Hypoxia and interleukin-1beta stimulate vascular endothelial growth factor production in human proximal tubular cells.
BACKGROUND
Vascular endothelial growth factor (VEGF) promotes angiogenesis and inflammatory reactions. VEGF mRNA is detectable in the proximal tubules of inflamed kidneys but not in normals. In other organs VEGF gene expression is induced by hypoxia and cytokines such as interleukin 1 (IL-1). To identify the cellular mechanisms in control of tubular VEGF production, we studied effects of hypoxia and IL-1beta in VEGF mRNA levels, VEGF secretion, and activity of the hypoxia-inducible dimeric transcription factor 1 (HIF-1alpha/beta) in human proximal tubular epithelial cells (PTECs) in primary culture.


METHODS
PTECs were grown in monolayers from human kidneys. Hypoxia was induced by incubation at 3% O2. VEGF mRNA was quantitated by competitive polymerase chain reaction following reverse transcription. VEGF was measured by enzyme-linked immunoassay. HIF-1alpha was demonstrated by Western blot analysis and HIF-1 DNA binding by gel shift assay.


RESULTS
Significant amounts of VEGF mRNA and VEGF protein were measured in PTEC extracts and culture media, respectively. Stimulation of VEGF synthesis at low O2 tension and following IL-1beta treatment was detectable at the protein level only. Nuclear HIF-1alpha protein levels and HIF-1 binding to DNA were also increased under these conditions.


CONCLUSIONS
PTECs in culture produce VEGF. One mechanism of induction appears to be increased DNA binding of HIF-1 to hypoxia-responsive elements in the VEGF gene promoter. In inflammatory diseases of the kidney, tubular cell-derived VEGF may contribute to microvascular leakage and monocyte extravasation.
El Awad
B
B
Institute of Physiology and Department of Internal Medicine I, Medical University of Lübeck, Germany.
Kreft
B
B
Wolber
E M
EM
Hellwig-Bürgel
T
T
Metzen
E
E
Fandrey
J
J
Jelkmann
W
W
D000818:Animals; D015153:Blotting, Western; D002273:Carcinogens / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular; D004268:DNA-Binding Proteins / Q000032:analysis / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000032:analysis / Q000235:genetics*; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D006801:Humans; D000860:Hypoxia / Q000503:physiopathology*; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007375:Interleukin-1 / Q000494:pharmacology*; D007687:Kidney Tubules, Proximal / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D008222:Lymphokines / Q000032:analysis / Q000235:genetics*; D051379:Mice; D009687:Nuclear Proteins / Q000032:analysis / Q000378:metabolism; D010100:Oxygen / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D002273:Carcinogens; D004268:DNA-Binding Proteins; D016228:Endothelial Growth Factors; C497442:HIF1A protein, human; C497443:Hif1a protein, mouse; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007375:Interleukin-1; D008222:Lymphokines; D009687:Nuclear Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D013755:Tetradecanoylphorbol Acetate; D010100:Oxygen