VEGF Production by Primary Human Renal Proximal Tubular Cells: Requirement of HIF-1, PI3-Kinase and MAPKK-1 Signaling

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Renal proximal tubular epithelial cells (PTEC) respond to hypoxia exposure or interleukin-1β (IL-1β) treatment with increased vascular endothelial growth factor (VEGF) production. With respect to O₂ deprivation, the hypoxia-inducible factor 1α/ β (HIF-1) is the most important transcription factor driving VEGF mRNA expression. HIF-1 is also activated by IL-1β 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-1β 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-1α by Western Blotting, EMSA and fluorescence microscopy. In addition, the activities of PI3K and MAPKK-1 were studied following hypoxia and IL-1β treatment of the cultures. The study shows that PI3K but not MAPKK-1 inhibition resulted in the loss of hypoxic and IL-1β induced HIF-1α accumulation, whereas VEGF synthesis was reduced by either intervention. Thus, PI3K signaling is required for HIF-1α 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.