背景:全球,杂草艾草(Artemisiavulgaris)的花粉是严重呼吸道过敏的主要原因,它的主要过敏原,第1条,是数百万患者的关键致病分子。已经制备了对主要T细胞表位和相应HLA具有特异性的人T细胞受体转基因的人源化小鼠。
目的:我们试图表征Artv1致敏患者和人源化小鼠的IgE表位,用于艾草过敏的分子免疫治疗。
方法:合成四种重叠的肽,所述肽包含代表全长Artv1序列的表面暴露的氨基酸,并用于搜索IgE对连续表位的反应性。对于间接映射,产生肽特异性兔抗体以阻断折叠的Artv1上表面暴露的表位的IgE。在临床定义的艾草过敏患者中进行了IgE反应性和嗜碱性粒细胞活化研究。通过圆二色性光谱法确定重组(r)Artv1和肽的二级结构。
结果:变应原吸入致敏的艾草过敏患者和人源化小鼠显示IgE反应性和/或嗜碱性粒细胞活化主要为折叠,完成艺术v1,但不展开,序贯肽表位。用肽特异性兔抗血清阻断过敏性患者IgE,在C-末端Artv1结构域中鉴定出迄今未知的主要构象IgE结合位点。
结论:鉴定第v1条上新的主要构象IgE结合位点,该位点可以用针对非IgE反应性第v1条肽的IgG阻断,是艾草过敏低变应原肽疫苗研制的重要基础。
BACKGROUND: Worldwide, pollen of the weed mugwort (Artemisiavulgaris) is a major cause of severe respiratory allergy, with its major allergen, Art v 1, being the key pathogenic molecule for millions of patients. Humanized mice transgenic for a human T-cell receptor specific for the major Art v 1 T-cell epitope and the corresponding HLA have been made.
OBJECTIVE: We sought to characterize IgE epitopes of Art v 1-sensitized patients and humanized mice for molecular immunotherapy of mugwort allergy.
METHODS: Four overlapping peptides incorporating surface-exposed amino acids representing the full-length Art v 1 sequence were synthesized and used to search for IgE reactivity to sequential epitopes. For indirect mapping, peptide-specific rabbit antibodies were raised to block IgE against surface-exposed epitopes on folded Art v 1. IgE reactivity and basophil activation studies were performed in clinically defined mugwort-allergic patients. Secondary structure of recombinant (r) Art v 1 and peptides was determined by circular dichroism spectroscopy.
RESULTS: Mugwort-allergic patients and humanized mice sensitized by allergen inhalation showed IgE reactivity and/or basophil activation mainly to folded, complete Art v 1 but not to unfolded, sequential peptide epitopes. Blocking of allergic patients\' IgE with peptide-specific rabbit antisera identified a hitherto unknown major conformational IgE binding site in the C-terminal Art v 1 domain.
CONCLUSIONS: Identification of the new major conformational IgE binding site on Art v 1, which can be blocked with IgG raised against non-IgE reactive Art v 1 peptides, is an important basis for the development of a hypoallergenic peptide vaccine for mugwort allergy.