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¹³N-Ammonia PET as a Measurement of Hindlimb Perfusion in a Mouse Model of Peripheral Artery Occlusive Disease

¹ Department of Nuclear Medicine, Clínica Universitaria, University of Navarra, Pamplona, Spain; ² MicroPET Research Unit CIMA-CUN, University of Navarra, Pamplona, Spain; ³ Hematology and Cell Therapy Service, Clínica Universitaria, University of Navarra, Pamplona, Spain; and ⁴ Foundation for Applied Medical Research (FIMA), University of Navarra, Pamplona, Spain

View larger version (95K): [in this window] [in a new window]   FIGURE 1.  Animal placement in scanner for small-animal PET studies. (A) Lines represent iliac–femoral arteries showing where arteries were cut and excised (arrowheads, I model; arrows, F model). (B) Position of animal on cradle, illustrating position of legs of animals.

View larger version (6K): [in this window] [in a new window]   FIGURE 2.  Left-to-right ratio of hindlimb perfusion (mean ± SD) in nonsurgical control animals (n = 5) as determined for dynamic PET studies. Data were acquired in list mode; framing was applied as described in the text and exported to PMOD software for numeric calculations. t = time.

View larger version (16K): [in this window] [in a new window]   FIGURE 3.  Comparison of spontaneous perfusion recovery along time between F model (white bars, n = 5) and I model (hatched bars, n = 10). NS = nonsignificant differences; RFV = relative flow values of ischemic vs. control hindlimb. *P < 0.05; **P < 0.01.

View larger version (20K): [in this window] [in a new window]   FIGURE 4.  13N-Ammonia small-animal PET images of spontaneous perfusion recovery in a representative animal after ligation of left iliac artery. Image in A shows a photograph of the hindlimbs of the animal to facilitate interpretation. Images in B–E correspond to the same animal studied 1, 7, 15, and 30 d after surgery. For this animal, perfusion of left hindlimb accounts for 10.3% (B), 28.6% (C), 51.0% (D), and 56.0% (E) of perfusion of nonsurgical contralateral hindlimb. Note spontaneous recovery of perfusion in left limb.

View larger version (117K): [in this window] [in a new window]   FIGURE 5.  (A and B) Analysis of necrosis (defined by presence of fat cells and "ghost" muscle cells devoid of a nucleus) on H&E–stained cross-sections (A) and analysis of fibrosis on sirius red–stained cross-sections (B) of quadriceps muscles in right nonischemic limb and left ischemic limb at 7, 15, and 30 d after iliac artery ligation. (C and D) Analysis of vascular bed at capillary level (BS-I) (C) and arteriolar level (-actin) (D) of quadriceps muscles in right nonischemic limb and left ischemic limb at 7, 15, and 30 d after iliac artery ligation. Scale bars: 200 µm (A and B), 50 µm (C), and x200 (D).

View larger version (15K): [in this window] [in a new window]   FIGURE 6.  (A and B) Quantification of necrotic (A) and fibrotic (B) area expressed as percentage vs. total muscle area of quadriceps muscle. (C and D) Quantification of microvascular vessels stained with BS-I (C) expressed as number of vessels by mm2 of quadriceps muscle and arteriolar vessels (D) expressed as area positive for -actin staining vs. total muscle area of quadriceps muscle. For histologic analysis, only animals in which left iliac artery was excised were used. Results represent mean ± SD. *P < 0.05; **P < 0.01.