ORIGINAL ARTICLE Uterine damage in swine following uterine artery embolization: comparison among gelatin sponge particles and two concentrations of N-butyl cyanoacrylate Tetsuo Sonomura • Nobuyuki Kawai • Akira Ikoma • Hiroki Minamiguchi • Takashi Ozaki • Kazushi Kishi • Hiroki Sanda • Kohei Nakata • Motoki Nakai • Yasuteru Muragaki • Morio Sato Received: 4 March 2013 / Accepted: 13 August 2013 / Published online: 25 August 2013 � Japan Radiological Society 2013 Abstract Purpose To compare the degree of uterine damage caused by uterine artery embolization (UAE) with gelatin sponge particles (GSPs) and N-butyl cyanoacrylate (NBCA) in swine. Materials and methods Fifteen swine were divided into three groups of five according to embolic material: group A (1-mm GSPs), group B (NBCA:Lipiodol = 1:1), and group C (NBCA:Lipiodol = 1:7). The uterine arteries were completely occluded bilaterally. The uteri were removed 3 days after embolization, and radiographs of the removed specimens were obtained in groups B and C to evaluate the distribution of the NBCA. The macroscopic necrosis rates of the uteri were calculated, and the uteri were evaluated histologically. Results Uterine necrosis rates were 4.9 ± 6.1, 1.3 ± 3.3, and 41.4 ± 28.8 % in groups A, B, and C, respectively, and were significantly higher in group C than in groups A (p = 0.0014) and B (p \ 0.001). Uterine necroses were found in all 9 of the uteri with distal distributions of NBCA, and in only 1 of the 11 uteri with proximal distributions of NBCA. Conclusions Dilute NBCA caused more damage to the uteri than GSPs and concentrated NBCA did. Distal embolization using NBCA caused large necroses. There- fore, proximal UAE using concentrated NBCA should be considered in clinical situations. Keywords Uterine artery embolization � Gelatin sponge particle � N-butyl cyanoacrylate � Swine Introduction Obstetric hemorrhage can be life-threatening. It has a high mortality rate, with massive hemorrhage commonly leading to a coagulopathic condition [1]. Emergency peripartum hysterectomy is associated with a high incidence of maternal morbidity and a fatality rate of 4 % [2]. Uterine artery embolization (UAE) is an effective treatment for obstetric hemorrhage because it is minimally invasive, has a high success rate, and preserves fertility [3–9]. The hemostasis rate of UAE with gelatin sponge particles (GSPs) for obstetric and gynecologic hemorrhage ranges from 73 to 97 % [4–6]. In a coagulopathic condition, however, the recurrent hemorrhage rate is higher for GSPs than for N-butyl cyanoacrylate (NBCA) [10, 11]. Therefore, UAE with varying concentrations of NBCA is performed clinically for postpartum hemorrhage in patients with coagulopathy and/ or pseudoaneurysms [12–16]. The goal of UAE is complete hemostasis without uterine damage; however, the safety of NBCA in these concentrations is unknown for UAE. To address this shortcoming, we compared the degree of uterine damage caused by UAE with GSPs and with two different concentrations of NBCA in swine. Materials and methods Study design Institutional review board approval was obtained for this animal research. Fifteen swine were divided into three T. Sonomura (&) � N. Kawai � A. Ikoma � H. Minamiguchi � K. Kishi � H. Sanda � K. Nakata � M. Nakai � M. Sato Department of Radiology, Wakayama Medical University, 811-1 Kimiidera, Wakayama 641-8510, Japan e-mail:
[email protected] T. Ozaki � Y. Muragaki Department of Pathology, Wakayama Medical University, 811-1 Kimiidera, Wakayama 641-8510, Japan 123 Jpn J Radiol (2013) 31:685–692 DOI 10.1007/s11604-013-0236-2 groups of five according to the embolic material used: group A (1-mm GSPs), group B (concentrated NBCA mixture, NBCA:Lp = 1:1), and group C (dilute NBCA mixture, NBCA:Lp = 1:7). Ten uteri were tested per group; because swine have a bicornuate uterus, each swine provided two uteri. In swine, the two horns of the uterus are separate and the bilateral uterine arteries have no communication with each other. This allowed us to evaluate each horn individ- ually. We compared the degree of uterine damage caused by UAE for three different embolic materials. Animal model Five healthy, nonpregnant adult domestic swine were allocated to each of the three groups (n = 15). Mean body weight (±SD) was 53.6 ± 3.0, 53.2 ± 2.8, and 54.8 ± 1.8 kg in groups A, B, and C, respectively. There was no significant difference in weight among the three groups. For 3 days, starting 7 days before the procedure, a total of 6000 IU chorionic gonadotropin was administered intra- muscularly to enlarge the uterus and the diameter of the uterine arteries. All experimental procedures were performed under general anesthesia, and medical management was provided by veterinarians. Continuous blood pressure, ventilation rate, and electrocardiographic monitoring were maintained throughout the procedure. Preparation of embolic material Porous GSPs with a diameter of 1 mm (Gelpart, Nippon Kayaku/Astellas, Tokyo, Japan) were mixed with 10 mL of contrast material (Iopamidol 370, Bracco, Milan, Italy; iodine concentration: 370 mg) and the resulting suspension was aspirated into a 1-mL syringe. NBCA was mixed with Lipiodol (Lp) to enable monitoring under fluoroscopy. We prepared two NBCA mixtures: con- centrated (NBCA:Lp = 1:1) and dilute (NBCA:Lp = 1:7), which were each aspirated into a 2.5-mL syringe. Uterine artery embolization Under sterile conditions, a 4F sheath (Medikit, Tokyo, Japan) was introduced via the right femoral artery using Fig. 1 Angiography (#A1). a Aortography shows the common internal iliac artery (arrow), which is different to that seen in the human anatomy. b Common internal iliac arteriography clearly shows bilateral uterine arteries (arrows). c Left uterine arteriography shows many tortuous branches crossing the uterus. d Image obtained after UAE with GSPs shows complete occlusion of the left uterine artery (arrow) 686 Jpn J Radiol (2013) 31:685–692 123 Seldinger’s technique. Aortography or common internal iliac arteriography [17] was then performed for anatomic mapping using a 4F pigtail catheter (Cook, Bloomington, IN, USA) or a 1CJ catheter (Terumo-Clinical Supply Co., Gifu, Japan) with 370 mgI/mL nonionic contrast material (Fig. 1a, b). A coaxial 2.2F Sirabe microcatheter (Piolax, Yokohama, Japan) was advanced into the bilateral uterine arteries using a GT microguidewire (reshaped type, Terumo, Tokyo, Japan), and selective embolization was performed using GSPs (Fig. 1c, d) and the two NBCA mixtures. Gelatin sponge particles were injected slowly and intermittently to achieve complete occlusion of the uterine artery. Just before the NBCA mixtures were injected, the microcatheter was flushed with a 5 % glucose solution to prevent polymerization within the microcatheter. The NBCA was injected slowly into the uterine artery through a microcatheter, and the microcatheter was immediately retracted to prevent adherence of the catheter tip to the artery wall. We avoided rapid injection of the mixture by using a flush of 5 % glucose solution to push mixture from inside the microcatheter out into the artery; this technique gave us more accurate control over injection of the mixture. Sacrifice The swine were euthanized 3 days after embolization. The uteri, ovaries, and afferent arteries were removed and then fixed in 5 % neutral buffered formalin. Radiographic evaluation In groups B and C, radiographs of the removed specimens were taken to evaluate the distribution of NBCA. When NBCA mixtures were seen on a radiograph in the me- sometrial area, this was defined as a proximal distribution. However, when the NBCA mixtures were seen on a radiograph in the intrauterine area, this was defined as a distal distribution. The number of distal branches con- taining NBCA entering the uterus was counted (Fig. 2). Gross inspection The excised specimens in all three groups were evaluated for uterine necrosis, which is visible as brown-colored areas (Fig. 3). Necrosis rates in the uteri were calculated using 1-cm grids. To do this, we used a picture of the resected uterus and placed a grid of 1-cm squares over the Fig. 3 Macroscopic uterine findings. a Uteri from group A (#A1), showing small necroses (arrows). b Uteri from group B (#B5); no necrosis is seen. c Uteri from group C (#C3), showing large necroses Fig. 2 Diagram of a distal distribution of NBCA in the uterus. When NBCA was observed entering the uterus on the radiograph obtained after the procedure, the number of distal branches was counted. In this example, three distal branches (arrows) enter the uterus Jpn J Radiol (2013) 31:685–692 687 123 picture. We could then calculate the necrosis rates by dividing the number of 1-cm grids with necrosis by the total number of 1-cm grids covering the uterus. Microscopic evaluation The sections were embedded in paraffin blocks. Each slice from the paraffin block was stained with hematoxylin and eosin and examined under light microscopy. The uterus and mesometrium were observed, and the internal short diam- eters of occluded arteries in which GSPs and NBCA were detected were measured. Statistical analysis All data were expressed as the mean and standard deviation (SD), and the statistical significance of each difference was determined by applying Student’s t test. A p value of\0.05 was considered statistically significant. Results Selective embolization of the bilateral uterine arteries was successfully performed in all 15 swine (total 30 emboli- zations). The volume of NBCA used in UAE (excluding the dead space volume in the microcatheter) was 0.11 ± 0.04 mL (range 0.06–0.2 mL) in group B, and 0.16 ± 0.10 mL (range 0.08–0.4 mL) in group C. No significant difference in injected NBCA volume was observed between groups B and C (p = 0.207). Uterine artery angiograms/internal iliac artery angiograms obtained after UAE revealed occlusion of the bilateral uterine arteries in all 15 swine. Distribution of NBCA in groups B and C NBCA distributions are summarized in Table 1. Radio- graphs of group B showed a proximal distribution in 8 of the 10 uteri (Fig. 4a) and a distal distribution in 2 of the 10 uteri. In each of these two uteri, only one distal branch had entered the uterus. Table 1 Distribution of NBCA Swine Group B Swine Group C Right Left Right Left P D P D P D P D #B1 ? - ? - #C1 ? 3a ? 3a #B2 ? - ? - #C2 ? - ? - #B3 ? - ? - #C3 ? 3a ? 5a #B4 ? 1a ? 1a #C4 ? - ? 4a #B5 ? - ? - #C5 ? 5a ? 3a P proximal distribution, D distal distribution a Number of distal branches containing NBCA that entered the uterus Fig. 4 Post-procedure radiographs of the uteri. a A proximal distribution of NBCA is seen in the mesometrium of a group B swine (#B5). b Proximal and distal distributions of NBCA mixtures are seen in the mesometrium and uteri of a group C swine (#C3). c Magnified image of b, showing five distal branches containing NBCA entering the uterus (arrows) 688 Jpn J Radiol (2013) 31:685–692 123 Radiographs of group C showed a proximal distribution in 3 of the 10 uteri and a distal distribution in 7 of the 10 uteri (Fig. 4b). In the uteri with a distal distribution, the number of distal branches embolized with NBCA that had entered the uterus was 3 in 4 of the uteri, 4 in 1 of the uteri, and 5 in 2 of the uteri (Fig. 4c). Uterine necrosis Uterine necrosis is summarized in Table 2. Uterine nec- roses were found in 6 of the 10 uteri in group A, in 2 of the 10 uteri in group B, and in 8 of the 10 uteri in group C. The necrosis rate was 4.9 ± 6.1 % in group A, 1.3 ± 3.3 % in group B and 41.4 ± 28.8 % in group C, and was signifi- cantly higher in group C than in groups A (p = 0.0014) and B (p \ 0.001) (Fig. 5). Uterine necroses were found in all 9 uteri with a distal distribution of NBCA, and in only 1 of the 11 uteri with a proximal distribution of NBCA. There was a linear relationship (c2 = 0.93) between necrosis rate and the number of distal branches embolized with NBCA. Histological evaluation Histopathological examination (summarized in Fig. 6) showed karyolysis, karyorrhexis, and massive hemorrhage in the necrotic portions. GSPs were detected in uterine arteries with diameters of 154 lm (#A4) to 900 lm (#A4) in group A. NBCA was detected in arteries with diameters of 37 lm (#B4) to 2000 lm (#B5) in group B, and in arteries with diameters of 34 lm (#C3) to 2000 lm (#C3) in group C. Inflammatory reactions such as neutrophilic and/or lymphocytic infiltration and foreign body reactions were not observed around or inside embolized arteries containing GSPs or any concentration of NBCA. Discussion Gelatin sponge particles 1 mm in diameter caused limited areas of necrosis in swine uteri and were detected in uterine arteries 154–900 lm in diameter, similar to the findings of Sone et al. [18], who also reported lymphocytic infiltration around and inside the embolized arteries. No inflammatory reactions were observed in the present study, which may be due to the sacrifice time: 3 days in our study, versus 1 week in the study of Sone et al. We clinically use NBCA mixtures with ratios of between 1:1 and 1:7 for acute arterial bleeding [10]. For the purpose of comparison, we decided to look at the differ- ences in uterine damage between these two extreme ratios. Uterine necroses were found in all 9 uteri with a distal distribution of NBCA and in only 1 of the 11 uteri with a proximal distribution of NBCA. Distal emboli- zation causes more damage to the organs because there is less development of collaterals [19, 20]. A distal distribution of NBCA was found in 2 of the 10 uteri in group B and in 7 of the 10 uteri in group C. A distal distribution of NBCA was more common in uteri em- bolized with the dilute NBCA mixture. Pollak et al. [21] Table 2 Necrosis rates Swine Group A Swine Group B Swine Group C Right (%) Left (%) Right (%) Left (%) Right (%) Left (%) #A1 1.4 2.6 #B1 0 0 #C1 43.3 45.2 #A2 0 0 #B2 0 0 #C2 0 14.0 #A3 4.0 12.5 #B3 0 0 #C3 49.3 53.9 #A4 15.2 12.1 #B4 2.3 10.3 #C4 0 72.2 #A5 0 0 #B5 0 0 #C5 86.4 50.0 Mean ± SD 4.9 ± 6.1 1.3 ± 3.3 41.4 ± 28.8 Fig. 5 Histogram of necrosis rates among the three groups. Necrosis rates were significantly higher in group C than in groups A (p = 0.0014) and B (p \ 0.001) Jpn J Radiol (2013) 31:685–692 689 123 reported an in vivo polymerization time of 1–4 s for NBCA and Lipiodol mixed in ratios of 1:1 up to 1:4, and found a linear relationship between the time and the ratio. Dilution of the NBCA prolongs the polymerization time [22, 23], thereby enabling the mixture to travel a greater distance. In the present study, there was a linear 690 Jpn J Radiol (2013) 31:685–692 123 relationship between necrosis rate and the number of distal branches embolized with NBCA, which is com- patible with previous findings that embolization involv- ing four or more vasa recta carried an increased risk of substantial ischemic bowel damage [20, 24]. We detected NBCA in the capillary vessels of the necrotic portions, which is compatible with the findings of previous reports [25, 26]. However, Kish et al. [27] reported that NBCA did not permeate to the capillary level. This discrepancy is likely to be caused by factors such as differences in the NBCA injection speed and differences in the target organs. Sadato et al. [28] reported no substantial differ- ences in the tissue reactions of vessels embolized with low- and high-concentration mixtures of NBCA and Ethiodol (1:4 and 1:1, respectively) in their experimental study of rabbit renal arteries. In our study, the findings in the necrotic portions were similar for dilute NBCA (NBCA:Lp = 1:7) and concentrated NBCA (NBCA:Lp = 1:1), even though the necrosis rates were different for the two concentrations. There is one report of a woman becoming pregnant after treatment with the UAE procedure performed using NBCA at a concentration of NBCA:Lp = 1:10 [15]. In the present study of swine, however, the weak NBCA concentration (NBCA:Lp = 1:7) caused severe damage to the uteri. This discrepancy in results may be due to anatomical differences between human and swine uteri. Human uteri have rich anastomosis between both uterine and ovarian arteries. Also, postpartum uteri have a large number of collaterals and a dilated vascular bed. Therefore, the risk of ischemic injury after NBCA embolization may be lower than that for swine uteri, and a large amount of NBCA may be needed in UAE for postpartum hemorrhage. We clinically use NBCA mixtures in UAE for life- threatening obstetric and gynecologic hemorrhages in co- agulopathic patients. Therefore, a limitation of our study is that the swine uteri were not hemorrhagic. It is very dif- ficult to create such conditions artificially and to then perform UAE. These limitations mean that NBCA mixtures will cause less damage in human uteri. However, these results are still applicable to human uteri in terms of the difference in safety between low-concentration and high-concentration NBCA mixtures and embolization sites. Conclusion The concentration of NBCA is an important consideration when performing UAE for clinical patients. Dilute NBCA is able to migrate across a large distance because it takes a long time to polymerize. This in turn results in distal embolization over an extensive area, with uterine damage caused by the resultant necroses. 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