72â¯h survival rate of LPS-induced ALI mice. In-vitro assays demonstrated that miR-150 alleviated LPS-induced A549 cell apoptosis, autophagy, and release of inflammatory cytokines. Further, AKT3 was a direct target of miR-150. Silencing of AKT3 partially reversed LPS-induced A549 cell injury, and enhanced the protective effects of miR-150. In addition, miR-150 or si-AKT3 effectively inhibited the phosphorylation levels of c-Jun N-terminal kinase (JNK) and nuclear factor-κB (NF-κB) (p65 and IκBα). In conclusion, miR-150 alleviated LPS-induced acute lung injury via directly targeting AKT3 expression or regulating JNK and NF-κB pathways, which may be a promising therapeutic strategy to treat ALI/ARDS.
Assuntos
Lesão Pulmonar Aguda , MicroRNAs/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Síndrome do Desconforto Respiratório , Células A549 , Lesão Pulmonar Aguda/sangue , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/imunologia , Adulto , Idoso , Animais , Feminino , Inativação Gênica , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos , Masculino , Camundongos Endogâmicos BALB C , MicroRNAs/sangue , Pessoa de Meia-Idade , NF-kappa B/imunologia , Proteínas Proto-Oncogênicas c-akt/genética , Síndrome do Desconforto Respiratório/sangue , Síndrome do Desconforto Respiratório/genética , Síndrome do Desconforto Respiratório/imunologiaRESUMO
Soybean allergy is a serious health risk to humans and animals; ß-conglycinin is the primary antigenic protein in soybean. Intestinal porcine epithelial (IPEC-J2) cells were used as an in vitro physiological model of the intestinal epithelium to study the effects of different concentrations of soybean antigen protein ß-conglycinin to identify the involved signaling pathways. The cells were divided into eight groups and either untreated or treated with different concentrations of ß-conglycinin, pyrrolidine dithiocarbamate (PDTC), Nω-nitro-l-arginine methyl ester hydrochloride (l-NAME), SP600125, and SB202190 either alone or in combination. The cells were incubated with 1, 5, and 10 mg·mL-1 ß-conglycinin or 5 mg·mL-1 ß-conglycinin and 1 µmol·L-1 nuclear factor κB (NF-κB) inhibitor (PDTC), inducible nitric oxide synthase inhibitor (l-NAME), c-Jun N-terminal kinase (JNK) inhibitor (SP600125), and p38 inhibitor (SB202190) for 24 h, separately; controls were left untreated. The mRNA, protein, and phosphorylation levels of NF-κB, p38, and JNK were higher in the treated groups than in the control group. ß-Conglycinin decreased tight junction distribution, destroyed the cytoskeleton of IPEC-J2 cells, and caused cell death. After the addition of the inhibitors, ß-conglycinin-induced IPEC-J2 cell damage was significantly reduced. ß-Conglycinin caused damage to IPEC-J2 cells via the mitogen-activated protein kinase/NF-κB signaling pathway. The results of this study are crucial for exploring the mechanisms underlying allergic reactions caused by soybean antigen proteins.
Assuntos
Antígenos de Plantas/imunologia , Células Epiteliais/imunologia , Hipersensibilidade Alimentar/imunologia , Globulinas/imunologia , Glycine max/imunologia , Proteínas Quinases Ativadas por Mitógeno/imunologia , NF-kappa B/imunologia , Proteínas de Armazenamento de Sementes/imunologia , Proteínas de Soja/imunologia , Animais , Humanos , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Quinases Ativadas por Mitógeno/genética , NF-kappa B/genética , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/imunologia , Fosforilação , Transdução de Sinais , Suínos , Junções Íntimas/genética , Junções Íntimas/imunologiaRESUMO
Sepsis is a potentially fatal systemic inflammatory response syndrome caused by infection. In this study, we evaluated the effects of MCP-induced protein 1 (MCPIP1), a recently discovered inflammation-related ribonuclease, on sepsis-induced acute lung injury (ALI) and investigated the underlying mechanisms. Cecal ligation puncture and lipopolysaccharide induction were performed on Sprague-Dawley rats and RAW264.7 cells, respectively, to establish sepsis-induced ALI models. The proteasome inhibitor MG132 used as an activator of MCPIP1 overexpression, and we showed that MG132 can indeed increase the expression of MCPIP1. MCPIP1 overexpression induced by MG132 alleviated sepsis-induced pathologic changes, water content and protein leakage in the lungs, and induction of systemic inflammatory mediators, and improved the 7-day mortality rate in the model rats. We also showed that MCPIP1 p showed romoted macrophage polarization from the M1 to the M2 type in sepsis-induced ALI. Furthermore, MCPIP1-enhanced M2 polarization was inhibited by an MCPIP1-targeting small interfering RNA (siMCPIP1) in RAW264.7 cells. Further mechanistic studies showed that the promotive effect of MCPIP1 on M2 polarization was related to the inhibition of c-Jun N-terminal kinase (JNK) and its downstream transcription factor c-Myc in the in vitro model. Conversely, siMCPIP1 transfection resulted in the recovery of JNK and c-Myc expression in LPS-treated cells. Taken together, these findings indicate that MCPIP1 plays a protective role in sepsis-induced ALI by modulating macrophage polarization through inhibition of the JNK/c-Myc signaling pathway. Our study presents a potentially novel therapeutic strategy for the treatment of lung injury involving the inflammatory cascade.
Assuntos
Lesão Pulmonar Aguda/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Proto-Oncogênicas c-myc/imunologia , Ribonucleases/imunologia , Sepse/imunologia , Lesão Pulmonar Aguda/etiologia , Animais , Leupeptinas/farmacologia , Masculino , Camundongos , Células RAW 264.7 , Ratos Sprague-Dawley , Sepse/complicações , Transdução de SinaisRESUMO
Diabetic cardiomyopathy (DCM) is a leading contributor to the increased morbidity and mortality rates associated with diabetes. Persistent inflammation has previously been reported to be involved in the pathogenesis of DCM. However, the exact underlying molecular mechanisms remain to be fully elucidated. In the present study, the role of spleen tyrosine kinase (Syk) and cJun Nterminal kinase (JNK) in NLR family pyrin domaincontaining 3 (NLRP3 inflammasome) activation in DCM were investigated in vivo and in vitro. Streptozotocin (65 mg/kg) was injected intraperitoneally into SpragueDawley rats to induce a rat model of diabetes. Neonatal rat cardiomyocytes and H9c2 cells were cultured to detect the expression of JNK, NLRP3 and its associated downstream molecules, following treatment with Syk/JNK inhibitor or Syk/JNKsmall interfering (si)RNA in high glucose (HG) conditions. It was revealed that the protein and mRNA expression levels of phospho (p)Syk, pJNK, NLRP3 and its associated downstream molecules, including interleukin (IL)1ß, were upregulated in vivo and in vitro. The JNK inhibitor significantly decreased the expression of NLRP3 and its downstream molecules in neonatal rat cardiomyocytes and H9c2 cells treated with HG. Furthermore, SyksiRNA and the Syk inhibitor markedly inhibited the HGinduced activation of JNK, followed by the downregulation of NLRP3 and its downstream molecules at the mRNA and protein levels in cells. Therefore, it was demonstrated that the HGinduced activation of NLRP3 was mediated by the activation of Syk/JNK, which subsequently increased the protein expression levels of mature IL1ß, suggesting that the Syk/JNK/NLRP3 signaling pathway serves a critical role in the pathogenesis of DCM.
Assuntos
Diabetes Mellitus Experimental/imunologia , Cardiomiopatias Diabéticas/imunologia , Inflamassomos/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Quinase Syk/imunologia , Animais , Linhagem Celular , Células Cultivadas , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/patologia , Cardiomiopatias Diabéticas/etiologia , Cardiomiopatias Diabéticas/patologia , Inflamação/complicações , Inflamação/imunologia , Inflamação/patologia , Masculino , Ratos Sprague-DawleyRESUMO
BACKGROUND: Clinically, liver fibrosis and cholestasis are two major disease entities, ultimately leading to hepatic failure. Although autophagy plays a substantial role in the pathogenesis of these diseases, its precise mechanism has not been determined yet. MATERIALS AND METHODS: Mouse models of liver fibrosis or cholestasis were obtained after the serial administration of thioacetamide (TAA) or surgical bile duct ligation (BDL), respectively. Then, after obtaining liver specimens at specific time points, we compared the expression of makers related to apoptosis (cleaved caspases), inflammation (CD68), necrosis (high-mobility group box 1), phospho-c-Jun N-terminal kinase (p-JNK), and autophagy (microtubule-associated protein light chain 3B and p62) in the fibrotic or cholestatic mouse livers, by polymerase chain reaction, Western blot analysis, immunohistochemistry, and immunofluorescence. RESULTS: Although cholestatic livers exhibited the tendency of progressively increasing the expression of most apoptosis-related markers (cleaved caspases), it was not prominent when it was compared with the tendency found in the livers of TAA-treated mice. Contrastingly, the necrosis-related factor (high-mobility group box 1) was significantly increased in the livers of BDL mice over time, reaching their peak values on day 7 after BDL. In addition, the inflammation-related factor (CD68) was highly expressed in BDL mice compared with TAA-treated mice over time. Autophagy marker studies indicated that autophagy was upregulated in fibrotic livers, whereas it was downregulated in cholestatic livers. We also observed mild to moderate activation of p-JNK in the livers of TAA-treated mice, whereas significantly higher p-JNK activation was detected in the livers of BDL mice. CONCLUSIONS: Unlike TAA-treated mice, BDL mice exhibited higher expression of the markers related with inflammation and necrosis, especially including p-JNK, while maintaining low levels of autophagic process. Therefore, obstructive cholestasis is characterized by higher p-JNK activation, which could be related with marked necrotic cell death resulting from extensive inflammation and little chance of compensatory autophagy.
Assuntos
Autofagia , Colestase/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Cirrose Hepática Experimental/imunologia , Fígado/patologia , Animais , Ductos Biliares/cirurgia , Colestase/etiologia , Colestase/patologia , Hepatócitos/imunologia , Hepatócitos/patologia , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Ligadura , Fígado/citologia , Fígado/imunologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/patologia , Masculino , Camundongos , Necrose/imunologia , Necrose/patologia , Fosforilação/imunologia , Tioacetamida/toxicidadeRESUMO
Acute lung injury (ALI) is served as a severe life-threatening disease. However, the pathogenesis that contributes to ALI has not been fully understood. Tumor necrosis factor receptor-associated factor 1 (TRAF1) interacts with multiple regulators, performing its diverse role in biological functions. However, the effects of TRAF1 on ALI remain unknown. In this study, we attempted to explore the role of TRAF1 in ALI progression. The findings suggested that TRAF1-knockout (KO) markedly attenuated LPS-induced severe mortality rate in murine animals. LPS-elicited histological alterations in pulmonary tissues were significantly alleviated by TRAF1-deletion. Additionally, TRAF1 knockout effectively attenuated lung injury, as evidenced by the reduced lung wet/dry (W/D) weight ratio, as well as decreased bronchoalveolar lavage fluid (BALF) protein levels and neutrophil infiltration. Meanwhile, TRAF1 deletion markedly lessened inflammation, oxidative stress and apoptosis in BALF and/or lung tissues. The levels of pro-inflammatory cytokines stimulated by LPS were down-regulated by TRAF1 ablation, along with the inactivation of nuclear factor κB (NF-κB). LPS-promoted reactive oxygen species (ROS) generation was decreased in TRAF1-KO mice, partly through the improvement of anti-oxidants. Apoptosis was also inhibited by TRAF1 deletion in lung tissues of LPS-challenged mice through the suppression of cleaved Caspase-3. Moreover, TRAF1 knockout significantly decreased c-Jun N-terminal kinase (JNK) activation and its down-streaming signal of c-Jun in pulmonary samples of LPS-induced mice. Importantly, the in vitro study suggested that promoting JNK activation markedly abrogated TRAF1 knockdown-attenuated inflammation, ROS production and apoptosis in LPS-exposed A549â¯cells. Therefore, our experimental results provided evidence that TRAF1 suppression effectively protected LPS-induced ALI against inflammation, oxidative stress and apoptosis through the suppression of JNK activity.
Assuntos
Lesão Pulmonar Aguda/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos/imunologia , Fator 1 Associado a Receptor de TNF/imunologia , Lesão Pulmonar Aguda/patologia , Animais , Linhagem Celular , Ativação Enzimática , Humanos , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos Endogâmicos C57BLRESUMO
During bacterial infection, granulocyte colony-stimulating factor (G-CSF) is produced and accelerates neutrophil production from their progenitors. This process, termed granulopoiesis, strengthens host defense, but Clostridium perfringens α-toxin impairs granulopoiesis via an unknown mechanism. Here, we tested whether G-CSF accounts for the α-toxin-mediated impairment of granulopoiesis. We find that α-toxin dramatically accelerates G-CSF production from endothelial cells in response to Toll-like receptor 2 (TLR2) agonists through activation of the c-Jun N-terminal kinase (JNK) signaling pathway. Meanwhile, α-toxin inhibits G-CSF-mediated cell proliferation of Ly-6G+ neutrophils by inducing degradation of G-CSF receptor (G-CSFR). During sepsis, administration of α-toxin promotes lethality and tissue injury accompanied by accelerated production of inflammatory cytokines in a TLR4-dependent manner. Together, our results illustrate that α-toxin disturbs G-CSF-mediated granulopoiesis by reducing the expression of G-CSFR on neutrophils while augmenting septic shock due to excess inflammatory cytokine release, which provides a new mechanism to explain how pathogenic bacteria modulate the host immune system.
Assuntos
Toxinas Bacterianas/toxicidade , Proteínas de Ligação ao Cálcio/toxicidade , Clostridium perfringens/patogenicidade , Gangrena Gasosa/genética , Fator Estimulador de Colônias de Granulócitos/genética , Lipopolissacarídeos/toxicidade , Receptores de Fator Estimulador de Colônias de Granulócitos/genética , Choque Séptico/genética , Fosfolipases Tipo C/toxicidade , Animais , Clostridium perfringens/genética , Clostridium perfringens/imunologia , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Feminino , Gangrena Gasosa/imunologia , Gangrena Gasosa/microbiologia , Gangrena Gasosa/mortalidade , Regulação da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos/imunologia , Hematopoese/efeitos dos fármacos , Hematopoese/genética , Hematopoese/imunologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/microbiologia , Receptores de Fator Estimulador de Colônias de Granulócitos/imunologia , Choque Séptico/imunologia , Choque Séptico/microbiologia , Choque Séptico/mortalidade , Transdução de Sinais , Análise de Sobrevida , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/imunologia , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologiaRESUMO
OBJECTIVE: The aim of this research is to explore the possible role of miR-152 in spinal cord injury and its underlying mechanism. MATERIALS AND METHODS: After a mouse model of spinal cord injury (SCI) was developed, Real Time-quantitative Polymerase Chain Reaction (RT-qPCR) was used to detect the expression of miR-152 and c-jun in the mouse. In addition, the expression levels of interleukin-1b (IL-1b), interleukin-18 (IL-18) and tumor necrosis factor-α (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). Subsequently, miR-152 was overexpressed and the levels of inflammation and c-jun after spinal cord injury were detected by Western blot. Furthermore, the grip strength of double forelimb, left forelimb or right forelimb of the mice was detected using a grip force test after miR-152 was overexpressed in the injured area of each group. RESULTS: By constructing a mouse model of spinal cord injury, we found that the expression of miR-152 in the injured area decreased with time; meanwhile, the inflammatory relative genes including IL-1b, IL18, TNF-α, and c-jun were significantly increased. However, miR-152 overexpression significantly reduced the levels of inflammation genes as well as the expression of c-jun. Besides, the strength of the forelimbs in the spinal cord injury mice was restored. CONCLUSIONS: MiR-152 could inhibit inflammatory responses and promote the recovery of the spinal cord injury through the c-jun N-terminal kinase pathway and it can be a target molecular for treating spinal cord injury.
Assuntos
Inflamação/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Transdução de Sinais/genética , Traumatismos da Medula Espinal/imunologia , Medula Espinal/fisiopatologia , Animais , Citocinas/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Membro Anterior/fisiopatologia , Humanos , Inflamação/imunologia , Inflamação/patologia , Inflamação/fisiopatologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Camundongos , Força Muscular/fisiologia , Recuperação de Função Fisiológica/genética , Recuperação de Função Fisiológica/imunologia , Transdução de Sinais/imunologia , Medula Espinal/patologia , Traumatismos da Medula Espinal/genética , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/fisiopatologiaRESUMO
Wnt5a signalling plays pathological roles in synovial inflammation and bone destruction. In the present study, we designed four human Wnt5a-based DNA recombinants and detected their effects on immunogenicity and anti-rheumatism in a collagen-induced arthritis (CIA) model. Histomorphometry and micro-CT scanning showed that the phWnt5a-NL was superior to other recombinants because it resulted in decreased severity of arthritis, histopathological scores of synovial inflammation and bone erosion in CIA mice. In addition, ELISA and TRAP staining showed that the phWnt5a-NL-immunized CIA mice had reductions in the serum concentrations of the rheumatoid-associated cytokines IL-1ß and RANKL and in osteoclastogenesis. Furthermore, flow cytometry showed that the phWnt5a-NL treatment increased the percentage of Treg cells. Finally, western blotting analysis showed that the phWnt5a-NL-immunization interrupted ß-catenin and JNK expression in osteoclast precursors derived from the CIA mice. The results suggest that depleting the carboxy-terminus in hWnt5a-based DNA recombinants may be beneficial for the treatment of chronic inflammatory disorders involving bone resorption.
Assuntos
Artrite Experimental/imunologia , Imunização/métodos , Proteínas Recombinantes/imunologia , Proteína Wnt-5a/imunologia , Animais , Artrite Experimental/diagnóstico por imagem , Artrite Experimental/patologia , Citocinas/sangue , Citocinas/imunologia , Humanos , Interleucina-1beta/sangue , Interleucina-1beta/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos DBA , Osteoclastos/citologia , Osteoclastos/imunologia , Osteoclastos/metabolismo , Osteogênese/imunologia , Proteínas Recombinantes/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Proteína Wnt-5a/genética , Proteína Wnt-5a/metabolismo , Microtomografia por Raio-X/métodosRESUMO
BACKGROUND: Mesenchymal stem cells (MSCs) play an anti-inflammatory role by secreting certain bioactive molecules to exert their therapeutic effects for disease treatment. However, the underlying mechanism of MSCs in chronic autoimmune liver diseases-primary biliary cholangitis (PBC), for example-remains to be elucidated. METHODS: Human umbilical cord-derived MSCs (UC-MSCs) were injected intravenously into 2-octynoic acid coupled to bovine serum albumin (2OA-BSA)-induced autoimmune cholangitis mice. Serum levels of biomarkers and autoantibodies, histologic changes in the liver, diverse CD4+ T-cell subsets in different tissues, and chemokine activities were analyzed. Moreover, we investigated galectin-9 (Gal-9) expression and its function in UC-MSCs. RESULTS: In this study, UC-MSC transplantation (UC-MSCT) significantly ameliorated liver inflammation, primarily by diminishing T helper 1 (Th1) and Th17 responses as well as modifying liver chemokine activities in experimental autoimmune cholangitis mice. Mechanistically, UC-MSCs significantly repressed the proliferation of CD4+ T cells and suppressed the differentiation of Th1 and Th17 cells, which was likely dependent on Gal-9. Furthermore, the signal transducer and activator of transcription (STAT) and c-Jun N-terminal kinase (JNK) signaling pathways were involved in the production of Gal-9 in UC-MSCs. CONCLUSIONS: These results suggest that Gal-9 contributes significantly to UC-MSC-mediated therapeutic effects and improve our understanding of the immunomodulatory mechanisms of MSCs in the treatment of PBC.
Assuntos
Doenças Autoimunes/terapia , Galectinas/genética , Imunomodulação , Cirrose Hepática Biliar/terapia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/imunologia , Animais , Doenças Autoimunes/induzido quimicamente , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Ductos Biliares/imunologia , Ductos Biliares/patologia , Técnicas de Cocultura , Células Epiteliais/imunologia , Células Epiteliais/patologia , Ácidos Graxos Monoinsaturados/química , Feminino , Galectinas/imunologia , Regulação da Expressão Gênica , Humanos , Imunoconjugados/administração & dosagem , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Cirrose Hepática Biliar/induzido quimicamente , Cirrose Hepática Biliar/imunologia , Cirrose Hepática Biliar/patologia , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/imunologia , Soroalbumina Bovina/administração & dosagem , Soroalbumina Bovina/química , Transdução de Sinais , Células Th1/imunologia , Células Th1/patologia , Células Th17/imunologia , Células Th17/patologia , Transplante Heterólogo , Cordão Umbilical/citologia , Cordão Umbilical/imunologiaRESUMO
Activation of p38 mitogen-activated protein kinase (MAPK) and c-Jun amino terminal kinase (JNK) is prominent in human crescentic glomerulonephritis. p38 and JNK inhibitors suppress crescentic disease in animal models; however, the upstream mechanisms inducing activation of these kinases in crescentic glomerulonephritis are unknown. We investigated the hypothesis that apoptosis signal-regulating kinase 1 (ASK1/MAP3K5) promote p38/JNK activation and renal injury in models of nephrotoxic serum nephritis (NTN); acute glomerular injury in SD rats, and crescentic disease in WKY rats. Treatment with the selective ASK1 inhibitor, GS-444217 or vehicle began 1 hour before nephrotoxic serum injection and continued until animals were killed on day 1 (SD rats) or 14 (WKY rats). NTN resulted in phosphorylation (activation) of p38 and c-Jun in both models which was substantially reduced by ASK1 inhibitor treatment. In SD rats, GS-444217 prevented proteinuria and glomerular thrombosis with suppression of macrophage activation on day 1 NTN. In WKY rats, GS-444217 reduced crescent formation, prevented renal impairment and reduced proteinuria on day 14 NTN. Macrophage activation, T-cell infiltration and renal fibrosis were also reduced by GS-444217. In conclusion, GS-444217 treatment inhibited p38/JNK activation and development of renal injury in rat NTN. ASK1 inhibitors may have therapeutic potential in rapidly progressive glomerulonephritis.