Figure 2

Iron rich ferritin is internalized by Caco2 and human neuroblastoma cells. (A) Reaction of DFO and PK treated brain homogenate for iron shows significant depletion in the DFO+ sample (panels 1-4). (B) Caco-2 monolayers were exposed to PK treated DFO- and DFO+ CJDH in the AP chamber. Following an overnight incubation, precipitated proteins from the AP and BL medium were subjected to Western blotting. Probing for ferritin shows significantly more transport of ferritin to the BL chamber of -DFO sample relative to the +DFO sample (lanes 2 and 4). (C) Immunostaining of filter inserts for ZO-1 shows intact monolayer of Caco-2 cells with tight junctions (panels 1 and 2). Bar: 10 μm. (D) Human neuroblastoma cells (M17) were exposed to 5 μl of -DFO and +DFO NH sample re-suspended in PBS. Immunostaining of fixed, permeabilized cells with anti-ferritin antibody followed by FITC-conjugated secondary antibody shows prominent reaction for ferritin in intracellular structures consistent with endosomes (panels 1 and 2). Reaction for ferritin is stronger in cells exposed to -DFO sample compared to +DFO exposed cells (panels 1 and 2). Bar: 10 μm. Inset shows enlarged image of the boxed area.