MDH2启动子区域低甲基化促进儿童食物过敏的机制研究

Hypersensitivity of the immune system to environmental agents such as pollen and food can cause allergic diseases, which collectively affect over one third of infants and children. We found CD14+ monocytes from cord blood of infants who developed food allergy secreted more inflammatory cytokines IL-1b, IL-6 and TNF-a upon stimulation with LPS, compared to those who did not develop food allergy. These inflammatory cytokines and the mucosal-derived cytokine TGF-b suppress IL-2 expression by CD4+ T cells, which promote the expression of large amount of IL-4 in T cells and lead to the development of IgE-mediated food allergy in children. To identify epigenetic mechanisms that govern immune deviation in the cord blood, we employed 450K methylation analysis on cord blood immune cells isolated from 5 food-allergic and 5 non-allergic infants. Promoter methylation at malate dehydrogenase 2 (MDH2) locus was significantly decreased in CD14+ monocytes in food-allergic infants. We hypothesize that promoter demethylation increases MDH2 expression and malate-aspartate shuttle (MAS), which in turn increases aerobic glycolysis, hypoxia-inducible factor (HIF1a) and inflammatory cytokine expression. We will: 1) validate whether MDH2 demethylation is a potential biomarker for food allergy, and 2) investigate whether MAS-HIF1a regulate inflammatory cytokine expression. This project will reveal potential novel early diagnostic biomarkers and mechanisms for allergic disease development in infants, and provide targets for therapy.

食物过敏是严重影响婴幼儿健康成长的重大医疗问题。我们已发表工作(Zhang et al, Sci Transl Med)表明CD14+单核细胞过强的炎症因子释放能力是诱发疾病的主要原因。但是导致单核细胞炎症应激能力升高的机制目前仍不明确。通过表观基因组和转录组筛选，近期预实验结果显示：食物过敏儿童单核细胞的苹果酸脱氢酶2（MDH2）的启动子发生了显著的去甲基化现象，MDH2表达增加，葡萄糖代谢通路活化。 那么，MDH2的启动子甲基化如何调控基因表达？ MDH2是否通过葡萄糖代谢调节炎症因子的分泌？我们将通过大量临床标本并结合MDH2功能抑制和基因敲除实验：（1）验证MDH2启动子甲基化与食物过敏的关系，（2）阐述启动子甲基化调控MDH2基因表达的机制，（3）明确MDH2调控炎症因子分泌的信号通路 。这将增加对婴幼儿过敏性疾病发病机制的了解，为疾病的早期预警和治疗提供新的分子标志物和靶标。

食物过敏是严重影响婴幼儿健康成长的重大医疗问题。我们已发表工作表明CD14+单核细胞过强的炎症因子释放能力是诱发疾病的主要原因。但是导致单核细胞炎症应激能力升高的机制目前仍不明确。本项目原拟验证MDH2过量表达通过增强苹果酸－天冬氨酸穿梭提高CD14+单核细胞的炎症应激能力促进食物过敏的科学假设。实验结果表明敲低MDH2降低单核/巨噬细胞的炎症反应能力。体内实验发现Mdh2全身敲除小鼠胚胎致死，而Mdh2+/-小鼠则表现出自发肥胖和肝脏巨噬细胞炎症因子分泌能力降低的现象。机制研究发现敲低MDH2表达促进脂滴合成，导致肝脏和皮下脂肪增加；但敲低MDH2却可以抑制炎症因子分泌，降低胰岛素耐受和非酒精性肝炎的发生（待发表）。本研究为MDH2相关肥胖及炎症相关疾病的治疗提供了理论依据。..除此之外，我们系统阐述了儿童肠道炎症（包含食物过敏）和新生儿肝脏自身免疫损伤的发生机制。我们发现cAMP第二信使通路缺失是导致儿童肠道炎症的共性机制，因此提出并验证了磷酸二酯酶抑制剂治疗肠道炎症的临床应用价值 (Cell, 2019)。我们发现东亚人群高发的NLRP3-R779C突变通过与去泛素化酶JOSD2结合而导致自发炎症 (JACI, 2021)。另外，我们阐述了围产期感染导致新生儿自身免疫性肝衰竭的机制，发现抑制B细胞活化可以有效改善肝脏自身免疫损伤（Cell, 2020 ）。..总之，在该项目的资助下，我们发表标注项目经费号论文11篇，包括Cell （2019，2020），Science Immunology （2021），Journal of Allergy & Clinical Immunology （2021， 2018）等。获得“胃肠道疾病的免疫机制及治疗药物”专利一项并开始进行专利转化工作，获得国家和地方级人才项目3项，国家及省部级各类项目17项，基本完成本项目的主要研究内容。