Role of mitochondrial oxidants in an in vitro model of sepsis-induced renal injury.
Oxidative stress has been implicated to play a major role in multiorgan dysfunction during sepsis. To study the mechanism of oxidant generation in acute kidney injury (AKI) during sepsis, we developed an in vitro model of sepsis using primary cultures of mouse cortical tubular epithelial cells exposed to serum (2.5-10%) collected from mice at 4 h after induction of sepsis by cecal ligation and puncture (CLP) or Sham (no sepsis). CLP serum produced a concentration-dependent increase in nitric oxide (NO) (nitrate + nitrite) release at 6 h and cytotoxicity (lactate dehydrogenase release) at 18 h compared with Sham serum treatment. Before cytotoxicity there was a decrease in mitochondrial membrane potential, which was followed by increased superoxide and peroxynitrite levels compared with Sham serum. The role of oxidants was evaluated by using the superoxide dismutase mimetic and peroxynitrite scavenger manganese(III)tetrakis(1-methyl-4-pyridyl)porphyrin tetratosylate hydroxide (MnTmPyP). MnTmPyP (10-100 μM) produced a concentration-dependent preservation of ATP and protection against cytotoxicity. MnTmPyP blocked mitochondrial superoxide and peroxynitrite generation produced by CLP serum but had no effect on NO levels. Although MnTmPyP did not block the initial CLP serum-induced fall in mitochondrial membrane potential, it allowed mitochondrial membrane potential to recover. Data from this in vitro model suggest a time-dependent generation of mitochondrial oxidants, mitochondrial dysfunction, and renal tubular epithelial cell injury and support the therapeutic potential of manganese porphyrin compounds in preventing sepsis-induced AKI.
Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.
Elina
E
Pathak
Lee Ann
LA
MacMillan-Crow
Philip R
PR
Mayeux
D058186:Acute Kidney Injury / Q000209:etiology* / Q000473:pathology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002432:Cecum / Q000293:injuries; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000187:drug effects; D016166:Free Radical Scavengers / Q000494:pharmacology; D066298:In Vitro Techniques; D007672:Kidney Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D008665:Metalloporphyrins / Q000494:pharmacology; D051379:Mice; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000502:physiology; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D016877:Oxidants / Q000009:adverse effects*; D018805:Sepsis / Q000150:complications* / Q000473:pathology; D013482:Superoxide Dismutase / Q000378:metabolism
D016166:Free Radical Scavengers; D008665:Metalloporphyrins; C502230:Mn(III) 5,10,15,20-tetrakis(N-methylpyridinium-2-yl)porphyrin; D009566:Nitrates; D009573:Nitrites; D016877:Oxidants; D009569:Nitric Oxide; D000255:Adenosine Triphosphate; D013482:Superoxide Dismutase