Tcf21+细胞在肝癌微环境中的分化命运及其在肝癌发生发展中的作用研究

Tumor-associated fibroblasts (TAF) are the main stromal cells in the tumor microenvironment and play an important role in tumor initiation, growth, invasion and metastasis. The cell origin of TAF in hepatocellular carcinoma (HCC) microenvironment lacks formal lineage tracing evidence and molecular markers, which greatly limits the targeted therapy of HCC microenvironment. Previously, Previously, the applicant found a group of Tcf21+ stromal cells around the hepatic portal vein and central vein. Preliminary experiments showed that Tcf21+ cells proliferated significantly before and after the occurrence of hepatocellular carcinoma, and were the precursor cells of TAF in hepatocellular carcinoma tissues. Blocking TGFβ signal in Tcf21+ cells decrease the hepatic fibrosis in precancerous microenvironment. Next, by using the techniques of lineage tracing, conditional gene knockout, immunohistochemistry and immunofluorescence, the applicant will systematically study the changes of signal expression of Tcf21+ cells during the establishment and development of hepatocellular carcinoma and their effects on the microenvironment of hepatocellular carcinoma, and elucidate the molecular mechanism of Tcf21+ cells regulating the occurrence and development of hepatocellular carcinoma. This work will help us to understand the role of TAF in regulating the microenvironment of hepatocellular carcinoma, and provide a basis for the design of targeted therapy for hepatocellular carcinoma microenvironment.

肿瘤相关成纤维细胞(tumor associated fibroblasts, TAF) 是肿瘤微环境中的主要间质细胞，在肿瘤的起始、生长、侵袭和转移中起着重要的作用。肝癌微环境中TAF的细胞起源至今缺乏直接的谱系示踪证据和分子标记，极大的限制了肝癌微环境的靶向治疗。申请人前期发现肝脏门静脉和中央静脉周围有一群Tcf21+间质细胞。初步的实验显示，Tcf21+细胞在肝癌发生前后显著扩增，是肝癌组织中TAF的前体细胞；在Tcf21+细胞中阻断TGFβ信号影响癌前微环境肝脏纤维化的发生。在下一步的工作中，申请人拟利用谱系示踪、条件性基因敲除、免疫组化和免疫荧光等技术系统性地研究Tcf21+细胞在肝癌建立和发展过程中的信号表达变化及其对肝癌微环境的影响，阐明其调控肝癌发生发展的分子机制。本项工作的开展将有助于我们深入理解TAF调节肝癌微环境的作用，为设计靶向肝癌微环境的治疗提供基础。

肝星状细胞被认为是肌成纤维细胞以及肿瘤相关成纤维细胞的前体细胞，在肝纤维化以及肝癌的发生发展过程中发挥着重要的作用。然而由于缺乏特异性的分子标记，之前对于肝星状细胞的研究主要基于体外实验，缺乏体内细胞示踪证据。根据拟定的目标，我们采用单细胞测序的方法鉴定到Tcf21可作为肝星状细胞的标记，并且构建了Tcf21-cre/ER；R26-tdTomato示踪小鼠，利用这个小鼠系统地分析Tcf21+细胞在成体小鼠肝脏中的组织定位，发现Tcf21+细胞标记肝星状细胞。同时还运用这个可诱导的谱系示踪小鼠观测到Tcf21+细胞在肝纤维化以及肝癌发生过程中分别分化形成肌成纤维细胞以及肿瘤相关成纤维细胞的过程，这是首次运用可诱导的谱系示踪的方法在体内观测到肝星状细胞到肿瘤相关成纤维细胞的分化过程。此外，我们还发现在Tcf21+细胞中阻断TGFβ信号能够抑制肝纤维的进程以及肿瘤的进程。本项目的完成首次在体内证明Tcf21+肝星状细胞来源的肿瘤相关成纤维细胞对肿瘤的促进作用，为靶向肿瘤相关成纤维细胞的治疗策略提供了坚实的基础。..