RESUMO
We have shown previously that T cells, tagged with biotinylated anti-CD3 antibody fragments, can exert avidin-dependent cytolytic activity on suitably biotinylated tumor cells in vitro. In this study, we demonstrate that avidin-driven CTL-tumor bridging in vivo leads to growth inhibition of murine tumors WEHI-164 fibrosarcoma and RMA lymphoma. The biodistribution of biotin-tagged 111In-labeled T cells demonstrated a selective avidin-dependent and time-dependent accumulation of radioactivity at tumor sites. The specificity of lymphocyte tumor localization was demonstrated by the concurrent time-dependent decrease of radioactivity in the blood and in all other organs. Furthermore, we documented a therapeutic effect of the adoptively transferred T cells, i.e., a significant delay of tumor growth at early stages. All of the experiments included a control group of mice, which received all of the reagents, except avidin. These avidin-minus mice showed no specific localization and no delay in tumor growth, indicating that avidin bridging was essential for T-cell activity at tumor sites.
Assuntos
Avidina/metabolismo , Biotina/metabolismo , Imunoterapia Adotiva/métodos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo , Animais , Anticorpos/imunologia , Anticorpos/metabolismo , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Biotinilação , Complexo CD3/imunologia , Citotoxicidade Imunológica , Feminino , Fibrossarcoma/imunologia , Fibrossarcoma/metabolismo , Fibrossarcoma/terapia , Linfoma de Células T/imunologia , Linfoma de Células T/metabolismo , Linfoma de Células T/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Transplante de NeoplasiasRESUMO
Gut-enriched Krüppel-like factor (GKLF) is a transcriptional regulator expressed in differentiated epithelia. We identified GKLF transcript as a regulated element in thymic epithelium of recombinase-deficient mice during thymus development induced by anti-CD3 antibody injection. This treatment recapitulates the organogenetic process depending on productive rearrangement of T cell receptor (TCR) beta gene with thymocytes expansion and acquisition of the CD4+8+ double positive phenotype. In wildtype mice, GKLF is expressed very early in embryogenesis and becomes intensely up-regulated in thymus epithelium at day 18 of gestation when TCR beta expressing cells have selectively expanded and express both CD4 and CD8. The results presented here suggest that thymocytes may regulate GKLF transcriptionally in the cortical epithelium at the developmental check-point controlled by TCR beta gene rearrangement. Furthermore, GKLF expression in hematopoietic stroma might suggest the thus far uncharacterised participation of this factor in hematopoiesis.
Assuntos
Proteínas de Ligação a DNA , Regulação da Expressão Gênica no Desenvolvimento , Células Estromais/fisiologia , Timo/metabolismo , Fatores de Transcrição/fisiologia , Células 3T3 , Animais , Anticorpos Monoclonais , Linhagem Celular , Regulação para Baixo , Epitélio/metabolismo , Imuno-Histoquímica , Separação Imunomagnética , Hibridização In Situ , Cinética , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like , Camundongos , Camundongos Endogâmicos BALB C , RNA/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ribonucleases/metabolismo , Análise de Sequência de DNA , Transdução de Sinais , Fatores de Tempo , Distribuição Tecidual , Fatores de Transcrição/análise , Fatores de Transcrição/metabolismo , Regulação para CimaRESUMO
There is still a considerable controversy about the usefulness of antiepileptic prophylaxis after traumatic brain injury. Overall incidence of posttraumatic fits and epilepsy's is well known, but an individual decision on prophylaxis requires knowledge about the individual risk. We performed a prospective observational study on 612 patients with traumatic brain injury of every degree of severity. Follow-up by phone call included 96.2% of the study population after 6 month and 91.2% after 36 month, respectively. The overall incidence for early fits (within 7 days after trauma), late fits (up to 36 month) and epilepsy (as defined by the International League Against Epilepsy) was 4.2%, 3.7% and 2.5%, respectively. These incidences increased according to the severity of the trauma, but the most powerful single predictor was intracranial hemorrhage. There was no significant difference related to the hemorrhage localisation. Development of epilepsy was much more common after late fits (48%) than after early fits (17%). These features established a hierarchy of risks for the development of epilepsy: no intracranial hemorrhage/no fit: 1% (4/437), intracranial hemorrhage/no fit: 8% (10/122), intracranial hemorrhage/early fit: 16% (3/19), intracranial hemorrhage/late fit: 53% (7/13). If prophylactic antiepileptic treatment is desired, but should be restricted to patients at high risk. These are patients with intracranial hemorrhage--are a well defined high risk group--when their first fit is a late fit.
Assuntos
Lesões Encefálicas/complicações , Lesões Encefálicas/fisiopatologia , Epilepsia/epidemiologia , Convulsões/epidemiologia , Epilepsias Parciais/epidemiologia , Epilepsia/etiologia , Epilepsia do Lobo Frontal/epidemiologia , Epilepsia do Lobo Temporal/epidemiologia , Seguimentos , Humanos , Incidência , Estudos Prospectivos , Fatores de Risco , Convulsões/etiologia , Índice de Gravidade de Doença , Fatores de TempoRESUMO
Thymus development depends on a complex series of interactions between thymocytes and the stromal component of the organ. To identify regulated genes during this codependent developmental relationship, we have applied an RNA fingerprinting technique to the analysis of thymus expansion and maturation induced in recombinase-deficient mice injected with anti-CD3 antibodies. This approach led us to the identification of a gene encoding a new member of the immunoglobulin superfamily, named epithelial V-like antigen (EVA), which is expressed in thymus epithelium and strongly downregulated by thymocyte developmental progression. This gene is expressed in the thymus and in several epithelial structures early in embryogenesis. EVA is highly homologous to the myelin protein zero and, in thymus-derived epithelial cell lines, is poorly soluble in nonionic detergents, strongly suggesting an association to the cytoskeleton. Its capacity to mediate cell adhesion through a homophilic interaction and its selective regulation by T cell maturation might imply the participation of EVA in the earliest phases of thymus organogenesis.
Assuntos
Moléculas de Adesão Celular/biossíntese , Mapeamento Cromossômico , Regulação da Expressão Gênica no Desenvolvimento , Timo/embriologia , Sequência de Aminoácidos , Animais , Células CHO , Moléculas de Adesão Celular/química , Moléculas de Adesão Celular/genética , Agregação Celular , Linhagem Celular , Cromossomos Humanos Par 11 , Clonagem Molecular , Cricetinae , Cruzamentos Genéticos , Primers do DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Embrião de Mamíferos , Desenvolvimento Embrionário e Fetal , Células Epiteliais/metabolismo , Células Epiteliais/fisiologia , Humanos , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência Molecular , Muridae , Proteínas Nucleares , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Biossíntese de Proteínas , Proteínas Recombinantes/biossíntese , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Timo/metabolismo , TransfecçãoRESUMO
Adhesive mechanisms are considered to be of crucial importance for blastocyst adherence to the uterine wall, as well as for the interactions between embryonal and decidual tissues during hemochorial placenta formation. Epithelial V-like Antigen (Eva) is a novel homophilic adhesion molecule of the immunoglobulin superfamily, which during mouse embryonic development is expressed by various differentiating epithelia. In the present paper we describe Eva expression during mouse trophoblast invasion and placental morphogenesis, analysing day 5.5 to 18.5 postcoitum (p.c.) placentas and deciduomas by in situ hybridization. Eva transcripts were detected in spongiotrophoblast cells from 7.5 to 18.5 days p.c. Expression was uniform at early stages, but after day 11.5, p.c. became limited to the invasive subpopulation of spongiotrophoblasts (known as glycogen cells). Trophoblast giant cells did not express Eva in any of the stages analysed. Besides trophoblasts, also early postimplantation decidua was positive for Eva transcripts. In decidual tissue, Eva expression was present at day 5.5 p.c., peaked at day 7.5 p.c., and declined on successive days. The expression pattern of Eva transcripts suggests that during mouse placenta formation, its protein product may play a role in the processes of trophoblast invasion, decidual response, and trophoblast-decidual interaction.
Assuntos
Moléculas de Adesão Celular/genética , Desenvolvimento Embrionário e Fetal/genética , Morfogênese/genética , Placenta/embriologia , Animais , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Hibridização In Situ , Camundongos , GravidezRESUMO
Physiological concentrations of urokinase plasminogen activator (uPA) stimulated a chemotactic response in human monocytic THP-1 through binding to the urokinase receptor (uPAR). The effect did not require the protease moiety of uPA, as stimulation was achieved also with the N-terminal fragment (ATF), while the 33 kDa low molecular weight uPA was ineffective. Co-immunoprecipitation experiments showed association of uPAR with intracellular kinase(s), as demonstrated by in vitro kinase assays. Use of specific antibodies identified p56/p59hck as a kinase associated with uPAR in THP-1 cell extracts. Upon addition of ATF, p56/p59hck activity was stimulated within 2 min and returned to normal after 30 min. Since uPAR lacks an intracellular domain capable of interacting with intracellular kinase, activation of p56/p59hck must require a transmembrane adaptor. Evidence for this was strongly supported by the finding that a soluble form of uPAR (suPAR) was capable of inducing chemotaxis not only in THP-1 cells but also in cells lacking endogenous uPAR (IC50, 5 pM). However, activity of suPAR require chymotrypsin cleavage between the N-terminal domain D1 and D2 + D3. Chymotrypsin-cleaved suPAR also induced activation of p56/p59hck in THP-1 cells, with a time course comparable with ATF. Our data show that uPA-induced signal transduction takes place via uPAR, involves activation of intracellular tyrosine kinase(s) and requires an as yet undefined adaptor capable of connecting the extracellular ligand binding uPAR to intracellular transducer(s).
Assuntos
Quimiotaxia de Leucócito/fisiologia , Receptores de Superfície Celular/metabolismo , Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Animais , Anticorpos Bloqueadores/farmacologia , Anticorpos Monoclonais/farmacologia , Linhagem Celular , Quimiotaxia de Leucócito/efeitos dos fármacos , Cricetinae , Endopeptidases/metabolismo , Ativação Enzimática , Humanos , Técnicas In Vitro , Camundongos , Monócitos/fisiologia , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-hck , Agregação de Receptores , Receptores de Ativador de Plasminogênio Tipo Uroquinase , Transdução de Sinais , Ativador de Plasminogênio Tipo Uroquinase/química , Ativador de Plasminogênio Tipo Uroquinase/farmacologiaRESUMO
Starting in November, 1993, until January, 1994, we performed a survey among 127 Neurosurgical Departments in Austria, Germany, and Switzerland concerning the practice of antiepileptic prophylaxis in head injured patients. Seventy seven percent of the 12-item multiple choice questionnaires were completed and returned. They indicate a variety of attitudes towards prophylaxis for seizures: in 12% of the responding institutions, antiepileptic prophylaxis is given to every brain trauma patient, in 36%, no prophylaxis is carried out. and in 52% some patients receive prophylaxis while others do not. Penetrating injuries, intracranial haemorrhages and electroencephalographic abnormalities were the most frequent reasons why prophylaxis was initiated. Phenytoin is by far the most popular drug, given usually for at least three months, and in most cases monitored by routine serum level observations. Nevertheless, about three out of four neurosurgeons conceded that a general renunciation of antiepileptic prophylaxis after brain trauma might be justified. There is no uniform way in which patients are informed about a possible risk of seizures, as it may be relevant, for instance, in respect of driving abilities.
Assuntos
Anticonvulsivantes/administração & dosagem , Lesões Encefálicas/complicações , Epilepsia Pós-Traumática/prevenção & controle , Atitude do Pessoal de Saúde , Áustria , Esquema de Medicação , Uso de Medicamentos , Alemanha , Humanos , Fatores de Risco , SuíçaRESUMO
During antigen recognition by T cells, CD4 and the T-cell receptor (TCR)/CD3/zeta complex are thought to interact with the same major histocompatibility complex II molecule in a stable ternary complex. Evidence has suggested that the association of CD4 with TCR/CD3/zeta requires the interaction of the protein tyrosine kinase p56lck with CD4. We have taken a biochemical approach to understand the mechanism by which p56lck and, in particular, its src homology (SH) 2 domain contributes to the association of CD4 with TCR/CD3/zeta during activation. We have previously shown that the p56lck SH2 domain binds directly to tyrosine-phosphorylated ZAP-70. Here we formally demonstrate the in vivo association of p56lck with the homologous protein tyrosine kinases Syk and ZAP-70 after CD3 stimulation of Jurkat cells. A tyrosine-phosphorylated peptide containing the sequence predicted to be optimal for binding to the SH2 domain of src family kinases specifically competes for this association, indicating that tyrosine-phosphorylated ZAP-70 and Syk bind to p56lck by an SH2-mediated interaction. We also show that the same peptide is able to compete for the activation-dependent TCR/CD4 association in Jurkat cells. Moreover, ZAP-70 and CD4 cocap only after CD3 stimulation in human T lymphoblasts. We propose that the interaction of the p56lck SH2 domain with zeta-associated tyrosine-phosphorylated ZAP-70 and/or Syk enables CD4 to associate with antigen-stimulated TCR/CD3/zeta complexes.
Assuntos
Antígenos CD/imunologia , Antígenos CD4/imunologia , Precursores Enzimáticos/metabolismo , Proteínas de Membrana/imunologia , Proteínas Tirosina Quinases/metabolismo , Complexo Receptor-CD3 de Antígeno de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/imunologia , Linhagem Celular , Eletroforese em Gel de Poliacrilamida , Precursores Enzimáticos/isolamento & purificação , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Cinética , Ativação Linfocitária , Proteína Tirosina Quinase p56(lck) Linfócito-Específica , Fosforilação , Fosfotirosina , Ligação Proteica , Proteínas Tirosina Quinases/isolamento & purificação , Quinase Syk , Células Tumorais Cultivadas , Tirosina/análogos & derivados , Tirosina/análise , Proteína-Tirosina Quinase ZAP-70RESUMO
Tyrosine phosphorylation of several intracellular proteins is observed very early during T cell activation. p56lck, a non-receptor src-like protein tyrosine kinase (PTK) which is associated with the intracellular domains of CD4 and CD8 co-receptors, has been implicated in these early signal transduction events. Furthermore, recent experiments indicate that the receptor phosphotyrosine phosphatase, CD45, might be important in the regulation of p56lck PTK activity and that its expression is required for the generation of second messenger molecules following TCR triggering. Here, using co-capping experiments and double indirect immunofluorescence microscopy in functional human T lymphocytes, a specific co-distribution of a significant fraction of p56lck with CD45, but not with several other cell surface proteins, has been revealed. This is the first demonstration of a physical interaction between a receptor phosphotyrosine phosphatase and a PTK under physiologically relevant conditions. In addition, after antibody-induced capping of CD4, both a co-localization of p56lck and CD4, and concomitantly a significant increase in intracellular phosphotyrosine at the sites of CD4 caps were observed. In strong contrast to these results, co-clustering of CD4 with CD45 did not result in any detectable intracellular phosphotyrosine at the cap sites. These data indicate that CD45 can act on CD4-associated phosphoproteins in viable human T lymphocytes. Further, this provides evidence that p56lck PTK is a substrate of CD45 phosphotyrosine phosphatase in vivo and thereby supports the idea that CD45 is an early regulator of T cell activation involved in the modulation of the coupling of receptor-triggered events to intracellular signalling pathways.
Assuntos
Antígenos Comuns de Leucócito/fisiologia , Ativação Linfocitária , Processamento de Proteína Pós-Traducional , Proteínas Tirosina Fosfatases/fisiologia , Proteínas Tirosina Quinases/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Transdução de Sinais , Linfócitos T/imunologia , Antígenos CD4/fisiologia , Linfócitos T CD4-Positivos/enzimologia , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Humanos , Capeamento Imunológico , Proteína Tirosina Quinase p56(lck) Linfócito-Específica , Fosforilação , Linfócitos T/enzimologiaRESUMO
The binding of antigen to the multicomponent T cell antigen receptor (TcR)-CD3 complex leads to the activation of several signal transduction pathways which result in T lymphocyte proliferation and lymphokine secretion by molecular mechanisms and catalytic molecules as yet poorly defined. One of the earliest events that follows the triggering of the antigen-specific TcR-CD3 complex is a rapid tyrosine phosphorylation of several intracellular substrates, suggesting stimulation of at least one protein tyrosine kinase (PTK). Since none of the seven TcR-CD3 subunits exhibits a recognizable kinase domain, it seems likely that the receptor complex is associated with an intracellular PTK. p59fyn and the T lymphocyte-specific p56lck are two intracellular, non-receptor, cell membrane-associated PTK of the src family expressed in T lymphocytes. Here, we show by double immunofluorescence microscopy a specific co-distribution of p59fyn, but not p56lck, with antibody-induced TcR or CD3 caps in intact human T lymphocytes. These findings provide direct evidence for a significant association of p59fyn with the TcR-CD3 complex under physiologically relevant conditions in functional T lymphocytes. They suggest that p59fyn is a crucial component of the TcR signal transduction machinery and that one of the earliest consequences of antigen recognition by the TcR is p59fyn-mediated phosphorylation of intracellular substrates on tyrosine residues.
Assuntos
Antígenos de Diferenciação de Linfócitos T/análise , Proteínas Tirosina Quinases/análise , Proteínas Proto-Oncogênicas/análise , Receptores de Antígenos de Linfócitos T/análise , Animais , Complexo CD3 , Humanos , Camundongos , Fosforilação , Proteínas Proto-Oncogênicas/fisiologia , Proteínas Proto-Oncogênicas c-fyn , Transdução de Sinais , Tirosina/metabolismoRESUMO
We have examined T cell recognition of a recombinant polypeptide (190L), corresponding to a 175-amino-acid-long conserved region of the major surface antigen (p190) of Plasmodium falciparum merozoites. We show that 190L contains a variety of T cell epitopes, and can be recognized in association with many different MHC class II molecules, including HLA-DR, DP, and DQ antigens. Most of the epitope-containing peptides are able to bind to more than one DR, and a single DR molecule can bind to different peptides. These findings, together with the fact that humans are generally heterozygous at the DR, DQ, and DP beta chain loci, suggest that MHC restriction should not be a major constraint in the development of malaria subunit vaccines.
Assuntos
Complexo Principal de Histocompatibilidade/imunologia , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Linfócitos T/imunologia , Vacinas Sintéticas/imunologia , Sequência de Aminoácidos , Animais , Formação de Anticorpos , Células Apresentadoras de Antígenos , Epitopos/imunologia , Genes MHC da Classe II , Humanos , Ativação Linfocitária/imunologia , Dados de Sequência Molecular , Polimorfismo Genético/imunologia , Proteínas Recombinantes/imunologiaRESUMO
The problems of genetic polymorphism and poor immunogenicity of malaria vaccine candidates are discussed, with emphasis on the Circumsporozoite (CS) and p190 proteins of Plasmodium falciparum. It may be possible to use conserved regions of these proteins to raise protective immune responses against non-polymorphic determinants. Better adjuvants or delivery systems will be a critical factor in development of an effective vaccine.
Assuntos
Antígenos de Protozoários/imunologia , Malária/imunologia , Plasmodium falciparum/imunologia , Vacinas Sintéticas , Sequência de Aminoácidos , Animais , Antígenos de Protozoários/genética , Antígenos de Protozoários/uso terapêutico , Clonagem Molecular/métodos , Humanos , Camundongos , Camundongos Endogâmicos , Dados de Sequência Molecular , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/uso terapêutico , Especificidade da Espécie , Linfócitos T/imunologiaAssuntos
Antígenos de Protozoários/imunologia , Epitopos/análise , Epitopos/imunologia , Antígenos HLA-D/imunologia , Plasmodium falciparum/imunologia , Engenharia de Proteínas/métodos , Linfócitos T/imunologia , Vacinas Sintéticas/imunologia , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Antígenos CD4/imunologia , Células Cultivadas , Células Clonais , Replicação do DNA , Humanos , Ativação Linfocitária , Malária Falciparum/imunologia , Dados de Sequência Molecular , Subpopulações de Linfócitos T/imunologia , Timidina/metabolismoRESUMO
In the case of the malaria CS protein we have shown that there is at least one T cell determinant which is able to bind to and be recognized by most human MHC class II molecules, while for the 190L polypeptide, derived from a conserved region of the p190 merozoite surface protein, we have identified several epitopes recognized by T cell clones in association with different HLA-class II isotypes and alleles. In addition, binding analysis of these epitopes indicated that most of the peptides are able to bind to multiple allelic forms of class II molecules. Although there are important obstacles to malaria vaccine development we believe that, in the light of these results, unresponsiveness in humans, caused by MHC restriction, might not be a major constraint in development of a subunit vaccine.
Assuntos
Antígenos de Protozoários/imunologia , Complexo Principal de Histocompatibilidade/imunologia , Malária/imunologia , Plasmodium falciparum/imunologia , Sequência de Aminoácidos , Animais , Antígenos H-2/imunologia , Antígenos HLA-D/imunologia , Humanos , Camundongos , Dados de Sequência Molecular , Proteínas de Protozoários/imunologia , Vacinas Protozoárias/imunologiaRESUMO
Although there are important obstacles to malaria vaccine development, we believe they might be overcome by a strategy of searching for conserved regions of a vaccine candidate that are recognized in association with many different HLA molecules and, if necessary, deliberately modifying the conserved sequences to improve their immunogenicity for T cells. This approach is illustrated by work on the circumsporozoite (CS) protein of Plasmodium falciparum, which covers the surface of the malaria sporozoite and is the best characterized of current vaccine candidates.
Assuntos
Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Epitopos/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários , Animais , Humanos , Sinais Direcionadores de Proteínas/genéticaRESUMO
The possibility of screening cDNA expression libraries with T cell clones was investigated. The model system was based on human T cell clones specific for the recombinant malaria protein 190L, which was expressed fused to beta-galactosidase in lambda gt11. Several membranes were tested for their capacity to bind antigen and stimulate T cell proliferation. Pretreatment of membranes with DMSO and/or sonication to release the antigen improved the sensitivity of the assay. Under optimal conditions, T cell proliferation in response to antigen bound to a low protein binder membrane was comparable to that observed with the antigen in solution. A dot-blot type apparatus was designed for screening large numbers of plaques with T cells. The technical problems of this approach, its requirements and possible applications are discussed.
Assuntos
DNA/análise , Epitopos/análise , Linfócitos T/imunologia , Células Clonais , HumanosRESUMO
The binding of several peptides derived from the Plasmodium falciparum circumsporozoite protein (CS protein) to the human major histocompatibility complex class II proteins HLA-DR5 and -DRw6 was examined in a competition assay. Fixed antigen-presenting cells (APCs) were incubated with various concentrations of each peptide and suboptimal concentrations of stimulator peptides. The binding of the CS peptides to DR5 or DRw6 proteins was then determined in a proliferation assay using two established DR5 or DRw6-restricted T-cell clones with specificity for the stimulator peptides as responder cells. One of five CS peptides, comprising together about 50% of the CS protein sequence, was found to compete with the binding of the stimulator peptides to DR5 and DRw6. The CS peptide CS-(378-398), binding to DR5 and DRw6, was then shown to be able to induce primary in vitro responses of T cells from donors with DR5 and DRw6 haplotypes. CS-(378-398)-induced T-cell clones responded not only to the homologous peptide but also to the native CS protein in the presence of appropriate APCs. The strategy we have applied is of considerable general interest for the engineering of vaccines against any pathogen, since it greatly facilitates the selection of appropriate T-cell epitopes to be incorporated in the vaccine.
Assuntos
Antígenos de Superfície/imunologia , Epitopos/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários , Linfócitos T/imunologia , Vacinas Sintéticas , Vacinas , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Ligação Competitiva , Antígenos HLA-DR/imunologia , Antígeno HLA-DR5 , Antígeno HLA-DR6 , Humanos , Ativação LinfocitáriaRESUMO
An ideal vaccine should elicit a long lasting immune response against the natural parasite, both at the T- and B-cell level. The immune response should occur in all individuals and be directed against determinants that do not vary in the natural parasite population. A major problem in designing synthetic peptide vaccines is that T cells generally recognize peptide antigens only in association with one or a few of the many variants of major histocompatibility complex (MHC) antigens. During the characterization of epitopes of the malaria parasite Plasmodium falciparum that are recognized by human T cells, we analysed a sequence of the circumsporozoite protein, and found that synthetic peptides corresponding to this sequence are recognized by T cells in association with many different MHC class II molecules, both in mouse and in man. This region of the circumsporozoite protein is invariant in different parasite isolates. Peptides derived from this region should be capable of inducing T-cell responses in individuals of most HLA-DR types, and may represent good candidates for inclusion in an effective anti-malaria peptide vaccine.
Assuntos
Antígenos de Protozoários/imunologia , Antígenos de Histocompatibilidade Classe II , Malária/imunologia , Plasmodium falciparum/imunologia , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Epitopos , Humanos , Camundongos , Dados de Sequência MolecularRESUMO
In order to characterize T cell epitopes in the Plasmodium falciparum circumsporozoite (CS) protein sequence, we isolated T cell clones, from non-immune donors, which reacted with synthetic peptides corresponding to two predicted CS protein T cell epitopes. Peptide CS.T3 (corresponding to a non-polymorphic region of the CS protein, residues 378-398) was recognized in association with either DR2 or DRw9 restriction elements. T cell clones recognizing CS.T3 also reacted with the sporozoite-derived CS protein. Peptide CS.T2 corresponds to a polymorphic region (residues 325-341) of the CS protein. Unlike the CS.T3-specific clones, the CS.T2-specific clones did not recognize the CS protein. Since the CS.T2 peptide includes residues which are polymorphic in different P. falciparum isolates, we investigated whether these residues were critical for recognition of the peptide. We show here that a single amino acid substitution at a position of the CS protein which shows genetic polymorphism affects recognition of the sequence by human T cells. The implications of these data for malaria vaccine development are discussed.