The Mechanisms Of Hypoxia-Induced Epithelial To Mesenchymal Transition In Hepatocellular Carcinoma Induces An Immunosuppressive Tumor Microenvironment

Obejective:Portal vein tumor thrombosis (PVTT) is a significant risk factor for metastasis in hepatocellular carcinoma (HCC) patients and is therefore associated with poor prognosis. But it is unclear about the mechanisms of PVTT induce the metastasis. We want to investigate the expression level of HIF-la in the sample of HCC patients with PVTT and its effect on the tumor microenvironment.Materials and methods:Tumor tissue samples were obtained from the Second Affiliated Hospital of Zhejiang University School of Medicine. Ninety patients with hepatocellular carcinoma underwent curative resection between 2007 and 2010, and samples from these patients were used for IHC and quantitative RT-PCR (qRT-PCR) to detect the expression of HIF-la, CCL20 and IDO. Furthermore, we analyzed the clinical relevance of elevated CCL20 levels in HCC patients. We cultured the hepatoma cell lines in the hypoxic incubator to investigate whether there was a change in the cytokine profile in the culture medium. To further identify the effect of the hypoxia-induced cytokines in the tumor microenvironments, we directly culture monocytes derived macrophages with hypoxic hepatoma cell lines.Results:We found the expression of HIF-1α was significantly increased in the tumor samples with PVTT. Moreover, HIF-1α induced the expression of CCL20 in hepatoma cells. We identified the hypoxia response element (HRE) on the promoter of CCL20 with ChIP assay. Meanwhile, we found CCL20 and IDO were also increased in the tumor samples with PVTT, in which the IDO mRNA was positively associated with the CCL20 mRNA level. We also demonstrated that the tumor-derived CCL20 upregulates IDO expression in the monocytes-derived macrophage. CCL20-induced expression of IDO might be a result of activation of phosphorylation and nuclear translocation of STAT1. This IDO+ subsets of macrophage can generate the immunosuppressive Treg-like cells in vitro. We analyzed the relationship between CCL20 and IDO expression in hepatocellular carcinoma patients with or without vascular invasion from National Center for Biotechnology Information Gene Expression Omnibus database. CCL20 levels significantly correlated with IDO in patients with vascular invasion. In contrast, no significant association was observed in patients without vascular invasion.Conclusion:Our findings suggest that the HIF-1α/CCL20/IDO axis in hepatocellular carcinoma is important for accelerating tumor metastasis through both the induction of EMT and the establishment of an immunosuppressive tumor microenvironment, warranting further investigation into the therapeutic effects of blocking specific nodes of this signaling network.