Heart failure (HF) is a major health and economic burden in developing countries and its prevalence is increasing [
This study was approved by the University of Paris René Descartes Ethics Committee and by the
Adult male White New Zealand rabbits weighing 3.8–4.4 kg were used in this study. During the procedure, animals were anaesthetised using 5 mg/kg ketamine-HCl and 20 mg/kg propofol via the ear vein and anticoagulated with heparin (300 IU/kg). An antiarrhythmic protocol was implemented, which included beta-blockers/amiodarone and lidocaine (1 mg/kg load followed by a 20
Three series of experiments were sequentially performed. Assessment of vascular access and coronary catheterization procedures: after having selected the right carotid artery as the vascular access entry site, 3 rabbits were used to test two devices (cathlons and fine needles); 2 animals were used for vessel catheterization and to select the optimal introducer sheath size. Four additional rabbits were then used for selecting the type and size of the guiding catheter for coronary catheterization and to determine the dilution of the contrast medium. Assessment of the coronary occlusion site: to determine the coronary artery branch most suitable for occlusion to induce a functionally relevant MI, 14 rabbits underwent release of free and droppable coils. They were divided into three groups according to the target territory: the left descending artery area ( Comparison of MI-inducing techniques: five different techniques of coronary occlusion were finally investigated: free coils (
For coils, a guiding catheter was positioned under fluoroscopic guidance at the coronary ostium and a 0.014-in. floppy wire was advanced into the coronary vessel. The wire was advanced into the dominant coronary artery branch, as determined by angiography (Figure
Hearts in which a myocardial infarct was macroscopically observed at autopsy underwent a histological analysis (Figures
Macroscopic views. Macroscopic views of a heart that underwent embolization of the middle segment of the right coronary artery; rear side view shows myocardial scarring of the right ventricle and the inferior wall of the left ventricle (a). The area of fibrosis is the inferior wall of the left ventricle and right ventricle (b). Heart that underwent embolization of the middle segment of the circumflex artery; apical section shows lateral wall transmural myocardial infarction (c).
Histological analysis of left ventricle lateral wall transmural infarction. Histological views of left ventricle lateral wall infarction focused on left ventricle anterior wall after mid segment circumflex artery alcohol injection (a) and of the interventricular septum focused on the basal segment after proximal segment right coronary artery alcohol injection (b) showing muscle and fibrosis. Heart was excised, cut, then embedded in OCT, flash frozen in liquid nitrogen, and stored at −80°C. Ten-micrometer cryosections were stained to confirm the presence of ischemic tissue.
Angiography of the coronary artery after mid segment alcohol injection. Selective angiography of the left main coronary artery, of the left anterior descending along the anterior interventricular sulcus reaching the apex of the heart, of a diagonal coronary artery, and of the circumflex coronary artery along the atrioventricular groove (a). Embolization of the right coronary artery ostium with free coil (b). Angiography of the right coronary artery after mid segment alcohol injection (c). The contrast product injected after alcohol injection stagnates in the necrotic area thus achieving an aspect of myography.
Data are given in percentages and means ± standard deviations. Comparisons between groups were made using the nonparametric ANOVA test for continuous variables. A
The safest micropuncture vascular access was achieved using needles and 0.018-in. wires. A 4-Fr micropuncture sheath was effective. It was advanced over the wire and exchanged for a 0.035-in. guidewire to support the placement of a 4-Fr internal mammary guide catheter (Cordis Corporation, Miami, FL, USA), with custom alteration for use in a rabbit. The guiding catheter was advanced retrogradely to the ascending aorta using fluoroscopy, and an angiography of the coronary artery was then performed to check for its adequate location. Dilution of the contrast medium was kept at 50% of the total volume to reduce the risk of ventricular arrhythmias.
Middle or proximal segment occlusion of the left anterior descending coronary artery led to severe heart failure with massive hemorrhagic infarction of the LV, while distal segment occlusion caused severe conduction disturbances requiring isoprenaline infusion. The occlusion of the branches (diagonal and septal branches) needed a custom device and led to only small nontransmural infarcted areas, without a relevant decrease in LVEF. Occlusion of obtuse marginal branches yielded a very limited, nontransmural LV infarction without functional consequences, as assessed by echocardiography. Only occlusion either of the mid segment of the right coronary artery or of the mid segment of the circumflex coronary artery was finally found to be the most reliable procedure for inducing an extensive infarction of the supplied LV area. Such infarcts led to an akinesia/hypokinesia of the right ventricle and of the inferior wall of the left ventricle (right coronary occlusion or circumflex coronary occlusion, resp.) with, in both cases, a 25% decrease in LVEF and only nonsustained ventricular arrhythmias. These data (summarized in Table
Distribution and follow-up depending on embolization site.
Embolization site |
Intraoperative events | 3-week survival |
Echographic assessment | Histologic assessment |
---|---|---|---|---|
LAD | ||||
Proximal segment ( |
VF | 0 | — | Transmural infarct |
Mid segment ( |
VF | 0 | — | Transmural infarct |
Distal segment ( |
ACD | 0 | — | Transmural infarct |
S. branch vessel ( |
— | 1 | No LV dysfunction | Nontransmural infarct |
CA | ||||
Proximal segment ( |
NSVT, CHF | 0 | — | Transmural infarct |
Mid segment ( |
NSVT | 2 | LV dysfunction | Transmural infarct |
Distal segment ( |
— | — | — | — |
S. branch vessel ( |
— | 2 | No LV dysfunction | Nontransmural infarct |
RCA | ||||
Proximal segment ( |
CHF | 0 | — | Transmural infarct |
Mid segment ( |
NSVT | 2 | LV and RV dysfunction | Transmural infarct |
Distal segment ( |
— | — | — | — |
LAD: left anterior descending; CA: circumflex artery; RCA: right coronary artery; LV: left ventricle; RV: right ventricle; VF: ventricular fibrillation; ACD: atrioventricular conduction disorder; NSVT: nonsustained ventricular tachycardia; CHF: congestive heart failure; S. branch vessel: secondary branch vessel.
Four rabbits underwent free coil embolization. Two coils deployed at the left main artery ostium and protruded into the aorta. One coil deployed at the circumflex artery proximal segment and one deployed at the right coronary artery ostium. All of these rabbits died from intraoperative refractory ventricular fibrillation. Histological examination revealed large left and right ventricle hemorrhagic necrosis, depending on the occlusion site (Tables
Distribution and follow-up depending on embolization device.
Embolization device |
Intraoperative events | 3-week survival |
Echographic assessment | Histologic assessment |
---|---|---|---|---|
Free coil | ||||
CA ( |
VF | 0 | — | Transmural infarct |
RCA ( |
VF | 0 | — | Transmural infarct |
Interlocking coil | ||||
CA ( |
VF | 1 | LVEF decrease <15% | Nontransmural infarct |
RCA ( |
VF | 1 | No dysfunction | No infarct |
Gelatin sponge | ||||
CA ( |
— | 3 | No dysfunction | Nontransmural infarct |
RCA ( |
— | 1 | No dysfunction | No infarct |
Balloon occlusion | ||||
CA ( |
NSVT | 2 | No dysfunction | No infarct |
RCA ( |
NSVT | 2 | No dysfunction | No infarct |
Alcoholization | ||||
CA ( |
VF | 4 | 15% decrease in LVEF | Transmural infarct |
RCA ( |
CHF | 2 | RV dysfunction |
Transmural infarct |
CA: circumflex artery; RCA: right coronary artery; LV: left ventricle; RV: right ventricle; VF: ventricular fibrillation; NSVT: nonsustained ventricular tachycardia; CHF: congestive heart failure.
Ultrasound assessment of alcoholization group.
Before embolisation | After embolisation |
|
Prob. > |
|
---|---|---|---|---|
DIVS (mm) | 3.8 ± 0.2 | 4.0 ± 0.2 | 2,354 | 0,155 |
DLVID (mm) | 17.4 ± 0.4 | 20.1 ± 1.1 | 29,236 | <0,001 |
DLVPW (mm) | 5.5 ± 0.2 | 5.5 ± 0.3 | 0,052 | 0,823 |
SIV (mm) | 4.1 ± 0.2 | 4.1 ± 0.2 | 0,220 | 0,648 |
SLVID (mm) | 12.1 ± 0.5 | 16.5 ± 0.9 | 104,824 | <0,001 |
SLVPW (mm) | 5.7 ± 0.2 | 5.7 ± 0.2 | 0,018 | 0,894 |
LVEF (Teichholz (%)) | 58.0 ± 2.9 | 43.5 ± 1.6 | 113,648 | <0,001 |
FS (%) | 30.5 ± 1.8 | 18.0 ± 0.9 | 240,384 | <0,001 |
LVESV (mL) | 2.9 ± 0.2 | 3.4 ± 0.1 | 19,736 | 0,001 |
LVEDV (mL) | 1.2 ± 0.2 | 1.8 ± 0.2 | 37,097 | <0,001 |
LVEF (biplane Simpson (%)) | 58.3 ± 2.6 | 47.0 ± 2.4 | 60,8421 | <0,001 |
SV (mL) | 1.7 ± 0.2 | 1.6 ± 0.2 | 1,680 | 0,223 |
DIVS: diastolic interventricular septum; DLVID: diastolic left ventricle inner diameter; DLVPW: diastolic left ventricle posterior wall; SIVS: systolic interventricular septum; SLVID: systolic left ventricle inner diameter; SLVPW: systolic left ventricle posterior wall; LVEF: left ventricle ejection fraction; FS: fractional shortening; LVESV: left ventricle end systolic volume; LVEDV: left ventricle end diastolic volume; SV: stroke volume.
Four rabbits underwent interlocking coil embolization in the mid segment of the target coronary artery. Two animals died from intraoperative refractory ventricular fibrillation during coil deployment in the circumflex and right coronary arteries, possibly due to the unwanted occlusion of the more proximal arterial segment due to excessively long coils. Two animals survived the procedure and until the end of the study. In both of these cases, the coils were too short, causing their migration towards the distality of the target vessel (the circumflex artery in one case and the right coronary artery in the other). Expectedly, this resulted in limited nontransmural or even undetectable infarct areas without changes in LVEF (Tables
Gelatin sponges were used in four procedures (three right coronary artery mid segment embolization and one circumflex mid segment embolization). No intraoperative or postoperative arrhythmias occurred and all animals survived until the end of the study. The likely reason was that only histologically limited nontransmural LV lateral wall infarcts were detectable after circumflex embolization while no myocardial damage could be identified in the case of right coronary artery embolization. No LVEF decrease could be echographically detected in any of these cases (Figure
Echographic assessment of left ventricular ejection fraction. Left ventricular measurements are made with the M-mode. Both ventricular diameters (systolic and diastolic) are measured from the leading edge to leading edge of each interface, and LVEF by Teichholz is then calculated (a). Ventricular volumes are calculated in the two orthogonal apical views: four-chamber (b) and (c)
Four animals underwent balloon occlusion, of the right coronary mid segment (two cases) and of the circumflex mid segment (two remaining cases). All animals survived despite the occurrence of nonsustained ventricular tachycardia which resolved after balloon deflation. Both echocardiographic and histological examinations failed to reveal any cardiac abnormality (Tables
Eight animals underwent alcohol injection in the mid segment of the right coronary artery and of the circumflex artery in five and three cases, respectively. All angiograms performed after alcohol injection showed coronary artery TIMI flow 0 or I and a pattern of myography in the infarcted area. One animal with circumflex artery alcoholization died from refractory ventricular fibrillation. Another rabbit which had undergone alcohol injection in the right coronary artery died two days after surgery from drug-refractory heart failure. However, the 6 remaining rabbits survived until the end of the study. Four of them were then found to have a macroscopically massive infarction of the right ventricle and of the LV inferior wall, which was confirmed by histological analysis. Two other rabbits had massive infarction of the left ventricle lateral wall. In all of these survivors, LVEF was significantly reduced from its baseline value (19,4 ± 2,4%; Tables
It is generally accepted that proof-of-concept studies designed to test the efficacy of new drug-, cell-, or other biologics-based therapies for HF can be reliably performed in rat and mouse [
Our review of the angiograms that we performed in our initial series of 14 rabbits did not yield results which matched those previously published [
Given this configuration, embolization of secondary branches of epicardial vessels such as marginal, septal, and diagonal branches only resulted in functionally inconsequent infarctions at the cost of complicated procedures while, contrariwise, embolization of the LAD was associated with much too extensive myocardial damage. We thus elected to focus on the dominant vessel; however the high sensitivity of the rabbit to ischemia required carefully controlling the level of its occlusion to ensure a substantial impairment of LV function, but with an acceptable mortality rate. Targeting a decrease in LVEF of 20% from baseline targeting was found to be a reasonable objective which could then be reproducibly achieved by locating the occlusion site at the mid segment of either the right coronary or the circumflex coronary artery.
These screening experiments then set the stage for the comparison of different infarction-inducing techniques which, to our knowledge, have not yet been reported. So far, the few studies which have used a closed-chest approach for inducing MI in rabbits have relied on the use of coils [
We acknowledge several limitations of the present study. Coils which were tested were those commercially available and we cannot exclude that customizing these devices would not have resulted in better outcomes. Second, functional assessments only consisted of gross measurements of EF and a more detailed analysis of the patterns of regional contraction and relaxation is clearly required to better characterize the hemodynamic effects of any coronary artery occlusion technique. Finally, the damage induced by alcohol injection may not accurately model that seen in patients with ischemic heart disease, although alcohol injection was found to depress LV function to a sufficient extent as to give room for detecting treatment effects for testing interventions. Despite these caveats, the present data can provide a useful benchmark for improving catheter-based techniques of coronary artery occlusion in rabbits and facilitate a broader use of this model for testing drugs or interventions designed to mitigate the consequences of ischemically induced LV dysfunction.
By following a stepwise approach, a minimally invasive, effective, and reproducible rabbit model of catheter-induced myocardial infarction has been developed which addresses the limitations of rodent experiments while avoiding the logistical and cost issues associated with large animal models.
The authors declare that there is no conflict of interests regarding the publication of this paper.