Faculty of Life Sciences, The University of Manchester, Michael Smith Building, Oxford Road, Manchester M13 9PT, UK.
Toxicology. 2013 Jul 5;309:30-8. doi: 10.1016/j.tox.2013.04.011. Epub 2013 Apr 23.
An association between protein allergenicity and resistance to pepsin digestion in the gastrointestinal tract has been proposed. However, although widely accepted, such an association is inconsistent with known labile allergens and resistant nonallergens. Given the central role of antigen presenting cells, and in particular dendritic cells (DC), in the development of allergic responses, the stability of allergens to intracellular processing may be more relevant than resistance to extracellular pepsin digestion. We have characterised the expression by DC of cathepsins (proteolytic enzymes), and compared the proteolytic activity of the most highly expressed cathepsin with pepsin for a range of 9 allergens and 4 putative nonallergens. Cathepsin expression in bone marrow-derived DC (BM-DC) derived from BALB/c strain mice was characterised by flow cytometry; cathepsins D, E and S were identified, with cathepsin D being the most highly expressed. Digestion studies revealed that the majority of allergens (5/9) were pepsin resistant, whereas non-allergens (3/4) were labile. If the generation of pepsin-resistant fragments was considered as a feature of allergenicity, this increased to 7/9 allergens and 4/4 nonallergens. In contrast, most of the proteins examined were resistant to cathepsin digestion, with significant digestion recorded for only 2/9 allergens and 2/4 non-allergens. Chemical reduction (to mimic intracellular reducing conditions) increased the susceptibility of proteins to digestion by cathepsins, but did not improve discrimination between allergens and nonallergens on this basis. These data confirm that there is a general relationship between resistance to digestion with pepsin and allergenicity. The relationship is not absolute, but the information gained from this characteristic does provide useful information in a weight of evidence approach for allergenicity assessment. The most abundant cathepsin detected in antigen processing BM-DC, cathepsin D, is not an appropriate substitute for pepsin. The hypothesis that pepsin stability may be a surrogate for stability to digestion within DC may still hold true, but consideration of a single enzyme in this context is possibly an oversimplification.
有人提出，蛋白质的变应原性与在胃肠道中抵抗胃蛋白酶消化之间存在关联。然而，尽管这种关联被广泛接受，但它与已知的不稳定变应原和抗性非变应原并不一致。鉴于抗原呈递细胞，特别是树突状细胞（DC）在过敏反应发展中的核心作用，变应原对内源性处理的稳定性可能比抵抗细胞外胃蛋白酶消化更为重要。我们已经描述了 DC 中组织蛋白酶（蛋白水解酶）的表达，并比较了一系列 9 种过敏原和 4 种假定非过敏原中最高度表达的组织蛋白酶与胃蛋白酶的蛋白水解活性。通过流式细胞术对 BALB/c 品系小鼠骨髓来源的树突状细胞（BM-DC）中的组织蛋白酶表达进行了特征描述；鉴定出组织蛋白酶 D、E 和 S，其中组织蛋白酶 D 的表达水平最高。消化研究表明，大多数过敏原（9 种中的 5 种）对胃蛋白酶具有抗性，而非过敏原（4 种中的 3 种）不稳定。如果将胃蛋白酶抗性片段的产生视为变应原性的一个特征，那么这一比例增加到 9 种过敏原中的 7 种和 4 种非过敏原。相比之下，大多数被检查的蛋白都能抵抗组织蛋白酶消化，只有 9 种过敏原中的 2 种和 4 种非过敏原中的 2 种记录到明显的消化。化学还原（模拟细胞内还原条件）增加了蛋白对组织蛋白酶消化的敏感性，但在此基础上并不能改善对过敏原和非过敏原的区分。这些数据证实，在胃蛋白酶消化抗性与变应原性之间存在普遍关系。这种关系并非绝对，但基于这种特征获得的信息确实为变应原性评估的综合证据方法提供了有用的信息。在抗原加工 BM-DC 中检测到的最丰富的组织蛋白酶，组织蛋白酶 D，并不是胃蛋白酶的合适替代品。胃蛋白酶稳定性可能是 DC 内消化稳定性的替代物的假设可能仍然成立，但在这种情况下考虑单一酶可能过于简单化。
