VPF/VEGF acts selectively on the vascular endothelium to enhance permeability, induce cell migration and division, and delay replicative senescence. To understand the changes in gene expression during endothelial senescence, we investigated genes that were differentially expressed in early vs. late passage (senescent) human dermal endothelial cells (HDMEC) using cDNA array hybridization. Early passage HDMEC cultured with or without VPF/VEGF overexpressed 9 and underexpressed 6 genes in comparison with their senescent counterparts. Thymosin beta-10 expression was modulated by VPF/VEGF and was strikingly down-regulated in senescent EC. The beta-thymosins are actin G-sequestering peptides that regulate actin dynamics and are overexpressed in neoplastic transformation. We have also identified senescent EC in the human aorta at sites overlying atherosclerotic plaques. These EC expressed senescence-associated neutral beta-galactosidase and, in contrast to adventitial microvessel endothelium, exhibited weak staining for thymosin beta-10. ISH performed on human malignant tumors revealed strong thymosin beta-10 expression in tumor blood vessels. This is the first report that Tbeta-10 expression is significantly reduced in senescent EC, that VPF/VEGF modulates thymosin beta-10 expression, and that EC can become senescent in vivo. The reduced expression of thymosin beta-10 may contribute to the senescent phenotype by reducing EC plasticity and thus impairing their response to migratory stimuli.