Inducible nitric oxide synthase and apoptosis in murine proximal tubule epithelial cells.
Since inducible nitric oxide synthase (iNOS) and proximal tubule injury are known to be critical determinants of lipopolysaccharide (LPS)-induced renal failure, the role of nitric oxide (NO) in proximal tubule cell apoptosis was examined. An 18-h treatment with a combination of LPS (5 microg/ml) and interferon-gamma (IFN-gamma, 100 units/ml) synergistically induced iNOS and produced a 20-fold increase in NO generation in the TKPTS murine proximal tubule cell line. NO generation by LPS + IFN-gamma was blocked by a specific iNOS blocker, L-N6-(1-iminoethyl)-lysine (L-NIL, 1 mM). To assess the role of iNOS-derived NO in proximal tubule cell apoptosis, annexin V- and propidium iodide-labeled cells were analyzed by flow cytometry. Neither the induction of iNOS nor its inhibition produced significant apoptotic cell death in TKPTS cells. Two exogenous NO donors were used to examine the role of NO more directly in proximal tubule apoptosis. Although both sodium nitroprusside (SNP), an iron-containing, nitrosonium cation donor, and S-nitroso-N-acetylpenicillamine (SNAP), a noniron-containing, NO generator, produced a concentration-dependent increase in NO generation, only SNP increased apoptotic cell death in TKPTS cells (5.9 +/- 0.7% in control cells vs. 21.6 +/- 3.8% in SNP [500 microM]-treated cells; n = 4-9; p < 0.01). SNP-mediated tubule cell apoptosis was not dependent on the activation of caspases or p53 but was possibly related to the generation of reactive oxygen species by SNP. Thus, in TKPTS cells induction of iNOS and generation of NO by LPS does not lead to tubular epithelial cell death.
Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Manish M
MM
Tiwari
Kurt J
KJ
Messer
Philip R
PR
Mayeux
D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology*; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D051379:Mice; D009569:Nitric Oxide / Q000096:biosynthesis*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D052247:Nitric Oxide Synthase Type II / Q000096:biosynthesis; D009573:Nitrites / Q000378:metabolism; D009599:Nitroprusside / Q000494:pharmacology; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism
D008070:Lipopolysaccharides; D020030:Nitric Oxide Donors; D009573:Nitrites; D017382:Reactive Oxygen Species; C110881:S-nitro-N-acetylpenicillamine; D009599:Nitroprusside; D009569:Nitric Oxide; D007371:Interferon-gamma; D052247:Nitric Oxide Synthase Type II; D010396:Penicillamine