板蓝根源甾类化合物靶向调控流感诱导的固有免疫过度炎症的机制研究

Emerging influenza viruses post threat to public health and chemical compounds of single target are not able to exhibit anti-inflammatory effect throughout the whole course of infection. Effective intervention to suppress cytokines storm induced by influenza virus should point to innate immune system. The traditional Chinese medicine Radix Isatidis shows advantage over chemical compounds in terms of immunomodulatory effects, which is supported by our previous studies showing both anti-viral and anti-inflammatory efficacy. We reported for the first time that steroids from Radix Isatidis inhibited RIG-I, blocked interferon generation and the subsequent signal transduction, as well as inhibited immune cells recruitment and inflammatory mediators’ expression. Thus, presumably steroids is one of the representative component in Radix Isatidis with modulatory effects on excessive immune response. The steroids probably acts on PRRs and components related to interferon signaling, resulting in suppressing cytokine storm. In this study, we will isolate strioids from Radix Isatidis, perform structural identification and optimization. Secondly, we will investigate the anti-inflammatory effects of strioids after which high-throughput sequencing and proteomics technologies will be applied to identify targets in innate immune networks. We will finally explore strioids’ effects and mechanism of action by using influenza virus infected mice model. Our study will elucidate, for the first time, the effects of strioids from Radix Isatidis in inhibiting excessive inflammatory response via regulating innate immune signaling pathways, which is in accord to the pharmacological nature of Radix Isatidis: clearing away heat and toxic material. This study will also provide valuable reference for exploration of other medicinal plant containing similar amount of strioids.

新发流感频发易造成巨大损失，化药靶点单一无法覆盖全病程抗炎治疗。控制炎症风暴关键是有效干预固有免疫致病因子。中药板蓝根比化药更具免疫调节特色，申请人证实不同成分兼具抗病毒抗炎，首次发现板蓝根源甾类含量高且可抑制模式识别受体识别(RIG-I)、阻断干扰素合成及信号转导、抑制免疫细胞募集和炎症因子等。据此推测甾类化合物是板蓝根调控过度免疫应答代表成分之一，通过干预模式识别受体启动的、与关键控制子干扰素相关的“炎症风暴”机制而抗炎。拟开展：①甾类分离提取、鉴定和结构优化；②筛选后以高通量测序及蛋白组学、代谢组学技术深入探讨流感致固有免疫炎症调控机制；③明确活性甾类化合物体内抗炎机制。将首次阐明板蓝根源甾类化合物干预固有免疫信号通路而调控流感致过度炎症机制，揭示板蓝根“清热解毒”物质基础，为其他含甾类药用植物的深度利用提供新的关键依据和借鉴。

流感是由流感病毒引起的一种多发急性呼吸道传染病，每年有超过50万人死于流感相关疾病。流感病毒感染引发宿主机体不同程度的免疫反应，机体早期大量募集炎症细胞至肺部引发过激的免疫应答是流感患者的主要免疫致病机制。本研究课题另辟蹊径从传统中医药挖掘可有效干预流感致过度固有免疫反应的药物，通过采用多种现代提取分离手段，分别从南北板蓝根中分离得到活性甾类化合物；转录组学和蛋白组学探索甾类化合物对调控病毒诱导活化固有免疫过度炎症的未知信号通路及作用机制；体内外实验深入阐明调节甾类化合物调控流感所致免疫炎症的体内外机制。最终我们从板蓝根中得到了β-谷甾醇、γ-谷甾醇、豆甾醇、过氧麦角甾醇4种具有生物学活性的甾类化合物，并且在5种常见的清热解毒中药中测定了β-谷甾醇的含量。转录组GO分析显示β-谷甾醇主要作用于单-多细胞生物、生物正向调控、细胞通讯、多细胞生物、单一生物信号等细胞过程，KEGG分析显示甾醇主要集中在细胞因子-细胞因子受体相互作用、TNF、IL-17、病毒蛋白与细胞因子及细胞因子受体的相互作用等信号通路。蛋白组Reactome富集分析提示，β-谷甾醇药理机制主要集中干扰素表达、止血、细胞凋亡等过程。本项目阐明了板蓝根源甾体化合物干扰先天免疫信号通路、调节流感引起过度炎症的机制，揭示了板蓝根“清热解毒”的物质基础，为甾体类药用植物的深入利用提供了新的关键依据。