Figure 8. The angiogenic activity of DFO-pretreated EPCs is compromised in hind-limb ischemic animals. (A) Laser Doppler perfusion images of hind-limb ischemic rats injected with human EPCs to test their angiogenic activity. The rats were imaged on day 0 and day 21 after being injected with PBS, EPCs or EPCs treated with 10 μM DFO for four days. PBS without EPCs served as a mock injection, while untreated right hind limbs served as the control. (B) Quantification of the ischemic area versus the normal perfusion area. The days on which the images were captured are indicated. * P < 0.05 compared with the PBS group, # P < 0.05 compared with the DFO-treated EPC group. (C) Damaged tissues were rescued through the injection of young EPCs but not DFO-treated EPCs. The degree of tissue damage was classified into three categories: normal (n=10, empty box), toe necrosis (n=11, grid box) or amputation (n=11, filled box). The percentage of amputated limbs was 10% higher in the DFO-treated EPC group (EPC+DFO) than in the EPC-injected group. (D) Representative images of staining for myocytes and capillaries. On day 21, tissues were sectioned and stained with laminin (blue) and Bandeiraea simplicifolia lectin 1 (BS-I, red) to visualize myocytes and capillaries, respectively. Due to tissue atrophy, the peripheries of the PBS-treated myocytes appeared to be smaller than those of the EPC-injected myocytes. (E) Quantification of capillary density, which was calculated as the number of capillaries divided by the number of myocytes. * P < 0.05 compared with the PBS group, # P < 0.05 compared with the DFO-EPC group. Scale bar, 50 μm.