Role of osterix in endothelin-1-induced downregulation of vascular endothelial growth factor in osteoblastic cells

Endothelin-1 (ET-1) is produced by vascular endothelial cells to play an important role during bone development, remodeling and repair. ET-1 promotes osteoblastic cell proliferation and differentiation, but has the unique effect of downregulating vascular endothelial growth factor (VEGF) and may thereby control angiogenesis during bone production. Our objectives were to identify the intracellular mechanisms by which ET-1 controls VEGF expression during osteoblastic proliferation and differentiation. ET-1 induced osteoblastic differentiation in rat SBMC-D8 osteoblastic cells, but downregulated expression of VEGF mRNA isoforms (VEGF120, 164 and 188) as demonstrated by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and by use of a luciferase reporter construct containing the promoter region of the VEGF gene. Co-transfection with the endothelin receptor A (ETRA) had the same effect. ET-1 and ETRA both upregulated the transcription factor osterix (Osx). RNA silencing of Osx resulted in an upregulation of VEGF. This study supports the novel inhibitory role for ET-1, via Osx, on VEGF synthesis in osteoblastic cells as a possible mechanism in the temporal and spatial feedback of angiogenesis to bone formation and resorption.