Ionizing Radiation Regulates Vascular Endothelial Growth Factor-A Transcription in Cultured Human Vascular Endothelial Cells Via the PERK/eIF2α/ATF4 Pathway

Abstract

The delivery of high-dose hypofractionated radiation to a tumor induces vascular damage, but little is known about the responses of vascular endothelial cells to high-dose radiation. We examined whether high-dose irradiation alters vascular endothelial growth factor (VEGF) signaling, which is a critical regulator of the functional integrity and viability of vascular endothelial cells. Human umbilical vein endothelial cells and human coronary artery endothelial cells were treated with 5, 10, 20, or 30 Gy ionizing radiation (IR). Expression values of VEGFA mRNA were analyzed by real-time polymerase chain reaction at 4 hours after irradiation and normalized to the average value of mock-irradiated human umbilical vein endothelial cell or human coronary artery endothelial cell controls. Irradiation with doses higher than 10 Gy causes an acute increase in VEGFA transcript levels, which was accompanied by activation of the PERK/eIF2α/activating transcription factor 4 (ATF4) pathway in human vascular endothelial cells. ATF4 knockdown with siRNA completely prevented the IR-induced upregulation of VEGFA transcripts, and chromatin immunoprecipitation assays demonstrated that ATF4 binding to the VEGFA locus was enriched in response to IR. Postirradiation treatment with an intracellular inhibitor of VEGF signaling significantly enhances high-dose IR-induced apoptosis in human vascular endothelial cells. Human vascular endothelial cells activate PERK/eIF2α/ATF4/VEGF signaling in response to high-dose IR to mitigate the apoptotic response. Thus, for cancer treatment, intracellular inhibitors of VEGF signaling could be employed to enhance stereotactic body radiation therapy-induced vascular damage, which would augment tumor cell death.