HBx和AFB1在卵圆细胞转化为肝癌中的作用及其机制研究

The cell arise of the hepatocellular carcinoma (HCC) are not very well understood. Whether HCC arise from stem cell that undergo a malignant transformation or from the de-differentiation of mature hepatocytes, however, has long been controversial. Hepatic oval cells (HOC) are considered to be the stem cells of the liver and have been linked to the development of HCC. Studies have demonstrated that chronic hepatitis B virus (HBV) infection and dietary aflatoxin B1 (AFB1) exposure are among the most important risk factors for the development of HCC. However, little research has been done to evaluate the role of oval cells in these two environmental factors on hepatocarcinogenesis. In this study, partial transformation of rat HOC (LE/6) were accomplished by transfected HBVx gene (HBx), and then transfected cells were implanted both intra-hepatically and subcutaneously into nude mice treated with AFB1 in vivo. We found the oval cells produced tumors (4/24 of the animals) in liver following transfection with HBx gene and treatment with AFB1. These intrahepatic tumors included HCC cells and mesenchymal cells. Further testing found that HBx activated Wnt and Notch signaling pathway, Wnt silence can not completely prevent the malignant transformation of oval cells, the Notch signal pathway may play more crucial role. This study aim to establish HBx collaborative AFB1 induced oval cell generate HCC, to clarify the role of HBx / Notch signal pathway in this transformation. These results provide aN evidence that oval cells have the capacity to generate HCC through the combined effects of the HBx and AFB1 in the liver microenvironment.

肝癌发病机制仍不清楚，对肝癌细胞的起源仍存在较大分歧。肿瘤干细胞的发现，提供了新的视角。卵圆细胞具有肿瘤干细胞的特征，与肝癌的发生关系密切。乙肝病毒（HBV）和黄曲霉毒素（AFB1）是肝癌的最常见病因，而且有协同作用。根据前期研究，推测乙肝病毒X基因（HBx）可能作用于卵圆细胞使其转化为肝癌；我们将表达HBx蛋白的卵圆细胞种植于裸鼠肝实质内，同时喂食AFB1作为诱癌剂，可在肝内形成肝癌肉瘤（含肝细胞癌和间质瘤），进一步检测发现HBx激活转染细胞中的Wnt和Notch信号通路，沉默Wnt并不能完全阻止卵圆细胞恶性转化，而Notch信号通路可能起到更关键作用。本研究将建立HBx协同AFB1诱导卵圆细胞转化为肝癌的动物模型，阐明Notch信号通路在其中的作用机制，为肝癌是否来源于卵圆细胞提供直接的实验证据，从而可以预防或降低肝癌的发生。

乙肝病毒（HBV）和黄曲霉毒素（AFB1）是肝癌的最常见病因，而卵圆细胞具有肿瘤干细胞的特征，与肝癌的发生关系密切。我们对HBx、AFB1、卵圆细胞、Notch信号通路与肝癌的关系进行了研究。本课题研究发现，在肝癌组织中Notch1的表达明显高于癌旁组织、肝硬化组织和正常肝组织。而且，Notch1和HBx在肝癌组织中的表达呈正相关。Notch信号通路随着卵圆细胞的增殖而激活。同时，卵圆细胞过表达HBx后激活MEK/ERK和PI3K/AKT通路上调cyclinD1促进卵圆细胞增殖。我们在肝癌细胞中也发现HBx通过Notch1通路调控DUSP1/ERK和PTEN/AKT信号通路。在使用AFB1对卵圆细胞进行长期小剂量的诱导后，卵圆细胞(WB-F344)Notch信号通路激活，增殖能力增加，而且发生了恶性转化。以上研究证实HBx和AFB1通过激活Notch信号通路诱导卵圆细胞恶性转化并参与肝癌的发生。体内成瘤实验表明肝卵圆细胞在HBx和黄曲霉素的共同作用下可部分分化成肝细胞癌。我们的研究首次将HBx、AFB1、卵圆细胞、Notch信号通路等串联起来，证实了Notch信号通路在肝癌发生发展中的重要作用，为肝癌的诊断和治疗提供了新的理论支持。