Premium
A Murine Model of Acute Hindlimb Ischemia/Reperfusion Injury
Author(s) -
TU HUIYIN,
LI YULONG,
TRAN PAUL
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.763.3
Subject(s) - hindlimb , medicine , ischemia , superoxide , tourniquet , reperfusion injury , anesthesia , chemistry , biochemistry , enzyme
15% of preventable combat fatalities are due to hemorrhage from extremity wounds, making emergency tourniquet one of the first‐line treatments. While lifesaving, tourniquets can cause serious ischemia‐reperfusion (IR) injury. Fundamental to IR injury is mitochondrial production of superoxide. We refined a murine model of acute hind limb IR to simulate the clinical use of tourniquet. Methods Groups of C57/BL6 mice underwent an IR protocol of three hours of ischemia and four hours of reperfusion. IR was achieved by placement and release of a rubber tourniquet around the hind limb, at the greater trochanter level. Muscle blood flow (MBF) was measured using a Doppler probe. Muscle necrosis was measured using TTC stain. Superoxide production was measured using chemiluminescence and DHE staining. MnSOD protein expression and activity were measured using Western blot and a modified nitrite method. Mitochondrial respiratory chain function was measured via spectrophotometry. Results Significant ischemia and microcirculatory dysfunction were achieved: MBF during IR was < 10% and < 41% of baseline, respectively (n=8), accompanied by >40% necrosis (n=9). Superoxide production was increased (0.05 vs 0.13, p<.05 sham vs IR)accompanied by concomitant increases in MnSOD activity and protein expression (1.51 vs 2.26 , p<.05,sham vs IR, n=8). Mitochondrial complexes I‐IV showed signs of dysfunction (Complex I‐ IV: 0.13, 0.31, 1.37, 2.76 sham vs 0.08, 0.21, 0.7, 1.97 IR, p<.05 ). Conclusions We have demonstrated an animal model of clinical tourniquet use. IR produces significant mitochondriopathy which is associated with elevated superoxide production and antioxidant activity. The model can be used to mechanistically evaluate antiischemic molecules in efforts to minimize the morbidity and mortality of battlefield injury.