Selective internalization of arachidonic acid by endothelial cells.
Ontology highlight
ABSTRACT:
SUBMITTER: Hall ER
PROVIDER: S-EPMC1148094 | biostudies-other | 1987 Jul
REPOSITORIES: biostudies-other
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Project description:Cell adhesion and migration are essential for the evolution, organization, and repair of living organisms. An example of a combination of these processes is the formation of new blood vessels (angiogenesis), which is mediated by a directed migration and adhesion of endothelial cells (ECs). Angiogenesis is an essential part of wound healing and a prerequisite of cancerous tumor growth. We investigated the effect of the amphiphilic compound arachidonic acid (AA) on EC adhesion and migration by combining live cell imaging with biophysical analysis methods. AA significantly influenced both EC adhesion and migration, in either a stimulating or inhibiting fashion depending on AA concentration. The temporal evolution of cell adhesion area was well described by a two-phase model. In the first phase, the spreading dynamics were independent of AA concentration. In the latter phase, the spreading dynamics increased at low AA concentrations and decreased at high AA concentrations. AA also affected EC migration; though the instantaneous speed of individual cells remained independent of AA concentration, the individual cells lost their sense of direction upon addition of AA, thus giving rise to an overall decrease in the collective motion of a confluent EC monolayer into vacant space. Addition of AA also caused ECs to become more elongated, this possibly being related to incorporation of AA in the EC membrane thus mediating a change in the viscosity of the membrane. Hence, AA is a promising non-receptor specific regulator of wound healing and angiogenesis.
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Project description:BACKGROUND:The interactions between luteal, vascular endothelial, immune cells and its products: steroids, peptide hormones, prostaglandins (PGs), growth factors and cytokines play a pivotal role in the regulation of corpus luteum (CL) function. Luteal endothelial cells undergo many dynamic morphological changes and their action is regulated by cytokines. The aims are: (1) to establish in vitro model for bovine luteal endothelial cells examination; (2) to study the effect of cytokines: tumor necrosis factor alpha (TNFalpha) and interferon gamma (IFNgamma) on cell viability, leukotrienes (LTs) and PG synthases, and endothelin-1 (EDN-1) mRNA, protein expression and their secretion in bovine immortalized luteal endothelial (EnCL-1) cells. METHODS:The primary cultures of bovine luteal endothelial cells were immortalized by transfection with vector carrying the Simian virus 40 T-antigen (SV40 T-ag) sequence. Expression of SV40 T-ag gene in EnCL-1 cells was confirmed by RT-PCR and immunofluorescence staining showed the presence of endothelial cell markers: VE-cadherin and von Willebrand factor. EnCL-1 cells were stimulated by TNFalpha with IFNgamma (50 ng/ml each) for 24 h. Cell viability, mRNA expression (real time RT-PCR), protein expression (western blotting) for LTC4 synthase (LTC4S), LTA4 hydrolase (LTA4H), PGE2 and PGF2alpha synthases and endothelin-1 (EDN-1), and levels of LTs (B4 and C4) and PGs (E2 and F2alpha) and EDN-1 in the medium (EIA) were evaluated. RESULTS:We received immortalized luteal endothelial cell line (EnCL-1). Cytokines did not change EnCL-1 cell viability but increased mRNA expression of LTC4S, LTA4H, PGE2 and PGF2alpha synthases and EDN-1. EDN-1/2/3, LTC4 and PGF2alpha synthases protein expression were elevated in the presence of TNFalpha/IFNgamma, and accompanied by increased EDN-1, LTC4 and PGF2alpha secretion. Cytokines had no effect on PGES and LTA4H protein expression, and PGE2 and LTB4 release. CONCLUSIONS:TNFalpha and IFNgamma modulate EnCL-1 cell function. Moreover, established EnCL-1 cell line appears to be a good model for investigating the molecular mechanisms related to cytokines action and aa metabolites production in cattle.
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