Effect of cellular senescence on the albumin permeability of blood-derived endothelial cells

Abstract

Cheung TM, Ganatra MP, Peters EB, Truskey GA. Effect of cellular senescence on the albumin permeability of blood-derived endothelial cells. Am J Physiol Heart Circ Physiol 303: H1374-H1383, 2012. First published September 28, 2012 doi:10.1152/ajpheart.00182.2012In this study, we tested the hypotheses that endothelial cells (ECs) derived from human umbilical cord blood (hCB-ECs) exhibit low permeability, which increases as hCB-ECs age and undergo senescence, and that the change in the permeability of hCB-ECs is due to changes in tight junction protein localization and the activity of exchange protein activated by cAMP (Epac)1. Albumin permeability across lowpassage hCB-EC monolayers on Transwell membranes was 10 times lower than for human aortic ECs (HAECs) (P Ͻ 0.01) but similar to in vivo values in arteries. Expression of the tight junction protein occludin and tyrosine phosphorylation of occludin were less in hCBECs than in HAECs (P Ͻ 0.05). More hCB-ECs than HAECs underwent mitosis (P Ͻ 0.01). hCB-ECs that underwent Ͼ44 population doublings since isolation had a significantly higher permeability than hCB-ECs that underwent Ͻ31 population doublings (P Ͻ 0.05). This age-related increase in hCB-EC permeability was associated with an increase in tyrosine phosphorylation of occludin (P Ͻ 0.01); permeability and occludin phosphorylation were reduced by treatment with 2 M resveratrol. Tyrosine phosphorylation of occludin and cell age influence the permeability of hCB-ECs, whereas levels of EC proliferation and expression of tight junction proteins did not explain the differences between hCB-EC and HAEC permeability. The elevated permeability in late passage hCB-ECs was reduced by 25-40% by elevation of membrane-associated cAMP and activation of the Epac1 pathway. Given the similarity to in vivo permeability to albumin and the high proliferation potential, hCB-ECs may be a suitable in vitro model to study transport-related pathologies and cell aging. vascular biology; cell aging; occludin; endothelial progenitor cell; permeability

Exchange protein directly activated by cAMP plays a critical role in bacterial invasion during fatal rickettsioses

“…Epac proteins function by responding to increased intracellular cAMP levels and activating the Ras superfamily small GTPases Ras-proximate 1 and 2 (Rap1 and Rap2). Accumulating evidence demonstrates that the cAMP/Epac1 signaling axis plays key regulatory roles in controlling various cellular functions in endothelial cells in vitro, including cell adhesion (18)(19)(20)(21), exocytosis (22), tissue plasminogen activator expression (23), suppressor of cytokine signaling 3 (SOCS-3) induction (24)(25)(26)(27), microtubule dynamics (28,29), cell-cell junctions, and permeability and barrier functions (30)(31)(32)(33)(34)(35)(36)(37). Considering the critical importance of endothelial cells in rickettsioses, we examined the functional roles of Epac1 in rickettsial pathogenesis in vivo, taking advantage of the recently generated Epac1 knockout mouse (38) and Epac-specific inhibitors (39, 40) generated from our laboratory.…”

Endothelial Cell Senescence Increases Traction Forces due to Age-Associated Changes in the Glycocalyx and SIRT1

“…The increased oxidative stress and EC replication can lead to localized senescence. Further, aging endothelium exhibits increased permeability to macromolecules 11 and increased sensitivity to oxidative stress. 10 A localized increase in the permeability of the endothelial layer to proteins 24 is one of the earliest events in atherosclerosis development and influences the progression of atherosclerosis.…”

“…9 Activation of SIRT1 reverses several age-associated phenotypes. 10, 11, 40, 66 In ApoE-knockout mice, overexpression of SIRT1 in ECs decreased atherosclerosis. 40 Furthermore, SIRT1 may regulate permeability through regulation of cell junction proteins, such as occludin.…”

“…40 Furthermore, SIRT1 may regulate permeability through regulation of cell junction proteins, such as occludin. 11 …”

“…In this study, we tested the hypothesis that age-associated changes in the EC glycocalyx or SIRT1 increase cell traction forces due to changes in actin localization and actin filament thickness. To test this hypothesis, we used human cord blood-derived ECs (hCB-ECs) as a model for cell aging because of their low, physiological permeability at low population doublings, 11 high proliferative potential in subconfluent cultures, 11 and similarity to arterial ECs. 3 We examined traction forces of aging ECs and assessed their dependence on activators and inhibitors of SIRT1 and heparan sulfate.…”