Xenogeneic human NK cytotoxicity against porcine endothelial cells is perforin/granzyme B dependent and not inhibited by Bcl-2 overexpression.
Because of organ shortages in clinical allotransplantation, the potential of pig-to-human xenotransplantation is currently being explored showing a possible critical role for natural killer (NK) cells in the immune response against xenografts. Therefore, we analyzed the cytotoxic pathways utilized by human natural killer cells (hNK) against porcine endothelial cells (pEC). Transmission electron microscopy of pEC cocultured with hNK cells showed both apoptotic and necrotic cell death, whereas soluble factors such as Fas ligand or TNFalpha did not induce apoptosis in pEC. NK lysis of pEC was abrogated by concanamycin A and ammonium chloride, reagents inhibiting the perforin/granzyme B (grB) pathway, but only partially blocked by caspase inhibition with z-VAD-fmk. Overexpression of bcl-2 protected pEC against apoptosis induced by staurosporine or actinomycin D, but failed to prevent hNK cell-mediated lysis. In conclusion, pEC are lysed in vitro by hNK cells via the perforin/grB pathway and are not protected from NK lysis by overexpression of bcl-2.
Laboratory for Transplantation Immunology, Department of Internal Medicine, University Hospital Zürich, Switzerland.
Ulrike B
UB
Matter-Reissmann
Pietro
P
Forte
Mårten K J
MK
Schneider
Luis
L
Filgueira
Peter
P
Groscurth
Jörg D
JD
Seebach
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015478:Antigens, Heterophile / Q000276:immunology*; D017209:Apoptosis; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured / Q000276:immunology; D018920:Coculture Techniques; D003546:Cysteine Endopeptidases / Q000502:physiology; D003602:Cytotoxicity, Immunologic; D003609:Dactinomycin / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000276:immunology* / Q000648:ultrastructure; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects; D053222:Fas Ligand Protein; D019254:Genes, bcl-2; D006084:Graft Rejection / Q000276:immunology*; D053804:Granzymes; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology* / Q000648:ultrastructure; D018942:Macrolides; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000494:pharmacology / Q000502:physiology*; D008854:Microscopy, Electron; D009336:Necrosis; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000494:pharmacology; D012697:Serine Endopeptidases / Q000502:physiology*; D019311:Staurosporine / Q000494:pharmacology; D013552:Swine / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D025801:Ubiquitin / Q000494:pharmacology
D000590:Amino Acid Chloromethyl Ketones; D000900:Anti-Bacterial Agents; D015478:Antigens, Heterophile; D004791:Enzyme Inhibitors; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D018942:Macrolides; D008562:Membrane Glycoproteins; D052899:Pore Forming Cytotoxic Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D025801:Ubiquitin; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D000643:Ammonium Chloride; D054353:Perforin; D003609:Dactinomycin; C036978:concanamycin A; C509213:GZMB protein, human; D053804:Granzymes; D012697:Serine Endopeptidases; D003546:Cysteine Endopeptidases; D019311:Staurosporine; D002118:Calcium