Endonuclease induced DNA damage and cell death in chemical hypoxic injury to LLC-PK1 cells.
Hypoxia is considered to result in a necrotic form of cell injury. We examined the role of endonuclease activation, considered a feature of apoptosis, in DNA damage and cell death in hypoxic injury in LLC-PK1 cells. Hypoxia in LLC-PK1 cells was induced using a combination of glucose deprivation and a mitochondrial inhibitor, antimycin A (10 microM). Chemical hypoxia caused DNA damage as measured by the alkaline unwinding assay and internucleosomal DNA fragmentation that preceded cell death. Incubating protein extract of cells subjected to chemical hypoxia with calf thymus DNA resulted in oligonucleosome length fragments, which were prevented by an endonuclease inhibitor, aurintricarboxylic acid. Chemical hypoxia resulted in an increased DNA degrading activity with a molecular mass of approximately 15 kDa. Endonuclease inhibitors, aurintricarboxylic acid and Evans blue, prevented antimycin A-induced DNA strand breaks, fragmentation and cell death. We conclude that endonuclease activation plays an important role in chemical hypoxic injury to LLC-PK1 cells.
Hagar
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Division of Nephrology, University of Arkansas for Medical Sciences, Little Rock, USA.
Ueda
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Shah
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D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000968:Antimycin A / Q000494:pharmacology; D001312:Aurintricarboxylic Acid / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D015687:Cell Hypoxia / Q000235:genetics; D004396:Coloring Agents / Q000494:pharmacology; D004249:DNA Damage / Q000502:physiology*; D004720:Endonucleases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005070:Evans Blue / Q000494:pharmacology; D018374:LLC-PK1 Cells / Q000166:cytology / Q000201:enzymology*; D013552:Swine
D000900:Anti-Bacterial Agents; D004396:Coloring Agents; D001312:Aurintricarboxylic Acid; D005070:Evans Blue; D000968:Antimycin A; D004720:Endonucleases