Microparticles Induce Cell Cycle Arrest Through Redox‐Sensitive Processes in Endothelial Cells: Implications in Vascular Senescence

Abstract

BackgroundChronic disease accelerates endothelial dysfunction in aging, a process associated with cell senescence. However, the mechanisms underlying this process are unclear. We examined whether endothelial cell (EC)-derived microparticles (MPs) facilitate EC senescence and questioned the role of reactive oxygen species in this process.Methods and ResultsSenescence was induced by sequential passaging of primary mouse ECs. Cells retained phenotypic characteristics of ECs from passage 4 through passage 21. Passage 21 ECs exhibited features of senescence, including increased staining of senescence-associated β-galactosidase (SA-βgal), a greater percentage of cells in G1/G0 phase of the cell cycle, and increased phosphorylation of p66Shc (P<0.05). Microparticle formation from passage 21 ECs was increased versus passage 4 ECs (∼2.2-fold increase versus passage 4, P<0.05), and the Rho kinase inhibitor fasudil blocked this increase. Exposure of passage 4 ECs to MPs shifted cells from a proliferating to a nonproliferating phenotype, as indicated by cell cycle analysis and increased senescence-associated β-galactosidase staining. MPs increased EC generation of O2•− (∼2.7-fold) and H2O2 (∼2.6-fold), effects blocked by apocynin (nicotinamide adenine dinucleotide phosphate oxidase inhibitor) and rotenone (mitochondrial oxidase inhibitor) but not by allopurinol (xanthine oxidase inhibitor). MPs increased expression of cell cycle proteins p 21 cip1 and p16ink4a and stimulated phosphorylation of p66Shc in ECs (P<0.05 versus untreated ECs). Pretreatment with the reactive oxygen species scavenger sodium 4,5-dihydroxybenzene-1,3-disulfonate (tiron) abrogated the prosenescent effects of MPs.ConclusionsMPs promote EC senescence through nicotinamide adenine dinucleotide phosphate oxidase- and mitochondrial-derived reactive oxygen species. Such redox-sensitive processes may be important in vascular dysfunction in aging. (J Am Heart Assoc. 2012;1:e001842 doi: 10.1161/JAHA.112.001842.)

Urinary Podocyte Microparticles Identify Prealbuminuric Diabetic Glomerular Injury

“…These data are consistent with our previous reports identifying ROCK as a determinant of endothelial MP formation and highlight the critical role of cytoskeletal reorganization in MP formation. 22,23 A key finding in the present study is that podocyte-derived MPs are present in urine at the earliest stages of diabetic renal injury. Indeed, in STZ-treated mice, podocyte MPs were detectable in urine in advance of albuminuria.…”

“…22,23 NTA Sizing and enumeration of extracellular vesicles were achieved by NTA using a Nanosight LM10 instrument (NanoSight Limited, Amesbury, UK) equipped with NTA 2.3 software. 30 NTA is a light-scattering technique that uses video analysis for sizing and enumeration of extracellular vesicles.…”

Elevated CSF and plasma microparticles in a rat model of streptozotocin-induced cognitive impairment

“…In addition, it has been reported that endothelial MPs increased the production of O 2 − in rat aortic rings [54] and the MPs could promote endothelial cells senescence through NADPH oxidase and mitochondria ROS production [55].Therefore, it is believed that MPs produce the pro-oxidative effects and regulate ROS production. It seems that this property is depending on the source of MPs and the manner in which they are formed [48].…”

Human Endothelial Colony-Forming Cells Protect against Acute Kidney Injury

“…23,30 A total of 50 mg protein was used for analysis, and results are expressed as arbitrary units/mg protein.…”