Abstract 120: Notch Signaling Negatively Regulates Vascular Endothelial Cell Senescence

Abstract

Accumulation of senescent vascular cells occurs in aged vessels, which leads to an increase of inflammation and a decline of regenerative potential, thereby promoting vascular dysfunction and atherosclerosis. However, the molecular mechanisms of these age-related changes remain unclear. The Notch pathway is a highly conserved signaling that controls cell fate determination and differentiation during the development of various tissues. In adults, Notch signaling has been reported to be essential for neovascularization and is implicated with the age-associated conditions such as cancer, neuronal disorder and impaired regeneration of skeletal muscle. Here, we show that Notch signaling has a crucial role in endothelial cell senescence. We found that inhibition of Notch signaling, using short hairpin RNA (shRNA) targeting Notch1, reduced the maximum population doublings of human umbilical vein endothelial cells (HUVEC), increased the activity of senescence-associated beta-galactosidase, and up-regulated the expression of aging-associated molecules such as p53, p21, and p16. Knockdown of the Notch ligand Jagged1 resulted in attenuation of Notch signaling, thereby inducing premature senescence in a way similar to knockdown of Notch1. In contrast, over-expression of Notch1 or Jagged1 extended the replicative lifespan of HUVEC and decreased the expression of aging-associated molecules. To elucidate the mechanism how inhibition of Notch signaling induces premature senescence in endothelial cells, we examined expression of various molecules by microarray analysis, and found that the expression of Id1 (inhibitor of DNA binding 1) and MKP1 (MAP kinase phosphatase 1) was significantly down-regulated by knockdown of Notch. Over-expression of Id1 improved Notch1 inhibition-induced premature senescence, which is associated with a decrease of p16 expression. Likewise, treatment with SB203580, an inhibitor of p38 MAPK, extended lifespan of Notch-deleted endothelial cells along with down-regulation of p16. These results suggest that Notch signaling and the downstream molecules (Id1 and p38) regulate endothelial cell senescence presumably via a p16-dependent pathway, and may be a new target of the therapy for age-associated vascular diseases.