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Improvement of allograft viability with organs procured from non–heart-beating donors in porcine liver transplantation
Improvement of Allograft Viability With Organs Procured From Non–Heart-Beating Donors in Porcine Liver Transplantation Y. Takada, K. Fukunaga, M. Gu, S. Ishiguro, H. Taniguchi, K. Seino, K. Yuzawa, M. Otsuka, and K. Fukao
W
ITH THE SHORTAGE of cadaveric donors, non– heart-beating donors (NHBDs) are a potential source of liver allografts. However, warm ischemic injury in the NHBD seriously affects liver allograft viability. In addition, the effects of warm ischemia are subsequently compounded by cold preservation and reperfusion injuries in liver transplantation.1 To establish the safety and efficacy of liver transplantation (LTx) from NHBDs, modalities to attenuate these ischemia and reperfusion injuries will be required. In the present study, pharmacologic modulations of graft viability procured from NHBDs were investigated in porcine liver transplantation.
MATERIALS AND METHODS Pigs weighing 17 to 20 kg were used after 24 hours of fasting. Orthotopic LTx was performed as previously described.2
Experiment 1 In the donors, 5 minutes after administration of 5000 IU of intravenous heparin, cardiac arrest was induced within a few
seconds by intravenous potassium chloride injection. The liver was subjected to in situ warm ischemia for 90 minutes. At the end of the warm ischemia period, the liver was harvested and stored for 4 hours using 4°C Euro-Collins solution. Pigs were divided into seven groups.3–5 Group 1 (n ⫽ 6) was the untreated, control group. In group 2 (n ⫽ 4), donors were intramuscularly injected with FK 506 (Fujisawa Pharmaceuticals, Osaka, Japan) at a dose of 0.3 mg/kg 18 hours before cardiac arrest. In group 3 (n ⫽ 4), the plateletactivating factor (PAF) antagonist, E5880 (Eisai Co, Tokyo, Japan), was administered to both donors and recipients at a dose of
From the Department of Surgery, Institute of Clinical Medicine, Tsukuba University, Tsukuba-city, Ibaraki, Japan. Supported in part by Grant-in-Aid for Scientific Research of the Ministry of Education, Science, Sports and Culture, Japan, and a grant from the Japan Society for the Promotion of Science (JSPS-RFTF96I00202). Address reprint requests to Dr Yasutsugu Takada, Department of Surgery, Institute of Clinical Medicine, Tsukuba University, Tennodai 1-1-1, Tsukuba-city, Ibaraki 305-8575, Japan. E-mail: [email protected].
Fig 1. Postoperative survival rate of recipients in the experiment 1.
© 2000 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010 Transplantation Proceedings, 32, 277–278 (2000)
0041-1345/00/$–see front matter PII S0041-1345(99)00956-2 277
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TAKADA, FUKUNAGA, GU ET AL
Fig 2. Serum AST concentrations at 4 hours after graft reperfusion in experiment 1. 0.3 mg/kg. In groups 4 (n ⫽ 4) and 5 (n ⫽ 6), the endothelin-1 (ET-1) antagonist, TAK-044 (Takeda Chemical Industries, Osaka, Japan), was administered to both donors and recipients at a dose of 3 and 10 mg/kg, respectively. In group 6 (n ⫽ 6), pigs were treated with both FK 506 and E5880. In group 7 (n ⫽ 4), pigs were treated with both TAK-044 (3 mg/kg) and E5880.
2.7%) than in the control group (59.2 ⫾ 5.9%, P ⬍ .05). Postoperative elevations of serum AST were attenuated in the treated group, although there were no significant differences between the two groups. DISCUSSION
Experiment 2 In donors, the respirator was stopped after administration of heparin and pancronium bromide. Cardiac arrest was induced in about 10 minutes. This model mimics the clinically controlled NHBD.1 Forty-five minutes after cessation of the respirator, the liver was flushed, harvested, and stored for 8 hours using 4°C UW solution. The pigs were divided into two groups: an untreated group (n ⫽ 6) and a drug-treated group (n ⫽ 6) in which TAK-044 was given in the UW solutions (10 mg/L) and to recipients (10 mg/kg).6
RESULTS Experiment 1
All recipient pigs in group 1 (control group) died within 12 hours after operation, whereas the pigs in the treated groups showed better survival (Fig 1). Especially in pigs treated with TAK 10 mg/kg (group 5), FK 506 ⫹ E5880 (group 6), and TAK 3 mg/kg ⫹ E5880 (group 7), recipient survival was significantly improved compared with control group. Serum AST concentrations at 4 hours after reperfusion of the graft (RPF) are demonstrated in Fig 2. In groups 5, 6, and 7, the postoperative increase in AST was attenuated significantly. Experiment 2
Postoperatively, the 7-day survival rate was 0% in the untreated group, and 67% in the TAK-treated group (P ⬍ .05). At 1 hour after RPF, the portal venous pressure was significantly lower in the treated group (12.2 ⫾ 0.5 cm H2O) than in the control group (15.0 ⫾ 1.1 cm H2O, P ⬍ .05). The 15-minute retention rate of indocyanine green (ICGR 15) was also significantly lower in the treated group (41.7 ⫾
In the experiment 1, it has been suggested that FK 506, the PAF antagonist E5880, and the ET-1 antagonist TAK-044 can improve the function of hepatic allografts subjected to prolonged (90-minute) warm ischemia in NHBDs. As a single agent, TAK-044, at the dose of 10 mg/kg, seemed to have the most protective effect among the three drugs. In experiment 2, effects of TAK-044 on graft function were evaluated in the model of controlled NHBD followed by a longer preservation time. Because donor pretreatment may have ethical implications, TAK-044 was administered only in the UW solution and to the recipients. As a result, treatment with TAK-044 alleviated the microcirculatory disturbances caused by ischemia-reperfusion injury, and improved graft function, leading to the better survival. The present study clearly demonstrates the feasibility of pharmacologic modulation of liver graft viability in organs procured from NHBDs. Because it has been shown that the protective effects of these drugs are synergistic, combined treatments with these agents have great potential for use in clinical LTx from NHBDs. REFERENCES 1. Cassavilla A, Ramirez C, Shapiro R, et al: Transplantation 59:197, 1995 2. Takada Y, Taniguchi H, Fukunaga K, et al: Transplantation 63:369, 1997 3. Takada Y, Taniguchi H, Fukunaga K, et al: Surgery 123:692, 1998 4. Fukunaga K, Takada Y, Taniguchi H, et al: Int Surg 83:226, 1998 5. Fukunaga K, Takada Y, Taniguchi H, et al: Transplantation 67:328, 1999 6. Fukunaga K, Takada Y, Mei G, et al: Am J Surg 178:64, 1999
W
ITH THE SHORTAGE of cadaveric donors, non– heart-beating donors (NHBDs) are a potential source of liver allografts. However, warm ischemic injury in the NHBD seriously affects liver allograft viability. In addition, the effects of warm ischemia are subsequently compounded by cold preservation and reperfusion injuries in liver transplantation.1 To establish the safety and efficacy of liver transplantation (LTx) from NHBDs, modalities to attenuate these ischemia and reperfusion injuries will be required. In the present study, pharmacologic modulations of graft viability procured from NHBDs were investigated in porcine liver transplantation.
MATERIALS AND METHODS Pigs weighing 17 to 20 kg were used after 24 hours of fasting. Orthotopic LTx was performed as previously described.2
Experiment 1 In the donors, 5 minutes after administration of 5000 IU of intravenous heparin, cardiac arrest was induced within a few
seconds by intravenous potassium chloride injection. The liver was subjected to in situ warm ischemia for 90 minutes. At the end of the warm ischemia period, the liver was harvested and stored for 4 hours using 4°C Euro-Collins solution. Pigs were divided into seven groups.3–5 Group 1 (n ⫽ 6) was the untreated, control group. In group 2 (n ⫽ 4), donors were intramuscularly injected with FK 506 (Fujisawa Pharmaceuticals, Osaka, Japan) at a dose of 0.3 mg/kg 18 hours before cardiac arrest. In group 3 (n ⫽ 4), the plateletactivating factor (PAF) antagonist, E5880 (Eisai Co, Tokyo, Japan), was administered to both donors and recipients at a dose of
From the Department of Surgery, Institute of Clinical Medicine, Tsukuba University, Tsukuba-city, Ibaraki, Japan. Supported in part by Grant-in-Aid for Scientific Research of the Ministry of Education, Science, Sports and Culture, Japan, and a grant from the Japan Society for the Promotion of Science (JSPS-RFTF96I00202). Address reprint requests to Dr Yasutsugu Takada, Department of Surgery, Institute of Clinical Medicine, Tsukuba University, Tennodai 1-1-1, Tsukuba-city, Ibaraki 305-8575, Japan. E-mail: [email protected].
Fig 1. Postoperative survival rate of recipients in the experiment 1.
© 2000 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010 Transplantation Proceedings, 32, 277–278 (2000)
0041-1345/00/$–see front matter PII S0041-1345(99)00956-2 277
278
TAKADA, FUKUNAGA, GU ET AL
Fig 2. Serum AST concentrations at 4 hours after graft reperfusion in experiment 1. 0.3 mg/kg. In groups 4 (n ⫽ 4) and 5 (n ⫽ 6), the endothelin-1 (ET-1) antagonist, TAK-044 (Takeda Chemical Industries, Osaka, Japan), was administered to both donors and recipients at a dose of 3 and 10 mg/kg, respectively. In group 6 (n ⫽ 6), pigs were treated with both FK 506 and E5880. In group 7 (n ⫽ 4), pigs were treated with both TAK-044 (3 mg/kg) and E5880.
2.7%) than in the control group (59.2 ⫾ 5.9%, P ⬍ .05). Postoperative elevations of serum AST were attenuated in the treated group, although there were no significant differences between the two groups. DISCUSSION
Experiment 2 In donors, the respirator was stopped after administration of heparin and pancronium bromide. Cardiac arrest was induced in about 10 minutes. This model mimics the clinically controlled NHBD.1 Forty-five minutes after cessation of the respirator, the liver was flushed, harvested, and stored for 8 hours using 4°C UW solution. The pigs were divided into two groups: an untreated group (n ⫽ 6) and a drug-treated group (n ⫽ 6) in which TAK-044 was given in the UW solutions (10 mg/L) and to recipients (10 mg/kg).6
RESULTS Experiment 1
All recipient pigs in group 1 (control group) died within 12 hours after operation, whereas the pigs in the treated groups showed better survival (Fig 1). Especially in pigs treated with TAK 10 mg/kg (group 5), FK 506 ⫹ E5880 (group 6), and TAK 3 mg/kg ⫹ E5880 (group 7), recipient survival was significantly improved compared with control group. Serum AST concentrations at 4 hours after reperfusion of the graft (RPF) are demonstrated in Fig 2. In groups 5, 6, and 7, the postoperative increase in AST was attenuated significantly. Experiment 2
Postoperatively, the 7-day survival rate was 0% in the untreated group, and 67% in the TAK-treated group (P ⬍ .05). At 1 hour after RPF, the portal venous pressure was significantly lower in the treated group (12.2 ⫾ 0.5 cm H2O) than in the control group (15.0 ⫾ 1.1 cm H2O, P ⬍ .05). The 15-minute retention rate of indocyanine green (ICGR 15) was also significantly lower in the treated group (41.7 ⫾
In the experiment 1, it has been suggested that FK 506, the PAF antagonist E5880, and the ET-1 antagonist TAK-044 can improve the function of hepatic allografts subjected to prolonged (90-minute) warm ischemia in NHBDs. As a single agent, TAK-044, at the dose of 10 mg/kg, seemed to have the most protective effect among the three drugs. In experiment 2, effects of TAK-044 on graft function were evaluated in the model of controlled NHBD followed by a longer preservation time. Because donor pretreatment may have ethical implications, TAK-044 was administered only in the UW solution and to the recipients. As a result, treatment with TAK-044 alleviated the microcirculatory disturbances caused by ischemia-reperfusion injury, and improved graft function, leading to the better survival. The present study clearly demonstrates the feasibility of pharmacologic modulation of liver graft viability in organs procured from NHBDs. Because it has been shown that the protective effects of these drugs are synergistic, combined treatments with these agents have great potential for use in clinical LTx from NHBDs. REFERENCES 1. Cassavilla A, Ramirez C, Shapiro R, et al: Transplantation 59:197, 1995 2. Takada Y, Taniguchi H, Fukunaga K, et al: Transplantation 63:369, 1997 3. Takada Y, Taniguchi H, Fukunaga K, et al: Surgery 123:692, 1998 4. Fukunaga K, Takada Y, Taniguchi H, et al: Int Surg 83:226, 1998 5. Fukunaga K, Takada Y, Taniguchi H, et al: Transplantation 67:328, 1999 6. Fukunaga K, Takada Y, Mei G, et al: Am J Surg 178:64, 1999