The combined effect of IDA and glutaraldehyde on the erythrocyte membrane proteins

A number of investigators have been focusing their attention on the encapsulation of antineoplastic drugs within erythrocytes to diminish their side-effects. Glutaraldehyde is often used as crosslinking agent to link the drugs (including idarubicin, IDA) to the cells. The previous studies indicated that in glutaraldehyde-treated human erythrocytes the elevated level of drugs was observed but also the various changes in the organization of the red cells were noted. In this study, we continue our investigations on the interaction of IDA and glutaraldehyde on the erythrocytes and now we concentrate on the effect of these compounds with the erythrocyte membrane proteins. For this purpose, SDS-gel electrophoresis of the cell proteins was carried out. Additionally, analysis of the disturbances of erythrocytes shape and size, accompanied by the application of flow cytometry and microscopy examination, were undertaken. The fluorimetric method was used to estimate content of IDA in supernatants, after erythrocyte membranes incubation with different glutaraldehyde concentrations. It was observed that glutaraldehyde caused in gradually dependent manner an increase of percent of IDA linked to the cell membrane proteins. After this incorporation, perturbations in the content of the proteins in the cell membrane were observed. The protein aggregates and changes in the level of spectrin, band 3 protein and small mass proteins were noted. The use of flow cytometry and microscopy technique demonstrated also disturbances in the shape and size of erythrocytes. For all tested concentrations of glutaraldehyde, the changes were statistically significant.