肿瘤相关巨噬细胞经不均一核糖核蛋白A/B介导上皮-间质转化促进肝癌侵袭转移

肿瘤相关巨噬细胞(TAM)是肿瘤免疫微环境中的重要成分，在肿瘤转移中发挥重要作用。前期工作中,我们发现TAM能提高肝癌细胞运动侵袭能力，且与上皮-间质转化(EMT)和不均一核糖核蛋白A/B表达上调密切相关。本课题拟观察TAM共培养对肝癌细胞EMT生物标志物表达及亚细胞定位的影响；通过shRNA干预不均一核糖核蛋白A/B表达，观察干预前后TAM对肝癌细胞EMT和侵袭转移的影响的变化，明确不均一核糖核蛋白A/B在TAM诱导肝癌EMT和侵袭转移中的作用；应用启动子芯片筛选不均一核糖核蛋白A/B调控的与EMT发生相关的关键基因；应用免疫印迹、荧光素报告基因分析TAM作用前后肝癌细胞内不均一核糖核蛋白A/B表达和转录活性的改变及其对关键基因转录调控的影响；同时分析关键基因在肝癌组织中的表达及其与肝癌转移复发间的关系，并对其进行体内、外功能分析。通过上述研究，为探讨肝癌治疗中新的干预靶点提供实验依据。

Tumor-associated macrophages (TAM) are a major component of the cancer-related microenvironment and play a central role in tumor metastasis. In our previous study, we found that hepatocellular carcinoma (HCC) cell lines exhibited higher invasive potentials when co-cultured with TAM; this effect may occur through the epithelial-mesenchymal transition (EMT) and up-regulation of hnRNP A/B. In this study, we intend to observe the impact of TAM on promoting the EMT process. The role of hnRNP A/B in TAM-induced HCC invasion and the EMT process will be investigated using short hairpin RNA (shRNA)-mediated down-regulation of hnRNP A/B in HCC cell lines with various metastatic potentials. Furthermore, we will use chromatin immunoprecipitation and gene promotor arrays to screen hnRNP A/B-regulated genes involved in the EMT process in HCC cells. Western blotting and luciferase assays will be respectively used to analyze hnRNP A/B expression and transcription activity in HCC cells co-cultured with TAM. We will then determine whether hnRNP A/B is involved in the transcriptional regulation of these genes in HCC after co-culture with TAM. In addition, the relationship between the expression of these genes and the invasive phenotype of HCC cells will be studied. We will determine whether these genes could be important independent factors in determining clinical outcomes of HCC. Using specific RNA interference or up-regulation of these genes in HCC cells, we will assess their roles in tumor invasion and in vivo metastasis. This study will provide a biological foundation for the discovery of new potential therapeutic targets for HCC.