From the Society for Vascular Surgery
The impact of ischemic intervals on
neuromuscular recovery in a porcine (Sus scrofa)
survival model of extremity vascular injury
Gabriel E. Burkhardt, MD,a,bShaun M. Gifford, MD,a,bBrandon Propper, MD,a,b
Jerry R. Spencer, RVT,aKen Williams, MSc,aLyell Jones, MD,aNathan Sumner, MD,a
Jerry Cowart, DVM,aand Todd E. Rasmussen, MD,a,cSan Antonio, Tex; and Bethesda, Md
Background: Despite advances in revascularization following extremity vascular injury, the relationship between time to
restoration of flow and functional limb salvage is unknown. The objectives of this study are to describe a large animal survival
model of hind limb ischemia/reperfusion and define neuromuscular recovery following increasing ischemic periods.
Methods: Sus scrofa swine (N ? 38; weight, 87 ? 6.2 kg) were randomized to iliac artery occlusion for 0 (Control), 1
(1HR), 3 (3HR), or 6 (6HR) hours, followed by vessel repair and 14 days of recovery. Additionally, one group
underwent iliac artery division with no restoration of flow (Ligation), and one group underwent iliac artery exposure only
without intervention (Sham). A composite physiologic measure of recovery (PMR) was generated to assess group
differences over 14 days of survival. PMR included limb function (Tarlov score) and electrophysiologic measures
(compound muscle action potential amplitude, sensory nerve action potential amplitude, and nerve conduction velocity).
Using the PMR and extrapolating the point at which recovery following ligation crosses the slope connecting recovery
after 3 and 6 hours of ischemia, an estimate of the ischemic threshold for the hind limb is made. These results were
correlated with peroneus muscle and peroneal nerve histology.
Results: Baseline physiologic characteristics were similar between groups. Neuromuscular recovery in groups with early
restoration of flow (Control, 1HR, 3HR) was similar and nearly complete (92%, 98%, and 88%, respectively; P > .45).
While recovery was diminished in both 6HR and Ligation, Ligation, rather than repair, exhibited greater recovery (68%
vs 53%; P < .05). These relationships correlated with the pathologic grade of degeneration, necrosis, and fibrosis (P <
.05). The PMR model predicts minimal and similar persistent loss of function in groups undergoing early surgical
restoration of flow (Control 8%, 1HR 1%, 3HR 12%; P > .45). In contrast, the Ligation group exhibited the greatest
degree of injury early in the reperfusion period, followed by more complete recovery and at a faster rate than 6HR.
Extrapolating from the PMR the point at which Ligation (68% recovery) crosses the slope connecting 3 hours (84%
recovery) and 6 hours (53% recovery) of ischemia estimates the ischemic threshold to be 4.7 hours. Restoration of flow
at ischemic intervals exceeding this are associated with less physiologic recovery than ligation.
Conclusion: In this model, surgical and therapeutic adjuncts to restore extremity perfusion early (1-3 hours) after
extremity vascular injury are most likely to provide outcomes benefit compared with delayed restoration of flow or
ligation. Furthermore, the ischemic threshold of the extremity after which neuromuscular recovery is significantly
diminished is less than 5 hours. Additional studies are necessary to determine the effect of other factors such as shock or
therapeutic measures on this ischemic threshold. (J Vasc Surg 2011;53:165-73.)
Clinical Relevance: Restoration of axial flow following extremity vascular injury is ideally addressed early and definitively.
However, in the setting of associated life-threatening and/or orthopedic injuries or prolonged evacuation, the impor-
tance of and decision to restore perfusion is guided by a paucity of data. This study provides new insight into the extent
of neuromuscular recovery that can be expected after progressive periods of extremity ischemia, to include ligation.
Acute limb ischemia following trauma is a common
cause of morbidity and mortality in trauma centers and
on the battlefield.1-3Rapid surgical intervention to re-
store tissue reperfusion is desirable but not always imme-
diately possible.4,5Early use of surgical adjuncts, such as
an attempt to limit ischemia/reperfusion injury.6-8While
clinical judgment of surgeons has successfully guided the
use of these adjuncts and the priority of revasculariza-
tion, a paucity of data exists to relate timing of restora-
tion of flow to eventual functional (ie, neuromuscular)
From the San Antonio Military Medicine Center Consortium, Wilford Hall
United States Air Force Medical Center, the 59th Clinical Research
Squadron, and Genesis Concepts & Consultants (under USAF Contract
No: FA7014-09-D-0008), Lackland Air Force Base;aThe University of
Texas HSC Department of Surgery;band The Uniformed Services Uni-
versity of the Health Sciences.c
Supported by funding from the Office of the Surgeon General of the United
States Air Force.
Competition of interest: none.
Presented at the 2010 Vascular Annual Meeting of the Society for Vascular
Surgery, June 10-13, 2010, Boston, Mass.
Reprint requests: Todd E. Rasmussen, MD, FACS, Lieutenant Colonel
USAF MC, San Antonio Military Vascular Surgery, Chief, Division of
Surgery/Flight CC, Wilford Hall, USAF Medical Center, 2200 Bergquist
Dr., Ste. 1, Lackland Air Force Base, Texas 78236 (e-mail: todd.
The editors and reviewers of this article have no relevant financial relationships
to disclose per the JVS policy that requires reviewers to decline review of any
manuscript for which they may have a competition of interest.
Published by Elsevier Inc. on behalf of the Society for Vascular Surgery.
Several small animal studies suggest that there is an
ischemic interval after which irreversible neuromuscular
injury occurs (ie, “ischemic threshold”), but this is incom-
pletely and variably defined.9-13Additionally, the impor-
tance of immediate (eg, vascular repair) compared with
delayed restoration of flow or even ligation and reperfusion
through native or collateral processes has been recognized
but incompletely characterized.14-17To date, models de-
scribing serial functional assessment after large, axial vessel
injury treated by repair or ligation are lacking.
In order to advance from the study of statistical to
quality or functional limb salvage, a translatable, large
animal survival model is necessary. Such a model would be
important to characterize the extent of anticipated neuro-
muscular recovery after incremental periods of ischemia
and provide insight into a threshold beyond which surgical
restoration of flow may not be beneficial. Such a model
would also allow eventual evaluation of factors (eg, shock),
which may move this threshold to a more or less favorable
position in the setting of extremity injury. The objective of
this study is to describe a large animal survival model of
hind limb ischemia and reperfusion. Additionally, the ob-
clinically relevant measures of neuromuscular recovery dur-
ing a 14-day reperfusion or outcomes period.
Experimental design. Institutional Animal Care and
Use Committee (IACUC) review and approval of this
protocol was obtained prior to initiation of the experiment.
All procedures were performed at an accredited animal
research facility (Lackland Air Force Base, Texas) in strict
and support of licensed veterinary staff. Female adolescent
(age 5-6 months) swine (Sus scrofa; Yorkshire/Landrace
Cross; John Albert Suppliers, San Antonio, Tex) were
arrived and housed at the research facility to allow acclima-
tion and observation for absence of disease 1 week prior to
initiation of the protocol. On the day of the study, induc-
tion of anesthesia was accomplished with intramuscular
ketamine and then maintained with inhaled isoflurane (2%-
4%). Animals were placed in the supine position on the
operating table and underwent sterile surgical scrub of the
abdomen and femoral regions. Retroperitoneal exposure of
the right iliac artery was accomplished using an established
model of limb previously described by this group (Fig 1).18
Occlusion of hind limb flow. To achieve hind limb
ischemia, the right external iliac artery of the animals was
exposed via a retroperitoneal (ie, “transplant”) incision and
dissection. A 5- to 6-cm segment of the right external iliac
artery was exposed, and two side branches (proximal and
distal) were ligated but not divided. The side branches
marked the proximal and distal extents of the arterial expo-
sure and were the lateral circumflex iliac and circumflex
femoral arteries, respectively. The proximal and distal ex-
tents of the iliac artery just beyond these side branches were
encircled with rubber vessel loops. The loops were pulled
taut to occlude flow to the extremity for the different
Fig 1. Experimental design. Central access via the right carotid artery and jugular vein for hemodynamic monitoring and
was created, and the vessel was occluded for 0 (Control), 1 (1HR), 3 (3HR), or 6 (6HR) hours before reconstruction via
patch angioplasty. Remaining groups underwent excision of a 3-cm segment of the external iliac artery (Ligation) as shown
in the postprocedure arteriogram or exposure of the external iliac artery without intervention (Sham).
JOURNAL OF VASCULAR SURGERY
166 Burkhardt et al
24. Katsanos K, Karnabatidis D, Diamantopoulos A, Kagadis GC, Rava-
zoula P, Nikiforidis GC, et al. Thrombin promotes arteriogenesis and
hemodynamic recovery in a rabbit hindlimb ischemia model. J Vasc
25. Waters RE, Terjung RL, Peters KG, Annex BH. Preclinical models of
human peripheral arterial occlusive disease: implications for investiga-
tion of therapeutic agents. J Appl Physiol 2004;97:773-80.
26. Buschmann IR, Voskuil M, van Royen N, Hoefer IE, Scheffler K,
Grundmann S, et al. Invasive and non-invasive evaluation of spontane-
ous arteriogenesis in a novel porcine model for peripheral arterial
obstructive disease. Atherosclerosis 2003;167:33-43.
27. Dick F, Li J, Giraud MN, Kalka C, Schmidli J, Tevaearai H. Basic
control of reperfusion effectively protects against reperfusion injury in a
realistic rodent model of acute limb ischemia. Circulation 2008;118:
Submitted Apr 23, 2010; accepted Jul 11, 2010.
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