Arterial Wall Temperature Following Coronary Stent Implantation in Pigs: The Role of Post-Stent Inflammation

Arterial endoprostheses (stents) are widely used to improve angioplasty outcome and successfully restore the reduced vascular lumen. However, the metallic nature of stents causes some degree of arterial wall inflammation. This inflammation is known from ex vivo studies,^1,2 and has been cited as a possible contributory cause in restenosis following stent implantation^3–5 as well as for increased local thrombogenicity.^6–8 Yet, it has not been studied in vivo. Such an in vivo study of post-stent inflammation would yield important information regarding the establishment and duration of this phenomenon, helping us to understand it better. Thermography of the arterial wall is a new method to assess wall tissue inflammation. Activated macrophages are the main cause of heat production in the inflamed wall areas.^1,9,10 The increased metabolism of the macrophages and the strong lytic activity at the inflammation site lead to augmented heat production that can be detected in vivo using thermography catheters. The correlation of an increased vascular wall temperature with macrophage activity has been previously demonstrated in vitro and in vivo.^1,9 Methods Study population. Twenty non-atherosclerotic “domestic crossbred pigs” (Sus Scrofa) of both sexes were used in these experiments. All animals were treated and cared for in accordance with the National Institute of Health guide for the care and use of laboratory animals. The domestic pigs (weight, 30–35 kg) were fed on a standard natural grain diet without lipid or cholesterol supplements. Pigs were divided into group A and group B, each consisting of 10 pigs. Each pig underwent coronary catheterization and stenting of the proximal right coronary artery (RCA). The temperature of the arterial wall was mapped in the proximal RCA before and immediately after stent implantation. Five days later, catheterization was performed again in the pigs of group A and the temperature was mapped in the stented and adjacent areas. Each animal was then sacrificed and the proximal RCA was harvested for light and electron microscopy as well as for histology. Eight days after stent implantation, catheterization was also performed in the pigs of group B and temperature was mapped in the stented and adjacent areas. As with group A, sacrifice and histological analyses followed the temperature study. Catheterization and stenting. For coronary catheterization and stenting, standard interventional procedures were used. All animals received a Freedom stent (Global Therapeutics, Inc.) with a diameter of 3.0 mm and a length of 18 mm. The vascular area to be stented was selected in the RCA at least 10 mm further from main bifurcations. The stent was inflated using a pressure of 12 atmospheres. Except for 250 mg acetyl salicylic acid, no other anti-inflammatory drugs were administered to the animals. Due to the effect of intravascular injection of contrast on the temperature measurements, all contrast injections were performed at least 3 minutes before any measurement, allowing the restoration of arterial temperature to normal levels. Arterial wall temperature measurements. To study the arterial wall temperature, we used the ThermoSense™ vascular thermographic system (Thermocore Medical Limited, United Kingdom). The system consists of a thermography catheter, a pullback device and an electronic console. The thermography catheter (Figures 1 and 2) is a 4 French (Fr) over-the-wire intravascular catheter that has a body length of 140 cm. The distal tip has 4 independent resiliently biased arms made of a superelastic material. At the end of each arm there is an accurate thermistor temperature microsensor. Each microsensor has an accuracy of 0.01 °C and a time constant of 150 ms. The arms can be held parallel to the catheter body covered by a sliding incorporated sleeve. When the sleeve is retracted (Figure 2), the arms are released to open to the maximum possible diameter, bringing the sensors in close contact with the vascular wall. The catheter can be used in 2 configurations: open and closed. The closed configuration (lowest possible profile) is used during the catheter insertion in the coronary artery, using standard interventional techniques, while the open configuration is used for the temperature study. The proximal part of the catheter is appropriately shaped to fit in a motorized pullback device, operating at adjustable speeds between 0.1 and 1.0 mm/s. We scanned the arterial wall with a pullback speed of 0.3 mm/s. All thermographic data collected from the 4 sensors during the automated pullback were sent to the computer-based console for analysis and storage. Although, due to its 4-sensor configuration, the thermography system provides 4 independent temperature measurements (at angles of 0°, 90°, 180° and 270°) at the circumference of the same arterial site, in this study we used the mean of the 4 temperature values per site. To verify the proper in vivo operation of the sensors, an injection of 10 cc NaCl 22 °C in the ostium of the RCA (Figure 3D) was performed. The reproducibility of the ThermoSense system has been tested and verified in vitro in externally heated coronary models with simulated blood flow (Cambridge Consultants Ltd. Laboratories, Science Park, Cambridge, United Kingdom). In addition, we verified the reproducibility of a thermal scan in vivo by repeating each scan. The console software automatically overlapped the 2 derived curves (temperature versus arterial length). Two thermal scannings were considered inconsistent if the variation between them was > 0.05 °C for the same arterial spot. In case of inconsistency, a third scan was performed in the area under question. Histology. The whole stented area plus 10 mm of area adjacent to the stent were harvested for histology. Every stent filament site was carefully examined for inflammatory cells, and inflammatory reaction was scored as follows: 0 = very rare or no appearance of histolymphocytes; 1 = sparsely located histolymphocytes around the stent filament; 2 = more densely located histolymphocytes covering the stent filament, but no lymphogranuloma and/or giant cell formations found; and 3 = diffusely located histolymphocytes, lymphogranuloma and/or giant cells, also invading the media. Electron microscopy was also used to evaluate the mechanical effect of the thermography catheter on the arterial wall, and it was performed on samples from the stented area, as well as from the proximal and distal neighboring areas. Procedure. Each pig was sedated with intramuscular azaperone 3 mg/kg (Stresnil, Janssen Pharmaceutics). General anesthesia was induced with intravenous ketamine (2 mg/kg, Ketalar, Parke Davis) and further intravenous bolus Diprivan (3.5 mg/kg, AstraZeneca SA). Intravenous infusion of Diprivan followed at a rate of 3 mg/kg/h. The pig was then intubated and ventilation (Mark 7A, Bird Cooperation, California) was started using a mixture of 20% volume of pure oxygen and 80% volume of room air. Ventilation was adjusted by frequent blood gas analyses in order to maintain a minimum PaO2 of 100 mmHg and physiologic PaCO2 and pH parameters. Continuous electrocardiographic, pressure and body-temperature monitoring was performed throughout the procedure. A carotid artery was surgically exposed and an 8 Fr arterial sheath was introduced over a 0.035´´ guidewire. Heparin 5,000 IU and 250 mg acetyl salicylic acid were administered intravenously as a bolus. Furthermore, 400 IU/h heparin were given as a continuous infusion during the procedure. An Amplatz 8 Fr guiding catheter was used to cannulate the ostium of the RCA. A Balance Middle Weight guidewire (Guidant Corporation, Inc., Temecula, California) was introduced in the RCA, until the distal segment. The thermographic catheter was then placed over the wire until the sensor markers were 60 mm from the coronary ostium. For correct positioning of the catheter, we used fluoroscopy and quantitative coronary angiography (QCA). The sensor sleeve was retracted, leading to sensor-arm opening and contact between the 4 sensors and the arterial wall. Automatic motorized pullback was immediately started, with temperature recordings from all sensors. The catheter was pulled back 60 mm with a speed of 0.3 mm/s. After saving the data on the console, the sensor arms were again closed using the sleeve and the catheter was repositioned at the beginning of the area of interest for a second pullback; this repetition was used to validate the reproducibility of our temperature measurements. The thermography catheter was then removed, and a 3.0 x 18 mm Freedom stent (Global Therapeutics Inc.) was expanded in the proximal RCA using a pressure of 12 atmospheres. The stent was placed with its distal end approximately 40 mm from the coronary ostium. Fluoroscopy and QCA were also used to guide stent implantation. Immediately after implantation, the thermographic catheter was repositioned and the thermography procedure was repeated. At the end of the procedure, pigs were randomized to group A or B. Following the 5- (group A) or 8-day (group B) waiting period, a second catheterization was performed and the pigs were thermally scanned and finally sacrificed. Statistical analyses. Statistical analyses were performed using the SAS statistical software (SAS version 8.2, SAS Institute). Data are presented as means ± the standard error of mean. The significance of the temperature difference was assessed with the repeated ANOVA test. Linear regression analyses were used to examine the relationship between inflammation score and temperature increment. The unpaired student’s t-test was used to compare any pair of mean group values. A p-value 3–5 and on acute or subacute stent thrombosis still have to be determined. Intravascular thermography seems to be a useful tool to spot and evaluate the degree of arterial wall inflammation. This knowledge can eventually help us improve intracoronary devices and decide on the best delivery approach. In the real clinical word, there are important cases where intravascular thermography could be useful. Increased macrophage infiltration and activity at the site of an atherosclerotic plaque has been correlated with the degree of the plaque stability.^7,8,11 Therefore, thermography might be a useful means of spotting dangerous vulnerable plaques. Since cardiac catherization of the patient is required for the application of such a diagnostic technique, the population that might benefit from it lays initially in the patients who already have clinical symptoms. Information on the form of the wall inflammation (localized or diffuse) and the degree of the thermal gradient inside the artery might help us understand and better treat the individual patient. One point of discussion is the slight drop of temperature in the stented area immediately after stent implantation. We were surprised to see this effect, which could be repeated in every animal. One possible explanation could be the fact that the stent, as a metal object, has a high specific thermity and therefore delays equalization with the temperature of the surrounding tissues. One could argue that the metallic mass of the stent is minimal and, being dipped in the relatively high volume of circulating blood, should immediately acquire its temperature. However, the driving force that tends to equalize temperature differences between blood and the stent is not stable; it degrades rapidly as the temperature of the stent rises, and therefore temperature equilibrium might be delayed for some time. Other explanations could involve the role of the stent-delivering balloon inflation (which is filled with cooler contrast). In order to investigate this issue further, we re-measured the temperature 30 minutes after the first measurement and found that initial differences were reduced to less than half. Another point of discussion is the effect of blood flow on the temperature measurements. Blood present in the artery has an equalizing effect on the temperature of the arterial wall. Like a cooling liquid, blood removes heat from the warmer wall areas and heats up areas with lower heat production. Our experiments in other animal models with induced flow disturbance showed a reproducible effect on the temperature of the arterial wall. However, the animal model in the current study includes only non-atherosclerotic pigs; therefore, blood flow was never disturbed during the temperature measurements. It is reasonable to assume that different stent types would cause different degrees of arterial wall inflammation. The strut design and the size of metallic surface coming in contact with the arterial wall play a role in causing an increased macrophage response. The same applies for different stent sizes. The inflation pressure for the stent delivery can also be linked to the degree of induced inflammation; stent coating and individual stent properties (drug elution, emitted radiation, etc.) that are found in some new stents might also alter the degree of wall inflammation. However, these parameters were outside the scope of the current study. We used 1 specific type of stent with the same dimensions in all animals and the inflation pressure was kept at 12 atmospheres. Temperature correlation between different animals for the purpose of quantification of temperature differences does not seem to offer any additional benefit. The background blood temperature of each animal differs according to its size, age and metabolic status. All the animals had a background temperature ranging between 36.1–37.2 °C, but still differed between them. For this reason, we considered temperature differences between different spots in the same coronary artery. The scanning speed of the thermography catheter pullback device is another issue of interest. The specific thermography catheter has a time constant of 150 ms, which means that it needs at least 150 ms contact time with a point to completely acquire its temperature. Bearing in mind that the sensor contact area is 0.25 mm2 and the shape of the contact is approximately circular, the diameter of the contact area is 0.56 mm. As a consequence, since the catheter must remain at least 150 ms on each point, in order to avoid undersampling and loss of data, the maximum pullback speed should be 12–14 however, these risks are minimized by following proper handling and operating procedures. Study limitations. Although the study addresses wall temperature and inflammation at 5 and 8 days, it does not give information on the exact time when temperature differences make their first appearance. In order to obtain this information, daily measurements of arterial wall temperature or more animal groups are required. The study showed the effect of stenting on the wall temperature in normal coronary arteries. However, the temperature response in atherosclerotic arteries, where temperature differences might already be present, still remains unknown. Unstable arterial plaques already have some degree of inflammation due to the activated macrophages.¹¹ In cases of stenting, it would be interesting to see the effect on plaque stability.¹⁵ As mentioned, the study is limited to a specific type of stent and balloon injury (12 atmospheres). The device and technique parameters (size of stent, inflation pressure, etc.) that affect the induced inflammation were not investigated. Conclusion This study proves that the temperature homogeneity of a normal coronary artery is temporarily disturbed after stent implantation. This disturbance is possibly due to the inflammation caused by the stent and its accompanying balloon injury. This wall inflammation plays a major role in the increased cell proliferation and early restenosis^3–5 as well as in local thrombotic activity. Due to its sensitivity and reproducibility, arterial wall thermography might be a useful tool to measure and quantify the extent of stent-induced inflammation in vivo. Further studies using arterial thermography are expected to evaluate and quantify the amount of inflammation caused by several intravascular techniques and devices. Acknowledgments: The authors would like to thank Thermocore Medical Limited for providing the thermography device and catheters used in the study.

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