Inflammation, Chimiokines et Immunopathologie, INSERM, Univ.Paris-Sud, Université Paris-Saclay, Châtenay-Malabry, France.
Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.
Allergy. 2018 Aug;73(8):1662-1672. doi: 10.1111/all.13418. Epub 2018 Feb 27.
There is in vitro evidence that T cells from allergic patients react to benzylpenicillin-human serum albumin (BP-HSA) bioconjugates. Our group has recently shown the existence of naïve CD4 T cells recognizing BP-HSA in healthy donors. However, BP-haptenated peptides from HSA participating in the immunization of allergic patients have never been identified. The purpose of the present study is to identify immunodominant BP-haptenated peptides from HSA involved in immunization of patients to BP and to refine the frequency calculation of naïve CD4 T cells recognizing BP.
Co-cultures were established with CD4 T cells from non-allergic donors and mature autologous dendritic cells (DCs) loaded with BP-HSA or BP-haptenated peptides from HSA. The CD4 T-cell response specific for BP-HSA or for individual BP-haptenated peptides was measured using an interferon-γ (IFN-γ) ELISpot assay. The frequency of BP-specific CD4 T cells was then calculated using the Poisson distribution. BP-HSA and BP-haptenated peptides recognition by allergic patients was evaluated on peripheral blood mononuclear cells (PBMCs) using a lymphocyte transformation test (LTT).
Results showed that BP-HSA and BP-haptenated peptides were recognized by naïve T cells from 15/16 and 13/14 tested healthy donors, respectively. Most donors responded to 3 peptides with BP covalently bound on lysines 159, 212, and 525. Two of these benzylpenicilloylated peptides (lysines 159 and 525) were also found to induce PBMCs proliferation in patients with allergic reaction to penicillins.
This study identifies and characterizes for the first time the BP-haptenated peptides from HSA involved in the immunization of patients to penicillins.
有体外证据表明，过敏患者的 T 细胞会对青霉素结合蛋白-人血清白蛋白（BP-HSA）结合物产生反应。我们的研究小组最近发现，健康供体中存在识别 BP-HSA 的初始 CD4 T 细胞。然而，参与致敏青霉素过敏患者的 BP 结合物的 HSA 肽段从未被鉴定过。本研究的目的是鉴定参与患者对青霉素免疫的 HSA 免疫显性 BP 结合肽，并细化识别 BP 的初始 CD4 T 细胞的频率计算。
用非过敏供体的 CD4 T 细胞与负载 BP-HSA 或 HSA 上 BP 结合物的成熟自体树突状细胞（DC）共培养。采用干扰素-γ（IFN-γ）ELISpot assay 检测针对 BP-HSA 或单个 BP 结合物的 CD4 T 细胞反应。然后使用泊松分布计算 BP 特异性 CD4 T 细胞的频率。使用淋巴细胞转化试验（LTT）评估 BP-HSA 和 BP 结合物在过敏患者外周血单核细胞（PBMC）中的识别情况。
结果表明，BP-HSA 和 BP 结合物可被 16 名/16 名和 14 名/14 名测试的健康供体的初始 T 细胞识别。大多数供体对 3 种与赖氨酸 159、212 和 525 共价结合的 BP 肽段有反应。在对青霉素过敏的患者中，这两种苄基青霉素酰化肽（赖氨酸 159 和 525）也被发现能诱导 PBMC 增殖。
本研究首次鉴定并描述了参与患者对青霉素免疫的 HSA 结合物肽段。
