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

Vascular endothelial growth factor ( VEGF )‐C is an important lymphangiogenic factor involved in the lymphangiogenesis of gallbladder carcinoma ( GBC ) and the lymph node metastasis of the tumor. Tumor necrosis factor ( TNF )‐α, a key inflammatory cytokine responding to chronic inflammation of GBC , has been reported to stimulate the expression of VEGF ‐C in some nonneoplastic cells. But whether TNF ‐α promotes the expression of VEGF ‐C in GBC has yet to be determined. Therefore, in the present study, the concentration of TNF ‐α and VEGF ‐C and the lymphatic vessel density ( LVD ) in the clinical GBC specimens were analyzed, and a linear correlation was found between the concentration of TNF ‐α and that of VEGF ‐C, the lymphatic vessel density ( LVD ); The transcription and protein level of VEGF ‐C in NOZ cell line were detected by real‐time polymerase chain reaction ( PCR ) and enzyme linked immunosorbent assay ( ELISA ), and TNF ‐α enhanced the expression of VEGF ‐C in NOZ cell lines in a dose and time‐dependent manner. Lymphatic tube formation in vitro was observed in a three‐dimensional coculture system consisting of HDLEC s 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 (Ch IP ). 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.