Abstract：
BACKGROUND: Antigen-specific memory B cells play a key role in the induction of desensitization and remission to food allergens in oral immunotherapy and in the development of natural tolerance (NT). Here, we characterized milk allergen Bos d 9-specific B cells in oral allergen-specific immunotherapy (OIT) and in children spontaneously outgrowing cow\'s milk allergy (CMA) due to NT.
METHODS: Samples from children with CMA who received oral OIT (before, during, and after), children who naturally outgrew CMA (NT), and healthy individuals were received from Stanford biobank. Bos d 9-specific B cells were isolated by flow cytometry and RNA-sequencing was performed. Protein profile of Bos d 9-specific B cells was analyzed by proximity extension assay.
RESULTS: Increased frequencies of circulating milk allergen Bos d 9-specific B cells were observed after OIT and NT. Milk-desensitized subjects showed the partial acquisition of phenotypic features of remission, suggesting that desensitization is an earlier stage of remission. Within these most significantly expressed genes, IL10RA and TGFB3 were highly expressed in desensitized OIT patients. In both the remission and desensitized groups, B cell activation-, Breg cells-, BCR-signaling-, and differentiation-related genes were upregulated. In NT, pathways associated with innate immunity characteristics, development of marginal zone B cells, and a more established suppressor function of B cells prevail that may play a role in long-term tolerance. The analyses of immunoglobulin heavy chain genes in specific B cells demonstrated that IgG2 in desensitization, IgG1, IgA1, IgA2, IgG4, and IgD in remission, and IgD in NT were predominating. Secreted proteins from allergen-specific B cells revealed higher levels of regulatory cytokines, IL-10, and TGF-β after OIT and NT.
CONCLUSIONS: Allergen-specific B cells are essential elements in regulating food allergy towards remission in OIT-received and naturally resolved individuals.
METHODS: Samples from children with CMA who received oral OIT (before, during, and after), children who naturally outgrew CMA (NT), and healthy individuals were received from Stanford biobank. Bos d 9-specific B cells were isolated by flow cytometry and RNA-sequencing was performed. Protein profile of Bos d 9-specific B cells was analyzed by proximity extension assay.
RESULTS: Increased frequencies of circulating milk allergen Bos d 9-specific B cells were observed after OIT and NT. Milk-desensitized subjects showed the partial acquisition of phenotypic features of remission, suggesting that desensitization is an earlier stage of remission. Within these most significantly expressed genes, IL10RA and TGFB3 were highly expressed in desensitized OIT patients. In both the remission and desensitized groups, B cell activation-, Breg cells-, BCR-signaling-, and differentiation-related genes were upregulated. In NT, pathways associated with innate immunity characteristics, development of marginal zone B cells, and a more established suppressor function of B cells prevail that may play a role in long-term tolerance. The analyses of immunoglobulin heavy chain genes in specific B cells demonstrated that IgG2 in desensitization, IgG1, IgA1, IgA2, IgG4, and IgD in remission, and IgD in NT were predominating. Secreted proteins from allergen-specific B cells revealed higher levels of regulatory cytokines, IL-10, and TGF-β after OIT and NT.
CONCLUSIONS: Allergen-specific B cells are essential elements in regulating food allergy towards remission in OIT-received and naturally resolved individuals.
摘要：
背景：抗原特异性记忆B细胞在口服免疫疗法中诱导脱敏和缓解食物过敏原以及天然耐受性（NT）的发展中起关键作用。这里,我们在口服过敏原特异性免疫疗法（OIT）和儿童中描述了牛奶过敏原Bosd9特异性B细胞的特点。
方法：来自接受口服OIT的CMA儿童的样本（之前，during,之后),自然超过CMA（NT）的孩子，健康个体来自斯坦福生物银行。通过流式细胞术分离Bosd9特异性B细胞并进行RNA测序。通过邻近延伸测定法分析Bosd9特异性B细胞的蛋白质谱。
结果：在OIT和NT后观察到循环牛奶过敏原Bosd9特异性B细胞的频率增加。牛奶脱敏受试者显示部分获得缓解的表型特征，表明脱敏是缓解的早期阶段。在这些最显著表达的基因中，IL10RA和TGFB3在脱敏OIT患者中高表达。在缓解组和脱敏组中，B细胞激活-，布雷格细胞-，BCR-信号-,分化相关基因上调。在NT,与先天免疫特征相关的途径，边缘区B细胞的发育，并且B细胞的抑制功能更为确立，可能在长期耐受性中起作用。对特定B细胞中免疫球蛋白重链基因的分析表明，IgG2在脱敏中，IgG1、IgA1、IgA2、IgG4和IgD缓解期，NT中以IgD为主。从过敏原特异性B细胞分泌的蛋白质显示更高水平的调节细胞因子，OIT和NT后的IL-10和TGF-β。
结论：过敏原特异性B细胞是调节OIT接受和自然解决的个体的食物过敏缓解的基本要素。
方法：来自接受口服OIT的CMA儿童的样本（之前，during,之后),自然超过CMA（NT）的孩子，健康个体来自斯坦福生物银行。通过流式细胞术分离Bosd9特异性B细胞并进行RNA测序。通过邻近延伸测定法分析Bosd9特异性B细胞的蛋白质谱。
结果：在OIT和NT后观察到循环牛奶过敏原Bosd9特异性B细胞的频率增加。牛奶脱敏受试者显示部分获得缓解的表型特征，表明脱敏是缓解的早期阶段。在这些最显著表达的基因中，IL10RA和TGFB3在脱敏OIT患者中高表达。在缓解组和脱敏组中，B细胞激活-，布雷格细胞-，BCR-信号-,分化相关基因上调。在NT,与先天免疫特征相关的途径，边缘区B细胞的发育，并且B细胞的抑制功能更为确立，可能在长期耐受性中起作用。对特定B细胞中免疫球蛋白重链基因的分析表明，IgG2在脱敏中，IgG1、IgA1、IgA2、IgG4和IgD缓解期，NT中以IgD为主。从过敏原特异性B细胞分泌的蛋白质显示更高水平的调节细胞因子，OIT和NT后的IL-10和TGF-β。
结论：过敏原特异性B细胞是调节OIT接受和自然解决的个体的食物过敏缓解的基本要素。
