Nitric oxide-mediated renal epithelial cell injury during hypoxia and reoxygenation.
The potent endothelial-derived vasodilator nitric oxide (NO) has been identified as a protective agent in acute renal failure. However, some recent studies have suggested a detrimental effect of NO on rat proximal tubules exposed to hypoxia and reoxygenation. We determined whether NO metabolites cause intracellular oxidation during hypoxia and reoxygenation and whether this oxidative stress is linked to irreversible cell injury. Primary cultures of rat proximal tubular epithelial cells were studied in a subconfluent stage and subjected to 60 min hypoxia and 30 min reoxygenation. Intracellular oxidation was assessed by monitoring the conversion of nonfluorescent dihydrorhodamine 123 (DHR) to fluorescent rhodamine 123 as a probe for the long-lived oxidant peroxynitrite. Hypoxia and reoxygenation produced a marked increase in cellular generation of oxidant species. Intracellular oxidation of DHR was reduced by approximately 40% when cells were also exposed to the NO synthase inhibitor L-NAME. Oxidation of DHR following hypoxia and reoxygenation was not affected by SOD or DMTU. A combination of SOD and L-NAME was no more effective than L-NAME alone. Hypoxia and reoxygenation produced substantial injury (as LDH release). There was a 40% reduction in LDH release when cells were pretreated with a NO synthase inhibitor. In summary, increased generation of NO capable of inducing intracellular oxidizing reactions and cell death occurred during renal hypoxia and reoxygenation.
Paller
M S
MS
Department of Medicine, University of Minnesota, Minneapolis 55455, USA. palle001@maroon.tc.umn.edu
Weber
K
K
Patten
M
M
D058186:Acute Kidney Injury / Q000378:metabolism; D000818:Animals; D015687:Cell Hypoxia / Q000502:physiology; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D007687:Kidney Tubules, Proximal / Q000378:metabolism / Q000473:pathology*; D008297:Male; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D018384:Oxidative Stress / Q000502:physiology; D010101:Oxygen Consumption / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013997:Time Factors
D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D019331:NG-Nitroarginine Methyl Ester