Notch受体对肝星状细胞和窦内皮细胞在肝纤维化形成和血管生成中的作用

The interaction between hepatic stellate cells (HSC) and sinusoidal endothelial cells (SEC) plays an important role in the hepatic fibrogenesis and angiogenesis.Notch receptor signaling is an important signal to regulate cell interaction and it also is associated with hepatic fibrogenesis.The present study suggest that there may have a potential relationship between Notch and fibrogenesis.Our preliminary study showed that intrahepatic myofibrobasts were found high expression of Notch ligands and receptors in patients with liver cirrhosis, in vitro experiments suggested that inhibition of Notch receptor signaling could reduce HSC activation and collagen synthesis.In order to explore the mechanism of the Notch receptor in the hepatic fibrogenesis and angiogenesis in chronic liver injury，this project intends to research the impact of down- and up-regulation Notch receptor signaling by adenovirus vector overexpression and siRNA gene silencing technologies on the activation, proliferation, migration, extracellular matrix synthesis of HSC, and the proliferation, tube formation, capillarization，ultrastructure of SEC from cell and animal levels.The cross-talks between HSC and SEC will also be studied. Finally, we dynamically study the relationship of Notch receptors expression with fibrogenesis and angiogenesis in the evolution of hepatic fibrosis from human level.From HSC, SEC and their interactions, animal and human level， we will clarify the role and mechanism of Notch receptors in the process of hepatic fibrogenesis and angiogenesis for exploring the feasibility of Notch recrptors as a therapeutic targetof liver fibrosis.

肝星状细胞（HSC）与窦内皮细胞（SEC）间相互作用在肝维化形成及血管发生过程中发挥重要作用，Notch受体信号是调节细胞间相互作用的重要信号并与纤维化的发生发展有关。我们前期研究发现肝内肌成纤维细胞可高表达Notch受体和配体，而抑制Notch受体信号能减少HSC活化和胶原合成。本课题拟从细胞和动物水平利用腺病毒载体过表达、siRNA基因沉默等技术研究上调和下调Notch受体信号对HSC活化、增殖、迁移、细胞外基质合成和SEC增殖、管腔形成、毛细血管化、超微结构的影响，以及Notch受体在HSC-SEC间所发挥调节作用。最后从慢性肝病肝纤维化演变过程中，探索慢性肝损伤过程中Notch受体在肝组织中的表达、定位及分布。从不同水平研究HSC、SEC及其相互作用，阐明Notch受体在肝纤维化形成及血管生成过程中的作用及机制，探索Notch受体作为肝纤维化治疗靶点的可行性。

Notch信号通路不仅在胚胎发育和肿瘤发生中起关键作用，而且越来越多的研究发现Notch 通路与纤维化疾病有关联。HSCs活化是肝纤维化发生中的关键事件；肝窦内皮细胞（SEC）是一类特殊化的血管内皮细胞，在肝脏的血管生成中发挥重要作用。本课题从细胞、动物水平研究Notch受体信号对HSC活化、增殖、迁移、细胞外基质合成和SEC血管生成的影响。我们发现HSC在体外转化为肌成纤维细胞过程中Notch信号通路是活化的；抑制Notch信号可降低HSC活化和增殖。SEC在体外培养过程中Notch信号通路也活化，抑制Notch信号可降低SEC毛细血管化。SEC与HSC共培养时，细胞种植比例依据两者在正常肝脏中的比例（3.2 : 1），抑制SEC Notch通路可降低HSC的活化、增殖。在建立大鼠四氯化碳肝纤维化模型和胆总管结扎模型的基础上，应用腺病毒转染Notch信号核结合蛋白RBPJκ的干扰RNA（AdshRBPJκ）。结果发现，AdshRBPJκ显著抑制大鼠肝脏Notch信号活化，并可改善大鼠肝脏血管生成及纤维化指标。基于慢性乙型肝炎患者肝活检标本的研究显示，在人体肝纤维化发生发展中，伴随血管生成的增加，且Notch信号通路是激活的。本研究表明Notch信号在肝纤维化形成及血管生成中发挥重要作用，抑制Notch信号活化可改善肝纤维化及血管生成，为靶向干预Notch 信号的抗肝纤维化治疗提供了理论和实验依据。