ASONUMA, KATSUHIRO

HYUN-SUONG

INOMATA, YUKIHIRO

KATAYAMA, TETSUO

KITAKADO, YASUHITO

MATSUOKA, SADAYUKI

OKAMURA, RYUJI

OZAWA, KAZUE

SHIMAMOTO, TAKESHI

TANAKA, KOICHI

UEMOTO, SHINJI

UTSUNOMIYA, HIROFUMI

Kyoto University Research Information Repository

Title<Original Articles>A New Bioabsorbable Material for RatVascular Cuff Anastmosis : Establishment for the Long-TermOrthotopic Liver Transplantation ModelAuthor(s)KITAKADO, YASUHITO; TANAKA, KOICHI; ASONUMA,KATSUHIRO; UEMOTO, SHINJI; MATSUOKA,SADAYUKI; UTSUNOMIYA, HIROFUMI; OKAMURA,RYUJI; KATAYAMA, TETSUO; INOMATA, YUKIHIRO;OZAWA, KAZUE; SHIMAMOTO, TAKESHI; HYUN-SUONGCitation日本外科宝函 (1992), 61(6): 445-453Issue Date1992-11-01URL http://hdl.handle.net/2433/203758RightType Departmental Bulletin PaperTextversionpublisherKyoto UniversityArch Jpn Chir 61(6), 445～453, Nov., 1992 A New Bioabsorbable Material for Rat Vascular Cuff Anastomosis: Establishment for the Long-Term Orthotopic Liver Transplantation Model YASUHITO KITAKADO, K01cH1 TANAKA, KATSUHIRO AsoNUMA, SHINJI UEMOTO, SADAYUKI MATSUOKA, HIROFUMI UTSUNOM!YA, RYUJI OKAMURA, TETSUO KATAYAMA, YUKIHIRO lNOMATA, KAzUE OzAWA, TAKESHr SHrMAMOTOへSuoNG-HvuNへYOSHITO lKADA * Second Department of Surgery, Faculty of Medicine, Kyoto University and *Research Center for Medical Polymers and Biomaterials, Kyoto University, 54 Kawaracho, Shogoin, Sakyoku, Kyoto, 606 Japan Received for Publication, Aug 21, 1992 Abstract Rat vascular anastomosis was performed using a newly synthesized bioabsorbable material (LA-GA copolymer) cuffs and the ordinary polyethylene cuffs. The LA-GA copolymer cuff which anastomosed the portal vein and the inferior vena cava were patent and developed no collateral veins even after 6 months, keeping the transplanted liver normal. By contrast, the polyethylene cuff anastomosed portal vein was completely occluded and the collateral veins were highly developed, with the transplanted liver showing the fatty degeneration ofhepatocytes and numerous regenerative nodules. It is concluded that the LA-GA copolymer cuff is a suitable material for the short and long term study of rat orthotopic liver transplantation. Introduction Cuff vascular anastomosis is an excellent method that is widely used in orthotopic liver transplantation1・2l. The polyethylene cuff employed in this method, however, has the disadvantages of foreign body reaction, fibrous tissue formation, incidence of immunological reaction and possible carcinogenesis after long-term use3・4・5・6l The result is that the venous anastomosis becomes occlud-ed and the blood supply to the transplanted liver has to be maintained by collateral veins. So few studies have been done on the long term orthotopic liver transplanted rats. It is necessary to establish a long-term orthotopic liver transplanted rat model. At the Kyoto University Research Center for Medical Polymers and Biomaterials, a new bioab-sorbable mater凶（LA-GAcopolymer) was synthesized from D,L lactic acid and glycolic acid. This new bioabsorbable material has ester-bonds and can easily be hydrolyzed. The metabolites produc-ed upon hydrolysis enter the metabolic pathways, and are excreted primarily as water and carbon dioxide. The rate of biodegradation is dependent on the molecular weight and composition ratio of Key words: Inferior vena cava anastomosis, Portal vein anastomosis, Orthotopic liver transplantation, LA-GA copolymer cu百， Polyethylenecu百索引用語下大静脈吻合，門脈吻合，同所性肝移植， LA-GA共重合体カフ，ポリエチレンカフPresent address: 2nd Department of Surgerγ，Faculty of Medicine, Kyoto University, 54 Shogoin, Kawaracho, Sakyoku, Kyoto, 606Japan. 446 日外宝第61巻第6号（平成4年1月）D,L-lactic acid and glycolic acid7・8・9) In this paper, the LA-GA copolymer cuff was used in the cuff-anastomosed rat inferior vena cava and in the orthotopic rat liver transplantation in order to examine the patency of the anastomosis and the histological change of the transplanted liver. The result was compared with the ordinary polyethylene cuffs. Materials and民1ethodsAnimal Male 九'¥Tistarrats weighing 300-380 g were used. LA-GA copolymer LA-GA copolymer was synthesized by the polycondensation ofD,L-lactic acid and glycolic acid 7,B,9). The copolymer has a molecular weight of 40-50 kDa, its composition ratio of D,L-lactic acid to glycolic acid is 8 : 2, and its iπvivo degradation time is about 4 months. The cuffs used for the por-ta! vein and inferior vena cava anastomosis had cuff bodies 2.0 mm and 2.5 mm long, 2.2 and 2.6 mm in outside diameter, and 2.0 and 2.4 mm in inside diameter, respectively. Iψrior vena cava anastomosis Iψrior vena cavagraft procurement: Under ether anesthesia, the abdominal cavity was opened, and the right renal vessels and lumbar vein were ligated and dissected. The inferior vena cava was removed from the lower liver margin to the left renal vein for venous grafting. Cuff preparation was performed in a 4°C iced saline bath. Both ends of the vessel were everted over the cuff and secured with a circumferential 7-0 nylon suture. Cu technique for inferior vena cava nastomosis: Under ether anesthesia, abdominal cavity was open-ed, and a right kidney was resected. The inferior vena cava was cross-clamped with a mosquito clip, transecting the midpoint between the left renal vein and the lower liver margin. Cuff anastomosis was completed and secured with a circumferential 7 0 nylon suture. Rats were sacrificed at 3, 5, 7, 9, 11, 13, 15 and 17 weeks, both for the LA-GA copolymer and the polyethylene cuff. Orthotopic liver transplaπtalion Orthotopic liver transplants were performed in male Wistar rats, using the modified cufftechni-que described by KAMADA et al.1・2l. Recipients were divided into the LA-GA copolymer and the polyethylene cuff groups. Rats were maintained on a diet of laboratory chow and water ad libitum postoperatively. Venography The patency of the anastomosed inferior vena cava and portal vein, and formation of collateral veins were examined by venography, which was performed as follows. Under ether anesthesia, a silicon catheter was inserted into the femoral vein or the branch of the superior mesenteric vein through which contrast material (5 to 10 ml of 60% urogra伍n)was injected and X rーayfilms were then taken. Light micτOSCO.りImmediately after venography, 10 to 20 ml of physiological saline followed by 10% for-maldehyde was perfused through the same catheter used for venography. The anastomosed inferior vena cava and portal vein and the transplanted liver were dissected free, removed, and processed for histology. The sections were prepared with hematoxylin and eosin. A New Bioabsorbable Material for Rat Vascular Cuff Anastomosis 447 Results Iψrior vena cava anastomosis with the LA -GA ゅのmercu.f 23 rats were anastomosed with the LA-GA copolymer cuff. All survived before sacrifice. In 3 rats, at 7, 11, 15 week, respectively, steno sis of the venous graft and formation of collateral veins were seen not because of the LA-GA copolymer itself but because of technical error, possibly due to the twisting of the venous gra氏. But tissue reaction such as granulation and fibrosis was absent and thrombus formation was not noted. Venography showed that neither stenosis nor occlusion were observed in the other 20 rats (Fig. 1A). The LA-GA copolymer cuff showed signs of being gradually absorbed, the time course for which is shown in Fig. 2. The LA-GA copolymer cuff became white-colored, softened and slightly swollen at 5 weeks and almost completely absorbed at 15 weeks. Microscopically, the LA-GA copolymer cuff was gradually reduced in thickness and completely disappeared at 17 weeks (Fig. 3A). l抗iferiorvena cava ana山 mosiswith the polyethylene cu.f 12 rats were anastomosed with polyethylene cu汀s. All survived before sacrifice. Venous anastomosis gradually became stenotic and was occluded after 9 weeks. When the anastomosed vessels were occluded, collateral veins highly developed (Fig. 1B). The polyethylene cuff came to be encapsulated by granulation and fibrosis. Microscopically, foreign body reaction to the polyethylene cuff and thrombus formation in the anastomosed vessel were seen (Fig. 3B). 0泊otopicliver transplantation with the LA-GA cop均mercu.f Orthotopic liver transplantation was performed with LA-GA copolymer cuffs in 5 rats. All rats Fig. 1A Venography of the LA-GA copolymer cu百anastomosedinferior vena cava at 17 weeks. Neither stenosis nor forma-tion of collateral veins were seen Fig. 1B Venography of the polyethylene cuff anastomosed inferior vena cava at 15 weeks. Note many collateral veins and occlusion of the anastomosis. 448 日外宝第61巻第6号（平成4年1月）survived until sacrificed. Macroscopically, the LA-GA copolymer cuffs had completely disappeared from the anastomotic site after 6 months. Venography of the anastomosed portal vein showed neither stenosis nor collateral vein formation (Fig. 4A). Microscopically, foreign-body reaction and thrombus formation around the anastomotic site were not observed. Histology revealed that the transplanted liver was normal. Neither hepatocyte damage nor fibrosis were observed (Fig. SA). Orthotopic liver traゆ lantationwith the polyethylene cuff Orthotopic liver transplantation was performed with polyethylene cuffs in 5 rats. The polyethylene cuff anastomosed portal veins were occluded and collateral veins were highly developed 5 week 1 5 week 1 7 week Fig. 2 Time course ofbiodegradation of the LA-GA copolymer cuf. At 5 weeks, it became white-colored, softened and slightly swollen. At 17 weeks, it was completely absorbed. The arrow shows LA-GA copolymer cuff A New Bioabsorbable MaterialおrRat Vascular Cuff Anastomosis 449 5w 17w Fig. 3A Histology of LA-GA copolymer cuff anastomosis. Note that there is litle foreign body reaction and no thrombus formation in the lumen. At 5 weeks, the LA-GA copolymer cuff (arrow) is the lucent area due to acetone treatment. At 17 weeks, it 1s no longer seen (20×， H & E) L=intravascular lumen ~ 弘、一一、一Fig. 3B Histolo幻rof polyethylene cuff anastomosis at 15 weeks. Note fibrosis around the cuff and thrombus forma-tion in the lumen (20×， H & E). T=thrombus, PE=polyethylene 450 日外宝第61巻第6号（平成4年11月）Fig. 4A Venography of the LA-GA copolymer cuff-anastomosed portal vein. Note there is neither stenosis nor col-lateral vem formation. Fig. 4B Venography of the polyethylene cuff-anastomosed po口al、rein. Portal vein was occluded and collateral veins were highly developed. A New Bioabsorbable Material for Rat Vascular Cuff Anastomosis Fig. 5A Histology of the transplanted liver with the LA-GA copolymer cuff anastomosis hepatocyte damage (200 x , H& E) Fig. 5B 451 452 日外宝第61巻第6号（平成4年1月）in al rats after 6 months (Fig. 4B). Foreign body reaction to the polyethylene cuff and thrombus for-mation in the anastomosed vessels were observed microscopically. Histology of the transplanted liver showed the fatty degeneration of hepatocytes and numerous regenerative nodules (Fig. SB). Discussion As our previous report and those of others10・11・12・13l reveal, the LA-GA copolymer cuff has cer-tam characteristic advantages for vascular anastomosis. Basically, the copolymer disappears from the rat tissues within 4 months and produces litle foreign body reaction. As a result, the anastomotic site is neither occluded nor constricted by the presence of the copolymer. IWASA et al. have performed rat microvascular anastomosis between the carotid artery and jugular vein using the LA-GA copolymer as an external splint and have confirmed the good patency and virtual absence of the foreign body reaction upon examination by angiography and microscopy10l. But the arterio-venous anastomosis is different from veno-venous anastomosis in certain respects, such as blood pressure and flow. In rat orthotopic liver transplantation, veno-venous anastomosis is important because the transplanted liver is usually not being supplied with blood from the hepatic artery. Various methods for venous anastomosis have been proposed. Among them, the cuff method is widely used because it is relatively easy to perform. Many researchers have reported on the long-term survival of rat orthotopic liver transplantation using the polyethylene cuff, but few studies have been done on the long-term orthotopic liver transplanted rat because of the disadvan-tages of the polyethylene cuff employed in the anastomosis between portal vein and vena cava. Our histological findings revealed that anastomosed vessels with the polyethylene cuff became gradually stenotic and were finally occluded and collateral veins were highly developed. The foreign body reac-tion to the polyethylene cuff led to the stenotic change of the cuff-anastomosed vessel lumen. The stenotic change resulted in the blood flow stasis and thrombus formation. Histological study of these transplanted livers showed liver cell damage which was probably due to the portal vein obstruction. Although KAMADA et al. described that the histology of the liver graft 438 days after liver transplanta-tion showed preservation of normal lobular architecture, they did not state how many of the or-thotopic liver transplanted rats kept normal liver architecture and the patency of the cuff-anastomos-ed portal vein. In our laboratory, a half of the orthotopic liver transplanted rats with polyethylene cuffs restored from this liver cell damage at the time of one year after liver transplantation probably due to the highly developed collateral veins. On the other hand, the LA-GA copolymer cuff-anastomosed inferior vena cava and portal vem were patent and no collateral veins were developed even after 6 months. The histology of the transplanted liver was kept normal. It is concluded that the polyethylene cuff is material suitable only for short term rat orthotopic liver transplantation. Using the LA-GA copolymer cuff, however, it is possible to perform either short-term or long-term rat orthotopic liver transplantation. References 1) Kamada N, Caine RY: Orthotopic liver transplantation in the rat. Transplantation 28: 47-50, 1979 2) Kamada N, Caine RY: A surgical experience wi出品vehundred thirty liver transplants in the rat. Surgery 93: 6←69, 1983 A New Bioabsorbable Material for Rat Vascular Cuff Anastomosis 453 3) Oppenheimer ET, Willhite M, Danishefesky I, et al: Furtherはudiesof polymers as carcinogenic agents m animals. Cancer Res 15: 333-340, 1955. 4) Carter RL, Roe FIC: Induction of sarcomas in rats by solid and fragmented polyethylene: experimental observa-tions and clinical implications. Bri J Cancer 23: 401-407, 1968. 5) Paiva T, Makinen]: Histopathological observation on polyethylene type materials in chronic ear surgerγ， Acta Otolaryngol 95: 139-146, 1983. 6) Tomoda K: Histopatholog:ical studies in the long-term evaluation of plastipore prostheses. Arch Otorhinolaryngol 242：・ 119-128,1985. 7) Ikada Y, et al: Polymer Reprints, Japan 30, 1688, 1981. 8) Jam出 diK, Hyon SH, Nakamura T, et al: In vitro and in vivo degradation ofpoly-L-lactide fibers. Biol噌caland biochemical performance ofbiomaterials. Elsevier Science Publishers B.V., Amsterdam, p. 227, 1986. 9) Hyon SH, Jamshidi K, Ikada Y: ACS symposium series: polymers as biomaterials. p. 51, 1985. 10) Iwasa H, Sato F, Shimabukuro H, et al: One knot microvascular anastomosis. J Neurosurg 66: 76←767, 1987. 11) Sumitomo K, Dohi K, Ikada Y, et d・ Applicationof " new synthetic absorbable cuff material to vascular anastomosis in liver grafting. Transplantation 46: 318-321, 1988. 12) Kita】cadoY, Tanaka K, Asonuma K, et al: A new bioabsorbable material for rat venous anastomosis. Transplantation 46: 620-622, 1988. 13) Kitakado Y, Tan心也 K,Asonuma K, et al: A new bioabsorbable material for rat orthotopic liver transplantat-ion. Eur Sur Res 23: 366 370, 1991 和文抄録新しい生体吸収材料カフによるラットの血管吻合一長期肝移植モデルの確立－京都大学医学部第二外科学講座北角泰人，田中紘一，阿曽沼克弘，上本伸二，松岡貞幸宇都宮裕文，岡村隆仁，片山哲夫，猪股裕紀洋，小津和恵京都大学生体医療工学研究センター島本偉志，玄 丞休，筏義人新しく合成された生体吸収材料（LA-GA共重合体）カフを用い，ラ y トの血管吻合を行ない，従来のポリエチレンカフと比較検討した.LAーGA共重合体カフで吻合した門脈，下大静脈は6ヶ月後でも側副血行路は認められずに開存し，移植肝は正常の組織像を保ってし、た．一方，ポリエチレンカフで吻合した門脈，下大静脈は完全に閉塞し，高度に発達した側副血行路が認められ，その結果移植肝には肝細胞の脂肪変性と多数の再生結節が認められた．以上の結果より， LAGA共重合体カフはラ、ソト同所性肝移植の短期及び長期の研究に適した生体材料だと結論付けられる．

<原著>新しい生体吸収材料カフによるラットの血管吻合 : 長期肝移植モデルの確立

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<原著>新しい生体吸収材料カフによるラットの血管吻合 : 長期肝移植モデルの確立

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