人脐带间充质干细胞外泌体通过LncRNA-CEP95调控NK细胞的表型转换减轻肝移植急性排斥损伤的作用研究

Acute rejection (AR), as a fatal factor in affecting graft survival, is still a key issue in the field of liver transplantation. Innate immune system is increasingly attracted attention in the initiation and development of AR. Previous studies by the applicant found that the account of intrahepatic NK cells and the change of ratio of NK cell subpopulation were closely related to the severity of AR. In addition, both mesenchymal stem cells and their exosomes (MSC-Exo) exhibited an effect on alleviating AR via regulating NK cell function (FASEB, 2018; Cell Prof, 2018). However, the underlying mechanism is still unclear. Furthermore, the applicant also found that MSC-Exo could induce the conversion of NK cell phenotype through activated the MYD88//NF-κB signaling pathway, and screened out the long non-coding RNA (LncRNA) by RNA-seq. The result showed that the LncRNA-CEP95, enriched in the MSC-Exo, is a miR-149-5p sponge and significantly regulates the MYD88 expression. Based on these findings, we hypothesized that whether MSC-Exo could alleviate AR by inducing the change of NK cell phenotype via LncRNA-CEP95/miR-149-5P/MYD88 axis in rat liver transplantation. In this study, we aim to provide a sufficient theoretical basis for the administration of MSC-Exo to alleviate AR after liver transplantation.

急性排斥损伤（AR）是影响移植物存活的关键问题，如何减轻AR是肝移植领域的重要课题。固有免疫在AR的作用愈受重视，申请人前期研究已证实肝脏内NK细胞数量及亚群比例与AR严重程度密切相关；而间充质干细胞及其外泌体（MSC-Exo）可调控NK细胞功能并减轻AR（FASEB J, 2018; Cell Prof, 2018），但其机制不明确。此外，申请人还发现MSC-Exo可激活MYD88/NF-κB通路诱导NK细胞表型转换，且高通量测序鉴定出MSC-Exo富集调控MYD88表达的LncRNA-CEP95/miR-149-5p轴。基于此提出科学假说：MSC-Exo内LncRNA-CEP95是否通过吸附miR-149-5p调控NK细胞表型减轻肝移植术后AR？本课题拟探讨MSC-Exo通过调控NK细胞表型转换从而减轻肝移植后AR的作用及其可能机制，为其在肝移植中的应用提供理论依据。

移植后急性排斥损伤（AR）是目前影响肝移植患者中长期生存的最主要原因。研究证实除了适应性免疫外，固有免疫也发挥重要作用。课题组前期研究证实肝脏内NK细胞数量及亚群比例与AR严重程度密切相关；而间充质干细胞（MSC）及其来源胞外囊泡（MSC-EVs）可调节NK细胞功能并减轻AR，但其具体机制并不十分清楚。本课题利用肝移植急性排斥损伤模型和临床研究，通过体内外实验探索MSCs及MSC-EVs通过免疫调节治疗肝移植术后AR的机制，为MSCs及MSC-EVs在肝移植领域临床转化提供理论依据。.本研究的主要目的为通过开展初期临床研究结合体内外实验研究MSCs及其来源MSC-EVs是否通过发挥免疫调节功能有效防治肝移植后免疫介导并发症发生发展的作用及探究其中可能的机制，为进一步推广MSCs及MSC-EVs在肝移植领域的临床转化应用提供理论基础。首先在本课题的支持下通过构建大鼠肝移植急性排斥模型，发现MSCs可有效改善肝移植急性排斥反应，并发挥免疫调节作用，抑制促炎因子水平以及调节肝脏驻留NK细胞的功能。机制上，我们发现MSCs发挥免疫调节作用减轻肝移植急性排斥反应主要是通过其分泌的胞外囊泡（MSC-derived extracellular vesicles, MSC-EVs）实现。并通过RNA-Seq及蛋白质谱都能高通量测序技术分析其内富集的miRNA、lncRNA及circRNA从而深入挖掘MSC-EVs发挥免疫调节作用的可能机制。进一步我们还开展了MSCs在肝移植领域的I/IIa临床研究。通过收集ABO血型不符（ABO-incompatible, ABO-i） 肝移植患者并在术后分别给予美罗华或MSCs治疗，发现MSCs不但与美罗华相类似同样具有抗抗体介导的体液排斥反应，而且还可有效预防移植术后严重感染和胆道并发症的发生。.本项目实施期间，申请人以第一作者/通讯作者发表SCI论文8篇，影响因子大于10分4篇，影响因子大于5分2篇。