Stress-induced premature senescence mediated by a novel gene, SENEX, results in an anti-inflammatory phenotype in endothelial cells

Abstract

Cellular senescence is a mechanism to inhibit the growth of mammalian cells after oncogenic activation, or in response to damage or stress. We describe here the identification of a novel gene, SENEX, that regulates stress induced premature senescence pathways in endothelial cells (ECs) involving p16 INK4a and retinoblastoma protein activation. Endogenous levels of SENEX remain unchanged during replicative senescence but are regulated by H 2 O 2 -mediated stress. In contrast to that previously described for senescence in other cell types, the SENEX induced senescent ECs are profoundly anti-inflammatory. The cells are resistant to tumor necrosis factor (TNF)␣-induced apoptosis, adhesion of neutrophils and mononuclear cells, and the surface (but not cytoplasmic) expression of endothelial leukocyte adhesion molecule 1 and vascular cell adhesion molecule 1. Furthermore they are resistant to thrombin induced vascular leak. Senescent ECs such as those lining atherosclerotic lesions may therefore function to limit the inflammatory response. SENEX is also essential for EC survival since depletion either ectopically by siRNA or by highdose H 2 O 2 treatment causes apoptosis. Together, these findings expand our understanding of the role of senescence in the vasculature and identify SENEX as a fulcrum for driving the resultant phenotype of the endothelium after activation. IntroductionCellular senescence together with apoptosis is viewed as a major pathway to control cell proliferation and suppress tumorigenesis. 1,2 Recent evidence suggests that the senescence program may have a broader role, as an active mechanism to limit disease progression. 3 The recognition and impact of senescence on the vascular system is only just emerging. Increased numbers of senescent endothelial cells (ECs) are found in mature atherosclerotic plaques, in vessels from diabetic patients, in postangioplastic restenotic vessels, in coronary vessels of patients with ischemic heart disease, and in hypertensive patients (reviewed in Voghel et al 4 ). Senescent ECs have also been identified in the tumor vasculature in glioma. 5 However, the causes and consequences of these senescent ECs in the different pathologies have not been clearly defined.The recognition of senescent cells relies on several specific criteria. The cells exit the cell cycle but remain viable, they exhibit a large flattened morphology, 6 and show accumulation of senescenceassociated ␤-galactosidase (SA-␤-gal) activity. 7 In addition, they show altered genetic profiles which are likely to be cell type specific. 8 There are 2 broad forms of senescence, replicative and stress induced. Replicative senescence (RS) is mediated through the shortening of telomeres that occurs during each cell division. This shortening eventually registers as DNA damage and triggers ataxia telangiectasia mutated kinase (ATM) activation and initiates a program of cell cycle arrest. 9 Stress induced premature senescence (SIPS) is induced by oncogene activity, 10 oxidative stress, 11 or suboptimal culture cond...

Accelerated senescence of cord blood endothelial progenitor cells in premature neonates is driven by SIRT1 decreased expression

“…We found that PT-associated senescence is not dependent on telomere shortening or activation of the p53/p21 WAF pathway. In- phenotype involving p16 INK4a without significant increase in p21 is consistent with previous observations in endothelial cells exposed to oxidative stress 28 and the possible independent role of these 2 pathways in senescence induction. 22 Interestingly, in healthy elderly, p16…”

ARHGAP18: an endogenous inhibitor of angiogenesis, limiting tip formation and stabilizing junctions

“…ARHGAP18 exists as 2 isoforms, with the smaller isoform originating from a downstream start codon. 23,26 Knockdown of ARHGAP18 with siRNAs resulted in the reduced expression of both isoforms (Fig. S1A).…”

“…23 HUVECs were seeded at 1.8 £ 10 6 cells in 10 cm dishes overnight and transduced with adenoviral particles in 2% (v/v) FCS media containing 8 mg/mL polybrene (Sigma-Aldrich) for 2 h. EC were used following 1 d transduction.…”

“…23,24 Relative to other RhoGAPs, ARHGAP18 is highly expressed in EC. 25 More importantly, ARHGAP18 was found to be downregulated in the early migration phase and upregulated in the later stabilization phase of tube formation.…”

“…Consistent with this, knockdown of ARHGAP18 resulted in the inability to form stable tubes. 23 Interestingly, overexpression of ARHGAP18 in EC results in the induction of premature senescence, although this is through a GAP-independent mechanism, 23 but dependent on caveolae formation (Powter et al, in press). Here, we describe ARHGAP18 as a novel negative regulator of sprouting by acting dualistically to limit TC formation and to maintain junctional integrity.…”