Notch1/Hes1-IL17A正反馈环路促Th17细胞分化介导银屑病发病机制的研究

IL-17A is the important pathogenic factors of psoriasis by mediating immune dysfunction and inflammatory effects. Our previous studies have found that expression levels of Notch1 and IL-17A were significantly increased and positively correlated in psoriatic patients. Notch1 inhibitor DAPT can alleviate the severity of psoriasis-like skin inflammation in mice, and decrease the expression and secretion of IL-17A. IL-17A intervention can also affect the expression of Notch1 target gene Hes1 in model mice. While, the underlying regulatory mechanism between Notch1 and IL-17A has not been fully clarified. Combined with our research results and literature reports, we hypothesize that IL-17A may active Notch1 signal and promote Hes1 expression through Act1, then inhibit the negative-regulatory factor PTEN to activate PI3K/Akt/mTORC1 pathway and promote Th17 cell differentiation and IL-17A secretion in a positive feedback loop, which further exacerbates the pathogenic effect of IL-17A in psoriasis. We will explore the mechanism of Notch1/Hes1-IL17A positive feedback loop by molecular and immunological technologies, and validate its effect on promoting Th17 cell differentiation to mediate the pathogenesis of psoriasis through genetic manipulation and pharmacological intervention of the key components in the cascade. Our study will provide new strategy and scientific data for the treatment of psoriasis by targetedly suppressing Notch1/Hes1-IL17A feedback loop.

IL-17A是导致银屑病免疫紊乱和炎症反应的重要致病因素。课题组前期研究发现，银屑病患者Notch1与IL-17A高表达且呈正相关，阻断Notch1信号可减轻模型鼠皮损、降低IL-17A表达；IL-17A干预亦影响模型鼠Notch1靶基因Hes1的表达，但Notch1与IL-17A间调控机制未明。结合文献资料，课题组提出假说：IL-17A通过Act1激活Notch1信号促Hes1表达，进而抑制PTEN并激活PI3K/Akt/mTORC1通路，促进Th17细胞分化及IL-17A分泌形成正反馈环路，加剧IL-17A在银屑病的免疫损伤作用。本课题拟通过分子生物学和免疫学技术探讨Notch1/Hes1-IL17A正反馈环路的分子机理，对信号轴关键组分进行基因修饰和药物干预，在体内外评估其促IL-17A分泌介导银屑病的致病效应，以探求靶向干预Notch1/Hes1-IL17A环路治疗银屑病的新策略。

背景：Th17细胞效应性细胞因子IL-17A是银屑病发病机制中的核心因素。Notch1 信号通路与 PI3K/AKT信号通路均参与Th17细胞分化及IL-17A分泌的调控。近来研究显示Notch1靶基因Hes1能够负向调控PI3K/AKT上游负向调控因子PTEN，项目组前期研究发现IL-17A信号通路接头蛋白Act1可以与Notch1的胞内功能域NICD1结合形成Act1-NICD1复合物，并激活Notch1信号通路，因此提出Notch1/Hes1-PI3K/AKT-IL17A正反馈环路，并探讨该正反馈环路在银屑病病理条件下对Th17细胞分化及IL-17A分泌的调控。主要研究内容：通过分子生物学及免疫学实验技术探讨Notch1/Hes1-PI3K/AKT-IL17A正反馈环路构成的分子机理，对反馈环路关键节点信号分子Notch1和AKT进行药物干预，以评价该正反馈环路在银屑病病理环境下的致炎效应。重要结果及关键数据：1.IL-17A剂量依赖性及时间依赖性促Th17细胞分化，Notch1、Hes1、AKT、mTORC1、S6K1、S6K2及IL-17A 等关键节点信号分子mRNA表达水平与Th17细胞比例变化趋势一致，PTEN mRNA表达水平与Th17细胞比例变化趋势相反。2.PI3K/AKT抑制剂LY294002体内实验显示LY抑制剂能够明显减轻银屑病样皮炎小鼠皮损及组织学炎症反应、脾脏及淋巴结肿大，降低皮损组织中Th17细胞比例以及各关键节点信号分子表达水平；体外实验证实LY能够时间依赖性降低模型鼠脾脏单个细胞中上述关键节点信号分子的表达水平。3.LY抑制剂显著增加E3泛素连接酶c-Cbl表达量，溶酶体抑制剂氯喹明显减轻LY抑制剂对正反馈环路各关键节点信号分子的抑制效应，并显著降低c-Cb1表达量，LY294002抑制剂经溶酶体途径增加Notch1单泛素化及降解。4.Notch1/Hes1抑制剂DAPT体内实验显示DAPT能够明显减轻银屑病样皮炎小鼠皮损及组织学炎症反应，降低皮损组织中Th17细胞及γδT17细胞比例以及Hes1, AKT, mTORC1及 IL-17A的表达水平。科学意义：银屑病病理条件下，Notch1及PI3K/AKT信号过度活化并正反馈环路，为银屑病免疫靶向治疗提供新的理论基础和实验基础，阻断该环路可能成为银屑病免疫治疗的潜在靶点。