The mechanism of pentachlorobutadienyl-glutathione nephrotoxicity studied with isolated rat renal epithelial cells

Abstract Isolated renal epithelial cells were used to study the mechanism of toxicity of pentachlorobutadienyl-glutathione (PCBG), a nephrotoxic glutathione conjugate of hexachlorobutadiene. The cytotoxicity of PCBG displayed a very steep dose-response relationship; at 10 μ m PCBG no toxicity was observed whereas 25, 50, and 100 μM PCBG all resulted in a similar degree of toxicity. In all cases, loss of cell viability was observed only after a 30-min lag period and reached a plateau of 50 to 60% nonviable cells between 90 and 100 min. Toxic doses of PCBG also resulted in the depletion of cellular thiols. Blocking PCBG metabolism by inhibition of γ-glutamyl transpeptidase [1-γ- l -glutamyl-2-(2-carboxyphenyl)hydrazine (anthglutin), 2 m m ]or renal cysteine conjugate β-lyase (aminooxyacetic acid, 0.5 m m ) resulted in complete protection against PCBG-induced cell damage. Exposure of isolated renal epithelial cells to 100 μ m PCBG resulted in the rapid formation of plasma membrane blebs which appeared to be associated with a loss of Ca 2+ from the mitochondrial compartment and an elevation of cytosolic Ca 2+ concentration as measured by Quin-2. PCBG treatment also resulted in the inhibition of cell respiration and a marked depletion of cellular ATP content, indicating additional mitochondrial effects of the toxin. Our results support a role for renal cysteine conjugate β-lyase in the metabolic activation of PCBG and suggest that PCBG-induced renal cell injury may be the result of selective effects on mitochondrial function.