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Downregulation of developmentally regulated endothelial cell locus-1 inhibits the growth of colon cancer.

Zou X, Qiao H, Jiang X, Dong X, Jiang H, Sun X - J. Biomed. Sci. (2008)

Bottom Line: Gene transfection of Del1-shRNA downregulated expression of Del1 in LS-174T cells in vivo and in vitro, but did not alter the proliferative or survival properties of cells in vitro.Both Del1-shRNA and VEGF-shRNA gene therapies exhibited anti-tumor activities and they also showed a synergistic effect in suppressing growth of colon tumors by anti-angiogenesis and anti-proliferation.Although further investigation to clarify the mechanisms explaining the role of Del1 in tumor growth, and the interaction between VEGF and Del1, is required, the results indicate that downregulation of Del1 presents a potent therapeutic strategy to combat colon cancer.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Hepatosplenic Surgery Center, Department of General Surgery, The First Clinical College of Harbin Medical University, Harbin 150001, PR China. zouxiaolongbs@163.com

ABSTRACT
Developmentally regulated endothelial cell locus-1 (Del1) is an embryonic angiogenic factor expressed in early embryonic endothelial cells, but recently has been found to be expressed in some forms of cancers including colon and breast cancers, and melanoma, and human cancer cell lines. Overexpression of Del1 accelerates tumor growth by enhancing vascular formation, implying Del1 may be a potential target for anti-angiogenic cancer therapy. The study aims to investigate whether downregulation of Del1 could inhibit the growth of tumors established in nude Balb/c mice by subcutaneous implantation of human LS-174T colon cancer cells. The shRNA expression vectors targeting human Del1, and vascular endothelial growth factor (VEGF) were constructed. Gene transfection of Del1-shRNA downregulated expression of Del1 in LS-174T cells in vivo and in vitro, but did not alter the proliferative or survival properties of cells in vitro. Gene transfection of VEGF-shRNA downregulated expression of both VEGF and Del1 in LS-174T cells in vivo and in vitro. Both Del1-shRNA and VEGF-shRNA gene therapies exhibited anti-tumor activities and they also showed a synergistic effect in suppressing growth of colon tumors by anti-angiogenesis and anti-proliferation. Although further investigation to clarify the mechanisms explaining the role of Del1 in tumor growth, and the interaction between VEGF and Del1, is required, the results indicate that downregulation of Del1 presents a potent therapeutic strategy to combat colon cancer.

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VEGF-shRNA downregulates expression of VEGF and Del1 in vitro. LS-174T cells were transfected with VEGF-shRNA and harvested 48 h later. (A) VEGF (upper row) and Del1 proteins (middle row) in lysates of untreated cells (lane 1) or cells transfected with control vector (lane 2), or VEGF-shRNA plasmid (lane 3) was Western blotted. The density of each band was measured and compared to that of the internal control β-actin (bottom row). (B, C) The expression of VEGF mRNA (200 bp) (B) or Del1 mRNA (350 bp) (C) was detected by RT-PCR in lysates of untreated LS-174T cells (lane 2) or cells transfected with control vector (lane 3), or Del1-shRNA plasmid (lane 4). GAPDH (500 bp) was amplified as a PCR internal control. Lane 1 denotes the DNA marker. * Indicates a significant difference at P < 0.05 in the band intensities compared to control.
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Figure 7: VEGF-shRNA downregulates expression of VEGF and Del1 in vitro. LS-174T cells were transfected with VEGF-shRNA and harvested 48 h later. (A) VEGF (upper row) and Del1 proteins (middle row) in lysates of untreated cells (lane 1) or cells transfected with control vector (lane 2), or VEGF-shRNA plasmid (lane 3) was Western blotted. The density of each band was measured and compared to that of the internal control β-actin (bottom row). (B, C) The expression of VEGF mRNA (200 bp) (B) or Del1 mRNA (350 bp) (C) was detected by RT-PCR in lysates of untreated LS-174T cells (lane 2) or cells transfected with control vector (lane 3), or Del1-shRNA plasmid (lane 4). GAPDH (500 bp) was amplified as a PCR internal control. Lane 1 denotes the DNA marker. * Indicates a significant difference at P < 0.05 in the band intensities compared to control.

Mentions: Give that VEGF-shRNA gene therapy downregulated expression of VEGF and Del1 in situ, we further investigated whether VEGF-shRNA has the same effects in vitro. The LS-174T cells were transfected with VEGF-shRNA expression plasmids and harvested 48 h later, lysed and subjected to Western blot analysis with Abs against VEGF and Del1, respectively. As shown in Figure 7A, VEGF-shRNA gene transfection significantly downregulated expression of both VEGF and Del1 proteins, which was further supported by the downregulation of VEGF mRNA (Figure 7B) and Del1 mRNA by RT-PCR (Figure 7C).


Downregulation of developmentally regulated endothelial cell locus-1 inhibits the growth of colon cancer.

Zou X, Qiao H, Jiang X, Dong X, Jiang H, Sun X - J. Biomed. Sci. (2008)

VEGF-shRNA downregulates expression of VEGF and Del1 in vitro. LS-174T cells were transfected with VEGF-shRNA and harvested 48 h later. (A) VEGF (upper row) and Del1 proteins (middle row) in lysates of untreated cells (lane 1) or cells transfected with control vector (lane 2), or VEGF-shRNA plasmid (lane 3) was Western blotted. The density of each band was measured and compared to that of the internal control β-actin (bottom row). (B, C) The expression of VEGF mRNA (200 bp) (B) or Del1 mRNA (350 bp) (C) was detected by RT-PCR in lysates of untreated LS-174T cells (lane 2) or cells transfected with control vector (lane 3), or Del1-shRNA plasmid (lane 4). GAPDH (500 bp) was amplified as a PCR internal control. Lane 1 denotes the DNA marker. * Indicates a significant difference at P < 0.05 in the band intensities compared to control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2666667&req=5

Figure 7: VEGF-shRNA downregulates expression of VEGF and Del1 in vitro. LS-174T cells were transfected with VEGF-shRNA and harvested 48 h later. (A) VEGF (upper row) and Del1 proteins (middle row) in lysates of untreated cells (lane 1) or cells transfected with control vector (lane 2), or VEGF-shRNA plasmid (lane 3) was Western blotted. The density of each band was measured and compared to that of the internal control β-actin (bottom row). (B, C) The expression of VEGF mRNA (200 bp) (B) or Del1 mRNA (350 bp) (C) was detected by RT-PCR in lysates of untreated LS-174T cells (lane 2) or cells transfected with control vector (lane 3), or Del1-shRNA plasmid (lane 4). GAPDH (500 bp) was amplified as a PCR internal control. Lane 1 denotes the DNA marker. * Indicates a significant difference at P < 0.05 in the band intensities compared to control.
Mentions: Give that VEGF-shRNA gene therapy downregulated expression of VEGF and Del1 in situ, we further investigated whether VEGF-shRNA has the same effects in vitro. The LS-174T cells were transfected with VEGF-shRNA expression plasmids and harvested 48 h later, lysed and subjected to Western blot analysis with Abs against VEGF and Del1, respectively. As shown in Figure 7A, VEGF-shRNA gene transfection significantly downregulated expression of both VEGF and Del1 proteins, which was further supported by the downregulation of VEGF mRNA (Figure 7B) and Del1 mRNA by RT-PCR (Figure 7C).

Bottom Line: Gene transfection of Del1-shRNA downregulated expression of Del1 in LS-174T cells in vivo and in vitro, but did not alter the proliferative or survival properties of cells in vitro.Both Del1-shRNA and VEGF-shRNA gene therapies exhibited anti-tumor activities and they also showed a synergistic effect in suppressing growth of colon tumors by anti-angiogenesis and anti-proliferation.Although further investigation to clarify the mechanisms explaining the role of Del1 in tumor growth, and the interaction between VEGF and Del1, is required, the results indicate that downregulation of Del1 presents a potent therapeutic strategy to combat colon cancer.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Hepatosplenic Surgery Center, Department of General Surgery, The First Clinical College of Harbin Medical University, Harbin 150001, PR China. zouxiaolongbs@163.com

ABSTRACT
Developmentally regulated endothelial cell locus-1 (Del1) is an embryonic angiogenic factor expressed in early embryonic endothelial cells, but recently has been found to be expressed in some forms of cancers including colon and breast cancers, and melanoma, and human cancer cell lines. Overexpression of Del1 accelerates tumor growth by enhancing vascular formation, implying Del1 may be a potential target for anti-angiogenic cancer therapy. The study aims to investigate whether downregulation of Del1 could inhibit the growth of tumors established in nude Balb/c mice by subcutaneous implantation of human LS-174T colon cancer cells. The shRNA expression vectors targeting human Del1, and vascular endothelial growth factor (VEGF) were constructed. Gene transfection of Del1-shRNA downregulated expression of Del1 in LS-174T cells in vivo and in vitro, but did not alter the proliferative or survival properties of cells in vitro. Gene transfection of VEGF-shRNA downregulated expression of both VEGF and Del1 in LS-174T cells in vivo and in vitro. Both Del1-shRNA and VEGF-shRNA gene therapies exhibited anti-tumor activities and they also showed a synergistic effect in suppressing growth of colon tumors by anti-angiogenesis and anti-proliferation. Although further investigation to clarify the mechanisms explaining the role of Del1 in tumor growth, and the interaction between VEGF and Del1, is required, the results indicate that downregulation of Del1 presents a potent therapeutic strategy to combat colon cancer.

Show MeSH
Related in: MedlinePlus