DOI:10.2214/AJR.06.0810
AJR 2007; 188:1028-1032
© American Roentgen Ray Society
Differences in Ablation Size in Porcine Kidney, Liver, and Lung After Cryoablation Using the Same Ablation Protocol
Sompol Permpongkosol1,2,
Theresa L. Nicol3,
Richard E. Link1,4,
Ioannis Varkarakis1,
Hema Khurana5,
Qihui Jim Zhai5,
Louis R. Kavoussi1,6 and
Stephen B. Solomon1,7
1 James Buchanan Brady Urological Institute, Johns Hopkins University School of
Medicine, Johns Hopkins Hospital, Baltimore, MD.
2 Present address: Department of Surgery, Faculty of Medicine, Ramathibodi
Hospital, Mahidol University, Bangkok, Thailand.
3 Department of Pathology, Johns Hopkins Bayview Medical Center, Baltimore,
MD.
4 Present address: Scott Department of Urology, Baylor College of Medicine,
Houston, TX.
5 Department of Pathology, The Methodist Hospital, Houston, TX.
6 Present address: Institute for Urology, North Shore LIJ Health System, Long
Island, New York, NY.
7 Present address: Department of Radiology, Memorial Sloan-Kettering Cancer
Center, 1275 York Ave., New York, NY.
Received June 19, 2006;
accepted after revision September 12, 2006.
Address correspondence to S.B. Solomon
(SolomonS{at}mskcc.org).
Abstract
OBJECTIVE. The purpose of our study was to assess the variation in
size of acute necrosis and the variation in thermal map measured during
cryoablation in multiple organs using the same ablation protocol for each
organ.
MATERIAL AND METHODS. Eight female pigs underwent one cryoablation
per organ of kidney, lung, and liver performed with open surgery with a 2.4-mm
cryoprobe. A 12- and 8-minute double-freeze cycle was used. Intratissue
temperatures were monitored using 16-gauge thermometers spaced at 5.0-mm
increments from the cryoprobe. The comparison of results among tissues was
performed using the multiple analysis of variance. The -20°C thermal
diameter was correlated with tissue damage. The kidneys, lungs, and liver were
removed and examined histologically for a pathologic complete coagulative
necrosis zone.
RESULT. A single 2.4-mm cryoprobe had a mean ice ball diameter in
kidney, lung, and liver of 38.5 ± 4.7, 35.5 ± 3.6, and 32.5
± 2.7 mm, respectively. A mean -20°C thermal diameter was achieved
at 24.07 ± 1.38 mm in kidney, 12.76 ± 3.0 mm in lung, and 8.8
± 3.7 mm in liver by means of regression analysis. The acute pathologic
complete coagulative necrosis zone size was 21.0 ± 1.56 mm (kidney),
11.6 ± 1.48 mm (lung), and 8.0 ± 1.20 mm (liver).
CONCLUSION. The inherent characteristics of different organs
manifest different ablation zone sizes during cryoablation despite the same
ablation protocol being used. This information should be factored into
planning for ablation procedures.
Keywords: ablation • coagulative necrosis • cryoablation • kidney • liver • lung • porcine studies
Introduction
Cryoablation is a promising technique for causing localized tissue
coagulative necrosis that has been applied clinically to various tissues
including liver, kidney, uterus, prostate, and lung
[1-3].
However, concerns regarding the effectiveness of this technique to kill tumor
cells in situ still exist. One of the challenges during cryoablation is the
need to space individual cryoprobes in a pattern that will guarantee
adequately low temperatures for coagulation in all areas of viable tumor. How
widely these probes must be spaced depends on the relationship of temperature
to distance from the cryoprobe. This relationship will differ in various
tissues on the basis of thermal conductivity and tissue characteristics,
including differences in freezing point or the temperature sink effect
[4-6].
Gage and Baust [4] reviewed the
mechanism of tissue injury in cryotherapy and suggested that tissue thermal
conductivity was seldom a factor in most solid organs because of the high
water content of tissue. However, the normal lung containing air has a lower
thermal conductivity, which may affect the ultimate necrosis zone size in this
particular organ [7,
8]. Appropriate spacing of
cryoprobes is critical to achieve the low temperatures (-20°C) necessary
to kill cells consistently [9].
Another potential limitation to thermal ablation results from the heat (or
cold) sink effect, in which nearby blood vessels deliver 37°C blood flow
to the area being heated or cooled
[10]. Therefore, during
cryotherapy, the size of the ablation zone, and consequently, the diameter of
coagulative necrosis, may vary from tissue to tissue depending on the inherent
characteristics of the various tissues
[11].
In this study, we sought to correlate the acute coagulative necrosis size
in different organs with the thermal map around a single cryoprobe to provide
rational guidance of ablation procedures in clinical practice today.
Materials and Methods
Animals
The animal protocol was approved by the institutional animal care and use
committee. Eight female pigs weighing 50-60 kg were anesthetized with 22 mg/kg
of ketamine (intramuscular) and 5-8 mg/kg of thiopental (Sodium Pentothol,
Abbott Laboratories) (IV) and then underwent general endotracheal anesthesia
with 2% isoflurane. At the conclusion of the procedure, euthanasia was
performed with an overdose of sodium pentobarbital. The professional
veterinarians and trained personnel provided comprehensive care for the
animals throughout the study period.
Cryotherapy
A small plastic guide was made with holes to accommodate a single 2.4-mm
cryoprobe (PERC-24, Endocare) assembled with five 16-gauge thermometer probes
(TempProbe, Endocare). The temperature probes were placed at sequentially
greater distances from the cryoprobe so that temperature recordings could be
made at 5, 10, 15, 20, and 25 mm from an argon-based cryoprobe
(Fig. 1). The guide was used to
keep the thermometers and cryoablation probe in position throughout the
procedure.
First, a midline laparotomy was performed, enabling access to both kidneys
and the liver. The intestine was covered with dry abdominal pads to prevent
freezing damage. The cryoablation probe and thermometers were inserted
vertically to the same depth (20 mm) under visual guidance. Positioning of the
thermometer probes was confirmed with sonography (Ultramark 9, ATL)
(Fig. 2). During cryoablation,
the leading edge of the ice ball zone can be identified as a hyperechoic
interface with posterior acoustic shadowing.
After the kidney ablations, cryoablation was performed at the left lobe of
the liver. Probes were placed far enough from the anterior liver edge to have
a sufficient thickness of parenchyma, but not too close to the hepatic hilum
or to the vena cava, in order to avoid major vascular or biliary injury. In
one pig enough room remained to accommodate two independent liver
cryoablations. The thermometer probes were fixed into the liver at precise
distances from the cryoprobe.
For the lung, a small, right anterior thoracotomy was performed in the
fifth intercostal space after complete ablation of kidney and liver. The
lingual segment of the left upper lung lobe was exposed. Special care was
taken to avoid compression of the pulmonary parenchyma and interference with
the blood flow to the surface of the lung in the region to be frozen, similar
to the procedure in the prior report of Rodgers et al.
[12]. Again, the cryoablation
probe and thermometers were inserted to the same depth (20 mm) under visual
guidance. Sonography was not used with the lung.
Cryoablation was performed with two freezing/thawing cycles. An initial
freezing was performed for 12 minutes with argon gas followed by an active
thawing with helium gas. This was then repeated with an 8-minute freeze and an
active thaw. This protocol is similar to that used for cryoablation in human
subjects of kidney cryotherapy
[13]. The temperature of the
cryoprobe was checked before and during ablation to ensure effective
cryoablation. The diameter of the ice ball was recorded by direct
visualization. Temperature measurements from each of the thermometer probes
were recorded at 0, 6, and 12 minutes for the initial freeze and at 4 and 8
minutes for the second freeze. After the probes were removed, the animals were
sacrificed, and the ablated kidney, liver, and lung were harvested
immediately. One animal experiment required approximately 3 hours to
complete.
Pathology
Before histologic processing, the organs were fixed in 10% buffered
formalin. After fixation, the cryoprobe insertion site served as the central
axis of dissection, the samples were serially sectioned, and the zone of
ablation was identified grossly. Ablation necrosis measurements were made from
the mounted 5-µm microscopic slides stained with H and E. Slides were
examined by pathologists for coagulative necrosis, hemorrhage, interface
width, and surrounding tissue damage. The zones of coagulative necrosis were
histologically mapped with a calibrated microscope (x100 magnification).
The radii of the complete necrosis zones were measured perpendicular to the
cryoprobe axis, as previously reported
[14].
Statistical Analysis and Terminology
Statistical analysis was performed with commercially available software.
Regression analysis was performed to obtain the -20°C thermal diameter.
The comparison of results among tissues was performed using the multiple
analysis of variance. We reported our studies using the standardization of
terminology and reporting criteria of imaging-guided tumor ablation provided
by the Society of Interventional Radiology
[15].
Results
Reproducible relationships between temperature and distance from the
cryoprobe were measured. Eight kidney, eight liver, and eight lung lesions
were evaluated. The mean lowest temperatures of the cryoprobe in kidney, lung,
and liver were -134.13°C ± 1.5°C, -134.88°C ±
1.13°C, and -133.13°C ± 1.96°C, respectively. No
significant difference was seen among the probes according to multiple
analysis of variance tests (p = 1.0 in kidney and lung, p =
0.66 in kidney and liver, p = 0.11 in liver and lung). The mean
lowest temperature measurements made at 5.0-mm increments from the cryoprobe
at the end of the second freeze cycle are displayed in
Table 1. Figure
3A,
3B,
3C shows extrapolated
temperature relationships with distance over the entire cryoablation procedure
in porcine kidney, lung, and liver. The correlation coefficients between the
lowest temperature and distance from the cryoprobe in kidney, lung, and liver
were 0.95, 0.92, and 0.90, respectively.
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TABLE 1: Mean Lowest Temperatures at Different Distances from Cryoprobe Center
and in Different Tissues at End of Second Freeze
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Fig. 3A —Temperatures during cryotherapy. Graphs show mean temperatures at
different time points during cryotherapy measured at varying distances from
cryoprobe in porcine kidney (A), lung (B), and liver (C)
during entire double freezing cycle. Thermometer probes were inserted at 5,
10, 15, 20, and 25 mm from axis of cryoprobes.
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Fig. 3B —Temperatures during cryotherapy. Graphs show mean temperatures at
different time points during cryotherapy measured at varying distances from
cryoprobe in porcine kidney (A), lung (B), and liver (C)
during entire double freezing cycle. Thermometer probes were inserted at 5,
10, 15, 20, and 25 mm from axis of cryoprobes.
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Fig. 3C —Temperatures during cryotherapy. Graphs show mean temperatures at
different time points during cryotherapy measured at varying distances from
cryoprobe in porcine kidney (A), lung (B), and liver (C)
during entire double freezing cycle. Thermometer probes were inserted at 5,
10, 15, 20, and 25 mm from axis of cryoprobes.
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With the use of repeated analysis of variance tests, the lowest temperature
in kidneys was significantly lower than those in ablated lung (p <
0.001) and liver (p = 0.005) at the same distance. Although the
temperature of lung decreased more than that of liver at the same distance
from the cryoprobe, no significant difference was seen in comparison analysis.
Using regression analysis, the temperature of -20°C was achieved at a mean
diameter of 24.07 ± 1.38, 12.76 ± 3.0, and 8.8 ± 3.7 mm
in kidney, lung, and liver, respectively.
The diameters of the ice ball were 38.5 ± 4.7, 35.5 ± 3.6,
and 32.5 ± 2.7 mm in kidney, lung, and liver, respectively. The mean
diameter of the histologic complete necrotic zone seen was 21.0 ± 1.56
mm (kidney), 11.6 ± 1.48 mm (lung), and 8.0 ± 1.2 mm (liver).
Table 2 compares the pathologic
ablation zone diameters in each organ with the -20°C temperatures.
Microscopic examination with H and E revealed coagulation necrosis with no
viable parenchyma in the ablated region (Fig.
4A,
4B,
4C). Cytoplasmic and
architectural details were destroyed in ablated foci. Coagulative necrosis was
present with resultant congestion.
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TABLE 2: Comparison of the Diameter of Ice Ball, –20°C Thermal
Diameter, and Diameter of Coagulative Necrosis in Different Tissues Created
with 2.4-mm Cryoprobe Using the Same Protocol
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Fig. 4A —Photomicrographs of ablated tissue. In kidney, clear demarcation is
seen between viable (R) and infarcted kidney (L). Area indicated by R shows
viable tubules and mild intervening interstitial fibrosis. Area indicated by L
shows tubules lined by nucleated, hypereosinophilic epithelium and desquamated
necrotic cells and debris in lumen.
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Fig. 4B —Photomicrographs of ablated tissue. In lung, right side of
photomicrograph displays normal lung architecture that has clear demarcation
in center. Left side shows infracted lung parenchyma, congested alveolar
capillaries, and compressed alveoli (focal) filled with amorphous pink
material.
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Fig. 4C —Photomicrographs of ablated tissue. In liver, area of infarction
(lower left field) with poorly stained and mummified hepatocytes and
occasional lysed nuclei is seen. Scattered congested sinusoids are also noted.
Hexagonal liver architecture is retained.
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Discussion
The clinical success of thermal ablation for hepatic tumors has led to the
application of this approach to tumors in other organs, including kidney and
lung [16]. However, each of
these organs has its own specific thermal characteristics. Protocols that have
been acceptable in the liver
[17], for instance, may not be
ideal for the kidney or lung. One of the specific differences between organs
is the degree and pattern of perfusion. A limitation to thermal ablation is
the thermal sink effect, in which normal blood flow to an organ serves to
limit the heating or cooling of the ablation probe
[18-21].
However, other tissue-specific factors—most notably thermal conductivity
and tissue composition, including differences in freezing point—might
play a role in the cryoablation effect in different organs
[4,
5]. For instance, the lung,
containing air, has a low thermal conductivity
[4], and air movement may
provide an additional thermal sink. For this reason, tissue-specific protocols
are needed to ensure adequate killing of tumor cells during thermal
ablation.
In our study, the same cryotherapy protocol applied to liver, lung, and
kidney showed varying temperature gradients that are most likely related to
these organ-specific differences. The liver showed the smallest ablation zone,
-20°C thermal diameter, and an acute pathologic coagulative necrosis zone,
whereas the kidney had the largest ablation of the tested organs. The
implications of these findings are that cryoablation in the liver may require
longer freeze cycles or more densely clustered cryoprobes for consistent
success. Operators should recognize that the expected size of tissue
coagulation in the liver for a given probe size may be smaller than that
observed in another organ.
One limitation of this study is that normal healthy porcine tissues were
ablated. The ablation efficacy observed in tumor tissue might differ because
of increased vascularity, increased cell density, or other alterations in
tissue composition as compared with healthy controls. In addition, the
procedures documented here were performed through a laparotomy, thereby
exposing the tissues to room temperature. Cryoablation performed
laparoscopically or percutaneously might result in slightly different
temperature relationships than those observed here because of the maintenance
of surrounding body temperature at 37°C. Last, thermocouples in this
experiment were placed linearly, whereas the cryoprobes create an oblong ice
ball that varies in diameter at different depths. Therefore, the thermal map
lines derived in these experiments may not have been taken at the ice balls'
greatest diameters.
In this study, direct visualization of the ice ball proved to be an
unreliable predictor of effectively low temperature during cryoablation. For
example, the diameter of tissue with a temperature of 
-20°C and the
acute pathologic coagulative necrosis zone was approximately 2 cm in the
kidney, whereas the visible ice ball extended to > 3.5 cm. Clinically, ice
ball growth is often monitored by sonography, CT, or MRI. However, operators
must recognize that the ice ball edge does not represent the coagulation
necrosis zone and that this coagulation zone may be significantly smaller than
the ablation zone, similar to revious reports
[9]. Although some suggest that
the mechanism and ultimate size of the coagulation may be also due to other
nonthermal mechanisms such as ischemia
[19,
21], the implications of our
work suggest that on the basis of temperature alone, expected zones of
coagulation would be smaller than the visualized ice ball seen by the
physician.
Interestingly, unpublished Endocare data (John Rewcastle, personal
communication) show that a single 2.4-mm cryoprobe would have a 28-mm thermal
diameter at -20°C in gelatin phantoms after 10 minutes of cryotherapy.
However, these phantom results do not appear to be applicable to tissue with
blood perfusion, because our data revealed a -20°C thermal diameter of
approximately 24.07 ± 1.38 mm in kidney, 12.76 ± 3.0 mm in lung,
and 8.8 ± 3.7 mm in liver. Our results confirm those of a prior study
by Schmidlin et al. [22] that
showed a -22°C freeze/thaw ablation zone with a 3.4-mm cryoprobe in renal
tissue had a roughly 20-mm diameter
[22].
Prior data have suggested that cooling tissue to -20°C during
cryotherapy leads to reliable coagulation, and our data confirmed these
findings in the acute setting. Acute pathologic analysis revealed coagulative
necrosis zones close to the -20°C isotherm. In the chronic setting, these
necrosis zones may be still larger because of nonthermal events such as
ischemia. These observations are similar to those of Rupp et al.
[14]. However, Seifert et al.
[23] have suggested that
-38°C is the effective temperature for destruction of colorectal liver
metastasis. This discrepancy may represent a difference between normal and
cancerous tissue and warrants additional investigation.
Moreover, although our study used placement of thermocouples to make
temperature measurements, a number of potential noninvasive methods for
thermal mapping might be more practical in clinical practice. Our study data
can serve as guidelines for ablation, but ideally an online system of thermal
monitoring would be preferred. Such a system would allow adjustment of the
ablation during the procedure. The uses of noninvasive thermal mapping with
MRI, CT, and sonography have all been investigated and may provide a practical
solution.
In conclusion, our study highlights the differences of the cryoablation
zone size in different organs. Although our data may be specific to
cryoablation, the general concept that an organ-specific ablation protocol is
necessary is probably a valid one for all thermal ablation energy sources.
Compared with in vitro experiments, these in vivo results document
significantly smaller-than-expected zones of necrosis. Moreover, potentially
clinically significant differences were observed, with liver having much
smaller zones of coagulative necrosis than kidney or lung. The implications of
these findings are that operators should modify their ablation protocols to
reflect the presence of the variability of this effect in different organs
during cryoablation.
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Abstract
Introduction
