RESUMEN
Recently, significant advances in our understanding of embryo implantation have been made using animal models, notably in the mouse and nonhuman primates. However, the determination of the molecular and cellular events that underpin the early stages of implantation in the human remains an intractable problem, in part due to the inaccessibility of early human implantation sites. In the absence of in vivo implantation sites, several experimental in vitro model systems have been developed recently that mimic the different stages of human embryo implantation that occur in vivo during the first few weeks of pregnancy. These include solid-phase assays of blastocyst attachment and trophoblast invasion, and two- and three-dimensional blastocyst-endometrial cell cocultures. An important feature of such models is that they allow functional studies to be performed and are not restricted to generating descriptive data. These models have the potential to make an important contribution to the development of new therapeutic and diagnostic strategies for implantation failure in the future, as well as toxicity testing. We describe the strengths and weakness of the models and some of the advances that have been made by the use of these in vitro models.
Asunto(s)
Técnicas de Cultivo de Célula , Técnicas de Cocultivo , Técnicas de Cultivo de Embriones , Implantación del Embrión , Investigaciones con Embriones , Embrión de Mamíferos/fisiología , Endometrio/fisiología , Blastocisto/fisiología , Adhesión Celular , Movimiento Celular , Colágeno/metabolismo , Combinación de Medicamentos , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Endometrio/citología , Endometrio/metabolismo , Células Epiteliales/fisiología , Matriz Extracelular/metabolismo , Femenino , Humanos , Laminina/metabolismo , Embarazo , Proteoglicanos/metabolismo , Células del Estroma/fisiología , Trofoblastos/fisiologíaRESUMEN
BACKGROUND: The major peanut allergen Ara h 2 consists of two isoforms, namely Ara h 2.0101 and Ara h 2.0201. The recently identified Ara h 2.0201 isoform contains an extra 12 amino acids including an extra copy of the reported immunodominant epitope DPYSPS. This study aimed to evaluate the IgE binding of the two Ara h 2 isoforms. METHODS: Ten clones of Ara h 2 were sequenced to assess the relative frequency of the Ara h 2 isoforms and to identify whether there was further variation in the Ara h 2 sequence. IgE binding to Ara h 2.0101 and Ara h 2.0201 was measured for 70 peanut-allergic children using an IgE DELFIA assay to quantitate specific IgE binding. A competition assay was used to measure whether Ara h 2.0201 contained IgE epitopes other than those found for Ara h 2.0101. RESULTS: The original Ara h 2.0101 sequence was found for 6/10 clones and Ara h 2.0201 was found for 2/10 clones. Ara h 2.0201 had the expected insertion of 12 amino acids as well as substitutions at positions 40 (40G) and 142 (142E). Two new isoforms were identified as different polymorphisms of position 142. One Ara h 2.01 clone (Ara h 2.0102) contained 142E and one Ara h 2.02 clone (Ara h 2.0202) contained 142D. A polymorphism that was previously identified by other investigators at position 77 (77Q or 77R) was not found for any of the 10 sequences. Although the level of IgE binding to Ara h 2.0201 of individual patients was frequently higher than the binding to Ara h 2.0101 (p < 0.01), there was a strong correlation in binding to both isoforms (r = 0.987, p < 0.0001) and when analyzed as a group the means were similar. Ara h 2.0101 was not as efficient at blocking reactivity to Ara h 2.0201 indicating there is an additional IgE specificity for the Ara h 2.0201 isoform. CONCLUSIONS: Ara h 2.0201 has similar but higher IgE binding than the originally sequenced Ara h 2.0101 isoform and contains other IgE specificities.
Asunto(s)
Arachis/inmunología , Glicoproteínas/genética , Glicoproteínas/inmunología , Inmunoglobulina E/inmunología , Hipersensibilidad al Cacahuete/inmunología , Isoformas de Proteínas/inmunología , Albuminas 2S de Plantas , Adolescente , Secuencia de Aminoácidos , Especificidad de Anticuerpos/inmunología , Antígenos de Plantas , Unión Competitiva , Niño , Preescolar , Glicoproteínas/química , Humanos , Lactante , Datos de Secuencia Molecular , Proteínas de Plantas , Reacción en Cadena de la Polimerasa , Polimorfismo Genético , Isoformas de Proteínas/química , Isoformas de Proteínas/genéticaRESUMEN
The examination of house dust mite extracts has indicated that over 30 different proteins can induce IgE antibody in patients allergic to the house dust mite. There are however dominant specificities especially the group 1 and 2 allergens which can account for much of the allergenicity of extracts. Of the 19 denominated allergens, the major IgE binding has been reported for the group 1, 2, 3, 9, 11, 14 and 15 allergens. The high-molecular-weight group 11, 14 and 15 allergens have only recently been described and although high IgE binding has been anticipated from immunoblotting, there is a need for considerable corroboration. Similarly, the study of the group 3 and 9 serine protease allergens has been incomplete. The group 4, 5, 7 and 8 allergens have shown intermediate IgE binding and the group 10 tropomyosins are of interest because of their potential cross-reactivity with allergen from disparate species. Although the progress with the production of recombinant group 1 allergens has been recent, many of the allergens can be produced as high IgE-binding polypeptides. The tertiary structure of the group 2 allergens has been determined from recombinant proteins and they are an excellent model for the investigation of modified allergens. An unexpected property of the group 1, 2 and 3 allergens has been the high degree of polymorphism found by cDNA analysis. It has however been possible to identify sequences to represent the variation in the natural allergens. The group 7 and 14 allergens show secondary modifications which vary in different extracts creating batch variation. While some estimate of the importance of allergens can be obtained from IgE binding, few analyses of T-cell responses have been made and these regulate both the development of, and the protection from sensitization.