Chronic exposure to cadmium results in proteinuria. To gain insights into the mechanism by which cadmium inhibits the protein transport in the renal proximal tubule, we investigated the effects of cadmium on the receptor-mediated endocytosis of albumin, using fluorescein isothiocyanate-labeled bovine serum albumin (FITC-albumin) as a model substrate and opossum kidney cell line (OK cell) as a proximal tubular cell model. Cell monolayers grown to confluence were treated with 100 microM CdCl(2) for 60 min at 37 degrees C, washed, and tested for FITC-albumin uptake (37 degrees C) and surface binding (4 degrees C). The amounts of FITC-albumin uptake and binding were quantified by fluorimetrically determining the cell-adherent fluorescence. Both the binding and uptake of FITC-albumin by OK cells appeared to be saturable and inhibitable by unlabeled albumin in the medium, indicating that specific receptor sites were involved. The uptake of FITC-albumin was inhibited by agents that interfere with the formation of endocytotic vesicle (hypertonic mannitol), endosomal acidification (NH(4)Cl), and vesicular trafficking (cytochalasin D and nocodazole), confirming that the uptake occurred via the process of receptor-mediated endocytosis. In cells treated with cadmium, the specific FITC-albumin uptake was significantly attenuated, and this was due to a reduction in V(max) and a rise in K(m). These changes in kinetic parameters were similar to those induced by NH(4)Cl. The binding of FITC-albumin to the apical surface of OK cells was inhibited by cadmium treatment, and this was attributed to a reduction in B(max). The values of K(d) and its pH dependency were not altered by cadmium treatment. The formation of endocytotic vesicles, as judged by fluid phase endocytosis of FITC-inulin, was not changed by cadmium treatment. These results indicate that the receptor-mediated endocytosis of albumin is impaired in cadmium-treated OK cells most likely due to a defect in endosomal acidification and the attendant fall in ligand-receptor dissociation, which impairs receptor recycling and the overall efficiency of endocytosis.
Copyright 1999 Academic Press.