Abstract
Purpose :
Activating transcriptional factor 3 (ATF3), a member of the ATF/CREB family of transcription factors reportedly takes part in various stress response. This study aimed to investigate whether ATF3 promotes angiogenesis in mice retina.
Methods :
To confirm the expression of ATF3 in vascular endothelial cells (ECs), we performed qPCR on CD31+ CD45- cells that were sorted from retina at postnatal 5 days (P5) and of oxygen induced retinopathy (OIR) model at P17 in C57BL/6 mice. qPCR was also performed in mRNA from Human Retinal Microvascular Endothelial Cells (HRMECs) which were stimulated by VEGF in vitro. Tube formation assays on Matrigel were tested using HRMECs with ATF3 knockdown. We generated EC specific conditional ATF3 knockout (ATF3-ECKO) mice which were obtained by crossing cdh5-BAC-CreERT2 mice and ATF3 flox mice, and then we analyzed postnatal angiogenesis in retina at P5 by immunohistochemical (IHC) analysis for ECs.
Results :
ATF3 mRNA expression are more than ten times higher in CD31+ CD45- ECs than CD31- CD45- non-ECs at P5 retinas (P<0.01). Three times higher expression of ATF3 was also detected in ECs from OIR mice retina compared to control normoxic mice at P17 (p<0.05). qPCR analysis also detected increased ATF3 expression in HRMECs when stimulated by VEGF in vitro (p<0.01). Tube-formation analysis using HRMECs with ATF3 knockdown cultured on Matrigel revealed that decrease of ATF3 expression caused less density of tubes (p<0.001). IHC analysis for CD31 showed that retinas from ATF3-ECKO mice showed less vascular progression length, branch point and tip cells compared to WT mice (P < 0.05, P < 0.01 and P < 0.001, respectively).
Conclusions :
ATF3 expression in retinal ECs were detected both in vitro and in vivo. Moreover, ATF3 deficient mice showed impaired angiogenesis during postnatal period. These results indicated that stress response transcriptional factor ATF3 potentially promotes angiogenesis in retina.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.