Inhibiting Cholesterol Synthesis Reduces the Binding and Toxicity of Amphotericin B against Rabbit Renal Tubular Cells in Primary Culture

Renal tubular cells are a target of amphotericin B (AmB) toxicity, but the mechanisms involved in the tubular cell-AmB interactions are unknown. Ketoconazole was selected to lower the cholesterol content of rabbit renal tubular cells in primary culture. The consequences of cholesterol depletion on AmB nephrotoxicity was investigated in vitro as the inhibition of Na^dependent phosphate uptake. After 1 h of exposure, AmB decreased phosphate uptake (490Zo, 770Zo, and 8207o inhibition with 5, 10, and 20 pMof AmB, respectively). Pretreatment of cells with ketoconazole (10 pM for 24 h) reduced by 5007o (P < .01) the phosphate uptake inhibition induced by AmB, decreased cellular cholesterol synthesis ^800Zo inhibition), and decreased AmB binding to cell membrane by 500Zo, as measured by the fluorescence extinction of a probe bound to tubular cell membrane. Incubation with exogenous exchangeable cholesterol again increased AmB binding to plasma membrane and restored AmB toxicity. These results demonstrate that the first step of AmB renal tubular toxicity is mediated by cellular cholesterol content and is parallel to the binding of AmB to cell plasma membrane.