| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Eur J Cardiothorac Surg 2006;30:492-498
© 2006 Elsevier Science NL
a Division of Thoracic and Cardiovascular Surgery, Hannover Medical School, Hannover, Germany
b Department of Pathology, Hannover Medical School, Hannover, Germany
c Clinic for Small Animals, University of Veterinary Medicine Foundation, Hannover, Germany
Received 17 November 2005; received in revised form 30 April 2006; accepted 31 May 2006.
* Corresponding author. Address: Division of Thoracic and Cardiovascular Surgery, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany. Tel.: +49 511 532 6581; fax: +49 511 532 5404. (Email: Khaladj.Nawid{at}mh-hannover.de).
 |
Abstract |
|---|
 Top Abstract 1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
Key Words: Hypothermic circulatory arrest • Selective cerebral perfusion • Perfusion temperature • Cerebral protection • Animal model
|
1. Introduction |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
Regarding the optimal temperature, different strategies are currently in use, mostly depending on empirical clinical data. Dr Bachet, one of the pioneers of SACP, as well as our group are using moderate hypothermia (26–28 °C) for HCA and cold temperatures for SACP (14 °C), achieving excellent clinical results with this regime [2,5]. In contrast to that, the Mount Sinai Group, affiliated with Dr Griepp, combined – based on their experimental data – cold core temperatures down to 11–14 °C with cold SACP [6,7].
Due to the potential detrimental side effects of extended extracorporal circulation times that are associated with deeper temperatures, several groups started to perform aortic arch surgery using warmer temperatures [8].
Based on an experimental porcine model from the Mount Sinai group, we investigated a modification, to perform SACP without additional cannulation of the carotid arteries [9]. In this study, we were able to demonstrate the advantages of cold SACP (20 °C) compared to HCA alone.
In the present study, the effects of profound (10 °C), deep (20 °C) and moderate (30 °C) SACP temperatures were compared with those of HCA alone. Among complex intraoperative neuromonitoring, post mortem analyses of brain tissue samples using standard histopathological evaluation and molecular genetic assessment were performed.
|
2. Materials and methods |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
2.2 Anesthesia, peri-operative management and neurophysiology
All animal received human care in compliance with the guidelines ‘Principles of Laboratory Animal Care’ formulated by the National Society for Medical Research and the ‘Guide for the Care and Use of Laboratory Animals’, published by the National Institute of Health (NIH Publication No. 88-23, revised 1996). The protocols for all experiments were approved by the Hannover Medical School Institutional Animal Care and Use Committee as well as the state Lower Saxony.
After pre-treatment with intramuscular azaperone (5 mg/kg) and atropine (0.5 mg), animals were anesthesized with intravenous disoprivan 1% (2 mg/kg). Endotracheal intubation was performed and the pigs mechanically ventilated with a FiO2 of 0.5 and isoflurane (1–2%). A positive endexperitatory pressure of 3–5 mmHg was frequently used; analgesia was achieved by continuous infusion of Fentanyl (1 µg kg–1 h–1). Intermittent intravenous injection of pancuroniumbromide (0.1 mg/kg) was performed to achieve paralysis. Ventilation parameters (rate, tidal volume) were adjusted to maintain the arterial carbon dioxide tension between 35 and 45 mmHg. All animals received 1 g of Ceftriaxon intravenously before intervention.
Urine output was measured with a Foley bladder catheter (8–10F) placed transurethral. Temperature probes were inserted in the rectum and the esophageous. For continuous monitoring of cardiac output and arterial pressure as well as for collection of arterial blood samples a PICCO-catheter (Pulsion Medical Systems AG, Munich, Germany) was placed into the right femoral artery after groin preparation (see below). For measuring of the central venous pressure and continuous venous infusion, a three-lumen catheter was advanced via the right femoral vein towards the right atrium.
After trepanation of the skull (0.7 cm burr hole), introduction of the intraparenchymal microtip pressure catheter (Codman ICP Express, Johnson and Johnson Professor Inc., Raynham, MA, USA) was performed, allowing continuous monitoring of the intracranial pressure (ICP). A 24G-catheter was placed into the sagittal sinus for blood sample collection.
Disposable subdermal needle electrodes for recording of SSEP of both median nerves and continous EEG-monitoring were placed in a standard fashion and connected with the neurophysiological device according to the manufactures direction (Preissler Medizintechnik GmbH, Augsburg, Germany). Analysis was performed by an investigator blinded to the experimental protocol.
2.3 General surgical technique and cardiopulmonary bypass
The surgical technique and cardiopulmonary bypass management have been described before [9]. Briefly, after preparation of the skull and the groin, a left thoracotomy was performed in the fourth intercostal space. After exploration of the supraaortic vessels and systemic heparinization (400 IU/kg), non-pulsative cardiopulmonary bypass (CPB) was instituted at a flow rate of 70–100 ml/kg after cannulation of the ascending aorta and pulmonary artery for venous return. Cooling was performed for a period of 45 min, reaching 20 °C body temperature. Blood gas analyses were carried out using alpha-stat principles. Methylprednisolone (20 mg/kg) was administered 30 min before HCA.
2.4 Selective antegrade cerebral perfusion (SACP)
The differences in pig anatomy necessary to modify perfusion technique in contrast to the human situation were described before [9]. After reaching 20 °C body temperature, aortic cross clamping was performed and CPB stopped. Passive venous drainage into the cardiotomy reservoir was allowed after institution of HCA. Dependence of the temperature for SACP, blood was allowed to circulate in the heart–lung-machine to reach the anticipated temperature. After 5 min, SACP was started with the corresponding temperature for 90 min. Perfusion was performed pressure controlled between 45 and 50 mmHg. Flow was indirectly controlled by near-infrared spectroscopy (NIRS) (INVOS, Somanetics, MA, USA).
2.5 Study protocol
Haemodynamic measurement was performed continuously during the whole experiment, blood samples (arterial, sagittal sinus) were collected and neuromonitoring recorded at the following time points:
2.6 Tissue harvesting
Right atrium was opened to allow passive drainage of blood and perfusate. The brain was flushed through the aortic cannula with 3 l of ice-cooled saline over 15 min to achieve tissue cooling and removal of red blood cells. Cross clamping was performed as for SACP. After perfusion, the scull was immediately opened and the entire brain and brainstem removed en bloc. All brains were cut into two pieces in the sagittal plane. Tissue samples were taken from the frontal cortex, hippocampus, medulla and cerebellum and fixed in 3.5% formalin or immediately snap frozen in liquid nitrogen and stored at –80 °C for further evaluation, respectively.
2.7 Tissue preparation and evaluation
2.7.1 Histopathology
After fixation of the tissue samples, transverse sections were embedded in paraffin, cut into 5 µm thick sections, and stained by using hematoxylin and eosin. Neuronal injury was semiquantitative evaluated at 200x magnification by a pathologist blinded to the experimental protocol for every area available (cortex, hippocampus, medulla and cerebellum). Damaged neurons were identified (edema, eosinophilic, necrotic, inflammatory, apoptotic cell types) and assigned to injury groups: 1–2 mild injury, 3–4 moderate injury, 5–6 severe injury, subdivided for focal and diffuse damage, respectively.
2.7.2 Molecular genetics
Extraction of total RNA was performed out the cortex by standard procedures according to the manufactures recommendation (RNeasy midi Kit, Quiagen, Hilden, Germany), followed by reverse transcription for further amplification (Omniscript RT Kit, Quiagen, Hilden, Germany). Primers were designed for house keeping genes (GAPDH, ß-actin) and for early ischemic brain markers according to literature review and our own results in a mouse model (HSP-72, c-fos, Cox-2) (Invitrogen, Karlsruhe, Germany), optimized for melting points and expected product length (Table 1
).
|
|
3. Statistics |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
|
4. Results |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
4.2 Experimental groups
A total of 20 animals were used for analysis (5 in each group). Animals that died before reaching the end of the experiment were excluded from the study (Section 4.1). Preoperative data (age, weight) showed no significant differences between groups. Basic haemodynamic data showed minor variations between groups in physiological parameters (heart rate, central venous pressure, mean arterial pressure and cardiac output). Due to interindividual variances in intracranial pressure, changes from baseline at each measurement time point were expressed. Arterial acid bases and blood gas parameters showed no significant differences among the groups, baseline sagittal sinus saturation levels were comparable.
4.3 Selective antegrade cerebral perfusion (SACP)
SACP was performed pressure controlled (50 mmHg). The flow rates among the groups vacilliated between 0.12 and 0.17 l/min. During the whole period of perfusion, the values where not statistically different (Table 2
).
|
|
|
|
|
|
|
|
5. Discussion |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
The advantage of antegrade cerebral perfusion for prolonged periods of HCA has been proven in clinical as well as experimental settings. Recent clinical reports describe side effects of SACP due to embolic events, triggered by unknown flow rates and driving pressures [10]. First experimental studies, dealing with these issues were focused on cerebral metabolism and neurological outcome [11].
Stimulated by the result of our last study, where we were able to demonstrate the superiority of SACP compared to HCA alone, we decided to compare different perfusion temperatures in the same setting [9]. Based on our previous experiences in this model, we decided to cool the animals to 20 °C core temperature before establishing SACP.
It has been shown that this temperature provides enough protection for other organs such as kidneys, liver and spinal cord despite the prolonged HCA time [12–14]. This admittedly relative long time of arrest is necessary, to produce enough brain damage to compare possible differences between the treatment groups [15,16]. For the same reason highly different SACP temperatures were applied in the experimental groups.
In the potpourri of neuromonitoring tools, the sagittal sinus saturation (SSS) is considered to be important to cover the reduction of the cerebral metabolism in the setting of HCA [17]. Silence of EEG signals were often used as sign of optimal suppression of the cerebral metabolism, but many studies showed that EEG signals can disappear not only due to hypothermia but also due to interaction with anaesthetic drugs [18].
The present study shows that 10 and 20 °C SACP provide sufficient cerebral protection over the 90 min period of SACP, indicated by stable or increased SSS, whereas in the 30 °C SACP group saturation decreases significantly until the end of SACP, never reaching baseline levels until elective sacrifice. The role of vascular resistance in this context cannot be answered in the current setting, but would further clarify whether higher flow rates are necessary to provide sufficient blood flow for the cerebrum.
In a recent study, we were able to demonstrate that intracranial pressure has been an important predictive tool of cerebral damage after hypothermic circulatory arrest. In this study a significant correlation between ICP and behavioural scores were found [12]. In another study, we showed the advantages of 20 °C SACP compared to HCA alone, which spurred us to evaluate the effects of different SACP temperatures [9].
In the current study, the ICP in the 30 °C SACP group was significant elevated through the entire perfusion and slightly increased during the reperfusion time. The other temperature groups remained close to baseline. In this context, several lines of evidence suggest that at least some of the injury may occur during reperfusion, raising the question if the extent of damage can be influenced by a modification in the reperfusion strategy [19]. On the other hand, the experimental set-up of cooling down to 20 °C with concomitant moderate brain perfusion may be in parts responsible for this finding. This set-up was chosen in accordance to our previous set-up and the fact, that 90 min HCA at warmer temperatures results in inadequate protection of the lower body (e.g. bowl, kidney, spinal cord, etc.). The problem of higher ICP at constant perfusion pressures and therefore a reduction of cerebral perfusion pressure may further aggravate a disturbance of the neurological integrity. These findings support Ehrlich's theory that cold reperfusion may be beneficial to improve cerebral outcome. But from the current study it seems also that perfusion with profound temperatures has a negative impact on ICP, not during SACP, but during reperfusion. Again, this may be due to a relatively high difference of core and brain temperatures during reperfusion. In this context, it has been shown previously, that profound temperatures are associated with a higher incidence of capillary leakage.
Despite the fact, that high ICPs during reperfusion indicate brain damage, it is interesting, that histopatholgical evaluation showed a reduction of edema and eosinophilic cells types of neurons in the groups treated with SACP, independent of the temperature. Since we have shown in the past that ICP is a powerful marker for neurological injury, we speculate that this finding may be due to a problem of sensitivity of our semiquantitative histopathological markers. But independent of these findings, the question remains, whether high ICPs are the result or the reflection of neurological damage.
To answer this question, more sophisticated tools are necessary to define the amount of cerebral injury. The idea of measuring immediate-early genes is not new in experimental brain research, especially in small animal models [20]. In these set-ups, the 72 kDa heat shock protein (HSP-72) has served as a useful indicator of ischemic stress after cerebral ischemia, but also excitatory stimuli or even heat [21]. An additional marker, expressed in the cerebrum and triggered by ischemic reperfusion injury is Cox-2, an isoform of cyclooxygenase (Cox). Cox-2 levels were shown to rise after cerebral ischemia triggered by several upstream response elements in various brain areas. The immediate-early gene c-fos is rapidly induced in neurons after seizures, hypoxia and global ischemia. The protein product of c-fos mRNA modulates transcription of several late-response genes. Some of these genes are associated with apoptosis, whereas others enhance cell survival. So far it is unknown whether c-fos expression is involved with cell survival or death, it is supposed to be a useful indicator of severely stressed neurons [22].
In our laboratory, HSP-72 has been established for the porcine model. The over proportional expression in the HCA and 30 °C SACP group is in our opinion another indicator for an imperfect cerebral protection in these animal groups.
It is surprising, that in contrast to our findings in small animals the expression of Cox-2 and c-fos levels showed no differences among treatment groups. This may be due to a different time course concerning the expression in the porcine model or the fact, that the amount of damage is not as high as in our small animal model.
|
6. Conclusion |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
On the basis of our experimental results, clinical trials with moderate temperatures for HCA and SACP should be reconsidered.
|
Appendix A |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
Dr M. Ehrlich (Vienna, Austria): You have used 100 min at 20 °C, which is quite a long time. Could you speculate that you would have achieved the same results, or even better ones, if you would have used a warmer temperature and a shorter HCA time, which would mimic more a clinical setting?
Dr Khaladj: We know that this is a very tough model, but to produce reliable results and to see potential differences between temperatures, we had to choose this long period of circulatory arrest and selective antegrade perfusion. Despite the fact that it is not really comparable to the clinical situation, we have the feeling that this model can provide further insight in the pathophysiology of cerebral ischemia and selective cerebral perfusion.
Dr T. Aybek (Frankfurt, Germany): I have one question about the method of the study. You use pressure-controlled perfusion. And what about flow at 10, 12 or 20 and 30 °C? That is the first question.
Dr Khaladj: The flows were not different in the groups. But you may be right that the flow in the 30 °C group was not high enough, since we had high intracranial pressure in this group. This may result in an insufficient cerebral perfusion pressure.
Dr Aybek: You use the same pressure at 30 °C and the same pressure at 20 °C?
Dr Khaladj: Yes.
Dr Aybek: And what were the steps? I understood that you cooled first to 12 °C and then make measurements and then warm up 20 °C.
Dr Khaladj: Actually, the body of all pigs were cooled to 20 °C core temperature. Then the blood temperature was adjusted to the anticipated temperature. During this short period of circulatory arrest (mimicing the introduction of our perfusion catheters in the clinical situation) perfusion was started according to the protocol.
Dr Aybek: If you cooled first to 20 °C and then rewarmed to 30 °C, I cannot say that the results are similar between 20 and 30 °C, because there are a lot of other experimental studies that show rewarming makes cerebral edema.
And another comment, you show that intracranial pressure is higher in 30 °C. What about if I use same temperature for coronary surgery, 30–32 °C, if I use CPB in your studies so that we have more intracranial pressure in 30 °C than 20 °C?
Dr Khaladj: One problem might be that we have cooled the body to 20 °C and then started perfusion with 30 °C. We agree, that this may aggravate cerebral edema. On the other hand we know from previous studies that high core temperatures in the porcine model of hypothermic circulatory arrest can cause major haemodynamic misbalance during reperfusion due to a complete loss of vascular resistance.
|
Acknowledgments |
|---|
We would like to thank Helmut Preissler from Preissler Medizintechnik GmbH, Augsburg, Germany for providing the neurophysiological devices for EEG and SSEP analysis.
The authors also thank Klaus Hoeffler (CCP) and Siegfried Bachmann for their valuable support, our research coordinators and technicians Astrid Diers-Ketterkatt, Petra Ziehme, Rosie Katt, Karin Peschel, Anja Giese for technical assistance and Kalle Napierski und Paul Zerbe for their care of the animals.
|
Footnotes |
|---|
Presented at the joint 19th Annual Meeting of the European Association for Cardio-thoracic Surgery and the 13th Annual Meeting of Thoracic Surgeons, Barcelona, Spain, September 25–28, 2005. |
References |
|---|
|
TopAbstract1. Introduction2. Materials and methods3. Statistics4. Results5. Discussion6. ConclusionAppendix AReferences |
|---|
This article has been cited by other articles:
|
|
 |
|
  H. Sasaki, K. J. Guleserian, R. Rose, C. Fotiadis, P. J. Boyer, and J. M. Forbess Hypothermic extracorporeal circulation in immature swine: a comparison of continuous cardiopulmonary bypass, selective antegrade cerebral perfusion and circulatory arrest Eur. J. Cardiothorac. Surg., December 1, 2009; 36(6): 992 - 997. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Allibhai, R. DiGeronimo, J. Whitin, J. Salazar, T. T.-S. Yu, X. B. Ling, H. Cohen, P. Dixon, and A. Madan Effects of moderate versus deep hypothermic circulatory arrest and selective cerebral perfusion on cerebrospinal fluid proteomic profiles in a piglet model of cardiopulmonary bypass J. Thorac. Cardiovasc. Surg., December 1, 2009; 138(6): 1290 - 1296. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. T. Strauch, P. L. Haldenwang, K. Mullem, M. Schmalz, O. Liakopoulos, H. Christ, J. H. Fischer, and T. Wahlers Temperature dependence of cerebral blood flow for isolated regions of the brain during selective cerebral perfusion in pigs. Ann. Thorac. Surg., November 1, 2009; 88(5): 1506 - 1513. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Pacini Invited commentary. Ann. Thorac. Surg., November 1, 2009; 88(5): 1514 - 1514. [Full Text] [PDF]
|
|
|
|
 |
|
 N. Khaladj, I. Ismail, M. Shrestha, S. Peterss, M. Pichlmaier, K. Kallenbach, A. Haverich, and C. Hagl Aortic root surgery in combination with hypothermic circulatory arrest: preserve or replace the aortic valve in the context of postoperative neurological outcome? A case match comparison Interactive CardioVascular and Thoracic Surgery, August 1, 2009; 9(2): 246 - 250. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Khaladj, S. Peterss, A. Haverich, and C. Hagl eComment: Selective antegrade cerebral perfusion and metabolicsuppression Interactive CardioVascular and Thoracic Surgery, June 1, 2009; 8(6): 653 - 653. [Full Text] [PDF]
|
|
|
|
|
|
O. Jonsson, G. Myrdal, V. Zemgulis, J. Valtysson, L. Hillered, and S. Thelin Selective antegrade cerebral perfusion at two different temperatures compared to hypothermic circulatory arrest - an experimental study in the pig with microdialysis Interactive CardioVascular and Thoracic Surgery, June 1, 2009; 8(6): 647 - 653. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Khaladj, M. Shrestha, S. Peterss, M. Strueber, M. Karck, M. Pichlmaier, A. Haverich, and C. Hagl Ascending aortic cannulation in acute aortic dissection type A: the Hannover experience Eur. J. Cardiothorac. Surg., October 1, 2008; 34(4): 792 - 796. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Khaladj, M. Shrestha, S. Meck, S. Peterss, H. Kamiya, K. Kallenbach, M. Winterhalter, L. Hoy, A. Haverich, and C. Hagl Hypothermic circulatory arrest with selective antegrade cerebral perfusion in ascending aortic and aortic arch surgery: A risk factor analysis for adverse outcome in 501 patients. J. Thorac. Cardiovasc. Surg., April 1, 2008; 135(4): 908 - 914. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Meybohm, G. Hoffmann, J. Renner, A. Boening, E. Cavus, M. Steinfath, J. Scholz, and B. Bein Measurement of Blood Flow Index During Antegrade Selective Cerebral Perfusion with Near-Infrared Spectroscopy in Newborn Piglets Anesth. Analg., March 1, 2008; 106(3): 795 - 803. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Khaladj, A. Haverich, and C. Hagl Should a Patient With Acute Aortic Dissection Type A Go to the Intensive Care Unit or Operating Room? Ann. Thorac. Surg., September 1, 2007; 84(3): 1069 - 1069. [Full Text] [PDF]
|
|
|
|
|
|
D. Pacini, A. Leone, L. Di Marco, D. Marsilli, F. Sobaih, S. Turci, V. Masieri, and R. Di Bartolomeo Antegrade selective cerebral perfusion in thoracic aorta surgery: safety of moderate hypothermia Eur. J. Cardiothorac. Surg., April 1, 2007; 31(4): 618 - 622. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Ates and A. U. Gullu Which temperature is better in acute type A aortic dissection? Eur. J. Cardiothorac. Surg., January 1, 2007; 31(1): 138 - 138. [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| ANN THORAC SURG | ASIAN CARDIOVASC THORAC ANN | EUR J CARDIOTHORAC SURG |
| J THORAC CARDIOVASC SURG | ICVTS | ALL CTSNet JOURNALS |
