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1.
Adv Healthc Mater ; 12(30): e2301673, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37505448

RESUMO

Chronic obstructive pulmonary disease (COPD) is a slow-progressing inflammatory lung disease that is associated with high mortality and disability. There is a lack of appropriate preclinical models of COPD, which hampers drug discovery efforts. Herein, a comparative inflammation-on-a-chip (IoC) is developed with a complete 3D interface without the formation of any micropillar and phaseguide structures that replicated chemoattractant-induced neutrophil transendothelial migration (NTEM), a key feature of COPD. The IoC model is used to evaluate the pharmacological effects of CXCR2 inhibitors (MK-7123, AZD5069, and SB225002) on the migration of neutrophil-like cells in the presence of plasma samples from patients with COPD. This is the first study to evaluate inhibitors of CXCR2-dependent NTEM in a comparative IoC model that mimics the physiological 3D microenvironment, consisting of an endothelial barrier, extracellular compartment, and inflammatory conditions. This IoC model will be useful to investigate COPD severity using patient samples, and will aid basic and translational research involving NTEM.


Assuntos
Neutrófilos , Doença Pulmonar Obstrutiva Crônica , Humanos , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Inflamação/tratamento farmacológico , Movimento Celular , Dispositivos Lab-On-A-Chip
2.
Blood ; 140(8): 889-899, 2022 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-35679477

RESUMO

Lung-resident neutrophils need to be tightly regulated to avoid degranulation- and cytokine-associated damage to fragile alveolar structures that can lead to fatal outcomes. Here we show that lung neutrophils (LNs) express distinct surface proteins and genes that distinguish LNs from bone marrow and blood neutrophils. Functionally, LNs show impaired migratory activity toward chemoattractants and produce high levels of interleukin-6 (IL-6) at steady state and low levels of tumor necrosis factor-α in response to lipopolysaccharide (LPS) challenge. Treating bone marrow neutrophils with bronchoalveolar lavage fluid or prostaglandin E2 induces LN-associated characteristics, including the expression of transglutaminase 2 (Tgm2) and reduced production of inflammatory cytokines upon LPS challenge. Neutrophils from Tgm2-/- mice release high levels of inflammatory cytokines in response to LPS. Lung damage is significantly exacerbated in Tgm2-/- mice in an LPS-induced acute respiratory distress syndrome model. Collectively, we demonstrate that prostaglandin E2 is a key factor for the generation of LNs with unique immune suppressive characteristics, acting through protein kinase A and Tgm2, and LNs play essential roles in protection of the lungs against pathogenic inflammation.


Assuntos
Dinoprostona , Neutrófilos , Animais , Líquido da Lavagem Broncoalveolar/química , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Citocinas/metabolismo , Dinoprostona/metabolismo , Lipopolissacarídeos , Pulmão/patologia , Camundongos , Neutrófilos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
3.
J Cell Mol Med ; 26(12): 3378-3386, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35502484

RESUMO

IM156, a novel biguanide with higher potency of AMP-activated protein kinase activation than metformin, has inhibitory activity against angiogenesis and cancer. In this study, we investigated effects of IM156 against polymicrobial sepsis. Administration of IM156 significantly increased survival rate against caecal ligation and puncture (CLP)-induced sepsis. Mechanistically, IM156 markedly reduced viable bacterial burden in the peritoneal fluid and peripheral blood and attenuated organ damage in a CLP-induced sepsis model. IM156 also inhibited the apoptosis of splenocytes and the production of inflammatory cytokines including IL-1ß, IL-6 and IL-10 in CLP mice. Moreover, IM156 strongly inhibited the generation of reactive oxygen species and subsequent formation of neutrophil extracellular traps in response to lipopolysaccharide in neutrophils. Taken together, these results show that IM156 can inhibit inflammatory response and protect against polymicrobial sepsis, suggesting that IM156 might be a new treatment for sepsis.


Assuntos
Armadilhas Extracelulares , Sepse , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Armadilhas Extracelulares/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/metabolismo , Sepse/metabolismo
4.
Adv Mater Interfaces ; 9(14): 2102046, 2022 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-35538927

RESUMO

A serological immunoassay based on enzyme-linked immunosorbent assay (ELISA) is a crucial tool for screening and identification of human SARS-CoV-2 seroconversion. Various immunoassays are developed to detect the spike 1 (S1) and nucleocapsid (NP) proteins of SARS-CoV-2; however, these serological tests have low sensitivity. Here, a novel microplate (MP) is developed on which a ZnO nanowire (NW) is fabricated by a modified hydrothermal synthesis method. This plate is coated with SARS-CoV-2 NP and used as a fluorescent immunoassay (FIA) to detect antibodies specific for SARS-CoV-2 NP. Compared with the bare MP, the ZnO-NW MP binds high levels (up to 5 µg mL-1) of SARS-CoV-2 NP tagged to histidine without any surface treatment. A novel serological assay based on the ZnO-NW MP is more sensitive than a commercial immunoassay, enabling early detection (within <5 days of a reverse transcription polymerase chain reaction-confirmed COVID-19 infection) of anti-SARS-CoV-2 NP IgG antibodies in asymptomatic patients with COVID-19. This is the first assay to detect early antibody responses to SARS-CoV-2 in asymptomatic patients. Therefore, this serological assay will facilitate accurate diagnosis of COVID-19, as well as estimation of COVID-19 prevalence and incidence.

5.
Viruses ; 14(2)2022 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-35215848

RESUMO

Coronavirus disease 2019 (COVID-19), the pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is characterized by symptoms such as fever, fatigue, a sore throat, diarrhea, and coughing. Although various new vaccines against COVID-19 have been developed, early diagnostics, isolation, and prevention remain important due to virus mutations resulting in rapid and high disease transmission. Amino acid substitutions in the major diagnostic target antigens of SARS-CoV-2 may lower the sensitivity for the detection of SARS-CoV-2. For this reason, we developed specific monoclonal antibodies that bind to epitope peptides as antigens for the rapid detection of SARS-CoV-2 NP. The binding affinity between antigenic peptides and monoclonal antibodies was investigated, and a sandwich pair for capture and detection was employed to develop a rapid biosensor for SARS-CoV-2 NP. The rapid biosensor, based on a monoclonal antibody pair binding to conserved epitopes of SARS-CoV-2 NP, detected cultured virus samples of SARS-CoV-2 (1.4 × 103 TCID50/reaction) and recombinant NP (1 ng/mL). Laboratory confirmation of the rapid biosensor was performed with clinical specimens (n = 16) from COVID-19 patients and other pathogens. The rapid biosensor consisting of a monoclonal antibody pair (75E12 for capture and the 54G6/54G10 combination for detection) binding to conserved epitopes of SARS-CoV-2 NP could assist in the detection of SARS-CoV-2 NP under the circumstance of spreading SARS-CoV-2 variants.


Assuntos
Anticorpos Monoclonais/metabolismo , Anticorpos Antivirais/metabolismo , Técnicas Biossensoriais/métodos , Epitopos/metabolismo , Proteínas do Nucleocapsídeo/metabolismo , SARS-CoV-2/imunologia , Proteínas Virais/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Epitopos/genética , Epitopos/imunologia , Humanos , Imunoensaio , Camundongos , Camundongos Endogâmicos BALB C , Proteínas do Nucleocapsídeo/genética , Proteínas do Nucleocapsídeo/imunologia , Peptídeos/imunologia , Peptídeos/metabolismo , Ligação Proteica , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Proteínas Virais/imunologia
6.
Biosens Bioelectron ; 203: 114034, 2022 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-35114464

RESUMO

Since the beginning of the COVID-19 pandemic, accumulating mutations have led to marked changes in the genetic sequence of SARS-CoV-2. Of these, mutations in the spike (S) protein can alter the properties of the virus, particularly transmissibility and antigenicity. However, it is difficult to detect antigenic variants of the SARS-CoV-2 S protein by immunoassay. Here, we developed an ACE2-based biosensor designed to detect both SARS-CoV-2 S1 mutations and neutralizing antibodies. In "binding mode", the biosensor works by detecting binding of the S protein to an immobilized ACE2 receptor. The ACE2-based biosensor was able to detect S1 proteins of the alpha (500 pg/mL) and beta variants (10 ng/mL), as well as wild-type S1 (10 ng/mL), of SARS-CoV-2. The biosensor distinguished wild-type SARS-CoV-2 S1 from the S1 alpha and beta variants via color differences. In addition, a slight modification to the protocol enabled the ACE2-based biosensor to operate in "blocking mode" to detect neutralizing antibodies in serum samples from COVID-19 patients. Therefore, the ACE2-based biosensor is a versatile test for detecting wild-type S1, S1 mutants, and neutralizing antibodies against SARS-CoV-2. This approach to targeting both the mechanism by which SARS-CoV-2 enters host cells and the subsequent adaptive immune response will facilitate the development of various biosensors against SARS-CoV-2.


Assuntos
Técnicas Biossensoriais , COVID-19 , Enzima de Conversão de Angiotensina 2 , Anticorpos Neutralizantes , Anticorpos Antivirais , COVID-19/diagnóstico , Humanos , Pandemias , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus
7.
Br J Pharmacol ; 178(17): 3463-3475, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33864386

RESUMO

BACKGROUND AND PURPOSE: Reduced bioavailability of NO, a hallmark of sickle cell disease (SCD), contributes to intravascular inflammation, vasoconstriction, vaso-occlusion and organ damage observed in SCD patients. Soluble guanylyl cyclase (sGC) catalyses synthesis of cGMP in response to NO. cGMP-amplifying agents, including NO donors and phosphodiesterase 9 inhibitors, alleviate TNFα-induced inflammation in wild-type C57BL/6 mice and in 'humanised' mouse models of SCD. EXPERIMENTAL APPROACH: Effects of the sGC stimulator olinciguat on intravascular inflammation and renal injury were studied in acute (C57BL6 and Berkeley mice) and chronic (Townes mice) mouse models of TNFα-induced and systemic inflammation associated with SCD. KEY RESULTS: Acute treatment with olinciguat attenuated increases in plasma biomarkers of endothelial cell activation and leukocyte-endothelial cell interactions in TNFα-challenged mice. Co-treatment with hydroxyurea, an FDA-approved SCD therapeutic agent, further augmented the anti-inflammatory effect of olinciguat. In the Berkeley mouse model of TNFα-induced vaso-occlusive crisis, a single dose of olinciguat attenuated leukocyte-endothelial cell interactions, improved blood flow and prolonged survival time compared to vehicle-treated mice. In Townes SCD mice, plasma biomarkers of inflammation and endothelial cell activation were lower in olinciguat- than in vehicle-treated mice. In addition, kidney mass, water consumption, 24-h urine excretion, plasma levels of cystatin C and urinary excretion of N-acetyl-ß-d-glucosaminidase and neutrophil gelatinase-associated lipocalin were lower in Townes mice treated with olinciguat than in vehicle-treated mice. CONCLUSION AND IMPLICATIONS: Our results suggest that the sGC stimulator olinciguat attenuates inflammation, vaso-occlusion and kidney injury in mouse models of SCD and systemic inflammation.


Assuntos
Anemia Falciforme , Doenças Vasculares , Anemia Falciforme/complicações , Anemia Falciforme/tratamento farmacológico , Animais , Humanos , Inflamação , Camundongos , Camundongos Endogâmicos C57BL , Guanilil Ciclase Solúvel
8.
Biosens Bioelectron ; 171: 112715, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33099241

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), a newly emerging human infectious disease. Because no specific antiviral drugs or vaccines are available to treat COVID-19, early diagnostics, isolation, and prevention are crucial for containing the outbreak. Molecular diagnostics using reverse transcription polymerase chain reaction (RT-PCR) are the current gold standard for detection. However, viral RNAs are much less stable during transport and storage than proteins such as antigens and antibodies. Consequently, false-negative RT-PCR results can occur due to inadequate collection of clinical specimens or poor handling of a specimen during testing. Although antigen immunoassays are stable diagnostics for detection of past infection, infection progress, and transmission dynamics, no matched antibody pair for immunoassay of SARS-CoV-2 antigens has yet been reported. In this study, we designed and developed a novel rapid detection method for SARS-CoV-2 spike 1 (S1) protein using the SARS-CoV-2 receptor ACE2, which can form matched pairs with commercially available antibodies. ACE2 and S1-mAb were paired with each other for capture and detection in a lateral flow immunoassay (LFIA) that did not cross-react with SARS-CoV Spike 1 or MERS-CoV Spike 1 protein. The SARS-CoV-2 S1 (<5 ng of recombinant proteins/reaction) was detected by the ACE2-based LFIA. The limit of detection of our ACE2-LFIA was 1.86 × 105 copies/mL in the clinical specimen of COVID-19 Patients without no cross-reactivity for nasal swabs from healthy subjects. This is the first study to detect SARS-CoV-2 S1 antigen using an LFIA with matched pair consisting of ACE2 and antibody. Our findings will be helpful to detect the S1 antigen of SARS-CoV-2 from COVID-19 patients.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas Biossensoriais/instrumentação , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Peptidil Dipeptidase A/química , Pneumonia Viral/diagnóstico , Glicoproteína da Espícula de Coronavírus/análise , Enzima de Conversão de Angiotensina 2 , Anticorpos Monoclonais/química , Técnicas Biossensoriais/economia , COVID-19 , Teste para COVID-19 , Técnicas de Laboratório Clínico/economia , Técnicas de Laboratório Clínico/instrumentação , Infecções por Coronavirus/economia , Desenho de Equipamento , Humanos , Imunoensaio/economia , Imunoensaio/instrumentação , Imunoconjugados/química , Pandemias , SARS-CoV-2 , Sensibilidade e Especificidade , Fatores de Tempo
9.
Complement Ther Med ; 52: 102481, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32951731

RESUMO

Sickle Cell Disease (SCD) is a chronic hemolytic disorder associated with frequent pain episodes, end organ damage and a shortened lifespan. Currently there exist no disease specific targeted therapies for the treatment of acute vaso-occlusive crisis (VOC) and management with analgesics and hydration is purely supportive. Improvement in understanding of disease pathophysiology has resulted in a great interest in disease modifying novel therapies and many are being evaluated in clinical trials. Here we report the results from the pre-specified mid-point analysis of the Phase 2 study of Intravenous Gamma Globulin (IVIG) for the treatment of acute VOC in patients with SCD and lessons learned.


Assuntos
Anemia Falciforme/tratamento farmacológico , Imunoglobulinas Intravenosas/uso terapêutico , Manejo da Dor/métodos , gama-Globulinas/uso terapêutico , Adolescente , Adulto , Anemia Falciforme/complicações , Criança , Método Duplo-Cego , Feminino , Humanos , Fatores Imunológicos/uso terapêutico , Masculino , Adulto Jovem
10.
Immunity ; 53(2): 417-428.e4, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32735844

RESUMO

Psychological stress has adverse effects on various human diseases, including those of the cardiovascular system. However, the mechanisms by which stress influences disease activity remain unclear. Here, using vaso-occlusive episodes (VOEs) of sickle cell disease as a vascular disease model, we show that stress promotes VOEs by eliciting a glucocorticoid hormonal response that augments gut permeability, leading to microbiota-dependent interleukin-17A (IL-17A) secretion from T helper 17 (Th17) cells of the lamina propria, followed by the expansion of the circulating pool of aged neutrophils that trigger VOEs. We identify segmented filamentous bacteria as the commensal essential for the stress-induced expansion of aged neutrophils that enhance VOEs in mice. Importantly, the inhibition of glucocorticoids synthesis, blockade of IL-17A, or depletion of the Th17 cell-inducing gut microbiota markedly reduces stress-induced VOEs. These results offer potential therapeutic targets to limit the impact of psychological stress on acute vascular occlusion.


Assuntos
Anemia Falciforme/patologia , Microbioma Gastrointestinal/imunologia , Interleucina-17/metabolismo , Estresse Psicológico/patologia , Células Th17/imunologia , Anemia Falciforme/psicologia , Animais , Bactérias/imunologia , Linhagem Celular , Vida Livre de Germes , Glucocorticoides/biossíntese , Fator Estimulador de Colônias de Granulócitos/metabolismo , Células HEK293 , Humanos , Sistema Hipotálamo-Hipofisário/metabolismo , Inflamação/imunologia , Inflamação/psicologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/imunologia
11.
Sci Rep ; 9(1): 14349, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31586128

RESUMO

The inflammasome is a specialized multiprotein oligomer that regulates IL-1ß production. Although regulation of the inflammasome is related to crucial inflammatory disorders such as sepsis, pharmacological inhibitors that effectively inhibit inflammasome activity are limited. Here, we evaluated the effects of a phospholipase D1 (PLD1)-selective inhibitor (VU0155069) against sepsis and inflammasome activation. VU0155069 strongly enhances survival rate in cecal ligation and puncture (CLP)-induced sepsis by inhibiting lung inflammation, leukocyte apoptosis, and the production of proinflammatory cytokines, especially IL-1ß. VU0155069 also significantly blocked IL-1ß production, caspase-1 activation, and pyroptosis caused by several inflammasome-activating signals in the bone marrow-derived macrophages (BMDMs). However, VU0155069 did not affect LPS-induced activation of signaling molecules such as MAPK, Akt, NF-κB, and NLRP3 expression in the BMDMs. VU0155069 also failed to affect mitochondrial ROS generation and calcium increase caused by nigericin or ATP, and subsequent ASC oligomerization caused by several inflammasome-activating signals. VU0155069 indirectly inhibited caspase-1 activity caused by LPS + nigericin in BMDMs independent of PLD1 activity. We demonstrated that a PLD1 inhibitor, VU0155069, shows anti-septic activity as well as inflammasome-inhibiting effects. Our results suggest that VU0155069 can be considered a novel inflammasome inhibitor.


Assuntos
Anti-Inflamatórios/farmacologia , Benzimidazóis/farmacologia , Inflamassomos/antagonistas & inibidores , Piperidinas/farmacologia , Sepse/tratamento farmacológico , Animais , Anti-Inflamatórios/uso terapêutico , Benzimidazóis/uso terapêutico , Caspase 1/metabolismo , Modelos Animais de Doenças , Humanos , Inflamassomos/imunologia , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Lipopolissacarídeos/imunologia , Camundongos , Camundongos Knockout , Fosfolipase D/antagonistas & inibidores , Fosfolipase D/genética , Piperidinas/uso terapêutico , Piroptose/efeitos dos fármacos , Piroptose/imunologia , Espécies Reativas de Oxigênio/metabolismo , Sepse/imunologia , Sepse/patologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia
12.
Sci Rep ; 9(1): 7242, 2019 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-31076618

RESUMO

We examined the role of phospholipase D2 (PLD2) on acetaminophen (APAP)-induced acute liver injury using a PLD2 inhibitor (CAY10594). 500 mg/kg of APAP challenge caused acute liver damage. CAY10594 administration markedly blocked the acute liver injury in a dose-dependent manner, showing almost complete inhibition with 8 mg/kg of CAY10594. During the pathological progress of acute liver injury, GSH levels are decreased, and this is significantly recovered upon the administration of CAY10594 at 6 hours post APAP challenge. GSK-3ß (Serine 9)/JNK phosphorylation is mainly involved in APAP-induced liver injury. CAY10594 administration strongly blocked GSK-3ß (Serine 9)/JNK phosphorylation in the APAP-induced acute liver injury model. Consistently, sustained JNK activation in the cytosol and mitochondria from hepatocytes were also decreased in CAY10594-treated mice. Many types of immune cells are also implicated in APAP-induced liver injury. However, neutrophil and monocyte populations were not different between vehicle- and CAY10594-administered mice which are challenged with APAP. Therapeutic administration of CAY10594 also significantly attenuated liver damage caused by the APAP challenge, eliciting an enhanced survival rate. Taken together, these results indicate that PLD2 is involved in the intrinsic response pathway of hepatocytes driving the pathogenesis of APAP-induced acute liver injury, and PLD2 may therefore represent an important therapeutic target for patients with drug-induced liver injury.


Assuntos
Acetaminofen/efeitos adversos , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fosfolipase D/antagonistas & inibidores , Fosforilação/efeitos dos fármacos , Animais , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
BMB Rep ; 51(8): 418-423, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30021674

RESUMO

Emergency granulopoiesis is a very important strategy to supply efficient neutrophil number in response to infection. However, molecular mechanism involved in this process remains unclear. Here, we found that administration of WKYMVm, an immune modulating peptide, to septic mice strongly increased neutrophil number through augmented emergency granulopoiesis. WKYMVm-induced emergency granulopoiesis was blocked not only by a formyl peptide receptor 2 (FPR2) antagonist (WRW4), but also by FPR2 deficiency. As progenitors of neutrophils, Lin-c-kit+Sca-1- cells expressed FPR2. WKYMVm-induced emergency granulopoiesis was also blocked by a phospholipase C inhibitor (U-73122). These results suggest that WKYMVm can stimulate emergency granulopoiesis via FPR2 and phospholipase C enzymatic activity. [BMB Reports 2018; 51(8): 418-423].


Assuntos
Hematopoese/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Oligopeptídeos/farmacologia , Receptores de Formil Peptídeo/metabolismo , Fosfolipases Tipo C/metabolismo , Animais , Interações Medicamentosas , Estrenos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/citologia , Neutrófilos/enzimologia , Neutrófilos/metabolismo , Pirrolidinonas/farmacologia , Receptores de Formil Peptídeo/fisiologia , Transdução de Sinais/fisiologia , Fosfolipases Tipo C/antagonistas & inibidores
14.
Exp Mol Med ; 49(12): e407, 2017 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-29217823

RESUMO

We investigated the effect of lysophosphatidic acid (LPA) in experimental acetaminophen (APAP)-induced acute liver injury. LPA administration significantly reduced APAP-challenged acute liver injury, showing attenuated liver damage, liver cell death and aspartate aminotransferase and alanine aminotransferase levels. APAP overdose-induced mortality was also significantly decreased by LPA administration. Regarding the mechanism involved in LPA-induced protection against acute liver injury, LPA administration significantly increased the glutathione level, which was markedly decreased in APAP challenge-induced acute liver injury. LPA administration also strongly blocked the APAP challenge-elicited phosphorylation of JNK, ERK and GSK3ß, which are involved in the pathogenesis of acute liver injury. Furthermore, LPA administration decreased the production of TNF-α and IL-1ß in an experimental drug-induced liver injury animal model. Mouse primary hepatocytes express LPA1,3-6, and injection of the LPA receptor antagonist KI16425 (an LPA1,3-selective inhibitor) or H2L 5765834 (an LPA1,3,5-selective inhibitor) did not reverse the LPA-induced protective effects against acute liver injury. The therapeutic administration of LPA also blocked APAP-induced liver damage, leading to an increased survival rate. Collectively, these results indicate that the well-known bioactive lipid LPA can block the pathogenesis of APAP-induced acute liver injury by increasing the glutathione level but decreasing inflammatory cytokines in an LPA1,3,5-independent manner. Our results suggest that LPA might be an important therapeutic agent for drug-induced liver injury.


Assuntos
Acetaminofen/efeitos adversos , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Fígado/efeitos dos fármacos , Lisofosfolipídeos/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Interleucina-1beta/genética , Isoxazóis/administração & dosagem , Fígado/lesões , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Propionatos/administração & dosagem , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/genética , Fator de Necrose Tumoral alfa/genética
15.
BMB Rep ; 49(9): 520-5, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27502013

RESUMO

We investigated the effects of two antimicrobial peptides (AMPs) isolated from Scolopendra subspinipes mutilans on neutrophil activity. Stimulation of mouse neutrophils with the two AMPs elicited chemotactic migration of the cells in a pertussis toxin-sensitive manner. The two AMPs also stimulated activation of ERK and Akt, which contribute to chemotactic migration of neutrophils. We found that AMP-stimulated neutrophil chemotaxis was blocked by a formyl peptide receptor (FPR) 1 antagonist (cyclosporin H); moreover the two AMPs stimulated the chemotactic migration of FPR1-expressing RBL-2H3 cells but not of vector-expressing RBL-2H3 cells. We also found that the two AMPs stimulate neutrophil migration in vivo, and that this effect is blocked in FPR1-deficient mice. Taken together, our results suggest that the two AMPs stimulate neutrophils, leading to chemotactic migration through FPR1, and the two AMPs will be useful for the study of FPR1 signaling and neutrophil activation. [BMB Reports 2016; 49(9): 520-525].


Assuntos
Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Quimiotaxia de Leucócito/efeitos dos fármacos , Receptores de Formil Peptídeo/metabolismo , Animais , Western Blotting , Linhagem Celular , Ciclosporina/farmacologia , Alcaloides Diterpenos , Medicamentos de Ervas Chinesas/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Toxina Pertussis/toxicidade , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Formil Peptídeo/antagonistas & inibidores , Receptores de Formil Peptídeo/genética , Transdução de Sinais/efeitos dos fármacos
16.
J Cell Biochem ; 117(4): 978-87, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26399466

RESUMO

A natural compound C23 H32 O4 Cl, ascochlorin (ASC) isolated from an incomplete fungus, Ascochyta viciae has been known to have several biological activities as an antibiotic, antifungal, anti-cancer, anti-hypolipidemic, and anti-hypertension agent. In this study, anti-inflammatory activity has been investigated in lipopolysaccharide (LPS)-induced murine macrophage RAW 264.7 cells, since ASC has not been observed on the inflammatory events. The present study has clearly shown that ASC (1-50 µM) significantly suppressed the production of nitric oxide (NO) and prostaglandin E2 (PGE2 ) and decreased the gene expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) in a dose-dependent manner. Moreover, ASC inhibited the mRNA expression and the protein secretion of interleukin (IL)-1ß and IL-6 but not tumor necrosis factor (TNF)-α in LPS-stimulated RAW 264.7 macrophage cells. In addition, ASC suppressed nuclear translocation and DNA binding affinity of nuclear factor-κB (NF-κB). Furthermore, ASC down-regulated phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) and p-p38. These results demonstrate that ASC exhibits anti-inflammatory effects in RAW 264.7 macrophage cells.


Assuntos
Alcenos/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Ciclo-Oxigenase 2/genética , Lipopolissacarídeos/antagonistas & inibidores , Macrófagos/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Fenóis/farmacologia , Transdução de Sinais/efeitos dos fármacos , Alcenos/isolamento & purificação , Animais , Anti-Inflamatórios não Esteroides/isolamento & purificação , Linhagem Celular , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/antagonistas & inibidores , Dinoprostona/biossíntese , Regulação da Expressão Gênica , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , NF-kappa B/antagonistas & inibidores , NF-kappa B/genética , NF-kappa B/metabolismo , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Fenóis/isolamento & purificação , Transporte Proteico , Saccharomycetales/química , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
17.
J Exp Med ; 212(9): 1381-90, 2015 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-26282875

RESUMO

We determined the function of phospholipase D2 (PLD2) in host defense in highly lethal mouse models of sepsis using PLD2(-/-) mice and a PLD2-specific inhibitor. PLD2 deficiency not only increases survival but also decreases vital organ damage during experimental sepsis. Production of several inflammatory cytokines (TNF, IL-1ß, IL-17, and IL-23) and the chemokine CXCL1, as well as cellular apoptosis in immune tissues, kidney, and liver, are markedly decreased in PLD2(-/-) mice. Bactericidal activity is significantly increased in PLD2(-/-) mice, which is mediated by increased neutrophil extracellular trap formation and citrullination of histone 3 through peptidylarginine deiminase activation. Recruitment of neutrophils to the lung is markedly increased in PLD2(-/-) mice. Furthermore, LPS-induced induction of G protein-coupled receptor kinase 2 (GRK2) and down-regulation of CXCR2 are markedly attenuated in PLD2(-/-) mice. A CXCR2-selective antagonist abolishes the protection conferred by PLD2 deficiency during experimental sepsis, suggesting that enhanced CXCR2 expression, likely driven by GRK2 down-regulation in neutrophils, promotes survival in PLD2(-/-) mice. Furthermore, adoptively transferred PLD2(-/-) neutrophils significantly protect WT recipients against sepsis-induced death compared with transferred WT neutrophils. We suggest that PLD2 in neutrophils is essential for the pathogenesis of experimental sepsis and that pharmaceutical agents that target PLD2 may prove beneficial for septic patients.


Assuntos
Regulação para Baixo , Armadilhas Extracelulares/metabolismo , Neutrófilos/metabolismo , Fosfolipase D/metabolismo , Receptores de Interleucina-8B/biossíntese , Sepse/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Armadilhas Extracelulares/genética , Quinase 2 de Receptor Acoplado a Proteína G/genética , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Lipopolissacarídeos/toxicidade , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Knockout , Infiltração de Neutrófilos/genética , Neutrófilos/patologia , Fosfolipase D/genética , Receptores de Interleucina-8B/genética , Sepse/induzido quimicamente , Sepse/genética , Sepse/patologia
18.
Biochem Biophys Res Commun ; 464(3): 774-9, 2015 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-26166823

RESUMO

Sepsis is a serious, life-threatening, infectious disease. In this study, we demonstrate that sucrose methyl 3-formyl-4-methylpentanoate (SMFM), a novel natural compound isolated from garlic (Allium sativum L.), markedly enhances survival rates by inhibiting lung inflammation in a cecal ligation and puncture (CLP) experimental polymicrobial sepsis model. SMFM strongly reduced bacterial colony units from peritoneal fluid in CLP mice by stimulating the generation of reactive oxygen species. Lymphocyte apoptosis in spleens from CLP mice was also markedly decreased by SMFM administration. SMFM also significantly inhibited the production of proinflammatory cytokines, such as TNF-α, interleukin-1ß (IL-1ß) and IL-6, in CLP mice. Lipopolysaccharide-stimulated production of TNF-α and IL-6 were also strongly inhibited by SMFM in mouse bone marrow-derived macrophages. Taken together, our results indicate that SMFM has therapeutic effects against polymicrobial sepsis that are mediated by enhanced microbial killing and blockage of cytokine storm.


Assuntos
Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Alho/química , Sepse/tratamento farmacológico , Sacarose/análogos & derivados , Animais , Carga Bacteriana/efeitos dos fármacos , Citocinas/biossíntese , Modelos Animais de Doenças , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/toxicidade , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Fitoterapia , Sepse/imunologia , Sepse/microbiologia , Sacarose/química , Sacarose/farmacologia
19.
Mar Drugs ; 13(6): 3936-49, 2015 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-26090845

RESUMO

For innate immune defense, lower animals such as fish and amphibian are covered with skin mucus, which acts as both a mechanical and biochemical barrier. Although several mucus sources have been isolated and studied for their biochemical and immunological functions, the precise mechanism(s) of action remains unknown. In the present study, we additionally found the eel skin mucus (ESM) to be a promising candidate for use in anti-tumor therapy. Our results showed that the viability of K562 cells was decreased in a dose-dependent manner by treatment with the isolated ESM. The cleaved forms of caspase-9, caspase-3 and poly adenosine diphosphate-ribose polymerase were increased by ESM. The levels of Bax expression and released cytochrome C were also increased after treatment with ESM. Furthermore, during the ESM mediated-apoptosis, phosphorylation levels of ERK1/2 and p38 but not JNK were increased and cell viabilities of the co-treated cells with ESM and inhibitors of ERK 1/2 or p38 were also increased. In addition, treatment with lactose rescued the ESM-mediated decrease in cell viability, indicating lactose-containing glycans in the leukemia cells acted as a counterpart of the ESM for interaction. Taken together, these results suggest that ESM could induce mitochondria-mediated apoptosis through membrane interaction of the K562 human leukemia cells. To the best of our knowledge, this is the first observation that ESM has anti-tumor activity in human cells.


Assuntos
Anguilla/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Muco/metabolismo , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/isolamento & purificação , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Células K562 , Lactose/metabolismo , Leucemia/tratamento farmacológico , Leucemia/patologia , Mitocôndrias/metabolismo , Polissacarídeos/metabolismo , Pele/metabolismo
20.
BMB Rep ; 48(6): 336-41, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25541055

RESUMO

Sepsis is a life-threatening, infectious, systemic inflammatory disease. In this study, we investigated the therapeutic effect of α-cubebenoate, a novel compound isolated from Schisandra chinensis against polymicrobial sepsis in a cecal ligation and puncture (CLP) experimental model. Administration of α-cubebenoate strongly enhanced survival in the CLP model. α-cubebenoate administration also markedly blocked CLP-induced lung inflammation and increased bactericidal activity by enhancing phagocytic activity and hydrogen peroxide generation in mouse bone marrow-derived macrophages and neutrophils. Expression of two important inflammatory cytokines, IL-1ß and IL-6, was strongly increased in the CLP model, and this was dramatically blocked by α-cubebenoate. Lymphocyte apoptosis and caspase-3 activation, which are associated with immune paralysis during sepsis, were markedly attenuated by α-cubebenoate. Taken together, our findings indicate that α-cubebenoate, a natural compound isolated from Schisandra chinensis, is a powerful potential anti-septic agent.


Assuntos
Sepse/tratamento farmacológico , Sesquiterpenos de Guaiano/farmacologia , Animais , Anti-Infecciosos Locais/isolamento & purificação , Anti-Infecciosos Locais/farmacologia , Apoptose/efeitos dos fármacos , Atividade Bactericida do Sangue/efeitos dos fármacos , Citocinas/biossíntese , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Linfócitos/efeitos dos fármacos , Linfócitos/patologia , Camundongos , Fagocitose/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Schisandra/química , Sepse/metabolismo , Sepse/patologia , Sesquiterpenos de Guaiano/isolamento & purificação
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