Donor pretreatment with gadolinium chloride improves early graft function and survival after porcine liver transplantation

Kupffer cell depletion by gadolinium chloride (GdCl₃) in rat livers has previously been proven to minimize hepatic ischemia/reperfusion injury after experimental liver transplantation (LTX). In the current study, we evaluated the effects of donor pretreatment with GdCl₃ on hepatic ischemia/reperfusion injury, macro- and microcirculation, and endotoxin clearance of the liver in a porcine model of experimental LTX. Two groups of 12 pigs were treated either with intravenous NaCl (0.9%; control) or GdCl₃ (20 mg/kg). Twenty-four hours after pretreatment, hepatic macrocirculation was quantified by Doppler flowmetry and liver parenchymous microcirculation by implanted thermodiffusion electrodes. The liver grafts were transplanted after 4–6 h of cold ischemia in University of Wisconsin (UW) solution. At 1 and 24 h after LTX, the perfusion values were re-evaluated and histology, biochemical (aspartate aminotransferase, AST) and functional parameters (partial thromboplastin time, prothrombin time, and bilirubin) were analyzed. Furthermore, endotoxin clearance of the liver was evaluated at all time points. In GdCl₃-treated animals 80% of the Kupffer cells were destroyed, and 24 h after LTX ischemia/reperfusion injury in treated grafts was significantly lower in comparison to controls, as shown by histology, AST levels (741±490 U/l in controls vs 379±159 U/l in treated grafts, P<0.05), survival (67% vs 92%), and enhanced macro- (total transhepatic blood flow [THBF]=112±22 ml/min per 100 g in controls vs 157±45 ml/min per 100 g in treated grafts, P0.05) and microcirculation (thermodiffusion [TD]=73±9 ml/min per 100 g in controls vs 90±16 ml/min per 100 g in treated grafts, P0.05). Despite destruction of the macrophage system in the liver, the transhepatic endotoxin gradient of treated livers was enhanced before and 1 h after transplantation (58% in controls vs 85% in treated grafts, P<0.05). Destruction of Kupffer cells of donors by pretreatment with GdCl₃ in pigs is effective in preventing liver graft dys- and nonfunction after LTX. Pretreatment with GdCl₃ does not diminish but increase hepatic endotoxin clearance.     

内皮素-1与肝脏缺血再灌注损伤的实验研究

目的：探索内皮素－1（ET－1）在肝脏血再灌注损伤中的作用。方法：选择雄性Wistar大鼠80只,分为正常对照组、缺血再灌注组1生理盐水组和ET－1抗体组、观察肝脏缺血60min再灌注3h后血浆ET－1、丙氨酸转氨酶（ALT）、透明质本酸（HA）、以及肝组织中ET－1和丙二醛（MDA）含量的变化,并观察肝组织病理学变化,同时,在缺血再灌注组选择第1、3、6、12和24h时相点观察ET－1的变化规律。结果：肝脏缺血再灌注后,血浆和肝组织中ET－1,血浆HA`ALT肝组织中MDA显著升高,而ET－1抗体组血浆ET－1、HA、ALT与缺血再灌注组相比显著降低（P＜0．01,P＜0．05）,同时,肝组织的瘀血程度和损伤程度显著改善。结论ET－1参与了肝脏缺血再灌注损伤,这种损伤与肝脏微循环障碍有关。     

High Dose Intravenous Immunoglobulin G Pretreatment: Effect on Lipid Peroxidation and Reperfusion Injury to the Liver

The aim of this study was to examine the effect of intravenous immunoglobulin G (IVIG) compared with that of gadolinium chloride (GdCl₃) in the inhibition of Kupffer cell activation on lipid peroxidation after severe total hepatic ischemia/reperfusion (I/R). Male Wistar rats (n = 40) were randomly divided into four equal groups: a sham-operation group, a control I/R group, and two I/R groups pretreated with either IVIG or GdCl₃ intravenously. Following 60 minutes of total hepatic ischemia and 120 minutes of reperfusion, the rats were sacrificed and liver and blood samples collected. Additional animals (n = 80) were followed up for survival rate determination. Results showed that I/R decreased the survival rate to 10%, increased the levels of aspartate (AST) and alanine (ALT) transaminase and lactate dehydrogenase (LDH) in serum to 2487, 2189, and 4236 IU/L, respectively, and increased malondialdehyde (MDA) levels in liver to 1.552 nmol/g compared with 1.114 nmol/g in the sham operation group. Pretreatment with GdCl₃ increased the survival rate to 50% and decreased the levels of AST, ALT, and LDH in serum to 1496, 1298, and 3245 IU/l, respectively. Pretreatment with IVIG increased the survival rate to 60% and decreased the levels of AST, ALT, and LDH in serum to 449, 367, and 1456 IU/l, respectively, and the levels of MDA in liver to 1.153 and 1.148 nmol/g for GdCl₃ and IVIG respectively. Histologic examination showed protection of liver parenchyma in the animals treated with GdCl₃ or IVIG.     

Protective effects of thromboxane A2 synthetase inhibitor (OKY-046) on hepatic ischemia-reperfusion injury in rats with obstructive jaundice

The effects of OKY-046, a thromboxane A₂ synthetase inhibitor, on complete hepatic ischemia with obstructive jaundice were studied in male Wistar rats in vivo. The objective of this study was to investigate the influence of OKY-046 (i) on the hepatic microcirculation, as measured by tissue partial oxygen pressure (tPO₂), and (ii) on the release of interleukin-8 (IL-8) in hepatic tissue after reperfusion. Fourteen days after bile duct clamping, the rats were subjected to 30-min warm complete liver ischemia and then reperfusion. The rats were divided into three groups; one group received no treatment (controls, group C), one group received OKY-046 from 15 min before hepatic ischemia to the end of the experiment (group OKY), and the third group received gadolinium chloride (group Gd), injected intravenously to evaluate the contribution of the Kupffer cell to IL-8 production. Group OKY maintained significantly higher tPO₂ levels 5, 10, and 15 min after declamping compared to group C (P<0.05). The IL-8 level in liver tissue in group OKY was lower than that in group C, but the difference was not significant. Group Gd exhibited the lowest IL-8 level of the three groups (P < 0.05). These findings demonstrated that OKY-046 improved the hepatic microcirculation during the reperfusion period, influenced the Kupffer cells in terms of the avoidance of hypoxia, and depressed the concentration of IL-8 in liver tissue.     

麦角新碱预处理对减轻大鼠移植肝缺血再灌注损伤的作用

目的 探讨使用外源性药物麦角新碱预处理对减轻大鼠移植肝缺血再灌注损伤的作用.方法 在大鼠的门静脉-左肾静脉搭桥、肝后下腔静脉内置管分流法自体原位肝移植模型中,于肝门阻断前10 min经大鼠尾静脉注射麦角新碱;观察移植肝缺血前和再灌注后5 min、30 min、2 h时血清一氧化氮(NO)和血浆内皮素1(ET1)水平以及NO/ET1的比值变化;测定血清丙氨酸转氨酶(ALT)酶学差异和肝组织内三磷酸腺苷(ATP)和丙二醛(MDA)含量变化;再灌注2 h取肝组织检测肝细胞、肝小叶超微结构.结果 应用麦角新碱预处理的大鼠移植肝缺血前门脉血浆中ET1升高(P＜0.01),但再灌注后5 min、30 min时,血浆中ET1水平降低(P＜0.05);而缺血前NO/ET1比值降低(P＜0.01),再灌注后5 min时,NO/ET1比值升高(P＜0.01);再灌注后ALT的升高有逐渐降低趋势;再灌注后2 h肝细胞内超微结构的损害程度减轻.结论 使用麦角新碱预处理能减轻大鼠移植肝缺血再灌注损伤.移植肝缺血再灌注损伤的靶细胞是肝血窦内皮细胞,NO/ET1比值平衡可能是影响移植肝微循环血流量变化的调节因素.     

Regulation of pro-inflammatory and anti-inflammatory cytokine responses by Kupffer cells in endotoxin-enhanced reperfusion injury after total hepatic ischemia

The effects of Kupffer cells on cytokine responses in endotoxin-enhanced reperfusion injury after total hepatic ischemia were investigated in this study. Male rats pretreated with either normal saline solution (NS group) or gadolinium chloride (GdCl(3)) to inhibit Kupffer cell function (GC group) were subjected to 60 min of hepatic ischemia. These animals received either normal saline solution or sublethal doses of endotoxin (1 mg/kg) at reperfusion. In the NS group, endotoxin administration induced an enhanced tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 production 1 h after reperfusion with a subsequent peak of macrophage inflammatory protein-2 (MIP-2) levels, which resulted in a 7-day survival rate of 30%. Despite endotoxin administration, GdCl(3) pretreatment significantly suppressed TNF-alpha and increased interleukin-10 production 1 h after reperfusion, which led to a decline in MIP-2 production and amelioration of functional and structural liver damage with a 7-day survival rate of 80%. Augmented pro-inflammatory and anti-inflammatory cytokine responses by Kupffer cells were associated with endotoxin-enhanced reperfusion injury after hepatic ischemia. Kupffer cell blockade has a potential to attenuate the insult via modulation of cytokine responses.     

Evaluation of microperfusion disturbances in the transplanted liver after Kupffer cell destruction using GdCl3: an experimental porcine study

Background

Organ function after liver transplantation is determined by ischemia-reperfusion injury. Destruction of Kupffer cells with gadolinium chloride (GdCl₃) has been shown to have a possible preventive effect on the extent of this injury, which can be extrapolated by analyzing the distribution of hepatic microperfusion. The aim of this study was to evaluate the protective effect of GdCl₃ on disturbances of microperfusion in the transplanted liver.

Methods

Landrace pigs were randomly divided into three groups. In the control group (CG; n = 6) a mapping of the native liver was conducted. For mapping, the four hepatic liver lobes were named from right to left with A to D and every lobe was divided into three vertical segments (cranial, medial, and caudal). In each of these 12 areas, microperfusion was quantified using a thermodiffusion probe (TD [mL/100 g/min]). The other two groups were considered as transplanted treated group (TTG; n = 10) and transplanted nontreated group (TnTG; n = 10). The TTG received an infusion of 20 mg/kg GdCl₃ intravenously 24 hours before organ harvesting. Then standardized orthotopic liver transplantation was performed. In TnTG, standardized orthotopic liver transplantation was carried out without prior GdCl₃ injection. In the recipients, the microperfusion of transplanted livers were mapped in both TnTG and TTG, in two different time points (1 hour [n = 5] and 24 hours (n = 5]) after reperfusion.

Results

A significant reduction of macrophages in the TTG livers in comparison to the CG and TnTG livers was observed (P < .05). However, the number of macrophages in CG and TnTG livers showed no significant difference (P > .05). Regarding liver microperfusion, in TnTG, a marked heterogeneity was detected in the livers after reperfusion. Significant differences between liver lobes (horizontal planes; P = .032) and vertical layers of intralobar liver parenchyma (P = .029) were observed. The same pattern was seen in TTG livers after reperfusion and a significant difference between horizontal (P = .024) and vertical layers (P = .018) of liver tissue were observed. Comparing intralobar regional flow data between vertical planes 24 hours after reperfusion still showed a prominent variation of hepatic tissue perfusion in TnTG livers (P = .028). Within the same horizontal layers, no significant differences between lobes were measured anymore (P = .16). Contrary to TnTG, in TTG, a homogenous pattern of regional liver tissue perfusion was recorded 24 hours after reperfusion. Comparison of TD data on the liver regions showed no significant microperfusion differences in either horizontal (P = .888) or vertical (P = .841) layers.

Conclusions

Application of GdCl₃ resulted in a significant reduction of Kupffer cells. Twenty four hours after transplantation microperfusion showed a homogeneous pattern, which constituted an earlier and better recovery of the transplanted liver. Therefore, destruction of Kupffer cells reduced ischemia-reperfusion injury and seemed to be responsible for the early recovery of microperfusion disturbances and thus for an improvement of graft function.     

Major hepatic trauma: warm ischemic tolerance of the liver after hemorrhagic shock

BACKGROUND: The management of severe hepatic trauma frequently involves exposing the liver to varying periods of warm ischemia. The ischemic tolerance of the liver, in the setting of hemorrhagic shock (HS) and trauma, is presently unknown. We tested the hypothesis that warm ischemic tolerance of the porcine liver will be decreased following resuscitation from HS. MATERIALS AND METHODS: Twenty-three Yorkshire pigs were divided into three groups: 1) hepatic ischemia alone (HI, n = 9); 2) hemorrhagic shock alone (HS, n = 3); and 3) hemorrhagic shock plus hepatic ischemia combined (HSHI, n = 11). Following reperfusion, a liver biopsy was obtained and serial blood chemistries were sampled. RESULTS: Post-operative day 7 mortality was increased in the HSHI group (7/11) compared to the HI (0/9) group, P = 0.038. Notably, deaths did not result from acute liver failure, but rather from intra-operative hemodynamic collapse shortly following hepatic reperfusion. In addition, the HSHI group experienced significantly elevated lactic acid, serum creatinine and liver enzyme levels. Analysis of the liver biopsy samples is consistent with a more severe liver injury in the HSHI group. CONCLUSIONS: The warm ischemic tolerance of the liver following resuscitation from HS is significantly decreased in this porcine model compared to HS or HI alone. Mortality was associated with acute intra-operative hemodynamic collapse occurring shortly after hepatic reperfusion.     

Prevention of ischemia-reperfusion-induced hepatic microcirculatory disruption by inhibiting stellate cell contraction using rock inhibitor

BACKGROUND: We demonstrated that hepatic stellate cells (HSCs) isolated from rat livers exposed to warm ischemia are significantly contractile when compared with HSCs from intact rat livers. This suggests that ischemia-reperfusion (IR)-induced impairment of sinusoidal microcirculation results, at least in part, from contraction of HSCs. METHODS: Rho-associated coiled-coil forming protein serine/threonine kinase (ROCK) is one of the key regulators of HSCs motility. Therefore we investigated whether Y-27632, a p160ROCK-specific inhibitor, has beneficial effects on warm IR injury in an in vivo rat partial liver IR model and a rat orthotopic liver transplantation model. RESULTS: After reperfusion following 90 min of warm ischemia, livers in untreated control rats had persistent congestion and impaired mitochondrial respiration, as demonstrated by increasing deoxy-hemoglobin and reduced cytochrome oxidase contents in the hepatic tissues using in vivo near-infrared spectroscopy. Serum levels of transaminase and endothelin (ET)-1 in these rats were markedly increased 1 hr after reperfusion. In contrast, when Y-27632 (3-30 mg/kg) was administered orally, hepatic tissue contents of deoxy-hemoglobin and cytochrome oxidase rapidly normalized. In such animals, the elevation of serum transaminase levels, but not that of ET-1 levels, was significantly suppressed. This is consistent with in vitro data demonstrating that Y-27632 causes HSCs to undergo relaxation even in the presence of ET-1. Moreover, in a rat orthotopic liver transplantation model, Y-27632 pretreatment dramatically improved the survival of recipients with liver grafts subjected to 45 min of warm ischemia. CONCLUSIONS: Y-27632 attenuates IR-induced hepatic microcirculation disruption by inhibiting contraction of HSCs.     

The effects of intraportal administration of prostaglandin E1 on liver ischemia and hepatectomy in rats

The effects of intraportal administration of prostaglandin E₁ (PGE₁) on portal venous flow, hepatic arterial flow, peripheral tissue blood flow, and systemic arterial flow before and after 60 min total liver ischemia followed by 70% partial hepatectomy in rats were investigated. Total liver ischemia was induced by occluding the hepatoduodenal ligament for 60 min. PGE₁ at a dose of 0.5 μg/kg/min was infused intraportally for 15 min before inducing hepatic ischemia (preischemic period) and for 60 min after ischemia (postischemic reperfusion period) in the treatment group. Normal saline was infused in the control group. Seventy percent partial hepatectomy was performed during ischemia. Serum biochemical analysis and liver tissue histology were carried out 1, 3, and 24 h, and 1 and 24 h after reperfusion respectively. One-week survival of the PGE₁ group was improved to 70% compared to that of the control group of 30%. Postischemia reperfusion values of portal and peripheral tissue blood flows in the PGE₁ group were 6.33 ± 0.600 ml/min and 27.2 ± 23.5 (arbitrary), and were significantly different from those of the control group of 4.34 ± 0.400 ml/min and 23.5 ± 5.54 (arbitrary), respectively. There was no significant difference in hepatic arterial flow between the two groups. Serum alkaline phosphatase decreased significantly in the prostaglandin group. Histological examination revealed a significant portal venous congestion in the control group 1 and 24 h after reperfusion. The extent of the sinusoidal congestion was also severe in the control group 24 h after reperfusion. It was concluded that PGE₁ has a protective effect against liver damage when the liver was injured by warm ischemia and reperfusion followed by partial resection. Received for publication on May 30, 1997; accepted on July 27, 1998     

Intramucosal pH and liver endotoxin clearance during experimental liver transplantation

The study was designed to assess the gastrointestinal ischaemia and the influence of the specific Kupffer cell toxin gadolinium chloride (GdCl₃) on the hepatic and extrahepatic endotoxin [lipopolysaccharide (LPS)] clearance during experimental orthotopic liver transplantation (OLT) in pigs. In eight pig liver transplantations, the donors received 20 mg/kg of GdCl₃ 24 h before explantation, while controls (n = 8) received normal saline. Gastric and sigmoid intramucosal pH (pHi), LPS and endotoxin-neutralising capacity (ENC) levels were measured in the portal vein and superior vena cava after laparatomy, at the end of the anhepatic phase and 1 h after reperfusion. During the anhepatic phase, the sigmoid pHi decreased significantly from 7.32 ± 0.02 to 7.29 ± 0.03 (P < 0.001) and was associated with a substantial increase of portal LPS. Following reperfusion, the systemic LPS concentrations were significantly lower in the pretreated group [39 ± 23 pg/ml (Control); 14 ± 7 (GdCl₃); P < 0.05] suggesting an improved liver LPS clearance [86 % (GdCl₃); 58.2 % (Control); P < 0.05]. This corresponded to an increased ENC in the pretreated group [118 ± 52 ENU/ml (GdCl₃) vs 81 ± 45 ENU/ml (Control); P < 0.05]. The anhepatic phase induced splanchnic ischaemia which correlated with portal endotoxaemia. Donor preconditioning with GdCl₃ leads to lower systemic LPS concentrations in the recipient and increases ENC values in the early phase after OLT. An improved hepatocellular LPS extraction and/or an activation of the extrahepatic reticulo-endothelial system as a result of GdCl₃ treatment is discussed.     

The roles of platelet-activating factor and endothelin-1 in renal damage after total hepatic ischemia and reperfusion

BACKGROUND: This study was designed to verify the involvement of platelet-activating factor (PAF) in renal damage associated with hepatic ischemia and reperfusion (HIR) injury through the release of endothelin (ET)-1 and to determine the modulating effect of a specific PAF receptor antagonist on these insults in rats. METHODS: Male rats pretreated with either normal saline as a vehicle (NS group) or intravenous TCV-309, a PAF receptor antagonist (TCV group), were subjected to 120 min of total hepatic ischemia under an extracorporeal portosystemic shunt. Plasma aspartate transaminase, creatinine, blood urea nitrogen, and ET-1 levels and the relative renal wet weight were determined under nonischemic conditions and at 1, 3, and 6 hr of reperfusion after hepatic ischemia. Changes in mean arterial blood pressure and renal tissue blood flow measurements in the kidney were determined throughout the experiment. RESULTS: Increased plasma aspartate transaminase, creatinine, blood urea nitrogen, and ET-1 levels and the relative renal wet weight after HIR in the NS group were significantly suppressed by TCV-309 pretreatment. Mean arterial blood pressure and renal tissue blood flow after HIR in the TCV group were significantly improved when compared with those in the NS group. These effects resulted in attenuation of structural hepatic and renal damage with the improvement of 7-day survival (62%). CONCLUSIONS: The present study demonstrates that renal damage as well as critical liver injury is produced after reperfusion following 120 min of total hepatic ischemia. A PAF receptor antagonist may be therapeutically useful to protect against these types of damage via indirect modulation of plasma ET-1 levels.     

Involvement of endothelin/ nitric oxide balance in hepatic ischemia/ reperfusion injury

Introduction: Endothelin (ET) and nitric oxide (NO) act as opponents in the regulation of the hepatic microcirculation. During ischemia/reperfusion (I/R) ET levels are increased, whereas no rise in NO levels is observed. This imbalance may be responsible for microcirculatory disturbances. The aim of this study was to restore the delicate ET/NO balance to maintain the integrity of the hepatic microcirculation and to reduce I/R injury. Methods: Ischemia was induced by crossclamping of the hepatoduodenal ligament for 30 min with portal decompression using a splenocaval shunt (56 Wistar rats, 200–250 g). Sham operation, ischemia and treatment groups with the endothelin receptor antagonist (ERA) bosentan (1 mg/kg body weight i.v.) and the NO donor l-arginine (400 mg/kg body weight i.v.) were performed. For assessment of the microcirculation, sinusoidal perfusion rate, diameters of sinusoids and postsinusoidal venules, leukocyte endothelium interactions and velocity of free-flowing leukocytes were investigated by means of in vivo microscopy 30–90 min after reperfusion. Local hepatic tissue pO₂ was measured prior to ischemia, 30 min and 60 min after reperfusion and aspartate aminotransferase (AST)/alanine aminotransferase (ALT) levels were investigated 2 h and 6 h after reperfusion. Results: After ischemia, sinusoidal and venular diameters were reduced to 76% and 85%, respectively, of sham operation group values (P<0.05), but were maintained at baseline in ERA (98/102%) and NO (102/105%) groups (P<0.05). Increased postischemic leukocyte sticking in sinusoids (144%) and venules (435%) was reduced by therapy to 110/253% (ERA) and 111/ 324% (NO), respectively (P<0.05). Perfusion rate was increased to 93% and 94% compared with 82% in the ischemia group (P<0.05). Concomitant with the improved microcirculation in therapy groups, local hepatic tissue pO₂ was improved 30 min after reperfusion in the ERA (11.0 mmHg) and the l-arginine group (11.5 mmHg) relative to the ischemic group (6.9 mmHg) (P<0.05). In addition, postischemic AST/ALT increase was reduced by therapy. Conclusion: Our results indicate that maintenance of ET/NO balance by blocking ET receptors, as well as providing a NO donor, protects the liver microcirculation and reduces hepatic I/R injury. Received: 14 August 1998 Accepted: 13 October 1998     

Different protection mechanisms after pretreatment with glycine or alpha-lipoic acid in a rat model of warm hepatic ischemia

BACKGROUND/AIM: Alpha-lipoic (LA) acid pretreatment has previously been described to reduce ischemia/reperfusion injury (IRI) after warm liver ischemia, whereas glycine pretreatment has been shown to be protective mostly in models of cold hepatic ischemia. The aim of this study was to determine whether glycine decreases IRI after warm hepatic ischemia. Furthermore we investigated whether doses of LA other than those used previously are also protective against IRI after warm hepatic ischemia. METHODS: Selective liver ischemia was maintained over a period of 90 min. In long-term as well as short-term experiments we studied IRI in several groups comparing animal survival as the pivotal endpoint. RESULTS: Animal survival was improved by glycine and 5,000 micromol LA, whereas all animals died within 3 days after pretreatment with 50 micromol LA. In the glycine group we observed a tendency towards decreased apoptosis-related cell death measured by the activity of caspase-3 in liver tissue and the percentage of TUNEL-positive hepatocytes in comparison to the untreated group. Serum alpha-glutathione S-transferase, lipid peroxidation, and caspase-3 activity as well as the percentage of TUNEL-positive hepatocytes and the percentage of liver necrosis were only significantly decreased by 5,000 micromol LA pretreatment. Liver tissue levels of tumor necrosis factor (TNF)alpha were reduced only in the glycine group whereas TNFalpha was increased in the untreated as well as the LA group. Levels of TNFalpha mRNA were upregulated in both the glycine- and LA-pretreated groups. CONCLUSION: Our data show that increased animal survival by glycine was accompanied by a reduced TNFalpha content in liver tissue. Protection by glycine is likely to result from a reduction in adverse TNFalpha effects. Administration of high-dose LA on the other hand led to a significant reduction in necrosis- and apoptosis-related cell death in IRI of the liver without a reduction in liver TNFalpha.     

山莨菪碱对大鼠肝脏缺血再灌注损伤保护作用的实验研究

目的 探索山莨菪碱对大鼠肝脏缺血再灌注损伤的保护作用。方法 选雄性Wistar大鼠160只,分为正常对照组、缺血再灌注组、生理盐水组和山莨菪碱组,观察了肝脏缺血60分钟及再灌注1、3、6、12及24小时后血浆内皮素－1（ET－1）、透明质酸（HA）和谷丙转氨酶（ALT）含量变化及肝组织病理学变化。结果 肝脏缺血再灌注后,血浆ET－1、HA和ALT含量均明显增高,同时肝脏瘀血很明显,肝脏缺血再灌注前用山莨菪碱后,血浆HA和ALT含量明显降低,同时肝组织瘀血减轻。结论 山莨菪碱可改善再灌注后的肝脏微循环障碍,对大鼠肝脏缺血再灌注损伤有保护作用。     

Effect of taurine on IRAK4 and NF-kappa B in Kupffer cells from rat liver grafts after ischemia-reperfusion injury

BackgroundThe aim of this study was to explore the protective mechanisms of taurine pretreatment against hepatic ischemia/reperfusion injury after liver transplantation.MethodsA Sprague-Dawley–to–Sprague-Dawley rat liver transplantation model was used in this study. At 0, 60, and 180 minutes after reperfusion, expression of interleukin-1 receptor–associated kinase-4 (IRAK-4) messenger ribonucleic acid and protein in Kupffer cells was determined by real-time polymerase chain reaction and Western blotting. The activity of nuclear factor κB in Kupffer cells was determined by electrophoretic mobility shift assay. The serum tumor necrosis factor–α level was detected by enzyme-linked immunosorbent assay. Serum transaminases, liver histology, and animal survival were also investigated.ResultsAt 60 and 180 minutes after reperfusion, levels of IRAK-4 messenger ribonucleic acid and protein, activities of nuclear factor κB, and levels of serum transaminases and tumor necrosis factor–α were all obviously elevated. However, changes in these parameters in rats treated with taurine were remarkably attenuated at the indicated time points.ConclusionsThese data suggest that taurine could protect against hepatic ischemia/reperfusion injury after liver transplantation, and the protective effects may be through downregulation of IRAK-4 and downstream nuclear factor κB and tumor necrosis factor–α expression in Kupffer cells.     

Hepatic stellate cell activation and hepatic fibrosis induced by ischemia/reperfusion injury

Objective

Warm ischemia causes severe allograft damage in liver transplantation. However, the long-term effects of ischemia/reperfusion injury (IRI) on fibrosis have not been fully elucidated. In this study, we used a partial warm hepatic ischemia mouse model to monitor fibrosis in the ischemic liver.

Materials and Methods

Male BALB/c mice were divided into ischemic and sham groups (n = 30/group). Via a midline laparotomy, an atraumatic clip was used to interrupt the arterial and the portal venous blood supply to the left liver lobe. After 90 minutes of partial hepatic ischemia, the clip was removed initiating hepatic reperfusion. Samples from normal, sham, and ischemic liver tissues were collected at intervals of 1, 5, 10, 15, 20, or 30 days after operation (n = 5 for each time point) for hematoxylin-eosin (H&E), Mallory's trichrome, and alpha-smooth muscle actin (α-SMA) immunohistochemical stains for fibrosis and activation of hepatic stellate cell (HSCs).

Results

IRI caused significant HSC activation in the ischemic liver tissues. Mallory's trichrome stain demonstrated that IRI caused hepatic parenchymal fibrosis near portal tracts and central veins. With prolonged reperfusion time hepatic parenchymal fibrosis was aggravated, showing the same pattern of HSC activation. IRI also caused increased portal tract fibrosis in ischemic liver tissues, especially around biliary tracts.

Conclusions

Hepatic IRI caused HSC activation, increasing hepatic parenchymal and portal tract fibrosis in ischemic liver tissues.     

ATP-MgCl2 produces sustained improvement in hepatic mitochondrial function and blood flow after hepatic ischemia

Recent studies have shown that infusion of ATP-MgCl₂ following hepatic ischemia significantly improved mitochondrial function and hepatic blood flow 1 hr after treatment. To determine if the improvement in the above parameters by ATP-MgCl₂ is short-lived or whether it persists for prolonged periods of time after treatment, hepatic ischemia in rats was produced for 90 min followed by reperfusion. The rats then received iv 0.5 ml of saline or ATP-MgCl₂ (12.5 μmole each). Twenty-four hours after reflow, hepatic blood flow was measured by H₂ polarography following which the animals were sacrificed and hepatic mitochondria isolated. The results indicated that 24 hr after reflow, mitochondrial state 3 respiration, respiratory control ratio, adenine nucleotide translocase activity, ATP synthetic activity, and hepatic blood flow were depressed by approximately 50% in animals which were treated with saline after hepatic ischemia. In addition, there was a fourfold increase in mitochondrial free fatty acid levels of such animals. Animals which were treated with ATP-MgCl₂ following hepatic ischemia showed significantly improved mitochondrial function (used as an index of cellular recovery) and hepatic blood flow. These results in conjunction with previous results suggest that infused ATP-MgCl₂ improves mitochondrial function and blood flow and that these effects persist even 24 hr after administration of ATP-MgCl₂. Thus, infusion of ATP-MgCl₂ following severe ischemia produces sustained improvement in cellular function.     

Advantage of ischemic preconditioning for hepatic resection in pigs

BACKGROUND: Ischemic preconditioning (IP) and intermittent inflow occlusion (IO) have provided beneficial outcomes in hepatic resection. However, comparison of these two procedures against warm hepatic ischemia-reperfusion injury has not been studied enough. MATERIALS AND METHODS: Pigs that had undergone 65% hepatectomy were subjected to Control (120 min continuous ischemia, n = 6), IP (10 min ischemia and 10 min reperfusion, followed by 120 min continuous ischemia, n = 6), and IO (120 min ischemia in the form of eight successive periods of 15 min ischemia and 5 min reperfusion, n = 6). We evaluated hepatocyte injury by aspartate aminotransferase, lactate dehydrogenase and hepaplastin test, hepatic microcirculation by hepatic tissue blood flow (HTBF) and endothelin (ET)-1, inflammatory response by tumor necrosis factor-alpha (TNF-alpha), and histopathology after reperfusion. RESULTS: IP prevented hepatocyte injury, HTBF disturbance, and hepatocyte necrosis in histopathology as well as IO. These two groups showed significantly better outcomes than Control. IP produced significantly less ET-1 and TNF-alpha than IO. CONCLUSIONS: IP ameliorated hepatic warm ischemia-reperfusion injury. Furthermore, IP gained more advantages in preventing chemokine production such as ET-1 and inflammatory response over IO. IP could take the place of IO for hepatectomy.     

The effect of FK506 on Warm Ischemia and reperfusion Injury in the Rat Liver

The protective effect of FK506 on hepatocytes against ischemia and reperfusion injury was examined by evaluating the following: the high energy phosphorus metabolism obtained using 31P magnetic resonance spectroscopy (³¹P-MRS) and the tissue blood flow of the liver in ischemia and the reperfusion process, mitochondrial glutamic oxaloacetic transaminase (m-GOT) and glutamic pyruvic transaminase (GPT), the survival rates of the animals, a histological study and immunohistological staining for intercellular adhesion molecule-1 (ICAM-1) in the liver after ischemia. The rats were treated with FK506 1 mg/kg/day i.m. for 4 days before testing. Ischemia was induced by clamping the hepatoduodenal ligament for 30 min. In³¹P-MRS, the recovery of the hepatic energy status after ischemia, evaluated by -ATP/inorganic phosphate (Pi), was significantly better in the FK506 group. It also coincided with the recovery of tissue blood flow monitored with a laser Doppler flowmeter. In the histological examination, the congestion observed in the periportal region of the control group was mild, while there was less induction of ICAM-1 in the endothelial cells of the portal veins and hepatic veins in the FK506 group. From these findings, we concluded that FK506 had a protective effect on hepatocytes against warm ischemia and reperfusion injury, and the mechanism for this could partially be attributed to improved tissue blood flow after ischemia by the modulation of immunological events.