RESUMO
BACKGROUND: Endotoxin-associated acute kidney injury (AKI), a disease characterized by marked oxidative stress and inflammation disease, is a major cause of mortality in critically ill patients. Mitochondrial fission and pyroptosis often occur in AKI. However, the underlying biological pathways involved in endotoxin AKI remain poorly understood, especially those related to mitochondrial dynamics equilibrium disregulation and pyroptosis. Previous studies suggest that heme oxygenase- (HO-) 1 confers cytoprotection against AKI during endotoxic shock, and PTEN-induced putative kinase 1 (PINK1) takes part in mitochondrial dysfunction. Thus, in this study, we examine the roles of HO-1/PINK1 in maintaining the dynamic process of mitochondrial fusion/fission to inhibit pyroptosis and mitigate acute kidney injury in rats exposed to endotoxin. METHODS: An endotoxin-associated AKI model induced by lipopolysaccharide (LPS) was used in our study. Wild-type (WT) rats and PINK1 knockout (PINK1KO) rats, respectively, were divided into four groups: the control, LPS, Znpp+LPS, and Hemin+LPS groups. Rats were sacrificed 6 h after intraperitoneal injecting LPS to assess renal function, oxidative stress, and inflammation by plasma. Mitochondrial dynamics, morphology, and pyroptosis were evaluated by histological examinations. RESULTS: In the rats with LPS-induced endotoxemia, the expression of HO-1 and PINK1 were upregulated at both mRNA and protein levels. These rats also exhibited inflammatory response, oxidative stress, mitochondrial fission, pyroptosis, and decreased renal function. After upregulating HO-1 in normal rats, pyroptosis was inhibited; mitochondrial fission and inflammatory response to oxidative stress were decreased; and the renal function was improved. The effects were reversed by adding Znpp (a type of HO-1 inhibitor). Finally, after PINK1 knockout, there is no statistical difference in the LPS-treated group and Hemin or Znpp pretreated group. CONCLUSIONS: HO-1 inhibits inflammation response and oxidative stress and regulates mitochondria fusion/fission to inhibit pyroptosis, which can alleviate endotoxin-induced AKI by PINK1.
Assuntos
Injúria Renal Aguda/genética , Heme Oxigenase (Desciclizante)/genética , Dinâmica Mitocondrial/genética , Proteínas Quinases/genética , Piroptose/genética , Choque Séptico/genética , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/enzimologia , Injúria Renal Aguda/patologia , Animais , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/enzimologia , Células Epiteliais/patologia , Regulação da Expressão Gênica , Técnicas de Inativação de Genes , Heme Oxigenase (Desciclizante)/antagonistas & inibidores , Heme Oxigenase (Desciclizante)/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Rim/efeitos dos fármacos , Rim/enzimologia , Rim/patologia , Lipocalina-2/genética , Lipocalina-2/metabolismo , Lipopolissacarídeos/administração & dosagem , Masculino , Dinâmica Mitocondrial/efeitos dos fármacos , Estresse Oxidativo , Proteínas Quinases/deficiência , Protoporfirinas/farmacologia , Piroptose/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Choque Séptico/induzido quimicamente , Choque Séptico/enzimologia , Choque Séptico/patologia , Transdução de Sinais , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The proline-specific enzymes dipeptidyl peptidase 4 (DPP4), prolylcarboxypeptidase (PRCP), fibroblast activation protein α (FAP) and prolyl oligopeptidase (PREP) are known for their involvement in the immune system and blood pressure regulation. Only very limited information is currently available on their enzymatic activity and possible involvement in patients with sepsis and septic-shock. The activity of the enzymes was measured in EDTA-plasma of patients admitted to the intensive care unit (ICU): 40 septic shock patients (sepsis-2) and 22 ICU control patients after major intracranial surgery. These data were used to generate receiver operating characteristic (ROC) curves. A survival analysis (at 90 days) and an association study with other parameters was performed. PRCP (day 1) and PREP (all days) enzymatic activities were higher in septic shock patients compared to controls. In contrast, FAP and DPP4 were lower in these patients on all studied time points. Since large differences were found, ROC curves were generated and these yielded area under the curve (AUC) values for PREP, FAP and DPP4 of 0.88 (CI: 0.80-0.96), 0.94 (CI: 0.89-0.99) and 0.86 (CI: 0.77-0.95), respectively. PRCP had a lower predicting value with an AUC of 0.71 (CI: 0.58-0.83). A nominally significant association was observed between survival and the DPP4 enzymatic activity at day 1 (p<0.05), with a higher DPP4 activity being associated with an increase in survival. All four enzymes were dysregulated in septic shock patients. DPP4, FAP and PREP are good in discriminating between septic shock patients and ICU controls and should be further explored to see whether they are already dysregulated in earlier stages, opening perspectives for their further investigation as biomarkers in sepsis. DPP4 also shows potential as a prognostic biomarker. Additionally, the associations found warrant further research.
Assuntos
Carboxipeptidases/sangue , Dipeptidil Peptidase 4/sangue , Gelatinases/sangue , Proteínas de Membrana/sangue , Serina Endopeptidases/sangue , Choque Séptico/sangue , Choque Séptico/enzimologia , Área Sob a Curva , Biomarcadores/sangue , Cuidados Críticos , Endopeptidases , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Prolina/metabolismo , Prolil Oligopeptidases , Estudos Prospectivos , Curva ROC , Choque Séptico/mortalidade , Choque Séptico/terapia , Análise de SobrevidaRESUMO
Ellipticine, a natural product from Ochrosia elliptica, has been broadly investigated for its anticancer effects. Although inflammation has been clearly identified as a key factor in the onset and progression of cancer, the relationship between ellipticine and inflammation remains unknown. Hence, the aims of the present study were to assess the effects of ellipticine on the inflammatory responses to lipopolysaccharide (LPS)-induced macrophages and to potentially identify the underlying mechanisms involved. Viability testing showed that ellipticine was not significantly toxic to Raw264.7 cells and actually conveyed protective effects to LPS-stimulated Raw264.7 cells and human peripheral blood monocytes by decreasing the secretion of inflammatory factors (TNF-α and IL-6). The results of western blot analysis and electrophoretic mobility shift assays showed that ellipticine markedly suppressed LPS-induced activation of the JNK/AP-1 (c-Fos and c-Jun) signaling pathway, but not ERK/p38/NF-κB pathway (p65 and p50) activation. Furthermore, ellipticine reduced the inflammatory response and mortality in a mouse model of LPS-induced endotoxic shock. Collectively, these data indicate that ellipticine may be a potential therapeutic agent for the treatment of inflammation-associated diseases.
Assuntos
Anti-Inflamatórios/farmacologia , Elipticinas/farmacologia , Inflamação/prevenção & controle , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Choque Séptico/prevenção & controle , Fator de Transcrição AP-1/metabolismo , Adulto , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Humanos , Inflamação/induzido quimicamente , Inflamação/enzimologia , Inflamação/imunologia , Interleucina-6/metabolismo , Lipopolissacarídeos , Macrófagos/enzimologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células RAW 264.7 , Choque Séptico/induzido quimicamente , Choque Séptico/enzimologia , Choque Séptico/imunologia , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The execution of shock following high dose E. coli lipopolysaccharide (LPS) or bacterial sepsis in mice required pro-apoptotic caspase-8 in addition to pro-pyroptotic caspase-11 and gasdermin D. Hematopoietic cells produced MyD88- and TRIF-dependent inflammatory cytokines sufficient to initiate shock without any contribution from caspase-8 or caspase-11. Both proteases had to be present to support tumor necrosis factor- and interferon-ß-dependent tissue injury first observed in the small intestine and later in spleen and thymus. Caspase-11 enhanced the activation of caspase-8 and extrinsic cell death machinery within the lower small intestine. Neither caspase-8 nor caspase-11 was individually sufficient for shock. Both caspases collaborated to amplify inflammatory signals associated with tissue damage. Therefore, combined pyroptotic and apoptotic signaling mediated endotoxemia independently of RIPK1 kinase activity and RIPK3 function. These observations bring to light the relevance of tissue compartmentalization to disease processes in vivo where cytokines act in parallel to execute diverse cell death pathways.
Assuntos
Caspase 8/metabolismo , Caspases/metabolismo , Infecções por Escherichia coli/enzimologia , Infecções por Escherichia coli/fisiopatologia , Choque Séptico/enzimologia , Choque Séptico/fisiopatologia , Animais , Apoptose , Proteínas Reguladoras de Apoptose/metabolismo , Caspase 8/genética , Caspases/genética , Caspases Iniciadoras , Células Cultivadas , Feminino , Inflamação/metabolismo , Inflamação/patologia , Fator Regulador 3 de Interferon/genética , Interferon beta/sangue , Interferon beta/metabolismo , Intestino Delgado/patologia , Peptídeos e Proteínas de Sinalização Intracelular , Lipopolissacarídeos/toxicidade , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Ligação a Fosfato , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Transdução de Sinais , Baço/patologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Berberine, a major isoquinoline alkaloid found in medicinal herbs, has been reported to possess anti-inflammatory effects; however, the underlying mechanisms responsible for its actions are poorly understood. In the present study, we investigated the inhibitory effects of berberine and the molecular mechanisms involved in lipopolysaccharide (LPS)-treated RAW 264.7 and THP-1 macrophages and its effects in LPS-induced septic shock in mice. In both macrophage cell types, berberine inhibited the LPS-induced nitric oxide (NO) production and inducible NO synthase (iNOS) protein expression, but it had no effect on iNOS mRNA transcription. Suppression of LPS-induced iNOS protein expression by berberine occurred via a human antigen R (HuR)-mediated reduction of iNOS mRNA stability. Molecular data revealed that the suppression on the LPS-induced HuR binding to iNOS mRNA by berberine was accompanied by a reduction in nucleocytoplasmic HuR shuttling. Pretreatment with berberine reduced LPS-induced iNOS protein expression and the cytoplasmic translocation of HuR in liver tissues and increased the survival rate of mice with LPS-induced endotoxemia. These results show that the suppression of iNOS protein expression by berberine under LPS-induced inflammatory conditions is associated with a reduction in iNOS mRNA stability resulting from inhibition of the cytoplasmic translocation of HuR.
Assuntos
Anti-Inflamatórios/farmacologia , Berberina/farmacologia , Proteína Semelhante a ELAV 1/metabolismo , Macrófagos/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/biossíntese , Estabilidade de RNA/efeitos dos fármacos , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/uso terapêutico , Berberina/administração & dosagem , Berberina/uso terapêutico , Linhagem Celular , Proteína Semelhante a ELAV 1/genética , Técnicas de Silenciamento de Genes , Humanos , Imuno-Histoquímica , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Óxido Nítrico Sintase Tipo II/genética , Ligação Proteica , Reação em Cadeia da Polimerase em Tempo Real , Choque Séptico/enzimologia , Choque Séptico/prevenção & controleRESUMO
BACKGROUND: Our aim was to investigate whether plasma glutathione reductase (GR) activity is well correlated with the erythrocyte-reduced glutathione (GSH)/glutathione disulfide (GSSG) ratio and is associated with the mortality of septic shock. MATERIALS AND METHODS: This study was conducted on male Sprague-Dawley rats and patients admitted to the intensive care unit with septic shock. To induce endotoxemia in rats, vehicle or lipopolysaccharide (LPS) at dosages of 5 or 10 mg/kg were injected into a tail vein. Animals were then euthanized 6 h post-LPS. Based on the 28-d mortality, the enrolled patients were divided into the survivors and nonsurvivors. We obtained blood samples from patients at admission (0 h) and 24 h after admission to the intensive care unit. RESULTS: In endotoxemic rats, the erythrocyte GSH/GSSG ratio, erythrocyte GR activity, and plasma GR activity in the 10 mg/kg of LPS group were lower than those in the sham and 5 mg/kg of LPS groups. In patients with septic shock, decrease in plasma GR activity at 24 h was independently associated with an increase in 28-d mortality (odds ratio, 0.828; 95% confidence interval, 0.690-0.992, P = 0.041). Plasma GR activity was correlated with erythrocyte GR activity (Spearman ρ = 0.549, P < 0.001) and the erythrocyte GSH/GSSG ratio (rho = 0.367, P = 0.009) at 24 h. CONCLUSIONS: Plasma GR activity was well correlated with erythrocyte GR activity and the erythrocyte GSH/GSSG ratio, and a decrease in plasma GR activity was associated with an increase in the mortality of septic shock patients.
Assuntos
Glutationa Redutase/sangue , Choque Séptico/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Biomarcadores/sangue , Western Blotting , Eritrócitos/metabolismo , Feminino , Glutationa/sangue , Dissulfeto de Glutationa/sangue , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Prognóstico , Ratos , Ratos Sprague-Dawley , Choque Séptico/sangue , Choque Séptico/enzimologia , Análise de SobrevidaRESUMO
BACKGROUND: Seven isoforms of histone deacetylase Class III have been reported - Sirtuin (SIRT) 1-7. We recently demonstrated that EX-527, an inhibitor of SIRT1, reduces mortality in a mouse model of lethal-cecal-ligationand- puncture (CLP)-induced septic shock. Our present study was aimed at determining whether selective inhibition of SIRT2, with AGK2, would decrease animal death and attenuate the inflammatory response in a septic model. METHODS: Experiment I: C57BL/6J mice were intraperitoneally given either AGK2 (82 mg/kg) in dimethyl sulfoxide (DMSO) or DMSO alone, and 2 h later subjected to CLP. Survival was monitored for 240 hours. Experiment II: mice treated the same way as Experiment I, were grouped into (i) DMSO vehicle, and (ii) AGK2, with sham mice (operating but without any treatment) serving as controls. Peritoneal fluid and peripheral blood were examined at 24 and 48 hours for cytokine production. Samples of blood at 48 h were also allocated to assess coagulability using Thrombelastography (TEG). Morphological changes of bone marrow were evaluated from long bones (femurs and tibias) with hematoxylin and eosin (H&E) staining. Bone marrow atrophy was quantified by a blinded pathologist. Experiment III: cytokines in supernatant of the cultured normal primary splenocytes were measured after the cells were stimulated by lipopolysaccharide and treated with or without AGK2 (10 µM) for 6 hours. RESULTS: AGK2 significantly reduced mortality and decreased levels of cytokines in blood (TNF-α: 298.3±24.6 vs 26.8±2.8 pg/ml, p=0.0034; IL-6: 633.4±82.8 vs 232.6±133.0 pg/ml, p=0.0344) and peritoneal fluid (IL-6: 704.8±67.7 vs 391.4±98.5 pg/ml, p=0.033) compared to vehicle control. Also, AGK2 suppressed the TNF-α and IL-6 production in the cultured splenocytes (TNF-α: 68.1±6.4 vs 23.9±2.8 pg/ml, p=0.0009; IL-6: 73.1±4.2 vs 49.6±3.0 pg/ml; p=0.0051). The TEG data showed that the mice subjected to CLP displayed prolonged fibrin formation and fibrin cross-linkage time, slower clot formation, decreased platelet function, and clot rigidity. AGK2 treatment was associated with dramatic improvements in fibrin cross-linkage and clot formation times, without a significant impact on the clot initiation parameters or platelet function. Additionally, AGK2 significantly attenuated the bone marrow atrophy (58.3±6.5 vs 30.0±8.2%, p=0.0262). CONCLUSION: Selective inhibition of SIRT2 significantly improves survival, and attenuates sepsis-associated "cytokine storm", coagulopathy, and bone marrow atrophy in a mouse model of lethal septic shock.
Assuntos
Furanos/administração & dosagem , Inibidores de Histona Desacetilases/administração & dosagem , Quinolinas/administração & dosagem , Choque Séptico/tratamento farmacológico , Sirtuína 2/antagonistas & inibidores , Animais , Atrofia/prevenção & controle , Medula Óssea/efeitos dos fármacos , Medula Óssea/patologia , Células Cultivadas , Citocinas/sangue , Avaliação Pré-Clínica de Medicamentos , Injeções Intraperitoneais , Lipopolissacarídeos/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Choque Séptico/sangue , Choque Séptico/enzimologia , Choque Séptico/imunologia , Sirtuína 2/metabolismoRESUMO
Endotoxin shock induction in mice is a commonly used animal model to evaluate the protective effect of biologically active reagents. After an lipopolysaccharides (LPS) stimulus, matrix metalloproteinase-8 (MMP-8) and matrix metalloproteinase-9 (MMP-9) are rapidly degranulated and released by neutrophils, aside other enzymes and effector molecules. MMPs cleave extracellular matrix components and cytokines, and such processes contribute to shock syndrome development. CPU1 and CPU2 are two peptide MMP inhibitors with different in vitro IC50 values to several key enzymes, including MMP-8 and MMP-9. In vivo work confirmed that CPU1 and CPU2 protected mice from endotoxin shock after intravenous and intraperitoneal injections. Furthermore, their minimal effective dose after an intravenous injection and the maximum time interval between intraperitoneal peptide injection (150 mg/kg) and intravenous LPS injection were determined. With the use of an indirect competitive ELISA, plasma CPU1 and CPU2 concentrations in different experimental settings were measured. In addition, the acuteness of MMP-9 release in the mouse circulation after an intravenous LPS injection was confirmed with the zymography technique. Our findings reinforce previous work with other inhibitors about a strict time window within which effective MMP inhibition is needed to obtain significant survival rate improvements and also show that, with strict pharmacokinetic monitoring, potent protease inhibitors may in the future become life-savers in shock conditions.
Assuntos
Metaloproteinase 8 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Peptídeos/farmacologia , Choque Séptico/prevenção & controle , Animais , Modelos Animais de Doenças , Feminino , Injeções Intraperitoneais , Injeções Intravenosas , Lipopolissacarídeos , Inibidores de Metaloproteinases de Matriz/administração & dosagem , Inibidores de Metaloproteinases de Matriz/sangue , Inibidores de Metaloproteinases de Matriz/farmacocinética , Camundongos , Peptídeos/administração & dosagem , Peptídeos/sangue , Peptídeos/farmacocinética , Choque Séptico/sangue , Choque Séptico/induzido quimicamente , Choque Séptico/enzimologiaAssuntos
Arginase/metabolismo , Arginina/metabolismo , Complexo CD3/metabolismo , Neoplasias do Colo/cirurgia , Complicações Pós-Operatórias/etiologia , Choque Séptico/etiologia , Linfócitos T/imunologia , Citoplasma/efeitos dos fármacos , Humanos , Choque Séptico/enzimologia , Transdução de Sinais/efeitos dos fármacosRESUMO
We previously demonstrated that 3,4,5-trihydorxycinnamic acid (THC), a derivative of hydroxycinnamic acids, possesses protective effect in lipopolysaccharide (LPS)-induced endotoxemia models. However, the effects of THC in LPS-induced septic kidney are still unclear. Therefore, the present study was carried out to examine the effects of THC in LPS-challenged septic kidney using mesangial cell line and Balb/c mice. THC pretreatment effectively inhibited LPS-induced macrophage infiltration and the secretion of pro-inflammatory cytokines in the kidney of LPS-challenged animals. Pretreatment of rat mesangial cells with THC significantly attenuated LPS-induced PGE2 production and COX-2 expression. THC also significantly suppressed LPS-induced expression of MCP-1 in LPS-activated septic kidney and rat mesangial cells. In addition, THC significantly attenuated LPS-induced degradation of IκB-α in LPS-induced rat mesangial cells. THC also increased the expression of heme oxygenase-1 (HO-1) in LPS-challenged septic kidney and mesangial cells. Multiple signaling pathways including p38 and AKT have been observed to be involved in the THC-induced activation of HO-1 expression. The present data clearly demonstrate that THC protects LPS-challenged septic kidney by decreasing macrophage infiltration and increasing HO-1 expression, suggesting that THC might be a valuable therapeutic agent for compromised kidney in sepsis.
Assuntos
Cinamatos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/biossíntese , Lipopolissacarídeos/toxicidade , Macrófagos/enzimologia , Proteínas de Membrana/biossíntese , Células Mesangiais/enzimologia , Choque Séptico/enzimologia , Animais , Linhagem Celular , Ciclo-Oxigenase 2/biossíntese , Dinoprostona/metabolismo , Macrófagos/patologia , Células Mesangiais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Choque Séptico/induzido quimicamente , Choque Séptico/patologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
We have previously demonstrated that a stable synthetic analog of 20-hydroxyeicosatetraenoic acid (20-HETE), N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), prevents vascular hyporeactivity, hypotension, tachycardia, and inflammation in rats treated with lipopolysaccharide (LPS) and mortality in endotoxemic mice. These changes were attributed to decreased production of inducible nitric oxide (NO) synthase (iNOS)-derived NO, cyclooxygenase (COX)-2-derived vasodilator prostanoids, and proinflammatory mediators associated with increased cyctochrome P450 (CYP) 4A1-derived 20-HETE and CYP2C23-dependent antiinflammatory mediator formation. The aim of this study was to determine whether decreased expression and activity of iNOS, soluble guanylyl cyclase (sGC), protein kinase G (PKG), COX-2, gp91(phox) (NOX2; a superoxide generating NOX enzyme), and peroxynitrite production associated with increased expression of COX-1 and CYP4A1 and 20-HETE formation in renal and cardiovascular tissues of rats contributes to the effect of 5,14-HEDGE to prevent vasodilation, hypotension, tachycardia, and inflammation in response to systemic administration of LPS. Mean arterial pressure fell by 28mmHg and heart rate rose by 47beats/min in LPS (10mg/kg, i.p.)-treated rats. Administration of LPS also increased mRNA and protein expression of iNOS and COX-2 associated with a decrease in COX-1 and CYP4A1 mRNA and protein expression. Increased NOS activity, iNOS-heat shock protein 90 complex formation (an index for iNOS activity), protein expression of phosphorylated vasodilator stimulated phosphoprotein (an index for PKG activity), gp91(phox), p47(phox) (NOXO2; organizer subunit of gp91(phox)), and nitrotyrosine (an index for peroxynitrite production) as well as cGMP (an index for sGC activity), 6-keto-PGF1α (a stable metabolite PGI2) and PGE2 levels (indexes for COX activity), and nitrotyrosine levels by LPS were also associated with decreased CYP hydroxylase activity as measured by 20-HETE formation from arachidonic acid in renal microsomes of LPS-treated rats. These effects of LPS, except iNOS mRNA and COX-1 protein expression, were prevented by 5,14-HEDGE (30mg/kg, s.c.; 1h after LPS). A competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (30mg/kg, s.c.; 1h after LPS) reversed the effects of 5,14-HEDGE, except iNOS and COX-1 mRNA and protein expression as well as expression of CYP4A1 mRNA. These results suggest that increased CYP4A1 expression and 20-HETE formation associated with suppression of iNOS/sGC/PKG pathway, COX-2, and gp91(phox) participate in the protective effect of 5,14-HEDGE against vasodilation, hypotension, tachycardia, and inflammation in the rat model of septic shock.
Assuntos
Lipopeptídeos/farmacologia , Óxido Nítrico Sintase Tipo II/metabolismo , Substâncias Protetoras/farmacologia , Choque Séptico/tratamento farmacológico , Choque Séptico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Moléculas de Adesão Celular/metabolismo , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Citocromo P-450 CYP2J2 , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Modelos Animais de Doenças , Guanilato Ciclase/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Ácidos Hidroxieicosatetraenoicos/farmacologia , Lipopolissacarídeos/farmacologia , Masculino , Glicoproteínas de Membrana/metabolismo , Proteínas dos Microfilamentos/metabolismo , NADPH Oxidase 2 , NADPH Oxidases/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Especificidade de Órgãos , Ácido Peroxinitroso/metabolismo , Fosfoproteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Distribuição Aleatória , Ratos , Ratos Wistar , Receptores Citoplasmáticos e Nucleares/metabolismo , Choque Séptico/enzimologia , Choque Séptico/genética , Guanilil Ciclase SolúvelRESUMO
Acute sepsis can be induced by cytokines such as TNF-α and biological products such as LPS. All of these agents cause systemic inflammation, which is characterized by hemodynamic shock and liver toxicity. However, the outcomes of different septic shock models were totally opposite in transglutaminase 2 knockout (TGase 2(-/-)) mice. The aim of our study was to clarify the role of TGase 2 in liver injury. Therefore, we explored the role of TGase 2 in liver damage using two different stress models: LPS-induced endotoxic shock and TNF-α/actinomycin D (ActD)-induced sepsis. TNF-α-dependent septic shock resulted in increased liver damage in TGase 2(-/-) mice compared with wild-type (WT) mice, and was accompanied by increased levels of caspase 3 and cathepsin D (CTSD) in the damaged liver. Conversely, LPS-induced septic shock resulted in ablation of inflammatory endotoxic shock in TGase 2(-/-) mice and decreased liver injury. We found that TGase 2 protected liver tissue from TNF-α-dependent septic shock by reducing the expression of caspase 3 and CTSD. However, TGase 2 differently participated in increased the hemodynamic shock in LPS-induced septic shock through macrophage activation rather than protecting direct liver damage. Therefore, these findings demonstrate that septic shock caused by different agents may induce different results in TGase 2(-/-) mice depending on the primary target organs affected.
Assuntos
Proteínas de Ligação ao GTP/genética , Choque Séptico/imunologia , Transglutaminases/genética , Animais , Apoptose , Catepsina D/metabolismo , Células Cultivadas , Dactinomicina/farmacologia , Proteínas de Ligação ao GTP/deficiência , Galactosamina/farmacologia , Lipopolissacarídeos/farmacologia , Fígado/enzimologia , Fígado/imunologia , Fígado/patologia , Ativação de Macrófagos , Masculino , Camundongos , Camundongos Knockout , Proteína 2 Glutamina gama-Glutamiltransferase , Choque Séptico/enzimologia , Choque Séptico/patologia , Transglutaminases/deficiência , Fator de Necrose Tumoral alfa/farmacologia , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Immunostimulating staphylococcal enterotoxin B (SEB) and related superantigenic toxins cause diseases in humans and laboratory animals by activating cells of the immune system. These toxins bind directly to the major histocompatibility complex (MHC) class II molecules on antigen-presenting cells and specific Vβ regions of T-cell receptors (TCR), resulting in hyperactivation of both T lymphocytes and monocytes/macrophages. Activated host cells produce excessive amounts of proinflammatory cytokines and chemokines, especially tumor necrosis factor α, interleukin 1 (IL-1), IL-2, interferon γ (IFNγ), and macrophage chemoattractant protein 1 causing clinical symptoms of fever, hypotension, and shock. The well-explored signal transduction pathways for SEB-induced toxicity downstream from TCR/MHC ligation and interaction of cell surface co-stimulatory molecules include the mitogen-activated protein kinase cascades and cytokine receptor signaling, culminating in NFκB activation. Independently, IL-2, IFNγ, and chemokines from activated T cells signal via the phosphoinositide 3-kinase (PI3K), the serine/threonine kinases, Akt and mammalian target of rapamycin (mTOR) pathways. This article reviews the signaling molecules induced by superantigens in the activation of PI3K/Akt/mTOR pathways leading to staphylococcal superantigen-induced toxicity and updates potential therapeutics against superantigens.
Assuntos
Toxinas Bacterianas/toxicidade , Enterotoxinas/toxicidade , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Staphylococcus/química , Superantígenos/toxicidade , Serina-Treonina Quinases TOR/metabolismo , Animais , Anticorpos Monoclonais/uso terapêutico , Toxinas Bacterianas/imunologia , Citocinas/antagonistas & inibidores , Citocinas/biossíntese , Citocinas/imunologia , Enterotoxinas/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Ligação Proteica , Receptores de Antígenos de Linfócitos T/metabolismo , Choque Séptico/tratamento farmacológico , Choque Séptico/enzimologia , Choque Séptico/imunologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Superantígenos/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologiaRESUMO
BACKGROUND: Puerperal group A streptococcus (GAS) infection, once the leading cause of postpartum sepsis, has been increasing again since the 1980s. Streptococcal toxic shock syndrome (STSS) is a serious complication characterized by rapidly spreading GAS infection, shock, and multiple organ failure. Immediate recognition and implementation of therapy is crucial for survival. Making informed decisions regarding surgical debridement, namely hysterectomy, based on clinical indicators is difficult for practitioners. OBJECTIVES: This article discusses the potential role of creatine kinase in the decision-making process for treatment of STSS, particularly with regard to hysterectomy. MATERIAL AND METHODS: A case report is presented. The literature was searched using the key words 'group A streptococcus', 'postpartum hysterectomy', 'creatine kinase', 'endomyometritis', and 'streptococcal toxic shock syndrome' in PubMed and the UptoDate database. Relevant articles published between 1991 and 2011 were evaluated. CONCLUSION: Decisions regarding hysterectomy in STSS management are difficult. A rise in CK levels in the serum may indicate involvement of the myometrium and may be an important parameter in the difficult decision of hysterectomy when treating STSS.
Assuntos
Creatina Quinase/sangue , Endometrite/cirurgia , Histerectomia , Complicações Infecciosas na Gravidez , Choque Séptico/cirurgia , Infecções Estreptocócicas/cirurgia , Streptococcus pyogenes/isolamento & purificação , Adulto , Antibacterianos/uso terapêutico , Biomarcadores/sangue , Terapia Combinada , Tomada de Decisões , Endometrite/enzimologia , Endometrite/microbiologia , Feminino , Humanos , Recém-Nascido , Doenças do Recém-Nascido/enzimologia , Doenças do Recém-Nascido/microbiologia , Laparoscopia , Masculino , Período Pós-Parto , Gravidez , Respiração Artificial , Choque Séptico/enzimologia , Choque Séptico/microbiologia , Infecções Estreptocócicas/enzimologia , Infecções Estreptocócicas/microbiologiaRESUMO
Histone deacetylases (HDACs) play a key role in homeostasis of protein acetylation in histone and nonhistone proteins and in regulating fundamental cellular activities. In this paper we review and discuss intriguing recent developments in the use of histone deacetylase inhibitors (HDACIs) to combat some critical conditions in an animal model of hemorrhagic and septic shock. HDACIs have neuroprotective, cardioprotective, renal-protective, and anti-inflammatory properties; survival improvements have been significantly shown in these models. We discuss the targets and mechanisms underlying these effects of HDACIs and comment on the potential new clinical applications for these agents in the future. This paper highlights the emerging roles of HDACIs as acetylation modulators in models of hemorrhagic and septic shock and explains some contradictions encountered in previous studies.
Assuntos
Inibidores de Histona Desacetilases/uso terapêutico , Histona Desacetilases/metabolismo , Choque Hemorrágico/tratamento farmacológico , Choque Séptico/tratamento farmacológico , Acetilação/efeitos dos fármacos , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Modelos Animais de Doenças , Epigenômica , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Histonas/metabolismo , Humanos , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Fosfatidilinositol 3-Quinases/efeitos dos fármacos , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Choque Hemorrágico/enzimologia , Choque Séptico/enzimologia , SuínosRESUMO
OBJECTIVE: Increasing evidence points to the role of mitochondrial dysfunction in the pathogenesis of sepsis. Previous data indicate that mitochondrial function is affected in monocytes from septic patients, but the underlying mechanisms and the impact of these changes on the patients' outcome are unknown. We aimed to determine the mechanisms involved in mitochondrial dysfunction in peripheral blood mononuclear cells from patients with septic shock. DESIGN: A cohort of patients with septic shock to study peripheral blood mononuclear cell mitochondrial respiration by high-resolution respirometry analyses and to compare with cells from control subjects. SETTING: Three intensive care units and an academic research laboratory. SUBJECTS: Twenty patients with septic shock and a control group composed of 18 postoperative patients without sepsis or shock. INTERVENTIONS: Ex vivo measurements of mitochondrial oxygen consumption were carried out in digitonin-permeabilized peripheral blood mononuclear cells from 20 patients with septic shock taken during the first 48 hrs after intensive care unit admission as well as in peripheral blood mononuclear cells from control subjects. Clinical parameters such as hospital outcome and sepsis severity were also analyzed and the relationship between these parameters and the oxygen consumption pattern was investigated. MEASUREMENTS AND MAIN RESULTS: We observed a significant reduction in the respiration specifically associated with adenosine-5'-triphosphate synthesis (state 3) compared with the control group (5.60 vs. 9.89 nmol O2/min/10(7) cells, respectively, p < .01). Reduction of state 3 respiration in patients with septic shock was seen with increased prevalence of organ failure (r = -0.46, p = .005). Nonsurviving patients with septic shock presented significantly lower adenosine diphosphate-stimulated respiration when compared with the control group (4.56 vs. 10.27 nmol O2/min/10(7) cells, respectively; p = .004). Finally, the presence of the functional F1Fo adenosine-5'-triphosphate synthase complex (0.51 vs. 1.00 ng oligo/mL/10(6) cells, p = .02), but not the adenine nucleotide translocator, was significantly lower in patients with septic shock compared with control cells. CONCLUSION: Mitochondrial dysfunction is present in immune cells from patients with septic shock and is characterized as a reduced respiration associated to adenosine-5'-triphosphate synthesis. The molecular basis of this phenotype involve a reduction of F1Fo adenosine-5'-triphosphate synthase activity, which may contribute to the energetic failure found in sepsis.
Assuntos
Leucócitos Mononucleares/citologia , Mitocôndrias/enzimologia , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Choque Séptico/sangue , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Estudos de Casos e Controles , Células Cultivadas , Metabolismo Energético , Feminino , Humanos , Unidades de Terapia Intensiva , Leucócitos Mononucleares/fisiologia , Masculino , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Monócitos/citologia , Monócitos/fisiologia , Valores de Referência , Choque Séptico/enzimologiaRESUMO
INTRODUCTION: Circulating lipopolysaccharides released from bacteria may activate both neutrophils and monocytes. The activated neutrophils release myeloperoxidase (MPO), a specific enzyme with strong oxidative activity. The aim of this study was to evaluate MPO enzyme activity in plasma of critically ill patients and to check the hypothesis that these concentrations in plasma would be higher in sepsis and systemic inflammatory conditions, as neutrophils release their contents before proliferating in response to stress. MATERIAL AND METHODS: Blood samples were collected from 105 critically ill patients admitted to the intensive care unit, consisting of those with systemic inflammatory response syndrome (n = 42), sepsis (n = 37), and septic shock (n = 26). Plasma MPO enzyme activity was determined by o-dianisidine-H(2)O(2) method, modified for 96-well plates. RESULTS: The plasma MPO enzyme activity in sepsis patients was significantly higher than that in the control group (mean, 2.4 ± 1.8 in sepsis and 1.86 ± 1.2 nmol per milligram protein per 10 minutes in systemic inflammatory response syndrome vs 0.32 ± 0.11 nmol per milligram protein per 10 minutes in healthy controls). Mean plasma lactate levels in sepsis (7.8 ± 1.2 mmol/L) and shock patients (9.5 ± 1.2 mmol/L) and cytokines like tumor necrosis factor-α, interleukin-8, and interleukin-1ß were simultaneously evaluated to establish onset of inflammation and sepsis. These results show that neutrophil activation occurring during inflammation and sepsis could be detected by plasma MPO concentration. CONCLUSION: The plasma MPO concentrations may be a marker of the neutrophil proliferation and severity of inflammation.
Assuntos
Estado Terminal , Peroxidase/sangue , Sepse/enzimologia , Síndrome de Resposta Inflamatória Sistêmica/enzimologia , Adulto , Biomarcadores/sangue , Feminino , Humanos , Unidades de Terapia Intensiva , Masculino , Neutrófilos/enzimologia , Choque Séptico/enzimologia , Estatísticas não ParamétricasRESUMO
G-protein-coupled receptor kinase 2 (GRK2) is a member of a kinase family originally discovered for its role in the phosphorylation and desensitization of G-protein-coupled receptors. It is expressed in high levels in myeloid cells and its levels are altered in many inflammatory disorders including sepsis. To address the physiological role of myeloid cell-specific GRK2 in inflammation, we generated mice bearing GRK2 deletion in myeloid cells (GRK2âµmye). GRK2âµmye mice exhibited exaggerated inflammatory cytokine/chemokine production, and organ injury in response to lipopolysaccharide (LPS, a TLR4 ligand) when compared to wild-type littermates (GRK2fl/fl). Consistent with this, peritoneal macrophages from GRK2âµmye mice showed enhanced inflammatory cytokine levels when stimulated with LPS. Our results further identify TLR4-induced NF-κB1p105-ERK pathway to be selectively regulated by GRK2. LPS-induced activation of NF-κB1p105-MEK-ERK pathway is significantly enhanced in the GRK2âµmye macrophages compared to GRK2fl/fl cells and importantly, inhibition of the p105 and ERK pathways in the GRK2âµmye macrophages, limits the enhanced production of LPS-induced cytokines/chemokines. Taken together, our studies reveal previously undescribed negative regulatory role for GRK2 in TLR4-induced p105-ERK pathway as well as in the consequent inflammatory cytokine/chemokine production and endotoxemia in mice.
Assuntos
Endotoxemia/enzimologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Sistema de Sinalização das MAP Quinases , Células Mieloides/enzimologia , Subunidade p50 de NF-kappa B/metabolismo , Choque Séptico/enzimologia , Animais , Células Cultivadas , Quimiocina CCL3/metabolismo , Endotoxemia/complicações , Endotoxemia/patologia , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Inflamação/complicações , Inflamação/enzimologia , Inflamação/patologia , Interleucina-10/metabolismo , Subunidade p40 da Interleucina-12/metabolismo , Lipopolissacarídeos/farmacologia , MAP Quinase Quinase Quinases/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Modelos Biológicos , Células Mieloides/efeitos dos fármacos , Células Mieloides/patologia , Subunidade p50 de NF-kappa B/antagonistas & inibidores , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Especificidade de Órgãos/efeitos dos fármacos , Choque Séptico/complicações , Choque Séptico/patologia , Análise de SobrevidaRESUMO
In this study, we investigated the protective effects of ligustilide against lipopolysaccharide (LPS)-induced endotoxic shock in Japanese white rabbits and attempted to elucidate the possible mechanism underlying these effects. Forty-two rabbits were randomized into 6 groups: normal group, LPS group, dexamethasone group (5 mg/kg), and 3 ligustilide groups (20, 40, and 80 mg/kg). After the rabbits had received a LPS infusion (0.3 mg/kg), dexamethasone and ligustilide were intravenously injected at the above-mentioned dosages. Heart rate (HR), mean arterial pressure (MAP), and rectal temperature (RT) were recorded throughout the experiment. Tumor necrosis factor- α (TNF- α), interleukin-1 ß (IL-1 ß), and nitric oxide (NO) levels were measured by radioimmunoassay every 30 minutes for the first hour and every 60 minutes thereafter until the end of the experiment. The serum levels of alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), γ-glutamyl transpeptidase (GGT), creatinine kinase (CK), lactate dehydrogenase (LDH), total protein (TP), creatinine (Scr), blood urea nitrogen (BUN), total bilirubin (T. BIL), and counts of formed elements of blood were measured at 0, 120, and 300 minutes after the administration of LPS. Hemorheology was assayed 300 minutes after the LPS injection. The vital organs were collected and weighed before histopathologic examination. A comparison between the LPS group and the ligustilide groups showed that ligustilide significantly inhibited the decline in MAP and RT and decreased the levels of TNF- α, IL-1 ß, and NO, but had no apparent effect on HR. Ligustilide also inhibited the increase in the levels of biochemical markers, such as ALT, AST, ALP, GGT, LDH, CK, BUN, and Scr, but showed no apparent effect on T. BIL and TP. Furthermore, ligustilide partly restored the function of injured vital organs, including the heart, liver, lungs, and kidneys. These results suggest that ligustilide protected the rabbits against LPS-induced endotoxic shock.
Assuntos
4-Butirolactona/análogos & derivados , Choque Séptico/prevenção & controle , 4-Butirolactona/farmacologia , Angelica sinensis/química , Animais , Bilirrubina/sangue , Pressão Sanguínea/efeitos dos fármacos , Proteínas Sanguíneas/química , Nitrogênio da Ureia Sanguínea , Creatina/sangue , Dexametasona/farmacologia , Relação Dose-Resposta a Droga , Feminino , Interleucina-1beta/sangue , Lipopolissacarídeos , Masculino , Óxido Nítrico/sangue , Coelhos , Choque Séptico/sangue , Choque Séptico/tratamento farmacológico , Choque Séptico/enzimologia , Fator de Necrose Tumoral alfa/sangueRESUMO
BACKGROUND: Despite global efforts to improve the treatment of sepsis, it remains a leading cause of morbidity and mortality in intensive care units. We have previously shown that suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, markedly improves survival in a murine model of lipopolysaccharide (LPS)-induced shock. SAHA has anti-inflammatory properties that have not been fully characterized. The liver plays an important role in the production of acute phase reactants involved in the inflammatory cascade and is also one of the major organs that can become dysfunctional in septic shock. The purpose of this study was to assess the effect of SAHA treatment on MAP kinases and associated inflammatory markers in murine liver after LPS-induced injury. METHODS: C57B1/6J mice were randomly divided into three groups: (A) experimental-given intraperitoneal (i.p.) SAHA (50 mg/kg) in dimethyl sulfoxide (DMSO) vehicle solution (n = 12); (B) control- given vehicle only (n = 12), and; (C) sham-given no treatment (n = 7). Two hours later, experimental and control mice were injected with LPS (20 mg/kg, i.p.) and experimental mice received a second dose of SAHA. Livers were harvested at 3, 24, and 48 h for analysis of inflammatory markers using Western Blot, Polymerase Chain Reaction (PCR), and Enzyme-Linked Immunosorbent Assay (ELISA) techniques. RESULTS: After 3 h, the livers of animals treated with SAHA showed significantly (P < 0.05) decreased expression of the pro-inflammatory MAP kinases phosphorylated p38, phosphorylated ERK, myeloperoxidase and interleukin-6, and increased levels of the anti-inflammatory interleukin-10 compared with controls. Phospho-p38 expression remained low in the SAHA treated groups at 24 and 48 h. CONCLUSION: Administration of SAHA is associated with attenuation of MAPK activation and alteration of inflammatory and anti-inflammatory markers in murine liver after a lethal LPS insult. The suppression of MAPK activity is rapid (within 3 h), and is sustained for up to 48 h post-treatment. These results may in part account for the improvement in survival shown in this model.