HBV相关性肝癌中Mig/CXCR3调控CD44+肝癌细胞跨内皮迁移及其分子机制研究

Hematogenous metastasis is an early event in hepatocellular carcinoma progression and the underlying mechanisms are still unclear. Metastasis is a multi-step cascade process in which transendothelial migration (TEM) of cancer cells into vessels is one of the rate-limiting procedures. It is known now that not all the cancer cells own the metastatic characteristics. Previous studies have demonstrated that cancer stem cells (CSCs) are proposed to initiate cancer propagation and metastasis, and CD44 is a putative HCC cancer stem-cell marker. There is increasing evidence that tumor microenvironment (including chemokine) plays a pivotal role in the dissemination and establishment of CSCs metastasis. For proof of principle, we have illustrated that Mig protein expressed higher in hepatitis B-related HCC serum and tissues than that in normal liver counterparts. In addition, the level of CXCR3 expression was positively correlated with HCC metastasis. Further, it was shown that stimulation with chemokine Mig lead to profound migration of the CD44 positive (CD44+) hepatocellular carcinoma cells through endothelial monolayer as compared with the control group. Yet the molecular mechanisms for tumor cells metastasis promoted by chemokine Mig have so far remained obscure. In the current proposed project, we will elucidate the correlation between Mig/CXCR3 and recurrence and prognosis in HCC patients. Moreover， we will try to investigate the imapact of Mig/CXCR3 in CD44+ hepatocellular carcinoma cells and endothelial cells in vitro, and to clarify the effect of the pathway on hepatoma recurrence and metastasis in vivo. In this proposal, we will explore the role and underlying molecular mechnisms of Mig/CXCR3 for TEM of CD44+ hepatocellular carcinoma cells, and eventually discover potential targets for early anti-metastasis intervention in hepatoma.

肝癌早期易发生血行转移。研究表明，跨内皮迁移（TEM）是肝癌细胞血行转移的必要步骤。并非所有癌细胞都具有恶性特征，肿瘤干细胞是肿瘤转移复发的根源，CD44是重要的肝癌干细胞标记物，肿瘤微环境中趋化因子对肿瘤干细胞迁移起主要调节作用。我们前期研究显示，乙肝相关性HCC患者血清及肝癌组织高表达趋化因子Mig，有转移者癌细胞亦高表达Mig受体（CXCR3）；Mig处理后实现TEM的CD44+细胞数量较对照组明显增加，这提示Mig在CD44+细胞TEM过程中发挥重要促进作用。但其分子机制尚无研究报道。本项目在此基础上，拟探讨Mig/CXR3与HCC患者转移及预后的关系；研究Mig/CXCR3通路对CD44+肝癌细胞及内皮细胞的调控；从动物水平证实其在肝癌转移中的作用。通过以上研究探讨Mig/CXCR3在CD44+肝癌细胞跨内皮屏障迁移中的分子机制及意义，为肝癌细胞早期转移寻找潜在的干预靶点。

原发性肝癌（HCC）是世界上第五大恶性肿瘤，病死率居我国恶性肿瘤的第二位，每年死于HCC的患者超过23万。肝癌易发生早期侵袭转移。与其他肿瘤相比较，HCC尤其乙肝相关性肝癌更易发生早期血行转移。研究表明，跨内皮迁移（TEM）是癌细胞血行转移的必要步骤。并非所有癌细胞都具有恶性特征，肿瘤干细胞是肿瘤转移复发的根源，CD133是重要的肝癌干细胞标记物，肿瘤微环境中趋化因子对肿瘤干细胞转移有重要调节作用。我们前期研究显示，乙肝相关性HCC患者体内趋化因子Mig高表达，有转移者其受体CXCR3亦高表达.本项目在此基础上，探讨Mig/CXCR3与HCC患者复发及预后的关系，研究Mig与受体结合后对CD133+肝癌细胞及内皮细胞的作用及其分子机制，并从动物水平证实Mig/CXCR3在肝癌转移复发中的作用。研究结果显示：1）CXCR3受体在肝癌组织中高表达，并与肿瘤大小，肿瘤分化程度，门脉侵袭和转移密切相关。在这项研究中，CXCR3-A剪接体，由Mig诱导可导致MAPK信号通路ERK1/2磷酸化水平的显著变化，并从而上调的MMP2和MMP9的表达，促进CD133+肝癌细胞的侵袭和转移。此外，过表达CXCR3-A可抑制24小时Mig诱导后的CD133 +肝癌细胞的黏附能力。2）检测了血管内皮细胞粘附分子Ve-cadherin在不同浓度Mig刺激时mRNA和蛋白水平的改变，在Mig50ng/ml和100ng/ml时有统计学上的差异。 CD133+肿瘤细胞和脐静脉内皮细胞（HUVEC）接触性共培养时可激活CD133+肿瘤细胞中NF-kB而上调接触性共培养基上清的Mig，从而促进CD133+肿瘤细胞的侵袭和转移，结果提示Mig在肿瘤细胞跨内皮迁移的接触过程中有潜在的促肿瘤转移的作用。综上所述，阐明Mig/CXCR3促进CD133+肝癌细胞转移和跨内皮屏障迁移中的作用及潜在机制，为肝癌细胞早期转移寻找潜在的干预靶点。