EGF regulation of proximal tubule cell proliferation and VEGF-A secretion

Proximal tubule cell (PTC) proliferation is critical for tubular regeneration and recovery from acute kidney injury. Epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF-A) are important for the maintenance of tubulointerstitial integrity and can stimulate PTC proliferation. We utilized HK-2 cells, an immortalized human PTC line, to characterize the EGF-dependent regulation of VEGF-A secretion and proliferation in PTCs. We demonstrate that EGF stimulates VEGF-A secretion via the EGF receptor (EGFR) and stimulates cell proliferation via activation of the VEGF receptor, VEGFR-2. EGFR activation promotes MAPK (ERK1/2) activation and HIF-1α expression, which are required for basal and EGF-stimulated VEGF-A secretion. EGF also stimulates the phosphorylation of P70S6 kinase (P70S6K), the downstream target of mTORC1. Rapamycin decreased basal and EGF stimulated HIF-1α and enhanced MAPK (ERK1/2) activation, while MAPK (ERK/12) inhibition downregulated HIF-1α expression and the phosphorylation of p70S6K. EGF stimulation of p70S6K was also independent of p-AKT. Inhibition of the mTORC1 pathway with rapamycin abolished phosphorylation of p70S6K but had no effect on VEGF-A secretion, indicating that EGF-stimulated VEGF-A secretion did not require mTORC1 pathway activation. We demonstrate evidence of a complex crosstalk between the MAPK/ERK and mTORC1 pathways, wherein MAPK (ERK1/2) activation stimulates p-P70S6K, while p-P70S6K activation seems to inhibit MAPK (ERK1/2) in EGF-treated HK-2 cells. Our results suggest that EGF stimulates MAPK (ERK1/2) in HK-2 cells, which in turn increases HIF-1α expression and VEGF-A secretion, indicating that VEGF-A mediates EGF-stimulated cell proliferation as an autocrine proximal tubular epithelial cell growth factor.