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Cellular effects of reactive intermediates: nephrotoxicity of S-conjugates of amino acids
Abstract:
Several cysteine S-conjugates are potent nephrotoxins and require enzymatic activation to produce cytotoxicity. Strategies based on the knowledge that renal cysteine conjugate beta-lyase is apparently a pyridoxal phosphate (PLP)-dependent enzyme have been exploited to test the hypothesis that a beta-lyase-dependent activation is required for the expression of cysteine S-conjugate-induced toxicity. First, the toxicity of the model conjugate S-(1,2-dichlorovinyl)-L-cysteine (DCVC) is blocked both in vivo and in isolated, renal proximal tubular cells by aminooxyacetic acid, an inhibitor of PLP-dependent enzymes. Second, the nonmetabolizable alpha-methyl analogue S-(1,2-dichlorovinyl)-DL-alpha-methylcysteine is not toxic. Third, to test the hypothesis that the toxicity of DCVC is associated with the metabolic formation of a reactive thiol, S-(1,2-dichlorovinyl)-L-homocysteine (DCVHC), which may undergo a PLP-dependent gamma-elimination reaction to produce an identical thiol, was studied. DCVHC is a potent nephrotoxin, and, similar to DCVC, its toxicity was blocked by aminooxyacetic acid and the alpha-methyl analogue S-(1,2-dichlorovinyl)-DL-alpha-methylhomocysteine was not toxic. Moreover, exposure of renal proximal tubular cells to propargylglycine, a suicide substrate for PLP-dependent enzymes that catalyze gamma-elimination reactions, blocked the toxicity of DCVHC. Fourth, the renal mitochondrial beta-lyase is localized in the outer membrane; therefore, although DCVC was toxic to mitochondria, no toxicity was produced in mitoplasts, which shows that a suborganelle site of activation is involved in the mitochondrial toxicity of DCVC. Finally, the toxicity of both DCVC and DCVHC was blocked by probenecid, indicating a role for the anion transport system. DCVC and DCVHC inhibit cellular and mitochondrial respiration, indicating that mitochondria are primary intracellular targets for nephrotoxic S-conjugates.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Nephrotoxic cysteine S-conjugates require enzymatic activation by pyridoxal phosphate (PLP)-dependent enzymes. Blocking these enzymes prevents toxicity, indicating PLP-dependent beta-lyase activation is crucial for S-conjugate-induced kidney damage.
Area of Science:
- Biochemistry
- Toxicology
- Nephrology
Background:
- Cysteine S-conjugates are potent nephrotoxins requiring enzymatic activation for cytotoxicity.
- Renal cysteine conjugate beta-lyase, a pyridoxal phosphate (PLP)-dependent enzyme, is implicated in this activation.
- Understanding this activation pathway is key to developing strategies against S-conjugate-induced nephrotoxicity.
Purpose of the Study:
- To test the hypothesis that beta-lyase-dependent activation is required for cysteine S-conjugate toxicity.
- To investigate the role of PLP-dependent enzymes in the nephrotoxicity of S-conjugates like DCVC and DCVHC.
- To identify the intracellular site of activation and the role of transport systems in S-conjugate toxicity.
Main Methods:
- Inhibition studies using aminooxyacetic acid (a PLP-dependent enzyme inhibitor) and propargylglycine (a gamma-elimination suicide substrate).
- Assessment of toxicity using nonmetabolizable alpha-methyl analogues of S-conjugates.
- Comparative toxicity studies in renal proximal tubular cells, mitochondria, and mitoplasts.
- Evaluation of probenecid's effect on S-conjugate toxicity.
Main Results:
- Toxicity of S-(1,2-dichlorovinyl)-L-cysteine (DCVC) and S-(1,2-dichlorovinyl)-L-homocysteine (DCVHC) was blocked by aminooxyacetic acid.
- Nonmetabolizable alpha-methyl analogues of DCVC and DCVHC were not toxic.
- Propargylglycine blocked DCVHC toxicity, and DCVC toxicity was localized to the mitochondrial outer membrane.
- Probenecid blocked the toxicity of both DCVC and DCVHC, and both inhibited cellular and mitochondrial respiration.
Conclusions:
- Beta-lyase-dependent activation is essential for the nephrotoxicity of cysteine S-conjugates.
- Mitochondria are primary intracellular targets for nephrotoxic S-conjugates, with activation occurring at the outer membrane.
- Anion transport systems play a role in the uptake or action of these nephrotoxins.