title: Partial attenuation of cytotoxicity and apoptosis by SOD1 in ischemic renal epithelial cells.
abstract: Reactive oxygen species (ROS) contribute significantly to apoptosis in renal ischemia-reperfusion (IR) injury, however the exact mechanisms are not well understood. We used novel lentiviral vectors to over-express superoxide dismutase 1 (SOD1) in proximal tubular epithelial (LLC-PK(1)) cells and determined effects of SOD1 following ATP depletion-recovery, used as a model to simulate renal IR. SOD1 over-expression partially protected against cytotoxicity (P < 0.001) and decreased superoxide (O(2) (*-)) in ATP depleted cells. The ATP depletion-mediated increase in nuclear fragmentation, an index of apoptosis and activation of caspase-3 was also partially blocked by SOD1 (P < 0.05). However, SOD1 over-expression was insufficient to completely attenuate caspase-3, indicating that ROS other than cytoplasmic O(2) (*-) are involved in ATP depletion mediated injury. To test the contribution of hydrogen peroxide, a subset of enhanced green fluorescent protein (EGFP) and SOD1 (serum free and injured) cells were treated with polyethylene glycol-catalase (PEG-catalase). As expected there was 50% reduction in cytotoxicity and caspase-3 in SOD1 cells compared to EGFP cells; catalase treatment decreased both indices by an additional 28% following ATP depletion. To test the role of mitochondrial derived superoxide, we also treated a subset of LLC-PK(1) cells with the mitochondrial antioxidant, MitoTEMPO. Treatment with MitoTEMPO also decreased ATP depletion induced cytotoxicity in LLC-PK(1) cells in a dose dependant manner. These studies indicate that both SOD1 dependent and independent pathways are integral in protection against ATP depletion-recovery mediated cytotoxicity and apoptosis, however more studies are needed to delineate the signaling mechanisms involved.
authors.affiliation: Division of Transplant Surgery, Medical College of Wisconsin, Milwaukee, WI 53226, USA.
authors.forename: Huan Ling
authors.initials: HL
authors.lastname: Liang
authors.forename: Jody
authors.initials: J
authors.lastname: Arsenault
authors.forename: Jordan
authors.initials: J
authors.lastname: Mortensen
authors.forename: Frank
authors.initials: F
authors.lastname: Park
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Johnson
authors.forename: Vani
authors.initials: V
authors.lastname: Nilakantan
mesh_terms: D000255:Adenosine Triphosphate / Q000172:deficiency; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002374:Catalase / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology*; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007511:Ischemia / Q000201:enzymology*; D007687:Kidney Tubules, Proximal / Q000098:blood supply* / Q000166:cytology*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D018374:LLC-PK1 Cells; D016086:Lentivirus / Q000235:genetics; D010880:Piperidines / Q000494:pharmacology; D015203:Reproducibility of Results; D013482:Superoxide Dismutase / Q000378:metabolism*; D000072105:Superoxide Dismutase-1; D013481:Superoxides / Q000378:metabolism; D013552:Swine; D013997:Time Factors
chemical_list: C528764:2,2,6,6-tetramethyl-1-piperidine; D010880:Piperidines; C000606290:SOD1 protein, human; C485184:enhanced green fluorescent protein; D013481:Superoxides; D049452:Green Fluorescent Proteins; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D007770:L-Lactate Dehydrogenase; D002374:Catalase; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D053148:Caspase 3
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