In vitro response of rat and human kidney lysosomes to aminoglycosides

Abstract

To study aminoglycoside nephrotoxicity, renal cortical lysosomes were prepared from rat kidneys and from healthy portions of five human kidneys removed for tumor. The renal cortex was homogenized in 1 mM EDTA with 0.3M sucrose, and the lysosomes were separated by differential centrifugation. Lysosomes were incubated in isotonic sucrose solution with various drug concentrations for 1 hr at 37°. They were resedimented adn the N-acetyl- β-glucosaminidase (NAG) activity was measured in the supernatant fraction and in the disrupted pellet. Incucation with four aminoglycosides at therapeutic plasma concentrations lowered the percentage of NAG released into the supernatant fraction in a dose-related fashion. Incubation with the polyamines spermine and spermidine also produced this effect, with spermine and gentamicin being additive. This apparent lysosomal stabilization at clinically achieved plasma concentrations was also observed after substituting isotonic glycine for sucrose in the incubation mixture. High concentrations of aminoglycoside consistent with those accumulated in the renal cortex of patients and rats produced a dose-dependent release of lysosomal NAG with a rank order of potency paralleling their clinically observed potential for producing nephrotoxicity. Rats were treated with 20 mg/kg gentamicin twice a day for 28 days producing kidneys resistant to aminoglycoside nephrotoxicity. Lysosomes prepared from these animals compared to saline-treated controls showed decreased response to gentamicin at 2 and 4μg/ml and to apermine. Human renal cortical lysososomes also exhibited aminoglycoside- and polyamine-induced changes in NAG release. We conclude taht the lysosome is a site of action for aminoglycoside nephrotoxicity. We propose that aminoglycoside stabilization of this lysosomal membrane may lead to eventual disruption of the proximal tubular lysosomal system and cell injury.