Mode of cytotoxic action of nephrotoxic agents: oxidative stress and glutathione-dependent enzyme.
OBJECTIVE
To investigate the cytotoxic action of nephrotoxic agents using an in vitro renal cell model, focusing on the cellular oxidative status and a specific glutathione (GSH)-dependent enzyme, glyoxalase I (Gly-I).


METHODS
Renal proximal tubular LLC-PK(1) cells were exposed to mercuric chloride, glycerol, cisplatin, gentamicin and cyclosporin A, and cell number/viability were determined. Oxidative stress was assessed by lipid peroxidation (LPO) assay, and Gly-I activity was measured by enzymatic method on a spectrophotometer.


RESULTS
Both mercuric chloride (30 microm) and glycerol (2.5%) were highly toxic to LLC-PK(1) cells, inducing >90% cell death within 24 h. The remaining agents led to slightly >50% growth inhibition at 72 h. The LPO levels at 3 h in cells exposed to mercuric chloride or glycerol were approximately 2.5 times higher than that in controls. N-acetylcysteine (NAC), a potent antioxidant and precursor for GSH, almost completely (>95%) prevented renal cell death from mercuric chloride or glycerol. Gly-I activity was dependent on NAC and closely associated with cell viability. A approximately 65% loss in Gly-I activity by mercuric chloride/glycerol led to >90% cell death, while restoring a basal activity of Gly-I with NAC was accompanied by complete cell viability.


CONCLUSIONS
The cytotoxic action of nephrotoxic agents appears to be triggered by oxidative stress, leading to Gly-I inactivation. As Gly-I plays a key role in cellular detoxification, its inactivation under oxidative stress probably becomes fatal to cells. However, cytoprotection provided with NAC is significant and might have implications in preventing renal cell injury mediated through nephrotoxic agents.
Louie
Brandon
B
Department of Urology, New York Medical College, Valhalla, NY 10595, USA.
Rajamahanty
Srinivas
S
Pyo
Paul
P
Choudhury
Muhammad
M
Konno
Sensuke
S
D000970:Antineoplastic Agents / Q000009:adverse effects; D000975:Antioxidants / Q000494:pharmacology*; D002470:Cell Survival; D002478:Cells, Cultured; D005978:Glutathione / Q000378:metabolism*; D005990:Glycerol / Q000009:adverse effects; D006801:Humans; D007674:Kidney Diseases / Q000139:chemically induced* / Q000201:enzymology / Q000503:physiopathology; D007687:Kidney Tubules, Proximal / Q000201:enzymology* / Q000503:physiopathology; D007791:Lactoylglutathione Lyase / Q000378:metabolism*; D008627:Mercuric Chloride / Q000009:adverse effects; D018384:Oxidative Stress / Q000502:physiology*
D000970:Antineoplastic Agents; D000975:Antioxidants; D008627:Mercuric Chloride; D007791:Lactoylglutathione Lyase; D005978:Glutathione; D005990:Glycerol