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Hypoxia-induced up-regulation of VASP promotes invasiveness and metastasis of hepatocellular carcinoma.

ABSTRACT: Rational: Patients with hepatocellular carcinoma (HCC) have a poor prognosis mostly due to intrahepatic as well as distal metastasis. Vasodilator-stimulated phosphoprotein (VASP), a regulator of actin cytoskeleton and cell migration, is overexpressed in HCC and correlated with its malignant features and poor prognosis. Very little is known about its function in HCC. Methods: qRT-PCR, Western blot and IHC were used to detect the VASP expression in tissues and cells. Transwell and wound healing assays were used to measure the migration and invasion of HCC cells. Immunoblotting and immunofluorescence were used for detection of epithelial-to-mesenchymal transition (EMT) progression in HCC cells. A lung metastasis mouse model was used to evaluate metastasis of HCC in vivo. The putative targets of miR-204 were disclosed by public databases and a dual-luciferase reporter assay. IP was used to show the interaction between VASP and CRKL. ChIP was used to analyze the binding of HIF-1? to VASP promoter region. Results: Our data involving both gain- and loss-of-function studies revealed that VASP activated AKT and ERK signaling and promoted HCC migration and invasion in vitro and in vivo by altering the EMT phenotype and expression of MMPs. We investigated the positive correlation between VASP and an adapter protein, CRKL. VASP dynamically co-localized at the SH3N domain of CRKL and mediated its function. Mechanistically, VASP overexpression at the transcriptional level was mediated by HIF-1? through direct binding to two hypoxia response elements (HRE) in the VASP promoter region. Furthermore, we identified hypoxia-induced down-regulation of miR-204, which functioned as the regulator of VASP overexpression at the post-transcriptional level. Also, hypoxia-activated p-Smad3 dependent TGF-? signaling indirectly promoted VASP expression. Conclusion: A variety of hypoxia-induced molecular mechanisms contributed to the upregulation of VASP at transcriptional and post-transcriptional levels. These mechanisms involved CRKL, HIF-1?, miR-204, and TGF-? activating the AKT and ERK signaling to promote EMT and expression of MMPs. Taken together, our results defined VASP as an oncogene of HCC pathogenesis and metastasis with the potential to serve as a prognostic biomarker.

SUBMITTER: Liu Z

PROVIDER: S-EPMC6160773 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Hypoxia-induced up-regulation of VASP promotes invasiveness and metastasis of hepatocellular carcinoma.

Liu Zhikui Z Wang Yufeng Y Dou Changwei C Xu Meng M Sun Liankang L Wang Liang L Yao Bowen B Li Qing Q Yang Wei W Tu Kangsheng K Liu Qingguang Q

Theranostics 20180909 17

<b>Rational:</b> Patients with hepatocellular carcinoma (HCC) have a poor prognosis mostly due to intrahepatic as well as distal metastasis. Vasodilator-stimulated phosphoprotein (VASP), a regulator of actin cytoskeleton and cell migration, is overexpressed in HCC and correlated with its malignant features and poor prognosis. Very little is known about its function in HCC. <b>Methods:</b> qRT-PCR, Western blot and IHC were used to detect the VASP expression in tissues and cells. Transwell and wo ...[more]

PMID: 30279729

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Project description:Rationale: Metastasis and recurrence are the primary reasons for the high mortality rate of human hepatocellular carcinoma (HCC) patients. However, the exact mechanism underlying HCC metastasis remains unclear. The Homeobox (HOX) family proteins, which are a highly conserved transcription factor superfamily, play important roles in cancer metastasis. Here, we report a novel role of HOXC10, one of the most upregulated HOX genes in human HCC tissues, in promoting HCC metastasis. Methods: The expression of HOXC10 and its functional targets was detected by immunohistochemistry in two independent human HCC cohorts. Luciferase reporter and chromatin immunoprecipitation assays were used to measure the transcriptional regulation of target genes by HOXC10. The effect of HOXC10-mediated invasion and metastasis were analyzed by Transwell assays and by an orthotopic metastasis model. Results: Elevated expression of HOXC10 was positively correlated with the loss of tumor encapsulation and with higher tumor-nodule-metastasis (TNM) stage and poor prognosis in human HCC. Overexpression of HOXC10 promoted HCC metastasis by upregulating metastasis-related genes, including 3-phosphoinositide-dependent protein kinase 1 (PDPK1) and vasodilator-stimulated phosphoprotein (VASP). Knockdown of PDPK1 and VASP inhibited HOXC10-enhanced HCC metastasis, whereas upregulation of PDPK1 and VASP rescued the decreased metastasis induced by HOXC10 knockdown. Interleukin-1? (IL-1?), which is the ligand of IL-1R1, upregulated HOXC10 expression through the c-Jun NH2-terminal kinase (JNK)/c-Jun pathway. HOXC10 knockdown significantly reduced IL-1?-mediated HCC metastasis. Furthermore, Anakinra, a specific antagonist of IL-1R1, inhibited IL-1?-induced HOXC10 upregulation and HCC metastasis. In human HCC tissues, HOXC10 expression was positively correlated with PDPK1, VASP and IL-1R1 expression, and patients with positive coexpression of HOXC10/PDPK1, HOXC10/VASP or IL-1R1/HOXC10 exhibited the poorest prognosis. Conclusions: Upregulated HOXC10 induced by IL-1? promotes HCC metastasis by transactivating PDPK1 and VASP expression. Thus, our study implicates HOXC10 as a prognostic biomarker, and targeting this pathway may be a promising therapeutic option for the clinical prevention of HCC metastasis.

Project description:Objective: To investigate the potential biomarkers for venous metastasis of hepatocellular carcinoma (HCC), and briefly discuss their target genes and the signaling pathways they are involved in. Materials and Method: The dataset GSE6857 was downloaded from GEO. Significantly differentially expressed miRNAs were identified using the R package "limma," After that, the survival analysis was conducted to discover the significance of these up-regulated miRNAs for the prognosis of HCC patients. Additionally, miRNAs which were up-regulated in venous metastasis positive HCC tissues and were significant for the prognosis of HCC patients were further verified in clinical samples using RT-qPCR. The miRNAs were then analyzed for their correlations with clinical characteristics including survival time, AFP level, pathological grade, TNM stage, tumor stage, lymph-node metastasis, distant metastasis, child-pugh score, vascular invasion, liver fibrosis and race using 375 HCC samples downloaded from the TCGA database. The target genes of these miRNAs were obtained using a miRNA target gene prediction database, and their functions were analyzed using the online tool DAVID. Results: 15 miRNAs were differentially expressed in samples with venous metastasis, among which 7 were up-regulated in venous metastasis positive HCC samples. As one of the up-regulated miRNAs, hsa-miR-210 was identified as an independent prognostic factor for HCC. Using RT-qPCR, it was evident that hsa-miR-210 expression was significantly higher in venous metastasis positive HCC samples (p = 0.0036). Further analysis indicated that hsa-miR-210 was positively associated with AFP level, pathological grade, TNM stage, tumor stage and vascular invasion. A total of 168 hsa-miR-210 target genes, which are mainly related to tumor metastasis and tumor signaling pathways, were also predicted in this study. Conclusion: hsa-miR-210 might promote vascular invasion of HCC cells and could be used as a prognostic biomarker.

Dataset Information

Hypoxia-induced up-regulation of VASP promotes invasiveness and metastasis of hepatocellular carcinoma.

Publications

Hypoxia-induced up-regulation of VASP promotes invasiveness and metastasis of hepatocellular carcinoma.

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