牛乳β-乳球蛋白过敏原表位及其潜在应用

过敏原表位是过敏原直接参与免疫反应的物质基础, 也是导致过敏反应的“元凶”。牛乳β-乳球蛋白是lipocalin家族的代表和常用模式蛋白, 它的表位信息比较完整, 拥有7个人血清IgE和6个IgG的线性表位、3个IgE结合的构象性表位以及7个T细胞表位。另外, 它的动物源血清IgE、IgG和T细胞表位也在不断被发现。牛乳β-乳球蛋白的表位信息为基于表位的应用提供了重要的支撑, 它们的应用主要涉及到预测食物过敏原的致敏性, 食物过敏的交叉反应, 食物过敏的诊断, 食物过敏的免疫治疗以及食物过敏原的检测等5个方面。     

利用细胞模型和动物模型筛选抗过敏菌株

食物过敏是机体对食物中的主要过敏原产生不正常免疫应答而导致的一种变态反应。相关研究表明食物过敏主要是由Th1/Th2细胞失衡所致,提示纠正Th1/Th2失衡可缓解食物过敏。近年来,利用益生菌特有的抗过敏作用缓解食物过敏是治疗食物过敏的新方向。但缺少筛选用于治疗或预防过敏反应的益生菌的方法,本研究利用细胞模型和动物模型筛选具有提高IFN-γ分泌水平,降低IL-4分泌水平的乳酸菌,可快速准确地筛选到具有抗过敏功能的菌株。     

肠道黏膜免疫细胞在食物致敏中的作用研究进展

食物过敏是食品安全的重要问题之一,且大部分食物过敏是在胃肠道消化过程中引发的。食物过敏原蛋白 经胃肠道消化后形成肽段,其中部分消化肽在通过肠上皮组织屏障进入肠道黏膜系统后,被递呈给相关免疫细胞, 从而引发肠道黏膜系统中一系列免疫应答反应。本文阐述了消化肽进入肠道黏膜后,肠上皮细胞、树突状细胞、B 淋巴细胞及CD4＋ T淋巴细胞在黏膜免疫致敏过程中所起的作用,为探讨食物过敏引发胃肠道免疫的工作机制提供 了理论依据。     

食物口服免疫耐受中调节性T细胞及其影响因素研究进展

食物过敏反应大致可分为致敏、激发和效应三个阶段。在致敏阶段,食品中的过敏原透过肠道黏膜免疫屏障,经抗原递呈细胞处理、T细胞活化等步骤,诱导机体产生特异性IgE并结合于肥大细胞和嗜碱性粒细胞表面;在激发和效应阶段,过敏原与IgE发生桥联反应,介导组胺、白三烯等免疫介质释放,它们与效应器官上的相应受体结合,会引起局部或全身过敏反应。食物过敏发生的根本原因是部分个体对特定过敏原口服免疫耐受的破坏,而诱导建立和重建口服免疫耐受是应对食物过敏较有前景的方法,或可突破该病目前尚无有效治愈方法的瓶颈。本文就口服免疫耐受建立和重建中起核心作用的调节性T细胞以及其密切相关因素如树突状细胞、调节性B细胞、肠道菌群及其代谢产物进行综述,以期为食物过敏的防治工作提供帮助。     

热处理对食物致敏性的影响及其体外细胞学评价

综述了热加工处理对典型食物过敏原致敏性的影响,重点介绍了体外细胞学评价方法在评价食物过敏原致敏性变化方面的一些研究进展。人们生活方式和环境的改变,以及日常食物丰富所致过敏诱发因素的增加,食物过敏发生比例大幅提高,食品过敏问题已成为目前国内外食品安全研究的热点之一。食品热加工可以改变食物过敏原的蛋白表位结构、聚合特性以及过敏原诱发淋巴细胞释放细胞因子的特性。由于食品基质、配料和处理过程的差异,加工过程中食品致敏性的变化尚无普遍规律可循。通过对典型食物过敏原的分离纯化、分子鉴定,可以追踪过敏原组分在食物加工过程中的改变规律。目前过敏效应分子IgE抗体的水平变化常常被作为致敏性评价的指标,酶联免疫分析和免疫印迹试验是常用的测定手段。然而,对于过敏反应阶段由IgE抗体与效应细胞上FcεR受体之间的结合引发系列炎症反应的部分,则无法通过这些方法进行判断。全面评价食物过敏原的致敏效应不但需要监测IgE抗体水平,也需要监测效应细胞致敏程度,以及释放的细胞因子水平。     

具有T细胞口服免疫耐受作用的β-乳球蛋白水解物的制备及鉴定

以牛乳中主要过敏原β-乳球蛋白为研究对象,制备和鉴定具有T细胞表位和口服免疫耐受性的水解物,旨在为牛乳过敏患者口服免疫治疗方法提供理论依据。采用生物信息学方法预测β-乳球蛋白的T细胞表位,通过质谱分析6种蛋白酶水解β-乳球蛋白水解物的氨基酸序列,用T细胞增殖实验鉴定水解物中多肽免疫耐受作用,体内实验测定小鼠血清特异性抗体(IgE、IgG₁、IgG_2a)、Th1细胞因子(IFN-γ、IL-17)、Th2细胞因子(IL-4、IL-5、IL-13)、组胺、几丁质酶-3样蛋白1的水平及小鼠脾脏细胞亚群的变化水平,验证β-乳球蛋白水解物的口服免疫治疗活性。研究结果表明:胰蛋白酶、复合蛋白酶和木瓜蛋白酶水解物具有口服耐受性,其中复合蛋白酶和木瓜蛋白酶水解物具有口服耐受性是创新性发现。中性蛋白酶水解物虽然含有T细胞表位,但是仍然具有致敏性。因此含有T细胞表位的水解物不一定具有口服免疫耐受性,需要体内实验验证。研究结论以期为新型抗过敏乳基料的开发和临床治疗牛乳过敏提供参考 数据。     

乳酸菌降低过敏食物致敏性的研究进展

食物过敏的患病率在近几十年内急剧上升,越来越多的证据表明机体内微生物的生态失调是引起食物过敏的主要原因,因此乳酸菌等益生菌对过敏症状的潜在预防和治疗效果引起了人们的广泛关注。大量研究表明乳酸菌不仅能够通过发酵降低食物过敏原的致敏性,而且有可能通过调节肠道菌群的平衡在宿主免疫系统的发育和调节中发挥作用,从而改变过敏性疾病的发病风险。本文综述了乳酸菌的分类和生物特性,乳酸菌发酵在降低牛奶、大豆、花生、小麦等主要食物过敏原中发挥的作用,乳酸菌发挥免疫调节作用的主要机制,包括改变抗原递呈细胞的表型、调节Th1/Th2平衡、诱导调节性T细胞、增强肠道屏障功能。以期通过乳酸菌及其发酵作用达到预防和/或治疗食物过敏的效果,为乳酸菌的研究及应用提供参考。     

免疫低下和食物过敏小鼠外周血中细胞因子的变化

建立小鼠免疫低下模型和食物过敏模型,采用ELISA法检测外周血中细胞因子(IL-4、IL-6、IL-10、IL-12、IFN-γ、TGF-β)的变化。结果表明：与空白对照组比较,腹腔注射环磷酰胺所建立免疫低下小鼠胸腺指数和脾脏指数均有显著降低(P≤0.05),且在第3天达到最低;经卵清蛋白致敏小鼠外周血IgE水平极显著升高(P≤0.01)。免疫低下小鼠外周血中IFN-γ(除第3天外)、TGF-β含量(除第3天外)和IFN-γ/IL-4比值均显著降低,而IL-10水平也有下降趋势;除第2天外IL-6水平显著升高,但IL-4水平只在第3天显著性增高,IL-12先是下降随后呈现增多的趋势;食物过敏小鼠的IL-4、IL-6、IL-10和TGF-β水平均有显著性增高,IFN-γ/IL-4比值有显著性下降,而IFN-γ和IL-12没有明显的变化。因此,免疫低下和食物过敏小鼠的Th1/Th2细胞平衡均向Th2细胞偏移,免疫低下对Th1细胞的影响大,食物过敏对Th2细胞的影响更明显。     

食物致敏原表位定位技术的研究进展

食物致敏原是引起食物过敏的元凶，多为蛋白质。抗原表位是在抗原分子中与抗体反应或被抗原受体识别，并引发机体免疫应答的特殊化学基团。从表位水平认识食物致敏原，能揭示食物过敏的物质基础，为解决食物过敏问题提供精准的靶标。本文基于表位结构和定位方法的不同，介绍了食物致敏原表位定位技术的发展，并进一步展望了致敏原表位信息对于改进食物过敏鉴定技术和低致敏食物加工方法的应用前景。     

肠道菌群与婴幼儿食物过敏相关性的研究进展

随着婴幼儿食物过敏的发病率逐年上升,食物过敏逐渐成为了全球性公共卫生问题。大量研究表明人体内肠道菌群与食物过敏有着密切的关系,在受到外界因素的干扰后,肠道菌群多样性和数量发生改变,进而引发机体对某些正常抗原的免疫耐受度降低,导致过敏反应。本文介绍了食物过敏婴幼儿和健康婴幼儿肠道菌群的差别,对肠道菌群在食物过敏中的调节机制(影响肠道细胞功能和调节屏障保护特性、产生信号分子进行免疫调控以及促进和维护机体口服耐受)进行综述,并对肠道菌群调节婴幼儿食物过敏的策略进行展望。     

免疫耐受在牛乳过敏中的研究进展

食物过敏是机体对过敏原产生的一种不良免疫反应,而免疫耐受是指机体对摄入的食物抗原不产生此类免疫应答。近年来,牛乳过敏的发生率越来越高,大部分牛乳过敏的儿童3岁之后出现了免疫耐受。基于此,介绍了牛乳过敏及引发免疫耐受的相关免疫细胞包括树突状细胞、调节性B细胞、调节性T细胞在免疫耐受中的作用及3种诱导免疫耐受的方法。     

TBa过敏原表位区段的融合表达及免疫活性分析

为了确定苦荞过敏原TBa(tartarybuckwheatallergen)的抗原表位及进一步了解荞麦过敏反应机制,本实验以苦荞过敏原TBacDNA序列为模板,设计引物,克隆苦荞过敏原TBa表位区段基因,分别构建TBa及两个表位区段原核表达载体,在大肠杆菌BL21(DE3)中进行表达并纯化,采用竞争ELISA对其免疫活性进行分析与比较。SDS-PAGE及WesternBlot鉴定和检测结果表明,目的蛋白在E.coliBL21(DE3)中可高效表达,其N端带有6个组氨酸标签。Ni2+-NTA琼脂糖柱亲和纯化得到了纯度较高的目的片段。ELISA实验结果显示,表达产物与荞麦食品过敏病人血清中的IgE具有特异的结合活性。本研究为揭示苦荞过敏蛋白结构与功能的关系奠定了基础。     

T lymphocytes and food allergy

Food allergy is a hypersensitivity reaction to normally harmless substances and involves humoral immune responses, mediated by immunoglobulin (IgE) synthesized by B lymphocytes, and cellular immune responses mediated by T lymphocytes. An IgE-mediated mechanism leads to clinical symptoms occurring immediately after food ingestion, e. g., "the oral allergy syndrome". For delayed reactions involving the gastrointestinal tract or the skin, the underlying immune mechanisms are less clear. In order to elucidate the cellular response to food allergens, human allergen-specific T cell cultures generated in vitro represent helpful tools. The majority of food allergen-specific CD4(+) T lymphocytes isolated from food-allergic individuals was found to synthesize high levels of IL-4 and IL-13, two cytokines required for initiation of IgE synthesis. Due to selective homing profiles, food-specific T cells seem also to be involved in defining the target organ of the allergic inflammation. Recent data provide evidence that in addition to IgE-mediated inflammation, food allergen-specific T lymphocytes may also cause inflammatory responses independently of IgE-mediated mechanisms.     

牛乳乳清中主要过敏原的B细胞表位定位

采用噬菌体展示技术结合生物信息学定位牛乳α-乳白蛋白和β-乳球蛋白的B细胞表位,分析不同类型表位之间的氨基酸组成和空间分布规律。结果表明,某些线性表位是构象性表位的组成部分,IgE与IgG表位存在共同区域。本研究得到的表位是牛乳过敏诊断和预后的候选生物标志物,也可用于指导于低致敏性乳制品的开发。     

Citrus Tachibana Leaf Extract Mitigates Symptoms of Food Allergy by Inhibiting Th2‐Associated Responses

Although the incidence of food allergy continues to rise, there have been no effective therapeutic strategies. Citrus fruits contain a number of bioactive flavonoids with immune‐regulatory functions. The objective of this study was to determine whether Citrus tachibana (fruit body with peel, leaves, and branch) can protect against the development of food allergy and the mechanism behind it, and to identify the active compound(s) responsible. We found that C. tachibana leaf extract (CLE) mitigated ovalbumin (OVA)‐induced food allergy symptoms including increased rectal temperature, diarrhea, and anaphylaxis. This mitigation was likely due to CLE‐mediated decreases in cytokine release from T‐helper 2 cells (Th2 cells) in mesenteric lymph nodes. Moreover, higher levels of CLE attenuated systemic Th2 cell–mediated responses in mouse splenocytes sensitized with OVA+Alum. This was evidenced by CLE‐mediated reductions in Th2 cytokine release, including interleukin (IL)‐4, IL‐5, and IL‐13, but not the Th1 cytokines IL‐12 and interferon (IFN)‐γ, which was attributable to decreased gene expression levels. We also identified kaempferol as the most potent compound for reducing Th2‐associated responses in splenocytes. The findings of this study suggest that CLE suppresses Th2‐cell–mediated immune responses, contributing to alleviation of food allergy symptoms, and that kaempferol is a flavonoid with potential antiallergenic activity that targets Th2 cell–induced responses.     

Maillard reaction in food allergy: Pros and cons

Food allergens have a notable potential to induce various health concerns in susceptible individuals. The majority of allergenic foods are usually subjected to thermal processing prior to their consumption. However, during thermal processing and long storage of foods, Maillard reaction (MR) often takes place. The MR is a non-enzymatic glycation reaction between the carbonyl group of reducing sugars and compounds having free amino groups. MR may sometimes be beneficial by damaging epitope of allergens and reducing allergenic potential, while exacerbation in allergic reactions may also occur due to changes in the motifs of epitopes or neoallergen generation. Apart from these modulations, non-enzymatic glycation can also modify the food protein(s) with various type of advance glycation end products (AGEs) such as N?-(carboxymethyl-)lysine (CML), pentosidine, pyrraline, and methylglyoxal-H1 derived from MR. These Maillard products may act as immunogen by inducing the activation and proliferation of various immune cells. Literature is available to understand pathogenesis of glycation in the context of various diseases but there is hardly any review that can provide a thorough insight on the impact of glycation in food allergy. Therefore, present review explores the pathogenesis with special reference to food allergy caused by non-enzymatic glycation as well as AGEs.     

Suppression of immune responses to β‐lactoglobulin in mice by the oral administration of peptides representing dominant T cell epitopes

BACKGROUND: The significance of oral tolerance in the treatment of adverse immune reactions such as allergic and autoimmune diseases has been noted. In the present study, peptides that could effectively induce oral tolerance to bovine β‐lactoglobulin (BLG), a milk allergen, were investigated in a murine model. RESULTS: The oral administration of peptides corresponding to the T cell epitope regions of BLG, i.e. p42–56, p62–76 and p139–154, apparently down‐regulated T cell proliferation to BLG. The in vitro cytokine production by the lymph node cells from the peptide‐fed mice cultured in the presence of the antigen was also analysed. It was found that p62–76 and p139–154 feeding suppressed the production of both Th1 and Th2 types. Interestingly, p139–154 feeding suppressed both T cell and antibody responses to BLG. Additionally, p139–154 feeding diminished BLG‐specific IgE and IgG1 antibody responses. CONCLUSION: The unique tolerogen peptide p139–154 that could suppress both T and B cell responses to BLG in a murine model was identified. These findings can be useful for the selection of an optimum tolerogenic peptide to prevent and treat milk and other food allergies. Copyright © 2007 Society of Chemical Industry     

虾制品致敏性诱发豚鼠相关细胞因子及Th1/Th2细胞平衡的变化

以消减致敏性南美白对虾的虾仁、虾肉和虾蛋白为样品,以pH 7.5的磷酸盐缓冲液为阴性对照,未处理的虾蛋白为阳性对照,建立豚鼠过敏模型,研究过敏豚鼠血清中相关细胞因子与食物过敏的相关性,并推断食物过敏对辅助性T（type 1/type 2 T-helper, Th1/Th2）细胞平衡的影响。收集过敏豚鼠的血清,采用酶联免疫吸附测定法检测血清中免疫球蛋白E（immunoglobulin E,IgE）、组胺（histamine,HIS）、相关细胞因子（白细胞介素（interleukin,IL）-1、IL-2、IL-3、IL-4、IL-6、IL-10、肿瘤坏死因子-α（tumor necrosis factor α,TNF-α）、干扰素-γ（interferon-γ,IFN-γ）质量浓度。结果表明,用未处理的虾蛋白致敏豚鼠,用致敏性消减程度不同的虾制品（虾蛋白、虾肉、虾仁）提取的蛋白激发,消减致敏性的虾蛋白、虾肉和虾仁提取蛋白激发后豚鼠血清中IgE含量分别为（3.905±0.120）、（4.813±0.188）、（5.199±0.327）U/mL,HIS质量浓度分别为（16.437±1.120）、（19.656±1.080）、（21.071±1.732）μg/mL,激发后血清中IgE和HIS质量浓度变化与致敏性程度呈正相关,致敏性越低,血清中IgE和HIS质量浓度越低。过敏豚鼠血清中IL-1、IL-2、IL-3、IL-4、IL-6、TNF-α质量浓度的变化与虾制品致敏程度呈正相关;同时,与IgE和HIS质量浓度变化规律具有一致性;血清中IL-10质量浓度变化与致敏性程度呈负相关;因此,这些细胞因子与食物过敏具有很好的相关性。过敏血清中IFN-γ没有呈现规律性变化,但IFN-γ/IL-4随着致敏性的增强而减小。因此,推测虾制品激发过敏豚鼠的Th1/Th2细胞平衡向Th2细胞偏移。