title: Stimulation of suicidal erythrocyte death by oridonin.
abstract: Oridonin triggers apoptosis of cancer cells and was suggested as anticancer agent. Oridonin is partially effective through mitochondrial depolarization and partially by modifying gene expression. Erythrocytes lack mitochondria and nuclei but may undergo eryptosis, a suicidal cell death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine exposure at the cell surface. Triggers of eryptosis include increase of cytosolic Ca(2+)-activity, ATP depletion and ceramide formation. The present study explored, whether oridonin triggers eryptosis. Cytosolic Ca(2+)-concentration was estimated from Fluo3-fluorescence, cell volume from forward scatter in FACS analysis, phosphatidylserine exposure from binding of fluorescent annexin V, hemolysis from hemoglobin release, ATP concentration utilizing a luciferin-luciferase assay and ceramide abundance utilizing fluorescent anti-ceramide antibodies. A 48 h exposure to oridonin (≥25μM) significantly increased cytosolic Ca(2+)-concentration, increased ceramide formation, decreased forward scatter and triggered annexin V-binding (the latter in >20% of the erythrocytes). Oridonin didn't decrease ATP concentration and hemolysed <5% of erythrocytes. The effects of oridonin on annexin V binding were partially reversed in the nominal absence of Ca(2+) and by the addition of amiloride (1mM). The present observations reveal a completely novel effect of oridonin, i.e. triggering of Ca(2+) entry and ceramide formation as well as suicidal death of erythrocytes.
authors.affiliation: Department of Physiology, University of Tuebingen, Gmelinstraße 5, D-72076 Tuebingen, Germany.
authors.forename: Kashif
authors.initials: K
authors.lastname: Jilani
authors.forename: Syed M
authors.initials: SM
authors.lastname: Qadri
authors.forename: Christine
authors.initials: C
authors.lastname: Zelenak
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D000255:Adenosine Triphosphate / Q000097:blood; D017304:Annexin A5 / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000097:blood; D002518:Ceramides / Q000097:blood; D045786:Diterpenes, Kaurane / Q000494:pharmacology*; D004912:Erythrocytes / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D010718:Phosphatidylserines / Q000097:blood
chemical_list: D017304:Annexin A5; D000970:Antineoplastic Agents; D002518:Ceramides; D045786:Diterpenes, Kaurane; D010718:Phosphatidylserines; C011959:oridonin; D000255:Adenosine Triphosphate; D002118:Calcium