IL-35通过GCN2-CHOP/mTORC1信号网络调控低氧状态下氨基酸代谢微环境介导的Treg细胞分化异常：ARDS失控性炎症的一个潜在治疗靶点

IL-35 can protect ARDS by regulating uncontrollable inflammation, and IL-35 is an important cytokine required to regulate the inflammatory suppressive function of Treg cells. Our previous studies showed that amino acid metabolism microenvironment, such as L-arginine decreased, can affect the Treg cell differentiation in ARDS. And we had illustrated that IL-35 can promote the differentiation from Naïve CD4 T cells into Treg cells. Simultaneously, the GCN2-CHOP/mTORC1 signaling network is one of the key regulatory element of T cell differentiation during arginine deficiency. Based on theories and results from the above, we proposed a scientific hypothesis that IL-35 could initiate protein translation and reduce cell death through regulation of the GCN2-CHOP/mTORC1 signaling network, and then promote Treg cells differentiation and suppress uncontrolled inflammatory reaction of ARDS through regulating the GCN2-CHOP/mTORC1 signaling. Naïve CD4 T cells, pulmonary vascular endothelial cells and other knockout mouse models, including IL-35(KO) mice, GCN2(KO) mice and B-NSG mice were used; adenovirus vector RNA interference, amino acid starvation, cells co-culture and adoptive transfusion of T cells were carried out to explore a new mechanism of ARDS, which could contribute to use the Treg cells as the metabolic check point to provide a promising strategy for identifying potential therapeutic targets of ARDS.

IL-35可通过调节失控性炎症对ARDS发挥保护作用，是调控Treg细胞炎症抑制功能所需细胞因子。我们前期研究表明：ARDS时氨基酸代谢微环境(L-精氨酸降低)影响Treg细胞分化，而IL-35能促进NaïveCD4T细胞向Treg细胞分化；且GCN2-CHOP/mTORC1信号网络是精氨酸缺乏时T细胞分化关键调控环节。基于此，我们提出“IL-35通过调控GCN2-CHOP/mTORC1信号网络，启动蛋白质翻译、减少细胞死亡，促进Treg细胞分化，抑制ARDS失控性炎症”的科学假说，并拟以NaïveCD4T、肺血管内皮等细胞(株)和IL-35(KO)、GCN2(KO)、B-NSG等基因敲除小鼠模型，腺病毒载体RNA干扰、氨基酸饥饿、细胞共培养、T细胞过继回输等技术与方法，从分子-细胞-动物-临床整体论证并阐明ARDS新机制，为以Treg细胞作为代谢检测点设计ARDS潜在治疗靶标提供新方向。

失控性炎症反应是引起ARDS中心环节。IL-35可通过调节失控性炎症对ARDS发挥保护作用，是调控Tregs炎症抑制功能所需细胞因子。课题组前期工作表明：IL-35能促进NaiveCD4T细胞向Tregs分化；ARDS代谢微环境（低Arg）影响Tregs分化，GCN2-CHOP/mTORC1信号网络是Arg缺乏时T细胞分化关键调控环节。提出“IL-35通过调控GCN2-CHOP/mTORC1信号网络，启动蛋白质翻译、减少细胞死亡，促进Tregs分化，抑制ARDS失控性炎症”科学假说。在项目资助下，课题组研究IL-35在ARDS中的作用及相关分子机制，通过4年工作：①阐明ARDS时IL-35异常表达、代谢微环境改变与Tregs分化间的关系，以及对ARDS发生发展的作用；②研究并揭示ARDS时，IL-35通过GCN2-CHOP/mTORC1等多种炎症代谢信号网络，调控Tregs分化异常，影响ARDS的分子机制；③通过naiveT细胞分化诱导模型，揭示ARDS时IL-35在Tregs命运决定中的调控作用。重要研究结果包括：确定IL-35是ARDS重要炎症靶标，对ARDS具重要保护性作用；证明提高CD4+/CD4+CD25+Foxp3+Tregs比例是IL-35发挥治疗性作用潜在机制；抗炎类天然化合物治疗ARDS与提高naiveCD4+T细胞向的Tregs分化，促进STAT5和IL-35表达有关，“IL-35/Tregs”是ARDS炎症调控关键环节；IL-35能通过促进naïveCD4+T细胞分化为Foxp3+Helios+Tregs来减轻ARDS，从而为抗ARDS治疗提供了一种新工具。确定并筛选出“D-精氨酸和D-鸟氨酸代谢”差异性代谢途径；低氧代谢影响Treg细胞分化，IL-35通过调控炎症代谢信号网络（GCN2-CHOP/mTORC1、JAK/STAT、NF-κB等）促进向Treg细胞分化；IL-35不能诱导低L-Arg条件naïveCD4+T细胞分化为Tregs；IL-35能通过JAK/STAT通路促进GS表达，抑制GSH从头合成，上调Tregs。本项目实施从转化医学角度阐述“以Tregs为代谢检测点设计ARDS新的潜在治疗靶标（IL-35/Tregs）”的问题。项目执行期间，主要发表 S C I 论文6篇，授权国家发明专利1项。