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K. Park, J.-X. Ma; Regulation of PEDF Expression in HepG2 by Progesterone via the SREBP Pathway. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4928.
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Despite the extensive studies of the role of pigment epithelium-derived factor (PEDF) in pathological alternations of diabetic retinopathy, the hormonal regulation of PEDF expression is largely unknown. Our recent studies showed that PEDF levels are elevated in the plasma of pregnant women, correlating with progesterone levels. The purpose of this study was to investigate whether progesterone up-regulates PEDF and elucidate the mechanism by which progesterone regulates PEDF expression.
To determine the up-regulation of PEDF by progesterone in vivo, Wistar rats received i.p. injections of progesterone or vehicle every 12 hr for 2 weeks. The progesterone effect was also evaluated in vitro using HepG2, a human cell line derived from the liver. The PEDF promoter activity was assayed by transient transfection of luciferase reporter constructs under the control of the PEDF promoter. Critical promoter region was identified by serial deletion and by the presence of transcription factor- binding sites. Binding of SREBP-1 to the PEDF gene promoter was tested in electrophoretic mobility shift assays (EMSAs).
ELISA and EIA assay showed that plasma progesterone levels and PEDF levels in pregnant woman were significantly correlated at different stages of pregnancy. The rats injected with a dose of 2 or 8 mg/kg of progesterone had significantly increased plasma PEDF concentrations, and increased PEDF protein and mRNA levels in the liver, compared to those injected with vehicle. In cultured HepG2 cells, progesterone up-regulated PEDF mRNA expression and PEDF secretion in a concentration-dependent manner. The PEDF promoter activity was induced by progesterone. As shown by cytohistochemistry and cell fractionation assay, progesterone induced nuclear translocation of SREBP-1. Deletion or mutation of the consensus SREBP-binding sites in the 5'-flanking sequence of the PEDF gene demonstrated that one of the SREBP-binding sites is essential for the progesterone regulation of the PEDF promoter activities. EMSA confirmed that the SREBP-binding site was bound by a nuclear protein. RNA interference (RNAi)-mediated knockdown of SREBP cleavage activating protein (SCAP) production decreased PEDF levels but only those of SREBP-1 had no effect on down-regulation of PEDF expression. SCAP was also identified as an essential factor for the regulation of PEDF transcription by progesterone.
Progesterone up-regulates PEDF transcription in the liver via activating SREBP.
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