基于Kupffer细胞Exosome传输miRNA探索ACLF发病机制及解毒化瘀颗粒的调控研究

Acute-on-chronic liver failure(ACLF) shows a high incidence and situation on prevention and treatment is severe. At present, the pathogenesis of ACLF is not clear. Kupffer cells, which are used to colonize the liver, are closely related to the pathogenesis of various types of liver diseases, and it has been proved that Kupffer cells can exert immune effects through the Exosome pathway. The preliminary project group has confirmed the efficacy of Jiedu Huayu Granule on the treatment of ACLF. Accordingly, the project team speculated that Kupffer cells participated in the pathological process through transmission of miRNA by Exosome patheay and Jiedu Huayu Granule could regulate the process to play a therapeutic role. To this end, the project aims to reveal the function, mechanism and target of regulation of Kupffer cells on Exosome transmission of miRNA in the pathological process of ACLF from three levels of molecular, cellular and animals in order to reveal 3 Kupffer cell Exosome transmission miRNA play in the ACLF pathological process and the regulation of the target through constructing ACLF rat model and using advance technology like miRNA chip, RNA interference and laser confocal microscope and so on. The completion of the project will be a new perspective to clarify the immune effect of Kupffer in the ACLF process from the Exosome approach and provide a scientific basis for the treatment with Traditional Chinese Medicine of this disease.

慢加急性肝衰竭（ACLF）呈高发病态势，防治形势严峻。目前 ACLF发病机制尚有待阐明。作为肝脏定植巨噬细胞的枯否细胞（Kupffer cell）与各类肝病发病密切相关，已有研究证实Kupffer细胞可通过外泌体（Exosome）途径发挥免疫效应。前期项目组已证实解毒化瘀颗粒治疗ACLF的疗效肯定。据此，项目组推测Kupffer细胞可通过Exosome途径传输miRNA参与ACLF发病进程，解毒化瘀颗粒可调控该进程发挥治疗作用。为此，该项目组拟通过构建ACLF大鼠模型，采用miRNA芯片、RNA干扰、激光共聚焦显微镜等先进技术，从分子、细胞及动物3个层面次第揭示Kupffer细胞Exosome传输miRNA在ACLF疾病进程中发挥的作用和机制及该方的调控靶位。项目的完成将从Exosome途径这一新视角来阐明Kupffer细胞在ACLF中发挥的免疫致病效应；也为中医药治疗本病提供科学依据。

慢加急性肝衰竭（ACLF）多由慢性肝炎发展而来，其进展迅速，病情危重，预后不良，病死率极高，目前ACLF发病机制尚有待阐明。Kupffer细胞是肝脏固有免疫系统的一个重要组成部分，具有很高的可塑性。已有研究证实Kupffer细胞可通过外泌体（Exosome）途径发挥免疫效应。项目组通过构建ACLF大鼠模型，解毒化瘀颗粒干预处理，提取Kupffer细胞Exosome，采用miRNA测序技术、mRNA测序技术、miRNA干扰、信号通路干扰、流式细胞仪及免疫组化等多种研究技术。研究明确了Kupffer细胞Exosome参与肝衰竭病理进展过程，中药复方解毒化瘀颗粒能通过影响Kupffer细胞Exosome miRNAs发挥其疗效作用，并建立其多维疗效机制网络；进一步发现解毒化瘀颗粒上调Exosome中miR-9a-5p表达减弱Hif信号通路过度活化，抑制肝细胞凋亡死亡，以发挥其治疗ACLF的疗效作用。研究成果为解毒化瘀颗粒治疗慢加急性肝衰竭的作用机制提供了依据，以利于解毒化瘀颗粒的深入研发和临床应用，也为中医药治疗本病提供科学依据。