VEGFR-1/Src/miR-21/MMP-9信号轴调控肝细胞癌侵袭转移的机制研究

Our previous results showed c-Src(Src) mediated the epithelial-mesenchymal transition(EMT) induced by vascular endothelial growth factor receptor-1(VEGFR-1) activation and contributed to migration and invasion of hepatoma cells in hepatocellular carcinoma(HCC). However, how Src mediates the EMT, what the targeting molecules of Src downstream are and whether it exists crosstalk between Src kinase pathway and other pathways or not remain unclear. Our previous study found the Src specific inhibitor saracatinib significantly down-regulated microRNA-21(miR-21) expression, miR-21 up-regulated matrix metalloproteinases-9(MMP-9). Therefore, we hypothesize overexpressing miR-21 and Src/miR-21/MMP-9 signaling axis are the central factors and signaling axis regulating EMT induced by VEGFR-1 activation in hepatocellular carcinoma, and then contributes to the recurrence and metastasis of HCC. To test our hypothesis, several methods such as the tissue microassay, gene clone, chromatin immunoprecipitaion assay (ChIP) and gene knockdown will be used to illustrate the role and mechanism of miR-21 and Src/miR-21/MMP9 in EMT induced by VEGFR-1 activation and provide the theoretical evidences for developing the new strategies preventing and treating the recurrence and metastasis of HCC.

我们研究发现，c-Src(Src)介导了血管内皮生长因子受体1(VEGFR-1)激活诱导的肝癌细胞上皮-间质转化(EMT)并促进肝癌细胞侵袭转移，但Src如何介导肝癌细胞EMT、Src的下游靶分子以及Src信号轴是否与其它通路存在交叉对话尚不明确。近期我们还发现Src特异性抑制剂saracatinib显著下调促肿瘤侵袭转移的micrRNA-21(miR-21)表达，且与基质金属蛋白酶(MMP-9)呈正相关。因此假设在HCC中高表达的miR-21及Src/miR-21/MMP9信号轴是调控VEGFR-1诱导的肝癌细胞EMT的中枢因子和信号轴，并促进HCC复发转移。本研究拟用组织芯片、分子克隆、染色质免疫沉淀和基因敲除等方法从分子、细胞和整体水平阐明miR-21及Src/miR-21/MMP-9在VEGFR-1激活诱导EMT中的作用及调控机制，为临床开发防治HCC复发转移的新策略提供理论依据。

肝细胞癌(Hepatocellular Carcinoma, HCC)是我国最常见的恶性肿瘤之一，已成为我国癌症相关死亡原因的第三位[1]。HCC复发转移是影响治愈率的最主要原因，但分子机制尚不完全清楚。近来研究人员在多种实体肿瘤细胞上发现“异位”表达的功能性血管内皮生长因子受体-1(Vascular Endothelial Growth Factor Receptor-1, VEGFR-1)，被其配体(VEGF-B和PIGF)特异性激活后，能够显著增强肿瘤细胞的侵袭转移能力，但确切的分子机制不清楚。我们研究发现，肝癌细胞中c-Src(Src)激酶介导了VEGFR-1活化诱导的上皮-间质转化(Epithelial-Mesenchymal Transition, EMT)，从而促进肝癌细胞侵袭转移(详见工作基础第一部分)，但是Src如何介导肝癌细胞EMT、Src的下游靶分子以及Src激酶通路是否与其它信号通路存在交叉对话尚不明确。阐明上述问题将为防治HCC复发转移和提高HCC治愈率、降低死亡率提供理论依据。本项目是我们团队成员以前研究工作的延续，是在Src介导VEGFR-1活化诱导肝癌细胞EMT的基础上进行下游分子机制的探讨。根据我们的前期研究结果和国内外研究现状，在预实验的基础上，拟运用组织芯片、分子克隆、细胞生物学、染色质免疫沉淀(ChIP)和基因敲除等多种方法，探讨miR-21在Src介导VEGFR-1激活诱导肝癌细胞EMT而促进HCC复发转移中的作用及调控机制，为临床开发防治HCC复发转移的新策略提供理论依据。