Fibroblast Growth Factor-2, But Not Vascular Endothelial Growth Factor, Upregulates Telomerase Activity in Human Endothelial Cells

Abstract

Objective-Telomerase plays a major role in the control of replicative capacity, a critical property for successful angiogenesis and maintenance of endothelial integrity. In this study, we examined the relationship between telomerase activity and endothelial cell proliferation as well as the regulation of this enzyme by fibroblast growth factor-2 (FGF-2) and vascular endothelial growth factor-A (VEGF). Methods and Results-Telomerase was repressed in endothelial cells freshly derived from intact endothelium, whereas activity was present during logarithmic growth in culture. In cultured human umbilical vein endothelial cells (HUVECs), mRNA levels of hTERT-the catalytic subunit of telomerase-and enzyme activity decreased reversibly on induction of quiescence. Treatment of quiescent HUVECs with FGF-2 restored telomerase activity in a time-and dose-dependent manner, whereas VEGF had no such effect, although both factors induced comparable mitogenic responses. FGF-2, but not VEGF, upregulated the mRNA levels for hTERT and for the hTERT gene transactivation factor Sp1. Key Words: telomerase Ⅲ fibroblast growth factor-2 Ⅲ endothelial cell Ⅲ Sp1 Ⅲ senescence I nduction of angiogenesis is a novel therapeutic strategy presently being evaluated for the treatment of human ischemic vascular disease. The success of this strategy depends on an adequate proliferative response of vascular endothelial cells at the site of ischemia. 1 Cellular replicative potential is in part limited by the integrity of telomeres, the physical ends of chromosomal DNA. 2 Telomeric DNA shortens by Ϸ50 to 200 base pairs with each round of cell division as a consequence of the inability of conventional DNA polymerases to replicate the 3Ј termini of the template strands. A large body of evidence indicates that critical shortening of one or more telomeres activates a DNA damage checkpoint that blocks additional replication and leads to cell senescence. However, more recently, the integrity of the T-loop-a higher-order structure formed at the end of the telomere by telomeric DNA and specialized proteins-rather than telomere length, per se, has been implicated as a major determinant of senescence. 3,4 In many actively proliferating cells, maintenance of replicative capacity and chromosome stability requires the activity of telomerase, a specialized reverse-transcriptase that adds hexanucleotide repeats of the sequence TTAGGG to the 3Ј ends of nuclear DNA, thus counteracting telomeric erosion. 3 Studies in various immortalized cell lines 5 and normal human lymphocytes 6 have shown that like other enzymes involved in chromosomal DNA synthesis, 7 telomerase activity levels are differentially regulated during periods of proliferation and quiescence. Human telomerase consists of at least 3 subunits. Of these, the RNA component hTR, which provides the template for telomere synthesis, and the telomeraseassociated protein hTEP1 are constitutively expressed. 8,9 The third component, human telomerase reverse transcriptase (hTERT), which contains the catalytic activit...

Sulfhydryl Angiotensin-Converting Enzyme Inhibitor Promotes Endothelial Cell Survival through Nitric-Oxide Synthase, Fibroblast Growth Factor-2, and Telomerase Cross-Talk

“…The mechanism whereby ACEI promote TERT, in the present conditions, seems to be related to the activation of the FGF-2 pathway, because its silencing or neutralization abrogates TERT expression. Although the evidence indicating down-regulation of telomerase by oxidative stress is overwhelming (Haendeler et al, 2004;Voghel et al, 2008;Erusalimsky and Skene, 2009), interventions that favor its up-regulation are limited to NO donor drugs (Vasa et al, 2000), statins (Haendeler et al, 2004), low-dose aspirin (Hu et al, 2008), estrogen (Grasselli et al, 2008), and FGF-2 (Kurz et al, 2003). The direct stimulation of human TERT expression by NO, either endogenously produced or delivered by NO donor agents as shown here, has recently been disputed (Erusalimsky and Skene, 2009).…”

Fibroblast growth factor/fibroblast growth factor receptor system in angiogenesis

“…Accordingly, the study of the transcriptional changes occurring in cultured endothelial cells revealed that, together with a cluster of angiogenesisrelated genes that were similarly modulated by FGF2 and VEGF, the two growth factors affected the expression of distinct subsets of transcripts [107,108]. Accordingly, FGF2, but not VEGF, induces the upregulation of telomerase activity in endothelial cells, thus preventing the early onset of senescence [109]. Distinct patterns of vascular morphology upon FGF2 or VEGF stimulation are described also in the quail embryo CAM assay [86].…”

Chronic oxidative stress compromises telomere integrity and accelerates the onset of senescence in human endothelial cells

“…Aliquots of the cleared lysate equivalent to 2×10 4 cells were assayed for telomerase activity by a modified telomeric repeat amplification protocol (TRAPeze, Intergen, Oxford, UK), as previously described (Kurz et al, 2003). The activity of each sample was calculated from the ratio of the intensity of the telomeric repeat ladder (starting at 50 bp) to the 36 bp internal standard.…”