BackgroundCholangiocarcinoma (CCA) is a form of cancer that easily aggress to contiguous structures. Vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2) are increased in majority species of cancers and suppress tumor progression by blocking VEGF/VEGFR2. Apatinib is a highly selective VEGFR2 antagonist which has inhibitive effect on antiapoptotic and cell growth in CCA. While, the effect of apatinib cell migration and invasion in CCA is still unknown.MethodsCCA cell lines QBC939 and TFK-1 were transfected with siKDR to establish the KDR function loss cell model, and recombined human VEGF (rhVEGF) protein was added into the culture medium to enhance the VEGF expression. RT-qPCR and western bloting were used to detect the mRNA and protein expression levels of VEGFR2 to investigate whether it was effectively repressed or activated with rhVEGF or apatinib treatment. Then, MTT, wound healing assay, and transwell matrix assay were applied to measure the effect of apatinib and rhVEGF on cell viability, migration and invasion, respectively.ResultsThe mRNA and protein expressions of VEGFR2 were significantly reduced with KDR RNAi in both QBC939 and TFK-1 cells, and rhVEGF treatment increased these expression levels (p < 0.05). Apatinib dramatically suppressed VEGF-mediated cell migration and invasion at the concentration of 100 nM treatment and significantly decreased the expression of metastasis-associated protein such as Slug, snail and MMP9. Moreover, all of these inhibiting effects of apatinib depended on the VEGFR2 existence. In addition, VEGFR2/RAF/MEK/ERK and PI3K/AKT signal pathways were enhanced by the introduction of rhVEGF, but were dramatically suppressed after the apatinib treatment.ConclusionApatinib inhibit VEGF-mediated cell migration and invasion in CCA cell lines via inhibiting the VEGFR2/RAF/MEK/ERK and PI3K/AKT pathways. It will be a potentially effective targeted drug for CCA.

Background:Hepatocellular carcinoma is a highly vascularized tumor, so it is critical to study its angiogenesis. Cancer-associated fibroblasts and enhancer of zeste homolog 2 play an important role in tumor angiogenesis and became significant hallmarks of cancer. But the relationship between enhancer of zeste homolog-2 and cancer-associated fibroblasts in response to angiogenesis and its precise mechanism remains unclear.Methods:The separation of cancer-associated fibroblasts was identified by immunofluorescence. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analysis was used to reveal the proliferation of human umbilical vein endothelial cells. Vascular endothelial growth factor level was quantified by enzyme-linked immunosorbent assay. The wound healing, transwell, and vascular tube formation assays were used to identify the capability of migration, invasion, and tube formation of human umbilical vein endothelial cells in vitro. The detection of tumor angiogenesis was also performed in vivo. Finally, the level of enhancer of zeste homolog-2 and vasohibin 1 were determined by quantitative real-time polymerase chain reaction and Western blotting.Results:In comparison to control and condition medium noncancerous fibroblasts groups, the condition medium cancer-associated fibroblasts could significantly promote the proliferation, migration, invasion, and angiogenesis of human umbilical vein endothelial cells. We found that cancer-associated fibroblasts promoted angiogenesis of human umbilical vein endothelial cells via vascular endothelial growth factor secretion in vitro and in vivo. The upregulation of enhancer of zeste homolog 2 by vascular endothelial growth factor inhibited the expression of vasohibin 1, thus promoting the proliferation and angiogenesis of human umbilical vein endothelial cells. Taken together, the cancer-associated fibroblasts of hepatocellular carcinoma regulate the enhancer of zeste homolog-2/vasohibin 1 pathway via vascular endothelial growth factor secretion, thereby promoting the proliferation and angiogenesis of human umbilical vein endothelial cells.Conclusion:Our study identified the relationship between cancer-associated fibroblasts and enhancer of zeste homolog-2 and confirmed the pivotal role of cancer-associated fibroblasts in angiogenesis of hepatocellular carcinoma. Cancer-associated fibroblasts promote angiogenesis of hepatocellular carcinoma by vascular endothelial growth factor–mediated enhancer of zeste homolog-2/vasohibin 1 pathway and may be a potentially useful therapeutic target for hepatocellular carcinoma.

No abstract

The aim of present investigation is to explore the molecular mechanisms of vasculogenic mimicry (VM) induced by hypoxia. Hepatocellular carcinoma cell lines were treated with CoCl2, and the VM-related parameters were assayed by real-time qPCR, Western blotting and immunofluorescence. Matrigel tube structure was also detected. We demonstrated that the expression of pMEK, MEK, pERK1/2 and ERK1/2 had a positive correlation with VM induced by hypoxia in MHCC97H while HepG2 signified VM under normoxia condition. PD98059 was negatively while epidermal growth factor positively participated in the increased tubes and area of VM. At the meaning time, the increased VM-related genes VE-cadherin, MMP2, MMP9, EphA2 and LAMC2 in hypoxia group were down-regulated by PD98059 in a dose-dependent manner. Furthermore, we elucidated that PKA, but not PKC, mediated the MEK/ERK pathway in a negative manner in VM. In conclusion, MEK/ERK pathway is positively involved in VM in hepatocellular carcinoma cell line, which was mediated by PKA negatively.