title: Triggering of erythrocyte cell membrane scrambling by ursolic acid.
abstract: Ursolic acid (1), a triterpenoid with pleotropic effects including inhibition of tumor growth, is well known to trigger apoptosis of nucleated cells. The effect is at least partially due to altered gene expression and mitochondrial dysfunction. Erythrocytes lack nuclei and mitochondria but, similar to nucleated cells, may undergo suicidal cell death or eryptosis, which is characterized by cell shrinkage and phospholipid scrambling of the cell membrane. Triggers of eryptosis include increase of cytosolic Ca2+ activity ([Ca2+]i), ceramide formation and/or ATP depletion. The present study has investigated whether or not 1 induces eryptosis. [Ca2+]i was estimated from Fluo-3 fluorescence, cell volume from forward scatter, phospholipid scrambling from annexin V binding, hemolysis from hemoglobin release, and cytosolic ATP concentration ([ATP]i) utilizing a luciferase assay and ceramide-utilizing fluorescent antibodies in FACS analysis. As a result, exposure of erythrocytes for 48 h to 1 (≥5 μM) did not significantly modify [ATP]i, but significantly increased [Ca2+]i, stimulated ceramide formation, decreased forward scatter, triggered annexin V binding, and elicited hemolysis. At 5 μM, 1 stimulated phospholipid scrambling in 10% and hemolysis in 2% of treated erythrocytes. Annexin V binding was blunted in the nominal absence of Ca2+. In conclusion, the food component ursolic acid stimulates suicidal death of erythrocytes, i.e., cells devoid of nuclei and mitochondria.
authors.affiliation: Department of Physiology, University of Tuebingen, Gmelinstraße 5, 72076 Tuebingen, Germany.
authors.forename: Kashif
authors.initials: K
authors.lastname: Jilani
authors.forename: Majed
authors.initials: M
authors.lastname: Abed
authors.forename: Christine
authors.initials: C
authors.lastname: Zelenak
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Lang
authors.forename: Syed M
authors.initials: SM
authors.lastname: Qadri
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D000255:Adenosine Triphosphate / Q000032:analysis / Q000378:metabolism; D000814:Aniline Compounds / Q000494:pharmacology; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000032:analysis; D002462:Cell Membrane / Q000378:metabolism*; D002518:Ceramides / Q000276:immunology; D004912:Erythrocytes / Q000187:drug effects*; D006461:Hemolysis / Q000187:drug effects; D015394:Molecular Structure; D014315:Triterpenes / Q000737:chemistry / Q000494:pharmacology*; D014966:Xanthenes / Q000494:pharmacology; D000097245:Ursolic Acid
chemical_list: D000255:Adenosine Triphosphate; D000814:Aniline Compounds; D017304:Annexin A5; D002118:Calcium; D002518:Ceramides; D014315:Triterpenes; D014966:Xanthenes; D000097245:Ursolic Acid; C059715:Fluo-3