力学参数－肝基质硬度调控血管内皮细胞VEGFR2表达参与肝癌新生血管生成新机制研究

Fibrosis-caused matrix stiffness increase of liver is the predominant clinical characteristic of HCC patients. Recent studies suggest that increased matrix stiffness of liver can promote cell proliferation, drug resistance and invasion/metastasis process in hepatocellular carcinoma. However, the role of mechanical parameters-liver matrix stiffness in angiogenesis of hepatocellular carcinoma is largely unknown. Our previous study found that there was a positive correlation between vascular density and increased matrix stiffness, meanwhile increased matrix stiffness upregulated the expression of VEGF in HCC cells and the expression of VEGFR2 in endothelial cells. These results indicate that the matrix stiffness of liver may be involved in the formation of new blood vessels of HCC via a synergic regulation between HCC cells and vascular endothelial cells. This is a Novel strategies to study the angiogenesis of HCC stimulated by matrix stiffness. This project is to discover matrix stiffness associated proteins which regulate the VEGFR2 expression in endothelial cells, and its detailed role in angiogenesis of HCC based on the cell-culture platform and buffalo rat HCC models with different liver matrix stiffness backgrounds established previously, as well as other technologies. The project will reveal a new mechanism that the angiogenesis of HCC regulated by liver matrix stiffness, this may supply a new target point and strategy for anti-angiogenic therapy of HCC.

肝纤维化导致的肝基质硬度（力学信号）增加是肝癌患者显著临床特征，近来研究发现其参与肝癌增殖、耐药、侵袭/转移等病理过程，然而肝基质硬度是否影响肝癌新生血管形成目前国内外尚未见报道。前期研究显示，肝基质硬度通过整合素β1-PI3K/Akt通路上调肝癌细胞VEGF表达，且显著上调血管内皮细胞VEGFR2表达，提示基质硬度可能协同影响肝癌细胞及血管内皮细胞进而促进肝癌血管生成，但血管内皮细胞具体调控机制仍不清晰。本项目依托已建立的不同肝基质硬度大鼠肝癌模型和不同基底硬度体外细胞培养平台，拟通过转录因子芯片、信号通路磷酸化芯片及siRNA抑制和慢病毒过表达等技术，对基质硬度调控血管内皮细胞VEGFR2表达的完整信号通路进行分析，并在细胞、动物和临床组织水平层面验证基质硬度对血管生成的影响。预期该项目将揭示基质硬度调控肝癌血管生成的新机制和关键调控点，为肝癌临床治疗提供新的靶点。

血管生成在肝癌的发生、发展及侵袭转移过程中起重要作用，它不仅为肿瘤组织提供氧和养料，也是肿瘤细胞播散转移的重要途径。中国肝癌患者90%有乙型肝炎感染背景，约80%以上的肝癌患者具有不同程度肝纤维化和肝硬化，肝纤维化/肝硬化导致的肝基质硬度（力学信号）增加是肝癌患者的显著背景，肝基质硬度调控肝癌血管生成的分子机制仍不清楚。本课题基于建立的不同肝基质硬度背景大鼠肝癌模型和体外不同基底硬度细胞培养体系平台，对基质硬度参与调控肝癌血管生成的分子机制进行探索。体内试验和体外试验结果显示，肝基质硬度增加与VEGFR2的表达呈正相关；转录因子芯片筛选出与基质硬度变化相关的差异转录因子，荧光素酶报告基因证实SP1参与调控VEGFR2的表达；基质硬度激活PI3K/Akt信号通路，抑制PI3K/Akt通路下调VEGFR2和SP1的表达；同时研究发现整合素αV表达与基质硬度增加呈负相关。本研究发现基质硬度可通过PI3K/Akt-SP1通路调控血管内皮细胞VEGFR2的表达，参与肝癌肿瘤血管生成的过程。本研究通过前期建立的不同基质硬度背景的体内和体外实验平台，阐明基质硬度力学信号调控血管内皮细胞VEGFR2表达的分子机制，完善了肝癌血管生成的调控机制，为深入研究基质硬度参与肝癌血管生成机制奠定良好基础和开发新的靶向药物提供新的依据。