Regulation of PDGF-B in endothelial cells exposed to cyclic strain

The present study was designed to examine the regulation by cyclic strain of endothelial cell (EC) platelet-derived growth factor-B chain (PDGF-B) expression. We demonstrate in this study that bovine aortic ECs subjected to 10% (but not 6%) average strain resulted in a 2.6-fold increase in PDGF-B steady state mRNA and immunoreactive protein. Nuclear runoff transcription assays confirmed the induction of PDGF-B transcripts. To address the regulation of PDGF-B gene expression by cyclic strain, we transfected bovine aortic ECs with a construct containing 450 bp of human PDGF-B promoter sequence coupled to chloramphenicol acetyltransferase (CAT), and found that subjecting these cells to 10% average strain resulted in a twofold increase in CAT activity by 4 hours. Analysis of nested 5' deletions of the promoter transfected into ECs demonstrated a 55% drop-off in activity between position -313 and -153, with no induction of activity with the - 101-bp minimal promoter. Since a shear stress response element (SSRE) is located at position -125, we tested the hypothesis that the SSRE site was necessary and/or sufficient for induction of PDGF-B activity with strain. Electromobility shift assays revealed that nuclear proteins from ECs exposed to strain for short intervals (30 minutes) bound to the PDGF-B SSRE. However, transfection of ECs with hybrid promoter constructs containing the SV40 sequence promoter downstream of the SSRE or the -153 PDGF-B promoter sequence bearing a mutation in the SSRE demonstrated that the SSRE was not necessary for inducible reporter gene expression in ECs exposed to cyclic strain.