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METHODS: To confirm the presence of an uptake mechanism, we performed overnight time lapse experiments with a low dose of FITC-BSA (10 µg/ml). To evaluate the uptake kinetic, we stimulated podocytes with BSA at different time (2, 4, 8 h) and doses (10 µg/ml, 100 µg/ml and 1 mg/ml) both at 37 and 4 C. Immunofluorescence was performed to evaluate the presence of ClC-5, Megalin, Cubilin, Dab2 and Amnionless in human podocytes cultivated in standard condition and stimulated with FITC-BSA (10 µg/ml) at different time points (2, 4, 8 and 24 h). Finally, we stimulated human podocytes with increasing concentrations of BSA (range 10 µg/ml-30 mg/ml), evaluating changes in gene expression of the components of this system by Real Time PCR at different time points (24, 48 and 72 h).

RESULTS: Albumin internalization in podocytes was detected starting from 2 to 15 h in 23±9% of cells observed. The uptake …