RESUMEN
In order to discuss first experiences with the implementation of the EU Regulation on In Vitro Diagnostic Medical Devices (IVDR) about one year after its entry into force, the German Association of the Scientific Medical Societies (AWMF e.V.) organized a full-day public webinar. Overall, it became clear that the implementation of the IVDR still poses significant challenges for laboratory medicine and pathology. Corrections at the political level and implementation with a sense of proportion are required. Before the long-term goal of the IVDR, i.e. the increase in patient safety, can be realized, the prevention of disadvantages for patients due to gaps in care must be strived for in the medium term by all parties involved.
Asunto(s)
Medicina , Humanos , Sociedades MédicasRESUMEN
A hallmark of T. gondii infections is passage of parasites across restrictive biological barriers--intestine, blood-brain barrier, blood-retina barrier and placenta-during primary infection or reactivation of chronic disease. Traversal of cellular barriers permits rapid dissemination of parasites to gain access to biologically restricted organs. This process involves active parasite motility and tightly regulated interactions between host cell receptors and parasite adhesins that facilitate paracellular transfer. Mounting evidence also suggests that parasites use migrating leukocytes as Trojan horses to disseminate in the organism while avoiding immune attack. Thus, the interaction of Toxoplasma with biological barriers is a determinant factor of human toxoplasmosis. The elucidation of determinants involved in the process of migration may reveal virulence factors and novel therapeutic targets to combat disease.
Asunto(s)
Células Epiteliales/parasitología , Toxoplasma/fisiología , Toxoplasma/patogenicidad , Animales , Ratones , Modelos Animales , VirulenciaRESUMEN
The obligate intracellular parasite Toxoplasma gondii chronically infects up to one-third of the global population, can result in severe disease in immunocompromised individuals, and can be teratogenic. In this study, we demonstrate that death receptor ligation in T. gondii-infected cells leads to rapid egress of infectious parasites and lytic necrosis of the host cell, an active process mediated through the release of intracellular calcium as a consequence of caspase activation early in the apoptotic cascade. Upon acting on infected cells via death receptor- or perforin-dependent pathways, T cells induce rapid egress of infectious parasites able to infect surrounding cells, including the Ag-specific effector cells.
Asunto(s)
Citotoxicidad Inmunológica , Perforina/metabolismo , Receptores de Muerte Celular/metabolismo , Linfocitos T Citotóxicos/inmunología , Toxoplasma/inmunología , Animales , Apoptosis , Calcio/metabolismo , Caspasas/metabolismo , Línea Celular , Humanos , Ratones , Ratones Congénicos , Perforina/genética , Linfocitos T Citotóxicos/parasitología , Receptor fas/metabolismoRESUMEN
Reactivation of infection in the central nervous system (CNS) with the opportunistic parasite Toxoplasma gondii is a major concern in chronically infected immunocompromised individuals. Yet, the pathophysiology associated with recrudescence of infection remains poorly characterized. The onset of acute reactivated Toxoplasma encephalitis in the murine model was assessed using bioluminescence imaging as a spatio-temporal indicator. An uneven distribution of recrudescence of infection in the CNS was found. Foci of recrudescence after immunosuppression were most commonly located in frontal and parietal cortex, whereas little infection was found in the cerebellum. Recrudescence was also more common in grey matter than in white matter. Pathology was exacerbated in mice deficient in interferon gamma receptors (IFN gamma R(-/-)) corroborating the importance of interferon gamma (IFN gamma) for control of CNS infection. Analysis of parasitic foci identified abundant leukocyte infiltration (CD45+, CD4+, CD8+, F4/80+ cells) in the vicinity of replicating parasites and microvasculature. This is the first report that addresses the suborganic localization of acute Toxoplasma encephalitis in the murine model. Collectively, the findings suggest that the localization of reactivation foci in the CNS, in conjunction with immune responses, influences the outcome of acute reactivated Toxoplasma encephalitis.
Asunto(s)
Sistema Nervioso Central/parasitología , Diagnóstico por Imagen/métodos , Luminiscencia , Toxoplasmosis Cerebral/inmunología , Animales , Sistema Nervioso Central/patología , Corteza Cerebral/patología , Lóbulo Frontal/patología , Huésped Inmunocomprometido , Interferón gamma/inmunología , Leucocitos/inmunología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Lóbulo Parietal/patología , Receptores de Interferón/deficiencia , Recurrencia , Receptor de Interferón gammaRESUMEN
The processes leading to systemic dissemination of the obligate intracellular parasite Toxoplasma gondii remain unelucidated. In vitro studies on human and murine dendritic cells (DC) revealed that active invasion of DC by Toxoplasma induces a state of hypermotility in DC, enabling transmigration of infected DC across endothelial cell monolayers in the absence of chemotactic stimuli. Infected DC exhibited upregulation of maturation markers and co-stimulatory molecules. While modulation of cell adhesion molecules CD11/CD18 was similar for Toxoplasma-infected DC and lipopolysaccharide (LPS)-matured DC, Toxoplasma-infected DC did not exhibit upregulation of CD54/ICAM-1. Induction of host cell migration in vitro required live intracellular parasite(s) and was inhibited by uncoupling the Gi-protein signalling pathway with pertussis toxin, but did not depend on CCR5, CCR7 or Toll/interleukin-1 receptor signalling. When migration of Toxoplasma-infected DC was compared with migration of LPS-stimulated DC in vivo, similar or higher numbers of Toxoplasma-infected DC reached the mesenteric lymph nodes and spleen respectively. Adoptive transfer of Toxoplasma-infected DC resulted in more rapid dissemination of parasites to distant organs and in exacerbation of infection compared with inoculation with free parasites. Altogether, these findings show that Toxoplasma is able to subvert the regulation of host cell motility and likely exploits the host's natural pathways of cellular migration for parasite dissemination.
Asunto(s)
Movimiento Celular , Células Dendríticas/parasitología , Toxoplasma/parasitología , Toxoplasmosis/parasitología , Traslado Adoptivo , Animales , Células de la Médula Ósea/inmunología , Células Cultivadas , Células Dendríticas/citología , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/metabolismo , Proteínas Fluorescentes Verdes , Humanos , Interleucina-1/metabolismo , Ganglios Linfáticos/parasitología , Ratones , Ratones Endogámicos C57BL , Receptores CCR5/metabolismo , Receptores CCR7 , Receptores de Quimiocina/metabolismo , Transducción de Señal , Bazo/parasitología , Toxoplasma/inmunología , Toxoplasma/patogenicidad , Toxoplasmosis/inmunologíaRESUMEN
Toxoplasma gondii infection can lead to life-threatening systemic disease in the immunocompromised individual and in the developing fetus. Despite intensive investigation in animal models of toxoplasmosis, the processes leading to systemic dissemination remain poorly characterized. In the present study, in vivo bioluminescence imaging (BLI) was applied to the Toxoplasma mouse model to study the dynamics of infection in real time. Photon emission analyses revealed rapid dissemination of parasites in the organism and dissemination to immunoprivileged organs (brain, eyes and testes). Spatio-temporal analysis by BLI in individual mice showed that the virulent RH strain (type I) and the non-virulent ME49/PTG strain (type II) disseminate widely, but the virulent RH strain (type I) exhibits a more dramatic expansion of parasite biomass. Assessment by BLI of the Toll/interleukin-1 receptor (TIR) signalling pathway in host resistance to T. gondii revealed that signal transduction to the adaptor protein MyD88 is probably mediated by Toll-like receptor(s) rather than by IL-1R or IL-18R signalling. However, TLR1(-/-), TLR2(-/-), TLR4(-/-), TLR6(-/-) and TLR9(-/-) animals did not exhibit increased susceptibility to infection. These results suggest that intricate mechanisms regulate TIR-mediated responses during Toxoplasma infection.
Asunto(s)
Glicoproteínas de Membrana/fisiología , Receptores de Superficie Celular/fisiología , Receptores de Interleucina-1/fisiología , Toxoplasma/patogenicidad , Toxoplasmosis Animal/inmunología , Proteínas Adaptadoras Transductoras de Señales , Animales , Antígenos de Diferenciación/genética , Caspasa 1/genética , Inmunidad Innata , Mediciones Luminiscentes , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Factor 88 de Diferenciación Mieloide , Receptores de Superficie Celular/genética , Receptores Inmunológicos/genética , Transducción de Señal , Receptor Toll-Like 1 , Receptor Toll-Like 2 , Receptor Toll-Like 4 , Receptores Toll-Like , Toxoplasmosis Animal/genética , Toxoplasmosis Animal/parasitologíaRESUMEN
HIV proteins contain a multitude of naturally processed cytotoxic T lymphocyte (CTL) epitopes that concentrate in clusters. The molecular basis of epitope clustering is of interest for understanding HIV immunogenicity and for vaccine design. We show that the CTL epitope clusters of HIV proteins predominantly coincide with hydrophobic regions, whereas the noncluster regions are predominantly hydrophilic. Analysis of the proteasomal degradation products of full-length HIV-Nef revealed a differential sensitivity of cluster and noncluster regions to proteasomal processing. Compared with the epitope-scarce noncluster regions, cluster regions are digested by proteasomes more intensively and with greater preference for hydrophobic P1 residues, resulting in substantially greater numbers of fragments with the sizes and COOH termini typical of epitopes and their precursors. Indeed, many of these fragments correspond to endogenously processed Nef epitopes and/or their potential precursors. The results suggest that differential proteasomal processing contributes importantly to the clustering of CTL epitopes in hydrophobic regions.
Asunto(s)
Cisteína Endopeptidasas/metabolismo , Productos del Gen nef/química , VIH-1/metabolismo , Complejos Multienzimáticos/metabolismo , Proteínas/química , Secuencia de Aminoácidos , Aminoácidos/química , Línea Celular , Epítopos , Productos del Gen nef/metabolismo , Humanos , Células Jurkat , Datos de Secuencia Molecular , Péptidos/química , Complejo de la Endopetidasa Proteasomal , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Homología de Secuencia de Aminoácido , Linfocitos T Citotóxicos/metabolismo , Productos del Gen nef del Virus de la Inmunodeficiencia HumanaRESUMEN
Yersinia enterocolitica mutant strains, including mutants deficient in the chaperone SycH resulting in a functional deficiency in tyrosine phosphatase (YopH), Mn-cofactored superoxide dismutase (SodA), iron-repressive protein 1 (IRP-1), and Yersinia adhesin A (YadA), were demonstrated to be highly attenuated in wild-type C57BL/6 mice. TNFRp55(-/-), IL-12p40(-/-), and IL-18(-/-) mutant mice, in which the Yersinia wild-type strain causes severe systemic infections, were used to investigate whether these Yersinia mutant strains would be attenuated in immunodeficient hosts. A plasmid-cured Yersinia mutant strain was unable to colonize any of the mutant mice tested. A SycH-deficient mutant strain colonized intestinal tissues of these mice but was attenuated for systemic infection in all of the mutant mice. Both YadA- and Irp-1-deficient Yersinia mutants were still attenuated in IL-12(-/-) and IL-18(-/-) mice but were pathogenic in TNFRp55(-/-) mice. By contrast, a Yersinia sodA mutant was highly pathogenic for TNFRp55(-/-) and IL-12p40(-/-) mice while interleukin-18 (IL-18) was dispensable. This finding demonstrates that certain virulence factors enable yersiniae to compete with distinct cytokine-dependent host defense mechanisms. Moreover, while gamma interferon mRNA expression did not reflect protective host responses in cytokine-deficient mice, IL-10 expression coincided with a heavy splenic bacterial load and was associated with progressive infection courses. We can thus segregate minor (SodA), intermediate (YadA and IRP-1), and major (YopH) virulence factors of Y. enterocolitica. Finally, we demonstrate that, even in immunocompromised hosts, Yersinia sycH and, with some restrictions, irp-1 mutants may be suitable for use as live carrier vaccines.
Asunto(s)
Citocinas/deficiencia , Ratones Noqueados , Mutación , Vacunas Atenuadas , Yersinia enterocolitica/patogenicidad , Animales , Proteínas Bacterianas/genética , Vacunas Bacterianas , Citocinas/genética , Susceptibilidad a Enfermedades , Humanos , Ratones , Ratones Endogámicos C57BL , Chaperonas Moleculares/genética , Virulencia , Factores de Virulencia/genética , Yersiniosis/inmunología , Yersiniosis/microbiología , Yersinia enterocolitica/genéticaRESUMEN
Recently, a novel 'two-step' model of pathogenicity has been described that suggests host-cell-derived vasculoproliferative factors play a crucial role in the pathogenesis of bacillary angiomatosis, a disease caused by the human pathogenic bacterium Bartonella henselae. The resulting proliferation of endothelial cells could be interpreted as bacterial pathogens triggering the promotion of their own habitat: the host cell. Similar disease mechanisms are well known in the plant pathogen Agrobacterium tumefaciens, which causes crown gall disease. There are notable similarities between the pathogenicity of A. tumefaciens leading to tumourous disease in plants and to the B. henselae-triggered proliferation of endothelial cells in humans. Here, we hypothesize that this pathogenicity strategy might be common to several bacterial species in different hosts owing to shared pathogenicity factors.