Abstract 11564: A Key Role of PGC-1α Co-transcription Factor, but not HIF1α Transcription Factor, in Production of Vascular Endothelial Growth Factor (VEGF) by a Novel Angiogenic Agent COA-Cl in Cultured Human Fibroblasts

Abstract

We have recently demonstrated a potent angiogenic effect of a newly developed adenosine-like agent termed COA-Cl (Figure). COA-Cl exerts tube forming activity in human umbilical vein endothelial cells in the presence of normal human dermal fibroblasts (NHDF). We thus hypothesized that COA-Cl induces production of VEGF, a key promoter of angiogenesis, from fibroblasts. The present study addressed this hypothesis and elucidated the mechanism underlying COA-Cl-induced VEGF production. ELISA revealed that COA-Cl (100 μM, 48 h) promoted secretion of VEGF into culture medium of NHDF from 16±15 to 274±52 pg/mL (p<0.01). COA-Cl also up-regulated the expression of VEGF mRNA by 2.1±0.3 fold (RT-PCR, p<0.05). Two transcriptional regulatory proteins have been identified as major activators of VEGF gene: HIF1α, a transcription factor associated with hypoxic insult, and PGC-1α, a co-transcription factor associated with metabolic stress that activates a transcription factor ERRα. Immunoblot and RT-PCR analyses revealed that COA-Cl (100 μM) markedly upregulated the expression of PGC-1α protein (2.1±0.3 fold) and mRNA (4.2±0.5 fold) (p<0.05, respectively). In contrast, COA-Cl had no effect on the expression of HIF1α protein/mRNA in both hypoxia (1% O 2 for 3h) and normoxia (20% O 2 ). Silencing PGC-1α gene by siRNA attenuated the ability of COA-Cl to promote VEGF secretion. HIF1α gene silencing was without effect. When PGC-1α and ERRα were co-transfected in COS-7 cells, COA-Cl (100 μM) upregulated expression of the endogenous VEGF mRNA (2.2±0.3 fold p<0.05 vs. vehicle). COA-Cl had no effect in COS-7 cells transfected with HIF1α. Together, these results demonstrate the ability of COA-Cl to induce VEGF gene expression and protein secretion in fibroblasts. The co-transcription factor PGC-1α, in concert with ERRα, plays a key role in the COA-Cl-induced VEGF production, potentially representing a novel means to induce therapeutic angiogenesis.