Senescence of aortic endothelial cells in vitro: Influence of culture conditions and preliminary characterization of the senescent phenotype

SUMMARYThe advent of cell culture-based methods for the establishment and expansion of human corneal endothelial cells (CEnC) has provided a source of transplantable corneal endothelium, with a significant potential to challenge the one donor-one recipient paradigm. However, concerns over cell identity remain, and a comprehensive characterization of the cultured CEnC across serial passages has not been performed. To this end, we compared two established CEnC culture methods by assessing the transcriptomic changes that occur during in vitro expansion. In confluent monolayers, low mitogenic culture conditions preserved corneal endothelial cell state identity better than culture in high mitogenic conditions. Expansion by continuous passaging induced replicative cell senescence. Transcriptomic analysis of the senescent phenotype identified a cell senescence signature distinct for CEnC. We identified activation of both classic and new cell signaling pathways that may be targeted to prevent senescence, a significant barrier to realizing the potential clinical utility of in vitro expansion.

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Effects of Dipyrone on Prostaglandin Production by Human Platelets and Cultured Bovine Aortic Endothelial Cells

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657338 ◽

1983 ◽

Vol 49 (02) ◽

pp. 132-137 ◽

Cited By ~ 16

Author(s):

A Eldor ◽

G Polliack ◽

I Vlodavsky ◽

M Levy

Keyword(s):

Endothelial Cells ◽

Arachidonic Acid ◽

Competitive Inhibition ◽

Inhibitory Effect ◽

Human Platelets ◽

Ionophore A23187 ◽

Aortic Endothelial Cells ◽

Bovine Aortic Endothelial Cells ◽

Aortic Endothelial

SummaryDipyrone and its metabolites 4-methylaminoantipyrine, 4-aminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoan- tipyrine inhibited the formation of thromboxane A2 (TXA2) during in vitro platelet aggregation induced by ADP, epinephrine, collagen, ionophore A23187 and arachidonic acid. Inhibition occurred after a short incubation (30–40 sec) and depended on the concentration of the drug or its metabolites and the aggregating agents. The minimal inhibitory concentration of dipyrone needed to completely block aggregation varied between individual donors, and related directly to the inherent capacity of their platelets to synthesize TXA2.Incubation of dipyrone with cultured bovine aortic endothelial cells resulted in a time and dose dependent inhibition of the release of prostacyclin (PGI2) into the culture medium. However, inhibition was abolished when the drug was removed from the culture, or when the cells were stimulated to produce PGI2 with either arachidonic acid or ionophore A23187.These results indicate that dipyrone exerts its inhibitory effect on prostaglandins synthesis by platelets or endothelial cells through a competitive inhibition of the cyclooxygenase system.

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Impact of cigarette versus electronic cigarette aerosol conditioned media on aortic endothelial cells in a microfluidic cardiovascular model

Scientific Reports ◽

10.1038/s41598-021-83511-7 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Om Makwana ◽

Gina A. Smith ◽

Hannah E. Flockton ◽

Gary P. Watters ◽

Frazer Lowe ◽

...

Keyword(s):

Endothelial Cells ◽

Human Monocyte ◽

Conditioned Media ◽

Electronic Cigarette ◽

Microfluidic Channels ◽

Adhesion Assay ◽

Monocyte Adhesion ◽

Aortic Endothelial Cells ◽

Aortic Endothelial

AbstractAtherosclerosis is a complex process involving progressive pathological events, including monocyte adhesion to the luminal endothelial surface. We have developed a functional in vitro adhesion assay using BioFlux microfluidic technology to investigate THP-1 (human acute monocytic leukaemia cell) monocyte adhesion to human aortic endothelial cells (HAECs). The effect of whole smoke conditioned media (WSCM) generated from University of Kentucky reference cigarette 3R4F, electronic cigarette vapour conditioned media (eVCM) from an electronic nicotine delivery system (ENDS) product (Vype ePen) and nicotine on monocyte adhesion to HAECs was evaluated. Endothelial monolayers were grown in microfluidic channels and exposed to 0–1500 ng/mL nicotine or nicotine equivalence of WSCM or eVCM for 24 h. Activated THP-1 cells were perfused through the channels and a perfusion, adhesion period and wash cycle performed four times with increasing adhesion period lengths (10, 20, 30 and 40 min). THP-1 cell adhesion was quantified by counting adherent cells. WSCM induced dose-dependent increases in monocyte adhesion compared to vehicle control. No such increases were observed for eVCM or nicotine. Adhesion regulation was linked to increased ICAM-1 protein expression. Staining of ICAM-1 in HAECs and CD11b (MAC-1) in THP-1 cells demonstrated adhesion molecule co-localisation in BioFlux plates. The ICAM-1 adhesion response to WSCM was downregulated by transfecting HAECs with ICAM-1 siRNA. We conclude that the BioFlux system is able to model human monocyte adhesion to primary human endothelial cells in vitro and WSCM drives the greatest increase in monocyte adhesion via a mechanism involving endothelial ICAM-1 expression.

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Effect of tumour angiogenesis factors on the membrane of capillary and aortic endothelial cells: an e.s.r. spin-label study

Journal of Cell Science ◽

10.1242/jcs.68.1.153 ◽

1984 ◽

Vol 68 (1) ◽

pp. 153-162

Author(s):

N.J. Dodd ◽

S. Kumar

Keyword(s):

Endothelial Cells ◽

Spin Label ◽

Plasma Membranes ◽

Detectable Effect ◽

Aortic Endothelial Cells ◽

Whole Cells ◽

Membrane Order ◽

Migration Factor ◽

Aortic Endothelial

Two distinct factors have been separated from an angiogenic extract of a rat Walker 256 carcinoma, one inducing proliferation and the other migration of capillary endothelial cells in vitro, but having no detectable effect on aortic endothelial cells. The influence of these factors on the order of plasma membranes of these cells was examined by electron spin resonance, using the lipophilic spin label 5-doxyl stearic acid. No detectable effect was observed on treating whole cells or isolated membranes with proliferation factor. In contrast, exposure of capillary endothelial cell membranes to migration factor caused a reduction of membrane order, particularly at temperatures above 30 degrees C. The migration factor had no detectable effect on membrane order of aortic endothelial cells.

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Effect of substrata and fibroblast growth factor on the proliferation in vitro of bovine aortic endothelial cells

Journal of Cellular Physiology ◽

10.1002/jcp.1041090109 ◽

1981 ◽

Vol 109 (1) ◽

pp. 69-81 ◽

Cited By ~ 41

Author(s):

D. Gospodarowicz ◽

G.-M. Lui

Keyword(s):

Endothelial Cells ◽

Growth Factor ◽

Fibroblast Growth Factor ◽

Fibroblast Growth ◽

Aortic Endothelial Cells ◽

Bovine Aortic Endothelial Cells ◽

Proliferation In Vitro ◽

Aortic Endothelial

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Clinopodium tomentosum (Kunth) Govaerts Leaf Extract Influences in vitro Cell Proliferation and Angiogenesis on Primary Cultures of Porcine Aortic Endothelial Cells

Oxidative Medicine and Cellular Longevity ◽

10.1155/2020/2984613 ◽

2020 ◽

Vol 2020 ◽

pp. 1-11

Author(s):

Irvin Tubon ◽

Chiara Bernardini ◽

Fabiana Antognoni ◽

Roberto Mandrioli ◽

Giulia Potente ◽

...

Keyword(s):

Endothelial Cells ◽

Primary Cultures ◽

Ethanolic Extract ◽

Total Phenolic ◽

Dependent Manner ◽

Aortic Endothelial Cells ◽

Phenolic Components ◽

Porcine Aortic Endothelial Cells ◽

Aortic Endothelial

Clinopodium tomentosum (Kunth) Govaerts is an endemic species in Ecuador, where it is used as an anti-inflammatory plant to treat respiratory and digestive affections. In this work, effects of a Clinopodium tomentosum ethanolic extract (CTEE), prepared from aerial parts of the plant, were investigated on vascular endothelium functions. In particularly, angiogenesis activity was evaluated, using primary cultures of porcine aortic endothelial cells (pAECs). Cells were cultured for 24 h in the presence of CTEE different concentrations (10, 25, 50, and 100 μg/ml); no viability alterations were found in the 10-50 μg/ml range, while a slight, but significant, proliferative effect was observed at the highest dose. In addition, treatment with CTEE was able to rescue LPS-induced injury in terms of cell viability. The CTEE ability to affect angiogenesis was evaluated by scratch test analysis and by an in vitro capillary-like network assay. Treatment with 25-50 μg/ml of extract caused a significant increase in pAEC’s migration and tube formation capabilities compared to untreated cells, as results from the increased master junctions’ number. On the other hand, CTEE at 100 μg/ml did not induce the same effects. Quantitative PCR data demonstrated that FLK-1 mRNA expression significantly increased at a CTEE dose of 25 μg/ml. The CTEE phytochemical composition was assessed through HPLC-DAD; rosmarinic acid among phenolic acids and hesperidin among flavonoids were found as major phenolic components. Total phenolic content and total flavonoid content assays showed that flavonoids are the most abundant class of polyphenols. The CTEE antioxidant activity was also showed by means of the DPPH and ORAC assays. Results indicate that CTEE possesses an angiogenic capacity in a dose-dependent manner; this represents an initial step in elucidating the mechanism of the therapeutic use of the plant.

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African Swine Fever Virus Infection of Porcine Aortic Endothelial Cells Leads to Inhibition of Inflammatory Responses, Activation of the Thrombotic State, and Apoptosis

Journal of Virology ◽

10.1128/jvi.75.21.10372-10382.2001 ◽

2001 ◽

Vol 75 (21) ◽

pp. 10372-10382 ◽

Cited By ~ 31

Author(s):

Isabelle Vallée ◽

Stephen W. G. Tait ◽

Penelope P. Powell

Keyword(s):

Endothelial Cells ◽

African Swine Fever Virus ◽

African Swine Fever ◽

Surface Expression ◽

Fever Virus ◽

Parp Cleavage ◽

Aortic Endothelial Cells ◽

Porcine Aortic Endothelial Cells ◽

Aortic Endothelial

ABSTRACT African swine fever (ASF) is an asymptomatic infection of warthogs and bushpigs, which has become an emergent disease of domestic pigs, characterized by hemorrhage, lymphopenia, and disseminated intravascular coagulation. It is caused by a large icosohedral double-stranded DNA virus, African swine fever virus (ASFV), with infection of macrophages well characterized in vitro and in vivo. This study shows that virulent isolates of ASFV also infect primary cultures of porcine aortic endothelial cells and bushpig endothelial cells (BPECs) in vitro. Kinetics of early and late gene expression, viral factory formation, replication, and secretion were similar in endothelial cells and macrophages. However, ASFV-infected endothelial cells died by apoptosis, detected morphologically by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling and nuclear condensation and biochemically by poly(ADP-ribose) polymerase (PARP) cleavage at 4 h postinfection (hpi). Immediate-early proinflammatory responses were inhibited, characterized by a lack of E-selectin surface expression and interleukin 6 (IL-6) and IL-8 mRNA synthesis. Moreover, ASFV actively downregulated interferon-induced major histocompatibility complex class I surface expression, a strategy by which viruses evade the immune system. Significantly, Western blot analysis showed that the 65-kDa subunit of the transcription factor NF-κB, a central regulator of the early response to viral infection, decreased by 8 hpi and disappeared by 18 hpi. Both disappearance of NF-κB p65 and cleavage of PARP were reversed by the caspase inhibitor z-VAD-fmk. Interestingly, surface expression and mRNA transcription of tissue factor, an important initiator of the coagulation cascade, increased 4 h after ASFV infection. These data suggest a central role for vascular endothelial cells in the hemorrhagic pathogenesis of the disease. Since BPECs infected with ASFV also undergo apoptosis, resistance of the natural host must involve complex pathological factors other than viral tropism.

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