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E.J. Duh, Z. Xu; TFPI–2 is Induced by Vascular Endothelial Growth Factor and Modulates Endothelial Cell Activity . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1836.
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Human tissue factor pathway inhibitor–2 (hTFPI–2) is a 32–kDa serine protease inhibitor that is associated with the extracellular matrix. Recently, hTFPI–2 has been found to have anti–angiogenic activity. The purpose of this study was to investigate the expression of hTFPI–2 in human retinal and umbilical vein endothelial cells and to study the regulation of hTFPI–2 expression in endothelial cells by vascular endothelial growth factor (VEGF). In addition, the effect of recombinant TFPI–2 on endothelial cell proliferation was investigated.
Expression of hTFPI–2 in human retinal endothelial cells (HRECs) and human umbilical vein endothelial cells (HUVECs) was assayed by real–time PCR and western blot analysis. In addition, the effect of VEGF on hTFPI–2 promoter activity was assessed using a luciferase reporter assay system. The effect of recombinant hTFPI–2 on endothelial cells was assessed using a cell proliferation assay.
VEGF induced a dose–dependent increase in hTFPI–2 mRNA and protein expression in endothelial cells. VEGF significantly increased hTFPI–2 (by up to 8–fold) at 12 and 24 hours. This up–regulation was abrogated by pharmacologic inhibition of the ERK signaling pathway. VEGF also significantly upregulated luciferase reporter activity driven by the hTFPI–2 promoter, indicating that VEGF activates hTFPI–2 transcription. Recombinant hTFPI–2 significantly blocked both VEGF– and FGF–2–induced endothelial cell proliferation.
These results indicate that VEGF significantly induces hTFPI–2 transcription in endothelial cells in an ERK–dependent fashion. In addition, hTFPI–2 dramatically blocks growth factor–stimulation of endothelial cell proliferation. Therefore, VEGF–upregulation of hTFPI–2 may represent a mechanism for feedback inhibition of angiogenesis. TFPI–2 could be a therapeutic target for modulation of retinal neovascularization.
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