Wheat sensitivity: Impact of wheat varieties and wheat growing conditions on innate immunity

非乳糜泻小麦敏感症(NCWS)是指非乳糜泻或小麦过敏引起的对摄入小麦的免疫反应。NCWS影响2%到10%的小麦消费对象，是由对小麦蛋白质的先天免疫反应引起的。最近，我们发现小麦淀粉酶-胰蛋白酶抑制因子(ATIS)家族是天然免疫的触发物。小麦ATI激活Toll样受体4(TLR4)-MD2-CD14复合体，导致树突状细胞和巨噬细胞单核细胞释放促炎细胞因子，并促进树突状细胞的成熟。相关的TLR4刺激活性仅限于含面筋的谷物，如小麦、黑麦和大麦。值得注意的是，我们证明营养性ATI以剂量依赖的方式加重实验性自身免疫和变态反应性疾病。因此，测定当前使用和替代的小麦品种的ATI含量、ATI生物活性和ATI基因调控对人类健康具有很高的相关性。到目前为止，只有一个小麦品种进行了广泛的转录组和蛋白质组分析，表明ATIS占总蛋白的2-4%，面筋蛋白占总蛋白的80%。此外，19个DNA区域编码了潜在的ATI。由于小麦基因和蛋白的表达高度依赖于小麦品种和生长条件，因此迫切需要对更多的小麦品种在不同条件下生长进行研究，以更好地了解ATI的表达规律。这将允许生产更健康的小麦产品，防止由于小麦敏感而导致的发病率增加和慢性炎症性疾病的恶化。我们的建议结合生物医学、小麦育种和小麦生长研究以及遗传学和蛋白质组学的专业知识，研究小麦ATI的作用和调节，以及它们在体外和体内对先天和适应性免疫的影响。包括现代优良品系和重要的老品种在内的160个小麦品种将在德国的3个不同的田间地点种植。收获的样品将接受小麦育种者定期选择的品质性状的检查，以ATI表达为重点的高级蛋白质组和遗传分析，以及基于细胞的体外TLR4刺激活性。选定的样本将用于活体喂养研究，研究食物基质对肠道和一般ATI诱导的先天性和获得性免疫激活的影响。结合深入的统计分析，这些数据将使我们能够表征在不同条件下生长的各种小麦的ATI表达和生物活性，以选择ATI表达和生物活性低的变异体和条件。这可能会促进免疫原性较低、因此更健康的小麦品系的生长。

Non-celiac wheat sensitivity (NCWS) is defined as an immunological reaction to ingestion of wheat that is not caused by celiac disease or wheat allergy. NCWS affects between 2% and 10% of wheat consuming subjects and is caused by an innate immune response to wheat proteins. Recently, we identified the family of wheat amylase-trypsin inhibitors (ATIs) as triggers of innate immunity. Wheat ATIs activate the toll like receptor 4 (TLR4)-MD2-CD14 complex, lead to the release of proinflammatory cytokines in dendritic cells>macrophages>monocytes and to the maturation of dendritic cells. Relevant TLR4 stimulating activity is confined to ATIs of gluten containing grains like wheat, rye and barley. Notably, we demonstrated that nutritional ATIs exacerbate experimental autoimmune and allergic diseases in a dose-dependent manner. Therefore, determination of ATI content, ATI bioactivity and of ATI-gene regulation in currently used and alternative wheat variants should be of high relevance to human health. To date, only a single wheat variety had been subjected to extensive transcriptome and proteome analysis, showing that ATIs represented 2-4% and gluten proteins >80% of total protein. Moreover, 19 DNA regions encoded potential ATIs. Since the expression of wheat genes and proteins is highly dependent on the wheat variety and growing conditions, studies with larger numbers of wheat varieties grown under different conditions are urgently needed, to better understand the regulation of ATI expression. This should allow the production of healthier wheat products, preventing enhanced morbidity due to wheat sensitivity and worsening of chronic inflammatory diseases.Our proposal combines the expertise of biomedicine, of wheat breeding and wheat growing research, and of genetics and proteomics, to study the role and the regulation of wheat ATIs and their in vitro and in vivo effects on innate and adaptive immunity. 160 wheat varieties comprising modern elite strains and important old varieties will be grown at 3 different field locations in Germany. The harvested samples will be subjected to examination of quality traits regularly selected by wheat breeders, advanced proteomic and genetic analyses focused on ATI expression, and to cell based in vitro TLR4 stimulatory activity. Select samples will be employed for in vivo feeding studies, addressing food matrix effects on intestinal and general ATI-induced innate and adaptive immune activation. Combined with thorough statistical analyses, the data will enable us to characterize ATI expression and bioactivity in a wide variety of wheats, grown under different conditions, to select for variants and conditions with low ATI expression and bioactivity. This will likely promote the growing of less immunogenic and therefore healthier wheat strains.