RESUMO
Sepsis-associated encephalopathy manifesting as delirium is a common problem in critical care medicine. In this study, patients that had delirium due to sepsis had significant cognitive impairments at 12-18 months after hospital discharge when compared with controls and Cambridge Neuropsychological Automated Test Battery-standardized scores in spatial recognition memory, pattern recognition memory, and delayed-matching-to-sample tests but not other cognitive functions. A mouse model of S. pneumoniae pneumonia-induced sepsis, which modeled numerous aspects of the human sepsis-associated multiorgan dysfunction, including encephalopathy, also revealed similar deficits in spatial memory but not new task learning. Both humans and mice had large increases in chemokines for myeloid cell recruitment. Intravital imaging of the brains of septic mice revealed increased neutrophil and CCR2+ inflammatory monocyte recruitment (the latter being far more robust), accompanied by subtle microglial activation. Prevention of CCR2+ inflammatory monocyte recruitment, but not neutrophil recruitment, reduced microglial activation and other signs of neuroinflammation and prevented all signs of cognitive impairment after infection. Therefore, therapeutically targeting CCR2+ inflammatory monocytes at the time of sepsis may provide a novel neuroprotective clinical intervention to prevent the development of persistent cognitive impairments.
Assuntos
Encéfalo/patologia , Disfunção Cognitiva/patologia , Citocinas/sangue , Inflamação/sangue , Monócitos/patologia , Encefalopatia Associada a Sepse/patologia , Adulto , Idoso , Animais , Anticorpos Monoclonais/uso terapêutico , Disfunção Cognitiva/microbiologia , Modelos Animais de Doenças , Feminino , Humanos , Inflamação/microbiologia , Interleucina-8/antagonistas & inibidores , Interleucina-8/imunologia , Microscopia Intravital , Masculino , Testes de Estado Mental e Demência , Camundongos , Microglia/fisiologia , Pessoa de Meia-Idade , Monócitos/metabolismo , Neutrófilos/patologia , Infecções Pneumocócicas/complicações , Receptores CCR2/antagonistas & inibidores , Receptores CCR2/imunologia , Receptores CCR2/metabolismo , Encefalopatia Associada a Sepse/sangue , Encefalopatia Associada a Sepse/microbiologiaRESUMO
BACKGROUND: Intravenous immunoglobulin (IVIg) therapy has been shown to have therapeutic benefit in more than 50 inflammatory and immune-related diseases; however, the potential benefit of IVIg in cardiovascular disease is more limited, in part because our understanding of the mechanisms underlying the effects of IVIg in innate immunity is incomplete. METHODS AND RESULTS: In this study, a systematic assessment of the role of IVIg in leukocyte recruitment was completed with an in vitro flow-chamber system and in vivo intravital microscopy in a feline ischemia-reperfusion model system. IVIg treatment of blood resulted in a profound decrease in recruitment of either immobilized P-selectin or E-selectin due to direct effects of IVIg on the leukocyte (not substratum). Similar results were observed on endothelium treated with histamine, which induces P-selectin-dependent rolling and beta2-integrin-dependent adhesion. IVIg reduced P-selectin glycoprotein ligand-1 (PSGL-1) antibody binding to PSGL-1 on leukocytes. Use of a beta2-integrin-dependent static assay to bypass selectin-dependent recruitment revealed some inhibitory effectiveness (60%), which suggests that the majority of the effects of IVIg were due to selectin inhibition, with some inhibition of integrin function. In vivo intravital microscopy revealed a potent inhibitory effect of IVIg on P-selectin-dependent rolling and beta2-integrin-dependent adhesion that led to reduced leukocyte recruitment and vascular dysfunction in postischemic microvessels. CONCLUSIONS: Our data demonstrate that IVIg has direct inhibitory effects on leukocyte recruitment in vitro and in vivo through inhibition of selectin and integrin function.
Assuntos
Moléculas de Adesão Celular/metabolismo , Imunoglobulinas Intravenosas/farmacologia , Traumatismo por Reperfusão/fisiopatologia , Animais , Antígenos CD18/metabolismo , Gatos , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Selectina E/metabolismo , Humanos , Migração e Rolagem de Leucócitos/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Selectina-P/metabolismo , Doenças Vasculares/prevenção & controleRESUMO
In the present study, we investigated whether inhaled nitric oxide (NO) was transported by plasma proteins, such as S-nitroso-albumin (SNO-Alb), in the feline circulation and whether this molecule delivers NO to the periphery under conditions of stress, specifically ischemia/reperfusion (I/R). A flow probe was interposed between the femoral and superior mesenteric artery for blood flow measurements, and a branch of the superior mesenteric vein was cannulated for arterial-venous sampling. In animals breathing room air, SNO-Alb was below detection level in arterial or venous blood. NO inhalation resulted in a significant arterial-venous gradient for SNO-Alb. Concomitant with this loss of SNO-Alb across the intestinal vasculature was an increase in nitrite (NO2-). However, this release of NO was not sufficient to alter intestinal blood flow. I/R during NO inhalation caused a very large increase in arterial SNO-Alb that permitted a 5-fold increase in SNO-Alb consumption and significant generation of NO2- within the postischemic intestinal vasculature. The increased SNO-Alb consumption was sufficient to dramatically improve intestinal blood flow. The very large burst of arterial SNO-Alb during I/R was completely blocked by the administration of superoxide dismutase, suggesting that oxidative stress contributed to the increased SNO-Alb formation. Our data suggest that inhaled NO can increase nitrosothiol production and these molecules may be a functional NO delivery system during cardiovascular disease.
Assuntos
Intestinos/irrigação sanguínea , Isquemia/metabolismo , Óxido Nítrico/farmacocinética , Soroalbumina Bovina/biossíntese , Administração por Inalação , Animais , Transporte Biológico , Gatos , Constrição , Artéria Mesentérica Superior , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/administração & dosagem , Doadores de Óxido Nítrico/farmacologia , Nitritos/sangue , Compostos Nitrosos , Oxirredução , Estresse Oxidativo , Proteínas Recombinantes de Fusão/farmacologia , Reperfusão , S-Nitrosoglutationa/farmacologia , Circulação Esplâncnica/efeitos dos fármacos , Superóxido Dismutase/farmacologia , Superóxidos/sangue , VasodilataçãoRESUMO
Patients with HIV infection exhibit deficits in bacterial and fungal clearance, and possibly depressed innate immunity. In this study, we observed that neutrophils from HIV-infected patients have a profound defect in chemotaxis in response to endogenous (IL-8) and bacterial (fMLP) chemoattractants, which was directly correlated with peripheral CD4(+) lymphocyte levels but not plasma viral load. A similar chemotactic defect was observed in the feline immunodeficiency virus (FIV) model of HIV infection. Intravital microscopy of FIV-infected animals revealed marked impairment in the in vivo recruitment of leukocytes; specifically integrin-dependent neutrophil adhesion and emigration induced by bacterial products. Treatment of FIV-infected animals with GM-CSF re-established both neutrophil recruitment (rolling, adhesion, and emigration) and in vitro chemotaxis to the levels seen in uninfected animals. This restoration of neutrophil responses was not due to GM-CSF-mediated priming. Rather, HIV and FIV infections resulted in defective neutrophil development, with an ensuing reduction in neutrophil granularity and chemotactic receptor expression. GM-CSF therapy restored neutrophil granularity, implying restoration of normal neutrophil development. Together, our findings underscore the fundamental defects in innate immunity caused by lentivirus infections, while also indicating that GM-CSF may be a potential immunorestorative therapy for HIV-infected patients.
Assuntos
Quimiotaxia de Leucócito , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Infecções por HIV/imunologia , Neutrófilos/imunologia , Animais , Gatos , Quimiotaxia de Leucócito/efeitos dos fármacos , Apresentação Cruzada , Grânulos Citoplasmáticos/ultraestrutura , Fator Estimulador de Colônias de Granulócitos e Macrófagos/uso terapêutico , Infecções por HIV/tratamento farmacológico , Humanos , Vírus da Imunodeficiência Felina , Interleucina-8/farmacologia , Infecções por Lentivirus/tratamento farmacológico , Infecções por Lentivirus/imunologia , Infecções por Lentivirus/veterinária , Lipopolissacarídeos/imunologia , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/ultraestruturaRESUMO
After an ischemic episode, reperfusion causes profound oxidative stress in the vasculature of the afflicted tissue/organ. The dysregulated accumulation of reactive oxygen species (ROS), such as superoxide, has been closely linked to the production and release of proinflammatory mediators, a profound increase in adhesion molecule expression by the vascular endothelium, and infiltration of neutrophils during ischemia-reperfusion (I/R). Superoxide dismutase (SOD) has been shown to protect tissues and organs against I/R-induced injury; however, the drug had to be continuously perfused or kidneys had to be occluded to prevent clearance. We used intravital microscopy, a system that allowed us to visualize neutrophil-endothelial interactions within the mesenteric postcapillary venules of cats subjected to I/R and tested the hypothesis that I/R-induced neutrophil recruitment was inhibited by treatment with SOD2/3. SOD2/3 is a chimeric fusion gene product that contains the mature SOD2 as well as the COOH-terminal "tail" of SOD3 and, unlike the three naturally occurring SODs (SOD1, SOD2, and SOD3), which bear a net negative charge at pH 7.4, SOD2/3 is positively charged and physiologically stable. Our results suggest that not only does SOD2/3 have a much greater efficacy in vivo than the native human SOD2, but its administration prevents I/R-induced neutrophil-endothelial cell interactions and microvascular dysfunction. Moreover, our data support the hypothesis that reactive oxidants mediate I/R-induced injury and that the chimeric recombinant SOD2/3 has the potential to be a therapeutic agent against this debilitating illness.
Assuntos
Vasos Sanguíneos/fisiopatologia , Endotélio Vascular/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Proteínas Recombinantes de Fusão/farmacologia , Traumatismo por Reperfusão/fisiopatologia , Superóxido Dismutase/genética , Superóxido Dismutase/farmacologia , Animais , Permeabilidade Capilar/efeitos dos fármacos , Gatos , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Endotélio Vascular/patologia , Hemodinâmica/efeitos dos fármacos , Hemodinâmica/fisiologia , Humanos , Cinética , Infiltração de Neutrófilos/fisiologia , Proteínas Recombinantes/farmacologia , Traumatismo por Reperfusão/patologiaRESUMO
Evidence indicates that the lentivirus, HIV, infection affects neutrophil response to bacteria and bacterial products in vitro. We used a novel model of rapid onset immunosuppression following infection with a similar lentivirus, feline immunodeficiency virus (FIV), in cats to examine neutrophil function within the microvasculature in vivo and to determine the steps that are impaired in the neutrophil recruitment cascade. In uninfected cats and cats infected neonatally with FIV, the mesentery was exteriorized, but remained autoperfused during intravital microscopy for 4 h. When the tissue was superfused with 10 micro g/ml of LPS for 4 h, intravital microscopy displayed a profound increase in neutrophil rolling at both 8 and 12 wk of age in uninfected cats. At 12 wk of age, FIV-infected animals showed a profound decrease in the number of rolling neutrophils. In vitro studies revealed that neutrophils from infected and uninfected animals rolled equally well on surrogate selectin substrata. In addition, in vivo neutrophil adhesion and emigration out of the vasculature were severely reduced, and in vitro neutrophil chemotaxis from FIV-infected animals was significantly impaired in response to fMLP or IL-8. However, FIV infection of neutrophils could not be detected. In summary, in vivo lentivirus infection with immunosuppression leads to a severe impairment in neutrophil rolling, adhesion, and emigration in response to bacterial stimulants potentially involving both endothelial and neutrophil dysfunction. These in vivo studies also indicate that neutrophil dysfunction should be taken into account when treating infections and tissue injury.