Intra-atrial Temperatures in Radiofrequency Endocardial Ablation: Histologic Evaluation of Lesions

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1 Intra-atrial Temperatures in Radiofrequency Endocardial Ablation: Histologic Evaluation of Lesions Teresa Santiago, MS, João Q. Melo, MD, PhD, Rosa H. Gouveia, MD, and Ana P. Martins, MD Instituto do Coração, and Cardiothoracic Surgery and Pathology Department, Hospital Santa Cruz, Carnaxide, Portugal Background. Because of the limited information on the effects of ablation in human tissues, we studied intraatrial temperatures during endocardial radiofrequency applications. We correlated the intra-tissue temperatures with the tissue thickness and with the histologic appearance of the lesions. Methods. Radiofrequency currents were delivered to human atrial tissue, simulating conditions in endocardial ablation during surgery at set temperature of 70 and 80 C, and intra-tissue temperatures were measured with thermocouples. Radiofrequency applications at 70 C were performed in patients undergoing mitral valve surgery and biopsy specimens were obtained. Samples from in vitro studies and from patients were assessed histologically. Results. The subepicardial temperatures were usually over 60 C in applications in vitro at 70 C and over 70 C in applications at 80 C. Values were higher when the interior of the tissue was warmer than its surface as a result of consecutive radiofrequency applications over the same area. Histologic examination of 12 in vitro samples showed that 10 had transmural lesions. Five of 10 samples from patients with mitral valve surgery had lesions confined to the endocardium, 3 had damaged variable portions of the myocardium, and 2 had transmural lesions. Conclusions. Although it is possible to obtain transmural lesions in vitro and in vivo with endocardial applications at 70 C, it is significantly more difficult to achieve transmural lesions in patients with mitral valve disease than in normal atrial tissue in vitro. Consecutive applications can raise the intra-tissue temperatures to values significantly higher than those used for application. Our findings suggest that the composition of the endocardium and of the myocardium is a major determinant in lesion formation. (Ann Thorac Surg 2003;75: ) 2003 by The Society of Thoracic Surgeons When radiofrequency (RF) electrical currents are delivered to biologic tissues, the energy is converted into heat in the close vicinity of the active electrode where the density of current lines is highest. Heat is then transmitted to the remaining tissue by conduction and dissipated by convection [1]. The temperature reached at different depths of the biologic tissue is therefore dependent on the RF settings and on the type of the application. Most RF generators used in therapeutic monopolar ablation work under temperature control, ie, the temperature is measured on the tissue surface by sensors in the catheter, and power is delivered to the tissue until the temperature set by the user has been attained. Although the temperature at the surface is known and maintained around the set value throughout the ablation, the same does not necessarily apply to the temperature inside the tissue. Different forms of RF application determine the physical conditions at the tissue surface. In endocardial applications during cardioplegic arrest, the atria are Accepted for publication Dec 11, Address reprint requests to Dr Santiago, Instituto do Coração, Av Prof Reynaldo dos Santos 27, Carnaxide, Portugal; teresa.santiago@incor.pt. cooled down to 4 to 10 C and there are heat exchanges between the endocardial and epicardial surfaces and the atmosphere. Therefore, the physical conditions at the point of RF application are different from those deeper in the myocardium. Although it might appear that the temperature inside the tissue is lower than at its surface, this is not necessarily true along the entire tissue depth. Power is applied, up to a set temperature, on a surface that is being cooled through contact with the air, and electrical current is conducted into areas of the tissue that are not in contact with the air. This may result in higher values of temperature below the tissue surface than the temperature at its surface that was set by the user. Deeper in the tissue the temperature will decrease to values on the opposite surface that are likely to depend on the thickness of the atrial wall, on its constitution, and on the physical conditions at that surface. There are several publications on electrode and tissue temperature using in vitro and in vivo experimental models [2 5], finite element models [6, 7], and in vitro studies of intra-tissue temperature during RF applications with irrigated tip catheters [8], all these models simulating percutaneous ablation. However, there are no data relating intra-tissue temperatures with histologic 2003 by The Society of Thoracic Surgeons /03/$30.00 Published by Elsevier Science Inc PII S (02)

2 1496 SANTIAGO ET AL Ann Thorac Surg TISSUE TEMPERATURE DURING ABLATION 2003;75: assessment of the lesions induced by surgical endocardial RF applications on human tissues. We studied the dependence of the temperatures measured subendocardially and subepicardially on the different settings of RF endocardial applications, and on the thickness of the atrial wall using human atrial tissue. The lesions were histologically studied to measure lesion depth and to assess the occurrence of transmural lesions. Similar applications were performed in patients during mitral valve surgery, a biopsy was performed, and the specimen was histologically assessed. Results were compared with those from in vitro studies to evaluate the possible influence of underlying disease of the atrial wall in patients with mitral valve disease. Material and Methods Radiofrequency Endocardial Applications In Vitro The thickness of 17 human atrial fragments from eight organ donors (4 women and 4 men) aged 20 to 72 years (mean, years) was measured at three points in a line with a digitizer (Threespace [Polhemus, Vermont, CA]) with an error of 0.3 mm. Each atrial fragment had an area of approximately 30 cm 2. T-type thermocouples (0.10 mm thick) were inserted subendocardially and subepicardially at the above referred points and connected to a specially modified Meca APM switch box (EPT Technologies, San Jose, CA) for filtering out the RF interference and for signal processing. The box was connected to a personal computer for data acquisition and real-time graphics display of subendocardial and subepicardial temperatures. Radiofrequency currents were delivered to the tissue between the catheter placed on the endocardial surface and the dispersive electrode. To create some distance between the active and dispersive electrodes and obtain a reasonable simulation of the conditions found in endocardial ablation during surgery, some gauze padding soaked in saline solution was placed between the epicardium and the dispersive electrode. The saline solution has an electrical conductivity similar to that of biologic tissues and is intended to create a conductive medium to close the electrical circuit between the active and dispersive electrodes. We used a malleable Thermaline catheter connected to an RF generator [9] (EPT Technologies). The seven electrodes (each 12 mm long with a diameter of 2.4 mm) can be selected in any number and combination and fired simultaneously. Endocardial applications were performed at set temperatures of 70 C (n 9) and 80 C (n 8) for 2 minutes, by placing the catheter active electrode over the three points where the temperatures were being recorded. The generator output was connected to another personal computer, and the variations of power, impedance, and temperature of the tissue surface (measured by the sensors in the catheter) were displayed in real time by means of EPT graphics software. It was therefore possible to compare the variation of the temperature at the endocardial surface with that of the temperatures measured simultaneously by the thermocouples placed subendocardially and subepicardially at the points of the atrial wall where its thickness had previously been measured. Radiofrequency Endocardial Applications in Mitral Patients Radiofrequency applications were performed at a set temperature of 70 C in 36 patients with mitral valve disease who had concomitant atrial fibrillation to achieve bilateral isolation of the pulmonary veins as previously described by the authors [10, 11]. Briefly, the heart was arrested with cold cardioplegic solution, and the left atrium was opened parallel to the interatrial septum, in front of the right pulmonary veins, after a core temperature of 30 C had been reached. The initial anterior opening was performed with a surgical incision, and an application of RF at 70 C for 2 minutes, performed posteriorly, completed the encircling lesion around the orifices of the pulmonary veins. Fragments of tissue approximately cm were removed from the zone of ablation. Samples from 10 patients, all women aged 47 to 75 years (mean, years) were observed histologically. All the remaining patients were excluded from the study owing to technical reasons. Informed consent was obtained from all patients. Histopathologic Assessment The samples were fixed in 10% buffered formalin, and fragments were serially taken from the whole line of the sample (containing a fragment of the RF-induced lesion) in sections that were perpendicular to the line and included the whole thickness of the atrial wall. After paraffin embedding, 2- m cuts were stained with histochemical dyes: hematoxylin and eosin, Gomori s trichrome, and elastic van Gieson (Verhoeff). The sections were analyzed under light microscopy by 2 observers. A metric eyepiece (with a precision of 0.02 mm) was used to measure the thickness of the left atrial wall, its layers, and the lesions caused by RF. Results Radiofrequency Endocardial Applications In Vitro Figure 1A shows an example of the temperatures, measured subendocardially and subepicardially, during endocardial RF application at set temperatures of 70 Cin three points where the atrial wall was 5.1, 3.4, and 3.1 mm thick. The subendocardial temperatures at those points were 5 to 10 C higher than the set temperature, which remained fairly constant throughout the application, as shown in Figure 1B. We found that the subepicardial temperatures were higher when, at the beginning of the application, the interior of the tissue was warmer than its surface (grad T 0) than when all the tissue was at the same temperature (grad T 0). The first situation always occurred when an application was aborted as a result of the generator shutting down and a second one followed

3 Ann Thorac Surg SANTIAGO ET AL 2003;75: TISSUE TEMPERATURE DURING ABLATION Fig 1. Endocardial radiofrequency application at 70 C on human atrial myocardium in vitro. (A) Subendocardial (send) and subepicardial (sepi) temperatures measured at three different points of human atrial myocardium. The initial temperature overshoot is related to the response time of the generator. (B) Variation in time of power, impedance, and of the temperature measured on the endocardial surface by sensors in the catheter. (over the same area) as soon as the temperature at the tissue surface was low enough for the generator to deliver power. For this reason the subepicardial temperatures and the depth of the lesions induced by RF applications at set temperatures of 70 and 80 C are shown in Table 1, split between the cases in which grad T 0 and grad T 0 at the beginning of the application. Values are presented as mean standard deviation except for lesion depth, where mean average deviation is used because of the small sample size. After a steady state had been reached, the subendocardial temperatures were always higher than the subepicardial ones and reached a maximum value of 87 C in applications at 70 C, and of 97 C in applications at 80 C. Subendocardial temperatures rose above the set values in 13 of 17 cases in which readings were available. Histologic evaluation was performed on 12 samples Seven pertained to applications at 70 C and five to applications at 80 C. All lesions showed damage of the muscle fibers, thinning, and collapse of the endocardial elastic fibers, and some had loss of substance at the endocardium myocardium interface, possibly as a result of liquefaction necrosis of previous loose fibrous or adipose tissue. Damage of muscle fibers was characterized by cytoplasm homogenization with total loss of cross striations, nuclear hyperchromasia, pyknosis, and illdefined cell membrane (histologic features of coagulation necrosis). These histologic features are identical to those observed in patients with mitral valve disease after RF endocardial ablations (Fig 2). All seven lesions obtained at 70 C were transmural, reaching the endocardium, the myocardium, and the whole epicardium. The lesions measured 1.35 mm to 3.50 mm (mean, mm) and were induced on walls with thickness that varied from 2.5 to 6.4 mm (mean, mm). The temperatures reached subepicardially were always above 60 C, except in applications on two atrial fragments in which temperatures of C and C were registered. Histologic results were available in the first case and confirmed that the lesion was transmural. Three of the five samples pertaining to applications at 80 C showed transmural lesions 2.50 to 2.75 mm (mean, mm) deep that damaged the three atrial layers and were induced on walls 3.0 to 4.0 mm (mean, mm) thick. The two remaining lesions, measuring 2.37 and 1.63 mm, were not transmural, showing damage of the whole endocardium and of 50% and 43% of the myocardium, respectively. These lesions were induced on atrial walls 4.6 and 4.0 mm thick, and the temperatures measured subepicardially were C and C, respectively. In the remaining cases the subepicardial temperatures were over 70 C. The histologic features of the lesions induced at 80 C are similar to those induced at 70 C. Radiofrequency Endocardial Applications in Mitral Patients Histologically the lesions showed similar features to the in vitro ones, but in patients with mitral valve disease (Fig 2), the myocardial interstitium at the damaged area contained hemorrhagic foci and thrombosis of the small vessels (Fig 3). The total depth of the lesions varied from 0.25 to 2.00 mm (mean, mm) in walls 1.25 to 4.50 mm thick (mean, mm). Five of the 10 lesions assessed did not reach the myocardium and were confined to the endocardium, 3 lesions showed 22% to 77% of damaged myocardium, and 2 lesions observed in left atrial walls thinner than the average (1.25 mm and 1.70 mm) were transmural without signs of wall rupture. Table 2 shows the thickness of the endocardium and the whole atrial wall, as well as the depth of the lesion at the myocardial layer, the lesion total depth, and the percentage of damaged myocardium. At 6 months follow-up 2 of the 5 patients with no myocardial damage were in normal sinus rhythm. Of the 5 patients with myocardial damage, 4 were in sinus

4 1498 SANTIAGO ET AL Ann Thorac Surg TISSUE TEMPERATURE DURING ABLATION 2003;75: Table 1. Tissue Temperatures and Lesion Depth in Radiofrequency Applications In Vitro Set T ( C) Initial Tissue T ( C) Atrial Thickness (mm) a Subepic T ( C) Lesion Depth (mm) b Transmural Lesions /1 (Grad T 0) /2 (n 6) /1 35 (Grad T 0) /2 (n 3) / (Grad T 0) /1 (n 6) /5 35 (Grad T 0) (n 2) a Measured before the radiofrequency application. b Measured under microscopy during histologic evaluation. Grad T temperature gradient; Subepic subepicardial; T temperature. rhythm, and 1 (with 38% myocardial damage) remained in atrial fibrillation. Comment The average thickness of the normal left atrial wall in humans is approximately 3 mm, and the composition of the subendocardial layer at the endocardium myocardium interface varies between fibrous tissue (loose or dense) and adipose tissue (Fig 4, right). However, both the thickness and the proportion of the three layers (endocardium, myocardium, and epicardium) vary from person to person, depending not only on age but also on related diseases, namely rheumatic fever, diabetes mellitus, hypertension, and tumors. Histologically a lesion induced by RF ablation can be regarded as transmural if it shows damage of the whole myocardium. Because endocardium damage is inevitable in endocardial RF ablation, the endocardium acts as an obstacle to the transmission of RF energy into the myocardium in the sense that part of the RF energy is absorbed in this layer. In endocardial RF application the epicardium does not need to be damaged for a lesion to be transmural. Radiofrequency Applications In Vitro Our in vitro studies show that it is possible to obtain transmural lesions with endocardial applications at 70 and 80 C in atria as thick as 6.4 mm (trabeculated area). However, it appears that the application temperature and the intra-tissue temperatures are not the sole factors that determine the lesion depth. Of all the lesions histologically assessed, the only nontransmural ones occurred at 80 C and had a depth in the range of those obtained at 70 C. In both nontransmural lesions the subepicardial temperatures were greater than 60 C ( C and C). Conversely, we had an application at 70 C that produced a transmural lesion with subepicardial temperatures of only C. Fig 2. Left atrial wall from patients with mitral valve disease. (Left) Collapse of the thickened subendocardial (A) and interstitial (B) elastic fibers (Verhoeff, 40). (Right) Liquefaction necrosis and tissue loss (C) at the endocardium/myocardium interface (Gomori s trichrome, 40). Fig 3. Arrows show thrombosis of small vessels at the myocardium interstitium of a patient with mitral valve disease in an area damaged by radiofrequency application (Gomori s trichrome, 400).

5 Ann Thorac Surg SANTIAGO ET AL 2003;75: TISSUE TEMPERATURE DURING ABLATION Table 2. Lesions and Wall Depth in Atria of Patients With Mitral Valve Disease Endocardium (mm) Myocardial Lesion (mm) %MD Total Depth (mm) Lesion/Atrial Wall Age (y) / / / a / / / b / / / / a This lesion did not reach the whole endocardium. b There was loss of tissue in a depth of 0.25 mm, which cannot be identified as part of the endocardial or myocardial layer. %MD percentage of damaged myocardium. We believe that the subepicardial temperatures were correctly measured during the applications at 80 C that led to the two nontransmural lesions. Actually, the thermocouples were inserted at three different points approximately 5 mm apart (to measure the tissue temperature under the area ablated by the same electrode) to compensate for the inevitable imprecision in their insertion. The fact that temperatures of more than 60 C were attained at the subepicardial level, without causing a transmural lesion, raises the issue of whether 50 C asthe temperature threshold between viable and nonviable tissue [1, 2, 12] is true for all biologic tissues. All the referred studies were performed in healthy canine ventricular myocardium and rabbit skeletal muscle and not in human myocardial tissue. Our findings also show that when the first application was aborted (because of generator shut down) and a Fig 4. (Left) Very thick left atrial wall of a patient with mitral valve disease (Verhoeff, 40). (Right) Normal left atrial wall of a 64-yearold man. Note the adipose tissue (B) at the endocardium myocardium interface (Verhoeff, 40) second application followed over the same area as soon as the temperature on the application surface was low enough for the generator to deliver power, the subepicardial temperatures reached higher values than they did when the RF application was uninterrupted. The endocardial surface, being in contact with the air, cools down faster than the remaining tissue, giving rise to a temperature gradient between this surface and the interior of the tissue. When a second application started, the interior of the tissue was warmer than the surface and its temperature rose to values that were higher than when all the tissue was at the same temperature. Therefore, consecutive endocardial applications can raise the intratissue temperatures (over which the surgeon has no information) to values significantly higher than those of application, eventually risking tissue carbonization at set temperatures that seem perfectly safe. However, it should be noted that the in vitro experiments do not duplicate but are rather a simulation of the in vivo applications. In fact, the microcirculation, which acts as a cooling factor, was not reproduced in our in vitro model; therefore, the intra-tissue temperatures may not rise as high above the set temperature during consecutive endocardial applications in surgical practice. Radiofrequency Applications in Mitral Patients The thickness of the left atrial wall of patients with mitral valve disease usually ranges from 1.7 to 5.3 mm. In these patients, the endocardium is always thicker than in normal subjects owing to a higher content in elastic fibers, collagen, fat tissue, and eventual smooth muscle hyperplasia. The thickness of the myocardium is also highly variable as a result of myocyte hypertrophy and fibrosis or lipomatosis of the interstitium (Fig 4, left). The thickness of the epicardium depends on its fat content. Histologic evaluation showed that half of the lesions induced at 70 C in patients with mitral valve disease did not reach the myocardium and were confined to the endocardium. Although this might be expected in patients whose atrial walls had a very thick endocardium, Table 2 shows that there is not a direct relation between the two factors. Patients with thin endocardium showed no lesion of the myocardium whereas patients with thicker endocardium had lesions that damaged considerable portions of the myocardium. In other words, the endocardial thickness is not the sole determinant to the depth of the myocardial lesion. When lesions obtained in vitro are compared with the in vivo ones, limitations of the in vitro model, such as the lack of microcirculation and the smaller distance between the active and return electrodes, must be taken into account. However, it is unlikely that these alone will account for the large difference in the average depth of the lesions obtained in vitro at 70 C ( mm) versus the ones induced in patients with mitral valve disease ( mm). Moreover, when the size of the lesion in the myocardial layer (believed to relate to electrical block) is considered, the difference is even greater ( mm in vitro versus mm in patients with mitral valve disease). It is worth noting that

6 1500 SANTIAGO ET AL Ann Thorac Surg TISSUE TEMPERATURE DURING ABLATION 2003;75: an important difference between the in vitro model and the in vivo results is that the former used atrial tissue from organ donors without previous disease, other than age-related alterations, whereas the in vivo results came from patients with mitral valve disease who had diseased atrial walls, namely thickened endocardium. These facts suggest that the composition and the thickness of the endocardium will determine to what extent the endocardium will act as a barrier to the transmission of energy into the myocardium, particularly in patients with mitral valve disease who have pathologic endocardium. Similarly the composition of the myocardium is likely to influence the formation of the lesion. From a biophysical point of view, it is likely that atrial tissues with different compositions will have different electrical properties, which in turn will affect the conduction of RF current in the tissue. When a biologic tissue is in the presence of an electrical field, a current (resulting from the movement of mobile ions within the aqueous biologic medium) will develop. This current is therefore related to the ionic content and ionic mobility of the particular tissue, which is expressed in terms of the tissue electrical conductivity. This measurement varies widely between biologic tissues and with the frequency of the applied field, and it has been shown to change with disease [13]. It is presumed that the primary mechanism of tissue injury caused by RF currents is thermal [1], and studies in guinea pigs have shown that hyperthermia causes significant changes in myocardial cellular electrophysiologic properties [14]. However, it is possible and has been suggested [2, 12] that besides an increase in temperature, the actual conduction of current through the tissue will cause alterations at the cellular level that are more difficult to quantify than the actual temperature increase and may influence the formation of the lesion. Moreover, the ischemia caused by the thrombosis of the microcirculation may be another mechanism of tissue injury. The presence and location of microcirculation thrombosis may explain why acute histologic examination does not necessarily correlate exactly with chronic histologic examination (R. Gouveia and associates, unpublished data). Our findings suggest that the thickness and the composition of the endocardium and of the myocardium play an important role in the formation of the myocardial lesion with endocardial RF applications. This may account for the large variability in the clinical results reported by different groups [15 17]. Because the histologic content varies from patient to patient, it is logical to assume that this feature is one of the reasons for such variability. The fact that 2 patients whose RF lesions were confined to the endocardium were in sinus rhythm at 6 months, whereas another patient with 38% damage of the myocardium remained in atrial fibrillation, suggests that a transmural lesion may not always be required for good clinical results. Conversely, previous results from our group showed that patients in sinus rhythm after surgery had evidence of electrical exclusion of the pulmonary veins [18, 19]. Nevertheless, the development of a definitive treatment of atrial fibrillation requires that the ideal ablation lines be defined. Until transmurality is achieved a scientific comparison between surgical techniques will remain a clinical challenge. This study was supported by Fundação para a Ciência e Tecnologia, Lisbon, Portugal. 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