Isolation and characterization of immortalized porcine aortic endothelial cell lines

Carrillo, Alberto J.; Chamorro, Sonia; Rodrı́guez-Gago, Manuel; Álvarez, Belén; Molina, Marı́a Jesús; Rodrı́guez-Barbosa, José Ignacio; Sánchez, Alicia; Ramı́rez, Pablo; Muñoz, Antonio; Domı́nguez, Javier; Parrilla, Pascual; Yélamos, José

doi:10.1016/s0165-2427(02)00170-8

Cited by 54 publications

(41 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Parental infectious clone EP#98/2 derived from CSFV strain Alfort Tübingen, the corresponding N pro deleted infectious clone EP # 96/2 and E rns C-H346D clone TF230/1 were a kind gift from Dr G. Meyers (FLI, Tübingen, Germany). Porcine aortic endothelial cells (AOC) (Carrillo et al, 2002) were cultured in RPMI with glutamax (Gibco) plus 10 % fetal calf serum (FCS), 50 IU penicillin ml 21 , 50 mg streptomycin ml 21 and 0.25 mg fungizone ml 21 (Fisher). Porcine kidney cells (PK15), were cultured in MEM with glutamax plus 10 % FCS, 50 IU penicillin ml 21 , 50 mg streptomycin ml 21 and 25 U nystatin ml 21 .…”

Section: Methodsmentioning

confidence: 99%

“…We first investigated whether CSFV infection could circumvent dsRNA-induced apoptotic signalling in a previously characterized porcine aortic endothelial cell line (AOC) (Carrillo et al, 2002). AOC were infected with CSFV strain Alfort 187, and 24 h post-infection they were treated with increasing concentrations of pIpC for 4 h. Activity of the effector caspases, caspases-3 and -7, measured using a luminescent caspase activity assay, increased proportionately with pIpC concentration and reached a plateau at and above 100 mg ml 21 in uninfected cells.…”

Section: Csfv Infection Prevents Pipc-induced Apoptosis Of Aoc Cellsmentioning

confidence: 99%

See 1 more Smart Citation

Classical swine fever virus infection protects aortic endothelial cells from pIpC-mediated apoptosis

Johns

Bensaude

Rocca

et al. 2009

Journal of General Virology

View full text Add to dashboard Cite

Classical swine fever virus (CSFV) causes severe disease in pigs associated with leukopenia, haemorrhage and fever. We show that CSFV infection protects endothelial cells from apoptosis induced by the dsRNA mimic, pIpC, but not from other apoptotic stimuli, FasL or staurosporine. CSFV infection inhibits pIpC-induced caspase activation, mitochondrial membrane potential loss and cytochrome c release as well as the pro-apoptotic effects of truncated Bid (tBid) overexpression. The CSFV proteins N pro and E rns both contribute to CSFV inhibition of apoptosis. We conclude that CSFV infection can inhibit apoptotic signalling at multiple levels, including at the caspase-8 and the mitochondrial checkpoints. By supporting viral replication, endothelial cells may promote CSFV pathogenesis. INTRODUCTIONClassical swine fever virus (CSFV), a pestivirus within the family Flaviviridae, is related in terms of its sequence and genome organization to other members of the flavivirus group that give rise to human diseases such as hepatitis C and dengue fever. CSFV is a positive-sense RNA virus, and is the causative agent of classical swine fever (CSF), a notifiable disease of pigs (Le Potier et al., 2006). Disease outbreaks are associated with considerable economic loss and adverse effects on animal welfare (Paton & GreiserWilke, 2003). In its most virulent form, CSFV infection causes severe immunopathological signs, including leukopenia, haemorrhage and fever, accompanied by high morbidity and mortality. However, within an infected population, some animals are found to recover or develop chronic disease (Le Potier et al., 2006). These observations suggest a complex interplay between the virus and the host immune system. CSFV infection is accompanied by depression of cellular immune defences (Ganges et al., 2008) and particularly innate responses mediated by interferon (Bensaude et al., 2004; La Rocca et al., 2005; Ruggli et al., 2005;Seago et al., 2007). In vivo, elevated levels of cell death by apoptosis are thought to account for the characteristic depletion of circulating leukocytes (Sanchez-Cordon et al., 2002;Summerfield et al., 2000Summerfield et al., , 2001. However, it has been found that apoptotic cells rarely contain detectable viral antigen, indicating that apoptotic cells are generally not CSFV-infected (Choi et al., 2004;Sato et al., 2000;Summerfield et al., 2001). This finding, together with the observation that acutely infected animals develop high viral loads, suggest a possible role for cell populations that are resistant to virus-induced apoptosis. These may include endothelial cells.Apoptosis is a controlled cell death programme employed by multicellular organisms to eliminate damaged, aberrant or infected cells (Benedict et al., 2002). A characteristic of apoptotic cell death is the activation of the caspase family of proteases. Caspases that function near the apex of cell death cascades are designated initiators and include caspases-8 and -9, whereas those involved in the terminal stages of cell death, such as ...

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Csfv Infection Prevents Pipc-induced Apoptosis Of Aoc Cellsmentioning

confidence: 99%

Classical swine fever virus infection protects aortic endothelial cells from pIpC-mediated apoptosis

Johns

Bensaude

Rocca

et al. 2009

Journal of General Virology

View full text Add to dashboard Cite

show abstract

“…Basal and inducible expression of PULBP and PMIC2 was also examined in AOC cells, a porcine aortic endothelial cell line (Carrillo et al, 2002). Although not a quantitative assay, 1-3, respectively), was analyzed by Southern blotting using a 437 bp fragment of PULBP cDNA as a probe as described in Section 2.…”

Section: Expression Of the Potential Porcine Nkg2d Ligands Pulbp Andmentioning

confidence: 99%

Molecular cloning and characterization of a porcine UL16 binding protein (ULBP)-like cDNA

Garcia-Borges

Phanavanh

Saraswati

et al. 2005

Molecular Immunology

View full text Add to dashboard Cite

UL16 binding proteins (ULBPs) are ligands for the NK cell activating receptor NKG2D. A cDNA encoding a porcine ULBP-like protein (PULBP) was cloned and the predicted amino acid sequence exhibited 35-52% identity to human ULBPs. Southern blot analysis suggested that there is only one ULBP-like gene in the pig genome. Transcripts of PULBP and another potential NKG2D ligand, MIC2, were detected by RT-PCR in a wide range of tissues. Recombinant PULBP-Fc and human ULBP2-Fc fusion proteins were made and used to examine PULBP binding to porcine PBMCs and a human NK cell line (NKL cells). PULBP-Fc bound to a subpopulation of porcine PBMCs but not NKL cells. Conversely, human ULBP2-Fc did not bind to porcine PBMCs but did stably interact with NKL cells.

show abstract

“…A recent study performed in vitro with porcine primary endothelial cell cultures has shown that the replication of CSF activates cell inflammatory responses but at the same time suppresses IFN production and apoptotic pathways [3]. The availability of an immortalized porcine aortic endothelial cell line (AOC cells) [6] prompted us to investigate the effect of the CSF virus infection on the expression of swine relevant markers, cytokines and procoagulant activity. AOC cells maintain most phenotypical and functional characteristics of primary endothelial cell cultures and constitute a useful model for the study of CSF pathogenesis in vitro.…”

Section: E Campos Et Almentioning

confidence: 99%

“…AOC cells, immortalized by transfection with a plasmid containing the complete genome of SV40 [6], were kindly provided by Dr Yelamos (Virgen de la Arrixaca Hospital, Murcia, Spain). These cells were plated onto flasks coated with 1% gelatin (Difco, USA) and cultured in RPMI 1640 (BioWhittaker, Verviers, Belgium), supplemented with 20% FCS and 2 mM L-glutamine.…”

Section: Cellsmentioning

confidence: 99%

In vitro effect of classical swine fever virus on a porcine aortic endothelial cell line

Campos¹,

Revilla²,

Chamorro³

et al. 2004

Vet. Res.

Self Cite

View full text Add to dashboard Cite

-The effect of classical swine fever (CSF) virus on some phenotypic and functional features of an established porcine aortic endothelial cell (AOC) line was investigated. AOC cells show most of the characteristics of primary endothelial cells, avoiding the alterations and senescence that these cells undergo after a few passages in culture. AOC cells were susceptible to CSF virus infection to a high degree, reaching 90% of CSF virus positive cells after 24 h of infection; however as with other porcine susceptible cells, no cytopathic effect could be observed. In these conditions none of the surface molecules studied, including SLA-II MHC antigens, adhesion or co-stimulatory molecules, were altered by virus infection after 24 or 48 h. Functionally CSF virus infection induced a decrease in the pro-coagulant activity of the AOC cells, determined by the increase in the clot formation time shown by the lysates of these cells. This contrasts with the increase observed in the expression of mRNA corresponding to IL-1α and IL-6, two proinflammatory and pro-coagulant cytokines, in CSF virus-infected AOC cells.classical swine fever / endothelial cells / cytokines / pro-coagulant activity

show abstract

Isolation and characterization of immortalized porcine aortic endothelial cell lines

Cited by 54 publications

References 37 publications

Classical swine fever virus infection protects aortic endothelial cells from pIpC-mediated apoptosis

Classical swine fever virus infection protects aortic endothelial cells from pIpC-mediated apoptosis

Molecular cloning and characterization of a porcine UL16 binding protein (ULBP)-like cDNA

In vitro effect of classical swine fever virus on a porcine aortic endothelial cell line

Contact Info

Product

Resources

About