title: VEGF production by primary human renal proximal tubular cells: requirement of HIF-1, PI3-kinase and MAPKK-1 signaling.
abstract: Renal proximal tubular epithelial cells (PTEC) respond to hypoxia exposure or interleukin-1beta (IL-1beta) treatment with increased vascular endothelial growth factor (VEGF) production. With respect to O2 deprivation, the hypoxia-inducible factor 1alpha/ beta (HIF-1) is the most important transcription factor driving VEGF mRNA expression. HIF-1 is also activated by IL-1beta and may thus be involved in the stimulation of VEGF production by this cytokine. However, the molecular mechanisms of HIF-1 dependent VEGF synthesis are poorly understood. Herein, human PTEC in primary culture were challenged by hypoxic incubation and/or IL-1beta treatment in absence or presence of specific phosphatidylinositol 3-kinase (PI3K) or mitogen activated protein kinase kinase-1 (MAPKK-1) inhibitors for assay of VEGF protein, VEGF mRNA and detection of HIF-1alpha by Western Blotting, EMSA and fluorescence microscopy. In addition, the activities of PI3K and MAPKK-1 were studied following hypoxia and IL-1beta treatment of the cultures. The study shows that PI3K but not MAPKK-1 inhibition resulted in the loss of hypoxic and IL-1beta induced HIF-1alpha accumulation, whereas VEGF synthesis was reduced by either intervention. Thus, PI3K signaling is required for HIF-1alpha accumulation and VEGF synthesis, whereas MAPKK-1 signaling is required for VEGF synthesis only. Furthermore, hypoxia alone was sufficient to activate PI3K in PTEC in contrast to MAPKK-1, whose activity was lowered in hypoxia.
authors.affiliation: Institute of Physiology, University of Lübeck, Germany. hellwig@physio.uni-luebeck.de
authors.forename: Thomas
authors.initials: T
authors.lastname: Hellwig-Bürgel
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Stiehl
authors.forename: Dörthe M
authors.initials: DM
authors.lastname: Katschinski
authors.forename: Jan
authors.initials: J
authors.lastname: Marxsen
authors.forename: Burkhard
authors.initials: B
authors.lastname: Kreft
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Jelkmann
mesh_terms: D015153:Blotting, Western; D015687:Cell Hypoxia / Q000187:drug effects; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005455:Fluorescent Antibody Technique; D006801:Humans; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009025:Morpholines / Q000494:pharmacology; D009687:Nuclear Proteins / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism*; D042442:Vascular Endothelial Growth Factors / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D002867:Chromones; D004268:DNA-Binding Proteins; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; D009025:Morpholines; D009687:Nuclear Proteins; D010100:Oxygen; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D014157:Transcription Factors; D042442:Vascular Endothelial Growth Factors; C497442:HIF1A protein, human; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one