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...

The formation of the vascular network requires a tightly controlled balance of pro-angiogenic and stabilizing signals. Perturbation of this balance can result in dysregulated blood vessel morphogenesis and drive pathologies including cancer. Here, we have identified a novel gene, ARHGAP18, as an endogenous negative regulator of angiogenesis, limiting pro-angiogenic signaling and promoting vascular stability. Loss of ARHGAP18 promotes EC hypersprouting during zebrafish and murine retinal vessel development and enhances tumor vascularization and growth. Endogenous ARHGAP18 acts specifically on RhoC and relocalizes to the angiogenic and destabilized EC junctions in a ROCK dependent manner, where it is important in reaffirming stable EC junctions and suppressing tip cell behavior, at least partially through regulation of tip cell genes, Dll4, Flk-1 and Flt-4. These findings highlight ARHGAP18 as a specific RhoGAP to fine tune vascular morphogenesis, limiting tip cell formation and promoting junctional integrity to stabilize the angiogenic architecture.

Cerebral cavernous malformations (CCMs) are vascular lesions predominantly developing in the central nervous system (CNS), with no effective treatments other than surgery. Lossof-function mutation in CCM1/krev interaction trapped 1 (KRIT1), CCM2, or CCM3/programmed cell death 10 (PDCD10) causes lesions that are characterized by abnormal vascular integrity. Vascular endothelial cadherin (VE-cadherin), a major regulator of endothelial cell (EC) junctional integrity is strongly disorganized in ECs lining the CCM lesions. We report here that microRNA-27a (miR-27a), a negative regulator of VE-cadherin, is elevated in ECs isolated from mouse brains developing early CCM lesions and in cultured ECs with CCM1 or CCM2 depletion. Furthermore, we show miR-27a acts downstream of kruppel-like factor (KLF)2 and KLF4, two known key transcription factors involved in CCM lesion development. Using CD5-2 (a target site blocker [TSB]) to prevent the miR-27a/VE-cadherin mRNA interaction, we present a potential therapy to increase VE-cadherin expression and thus rescue the abnormal vascular integrity. In CCM1-or CCM2-depleted ECs, CD5-2 reduces monolayer permeability, and in Ccm1 heterozygous mice, it restores dermal vessel barrier function. In a neonatal mouse model of CCM disease, CD5-2 normalizes vasculature and reduces vascular leakage in the lesions, inhibits the development of large lesions, and significantly reduces the size of established lesions in the hindbrain. Furthermore, CD5-2 limits the accumulation of inflammatory cells in the lesion area. Our work has established that VE-cadherin is a potential therapeutic target for normalization of the vasculature and highlights that targeting miR-27a/VE-cadherin interaction by CD5-2 is a potential novel therapy for the devastating disease, CCM.