VEGF production by primary human renal proximal tubular cells: Requirement of HIF-1, PI3-kinase and MAPKK-1 signaling

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 O-2 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. Copyright (C) 2005 S. Karger AG, Basel.