固有淋巴样细胞22(ILC22)在乙型肝炎病毒感染介导免疫炎症损伤中的作用

Innate lymphoid cells 22 (ILC22) comprise a heterogeneous population of ILCs which are dependent on the transcription factor RORγt and mainly secrete a plenty of interleukin (IL)-22. Recent studies have demonstrated that Notch signaling takes part in the regulation of ILC22 differentiation. Moreover, ILC22 plays an immunoregulatory role in different immune and inflammatory diseases, such as psoriasis, colitis, and acute hepatitis. However, there are no reports about ILC22 in the immune and inflammatroy response to hepatitis B virus (HBV) infection. Our previous studies revealed that HBV infection induced the elevation of hepatic ILC22 in mouse model. Furthermore, IL-22 plays a proinflammatory role and leads to immuno-inflammatory damage of liver in the immune reponse to HBV infection. Thus, the current results indicate that ILC22 may take part in HBV-mediated immune response. Thus, in the present study, we will examine the hypothesis that ILC22 plays an important role in the immuno-inflammtory response to HBV. To evaluate the hypothesis, we will carry out the experiments to determine the regulatory role of HBV infection in ILC22 differentiation, and to clarify the role of Notch signaling in regulation of ILC22 as well as mechanisms of ILC22 induced inflammatory damage to HBV infection in HBV transgenic mice and hydrodynamic injection mice models. These experiments are important, as they may identify ILC22 as a new subset that regulates HBV related liver inflammation and immune response. A complete understanding of these processes may also lead to improved therapies for the liver injury caused by HBV infection, which would have the potential to prevent liver cirrhosis and hepatocellular carcinoma.

ILC22是一类新命名的RORγt(+)固有淋巴样细胞亚群，通过分泌IL-22发挥生物学效应。新近研究发现，Notch信号通路参与ILC22分化调控，且ILC22在多种免疫和炎症性疾病（如银屑病、结肠炎和急性肝炎）中发挥重要免疫调控作用。但ILC22是否参与HBV感染免疫和炎症应答尚未见报道。我们前期研究发现，HBV感染可导致小鼠肝脏中ILC22水平明显升高，而ILC22的效应细胞因子IL-22在HBV感染中发挥促进肝脏炎症损伤的作用，提示ILC22可能参与了HBV介导的免疫炎症应答。因此，本研究以HBV感染者和小鼠模型为研究对象，分析HBV感染及抑制Notch信号通路对ILC22分化的影响，探讨ILC22在HBV感染介导肝脏炎症损伤中的作用，以期揭示ILC22在HBV感染炎症应答中的免疫调控作用，为全面阐明HBV感染诱导肝脏炎症应答的机制提供实验依据。

固有淋巴样细胞22（ILC22）属于ILC3亚群，通过分泌IL-22发挥生物学功能。Notch信号通路参与ILC22的分化调控，且ILC22在多种免疫和炎症性疾病中发挥重要免疫调控作用。项目负责人既往的研究已发现，Notch信号通路参与了分泌IL-22的CD4+T细胞的分化调控，Notch-IL-22信号通路参与了HBV感染的炎症应答。但Notch信号通路是否可调控ILC22的分化、ILC22在HBV感染的免疫炎症应答中是否发挥相应作用尚不清楚。因此，本研究以高压尾静脉注射HBV质粒构建的小鼠模型和HBV感染者为研究对象，分析肝脏内和外周血ILC22水平变化，并在体内和体外抑制Notch信号通路，分析Notch信号通路对ILC22分化的影响和相关机制。体内实验方面，成功构建了高压尾静脉注射HBV质粒诱导小鼠“感染”HBV，构建小鼠模型，HBV感染后小鼠肝脏中Notch1和Notch2的水平升高，虽然肝脏浸润的分泌IL-22的CD4+T细胞数量在HBV感染后未见明显变化，但肝脏浸润的NKp46+ILC22和LTi4细胞数量明显增加，CD4-ILC22数量显著降低，同时伴有多种非特异性炎症细胞（辅助性T细胞、杀伤性T细胞、NK细胞、NKT细胞）向肝脏募集浸润增加，肝脏内IL-22、IFN-γ、TNF-α、CXCL9、CXCL10 mRNA相对表达量升高。抑制Notch信号通路对于HBV抗原表达无显著影响，但可使NKp46+ILC22和LTi4细胞数量在肝脏内显著降低，同时向肝脏募集浸润的炎症细胞、促炎细胞因子、趋化因子水平亦显著降低。体外实验方面，抑制Notch信号通路可诱导非特异性CD4+T细胞分泌IL-22水平下降，同时，鉴定了人类外周血中ILC亚群，人ILC亚群的表型为Lin-CD45+CD127+CD161+，进一步根据CD117和CRTH2的表达对ILC1、ILC2和ILC3亚群进行分析，在Lin-CD45+CD127+CD161+ILC中，ILC1表型为CRTH2-CD117-，ILC2表型为CRTH2+，ILC3表型为CRTH2-CD117+。本研究结果提示，HBV感染可影响ILC22分化，诱导肝脏炎症应答，说明ILC22在HBV感染应答中发挥了重要作用，为抗HBV治疗提供了新的思路和治疗靶点。