TL1A调控巨噬细胞极化和募集在NASH小鼠肝脏炎症发生中的机制研究

Non-alcoholic steatohepatitis (NASH) is one of the most common chronic liver diseases in China. And the morbidity is increasing gradually. M1/M2 macrophage balance distruction and recruitment mononuclears to liver play key roles in NASH. The latest studies have proven that TL1A, a recruit of tumor necrosis factor superfamily, is increased in macrophages in many inflammatory diseases, such as inflammatory bowel disease, primary biliary cholangitis and liver fibrosis. One recent research had shown that the TL1A-/-mice showed weight reduced, abdominal adipose tissue and the level of liver leptin which was one of the key fat cytokines involved in NASH decreased. Consequently, our hypothesis is that TL1A may participate in NASH by regulating macrophages. In this study, NASH model will be established using FMS-TL1A-GFP-transgenic mice and WT mice to investigate the effects of TL1A on liver inflammation, polarization and recruitment of macrophages in murine NASH. Moreover, bone marrow-derived macrophages and Kuffer cells with high expression of TL1A will be isolated, and intervene to nonalcoholic fatty primary liver cells to explore the influence of TL1A to hepatocellular injury and apoptosis. This study may provide new research evidence on the pathogenesis and the effective prevention and therapeutic strategy of NASH.

非酒精性脂肪性肝炎(NASH)是我国最常见的慢性肝病之一，且发病率呈逐年上升趋势。研究证实巨噬细胞向肝脏募集并向M1型极化所介导的慢性炎症反应在NASH进展中发挥关键作用。肿瘤坏死因子样配体1A(TL1A)是肿瘤坏死因子家族成员之一，参与炎症性肠病、原发性胆汁性胆管炎、肝纤维化等多种炎症相关疾病，并在上述疾病靶器官巨噬细胞中表达升高。此外，有研究发现小鼠TL1A基因敲除后出现体重下降、腹部脂肪组织减少，提示TL1A可能参与NASH发生。本课题采用髓系细胞高表达TL1A的转基因型小鼠和具有相同遗传背景的野生型小鼠构建NASH模型，探讨TL1A在肝组织和巨噬细胞中的表达变化及对巨噬细胞极化和募集的影响；同时应用上述两种小鼠的巨噬细胞分别与体外诱导的非酒精性脂肪肝细胞共培养，探讨巨噬细胞高表达TL1A对肝细胞损伤及凋亡的影响，为揭示NASH的发病机制、寻求防治NASH的新靶标提供实验依据。

非酒精性脂肪性肝炎(NASH)是我国最常见的慢性肝病之一，且发病率呈逐年上升趋势。研究证实巨噬细胞向肝脏募集并向M1型极化所介导的慢性炎症反应在NASH进展中发挥关键作用。肿瘤坏死因子样配体1A(TL1A)是肿瘤坏死因子家族成员之一，参与炎症性肠病、原发性胆汁性胆管炎、肝纤维化等多种炎症相关疾病，并在上述疾病靶器官巨噬细胞中表达升高。.本课题利用高糖高脂（HFHF）饮食及蛋氨酸-胆碱缺乏（MCD）饮食诱导构建2种NASH模型，证实TL1A在NASH小鼠的肝组织和肝脏巨噬细胞中表达均升高；利用髓系细胞高表达TL1A的转基因型小鼠构建NASH模型，发现转基因小鼠的肝组织脂肪变性和炎症情况、肝细胞凋亡情况、血脂和转氨酶水平均比野生型小鼠的NASH模型更重；巨噬细胞在肝组织中的募集情况、炎症因子（TNF-α、IL-1β和IL-6）表达情况均比野生型小鼠更多；检测肝组织中的CCL2、CXCL1及其相应受体CCR2、CXCR2，也均比野生型小鼠表达更多。上述结果提示NASH小鼠巨噬细胞TL1A表达升高，通过促进巨噬细胞募集、向M1型极化、促炎因子表达升高，加重肝损伤。体外实验，提取2种模型小鼠的巨噬细胞进行体外刺激，发现高表达TL1A的巨噬细胞更容易向M1型极化并且释放促炎因子更多；将2种巨噬细胞（骨髓来源的巨噬细胞和枯否细胞)并体外诱导分化为M1型巨噬细胞，分别与肝细胞共培养，高表达TL1A的巨噬细胞更容易加重肝细胞脂肪变性；进一步研究证实转基因小鼠NF-κB p65蛋白表达较WT小鼠明显升高，同时IκBα蛋白表达显著下降，提示TL1A可能通过NF-κB信号通路活化巨噬细胞，促进巨噬细胞向M1型极化。.TL1A在炎症性肠病中的研究较为深入，目前已经有TL1A抗体用于IBD的临床试验，相信很快就会应用于临床。通过研究TL1A在NASH中的发病机制，也将为NASH的治疗提供靶点，为将来的难治性NASH的治疗提供理论基础。