Characterization of the B-cell receptor repertoires in peanut allergic subjects undergoing oral immunotherapy
Characterization of the B-cell receptor repertoires in peanut allergic subjects undergoing oral immunotherapy
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DOI：
10.1038/s10038-017-0364-0
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发表时间：
2018-02-01
影响因子：
3.5
通讯作者：
Nakamura, Yusuke
中科院分区：
文献类型：
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作者：
Kiyotani, Kazuma;Mai, Tu H.;Nakamura, Yusuke
B细胞受体（BCR）在获得性免疫中发挥着关键作用，因为它们产生高度多样化的免疫球蛋白库来识别多种抗原。为了更好地了解免疫反应，建立一种定量和快速的方法来全面分析BCR库是至关重要的。在这里，我们开发了“Bcrip”，一种新的方法来表征BCR库的测序数百万的BCR cDNA使用下一代测序仪。使用这种方法和定量实时PCR，我们分析了共17个花生过敏受试者的外周血样本的表达水平和剧目之前和之后接受口服免疫治疗（OIT）或安慰剂。通过我们的方法，我们成功地鉴定了所有可变（V）、连接（J）和恒定（C）区，平均占总读段的79.1%，并且这些VJC映射的读段中的99.6%包含与我们旨在分析的同种型对应的C区。在17例花生过敏受试者的外周血样本中，我们观察到OIT后花生过敏受试者中某些免疫球蛋白重链α（IGHA）和IGH γ（IGHG）克隆的寡克隆富集（分别为P = 0.034和P = 0.027）。这种新开发的BCR测序和分析方法可用于研究各种研究领域的B细胞库，包括食物过敏以及自身免疫和传染病。
B-cell receptors (BCRs) play a critical role in adaptive immunity as they generate highly diverse immunoglobulin repertoires to recognize a wide variety of antigens. To better understand immune responses, it is critically important to establish a quantitative and rapid method to analyze BCR repertoire comprehensively. Here, we developed "Bcrip", a novel approach to characterize BCR repertoire by sequencing millions of BCR cDNA using next-generation sequencer. Using this method and quantitative real-time PCR, we analyzed expression levels and repertoires of BCRs in a total of 17 peanut allergic subjects' peripheral blood samples before and after receiving oral immunotherapy (OIT) or placebo. By our methods, we successfully identified all of variable (V), joining (J), and constant (C) regions, in an average of 79.1% of total reads and 99.6% of these VJC-mapped reads contained the C region corresponding to the isotypes that we aimed to analyze. In the 17 peanut allergic subjects' peripheral blood samples, we observed an oligoclonal enrichment of certain immunoglobulin heavy chain alpha (IGHA) and IGH gamma (IGHG) clones (P = 0.034 and P = 0.027, respectively) in peanut allergic subjects after OIT. This newly developed BCR sequencing and analysis method can be applied to investigate B-cell repertoires in various research areas, including food allergies as well as autoimmune and infectious diseases.