title: Molecular mechanism of HIF-1-independent VEGF expression in a hepatocellular carcinoma cell line.
abstract: Hypoxia-inducible factor-1 (HIF-1) is a master transcription factor that plays a central role in the hypoxic expression of various genes. Vascular endothelial growth factor (VEGF), a known target gene of HIF-1α, has been shown to be induced by hypoxia through a HIF-1α-independent pathway. HIF-1α dominant-negative lentiviral vectors were introduced to decrease the expression of HIF in Hep3B cells. Cells were incubated under normoxic or hypoxic conditions. We performed a VEGF enzyme-linked immunosorbent assay (ELISA) using cell culture supernatants, and Western blotting using cell lysates. To validate signaling via HIF-1-dependent or HIF-1-independent pathways, we treated cells with an extracellular signal-regulated kinase (ERK) kinase inhibitor, a phosphoinositide 3-kinase (PI3K) inhibitor, and transfected cells with siSP1. HIF-1α protein expression was induced and the levels of VEGF increased under hypoxic conditions. Cells were transfected with siHIF-1α and incubated under normoxic or hypoxic conditions. We found that a significant amount of VEGF was produced by a HIF-1-independent pathway. PI3K inhibitor treatment and siSP1 transient transfection decreased VEGF expression in siHIF-1α-transfected cells. Therefore, VEGF regulation in Hep3B cells is primarily controlled by the Akt/PI3K and SP1 pathways and is independent of HIF-1 under hypoxic conditions.
authors.affiliation: Department of Surgery, Korea University College of Medicine, Korea University Guro Hospital, Guro-gu, Seoul 152-703, Republic of Korea.
authors.forename: Sae Byeol
authors.initials: SB
authors.lastname: Choi
authors.forename: Jong Bae
authors.initials: JB
authors.lastname: Park
authors.forename: Tae-Jin
authors.initials: TJ
authors.lastname: Song
authors.forename: Sang Yong
authors.initials: SY
authors.lastname: Choi
mesh_terms: D060887:B7 Antigens; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004797:Enzyme-Linked Immunosorbent Assay; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
chemical_list: D060887:B7 Antigens; D051793:Hypoxia-Inducible Factor 1; D008565:Membrane Proteins; C467484:VEGFA protein, human; C000588758:VSIR protein, human; D042461:Vascular Endothelial Growth Factor A; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases