In cultured human umbilical vein endothelial cells (HUVECs), fibroblast growth factor-2 (FGF-2), but not vascular endothelial growth factor-A (VEGF-A), upregulates telomerase activity. Here, we examined the functional significance of this differential regulation on the replicative life span of HUVECs. HUVECs were serially passaged until senescence under four different conditions: (1) EGM-2, a medium containing both VEGF-A and FGF-2; (2) basal medium (BM), consisting of EGM-2 devoid of FGF-2 and VEGF-A; (3) BM supplemented with FGF-2; and (4) BM supplemented with VEGF-A. Cells cultured in BM demonstrated decreased growth rate and ceased to proliferate at approximately 15 population doublings (PDs), whereas those cultured with VEGF-A alone initially proliferated vigorously but arrested growth abruptly at a PD level comparable with cultures grown in BM. In contrast, cells maintained in EGM-2 or in BM/FGF-2 attained a normal replicative life span (approximately 40 PDs). These differences in replicative behavior were reflected by the early appearance of a senescent phenotype in cultures grown in BM or BM/VEGF-A. HUVECs grown in the presence of VEGF-A alone have a decreased life span compared with cultures maintained with FGF-2. This suggests that the upregulation of telomerase activity by FGF-2, an effect not achieved with VEGF-A, plays a functional role in preventing the early onset of senescence.