Regulation ofEPC-1/PEDF in normal human fibroblasts is posttranscriptional

Abstract

The EPC-1 (early population doubling level cDNA-1) gene, also known as pigment epithelium-derived factor, encodes a protein belonging to the serine protease inhibitor (serpin) superfamily that has been reported to inhibit angiogenesis and proliferation of several cell types. We have previously reported that the EPC-1 mRNA and the secreted EPC-1 protein are expressed at levels more than 100-fold higher in early passage, G(0), WI-38 cells compared to either proliferating or senescent WI-38 fibroblasts. To examine the molecular mechanisms that regulate changes in EPC-1 gene expression in WI-38 cells, we isolated and characterized the human EPC-1 gene and determined the mRNA cap site. Transcriptional assays showed no change in the transcription rates of EPC-1 between young proliferating, quiescent, and senescent WI-38 cells. These results suggest posttranscriptional regulation of the EPC-1 gene. Reverse transcriptase polymerase chain reaction measurements (of hnRNA) indicate regulation at the hnRNA level. The regulation of the EPC-1 gene at the level of hnRNA can explain the observed slow increase in the steady-state EPC-1 mRNA levels when cells become quiescent. The reduction of EPC-1 mRNA levels that occurs when cells exit G(0) and are induced to proliferate can be accounted for by a reduction of the EPC-1 mRNA stability in stimulated cells as compared to quiescent cells.

Pigment epithelium-derived factor as a multifunctional regulator of wound healing

“…There is evidence that secreted PEDF is chemotactic for FBs (50) and serves as a regulator of FB cell cycle causing quiescence (43). It is thus feasible that PEDF, which is produced by quiescent FBs (15,44) (54). VEGF binds to the provisional ECM and promotes EC proliferation and migration, leading to angiogenesis, as well as EC permeability and leakiness of wound vasculature (22).…”

“…While various tumors (58) and ischemic hearts (49) express relatively low levels of PEDF, a recent study showed that high systemic levels of PEDF may be etiologic for impaired healing in diabetics due to excessive antiangiogenesis (45). PEDF is produced by quiescent FBs (15,44) in normoxic conditions (49) and by keratinocytes (KCs) (14), and it is readily secreted into the ECM. Analyses of its amino acid sequence (34) and its crystal structure (54) have revealed distinct binding sites for ECM components collagen-1 (35,39) and glycosaminoglycans (3), including heparin (59,66) and hyaluronan (6).…”

Identification of molecules derived from human fibroblast feeder cells that support the proliferation of human embryonic stem cells

“…PEDF is also considered to be involved in the maintenance of adult 'niches' of NSCs and to promote their proliferation via the activation of signalling pathways, while a blockade of endogenous PEDF decreases the proliferation [12]. Most interestingly, a few independent studies have linked alterations of PEDF expression in human fibroblast and fibroblast-like cells undergoing replicative senescence [13][14][15], which illustrates that senescent cells may not be able to produce PEDF transcripts. Previous studies have identified PEDF in a mouse and human fibroblast cell-conditioned medium, suggesting that this protein could also be a potential regulator of hESC.…”

Endogenous Inhibitors of Angiogenesis

“…Its antiangiogenic activity is selective, in that PEDF targets only new vessel growth but spares existing ones, and it is reversible. It is a protein that is highly up-regulated in the G o phase of early-passage cells compared with rapidly proliferating or senescent cells and thus is also linked to both cell cycle and cell senescence (168,169).…”