LGR4促进非酒精性脂肪性肝病相关肝癌发生及机制研究

Primary liver cancer is the second lethal cancer worldwide. With the decreasing viral hepatitis epidemic and fast growing prevalence of non-alcoholic fatty liver disease(NAFLD), NAFLD has become an increasingly important risk factor of primary liver cancer. We found that LGR4 mutant mice were resistant to NAFLD, some elderly wild type mice showed spontaneous liver cancer formation while LGR4 mutant mice resisted hepatocarcinogenesis . LGR4 liver specific knockout mice were reported to have reduced liver regeneration capacity after partial hepatectomy. Our previous work found that LGR4 mutant mice had reduced liver weight and reduced ZNRF3 level. As a negative regulator of canonical Wnt signaling，E3 ligase ZNRF3 was reported to participate in multiple kinds of cancer. We proposed that LGR4 potentiates primary liver cancer formation via modulating ZNRF3 either through gene expression or protein binding . In this project, we will use LGR4 liver specific knockout mice as well as other biomolecular techniques to further investigate the role of LGR4 in hepatocarcinogenesis in vivo and ex vivo，elucidating the mechanism of a novel candiate to regulate NAFLD related hepatocarcinogenesis .

肝癌是世界第二致死性肿瘤性疾病。随着病毒性肝炎发病率下降以及非酒精性脂肪性肝病的流行，非酒精性脂肪性肝病在肝癌发病中的地位越发凸显。我们前期研究发现LGR4基因敲除小鼠抵抗非酒精性脂肪肝的发生。部分野生小鼠肝脏自发成瘤而LGR4敲除小鼠抵抗肝癌发生。研究发现LGR4肝脏特异敲除小鼠肝脏部分切除后增殖减低。前期研究提示LGR4缺失小鼠肝脏重量减轻，Wnt信号通路负性调控因子泛素化连接酶ZNRF3下调。ZNRF3参与多种肿瘤的发生。我们提出科学假设：LGR4通过负性调节泛素化连接酶ZNRF3促进Wnt信号通路激活参与非酒精性脂肪肝相关肝癌发生。可能机制包括转转录调控和蛋白相互作用。本研究中我们以LGR4肝脏特异性敲除小鼠为主要研究模型，从体内、体外以及临床组织样本水平证明LGR4参与非酒精性脂肪肝相关肝癌发生，揭示LGR4通过抑制泛素化连接酶ZNRF3功能促进NAFLD相关肝癌发生的分子机制。

肝细胞癌是中国重要的疾病负担。非酒精性脂肪性肝病已成为发病率最高的慢性肝病，存在进展为肝硬化甚至肝癌风险，但相关机制需要深入研究。我们在前期研究的基础上发现LGR4参与非酒精性脂肪性肝病相关肝癌的发生发展。首先，在四氯化碳诱导的肝纤维化模型中，LGR4肝脏特异性敲除小鼠抵抗肝纤维化发生。在DEN联合高脂饮食肝癌模型中，LGR4肝脏特异性缺失小鼠肝癌结节数目明显少于对照组。在体外肝癌细胞系中，在LGR4高表达的细胞系中敲低LGR4后肿瘤细胞增殖明显降低，侵袭转移能力显著降低，干细胞相关基因表达下调，蛋白质印记分析显示PI3K-AKT信号通路参与其中，但具体的机制在进一步研究。此外，研究组还发现肿瘤-睾丸相关连环蛋白α2（CTNNA2）在肝癌患者的癌组织中表达高于癌旁组织，组织连环蛋白α2蛋白表达水平与患者血清AFP水平和门脉癌栓受累情况相关。网络分析提示肝癌中 CTNNA2基因和蛋白的功能调控可能与自身磷酸化 、启动子区 DNA 甲基化及与其他多种蛋白相互作用相关。