Phosphatidylserine exposure during early primary necrosis (oncosis) in JB6 cells as evidenced by immunogold labeling technique.
Apoptotic cell death is characterized by the early exposure of phosphatidylserine (PS) at the outer surface of the plasma membrane. The aim of the present study was to examine whether PS exposure also occurs during oncosis (early primary necrosis) and to localize PS at the subcellular level, applying a pre-embedding immunogold labeling technique with biotin conjugated annexin V. The issue was addressed by using caspase-8 deficient, Bcl-2 overexpressing JB6 cells, which die by oncosis when stimulated with synthetic dsRNA. We observed by fluorescence microscopy that oncotic cells with preserved plasma membrane integrity showed PS exposure (annexin+/propidium iodide-). The data was confirmed on the ultrastructural level and PS was localized in oncosis at the outer leaflet of the continuous plasma membrane with preserved trilamellar structure. In postoncotic necrotic cells the immunogold labels were found on the plasma membrane and on the intracellular membranes of the cells, which underwent plasma membrane disruption. In conclusion, this study reveals that PS externalization occurs not only in apoptosis but also in oncosis at least in our cell model system.
Krysko
O
O
Laboratory of Clinical Chemistry, Faculty of Pharmaceutical Sciences, K.U. Leuven, Belgium.
De Ridder
L
L
Cornelissen
M
M
D017304:Annexin A5 / Q000737:chemistry; D053181:Caspase 8; D020169:Caspases / Q000172:deficiency; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D019169:Jurkat Cells; D046529:Microscopy, Electron, Transmission; D008856:Microscopy, Fluorescence; D009336:Necrosis / Q000378:metabolism* / Q000473:pathology; D010718:Phosphatidylserines / Q000032:analysis / Q000378:metabolism*; D011070:Poly I-C / Q000494:pharmacology; D011419:Propidium / Q000737:chemistry; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000494:pharmacology; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D017304:Annexin A5; D010718:Phosphatidylserines; D019253:Proto-Oncogene Proteins c-bcl-2; D012330:RNA, Double-Stranded; D011419:Propidium; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases; D011070:Poly I-C