不同保存液保存小肠的效果评估

目的 评价乳酸林格（ＬＲ）液、Ｅｕｒｏ－Ｃｏｌｌｉｎｓ（ＥＣ）液、高渗枸橼酸嘌呤（ＨＣ－Ａ）液和武汉医学院器官１号（ＭＷＯ－１）液保存猪小肠的效果。方法 根据不同保存液将动物随机分成４组,每组又根据保存时间不同分成３个亚组,供肠保存２后行自体移植。结果 小肠保存１０小时后移植,术后３周各组受体的存活率均为１００％;保存１８小时,ＬＲ液组、ＥＣ液组、ＨＣ＝Ａ液组和ＷＭＯ－１液组受体的存活率分别为６０     

HX—3液和UW液保存大鼠肝脏效果的比较

采用大鼠肝脏非循环离体灌注模型比较自制的ＨＸ－３液和ＵＷ液对大鼠肝脏的保存效果。实验结果显示，经ＨＸ－３液原位灌洗并保存４８小时的肝脏肝组织含水量正常，而同等条件下换用ＵＷ液，肝组织的含水量虽无明显变化，但都低于正常值；随着保存时间的延长，两组肝窦内皮细胞死亡率逐渐上升，但在２４小时以内两组肝窥内皮细胞死亡率的差异不显著．     

长征－1号多器官保存液对兔肾脏冷冻保存的连续形态学观察

自１９９２年始研制成功长征－１号多器官保存液（简称ＣＺ－１液）。在完成对兔心脏及肝脏的冷冻保存后的形态学变化观察后，又做了对兔肾脏冷冻保存的形态学改变观察，并与ＵＷ液和ＨＣ－Ａ液作了比较。结果表明：冷冻保存７２小时以内ＣＺ－１液组肾脏组织光镜和电镜下形态学改变与ＵＷ组相比无明显区别，而且ＣＺ－１液组冷冻保存肾脏组织的变化以及细胞和细胞器的变性均缓于ＨＣ－Ａ组。由于ＨＣ－Ａ液已被证实能有效地保存犬肾在７２小时内，临床保存供肾在５７小时以内，ＵＷ液已经被证实能有效地保存供肾在７２小时以内，因此认为：从光镜和电镜下肾脏组织观察来看，ＣＺ－１液能保存兔肾脏在７２小时之内。     

血栓素A2合成抑制剂—川芎嗪改善大鼠胰腺保存效果的实验研究

本实验研究了川芎嗪加入本所自制的ＷＭＯ－１号器官保存淮对大鼠胰腺保存效果的影响。结果表明，ＷＭＯ－１号保存液对大鼠胰腺的确切保存时限为１２小时，１８小时保存存活率仅为１／３，加入川芎嗪要使保存时限延长至１８小时，２４小时保存存活率为１．３。证明川芎嗪确能改善胰腺保存。     

自制CZ—1液对兔心肌组织低温延时保存的生化学指标观察

研制出我国自制的多种改良的ＵＷ液，是摆在移植界的一大课题。我院于１９９２年开始研制成长征－１号多器官保存液（简称ＣＺ－１液），同时对心肌组织０、６、１２、１８、２４小时冷冻保存后的生化学指标作了观察。结果表明：与ＵＷ液比较，ＣＺ－１液０、１２、１８、２４小时冷冻保存兔心肌组织ＡＴＰ含量无明显差别。而ＣＺ－１液冷冻保存心肌线粒体Ｃａ＾２＋超载明显缓于ＵＷ液。文章认为从生化指标变化来说，我们自的ＣＺ－     

长征—1号多器官保存液对兔肾脏冷冻保存的连续形态学观察

自１９９２年开始研制成功长征１号多器官保存液（简称ＣＺ－１液）在完成对兔心脏及肝脏的冷冻保存后的形态学变化观察后，又做了对兔肾脏冷冻保存的形态学改变观察，并与ＵＷ液和ＨＣ－Ａ液作了比较，结果表明，冷冻保存７２小时以内ＣＺ－１液组肾脏组织光镜和电镜下形态学改变与ＵＷ组相比无明显区别，而且ＣＺ－１液组冷冻保存肾脏组织的变化以及细胞和细胞器的变性均缓于ＨＣ－Ａ组，由于ＨＣ－Ａ液已被证实能有效地保存犬肾在     

自制HYD液对大鼠肝脏低温保存后生化功能影响的研究

目的 研制自制ＨＹＤ液对大鼠肝脏低温保存后生化功能的影响。方法 采用大鼠肝脏非循环离体灌注模型，比较ＨＹＤ液，ＵＷ液和乳酸林格液（ＬＲ液）对大鼠肝脏６，１２，２４，３０，３６ｈ保存后生化功能的影响。结果 保存１２ｈ的肝脏，其各项生化功能ＨＹＤ组明显优于ＬＲ组。三磷酸腺苷，磷酸腺苷含量及Ａｔｋｉｎｓｏｎ能荷（ＡＥＣ），ＨＹＤ组略高于ＵＷ组，且保存３６ｈ差异有显著性。     

温度对WMO-1号液保存犬肾的影响

目的 探讨温度对ＷＭＯ－１号液保存犬肾的影响。方法 采用保存－自体移植模型,比较了不同温度条件下ＷＭＯ－１号液保存犬肾２４小时后的肾功能和形态学变化。结果 保存肾的损害程度与保存温度有关。１５℃保存２４小时不足以防止肾脏缺血损害。５－８℃组术后最大血肌酐值和平均血肌酐值均显著低于１－４℃组和９－１２℃组,形态学变化亦较轻。结论 ＷＭＯ－１号液保存犬肾的最适宜保存温度为５－８℃,保存温度过低或过高均     

川芎嗪对犬肾保存的影响

通过犬自体肾移植模型研究了一定浓度的川芎嗪加入ＷＭＯ－１号液对犬肾保存效果的影响。结果表明，单纯ＷＭＯ－１号液保存７２小时和加川芎嗪保存７２小时的肾行自体移植均难存活；加川芎嗪保存４８小时组术后血肌酐的平均值低于单纯ＷＭＯ－１号液保存４８小时组，且血肌酐恢复正常的时间也较单纯ＷＭＯ－１号液组早（Ｐ＜０．０５），说明该实验浓度的川芎嗪不能延长肾保存时间，但有利于移植肾功能的早期恢复。     

长征-1号多器官保存液低温保存大鼠肺脏并移植的实验研究

目的 通过大鼠原位肺移模型,研究长征－１号多器官保存液保存肺的效果。方法 将大鼠随机分为３组;对组照;以平衡液灌洗供肺,并立即移植;ＣＺ－１液组;以ＣＺ－１液灌洗供肺,保存８小时后移植;Ｅｕｒｏ－Ｃｏｌｌｉｎｓ液组,以Ｅｕｒｏ－Ｃｏｌｌｉｎｓ液灌洗供肺,保存８小时后移植。     

Liver Transplantation Using University of Wisconsin or Celsior Preserving Solutions in the Portal Vein and Euro-Collins in the Aorta

Introduction

Orthotopic liver transplantation (OLT) is today the gold standard treatment of the end-stage liver disease. Different solutions are used for graft preservation. Our objective was to compare the results of cadaveric donor OLT, preserved with the University of Wisconsin (UW) or Celsior solutions in the portal vein and Euro-Collins in the aorta.

Methods

We evaluated retrospectively 72 OLT recipients, including 36 with UW solution (group UW) and 36 with Celsior (group CS). Donors were perfused in situ with 1000 mL UW or Celsior in the portal vein of and 3000 mL of Euro-Collins in the aortia and on the back table managed with 500 mL UW or Celsior in the portal vein, 250 mL in the hepatic artery, and 250 mL in the biliary duct. We evaluated the following variables: donor characteristics, recipient features, intraoperative details, reperfusion injury, and steatosis via a biopsy after reperfusion. We noted grafts with primary nonfunction (PNF), initial poor function (IPF), rejection episodes, biliary duct complications, hepatic artery complications, re-OLT, and recipient death in the first year after OLT.

Results

The average age was 33.6 years in the UW group versus 41 years in the CS group (P = .048). There was a longer duration of surgery in the UW group (P = .001). The other recipient characteristics, ischemia-reperfusion injury, steatosis, PNF, IPF, rejection, re-OLT, and recipient survival were not different. Stenosis of the biliary duct occured in 3 (8.3%) cases in the UW group and 8 (22.2%) in the CS (P = .19) with hepatic artery thrombosis in 4 (11.1%) CS versus none in the UW group (P = .11).

Conclusion

Cadaveric donor OLT showed similar results with organs preserved with UW or Celsior in the portal vein and Euro-Collins in the aorta.     

Rescue of the Cold Preserved Rat Liver by Hypothermic Oxygenated Machine Perfusion

The aim of the study was to investigate whether hypothermic oxygenated liver perfusion after cold liver preservation resuscitated metabolic parameters and whether this treatment had a benefit for liver viability upon reperfusion.
We preserved rat livers either by cold storage (UW) for 10 h, or by perfusion for 3 h (oxygenated modified UW) after 10 h cold storage. We assessed viability of livers after preservation and after ischemic rewarming + normothermic reperfusion ex vivo . Ten hour cold storage reduced mitochondrial cytochrome oxidase and metabolically depleted the livers. Oxygenated perfusion after cold storage resulted in uploaded cellular energy charge and oxidized mitochondrial cytochrome oxidase. Reperfusion after 10 h cold storage increased formation of superoxid anions, release of cytosolic LDH, lipid peroxidation, caspase activities and led to disruption of sinusoidal endothelial cells. In contrast, reperfusion after 10 h cold storage + 3 h hypothermic oxygenated perfusion resulted in no changes of lipid peroxidation, bile flow, energy charge, total glutathione, LDH release and of caspase activation, as compared to fresh resected livers.
This study demonstrates, that a metabolically depleted liver due to cold storage can be energy recharged by short-termed cold machine perfusion. The machine perfused graft exhibited improved viability and functional integrity.     

冷保存供肝肝移植大鼠肝窦内皮细胞损伤机制的研究

目的 探寻肝移植大鼠肝窦内皮细胞(SEC)损伤的详细过程、方式及机制,为冷保存再灌注损伤的保护研究开辟新的途径.方法 雄性SD大鼠随机分为假手术组(n=6)、UW 1 h肝移植组(n:48)、Uw 12 h肝移植组(n=48).大鼠原位肝移植采用双袖套法,分别于术后不同时相点采取血液及组织标本,检测血清丙氨酸氨基转移酶(ALT)及透明质酸(HA)水平;HE染色观察肝脏病理学变化;TUNEL法检测凋亡,免疫组化法检测Bcl-2及Cleaved Caspase-3的表达状况.结果 UW 1 h、UW 12 h组肝移植后血清ALT、HA均较假手术组明显升高(P<0.05),UW 12 h组又明显高于Uw 1 h组(P<0.05).UW 12 h组ALT水平于术后6 h达高峰,而HA水平却在术后1 h、24 h呈双峰表现.Uw 12 h组首先出现SEC的凋亡继而出现肝细胞的坏死,且UW 12 h组细胞凋亡指数(apoptosis index,AI)明显高于UW 1 h组(P<0.01).两组大鼠SEC的AI均于术后6 h达高峰,与血中ALT的高峰时相点一致.肝移植术后Bcl-2表达明显减弱(P相似文献   

Comparison of HTK- and UW-solution for liver preservation tested in an orthotopic liver transplantation model in the pig

The aim of this experimental study was to compare the preservation potency of University of Wisconsin (UW) and HTK (Bretschneider) solutions in an orthotopic liver transplantation (OLT) model in pigs. Livers were harvested using an in situ perfusion technique, where organs were flushed with the solution being tested, stored on ice — cold storage (CS) — for 2 or 24 h and then transplanted. Parameters monitored were liver enzymes in serum, hepatic water content, high energy phosphates, nuclear magnetic resonance (NMR) relaxation time T2, light microscopy and bile production. CS for 24 h is an extreme in pig liver preservation and is not compatible with animal survival. Biopsies showed drastic morphological changes and grafts did not produce bile in either group. (Bile production 2 h CS: HTK, 5.6 ± 1.8 ml/h; UW, 4.7 ± 2.3 ml/h) Enzyme release after reperfusion (ASGOT, ?LDH) was higher in long-term preservation. Hepatic tissue water content significantly decreased during CS in UW preserved livers. Edema alter reperfusion (?H₂0: HTK 24 h = + 5.6%, UW 24 h= + 4.8%) and regeneration capacity after reperfusion (UW 2 h = 63%, HTK 2 h = 55%, UW 24 h = 30%, HTK 24 h = 30%) were not significantly different. However, we did not observe major differences in preservation potency between the solutions tested. Differences were correlated, rather, with length 9 time of CS, than with the solution used. Therefore, HTK solution seemed to be a low potassium containing alternative to UW solution.     

Greater Hemodynamic Instability With Histidine-Tryptophan-Ketoglutarate Solution Than University of Wisconsin Solution During the Reperfusion Period in Living Donor Liver Transplantation

Objective

University of Wisconsin (UW) and histidine-tryptophan-ketoglutarate (HTK) solutions are the 2 most commonly used liver preservation solutions. The aim of this study was to compare cardiovascular stability, acid-base status, and potassium concentrations between patients who received grafts preserved in either UW or HTK solution in orthotopic liver transplantation (OLT).

Patients and Methods

In this retrospective study, 87 patients who underwent living donor OLT were divided into 2 groups: UW (n = 28) and HTK (n = 59). Group HTK was subdivided into group NF-HTK (n = 31; nonflushed before reperfusion) and group F-HTK (n = 28; flushed before reperfusion). We determined mean arterial pressure (MAP) and heart rate every minute for 5 minutes after reperfusion and the maximum change in these values and incidence of postreperfusion syndrome (PRS). Body temperature, cardiovascular and acid-base parameters, as well as potassium concentrations were compared at 5 minutes before and 5 and 30 minutes after reperfusion.

Results

The maximum decreases in MAP within 5 minutes after reperfusion were significantly greater in both the NF-HTK and the F-HTK groups. The rate of PRS was significantly greater in the NF-HTK compared with the UW group. Flushing with HTK solution decreased the rate of PRS; there was no significant difference between the F-HTK and UW groups. All serial changes in body temperature, cardiovascular and acid-base parameters, as well as potassium concentrations were similar among the 3 groups.

Conclusions

The incidence of PRS was greater using HTK compared with UW solution during the reperfusion period. Therefore, careful hemodynamic management is advised when using HTK solution.     

Apoptosis and regeneration of sinusoidal endothelial cells after extended cold preservation and transplantation of rat liver

BACKGROUND: Sinusoidal endothelial cells (SECs) are particularly susceptible to cold ischemia-reperfusion (I/R) injury. We have examined the process of injury and recovery of graft after cold-preserved liver transplantation, with special focus on the proliferation of SECs and regulatory mechanisms involved. METHODS: Male SD rats were divided into two groups according to length of cold preservation time in University of Wisconsin [UW] solution of graft: UW1h group and UW12h group. Graft function, incidence of apoptosis, proliferation of SECs and the expression of related regulatory factors were assessed after orthotopic liver transplantation (OLT). RESULTS: SECs are more sensitive to apoptosis induced by cold I/R injury compared with hepatocytes. Using bromodeoxyuridine and rat endothelial cell antigen-1 double immunostaining assay, SECs exhibited a delayed proliferation in comparison with hepatocytes, reaching a peak at 72 hr in UW1h group and 96 hr in UW12h group, respectively. Vascular endothelial growth factor increased at 24 hr after reperfusion, and peaked at 72 hr in both groups. Flt-1 and flk-1 expression was found to be mainly limited to SECs, with a peak in expression occurring between 72 and 96 hr, which coincided with the peak in SEC proliferation in UW1h group. However, flt-1 was found to be reduced significantly at any time throughout the experiments in UW12h group compared to sham. CONCLUSION: The delayed recovery of rat liver after extended cold preservation and transplantation correlates with a retarded regeneration of SECs due to increased apoptosis and reduced expression of flt-1. These results suggest that SECs play an important role in cold-preserved liver transplantation.     

肝移植术中应用S-腺苷-L-蛋氨酸对供肝热缺血损伤的影响

目的探讨术中S-腺苷-L-蛋氨酸（SAMe）加入UW液和血浆冲洗液对热缺血损伤供肝及其恢复的影响。方法建立10min热缺血大鼠肝移植模型，分为A组：UW液灌注＋乳酸钠林格氏液冲洗、B组：UW液灌注＋血浆冲洗、C组：SAMe加入UW液灌注＋血浆冲洗和D组：UW液灌注＋SAMe加入血浆冲洗4组，观察肝组织组织病理学变化和电子显微镜下超微结构变化，并检测血清AST和透明质酸。结果C组和D组术后24h血清AST均低于B组（P〈0．05）。A组术后3h和24h血清HA高于B组（P〈0．05），B组复流后3h及24h血清HA均高于C组和D组（P〈0．05）。组织病理学表现B组复流后3h和24h肝细胞损伤和微循环紊乱较C组和D组明显；超微结构表现，A组复流后3h线粒体肿胀，肝窦内皮细胞肿胀，细胞核不规则，可见内皮细胞凋亡，大部分区域肝窦状隙明显狭窄，内皮层结构模糊，红细胞淤积，受压变形，白细胞附壁，可见内皮层完整性破坏；复流后24h，可见线粒体嵴断裂，核融解。B组内皮细胞损伤较A组轻，C组和D组超微结构表现微循环紊乱和肝细胞损伤表现较B组轻。结论供肝切取术中UW液中加入SAMe灌注保存，血浆冲洗液中加入SAMe可改善热缺血供肝微循环，减轻缺血再灌注损伤，并减轻肝细胞热缺血损伤，有利于10min热缺血供肝功能的恢复。     

Advantages of normothermic perfusion over cold storage in liver preservation

BACKGROUND: To minimize the ischemia-reperfusion injury that occurs to the liver with the current method of preservation and transplantation, we have used an extracorporeal circuit to preserve the liver with normothermic, oxygenated, sanguineous perfusion. In this study, we directly compared preservation by the standard method of simple cold storage in University of Wisconsin (UW) solution with preservation by perfusion. METHODS: Porcine livers were harvested from large white sows weighing between 30 and 50 kg by the standard procedure for human retrieval. The livers were preserved for 24 hr by either cold storage in UW solution (n=5) or by perfusion with oxygenated autologous blood at body temperature (n=5). The extracorporeal circuit used included a centrifugal pump, heat exchanger, and oxygenator. Both groups were then tested on the circuit for a 24 hr reperfusion phase, analyzing synthetic function, metabolic capacity, hemodynamics, markers of hepatocyte and reperfusion injury, and histology. RESULTS: Livers preserved with normothermic perfusion were significantly superior (P=0.05) to cold-stored livers in terms of bile production, factor V production, glucose metabolism, and galactose clearance. Cold-stored livers showed significantly higher levels of hepatocellular enzymes in the perfusate and were found to have significantly more damage by a blinded histological scoring system. CONCLUSIONS: Normothermic sanguineous oxygenated perfusion is a superior method of preservation compared with simple cold storage in UW solution. In addition, perfusion allows the possibility to assess viability of the graft before transplantation.     

Endothelial nitric oxide synthase protects transplanted mouse livers against storage/reperfusion injury: Role of vasodilatory and innate immunity pathways

Endothelial nitric oxide synthase (eNOS) plays a role in microcirculatory and immunomodulatory responses after warm ischemia/reperfusion. We hypothesized that eNOS is essential to maintain microcirculation, attenuate macrophage infiltration and decrease graft injury after liver transplantation. Liver transplantation was performed after 18 hours of cold storage in University of Wisconsin (UW) solution from wildtype and eNOS-deficient (B6.129P2-Nos3(tm/Unc)/J) donor mice into wildtype mice. Serum ALT, necrosis by histology, apoptosis by TUNEL, and macrophage infiltration by immunostaining against F4/80 antigen were determined 2 to 8 hours after implantation. Hepatic microcirculation was investigated after 4 hours by intravital confocal microscopy following injection of fluorescein-labeled erythrocytes. After sham operation, livers of wildtype and eNOS-deficient mice were not different in ALT, necrosis, apoptosis, macrophage infiltration, and microcirculation. After transplantation, ALT increased >3 times more after transplantation of eNOS-deficient livers than wildtype livers. Necrosis was >4 times greater, and TUNEL and F4/80 immunostaining in nonnecrotic areas were 2 and 1.5 times greater in eNOS-deficient donor livers, respectively. Compared with wildtype and eNOS sham-operated mice, sinusoidal blood flow velocity increased 1.6-fold after wildtype transplantation, but sinusoidal diameter was not changed. After transplantation of eNOS-deficient livers, blood flow velocity and sinusoidal diameter decreased compared with transplanted wildtype livers. These results indicate that donor eNOS attenuates storage/reperfusion injury after mouse liver transplantation. Protection is associated with improved microcirculation and decreased macrophage infiltration. Thus, eNOS-dependent graft protection may involve both vasodilatory and innate immunity pathways.     

c-Jun N-terminal kinase mediates hepatic injury after rat liver transplantation

BACKGROUND: Orthotopic liver transplantation (OLT) requires cold ischemic storage followed by warm reperfusion. Although c-Jun N-terminal kinase (JNK) is rapidly activated after OLT, the functional consequences of JNK activation are unknown. The aim of this study was to address the role of JNK after OLT using the selective JNK inhibitor CC-401. METHODS: Donors, recipients, or stored liver explants were treated with vehicle or JNK inhibitor before OLT by an arterialized two-cuff method with 40 hours of cold storage. Recipients were assessed for 30-day survival, and graft injury was assessed over time by hepatic histology, serum transaminases, caspase 3 activation, cytosolic cytochrome c, and lipid peroxidation. RESULTS: Survival after OLT increased after donor plus storage and storage only treatment with JNK inhibitor (P<0.05). Treatment of recipient only did not improve survival. Increased survival correlated with improved hepatic histology and serum aspartate aminotransferase levels. JNK inhibition significantly decreased nonparenchymal cell killing at 60 minutes after reperfusion (P<0.05) and pericentral necrosis at 8 hours after reperfusion (P<0.01). JNK inhibition decreased cytochrome c release, caspase 3 activation (P<0.05), and lipid peroxidation (P<0.05). JNK inhibition also transiently blocked phosphorylation of c-Jun at 60 minutes after reperfusion (P<0.05) without affecting other MAPK signaling, including p-38 and Erk activation. CONCLUSIONS: JNK inhibition decreases hepatic necrosis and apoptosis after OLT, suggesting that JNK activation promotes cell death by both pathways. Inhibition of JNK may be a new therapeutic strategy to prevent liver injury after transplantation.