(13)N-ammonia PET as a measurement of hindlimb perfusion in a mouse model of peripheral artery occlusive disease.
Peripheral arterial occlusive disease (PAOD) is a leading cause of mortality and morbidity in the western world. The development of noninvasive methods for assessment and comparison of the efficacy of novel therapies in animal models is of great importance.


METHODS
Hindlimb ischemia was induced in nude mice by ligation and excision of the left femoral artery (n = 5) or the left iliac artery (n = 10). Assessment of limb perfusion was performed by small-animal PET analysis after intravenous injection of (13)N-ammonia between 24 h and 30 d after surgery using the ratio of perfusion between the left limb (ischemic) and the right limb (control). Activity concentration per area unit was calculated in regions of interest placed on 1-mm-thick images for numeric calculations, and the iliac and the femoral models were compared. In addition, histopathologic studies were performed to assess the degree of necrosis (hematoxylin-eosin) and fibrosis (sirius red). Immunohistochemistry analyses for identification of arterioles (alpha-smooth muscle actin) and endothelium-capillaries-(Bandeiraea simplicifolia I [BS-I] lectin) were also performed.


RESULTS
Perfusion in both hindlimbs of control animals was similar (median of the left-to-right ratio = 0.99). Twenty-four hours after ischemia, perfusion of the ischemic limb (% mean +/- SD) was 33.3 +/- 10.6 and 22.1 +/- 9.9 in the femoral and iliac models, respectively. Spontaneous recovery of perfusion in the hindlimb that underwent surgery was significantly lower in the iliac model at day +15 (73.2 +/- 15.5 vs. 51.9 +/- 11.3; P < 0.01). Fibrosis increased progressively until day +30, whereas muscle necrosis was maximal at day +7 with a moderate reduction by day +30. In accordance with this positive effect, there was a statistically significant increase in the area covered with smooth muscle-coated vessels (arterioles) at day +30 in comparison with day 7 (P < 0.05). In addition, a correlation between (13)N-ammonia uptake and the amount of necrosis (r = -0.73; P = 0.06) and fibrosis (r = -0.67; P = 0.05) at day +30 was found.


CONCLUSIONS
(13)N-Ammonia imaging allows semiquantitative evaluation of hindlimb perfusion in surgical mouse models of acute hindlimb ischemia. Although spontaneous perfusion recovery is observed in both models, the iliac model shows a substantially lower recovery and is hence better suited for assessment of new therapeutic strategies for acute hindlimb ischemic disease.
Peñuelas
Iván
I
Department of Nuclear Medicine, Clínica Universitaria, University of Navarra, Pamplona, Spain. ipenuelas@unav.es
Aranguren
Xabier L
XL
Abizanda
Gloria
G
Martí-Climent
Josep María
JM
Uriz
Maialen
M
Ecay
Margarita
M
Collantes
Maria
M
Quincoces
Gemma
G
Richter
José A
JA
Prósper
Felipe
F
D000641:Ammonia; D000818:Animals; D001157:Arterial Occlusive Diseases / Q000000981:diagnostic imaging* / Q000473:pathology; D005355:Fibrosis; D006614:Hindlimb / Q000098:blood supply*; D007511:Ischemia / Q000000981:diagnostic imaging / Q000473:pathology; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009336:Necrosis; D009590:Nitrogen Radioisotopes; D016491:Peripheral Vascular Diseases / Q000000981:diagnostic imaging* / Q000473:pathology; D049268:Positron-Emission Tomography / Q000379:methods; D012039:Regional Blood Flow
D009590:Nitrogen Radioisotopes; D000641:Ammonia