Generation and characterization of porcine aortic endothelial cell lines: a tool to study pig endothelial cell/human immune interactions
Introduction The establishment of cell lines allow reproducible 'in vitro' studies and stable genetic modifications which are far more difficult to perform using primary cells. The aim of this study was to establish immortalized porcine aortic endothelial cell lines (AOCs) and assess their suitability to study the interactions between the human immune system and the porcine endothelial cells.
Material and methods Porcine aortic endothelial cells (PAEC) were transfected with the pRNS-1 plasmid (a gift from Dr Ozer; New Jersey Medical School) containing the SV40 genome using Lipofectamine. Eighteen AOCs cell lines were selected and cloned in the presence of G418 selection medium on the basis of their morphology and expression of CD31 (PECAM-1). Immortal AOCs cell lines were compared to PAEC for constitutive and induced expression of MHC Class I, MHC Class II, CD31. E-selectin, VCAM-1, CD11a, CD18 (β2-integrin), CD29 (β1-integrin), CD49d (α4-integrin), CD41/61, CD44, CD45, CD46 and the CD80/86 coestimulatory molecules by flow cytometry.
Results PAEC and the majority of AOCs cell lines displayed similar surface phenotype. Thus, we have observed that constitutive expression of MHC Class I, CD31, CD29, CD41/61, CD46 were conserved between AOCs cell lines and PAEC. Interestingly, an increased expression of CD80/CD86 coestimulatory molecules in the AOCs cell lines was observed when compared to PAEC. However, significant differences was observed in the ability to respond to activation between the cell lines. We have identified several cell lines capable of up-regulate SLA-DR and SLA-DQ molecules upon activation along with the constitutive expression of the coestimularoty CD80/86 molecules. In addition, these cell lines were found appropiate for genetic engineering since they could be easily transfected with foreign genes.
Conclusions This panel of porcine aortic endothelial cell lines would represent important tool to study pig endothelial cell/human immune system interactions and to test new gene therapy approaches in xenotransplantation.