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MiR200-upregulated Vasohibin 2 promotes the malignant transformation of tumors by inducing epithelial-mesenchymal transition in hepatocellular carcinoma.

Xue X, Zhang Y, Zhi Q, Tu M, Xu Y, Sun J, Wei J, Lu Z, Miao Y, Gao W - Cell Commun. Signal (2014)

Bottom Line: VASH2 knockdown in HepG2 cells inhibited epithelial-mesenchymal transition (EMT), but VASH2 overexpression in Hep3B cells promoted EMT.Western blot analyses showed that VASH2 promoted EMT through the ZEB1/2 pathway.VASH2 promoted invasion, reduced apoptosis and increased the proportion of stem cells in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of General Surgery, the First Affiliated Hospital with Nanjing Medical University, 300# Guangzhou Road, Nanjing, 210029, China. xfxue@suda.edu.cn.

ABSTRACT

Background: Hepatocellular carcinoma (HCC) typically relies on tumor transformation and angiogenesis for its malignant behavior, including growth and metastasis. Previously, we reported that Vasohibin2 (VASH2) is preferentially expressed in hepatocellular carcinoma (HCC) tumor tissues and promotes angiogenesis. Here, we further investigated the role of VASH2 in HCC tumor progression.

Results: Bioinformatics analyses and luciferase reporter gene assays confirmed the post-transcriptional regulation of VASH2 by miR-200a/b/c. We then used HepG2 and Hep3B cells, two representative hepatic cancer cell lines, to examine the role of VASH2 in tumors. VASH2 knockdown in HepG2 cells inhibited epithelial-mesenchymal transition (EMT), but VASH2 overexpression in Hep3B cells promoted EMT. Western blot analyses showed that VASH2 promoted EMT through the ZEB1/2 pathway.

Conclusion: VASH2 promoted invasion, reduced apoptosis and increased the proportion of stem cells in vitro and in vivo. These results indicated that VASH2 expression in HCC cells promotes the malignant transformation of tumors by inducing EMT.

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Related in: MedlinePlus

The effect of VASH2 expression on cell phenotype and marker change. (A) Cell phenotype changes resulting from different levels of VASH2 expression. (B) Western blot analysis showed that the levels of ZEB1/2, an EMT marker and core transcriptional factor, respectively, were affected by changes in VASH2 expression. (C) Immunofluorescence (IF) confirmed that the EMT markers changed with VASH2 expression. Magnification of 20 × .
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Fig3: The effect of VASH2 expression on cell phenotype and marker change. (A) Cell phenotype changes resulting from different levels of VASH2 expression. (B) Western blot analysis showed that the levels of ZEB1/2, an EMT marker and core transcriptional factor, respectively, were affected by changes in VASH2 expression. (C) Immunofluorescence (IF) confirmed that the EMT markers changed with VASH2 expression. Magnification of 20 × .

Mentions: HepG2 cells predominantly exhibit a mesenchymal phenotype. VASH2 knockdown in HepG2 cells caused the cells to change from a spindle-shaped morphology to an epithelioid-like morphology. We examined the HepG2-shVASH2 and HepG2-shcont cells at 1 and 3 days after subculture (Figure 3A). The HepG2-shVASH2 cells acquired the epithelioid-like phenotype and proliferated in clusters, and the HepG2-shcont cells exhibited a spindle-shaped morphology and grew separately. VASH2 overexpression in Hep3B cells did not significantly affect their phenotype (data not shown). We further detected EMT markers, such as vimentin and E-cadherin, in these cells. The results (Figure 3B) showed that VASH2 overexpression increased vimentin expression and decreased the level of E-cadherin. Moreover, VASH2 knockdown decreased vimentin expression and increased the level of E-cadherin. We further detected the core transcriptional factor, ZEB1/2, in EMT by western blot. The results (Figure 3B) showed that ZEB1/2 was upregulated with VASH2 overexpression and downregulated with VASH2 knockdown. Also promoter luciferase reporter gene assay was used to elucidate the mechanism between VASH2 and ZEB1/2. The results (Additional file 1: Figure S1) illustrated VASH2 activated ZEB1/2 promoter activity so that increased ZEB1/2 expression, in consistent with western blot.Figure 3


MiR200-upregulated Vasohibin 2 promotes the malignant transformation of tumors by inducing epithelial-mesenchymal transition in hepatocellular carcinoma.

Xue X, Zhang Y, Zhi Q, Tu M, Xu Y, Sun J, Wei J, Lu Z, Miao Y, Gao W - Cell Commun. Signal (2014)

The effect of VASH2 expression on cell phenotype and marker change. (A) Cell phenotype changes resulting from different levels of VASH2 expression. (B) Western blot analysis showed that the levels of ZEB1/2, an EMT marker and core transcriptional factor, respectively, were affected by changes in VASH2 expression. (C) Immunofluorescence (IF) confirmed that the EMT markers changed with VASH2 expression. Magnification of 20 × .
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4195883&req=5

Fig3: The effect of VASH2 expression on cell phenotype and marker change. (A) Cell phenotype changes resulting from different levels of VASH2 expression. (B) Western blot analysis showed that the levels of ZEB1/2, an EMT marker and core transcriptional factor, respectively, were affected by changes in VASH2 expression. (C) Immunofluorescence (IF) confirmed that the EMT markers changed with VASH2 expression. Magnification of 20 × .
Mentions: HepG2 cells predominantly exhibit a mesenchymal phenotype. VASH2 knockdown in HepG2 cells caused the cells to change from a spindle-shaped morphology to an epithelioid-like morphology. We examined the HepG2-shVASH2 and HepG2-shcont cells at 1 and 3 days after subculture (Figure 3A). The HepG2-shVASH2 cells acquired the epithelioid-like phenotype and proliferated in clusters, and the HepG2-shcont cells exhibited a spindle-shaped morphology and grew separately. VASH2 overexpression in Hep3B cells did not significantly affect their phenotype (data not shown). We further detected EMT markers, such as vimentin and E-cadherin, in these cells. The results (Figure 3B) showed that VASH2 overexpression increased vimentin expression and decreased the level of E-cadherin. Moreover, VASH2 knockdown decreased vimentin expression and increased the level of E-cadherin. We further detected the core transcriptional factor, ZEB1/2, in EMT by western blot. The results (Figure 3B) showed that ZEB1/2 was upregulated with VASH2 overexpression and downregulated with VASH2 knockdown. Also promoter luciferase reporter gene assay was used to elucidate the mechanism between VASH2 and ZEB1/2. The results (Additional file 1: Figure S1) illustrated VASH2 activated ZEB1/2 promoter activity so that increased ZEB1/2 expression, in consistent with western blot.Figure 3

Bottom Line: VASH2 knockdown in HepG2 cells inhibited epithelial-mesenchymal transition (EMT), but VASH2 overexpression in Hep3B cells promoted EMT.Western blot analyses showed that VASH2 promoted EMT through the ZEB1/2 pathway.VASH2 promoted invasion, reduced apoptosis and increased the proportion of stem cells in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of General Surgery, the First Affiliated Hospital with Nanjing Medical University, 300# Guangzhou Road, Nanjing, 210029, China. xfxue@suda.edu.cn.

ABSTRACT

Background: Hepatocellular carcinoma (HCC) typically relies on tumor transformation and angiogenesis for its malignant behavior, including growth and metastasis. Previously, we reported that Vasohibin2 (VASH2) is preferentially expressed in hepatocellular carcinoma (HCC) tumor tissues and promotes angiogenesis. Here, we further investigated the role of VASH2 in HCC tumor progression.

Results: Bioinformatics analyses and luciferase reporter gene assays confirmed the post-transcriptional regulation of VASH2 by miR-200a/b/c. We then used HepG2 and Hep3B cells, two representative hepatic cancer cell lines, to examine the role of VASH2 in tumors. VASH2 knockdown in HepG2 cells inhibited epithelial-mesenchymal transition (EMT), but VASH2 overexpression in Hep3B cells promoted EMT. Western blot analyses showed that VASH2 promoted EMT through the ZEB1/2 pathway.

Conclusion: VASH2 promoted invasion, reduced apoptosis and increased the proportion of stem cells in vitro and in vivo. These results indicated that VASH2 expression in HCC cells promotes the malignant transformation of tumors by inducing EMT.

Show MeSH
Related in: MedlinePlus