A novel model of murine chronic limb ischemia using partial femoral artery ligation

OBJECTIVES: Current hind limb peripheral artery disease (PAD) animal models, which claim to mimic chronic limb ischemia, are examples of acute limb ischemia because of total ligation of the femoral artery. In this study, we have created a novel animal model using partial femoral artery occlusion to mimic chronic limb ischemia with decreased blood flow but not total occlusion. We hypothesize that this model will help us better understand chronic PAD with decreased, but not completely obstructed, blood perfusion and facilitate future investigation of therapeutic strategies for chronic obstructive vascular disease. MATERIALS AND METHODS: Male C57BL/6J mice, 10–12 weeks old, were anesthetized using intraperitoneal injections of ketamine (150 mg/kg)/xylazine (10 mg/kg). In the partial hind limb ischemia model (PHLI) ( n = 16), the lumen of the Femoral artery was narrowed by applying a 10-0 nylon suture through the mid-diameter and ligating half of the lumen. In the complete hind limb ischemia model ( n = 16), the femoral artery was completely tied off. Daily laser Doppler blood flow measurements were employed to follow changes in limb perfusion up to 25 days postexperiments. The perfusion difference between the ischemic and the control limbs was measured. Six mice from each group were randomly euthanized on day 7, and their calf muscles underwent microscopic evaluations by a board-certified histopathologist blinded to the specimen groups. The amount of cell necrosis, inflammatory cell infiltration, and interstitial fibroblast proliferation in each specimen was averaged and scored from 0 to 5 based on the severity of the histological changes. RESULTS: Following surgery, the mice in both the PHLI and CHLI groups presented with persistent limping, and the involved hind limbs were found to be pale and cool to the touch. Comparison of the laser Doppler perfusion between the experimental and control hind limbs with both PHLI and CHLI showed that there was a significant reduction in blood flow in the experimental hind limb following the procedures. During the 1 st week of the experiment, mice in the PHLI group had significantly higher blood flow to their experimental hind limbs compared with the CHLI group (day 3, P < 0.05; day 7, P < 0.05). This difference gradually disappeared, and after 15 days, there was no difference in the hind limb perfusion of the mice between PHLI and CHLI. The histological evaluation of the experimental hind limbs in both groups showed evidence of ischemia-related coagulative necrosis of the gastrocnemius samples but the difference in histological scoring did not reach statistical significance. CONCLUSION: A partial hind limb ischemia mouse model mimicking chronic limb ischemia with partially decreased blood flow was successfully established. This new model was successful in the creation of a low-flow state during the experiments in the partial ischemia group. The partial ischemia model presented here more closely mimics human PAD with a reduced perfusion instead of acute total occlusion.