RESUMEN
BACKGROUND: Active immunization against Aß was reported to have a therapeutic effect in murine models of Alzheimer's disease. Clinical Aß vaccination trial AN1792 was interrupted due to the development in 6 % of the patients of meningoencephalitis likely involving pro-inflammatory CD4(+) T cells. However, the potential implication of auto-aggressive anti-Aß CD8(+) T cells has been poorly investigated. METHODS: Potential MHC-I-restricted Aß-derived epitopes were first analyzed for their capacity to recruit functional CD8(+) T cell responses in mouse models. Their impact on migration of CD8(+) T cells into the brain parenchyma and potential induction of meningoencephalitis and/or neuronal damage was investigated upon vaccination in the APPPS1 mouse model of AD. RESULTS: We identified one nonamer peptide, Aß33-41, which was naturally processed and presented in association with H-2-D(b) molecule on neurons and CD11b(+) microglia. Upon optimization of anchor residues for enhanced binding to H-2-D(b), immunization with the modified Aß33-41NP peptide elicited Aß-specific IFNγ-secreting CD8(+) T cells, which are cytotoxic towards Aß-expressing targets. Whereas T cell infiltration in the brain of APPPS1 mice is dominated by CD3(+)CD8(-) T cells and increases with disease evolution between 4 and 7 months of age, a predominance of CD3(+)CD8(+) over CD3(+)CD8(-) cells was observed in 6- to 7-month-old APPPS1 but not in WT animals, only after vaccination with Aß33-41NP. The number of CD11b(+) mononuclear phagocytes, which significantly increases with age in the brain of APPPS1 mice, was reduced following immunization with Aß33-41NP. Despite peripheral activation of Aß-specific CD8(+) cytotoxic effectors and enhanced infiltration of CD8(+) T cells in the brain of Aß33-41NP-immunized APPPS1 mice, no clinical signs of severe autoimmune neuroinflammation were observed. CONCLUSIONS: Altogether, these results suggest that Aß-specific CD8(+) T cells are not major contributors to meningoencephalitis in response to Aß vaccination.
Asunto(s)
Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/terapia , Péptidos beta-Amiloides/metabolismo , Encefalitis/etiología , Encefalitis/patología , Inmunoterapia Activa/efectos adversos , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/inmunología , Péptidos beta-Amiloides/inmunología , Precursor de Proteína beta-Amiloide/genética , Animales , Anticuerpos/análisis , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Modelos Animales de Enfermedad , Epítopos de Linfocito T/inmunología , Antígeno HLA-A2/genética , Antígeno HLA-A2/metabolismo , Antígeno HLA-DR1/genética , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microglía/inmunología , Microglía/metabolismo , Mutación/genética , Fragmentos de Péptidos/inmunología , Presenilina-1/genéticaRESUMEN
Recent reports suggest that immunity against the prion protein (PrP) retards transmissible spongiform encephalopathies progression in infected mice. A major obstacle to the development of vaccines comes from the fact that PrP is poorly immunogenic, as it is seen as self by the host immune system. Additional questions concern the immune mechanisms involved in protection and the risk of eliciting adverse reactions in the central nervous system of treated patients. Peptide-based vaccines offer an attractive strategy to overcome these difficulties. We have undertaken the identification of the immunogenic regions of PrP, which trigger helper T cells (Th) associated with antibody production. Our results identify two main regions, one between the structured and flexible portion of PrP (98-127) and a second between alpha 1 and alpha 2 helix (143-187). Peptides (30-mer) corresponding to these regions elicit class II-restricted Th cells and antibody production against native PrP and could therefore be of potential interest for a peptide-based vaccination.
Asunto(s)
Epítopos de Linfocito T , Proteínas PrPC/química , Proteínas PrPC/inmunología , Subgrupos de Linfocitos T/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Secuencia de Aminoácidos , Animales , Citocinas/biosíntesis , Ensayo de Inmunoadsorción Enzimática , Técnica del Anticuerpo Fluorescente , Antígenos de Histocompatibilidad Clase II , Ratones , Datos de Secuencia Molecular , Biblioteca de Péptidos , Péptidos/inmunología , Proteínas Recombinantes/inmunología , TransfecciónRESUMEN
In prion diseases, PrP(c), a widely expressed protein, is transformed into a pathogenic form called PrP(Sc), which is in itself infectious. Antibodies directed against PrP(c) have been shown to inhibit PrP(c) to PrP(Sc) conversion in vitro and protect in vivo from disease. Other effectors with potential to eliminate PrPSc-producing cells are cytotoxic T cells directed against PrP-derived peptides but their ability to protect or to induce deleterious autoimmune reactions is not known. The natural tolerance to PrP(c) makes difficult to raise efficient adaptive responses. To break tolerance, adenovirus (Ad) encoding human PrP (hPrP) or control Ad were administered to wild-type mice by direct injection or by transfer of Ad-transduced dendritic cells (DCs). Control Ad-transduced DCs from Tg650 mice overexpressing hPrP were also used for immunization. DC-mediated but not direct administration of AdhPrP elicited antibodies that bound to murine native PrP(c). Frequencies of PrP-specific IFNgamma-secreting T cells were low and in vivo lytic activity only targeted cells strongly expressing hPrP. Immunohistochemical analysis revealed that CD3(+) T cell infiltration was similar in the brain of vaccinated and unvaccinated 139A-infected mice suggesting the absence of autoimmune reactions. Early splenic PrP(Sc) replication was strongly inhibited ten weeks post infection and mean survival time prolonged from 209 days in untreated 139A-infected mice to 246 days in mice vaccinated with DCs expressing the hPrP. The efficacy appeared to be associated with antibody but not with cytotoxic cell-mediated PrP-specific responses.
Asunto(s)
Adenoviridae , Células Dendríticas/inmunología , Vectores Genéticos , Priones/inmunología , Scrapie/inmunología , Adenoviridae/genética , Adenoviridae/inmunología , Secuencia de Aminoácidos , Animales , Vectores Genéticos/genética , Vectores Genéticos/inmunología , Humanos , Inmunización , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Datos de Secuencia Molecular , Priones/genética , Scrapie/patología , Scrapie/prevención & control , Alineación de Secuencia , Tasa de SupervivenciaRESUMEN
Abs to the prion protein (PrP) can protect against experimental prion infections, but efficient Ab responses are difficult to generate because PrP is expressed on many tissues and induces a strong tolerance. We previously showed that immunization of wild-type mice with PrP peptides and CpG oligodeoxynucleic acid overcomes tolerance and induces cellular and humoral responses to PrP. In this study, we compared Ab and T cell repertoires directed to PrP in wild-type and PrP knockout (Prnp o/o) C57BL/6 mice. Animals were immunized with mouse PrP-plasmid DNA or with 30-mer overlapping peptides either emulsified in CFA or CpG/IFA. In Prnp o/o mice, Abs raised by PrP-plasmid DNA immunization recognized only N-terminal PrP peptides; analyses of Ab responses after PrP peptide/CFA immunization allowed us to identify six distinct epitopes, five of which were also recognized by Abs raised by PrP peptides/CpG. By contrast, in wild-type mice, no Ab response was detected after PrP-plasmid DNA or peptide/CFA immunization. However, when using CpG, four C-terminal peptides induced Abs specific for distinct epitopes. Importantly, immune sera from Prnp o/o but not from wild-type mice bound cell surface PrP. Abs of IgG1 and IgG2b subclasses predominated in Prnp o/o mice while the strongest signals were for IgG2b in wild-type mice. Most anti-PrP Th cells were directed to a single epitope in both Prnp o/o and wild-type mice. We conclude that endogenous PrPC expression profoundly affects the Ab repertoire as B cells reactive for epitopes exposed on native PrPC are strongly tolerized. Implications for immunotherapy against prion diseases are discussed.
Asunto(s)
Linfocitos B/inmunología , Priones/inmunología , Secuencia de Aminoácidos , Animales , Autoanticuerpos/biosíntesis , Mapeo Epitopo , Epítopos/genética , Femenino , Tolerancia Inmunológica , Inmunización , Inmunoglobulina G/biosíntesis , Inmunoterapia , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Datos de Secuencia Molecular , Enfermedades por Prión/inmunología , Enfermedades por Prión/terapia , Priones/genética , Linfocitos T/inmunologíaRESUMEN
The absence of a detectable immune response during transmissible spongiform encephalopathies is likely due to the fact that the essential component of infectious agents, the prion protein (PrP), is a self Ag expressed on the surface of many cells of the host. To overcome self-tolerance to PrP, we used 30-mer PrP peptides previously shown to be immunogenic in Prnp(-/-) mice, together with CFA or CpG-oligodeoxynucleotides (CpG) in IFA. Generation of anti-PrP T and B cell responses was analyzed in the spleen, lymph nodes, and serum of immunized C57BL/6 wild-type mice. Immunization with PrP peptides emulsified in CFA did not trigger an immune response to PrP. When CpG were used, vaccination with peptides P143-172 and P158-187 generated IFN-gamma-secreting splenic T cells, and only P158-187 significantly stimulated IL-4-secreting T cells. Both peptides induced few Ab-producing B cells, and low and variable serum Ab titers. In contrast, immunization with peptide P98-127 did not induce significant levels of T cell responses but elicited specific peptide Abs. T cell epitope mapping, performed using 15-mer peptides covering PrP segment 142-182, revealed that an immunogenic motif lies between positions 156 and 172. These results demonstrate that T and B cell repertoires against PrP can be stimulated in C57BL/6 when adjuvant of the innate immunity such as CpG, but not CFA, is added to PrP peptides, and that the pattern of immune responses varies according to the epitope.