Transgenic expression of HLA-E single chain trimer protects porcine endothelial cells against human natural killer cell-mediated cytotoxicity.
BACKGROUND
The susceptibility of porcine endothelial cells (pEC) to human natural killer (NK) cells is related to the failure of human major histocompatibility complex (MHC)-specific killer inhibitory receptors to recognize porcine MHC class I molecules. The aims of this study were (i) to assess the protection of pEC against xenogeneic NK-mediated cytotoxicity afforded by the stable expression of HLA-E single chain trimers (SCT) composed of a canonical HLA-E binding peptide antigen, VMAPRTLIL, the mature human beta2-microglobulin, and the mature HLA-E heavy chain, and (ii) to test whether HLA-E expression on pEC and porcine lymphoblastoid cells affects the adhesion of human NK cells.


METHODS
Porcine EC lines expressing different levels of HLA-E SCT were generated by Ca(2)PO(4)-transfection followed by limiting dilution cloning. Surface expression of HLA-E was measured by flow cytometry. Susceptibility of transfected pEC lines against human NK cells was tested in (51)Cr-release cytotoxicity assays. Interactions between human NK cells and HLA-E positive pEC or porcine lymphoblastoid cells were further addressed in adhesion and conjugation assays.


RESULTS
The level of protection of pEC from human NK-mediated cytotoxicity correlated with the intensity of surface HLA-E expression. Furthermore, the HLA-E SCT-mediated protection was specifically reversed by blocking the HLA-E specific NK inhibitory receptor CD94/NKG2A. HLA-E expression does neither affect the adhesion of human NK cells to pEC nor the heteroconjugate formation between human NK and porcine 13271.10 cells.


CONCLUSIONS
Stable surface expression of HLA-E on pEC was achieved in the absence of extrinsic peptide pulsing and provided partial protection from human NK cytotoxicity. Though insufficient to inhibit xenogeneic NK cell reactivity completely, transgenic HLA-E expression on pig organs might contribute to a successful application of clinical xenotransplantation in combination with other protective strategies.
Lilienfeld
Benjamin G
BG
Laboratory for Transplantation Immunology, Department of Internal Medicine, University Hospital Zürich, Zürich, Switzerland.
Crew
Mark D
MD
Forte
Pietro
P
Baumann
Bettina C
BC
Seebach
Jörg D
JD
D000818:Animals; D000910:Antibodies, Heterophile / Q000276:immunology; D002448:Cell Adhesion / Q000502:physiology; D002450:Cell Communication / Q000276:immunology / Q000502:physiology; D002460:Cell Line; D003602:Cytotoxicity, Immunologic / Q000276:immunology* / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology / Q000276:immunology* / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology*; D006680:HLA Antigens / Q000235:genetics / Q000502:physiology*; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology* / Q000502:physiology; D051920:NK Cell Lectin-Like Receptor Subfamily D / Q000276:immunology; D013552:Swine; D014162:Transfection; D019076:Transgenes / Q000235:genetics / Q000502:physiology*; D014183:Transplantation, Heterologous / Q000379:methods; D000097625:HLA-E Antigens
D000910:Antibodies, Heterophile; D006680:HLA Antigens; D015395:Histocompatibility Antigens Class I; D051920:NK Cell Lectin-Like Receptor Subfamily D; D000097625:HLA-E Antigens