TGF-β1信号通路介导成纤维细胞促进移植肝细胞血管化的研究

The hepatocyte transplantation shows a promising prospect, however as the consequence of insufficient vascularization, it is difficult for transplanted hepatocyte to have long-term survival and proliferation. As the most crucial component in microenvironment of tumor, actived fibroblast, induced by cytokine such as TGF-β1 secreted by tumor, plays an important role in the process of neovascularization and proliferation in tumor cells. Given this, fibroblast can be regarded as a significant medium to solve the problem of insufficient vascularization after hepatocyte transplantation. This research is planned to modify fibroblast with TGF-β1 gene: conduct experiment in vitro and vivo to perceive and verify the function that the actived fibroblast, induced by TGF-β1, can stimulate angiogenesis; and then co-transplanted the fetal hepatic cells and fibroblasts, modified by TGF-β1 gene, into the liver of SD rats. By cognizing the relationship between TGF-β1 signaling and actived fibroblast, the condition actived fibroblast responding to factors which stimulate angiogenesis such as HGF,COX-2,VEGF-A,SDF-1 and MMP-9, and the vascularization of transplanted cells, the experiment is aimed to investigate the effect and mechanism of the activated fibroblasts induced by TGF-β1 signal pathway in the vascularization and proliferation of transplanted liver cells, to lay a foundation as to solve the difficulty in terms of long-term survival and proliferation of transplanted hepatocyte.

肝细胞移植前景广阔，但血管化不足是移植肝细胞长期存活及增殖的主要障碍。作为肿瘤中最重要的微环境成分，成纤维细胞被肿瘤分泌的TGF-β1等细胞因子激活后对肿瘤新生血管的形成发挥至关重要的作用，故成纤维细有望成为解决肝细胞移植后血管化不足问题的重要媒介。本研究拟将TGF-β1基因修饰成纤维细胞，先通过体外及体内实验了解并证实TGF-β1途径激活的成纤维细胞具有促血管生成作用，再将TGF-β1基因修饰的成纤维细胞与胚胎肝细胞联合移植到SD大鼠肝脏。旨在通过了解TGF-β1信号与成纤维细胞激活的关系、激活的成纤维细胞表达HGF、COX-2、VEGF-A、SDF-1、MMP-9等促血管生成相关因子的情况、移植肝细胞的血管化情况，探讨TGF-β1途径介导成纤维细胞促进移植肝细胞血管化的作用及机制，为解决移植肝细胞长期存活及增殖困难奠定基础。

肝细胞移植前景广阔，但解决移植的肝干细胞定向分化及血管化问题是困扰移植肝细胞功能发挥及长期存活的关键。本课题主要从细胞实验、动物胚胎肝脏发育模型、以及动物肝脏大部分切除模型这几个层面，研究肝脏干细胞向胆管细胞及肝细胞定向分化过程中的不同机制和信号通路，以及研究肝细胞增殖过程中的血管化机制和相应的信号通路。另外，利用本课题经费探索了肿瘤在免疫选择压力下的基因进化过程，并探索了miR-34a在促进肝细胞表达MICB的作用机制。研究发现TGFβ介导的Jagged1-Notch-Sox9 信号通路在胚胎肝脏胆管定向分化过程中发挥关键作用；证实TGFβ信号通过Jagged1/Notch通路调节肝脏星状细胞向胆管细胞分化；揭示了肝星状细胞通过TGF-β1/Jagged1 信号轴调节肝脏祖细胞的定向分化；研究发现肝脏星状细胞通过TGF-β1-Jagged1/VEGFA 信号通路促进肝脏再生过程中的血管生成；还发现在不同的培养条件下肝脏祖细胞可以向不同的方向分化；还发现TGFβ信号可以赋予肝脏星状细胞祖细胞的功能。另外，研究发现MicroRNA-34a能够促进肝细胞MICB的表达，发现了胰腺神经内分泌瘤在免疫选择压力下的肿瘤进化过程。在此项目资助下，通过这些研究已经发表了7篇SCI论文，影响因子共达24分；另外还有1篇论文已经投稿，另1篇论文正在撰写中（预计这2篇论文SCI影响因子共达10分左右）。此外，还有多篇CSCD核心期刊论文发表。通过这些研究，发现了促进肝干细胞向肝细胞及胆管细胞定向分化、以及移植肝细胞增殖后血管化的重要机制，为下一步探索肝干细胞移植定向分化为类肝脏器官、以及移植肝细胞血管化奠定了坚实的基础。