Nephrotoxicity endocytosis by of Bence-Jones proteins: Correlation with BHK cells and intracellular movement

The aim of this investigation was to evaluate the endocytosis of two Bence- Jones proteins by renal cells in order to elucidate the interference of the ir physical and chemical characteristics on nephrotoxicity. Bence-Jones pro teins (AK and GL) were purified and isolated from the urine of two patients with multiple myeloma. The isotype of both proteins was characterised as b eing human monoclonal lambda light chain. The AK protein presented mainly a n Ip>7.0, a high content of galactose and a low amount of sialic acid molec ules. On the other hand, the GL protein presented a single band with an Ip of 4.3, a higher level of sialic acid and a reduced amount of galactose, in comparison with the AK protein. Baby Hamster Kidney (BHK) cells were maint ained in culture in bottles at 37 degreesC, using DMEM culture media supple mented with 10% of calf serum with a pH of 7.4. Once the monolayer was obse rved to be confluent, the BHK cells were incubated with the two proteins, d issolved in a serum-free medium for 1, 5, 15, 30, 60 minutes and 24 hours. Control cells were established omitting the incubation with Bence-Jones pro teins, but maintaining all of the other conditions. After, this the cells w ere washed, trypsinised, centrifuged and fixed in a solution of 4% paraform aldehyde and 0.5% glutaraldehyde on a 0.1 M, pH 7.4 phosphate buffer. Cells were processed for immunocytochemical reactions by using protein A coupled with colloidal gold and further silver enhancement. Semi-thin sections of the pellets were obtained and submitted to the cytochemical reactions. Dete ction of labelling was made by using light microscopy. It was observed that GL protein tended to be directed towards a perinuclear position, whereas t he AK protein tended to suffer lysosomal deviation, suggesting that there i s a direct contribution of physical and chemical characteristics on intrace llular direction taken by Bence-Jones proteins.