Inhibition of AQP-1 trafficking and sorting by MG132. (A) Freshly obtained mouse reticulocytes (0 hour; left lane) were cultured for 24 hours with the proteasome inhibitor MG132 (20 μM; 24 hours; right lane). Freshly obtained mouse reticulocytes were also cultured with an equivalent volume of the solvent, DMSO (24 hours; middle lane). (B) Exosomes were isolated from the culture medium after 24 hours and resuspended in the same volume of Laemmli buffer.²³  Cellular and exosomal proteins were separated by SDS-PAGE and immunoblotted for the proteins indicated on the right. In each lane, 10⁶ cells were loaded. (C) After inhibition of endocytosis, the cells were allowed to mature in the presence or absence of MG132 for 24 hours. Exosomes were then collected, and the compositions of both cells and the vesicles released from them were analyzed by SDS-PAGE and immunoblotted for the same proteins.