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Cardiopulmonary bypass model in the rat: A new minimal invasive model with a low flow volume

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Guillaume Lebreton at Hôpital La Pitié Salpêtrière (Groupe Hospitalier "La Pitié Salpêtrière - Charles Foix")
Guillaume Lebreton
Fabienne Tamion
Fabienne Tamion
Fabienne Tamion
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Jean Paul François Bessou at Centre Hospitalier Universitaire Rouen
Jean Paul François Bessou
Fabien Doguet
Fabien Doguet
Fabien Doguet
  • This person is not on ResearchGate, or hasn't claimed this research yet.
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Abstract and Figures

Numerous cardiopulmonary bypass (CPB) models in the rat have already been described, but these models often have an important mortality and differ a lot from human clinical conditions thus making them hardly usable. The CPB model in the rat we describe allows a femoro-femoral support CPB with a low priming volume, minimal surgical approach and excellent peroperative survival. This CPB model in the rat allows evaluating extracorporeal circulation effects.
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Cardiopulmonary bypass model in the rat: a new minimal invasive
model with a low flow volume
†
Guillaume Lebretona,b,*, Fabienne Tamionb, Jean-Paul Bessoub,c and Fabien Doguetb,c
a
Department of Thoracic and CardioVascular Surgery, University Hospital, CHU Pierre Zobda Quitmann, Martinique, France
b
INSERM U644, University Medical School, Rouen, France
c
Department of Thoracic and CardioVascular Surgery, University Hospital, Rouen, France
* Corresponding author. Tel: +596-5-96552271; e-mail: guillaumelebreton@live.fr (G. Lebreton).
Received 16 January 2011; received in revised form 2 August 2011; accepted 5 September 2011
Abstract
Numerous cardiopulmonary bypass (CPB) models in the rat have already been described, but these models often have an important
mortality and differ a lot from human clinical conditions thus making them hardly usable.
The CPB model in the rat we describe allows a femoro-femoral support CPB with a low priming volume, minimal surgical approach
and excellent peroperative survival. This CPB model in the rat allows evaluating extracorporeal circulation effects.
Keywords: Cardiopulmonary bypass •Experimental surgery •Animal model
INTRODUCTION
Numerous CPB models in the rat have already been described
[1–7] with various cannulation sites, circuits, priming volumes
(12–120 ml), products used for priming, temperature and out-
flows. Often, these models differ from human clinical conditions
thus making them hardly usable.
We here describe a model allowing a femoro-femoral support
CPB with a low priming volume, minimal surgical approach and
excellent peroperative survival.
TECHNIQUE
This CPB model was performed in Wistar male rats (450–500 g),
divided into two groups of 10 rats each (Sham group, femoral
cannulation without CPB and CPB group, femoral cannulation
with CPB). All rats were housed in individual cages and received
care in accordance with the ‘Guide for the Care and Use of
Laboratory Animals’(www.nap.edu/catalog/5140.html).
Extracorporeal circulation (Fig. 1)
We use a 16 G catheter (Introcan Safety BRAUN
®
) as a venous
cannula and a 22 G one as an arterial cannula. Arterial and
venous tubings as well as the pump are in PVC (internal diam-
eter: 2.5 mm. length = 20 cm). ‘Cardiotomy’reservoir is made of
a sterile 5 ml syringe (TERUMO
®
) connected to two consecutive
3-way taps, screwed to the pump. We use an occlusive roller
pump (GAMBRO®).
The oxygenator is a ‘Micro 1’(Kewei Medical Instrument Inc,
Shenzhen, China) with a 0.05 m
2
exchange surface which has
already been the subject of several publications [5–7] validating
its oxygenation capacities. The circuit is aseptically set-up and
‘freed of air bubbles’with Gelofusine
®
at 4% (BRAUN). Circuit
priming volume is 10 ml (Gelofusine
®
4%).
Operative protocol
After anaesthesia (intra-peritoneal injection of Chlorpromazine
Chlorhydrate (2 mg/kg) and Ketamine (80 mg/kg)), a heparinized
3F catheter is introduced into the left carotid. This approach is
used as an injection access and allows for a continuous pressure
monitoring. Heparin (500 UI/kg) and pancuronium bromide
(1 mg/kg) are injected through this access.
After orotracheal intubation (Arkansas no.18 cannula), con-
tinuous mechanical ventilator support is set-up at a 75 cycles/
min rate, a 10 ml/kg circulating volume and a 100% FiO
2
outside
CPB (21% during CPB). Further to intubation, a pancuronium
bromide injection is achieved through the left carotid.
The femoral vein is cannulated with a 16G catheter, and the
artery with a 22G catheter. The rat is placed upon a hotplate
with a 30° proclive positioning to ease venous drainage.
In the CPB group, cannulae are connected to the circuit after
a 2 ml Gelofusine® filling. CPB outflow is progressively increased
to 100 ml/kg/min. Vascular filling is achieved with Gelofusine®,
in compensation for diuresis, in order to maintain a MAP ≥70
mmHg and an adequate venous drainage for CPB. Rats not sub-
jected to CPB (sham group) were monitored for 150 min after
cannulation before being sacrificed.
†
Presented at the 60th International Congress of the European Society for
Cardiovascular and Endovascular Surgery, Moscow, Russia, 20–22 May 2011.
© The Author 2012. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery. All rights reserved.
Interactive CardioVascular and Thoracic Surgery 14 (2012) 642–644 BRIEF COMMUNICATION
doi:10.1093/icvts/ivr051 Advance Access publication 17 February 2012
During the entire experiment, tensional, rhythm, thermal and
diuresis monitoring were performed (Fig. 1). Diuresis was mea-
sured with a graded collector set against the urinary meatus.
Breathing and oxygenator parameters (O
2
outflow) were adjusted
according to gasometry so as to obtain a pH = 7.35, a PaO
2
=30
KPa and a PaCO
2
= 5 KPa. Arterial gasometry monitoring was
performed 20 min after femoral canulation and then after every
breathing parameter modification and at CPB discontinuation.
Blood samples (2 ml) were collected at cannulation (T0) and
at the time of sacrifice of the rats (T1 = T0 + 150 min), and imme-
diately centrifuged at 500 × g. Plasma samples were stored at
−80°C for measurement of serum tumour necrosis factor
(TNF)-αusing an ultrasensitive kit specific for rat TNF-α
(Cytoscreen; Biosource International, USA). The assay was per-
formed by measuring optical density at 450 nm. Each sample
was measured in duplicate and compared with a known concen-
tration range of TNF-α. The limit of detection was 0.7 pg/ml.
The rats were euthanized at the end of the study.
Data analysis
All results are presented with a mean ± standard deviation.
Results were compared with the Student’st-test. A P< 0.05 value
is considered as statistically significant. Statistical analysis was
performed with Statview
®
computer program.
RESULTS
None of the rats died during the procedure.
Figure 1: The cardiopulmonary bypass circuit and its monitoring. 1, blood
pressure and electrocardiogram; 2, temperature; 3, diuresis; 4, CPB flow
pump.
Figure 2: Blood pressure, heart rate and TNF-αvalues during the procedure.
EXPERIMENTAL
G. Lebreton et al. / Interactive CardioVascular and Thoracic Surgery 643
Mean arterial pressure analysis during the experiment (Fig. 2)
reveals a higher MAP (P< 0.0001) at post-cannulation 20th,
40th and 60th min in the CPB group compared with the sham
group. Observed heart rates do not significantly vary between
the two groups. Vascular filling and diuresis are significant
(P< 0.0001) in the CPB group.
TNF-αvalues (Fig. 2) at T1 are significantly higher in the CPB
group (P< 0.0001), and above those measured at T0 for both
groups in which they are very low.
DISCUSSION
Studying inflammation process requires reduced surgical aggres-
sion, which led us to use a femoral cannulation (closed chest).
Consequently, the observed inflammatory response is induced
by the one caused by blood contact with CPB circuit. Indeed, in
human clinical evaluation, the role played by the surgical trauma
in complement activation has been well established [8]. In our
model, it was mild.
Furthermore, our model offers the advantage of a low priming
volume not requiring transfusion and without raising the issue of
venous drainage. To avoid this problem, we used rats weighing
between 450 and 500 g (larger volemia). Besides, diuresis moni-
toring enables compensating it with the inflow. Lastly, placing
the rat in a 30° proclive position 15 cm above cardiotomy reser-
voir level improves venous drainage and a stable 100 ml/kg/min
CPB output is obtained.
Since it is a circulatory assistance without cardiac arrest,
we observe increased mean pressures during extracorporeal
circulation. MAP difference after CPB is probably due to
the filling variation between the control and the CPB groups,
as well as to the returned part of circuit volume at the end of CPB.
CONCLUSION
This CPB model in the rat allows evaluating extracorporeal circu-
lation effects. It has the advantages of CPB circuit low volume,
peripheral cannulation and reduced surgical aggression together
with low mortality.
Conflict of interest: none declared.
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G. Lebreton et al. / Interactive CardioVascular and Thoracic Surgery644
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... Choosing an appropriate anesthetic and administration method depends on the investigator's preference. The most common methods of administration are intraperitoneal injection and inhalation anesthesia [1,2,7,10,12,13,[17][18][19][20][21][22]24,25,27,29,31,38]. Ketamine, sodium pentobarbital, fentanyl, pancuronium, and isoflurane are generally used for anesthesia induction. ...
... The design and proper placement of the venous cannula result in adequate drainage flow. Several researchers have used modified multi-orifice 16-gauge angiocatheters as drainage cannulas [10,12,17,19,22,24,25,38]. However, the sharp end of the angiocatheter may damage the vessel or heart, making it dangerous to apply, especially for early-career researchers [13]. ...
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Article
  • Jan 2008
Aims: CPB is an essential component of cardiac surgery, with still unknown device/patient interactions. In order to evaluate the response of CPB to hemodynamic, biochemical, inflammatory, as well as thermo- pharmacodynamic interactions, a novel miniaturized oxygenator with controlled and standardized specifications has been developed together with an improved surgical central cannulation technique. Methods: A hollow-fibre small priming volume (6.3ml) oxygenator was manufactured according to specifications resulting from engineering, heart surgery and perfusionist expertise (Dideco-Sorin Group, Italy) with the following characteristics: Gas Exchange Surface-450cm2, Heat Exchange Surface-16cm2. The oxygenator was tested in vitro and in vivo in 5 anaesthetised, ventilated, open-chest rats using a miniaturized roller pump and heat exchanger. Pressures were monitored in the animal, before and after the oxygenator. Central venous cannulation through the right atrium, and aortic cannulation, through the carotid artery, were used. Results:In vitro: blood oxygenation increased 10-fold (from room air to 100% FIO2) and PCO2 removal was 2.5-fold. In vivo: CPB was performed without blood prime for 60mins (no ventilation) maintaining stable haemodynamics. A maximal blood flow rate of 124ml/min/kg was obtained. Arterio-venous PO2 gradients were 10-fold (FIO2@100%) with only small variations when changing blood flow rates. Conclusions: The results obtained with this new, standardized and miniaturized hollow fibre oxygenator, new cannulation technique and CPB circuit, achieves optimal gas transfer with small asanguinous priming volumes. This study opens new potentials for various CPB-related study protocols in the small animal.
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Validation of a Rat Model of Cardiopulmonary Bypass With a New Miniaturized Hollow Fiber Oxygenator
Article
  • Sep 2008
Cardiopulmonary bypass (CPB) is an essential component of cardiac surgery, with still unknown device/patient interactions. To evaluate the response of CPB to hemodynamic, biochemical, inflammatory, as well as thermo-pharmacodynamic interactions, a novel miniaturized oxygenator with controlled and standardized specifications has been developed together with an improved surgical central cannulation technique. A hollow-fiber small priming volume (6.3 ml) oxygenator was manufactured according to specifications resulting from engineering, heart surgery and perfusion expertise (Dideco-Sorin Group, Italy) with the following characteristics: Gas Exchange Surface--450 cm2, and Heat Exchange Surface--16 cm2. The oxygenator was tested in vitro and in vivo in five anesthetized, ventilated, open-chest rats using a miniaturized roller pump. Pressures were monitored in the animal before and after the oxygenator. Central venous cannulation through the superior vena cava and aortic cannulation through the carotid artery were used. In vitro: blood oxygenation increased 10-fold (from room air to 100% O2) and PCO2 removal was 2.5-fold. In vivo: CPB was performed without blood prime for 90 minutes (no ventilation) maintaining stable hemodynamics. A maximal blood flow rate of 124 ml/min/kg was obtained. Arterio-venous PO2 gradients were 10-fold (O2 100%) with only small variations when changing blood flow rates. This new, standardized and miniaturized hollow fiber oxygenator, new cannulation technique and CPB circuit achieved optimal gas transfer with small asanguinous priming volumes. This study opens new potentials for various CPB-related study protocols in the small animal.
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Complement activation in coronary artery bypass grafting patients without cardiopulmonary bypass - The role of tissue injury by surgical incision
Article
  • Nov 1999
Complement activation is a trigger in inducing inflammation in patients who undergo coronary artery bypass grafting (CABG) and is usually thought to be induced by the use of cardiopulmonary bypass (CPB). In this study, we examined whether tissue injury caused by chest surgical incision per se contributes to complement activation in CABG patients. Prospective study. Thorax center in university hospital. Twenty-two patients undergoing CABG without CPB were prospectively divided into two groups: a small chest incision via an anterolateral thoracotomy representing a minimized tissue injury (lateral group, n = 8), and a conventional median sternotomy representing a large tissue injury (median group, n = 14). Biochemical markers indicating complement activation as well as systemic inflammatory response were determined before, during, and after the operation. Plasma concentrations of complement 3a increased in both the lateral and median groups right after chest incision (p < 0.01 and p < 0.05, respectively) and by the end of operation increased only in the median group (p < 0.01). The terminal complement complex 5b-9 did not increase in the lateral group, but it did increase in the median group both after incision and by the end of the operation (p < 0.05 and p < 0.05, respectively). During surgery, complement 4a did not increase, suggesting that it is the alternative rather than the classic pathway that is involved in complement activation by tissue injury. Postoperatively, interleukin-6 production was greater in the median group (p < 0.01) than the lateral group (p < 0.05), suggesting a more pronounced inflammatory response to a larger chest incision. Tissue injury caused by surgical incision contributes to complement activation in CABG patients who are operated on without CPB. A small anterolateral thoracotomy is associated with reduced complement activation in comparison with a median sternotomy.
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A literature review of cardiopulmonary bypass models for rats
Article
  • Dec 1999
Cardiopulmonary bypass (CPB) has improved a great deal since its first applications in the early 1950s. If improvements are to be continued, a preclinical model of CPB for small animals is desirable, mainly because of convenience of equipment and low costs. We review the different models of CPB for rats that have been designed, discuss their characteristics and points where improvements may be made. We give suggestions and requirements for a new up-to-date model that could be a useful tool in continued research on the pathophysiology and therapeutic strategies of CPB.
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A recovery model of partial cardiopulmonary bypass in the rat
Article
  • Jun 2001
This study was undertaken to develop a recovery model of cardiopulmonary bypass (CPB) in rats. Twenty male Wistar rats (475-550 g) were anaesthetized, mechanically ventilated and the femoral vessels cannulated. The extracorporeal circulation circuit comprised a roller pump, a venous reservoir and a modified Capiox 308 paediatric membrane oxygenator. Priming consisted of 20 ml of fresh homologous blood and 15 ml of colloid. Anticoagulation was achieved with heparin (500 IU/kg). Blood gas analysis, blood pressure monitoring and survival studies were performed in CPB ( n=10) and Sham ( n=10) rats. Partial CPB was always easily established and was conducted at a flow rate of 100 ml/kg/min for 90 min. Blood gas analysis and blood pressure data did not differ between the two groups. All CPB rats survived and the 3-week follow-up period remained uneventful. The rat model of CPB was easy to perform and was associated with excellent survival. This recovery model should allow us to study the pathophysiological processes underlying post-CPB multiple organ dysfunction.
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Changes in mesenteric vascular reactivity and inflammatory response after cardiopulmonary bypass in a rat model
Article
  • Jul 2004
  • ANN THORAC SURG
Mesenteric ischemia and acidosis leading to intestinal ischemia has been observed during cardiopulmonary bypass (CPB) despite normal flow in the mesenteric vessels. The aim of this study was to assess mesenteric endothelium-dependent reactivity and vasoconstrictor responses of small mesenteric arteries in a rat model of CPB without aortic cross-clamping. After femoral cannulation a partial 90 minutes CPB was performed with hemodynamics and blood gas parameters monitoring. Blood samples and segments of small mesenteric arteries were obtained in rats sacrificed 2.5 hours (CPBH2.5) or 6 hours (CPBH6) after femoral cannulation. Sham surgery (sham H2.5, sham H6) was performed with femoral cannulation only. Segments of small mesenteric arteries were placed in a myograph in order to assess the contractile response to phenylephrine (with or without NO synthase inhibitor) or the endothelium-dependent relaxation to acetylcholine. Systemic inflammation was evaluated by measuring plasma concentrations of TNFalpha. Pulmonary and intestinal infiltration of activated leukocytes was assessed by immunohistochemistry. CPB induced increased contractile response to phenylephrine which persisted after blockade of NO synthesis as well as transient impairment of endothelium-dependent relaxations. CPB also led to early and marked release of TNFalpha. CPB was responsible for mesenteric endothelial dysfunction and direct increase in the contractile response to alpha1-adrenergic agonist with increased systemic inflammatory response. This phenomenon might contribute to an increase in the risk of mesenteric ischemic events during cardiac surgery especially when vasopressor agents are used.
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Protective Effects of Melatonin and N-Acetylcysteine on Hepatic Injury in a Rat Cardiopulmonary Bypass Model
Article
  • Oct 2007
An increasing number of patients were undergoing cardiac surgery with cardiopulmonary bypass (CPB) and more attention had been paid to hepatic injury after CPB. This study was designed to test the hypothesis that melatonin and N-acetylcysteine (NAC) could attenuate hepatic injury induced by CPB in rats. Male Sprague Dawley rats were randomly divided into four groups: sham, control (CPB + placebo), NAC (CPB + 250 mg/kg N-acetylcysteine), and melatonin (CPB + 20 mg/kg melatonin). Blood samples were collected at the beginning, at the end of CPB, and at 0.5, 1, 2, 3, and 24 h postoperation. Liver samples were harvested at 24 h after the operation. In the control group, the levels of serum liver enzymes and tumor necrosis factor-alpha, activities of inducible nitric oxide synthase, malondialdehyde, and myeloperoxidase in liver tissue were significantly increased. In addition, swollen hepatocytes, vacuolization, and congestion in sinusoids were observed. These changes were markedly reversed in both NAC and melatonin groups. Furthermore, the glutathione content and liver antioxidative enzymes activities were significantly decreased in the control group compared with the sham group. However, the levels of these antioxidants were markedly elevated after NAC or melatonin treatment compared with placebo treatment. Our findings showed that NAC and melatonin had acceptably beneficial effects against the CPB-induced hepatic injury.
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Work in progress report - Cardiopulmonary bypass: Hepatic injury in a rat cardiopulmonary bypass model
Article
  • Mar 2008
  • Interact Cardiovasc Thorac Surg
An increasing number of patients were undergoing cardiac surgery with cardiopulmonary bypass (CPB) and more attention had been paid to hepatic injury after CPB. This study was designed to study how CPB could induce and aggravate the hepatic injury in a rat model. Male Sprague-Dawley rats were randomly divided into two groups (n=12): sham and CPB groups. Blood samples were collected at the beginning, at the cessation of CPB, and at 0.5, 1, 2, 3 and 24 h post-operation. Liver samples were harvested at 24 h after operation. In CPB group, the levels of serum liver enzymes and tumor necrosis factor-alpha, activities of inducible nitric oxide synthase, malondialdehyde and myeloperoxidase in liver tissue were significantly increased. In addition, swollen hepatocytes, vacuolization and congestion in sinusoids were observed. On the contrary, the activities of liver antioxidative enzymes and the concentration of glutathione (GSH) decreased remarkably. All results indicated that CPB would induce and aggravate hepatic injury by facilitating oxidative stress and the systemic inflammatory response.
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Simvastatin Suppresses Lung Inflammatory Response in a Rat Cardiopulmonary Bypass Model
Article
  • Dec 2007
Inflammatory response in the lungs is a well-known complication after cardiopulmonary bypass (CPB). The main aims of our study were to explore whether pretreatment with simvastatin would inhibit toll-like receptor 4 expression and suppress lung inflammatory response in a rat CPB model. Male Sprague-Dawley rats were divided into four groups (n = 6 each): sham group; CPB (control group); CPB plus low-dose simvastatin (5 mg/kg daily [L-Sim group]); and CPB plus high-dose simvastatin (10 mg/kg daily [H-Sim group]). Blood samples were collected at the beginning and at the termination of CPB, and at 1, 2, 4, and 24 hours after operation. The bronchoalveolar lavage fluid and lungs were harvested 24 hours postoperatively. The simvastatin-treated groups had significantly higher ratios of PaO(2)/FiO(2) and lower values of respiratory index than the control group. We observed that simvastatin reduced CPB-induced toll-like receptor 4 and nuclear factor-kappaB expressions in CPB groups (p < 0.01, versus control group). The levels of interleukin-6, tumor necrosis factor-alpha, and monocyte chemotactic protein-1 in serum, bronchoalveolar lavage fluid, and lung tissues increased in CPB groups, whereas pretreatment with simvastatins reduced these inflammatory marks in a dose-dependent manner (p < 0.01, versus control group). Furthermore, pretreatment with simvastatin had a lower lung injury score (p < 0.05, versus control group). These findings suggest that simvastatin inhibited CPB-induced toll-like receptor 4 upregulation and nuclear factor-kappaB activation, efficaciously reduing the pulmonary inflammatory response after CPB.
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