Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury

Summary

Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl₃) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions.

Abstract

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, we have translated a vascular injury model from a small animal into a large animal (canine), with slight modifications that can be used for pre-clinical screening of prophylactic and thrombolytic agents. In addition to the surgical methods, the modified protocol describes the step-by-step methods to assess carotid artery canalization by angiography, detailed instructions to process both the brain and carotid artery for histological analysis to verify carotid canalization and cerebral hemorrhage, and specific parameters to complete an assessment of downstream thromboembolic events by utilizing magnetic resonance imaging (MRI). In addition, specific procedural changes from the previously well-established small animal model necessary to translate into a large animal (canine) vascular injury are discussed.

Introduction

Stroke therapy is largely modeled after coronary artery disease treatment, mainly because interventions in cardiovascular disease have responded well to drug therapy and endovascular interventions¹. These treatments, however, have not successfully translated to cerebral infarction. The difficulties with the current stroke treatment are that the recombinant tissue plasminogen activator (rTPA) cannot be reversed, and that administration carries a significant 6.4% risk of hemorrhagic conversion^2,3,4. The resulting morbidity and mortality limits its use to....

Protocol

The investigations described conform to the Guidelines for the Care and Use of Laboratory Animals of the National Institutes of Health and were approved by The Ohio State University Institutional Animal Care and Use Committee (#2015A00000029). All surgical manipulations were performed under deep anesthesia and the animals did not experience pain at any stage during the procedure. All experiments described were non-recovery.

1. Preparation

1. Freshly prepare 50 mL of ferric chloride (FeCl₃) at 50% (w/v) diluted in deionized water.
2. Cut the umbilical tape into 5.5 cm pieces and soak in the freshly prepared 50% FeC....

Results

Following the detailed procedures herein will result in the development of a model that can be used for prophylactic or thrombolytic assessment of occlusive arterial interventions. Figure 1A shows baseline flow velocity and the resulting blood flow velocity before, during, and after treatment recorded by a commercial software. Data from this recording can be used to determine the percent of re-perfusion with carotid artery injury and treatment in this canine .......

Discussion

The FeCl₃ induced vascular injury model is widely used to study thrombosis in small animals and is easy to translate into a large animal, pre-clinical model with a multitude of advantages. Slight modifications to adapt the protocol into a canine allow the utilization of both magnetic resonance imaging to assess stroke and hemorrhage volumes after a pharmacological intervention and angiography to assess vessel canalization before, during, and after treatment. Other thrombotic large animal models have not studie.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
1/8” umbilical tape	Jorgensen Laboratories Inc.,	#J0025UA	for ferric chloride application
4% paraformaldehyde in PBS	Alfa Aesar	AAJ61899AP
10% neutral buffered formalin	Richard-Allan Scientific	5701
2% 2,3,5-triphenyltetrazolium chloride (TTC in PBS, pH 7.4)	Sigma Aldrich	T8877
ADP/Collagen cartridges	Siemens Diagnostics	B417021A
4.5 ml 3.2% sodium citrate blood vacutainer	Becton Dickinson	BD 369714
4.5 ml lithium heparin vacutainer	Becton Dickinson	BD 368056
EDTA K3 vacutainers	Becton Dickinson	BD455036
Doppler flow probe	Transonic Systems Inc	MA2.5PSL
Hematoxylin 560	Surgipath	3801570
Eosin	Surgipath	3801602
LabChart Software	ADInstruments Inc.
Prisma Fit 3 tesla (3T) magnet	Siemen's Diagnostics
Sodium heparin for injection (to coat blood gas syringe)	NovaPlus	402525D
HUG-U-VAC positioning system	DRE Veterinary	1320