白细胞衍生趋化因子2（LECT2）结合肝窦内皮细胞膜受体Tie1促进肝纤维化进展

Liver fibrosis is the key step of the progression of chronic liver disease to the end stage, which lacks early non-invasive diagnosis and effective therapeutic target. Our previous work found that the level of LECT2 in serum and liver tissue of patients with cirrhosis is significantly increased. Loss of LECT2 alleviate the degree of fibrosis in mice induced by CCL4. Interestingly, we also found that LECT2 is not expressed in hepatic stellate cells and macrophages, but highly expressed in vascular endothelial cells and surrounding hepatocytes, which binding to endothelial cell membrane receptor Tie1. Based on the above findings, we speculate that LECT2 could combine with Tie1 and promote liver fibrosis by regulating angiogenesis. To test this hypothesis, we first confirm the role of LECT2 in promoting liver fibrosis and influencing angiogenesis by over-expressing LECT2 in vivo. Secondly, we plan to use in vitro experiments to confirm the direct binding of LECT2 to Tie1 by GST pull-down method, and explore the mechanism of LECT2 binding to Tie1 to regulate angiogenesis by transwell and tube formation assay. Last, based on the above findings, LECT2 will be used as a target for therapeutic exploration. This proposal may clarify the molecular mechanism of LECT2 combined with Tie1 in regulating angiogenesis and promoting liver fibrosis. This project also may provide some evidence to developing a new therapeutic strategy in liver fibrosis through targeting LECT2.

肝纤维化是慢性肝病进展至终末期的核心环节，临床缺乏早期无创性诊断和有效治疗靶点。前期研究发现肝硬化患者血清和肝组织中LECT2的含量显著升高，LECT2敲除小鼠能减缓CCL4诱导的肝纤维化；机制研究表明LECT2不表达于肝星状细胞和巨噬细胞，而在血管内皮细胞及其周围肝细胞中高表达，并能结合内皮细胞膜受体Tie1。据此，我们推测：LECT2可结合肝窦内皮细胞膜受体Tie1，通过调控血管生成进而促进肝纤维化。本课题拟体内过表达LECT2验证其促进肝纤维化的作用和对血管生成的影响；体外运用GST pull-down实验证实LECT2直接结合Tie1，并通过细胞侵袭实验和小管形成实验探索LECT2结合Tie1调控血管生成的机制；最后以LECT2为靶点进行治疗性探索。课题的实施将阐明LECT2结合Tie1调控血管生成进而促进肝纤维化的分子机制，为探索LECT2作为新的肝纤维化治疗靶点提供理论依据。

肝纤维化是慢性肝病进展至终末期的核心环节，临床缺乏早期无创性诊断和有效治疗靶点。前期研究发现肝硬化患者血清和肝组织中LECT2的含量显著升高，LECT2敲除小鼠能减缓CCL4 诱导的肝纤维化，LECT2过表达小鼠能够加重肝纤维化。机制研究表明LECT2能够和血管内皮细胞膜受体Tie1结合通过调控血管生成参与肝脏的损伤与修复，持续的损伤与修复会导致肝纤维化，因此我们推测 LECT2 参与肝纤维化的进展，并可作为临床诊断与评估肝硬化的潜在指标。此前还没有研究关注LECT2在慢性乙型肝炎（Chronic hepatitis B，CHB）患者肝纤维化中的作用。本研究纳入227例CHB患者，分为训练组（n=147）和验证组（n=80）。通过检测LECT2在CHB患者血清中的含量以及肝组织中蛋白和mRNA的表达水平，分析LECT2与肝纤维化程度的相关性。采用ROC曲线评估LECT2预测肝纤维化程度的特异性和敏感性，并与经典的APRI和FIB-4指标进行比较，结果证实血清LECT2含量可以作为评估CHB患者肝纤维化程度的可靠生物标志物。在研究的过程中我们发现King指数作为一种肝纤维化无创诊断模型，因其计算简单、可重复率高，可用于CHB患者慢性病程中对肝纤维化程度进行动态、连续性检测。.在进一步研究中我们发现LECT2与肝细胞癌的预后密切相关，通过TCGA数据库的分析和临床样本的验证及随访，结果表明LECT2在HCC中的低表达与预后不良密切相关，具有潜在的临床应用价值。此外，我们对细胞分裂周期相关蛋白4（CDCA4）在人类肿瘤中的致癌作用进行泛癌分析，发现CDCA4可作为肿瘤免疫浸润和预后不良的生物标志物；我们还鉴定了m6A相关的lncRNA特征，并构建了模型来评估肝细胞癌患者的预后，为肝细胞癌诊疗提供新的思路。