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1 Cardiovascular Mechanics and Biofluid Dynamics, Institute Biomedical Technology, Ghent University, Belgium 2 Haemodynamic Research Center (HemoLiège)3 Department of Pharmacy, University of Namur, Belgium4 Department of Medicinal Chemistry5 Thermodynamics of Irreversible Processes, University of Liège, Belgium
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Abstract |
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(Received 10 July 2006;
accepted after revision 9 October 2006; first published online 12 October 2006)
Corresponding author L. Lanoye: Hydraulics Laboratory, Institute Biomedical Technology, St-Pietersnieuwstraat 41, 9000 Gent, Belgium. Email: lieve.lanoye{at}ugent.be
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Introduction |
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Reperfusion injuries have mostly been assessed at the cellular level, and the direct causes of necrosis and apoptosis have been studied at the molecular level. Many different mechanisms of reperfusion-induced cell damage and cell death have been identified. For example, oxidants and free radicals play an important role in reperfusion injury (Zweier & Talukder, 2006). In addition, cell damage caused by Ca2+ overload-induced contracture and reoxygenation-induced rigor contracture is observed (Piper et al. 2004).
Nonetheless, it is equally important to study the overall cardiovascular changes that result from the ischaemia–reperfusion-induced mechanisms. Some more recent studies have focused on the influence of ischaemia and reperfusion on circulatory parameters in pigs (Kamimura et al. 2003; Khalil et al. 2005; Metzsch et al. 2006). In all these studies, however, only a limited haemodynamic analysis was performed, and the studies did not incorporate pressure–volume analysis or ventriculo-arterial coupling, which enable assessment of the ‘matching’ between cardiac and arterial parameters that results in a working point of the cardiovascular system (producing a certain cardiac output and blood pressure) and its associated energetic cost.
The primary aim of this study was to gain insight into and document the wide range of alterations in haemodynamics induced by prolonged ischaemia and subsequent reperfusion. We chose to perform the experiment in a large animal model with coronary structure and function closely resembling those of humans. Focusing on ventriculo-arterial function and coupling, we analysed our data in terms of the ‘classical’ ventriculo-arterial elastance, as well as in terms of a recently proposed coupling concept using both a stiffness and a temporal coupling index.
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Methods |
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All experimental procedures and protocols used in this investigation were reviewed and approved by the Ethics Committee of the Medical Faculty of the University of Liège. All procedures conformed to the Guiding Principles in the Care and Use of Animals of the American Physiological Society and were performed according to the Guide for the Care and Use of Laboratory Animals (NIH publication no. 85–23, revised 1996).
Experiments were performed on six healthy pure pietran pigs of either sex (20–28 kg). The animals were premedicated with intramuscular administration of ketamine (20 mg kg–1) and diazepam (1 mg kg–1). Anaesthesia was then induced and maintained by a continuous infusion of sufentanil (0.5 µg kg–1 h–1) and sodium pentobarbitone (3 mg kg–1 h–1). Spontaneous movements were prevented by pancuronium bromide (0.1 mg kg–1). After endotracheal intubation through a cervical tracheostomy, the pigs were connected to a volume-cycled ventilator (Evita 2, Dräger, Lübeck, Germany) set to deliver a tidal volume of 10 ml kg–1 at a respiratory rate of 20 min–1. Measurements of end-tidal partial pressure of CO2 (Capnomac, Datex, Helsinki, Finland) were used to monitor the adequacy of ventilation. Respiratory settings were adjusted to maintain end-tidal CO2 in the range of 35–40 mmHg (4.67–5.33 kPa). Arterial oxygen saturation was closely monitored and maintained above 95% by adjusting the fractional inspired O2 as necessary. Core temperature was measured with a rectal probe and maintained at 37°C by means of a heating blanket. A standard lead electrocardiogram was used for the monitoring of heart rate (HR).
The chest was entered through median sternotomy, the pericardium was incised and sutured to the chest wall to form a cradle for the heart, and the root of the aorta was dissected clear of adherent fat and connective tissue. A combined conductance–micromanometer catheter (CD Leycom, Zoetermeer, The Netherlands) was inserted through the right carotid artery and advanced into the left ventricle. A micromanometer-tipped catheter (Sentron pressure measuring catheter, Cordis, Miami, FL, USA) was inserted through the right femoral artery and advanced into the ascending aorta. A 14 mm diameter perivascular flow probe (Transonic Systems Inc., Ithaca, NY, USA) was closely adjusted around the aorta 2 cm downstream to the aortic valve. The micromanometer-tipped catheter was manipulated so that the pressure sensor was positioned just distal to the flow probe. Right atrial pressure was measured with a micromanometer-tipped catheter inserted into the cavity through the superior vena cava. A 2.0 mm Fogarty balloon catheter (Baxter Healthcare Corp., Oakland, CA, USA) was advanced into the inferior vena cava through a right femoral venotomy. Inflation of this balloon produced a leftward shift in pressure–volume (P–V) loops by reducing venous return. Thrombus formation along the catheters was prevented by administration of 100 i.u. kg–1 of heparin sodium intravenously just before the catheter insertion.
A segment of the left anterior descending (LAD) coronary artery was isolated distal to the first diagonal branch for later application of a surgical clamp. An electromagnetic flow probe (Transonic Systems Inc.) was placed around the LAD coronary artery 3 cm distal to the site of occlusion, to verify the occlusion (complete loss of coronary artery blood flow). In case of occurrence of ventricular fibrillation, animals were defibrillated at 1 J kg–1 using internal paddles.
Experimental protocol
The animals were continuously infused intravenously with Ringer lactate solution (5 ml kg–1 h–1), and, when necessary, with hydroxyethyl starch (6%) to maintain central venous pressure up to 6–7 mmHg over 30 min. After placement of all instrumentation and verification of their proper functioning, animals were allowed to equilibrate for an additional 30 min, after which baseline (BL) recordings were made. Thereafter, the LAD coronary artery was occluded at the site of previous dissection for 1 h, and measurements were repeated before coronary perfusion was restored (T60). Subsequent measurements were repeated after 2 (T180) and 4 h of reperfusion (T300; Fig. 1A).
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The conductance catheter was connected to a Sigma-5 signal-conditioner processor (CD Leycom). Each ultrasonic flow probe was connected to a flowmeter (HT 207, Transonic Systems Inc.), and each micromanometer-tipped catheter to the appropriate monitor (Sentron pressure monitoring). All analog signals and the ventricular pressure–volume loops were displayed on screen for continuous monitoring. The analog signals were continuously converted to digital form with appropriate software (Codas, DataQ Instruments Inc., Akron, OH, USA) at a sampling frequency of 200 Hz.
Ventricular systolic function and energetics.
Ventricular systolic function was assessed through different parameters. Left ventricular volumes were inferred using the dual field conductance catheter technique (Baan et al. 1984; Steendijk et al. 1993). Calibration of the conductance signal to obtain absolute volume was performed by the hypertonic saline method (Baan et al. 1984). Therefore, a small volume (1–2 ml) of 10% NaCl solution was injected into the pulmonary artery during continuous data acquisition. Stroke volume (SV) was calculated from steady-state recordings as the difference between end-diastolic (EDV) and end-systolic volume (ESV). Cardiac output (CO) was calculated as the product of heart rate (HR) and SV. Ejection fraction (EF) was calculated as SV/EDV. Left ventricular contractile function was assessed by the end-systolic pressure–volume relation (ESPVR) (Suga et al. 1979), and the preload recruitable stroke work (PRSW) relation (Glower et al. 1985). Both indices require pressure–volume loops (PV loops) measured under gradual preload decline obtained through inhibition of venous return (Fig. 1D). All measurements were taken immediately after the animal was disconnected from the ventilator to sustain end expiration. The caval occlusion was limited to a few seconds to avoid reflex responses. The instantaneous pressure–volume relationship was considered in terms of a time-varying elastance E(t), defined by:
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| (1) |
Myocardial energetics were assessed by computation of pressure–volume area (PVA) using steady-state measurements. In the time-varying elastance model of the ventricle, the total energy generated by each contraction is represented by the total area under the end-systolic pressure–volume relation line and the systolic segment of the pressure–volume loop, and above the end-diastolic pressure–volume relation curve, and denoted by PVA (Suga et al. 1979). Pressure–volume area is the sum of SW, or the energy that the ventricle delivers to the blood at ejection, and potential energy (PE), necessary to overcome the viscoelastic properties of the myocardium itself. It has been demonstrated that PVA is highly correlated with myocardial oxygen consumption (Denslow, 1996). Mechanical efficiency is defined as the SW/PVA ratio.
Arterial function.
Arterial properties were assessed from ascending aortic pressure and flow measurements during steady-state conditions and represented with a four-element windkessel model (WK4; see Fig. 1B; Lambermont et al. 1998). In this WK4 model, the resistor R2 represents the resistive properties of the systemic bed, which reside primarily in the arteriolar system. The capacitor, C, placed in parallel with R2, represents the compliant properties of the systemic vessels. The resistor R1 represents the characteristic impedance, the level of which depends predominently on the elastic properties of the proximal aorta. Finally, an inductance, L, is introduced to take blood inertia into account. Furthermore, L restores positive phase angles at high frequencies of the impedance spectrum (Grant & Paradowski, 1987). The values of R1, R2, C and L were estimated by a previously described method (Lambermont et al. 1998). Effective arterial elastance (Ea) was calculated according to the equation (Sagawa et al. 1988):
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| (2) |
Ventriculo-arterial coupling. Ventriculo-arterial (VA) coupling was first studied through the ratio Ees/Ea, where Ees is the ventricular end-systolic stiffness. It has, however, been demonstrated by Stergiopulos et al. (1996) and Segers et al. (2002) that VA coupling can be described in more detail using two dimensionless parameters, i.e. a ‘stiffness coupling index’, calculated as the ratio of Ees and arterial stiffness (1/C), yielding EesC, and the ‘temporal coupling index’, calculated as the ratio of the time constant of the arterial system (R2C) and the heart period (T), R2C/T. We applied both the ‘conventional’ VA coupling approach, as well as this newer approach with the stiffness and temporal coupling indices.
Histopathological data collection
At the end of the protocol, the animal was killed with an intravenous injection of pentobarbitone (100 mg kg–1). The heart was then rapidly harvested, rinsed with a cold isotonic saline solution and sectioned in five slices (0.6 cm thick) parallel to the atrioventricular groove from apex to base. Transverse tissue slices were rinsed with a cold isotonic saline solution and placed in 10% neutral-buffered formaldehyde. After 3 days of fixation, three or four tissue blocks were taken from each slice at standardized locations and routinely processed for paraffin histology. The tissue blocks from myocardium were cut at 6 µm, and each section was stained with Haematoxylin and Eosin (H&E) and Masson's trichrome. The stained sections were examined at magnifications of x25, x100, x200 and x400 to study the distribution of infarction. Photomicrographs were taken using a Zeiss photomicroscope (Axioscope 2 plus Sony 3CCD camera, 1024–768 pixels definition, Thornwood, NY, USA). In addition, tissue blocks of slice S3 were processed for immunohistochemical staining of desmin to investigate lesions of the cardiac muscle. Polyclonal antibodies to myoglobin (Dako, Glostrup, Denmark) were diluted 1:400. Monoclonal antibodies to desmin (clone D33, Biomeda, Foster City, CA, USA) were diluted 1:20. These methods to detect myocardial lesions were developed previously in other experimental protocols in the pig (Rolin et al. 2003; Dogné et al. 2005; Kolh et al. 2006).
Statistical analysis
Differences between the four measurements at baseline, after 1 h of ischaemia and after 2 and 4 h of reperfusion were tested with a general linear model for repeated measurements using SPSS software (SPSS Inc., Chicago, IL, USA). The model first revealed statistically significant differences in the evolution with time of the different parameters, with the level set at P = 0.05. If differences were present, post hoc tests were performed to detect at which instances in time the data were different. The Sidak confidential interval adjustment was used.
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Results |
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Ligation of the LAD coronary artery induced an increase in HR from 97.9 ± 15.6 beats min–1 at BL to 106.4 ± 12.8 beats min–1 at T60 (P < 0.001). Cardiac output and mean aortic pressure (MAP) were not significantly affected by an hour of ischaemia, while EF decreased from 54.5 ± 11.3 (BL) to 45.4 ± 9.0% (T60; P = 0.033).
End-diastolic volume did not change significantly between BL and T60, but there was a trend towards elevated ESV (35.8 ± 16.2 ml at BL versus 42.9 ± 13.1 ml at T60, P = 0.074). Over the same time interval, SV decreased from 39.2 ± 11.6 ml to 34.0 ± 3.8 ml (P = 0.001).
Neither end-systolic elastance nor Vd changed significantly between baseline and T60. The slope (PRSWs) and the intercept (PRSW0) of the preload recruitable stroke work curve were not significantly affected by the ischaemia either (Fig. 2).
Stroke work decreased from 3600 ± 649 to 3200 ± 342 mmHg ml (P = 0.002), while pressure–volume area and mechanical efficiency (SW/PVA) did not significantly change between BL and T60.
As for vascular function, we observed that the only four-element windkessel parameter that demonstrated significant changes was characteristic impedance R1, which decreased from 0.077 ± 0.018 to 0.060 ± 0.012 mmHg s ml–1 (P = 0.001) between BL and T60. Peripheral resistance R2, C and L did not change significantly (Fig. 3). Effective arterial elastance (Ea) increased from 2.75 ± 1.45 to 3.35 ± 1.60 mmHg ml–1, between baseline and T60 (P = 0.007).
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Effects of sustained coronary reperfusion (T180 and T300 versus T60)
Heart rate increased further after reperfusion (from 106.4 ± 12.8 beats min–1 at T60 to 120.6 ± 14.7 beats min–1 at T180; P = 0.001), while sustained reperfusion (T300) did not lead to a further increase. Compared with ischaemia (T60), reperfusion induced a decrease in CO (from 59.8 ± 6.4 ml s–1 at T60 to 44.6 ± 7.0 ml s–1 at T180; P < 0.001), and CO remained at this level up to T300. Ejection fraction decreased (from 45.4 ± 9.0% at T60 to 31.1 ± 5.0% at T180; P < 0.001) and remained at this low level during sustained reperfusion. Mean arterial pressure decreased from 102.7 ± 22.5 mmHg at T60 to 88.1 ± 19.6 mmHg at T180 (P = 0.024) and to 78.4 ± 21.4 mmHg at T300 (P = 0.021).
While end-diastolic volume remained unchanged, ESV increased from 42.9 ± 13.2 ml at T60 to 50.9 ± 12.3 ml at T180 (P = 0.014) and did not further increase at T300. Coronary reperfusion induced a decrease in SV from 34.0 ± 3.8 ml at T60 to 22.2 ± 2.5 ml at T180 (P < 0.001). Stroke volume did not change further between T180 and T300.
End-systolic elastance (Ees) showed an increasing trend from 1.66 ± 0.79 mmHg ml–1 at T60 to 2.19 ± 0.97 mmHg ml–1 at T180 (P = 0.063) and did not show a further increase with sustained reperfusion. The dead volume (Vd) increased significantly from –18.70 ± 10.77 ml at T60 to 4.08 ± 13.46 ml at T180 (P = 0.005), but did not increase further with sustained reperfusion. The preload recruitable stroke work slope and intercept, in contrast, were not significantly influenced by coronary reperfusion.
Stroke work decreased as a result of early reperfusion, from 3200 ± 342 mmHg ml at T60 to 1889 ± 442 mmHg ml at T180 (P < 0.001), and did not change with sustained reperfusion. In the same interval, PVA and SW/PVA were not significantly affected by reperfusion.
Applying the four-element windkessel model, we observed that characteristic impedance R1 increased significantly, from 0.060 ± 0.012 mmHg s ml–1 at T60 to 0.073 ± 0.023 mmHg s ml–1 at T180 (P = 0.014), after which R1 remained constant (Fig. 3). Peripheral resistance R2 showed a similar pattern, increasing from 1.71 ± 0.78 mmHg s ml–1 at T60 to 2.16 ± 0.89 mmHg s ml–1 at T180 (P = 0.044) and remaining at an elevated level with sustained reperfusion, while global vascular compliance C and inertance L remained unchanged. Effective arterial elastance (Ea) increased further to 4.76 ± 2.11 mmHg ml–1 at T180 (P = 0.020), and remained constant at T300.
When looking at the left ventriculo-arterial coupling (Fig. 4), no statistical changes were observed for the ratio Ees/Ea from T60 to T300, whereas the ratio R2C/T increased further from 2.500 ± 0.417 at T60 to 3.261 ± 0.678 at T180 (P = 0.023), after which this parameter remained constant. The product EesC remained unchanged during the whole period.
Histopathological examination of infarcted myocardium
Histopathological examination revealed that main ischaemic changes were observed in the anterior left ventricle, corresponding to the area supplied by the LAD coronary artery. Reperfused myocardium displayed altered muscle cells, showing a waviness aspect which expresses prominent myofibrillar contraction bands, anisokaryosis and accumulation of interstitially infiltrated inflammatory cells as neutrophils and lymphocytes (Fig. 5A). Immunohistochemical detection of desmin revealed, mainly in the anteroseptal region, blurred zones where it was difficult to clearly delineate a necrotic zone from healthy myocardium owing to a motley appearance (Fig. 5B). Clear zones of patchy necrosis were also observed. The area not supplied by the LAD artery (posterior region) showed myofibrils with normal morphology.
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Discussion |
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Pigs were used in this study because of the similarity between the hearts of pigs and humans concerning the anatomical distribution of coronary arteries, the lack of collateral circulation and the heart-to-body weight ratio (White & Bloor, 1981; Schaper et al. 1988). Open-chest preparation was used because it offers the advantage of a very strictly controlled induction of myocardial ischaemia and reperfusion using a clamp.
Ventricular systolic function and energetics
We assessed LV systolic function by the slope and intercept of the ESPVR and the PRSW, both of which have been demonstrated to be relatively insensitive to changes in preload and afterload (Suga et al. 1979; Glower et al. 1985). Our data illustrate some of the difficulties associated with the interpretation of these indices. An increase in contractility results in an ESPVR that shifts to the left and upwards (Suga et al. 1979), i.e. a decrease of Vd combined with an increase of end-systolic elastance, Ees. We anticipated that contractility would be depressed following myocardial ischaemia, but this was not reflected by the slope of the ESPVR, which increased throughout the protocol. The dead volume, however, shifted to the right, indicating a loss of contractile function (Fig. 2).
To better elucidate the net effect of concomitant changes in slope and intercept of the ESPVR, we calculated the maximal realizable pressure, Pmax, the pressure that would be generated by the LV in a sustained isovolumic contraction at EDV: Pmax = Ees(EDV – Vd). Figure 6A shows that an increase in Ees results in an increase in Pmax (roughly equivalent to an increase in contractility), while an increase in Vd results in a decrease in Pmax (roughly equivalent to a decrease in contractility). Clearly, increases in Ees as well as Vd, as seen in this study, have opposite effects on the maximal Pmax. In Fig. 6B, we used averaged ESPVR values and calculated Pmax using EDV data at BL, T60, T180 and T300. The increase in Vd and increase in Ees counterbalance, and Pmax remains unchanged. This finding is in line with the observation that the slope of the PRSW was virtually unchanged throughout the experiment. Note, however, that this does not imply that LV systolic function was unchanged. The higher Ees may be the result of elevated sympathetic tone in response to the ischaemia, using the functional reserve of the LV to withstand the effects of ischaemia and making LV function more sensitive to changes in volume and afterload. Also, there was a trend for an increase in the intercept of the PRSW. Since the end-diastolic volume did not change, this rightward shift of the PRSW with ischaemia and reperfusion will result in a drop of stroke volume, which was observed, and thus a drop of inotropy. These findings illustrate that when studying LV function, both the slope and the intercept of the ESPVR and PRSW should be taken into account to reveal a complete picture of LV function.
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Histopathological data collection
So far, we have only discussed ‘macroscopic’ findings and effects on global cardiovascular function, and we found that reperfusion following ischaemia does not restore function. The sustained haemodynamic decline that is observed is in agreement with the histological findings, in which we also observed features of irreversible damage despite the reperfusion. The question is whether the damage results from the ischaemia or from the ischaemia and subsequent reperfusion (true reperfusion injury). In two (additional) animals that we killed after 1 h of ischaemia, we found signs of necrosis within the area at risk, suggesting that 1 h of ischaemia induced by LAD ligation is already sufficient to induce severe lesions in pigs. This is in accordance with the findings of Taylor et al. (1984), who also demonstrated that 1 h of ischaemia was sufficient to produce severe myocardial lesions in pigs. Like Taylor and co-workers, we believe that this probably results from the absence of a collateral circulation in the pig heart. We should nevertheless note that beyond the signs of necrosis observed, clear delineation between the reversible and the irreversible areas was not obvious even with immunohistochemical analyses, owing to the motley appearance of the staining in the myocardium from the anteroseptal region.
Arterial function
Besides LV function, we also addressed arterial function changes. Myocardial ischaemia (T60) does not affect systemic vascular resistance (R2) which, together with a preserved cardiac output, results in an unaffected mean arterial blood pressure. However, an initial decrease of characteristic impedance R1 is seen. Since R1 depends merely on the elastic properties of the proximal aorta, this decrease, together with an unaffected mean aortic pressure, is suggestive of a (small) dilatation of the proximal aorta and/or decrease in aortic stiffness. With reperfusion (T180 and T300), R2 increases, but because of the decrease in cardiac output, mean blood pressure decreases, resulting in higher R1 (compared with T60) as the aorta will be less distended. Given the fact that sympathetic tone is likely to be elevated in these conditions, the higher R1 may not be solely the effect of passive changes in aortic cross-section and function, but may also be modulated by changes in aortic vascular smooth muscle tone.
Ventriculo-arterial coupling
The coupling between the ventricle and the arterial system is commonly studied in terms of Ees/Ea. It has been argued, however, that Ees/Ea is not a very sensitive parameter (Segers et al. 2002). Another problem involves the use of Ea as representing arterial function, since Ea cannot be considered as a ‘pure’ arterial function parameter (Segers et al. 2002). In this study, ischaemia and reperfusion induced an alteration in ventricular and arterial function, yet the change in ventriculo-arterial coupling expressed using Ees/Ea was insignificant. This preservation of Ees/Ea may result from the fact that coupling is preserved, but it may also be a demonstration of the relative insensitivity of the parameter.
In this respect, it may be relevant to address coupling using the approach first advocated by Stergiopulos et al. (1996), who considered two independent coupling parameters, R2C/T and EesC, in order to assess the ventriculo-arterial coupling in a more elaborate model of the interaction between the LV and the arterial tree. The temporal coupling index R2C/T relates the characteristic time periods of both the ventricle (T) and the arterial tree (R2C). The product R2C is the time constant of the arterial windkessel, and determines the time constant with which the pressure decays during diastole. In all mammals, R2C/T is similar, which implies that the systolic-to-diastolic pressure difference (the pulse pressure) is constant. An increase in heart rate (smaller T) and R2C/T allows less time for the arterial pressure to decay in diastole, with higher diastolic pressures (coronary perfusion pressure) as a consequence. In a setting of myocardial ischaemia, elevated coronary perfusion pressures may be beneficial. The stiffness coupling index, EesC, is in fact the ratio of the ventricular stiffness Ees and the arterial stiffness, 1/C. This ratio may be more meaningful than the usually used Ees/Ea because Ea is not a pure arterial parameter (Segers et al. 2002) and better describes the compliance mismatch between the arterial tree and the ventricle. In this study, the ratio is not significantly affected by ischaemia or by reperfusion, so there are no significant relative compliance changes of the arterial tree compared with the maximal compliance of the left ventricle. At this point, it is hard to tell which of the two coupling frameworks is more appropriate. It is certain that, although Ea has the dimensions of stiffness, it is not a measure of arterial stiffness (Segers et al. 2002). It should also be pointed out that the Ees/Ea framework evolved from the analysis of cardiovascular function in the pressure–volume plane, where the dimension ‘time’ is not taken into account. Changes in heart rate are therefore not explicitly accounted for by Ees/Ea (although they are implicitly included in Ea). As such, the framework as proposed by Stergiopulos et al. (1996) may be better suited to detect the effects of subtle changes in arterial stiffness and timing, and to place these changes in a context of ventriculo-arterial coupling.
Limitations
It is obvious that this study has some stringent limitations. Firstly, since this study mainly focused on the haemodynamic alternations observed during ischaemia and reperfusion, only limited histopathological data were collected. Nevertheless, it is important to underline that oxidants and free radicals generated in the ischaemic and reperfused heart are key mediators in the pathogenesis of the injury (Zweier & Talukder, 2006). However, since accumulation of interstitially infiltrated inflammatory cells was observed at the histological level, no more complementary analyses were realized at the molecular level because biochemical dosages, histological analyses and electron microscopy data presented irreversible injury.
Secondly, our data indicate that in a pig model, 1 h of ischaemia may be too long when studying reperfusion injury. Confronting the histological findings with the haemodynamic data, it is hard to attribute the decline in cardiac function solely to reperfusion injury. It is likely that we observed a dynamic process with a complex interaction between myocardial stunning, myocardial hibernation and progressive necrosis of myocardial areas where irreversible damage has occurred. Acute experiments in an open-chest setting are probably too short to capture the time scale within which stabilization of events (and cardiac function) occurs. However, close monitoring of cardiovascular function, as we did in this experiment, is extremely demanding in a more ‘chronic’ experimental setting, and it is also not easy to differentiate the effect of the animal preparation and surgical interventions from the impact of ischaemia and subsequent reperfusion. Also, further exploration of ischaemia–reperfusion injury would require the necessary control experiments to differentiate effects of ischaemia from effects of ischaemia–reperfusion. In this context, we refer to previous work of Kolh et al. (2003, 2005) where, in a setting of sustained occlusion, it was shown that haemodynamic alterations took place primarily during the first half-hour of ischaemia, followed by a plateau phase that was maintained for 3 h. These findings thus support the assumption that the changes observed in the present study are not solely due to ischaemia, and that reperfusion injuries may play a role in the haemodynamic decline.
Conclusion
In conclusion, we have demonstrated that 1 h of ischaemia in a pig model induces haemodynamic alterations which are not reversed by reperfusion, but rather sustained and worsened. It is unlikely that these alterations are attributable to ischaemic injury alone, indicating that acutely ischaemic pigs provide a valuable large animal model for further study of pharmacological treatments to minimize reperfusion injuries. We have further demonstrated that studying ventriculo-arterial coupling through both a stiffness and temporal coupling index may provide more detailed insights than the more commonly used Ees/Ea ratio.
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P < 0.06 versus baseline. Indicated P values illustrate the statistically significant differences in the evolution with time of the different parameters.
P < 0.05 versus T180 measurement. Indicated P values illustrate the statistically significant differences in the evolution with time of the different parameters.
