Selective cytotoxicity of 3-amino-l-tyrosine correlates with peroxidase activity |
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Authors: | John G Bruno Terence S Herman V Lorenzo Cano Lucy Stribling Johnathan L Kiel |
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Institution: | (1) Department of Radiology, Division of Radiation Oncology, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, 78284 San Antonio, Texas;(2) Veridian Corporation, P.O. Box 35505, 78235 Brooks Air Force Base, Texas;(3) Air Force Research Laboratory, U.S. Air Force, Human Effectiveness Directorate, 78235 Brooks Air Force Base, Texas;(4) U.S. Air Force, AFRL/HEDB, 8315 Hawks Road, Building 1162, 78235 Brooks AFB, Texas |
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Abstract: | In the presence of 3-amino-L-tyrosine (3-AT), abundant brown pigment forms in human HL-60 cells, but not in a variety of other cell lines, which are reported to be lower in mean myeloperoxidase (MPO) content than HL-60. Cells were assessed for peroxidase activity with an ABTS-based colorimetric assay and compared to values obtained with known amounts of human myeloperoxidase. HL-60 cells were estimated to contain the equivalent of 37.1 ng myeloperoxidase/10(6) cells versus 26.1 and 5.0 ng/10(6) cells for human K562 and murine RAW 264.7 cell lines, respectively. HL-60 cells exhibited a nearly 60% inhibition of proliferation and > 70% reduction in cell viability after 4 d of culture in the presence of 100 microg 3-AT per ml. Higher concentrations of 3-AT (up to 400 microg/ml) for 4 d reduced HL-60 proliferation by 80% and decreased viability to 1-3%. Comparable levels of cytotoxicity were achieved in KG-1 cells after 7 d with 200 or 400 microg 3-AT per ml. K562 cells exhibited a 40% reduction in cell number after 7 d with 400 microg 3-AT per ml, but concentrations less than 400 microg/ml did not significantly affect K562 proliferation. K562 viability remained unchanged with doses of 3-AT up to 400 microg/ml. RAW 264.7 cells exhibited unchanged viability and proliferation in the presence of 3-AT at concentrations up to 400 microg 3-AT per ml. K562, KG-1, and RAW 264.7 cells exhibited no evidence of brown pigment formation in the presence of 3-AT and medium containing 10% fetal bovine serum. However, RAW 264.7 cells that were converted to protein-free medium and exposed to 3-AT exhibited intense brown pigment in some cell nuclei. A high percentage of HL-60 cells treated with 3-AT exhibited membrane blebbing, pyknosis, and nuclear fragmentation, which was not observed among other 3-AT-treated cell lines. A mechanism involving toxic intermediates of peroxidase-mediated "aminomelanin" formation is hypothesized. |
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Keywords: | apoptosis HL-60 leukemia melanin myeloperoxidase pyknosis |
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