ANGPTL3/8复合物调控NF-kB活化和炎症的功能及机制研究

NF-kB-mediated inflammation is greatly associated with many major human diseases. ANGPTL3 and 8, two members of the Angiopoietin-like protein family, are known to be involved in the regulation of lipid metabolism in the form of secreted protein complex. However, it is not clear whether such ANGPTL3/8 complex play any roles in the regulation of inflammation. Previously, supported by Youth Scientific Fund of NSFC, the applicant reported for the first time that the intracellular ANGPTL8 inhibited NF-kB activation by mediating the selective autophagic degradation of IKKg (Nature Communications, 2017). Furthermore, our pilot study demonstrated that, similar to the secreted form, the overexpressed intracellular ANGPTL3 and 8 interact with each other. Besides, ANGPTL3/8 regulated IL-1b- but not TNFa- induced NF-kB activation by targeting IL-1RAcP. The applicant thus deduced that “ANGPTL3 and 8 could regulate different molecular targets under various inflammatory stimulations, either in the complex or individual protein form”. Toll like receptors (TLRs), whose structures are similar to Interleukin 1 receptor accessory protein(IL-1RAcP), are vital receptors that mediate many pathogen associated molecular patterns (PAMPs)-induced NF-kB activation and inflammation. Therefore, by using studies at the cellular and animal levels, the applicant proposes to further investigate 1）the formation and function(s) of ANGPTL3/8 complex in the IL-1RI/TLRs signaling; 2）the mechanisms of ANGPTL3/8 in the regulation of molecular targets such as IL-1RAcP; 3）the physiological effects of Angptl3/8 in the IL-1b/TLRs ligands-induced sepsis. This study will provide new insights in the mechanisms of the crosstalk between inflammatory and metabolic signal pathways.

NF-kB介导的炎症与多种重大疾病密切相关。类血管生成素家族的两个成员ANGPTL3/8（简称A3/8）能以分泌型复合物形式调控脂质代谢，但其在炎症中功能不详。申请人在青年基金支持下首次发现胞内的A8通过介导IKKg降解抑制NF-kB活化（Nature Communications，2017）。并通过预研究发现过表达A3/8在胞内也能结合；且A3/8以复合物形式靶向IL-1RAcP调控IL-1b而非TNFa介导的NF-kB活化。申请人据此推测“A3/8能以单独或结合形式，差异调控不同的NF-kB活化途径”。鉴于TLRs介导病原体诱发的炎症反应且与IL-1RAcP结构类似，故申请人将在细胞及动物水平研究A3/8复合物①在IL-1RI/TLRs通路的形成特征及调控功能；②对IL-1RAcP等靶标的调控机制；③对Il-1b及TLRs配体诱导的败血症的影响，以期丰富代谢与炎症通路的分子交谈机制。

NF-κB介导的炎症与多种重大疾病密切相关。类血管生成素家族的两个成员ANGPTL3/8（简称A3/8）能以分泌型复合物形式调控脂质代谢，并已成为降血脂药物的重要靶标，但其在炎症中功能不详。申请人在自然科学基金青年基金项目支持下首次发现胞内的A8通过介导IKKγ降解抑制NF-κB活化（Nature Communications，2017），并通过前期研究初步发现过A3能抑制白介素（IL-1β）诱导的NF-κB活化。本项目中，申请人以A3为研究重点，探究其在NF-κB活化过程中的调控机制以及A8是否参与该调控。实验结果表明：在原代永生化肝细胞及髓系细胞中，A3能抑制IL-1β而非LPS诱导的NF-κB活化，而敲除A3则具有相反效应，且该调控主要基于胞内形式的A3。机制实验表明，A3能与白介素受体（IL1R1）及其辅助受体（IL1RAP）持续结合，且主要通过其胞内的C端FLD结构域，抑制IL1R1相关受体复合物的组装。共定位实验表明这种抑制组装的作用可能与三者共同的蛋白分选及转运相关。肝脏是一个随时受到炎症与代谢挑战的器官，A3对IL-1β诱导的NF-κB活化的这种持续抑制作用可能与肝稳态的维持相关。但目前A8对该调控的贡献还需继续研究。综上，本项目揭示了胞内A3蛋白的功能，也发现了一种差异调控不同刺激下NF-κB活化的新机制，并丰富了脂质代谢与炎症反应的分子交谈机制。