Cytosine arabinoside induces programmed endothelial cell death through the caspase-3 pathway.
The anti-cancer effects of cytosine arabinoside (ARA-C) are well known. However, effects on nonmalignant cells have not been elucidated and may be important to understanding treatment-related toxicity. The purpose of this study was to examine the effect of ARA-C on nondividing vascular endothelial cells. The objectives were to determine the effects of ARA-C on cell viability and to ascertain whether ARA-C caused apoptosis in cultured vascular endothelial cells and hydrocortisone blunted caspase-3-induced apoptosis. Endothelial cells were cultured until confluent and mitotically quiescent then exposed to ARA-C (10(-7)to 10(-3) M) for 1 to 4 days. Some experiments involved cotreatment with hydrocortisone (10(-11),10(-10),10(-4), and 10(-3) M). Light microscopy and the colorimetric MTS assay were used to measure viability. Fluorescent annexin-V and DNA fragmentation assays were used to measure apoptosis, and a fluorescence-based enzymatic assay was used to measure caspase-3 activity, which is one pathway involved in the apoptosis cascade. Two-way ANOVA or the appropriate nonparametric test was used to determine statistical significance in studies of viability and apoptosis. Oneway ANOVA was used to determine statistical significance for caspase-3 activity. Viability was decreased with higher concentrations of ARA-C and increased days of treatment. The percentage of apoptotic cells increased with higher concentrations of ARA-C and increased days of treatment. ARA-C-treated samples showed DNA fragmentation, indicative of apoptosis. Caspase-3 activity increased after ARA-C addition; hydrocortisone blunted this increase. ARA-C caused apoptosis in nondividing endothelial cells in culture. Hydrocortisone may protect against ARA-C-induced apoptosis by reducing caspase-3 activity.
College of Nursing, University of Arizona, Tucson 85721-0203, USA. kmoore@nursing.arizona.edu
Ida M Ki
IM
Moore
Carrie J
CJ
Merkle
Petra
P
Miketova
Renee K
RK
Salyer
Bonny J
BJ
Torres
Richard C
RC
Schaeffer
David W
DW
Montgomery
D000704:Analysis of Variance; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology; D000964:Antimetabolites, Antineoplastic / Q000009:adverse effects*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects* / Q000502:physiology; D002417:Cattle; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003072:Cognition Disorders / Q000139:chemically induced; D003124:Colorimetry; D003561:Cytarabine / Q000009:adverse effects*; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D008859:Microscopy, Polarization; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy; D018709:Statistics, Nonparametric
D000893:Anti-Inflammatory Agents; D000964:Antimetabolites, Antineoplastic; D003561:Cytarabine; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D006854:Hydrocortisone