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Tumor necrosis factor-α promotes the lymphangiogenesis of gallbladder carcinoma through nuclear factor-κB-mediated upregulation of vascular endothelial growth factor-C.

Du Q, Jiang L, Wang X, Wang M, She F, Chen Y - Cancer Sci. (2014)

Bottom Line: Lymphatic tube formation in vitro was observed in a three-dimensional coculture system consisting of HDLECs and NOZ cell lines, and lymphatic vessels of GBC in nude mice model was detected by immunohistochemistry.TNF-α promoted the tube formation of lymphatic endothelial cells in vitro and the lymphangiogenesis of GBC in nude mice; The nuclear factor (NF)-κB binding site on the VEGF-C promoter was identified using Site-directed mutagenesis, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP).Taken together, TNF-α can upregulate the expression of VEGF-C and promote the lymphangiogenesis of GBC via NF-κB combining with the promoter of VEGF-C.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepatobiliary Surgery, The Affiliated Union Hospital of Fujian Medical University, Fuzhou, China; Key Laboratory of Ministry of Education for Gastrointestinal Cancer, Fuzhou, China.

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Tumor necrosis factor-α (TNF-α) promoted lymphangiogenesis in the orthotopic xenograft model of gallbladder carcinoma (GBC). (a) Three groups of orthotopic xenograft model of gallbladder carcinoma were separately established using three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/Ctrl VEGF-C siRNA (left image). After the treatment with a daily dose of TNF-α (2 μg/kg) for 2 weeks, the tumors were excised. As shown in the middle image, the tumor of the orthotopic xenograft model demonstrated invasive growth with liver and LN metastasis (yellow arrow), In the right image of H-E staining of LN, lymphoid follicle (white arrow) and invasive tumor cells (green arrow) could be observed. (b). Lymphatic vessels (marked by LYVE-1) of the orthotopic xenograft tumors were detected by immunohistochemistry. The brown tubular structures (indicated by red arrows) were lymphatic vessels. (c). Lymphatic vessel number of the orthotopic xenograft tumors. TNF-α increased the lymphatic vessels number of NOZ and NOZ/Ctrl group, while the knockdown of VEGF-C decreased this effect. (*P < 0.05).
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fig05: Tumor necrosis factor-α (TNF-α) promoted lymphangiogenesis in the orthotopic xenograft model of gallbladder carcinoma (GBC). (a) Three groups of orthotopic xenograft model of gallbladder carcinoma were separately established using three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/Ctrl VEGF-C siRNA (left image). After the treatment with a daily dose of TNF-α (2 μg/kg) for 2 weeks, the tumors were excised. As shown in the middle image, the tumor of the orthotopic xenograft model demonstrated invasive growth with liver and LN metastasis (yellow arrow), In the right image of H-E staining of LN, lymphoid follicle (white arrow) and invasive tumor cells (green arrow) could be observed. (b). Lymphatic vessels (marked by LYVE-1) of the orthotopic xenograft tumors were detected by immunohistochemistry. The brown tubular structures (indicated by red arrows) were lymphatic vessels. (c). Lymphatic vessel number of the orthotopic xenograft tumors. TNF-α increased the lymphatic vessels number of NOZ and NOZ/Ctrl group, while the knockdown of VEGF-C decreased this effect. (*P < 0.05).

Mentions: To study the effect of TNF-α on lymphangiogenesis in vivo, we established three orhtotopic xenograft models of GBC in nude mice, which were respectively inoculated the abovementioned three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/VEGF-C siRNA in nude mice. Two weeks after inoculation, a daily dose of TNF-α (2 μg/kg) was injected into the abdominal cavity for 2 weeks. The lymphatic vessels of tumors were observed by immunohistochemistry. Since the D2-40 antibody against lymphatic vessel marker podoplanin does not recognize the murine antigen, mouse lymphatic vessels were detected using LYVE-1.31 The intratumoral lymphatic vessels of the tumors in the nude mice models were hardly to be observed, so all of the lymphatic vessels are from peritumor. As shown in Figure5, TNF-α increased the LVD (14.33 ± 1.35 vs 9.89 ± 0.80, P < 0.05) of orhtotopic xenograft tumors. In the VEGF-C siRNA group, TNF-α also increased the LVD (7.44 ± 0.44 vs 5.89 ± 0.29, P < 0.05) of the tumors, but the increase rate (26.37% ± 4.85%) was lower than that of control (44.19% ± 3.85%) (P < 0.05). Meanwhile, the lymph node metastasis rates of orhtotopic xenograft tumors were increased by TNF-α (Table2).


Tumor necrosis factor-α promotes the lymphangiogenesis of gallbladder carcinoma through nuclear factor-κB-mediated upregulation of vascular endothelial growth factor-C.

Du Q, Jiang L, Wang X, Wang M, She F, Chen Y - Cancer Sci. (2014)

Tumor necrosis factor-α (TNF-α) promoted lymphangiogenesis in the orthotopic xenograft model of gallbladder carcinoma (GBC). (a) Three groups of orthotopic xenograft model of gallbladder carcinoma were separately established using three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/Ctrl VEGF-C siRNA (left image). After the treatment with a daily dose of TNF-α (2 μg/kg) for 2 weeks, the tumors were excised. As shown in the middle image, the tumor of the orthotopic xenograft model demonstrated invasive growth with liver and LN metastasis (yellow arrow), In the right image of H-E staining of LN, lymphoid follicle (white arrow) and invasive tumor cells (green arrow) could be observed. (b). Lymphatic vessels (marked by LYVE-1) of the orthotopic xenograft tumors were detected by immunohistochemistry. The brown tubular structures (indicated by red arrows) were lymphatic vessels. (c). Lymphatic vessel number of the orthotopic xenograft tumors. TNF-α increased the lymphatic vessels number of NOZ and NOZ/Ctrl group, while the knockdown of VEGF-C decreased this effect. (*P < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4462363&req=5

fig05: Tumor necrosis factor-α (TNF-α) promoted lymphangiogenesis in the orthotopic xenograft model of gallbladder carcinoma (GBC). (a) Three groups of orthotopic xenograft model of gallbladder carcinoma were separately established using three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/Ctrl VEGF-C siRNA (left image). After the treatment with a daily dose of TNF-α (2 μg/kg) for 2 weeks, the tumors were excised. As shown in the middle image, the tumor of the orthotopic xenograft model demonstrated invasive growth with liver and LN metastasis (yellow arrow), In the right image of H-E staining of LN, lymphoid follicle (white arrow) and invasive tumor cells (green arrow) could be observed. (b). Lymphatic vessels (marked by LYVE-1) of the orthotopic xenograft tumors were detected by immunohistochemistry. The brown tubular structures (indicated by red arrows) were lymphatic vessels. (c). Lymphatic vessel number of the orthotopic xenograft tumors. TNF-α increased the lymphatic vessels number of NOZ and NOZ/Ctrl group, while the knockdown of VEGF-C decreased this effect. (*P < 0.05).
Mentions: To study the effect of TNF-α on lymphangiogenesis in vivo, we established three orhtotopic xenograft models of GBC in nude mice, which were respectively inoculated the abovementioned three NOZ cell lines: NOZ, NOZ/Ctrl and NOZ/VEGF-C siRNA in nude mice. Two weeks after inoculation, a daily dose of TNF-α (2 μg/kg) was injected into the abdominal cavity for 2 weeks. The lymphatic vessels of tumors were observed by immunohistochemistry. Since the D2-40 antibody against lymphatic vessel marker podoplanin does not recognize the murine antigen, mouse lymphatic vessels were detected using LYVE-1.31 The intratumoral lymphatic vessels of the tumors in the nude mice models were hardly to be observed, so all of the lymphatic vessels are from peritumor. As shown in Figure5, TNF-α increased the LVD (14.33 ± 1.35 vs 9.89 ± 0.80, P < 0.05) of orhtotopic xenograft tumors. In the VEGF-C siRNA group, TNF-α also increased the LVD (7.44 ± 0.44 vs 5.89 ± 0.29, P < 0.05) of the tumors, but the increase rate (26.37% ± 4.85%) was lower than that of control (44.19% ± 3.85%) (P < 0.05). Meanwhile, the lymph node metastasis rates of orhtotopic xenograft tumors were increased by TNF-α (Table2).

Bottom Line: Lymphatic tube formation in vitro was observed in a three-dimensional coculture system consisting of HDLECs and NOZ cell lines, and lymphatic vessels of GBC in nude mice model was detected by immunohistochemistry.TNF-α promoted the tube formation of lymphatic endothelial cells in vitro and the lymphangiogenesis of GBC in nude mice; The nuclear factor (NF)-κB binding site on the VEGF-C promoter was identified using Site-directed mutagenesis, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP).Taken together, TNF-α can upregulate the expression of VEGF-C and promote the lymphangiogenesis of GBC via NF-κB combining with the promoter of VEGF-C.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepatobiliary Surgery, The Affiliated Union Hospital of Fujian Medical University, Fuzhou, China; Key Laboratory of Ministry of Education for Gastrointestinal Cancer, Fuzhou, China.

Show MeSH
Related in: MedlinePlus