RESUMEN
The prevalence of serum antibodies against Clostridium difficile (CD) toxins A and B in healthy populations have prompted interest in evaluating the therapeutic activity of intravenous immunoglobulin (IVIg) in individuals experiencing severe or recurrent C. difficile infection (CDI). Despite some promising case reports, a definitive clinical role for IVIg in CDI remains unclear. Contradictory results may be attributed to a lack of consensus regarding optimal dose, timing of administration and patient selection as well as variability in specific antibody content between commercial preparations. The purpose of this study was to investigate retrospectively the efficacy of three commercial preparations of IVIg for treating severe or recurrent CDI. In subsequent mechanistic studies using protein microarray and toxin neutralization assays, all IVIg preparations were analysed for specific binding and neutralizing antibodies (NAb) to CD antigens in vitro and the presence of anti-toxin NAbs in vivo following IVIg infusion. A therapeutic response to IVIg was observed in 41% (10 of 17) of the CDI patients. Significant variability in multi-isotype specific antibodies to a 7-plex panel of CD antigens and toxin neutralization efficacies were observed between IVIg preparations and also in patient sera before and after IVIg administration. These results extend our current understanding of population immunity to CD and support the inclusion of surface layer proteins and binary toxin antigens in CD vaccines. Future strategies could enhance IVIg treatment response rates by using protein microarray to preselect donor plasma/serum with the highest levels of anti-CD antibodies and/or anti-toxin neutralizing capacities prior to fractionation.
Asunto(s)
Anticuerpos Antibacterianos/uso terapéutico , Anticuerpos Neutralizantes/uso terapéutico , Toxinas Bacterianas/inmunología , Enterocolitis Seudomembranosa/terapia , Inmunoglobulinas Intravenosas/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Células CACO-2 , Clostridioides difficile , Humanos , Persona de Mediana Edad , Estudios Retrospectivos , Reino UnidoRESUMEN
An in-depth understanding of biochemical processes occurring within biological systems is key for early diagnosis of disease and identification of appropriate treatments. Nanobiophotonics offers huge potential benefits for intracellular diagnostics and therapeutics. Intracellular sensing using fluorescent nanoparticles is a potentially useful tool for real-time, in vivo monitoring of important cellular analytes. This work is focused on synthesis of optical chemical nanosensors for the quantitative analysis of pH inside living cells. The structure of the nanosensor comprises a biofriendly silica matrix with co-encapsulated Texas Red, acting as a reference dye, and pH-sensitive fluorescein isothiocyanate enabling ratiometric quantitative environmental detection. In order to obtain silica-based nanoparticles -70 nm in size, a modified sol-gel-based Stöber method was employed. The potential of these nanosensors for intracellular pH monitoring is demonstrated inside a live human embryonic kidney cell line whereby a significant change in fluorescence is observed when the cell pH is switched from acidic to basic. High loading efficiencies of nanoparticles into the cells is seen, with little effect on cell morphology even following extended nanoparticle exposure (up to 72 h). Nanoparticle incubation time and the fast response of the nanosensor (-2 s) make it a very powerful tool in monitoring the processes occurring within the cytosol.
Asunto(s)
Técnicas Biosensibles/métodos , Espacio Intracelular/metabolismo , Nanopartículas/química , Dióxido de Silicio/química , Calibración , Muerte Celular , Células HEK293 , Humanos , Concentración de Iones de Hidrógeno , Luz , Microscopía Fluorescente , Nanopartículas/ultraestructura , Dispersión de Radiación , Espectrometría de Fluorescencia , Factores de Tiempo , Receptor Toll-Like 4/metabolismoRESUMEN
Conjugated linoleic acids (CLA) affect atherogenesis, but mechanisms are not well understood. We explored how two isomers of CLA, cis9, trans11-CLA and trans10, cis12-CLA, affected lipid and glucose metabolism, as well as hepatic protein expression, in apolipoprotein E knockout mice. After 12 wk of intervention, plasma triglyceride, NEFA, and glucose concentrations were significantly higher in the trans10, cis12-CLA group, whereas plasma triglyceride, NEFA, glucose, and insulin concentrations were significantly lower in the cis9, trans11-CLA group, compared with control mice consuming linoleic acid. Proteomics identified significant up- or down-regulation of 113 liver cytosolic proteins by either CLA isomer. Principal component analysis revealed that the treatment effect of cis9, trans11-CLA was mainly explained by the up-regulation of different posttranslational forms of heat shock protein 70 kD. In contrast, the treatment effect of trans10, cis12-CLA was mainly explained by up-regulation of key enzymes in the gluconeogenic, beta-oxidation, and ketogenesic pathways. Correlation analysis again emphasized the divergent effects of both CLA isomers on different pathways, but also revealed a linkage between insulin resistance and increased levels of hepatic serotransferrin. Thus, our systems biology approach provided novel insights into the mechanisms by which individual CLA isomers differentially affect pathways related to atherogenesis, such as insulin resistance and inflammation.
Asunto(s)
Apolipoproteínas E/genética , Ácido Linoleico/química , Ácidos Linoleicos Conjugados/metabolismo , Proteómica/métodos , Alimentación Animal , Animales , Aterosclerosis/patología , Glucemia/metabolismo , Western Blotting , Composición Corporal , Peso Corporal , Citosol/metabolismo , Dieta , Ácidos Grasos/metabolismo , Ligamiento Genético , Glucosa/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Inflamación , Insulina/metabolismo , Resistencia a la Insulina , Hígado/metabolismo , Masculino , Ratones , Ratones Noqueados , Oxígeno/metabolismo , Perfusión , Análisis de Componente Principal , Biología de Sistemas , Triglicéridos/metabolismoRESUMEN
Immunization with the whole-cell pertussis vaccine (Pw), while effective at preventing whooping cough in infants, has been associated with local, systemic, and neuronal reactions, including fevers and convulsions in children. In contrast, the new acellular pertussis vaccines (Pa) have a considerably improved safety profile. The lack of an appropriate animal model has restricted investigations into the mechanisms by which neurological reactions are induced by vaccination. Here we describe a novel murine model wherein seizure-like behavioral changes are induced following parenteral administration of Pw. The proinflammatory cytokine interleukin-beta (IL-1beta), production of which has been associated with many neurodegenerative conditions, was significantly increased in the hippocampus and hypothalamus of vaccinated animals. Accompanying this change was a decrease in release of the inhibitory neurotransmitters gamma-aminobutyric acid and adenosine in the hippocampus. Seizure-like behavioral changes were significantly reduced following inhibition of IL-1beta production by the administration of an inhibitor of IL-1beta-converting enzyme and were almost completely abrogated in IL-1 receptor type I knockout mice. These results suggest a causal relationship between IL-1beta induction and convulsive behavior following Pw vaccination. Significantly, Pa neither increased IL-1beta nor induced behavioral changes in mice, but did induce the anti-inflammatory cytokine IL-10. In contrast, administration of active pertussis toxin and lipopolysaccharide, residual in Pw but absent from Pa, also induced convulsive activity. Our findings provide the first direct evidence of an immunological basis for pertussis vaccine reactogenicity and suggest that active bacterial toxins are responsible for the neurologic disturbances observed in children immunized with Pw.
Asunto(s)
Bordetella pertussis/inmunología , Interleucina-1/inmunología , Neuronas/inmunología , Vacuna contra la Tos Ferina/inmunología , Animales , Encéfalo/inmunología , Modelos Animales de Enfermedad , Femenino , Fiebre/etiología , Hipocampo/inmunología , Interleucina-1/biosíntesis , Interleucina-1/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Convulsiones/inmunología , Vacunación , Vacunas Acelulares/inmunología , Tos Ferina/prevención & control , Ácido gamma-Aminobutírico/metabolismoRESUMEN
Neurological side effects are a major cause of concern following immunization with a number of vaccines, especially the whole cell pertussis vaccine (Pw). In this study we report that IL-1beta concentrations were significantly increased in the hippocampus following subcutaneous (s.c.) injection of Pw, and that this was accompanied by increased activity of the stress-activated kinase, c-Jun-N-terminal kinase (JNK) and a decrease in glutamate release. These effects were mimicked by s.c injection of active pertussis toxin (PT) or lipopolysaccharide (LPS). Incubation of hippocampal synaptosomes in the presence of Pw, PT or LPS also resulted in increased JNK activation and decreased glutamate release, effects which were mimicked by IL-1beta and blocked by the IL-1 receptor antagonist (IL-ra). Our observations are consistent with the hypothesis that IL-1beta induced by active bacterial toxins present in vaccine preparations, mediate the neurochemical and perhaps the neurological effects of Pw.
Asunto(s)
Hipocampo/inmunología , Hipocampo/metabolismo , Interleucina-1/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos , Vacuna contra la Tos Ferina/inmunología , Animales , Química Encefálica/inmunología , Femenino , Expresión Génica/inmunología , Ácido Glutámico/metabolismo , Inmunización , Técnicas In Vitro , Proteína Antagonista del Receptor de Interleucina 1 , Interleucina-1/genética , Lipopolisacáridos/inmunología , Lipopolisacáridos/farmacología , MAP Quinasa Quinasa 4 , Ratones , Ratones Endogámicos BALB C , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Toxina del Pertussis , Fosforilación , ARN Mensajero/análisis , Sialoglicoproteínas/farmacología , Sinaptosomas/enzimología , Sinaptosomas/inmunología , Factores de Virulencia de Bordetella/inmunología , Factores de Virulencia de Bordetella/farmacologíaRESUMEN
Parenteral injection of endotoxin has been used as a model to examine the role of pro-inflammatory cytokines in the centrally controlled responses to Gram-negative bacterial infection. However, the events that occur following mucosal exposure to live bacteria have received little attention. In this study, we have used a murine model to demonstrate that respiratory infection with Bordetella pertussis, which is associated with a number of systemic complications including fever, seizure and encephalopathy in children, resulted in persistent expression of mRNA transcripts for IL-1beta and TNFalpha and transient expression of IL-6 in the hippocampus and hypothalamus. These changes correlated with elevated levels of cytokine protein in the same brain areas. The results demonstrate that infection at a mucosal surface can result in the induction of pro-inflammatory cytokine production in the brain and suggest that these locally synthesized mediators may contribute to the centrally controlled clinical manifestations of B. pertussis infection.