Comparison of DiOC(6)(3) uptake and annexin V labeling for quantification of apoptosis in leukemia cells and non-malignant T lymphocytes from children.
Early during apoptosis, there is a reduction in mitochondrial transmembrane potential (MTP) and externalization of phosphatidylserine (PS) in cell membrane prior to eventual cell death. Flow cytometric detection techniques targeting these changes, reduction of DiOC(6)(3) uptake upon the collapse of MTP and annexin V binding to PS have been successfully used to detect apoptotic cells. These methods have given comparable results when cell lines were used. We compared the two different techniques, DiOC(6)(3) uptake and Annexin V-propidium iodide co-labeling in the quantification of cytarabine, vincristine and daunorubicin induced apoptosis on three leukemia cell lines (HL-60, CEM, U937), and bone marrow blasts from 26 children with acute myeloid leukemia, 14 with T cell acute lymphoblastic leukemia. Anti-Fas-induced apoptosis in culture-grown peripheral blood T lymphocytes on 18 samples from 9 children with non-malignant conditions were also studied by these techniques. Our results showed that there is a correlation (P < 0. 05) between the apoptosis rates measured by these two techniques for drug-induced apoptosis in myeloid and lymphoid blasts, and for anti-Fas mAb-induced apoptosis in T lymphocytes. This data suggests that reduction of the MTP and PS externalization may be common to many apoptotic pathways and techniques targeting either of these changes may be used in quantification of apoptosis in different clinical samples.
Ozgen
U
U
Division of Hematology/Oncology, Barbara Ann Karmanos Cancer Institute, Children's Hospital of Michigan, Wayne State University, Detroit, Michigan, USA.
Savaşan
S
S
Buck
S
S
Ravindranath
Y
Y
D017304:Annexin A5; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis; D002232:Carbocyanines; D002648:Child; D003561:Cytarabine / Q000494:pharmacology; D003630:Daunorubicin / Q000494:pharmacology; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes; D018922:HL-60 Cells; D006801:Humans; D007202:Indicators and Reagents; D007938:Leukemia / Q000097:blood; D011419:Propidium; D013601:T-Lymphocytes / Q000187:drug effects*; D014407:Tumor Cells, Cultured / Q000187:drug effects*; D020298:U937 Cells; D014750:Vincristine / Q000494:pharmacology
D017304:Annexin A5; D000903:Antibiotics, Antineoplastic; D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D002232:Carbocyanines; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D003561:Cytarabine; D011419:Propidium; C007392:3,3'-dihexyl-2,2'-oxacarbocyanine; D014750:Vincristine; D003630:Daunorubicin