L-carnitine could not improve hepatic warm ischemia-reperfusion injury despite ameliorated blood flow

BACKGROUND: Carnitine is applied to ameliorate ischemia-reperfusion (I/R) injury of several organs. However, application to hepatic I/R injury is not frequently reported. The aim of this study was to elucidate the effect of exogenous carnitine administration to ameliorate the warm hepatic I/R injury. MATERIALS AND METHODS: Male Wistar rats were divided into two groups, a carnitine group (Car);100 mg/kg of l-carnitine administration and a control group (C); vehicle administration. Thirty minutes after administration, the left hepatic lobes were given 60-min ischemia and then reperfused. Plasma alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutamate dehydrogenase (GLDH), tumor necrosis factor (TNF)-alpha, and lipoperoxides (LPO) were measured. Hepatic adenosine triphosphate (ATP) concentration was also measured. The hepatic blood flow was estimated using a Laser Doppler. The presence of apoptosis in the livers was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. RESULTS: In group Car, the blood flow of the left hepatic lobes was better recovered during the reperfusion period than in group C (P < 0.0001). Plasma levels of ALT, AST, GLDH, and TNF-alpha at 1 h after reperfusion were not significantly different between the groups. Although there were no statistical significances, ALT, AST, and TNF-alpha levels in group Car at 24 h after reperfusion tended to be higher than in group C. Plasma LPO levels were not different between the two groups. Also hepatic ATP concentration was not different between the two groups. TUNEL positive liver cells were visible only in group Car at 24 h after reperfusion, but not in the controls. CONCLUSIONS: Although carnitine administration improved the hepatic blood flow during the reperfusion period, we could not demonstrate a protective effect to the hepatic warm I/R injury.     

Effects of ischemic preconditioning on regenerative capacity of hepatocyte in the ischemically damaged rat livers

BACKGROUND: Liver regeneration after partial hepatectomy is regulated by several factors that activate or inhibit hepatocyte proliferation. A short period of ischemia-reperfusion (IR), called ischemic preconditioning (IPC), protects the liver against subsequent sustained ischemic insults. The present study investigated the effects of IPC on liver regeneration after partial hepatectomy under IR in rats. MATERIALS AND METHODS: Male Wistar rats were subjected to 45 min of total hepatic ischemia, and 70% hepatectomy was performed just before reperfusion. Animals were pre-treated with either IPC (10/15 min) (IPC + PHx group) or not (ischemia + PHx). The survival rate, serum transaminases, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-6 levels, hepatocyte proliferation and histological change of the remnant liver were measured in both groups and compared with non-ischemic controls subjected to 70% hepatectomy alone (PHx group). RESULTS: The survival rate was significantly better in the IPC + PHx group than in the ischemia + PHx group. Furthermore, IPC reduced liver injury determined by liver histology and serum transaminases. There was an early rise in serum TNF-alpha and IL-6 levels in the ischemia + PHx group. Compared with non-ischemic controls, IPC significantly decreased TNF-alpha, but not IL-6 during the late (24 and 48 h) phases of reperfusion. Rats subjected to 70% hepatectomy and 45 min of hepatic ischemia showed significantly reduced hepatocyte proliferation (mitotic index, proliferating cell nuclear antigen, and relative liver weight) when compared with animals subjected to hepatectomy alone. However, hepatocyte proliferation was markedly increased in rats pretreatment with IPC when compared with ischemic controls. CONCLUSION: These results suggest that ischemic pre-conditioning ameliorates the hepatic injury associated with ischemia-reperfusion and has a stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective modality. Il-6 appears to be key mediator in promoting regeneration after combined ischemia and hepatic resection.     

Vitamin E Succinate Reduces Ischemia/Reperfusion Injury in Steatotic Livers

Steatotic livers represent a growing proportion of marginal organs available for transplantation. These livers are highly prone to primary nonfunction following transplantation and are therefore routinely turned down for surgery. Given the elevated levels and sensitivity for reactive oxygen species (ROS) in these livers, we evaluated whether pretreatment with a targeted ROS scavenger, vitamin E succinate, increased survival and decreased injury after ischemia/reperfusion (I/R). For this study, ob/ob mice received 50 IU/d vitamin E succinate in supplemented vs control chow for 7 days, and were subjected to 15 minutes of total hepatic ischemia and 24 hours of reperfusion. Treatment resulted in a 5-fold decrease in serum alanine aminotransferase (ALT) levels after reperfusion, mirrored by significant decreases in hepatocellular necrosis. These results suggested that targeted antioxidants such as vitamin E succinate may prove to be highly applicable for the pretreatment of steatotic donor livers, increasing their tolerance for I/R and the transplantation process.     

Impact of endothelin-1 on microcirculatory disturbance after partial hepatectomy under ischemia/reperfusion in thioacetamide-induced cirrhotic rats

BACKGROUND: Endothelin (ET)-1 contributes to hepatic ischemia and reperfusion (HIR) injury in normal liver. This study was conducted to clarify the role of ET-1 in HIR injury in cirrhotic state. MATERIALS AND METHODS: Using thioacetamide-induced cirrhotic rats with spontaneous portosystemic shunt, we determined the changes in plasma aspartate aminotransferase (AST) levels, plasma and hepatic ET-1 values, 7-day survival rates, and hepatic oxygen saturation (SO(2)) by time-resolved spectroscopy as an indicator of hepatic microcirculation under intermittent or continuous total hepatic ischemia with subsequent partial hepatectomy. RESULTS: Hepatic ET-1 levels in cirrhotic rats were significantly higher than those in noncirrhotic rats. Plasma and hepatic ET-1 levels at 1, 3 and 6 h of reperfusion after intermittent hepatic ischemia were significantly lower than those after continuous hepatic ischemia. In cirrhotic animals subjected to intermittent hepatic ischemia, the elevation of plasma AST levels at 1, 3 and 6 h of reperfusion and the decline in hepatic SO(2) at the end of 60-min hepatic ischemia and after reperfusion were significantly suppressed when compared with those subjected to continuous hepatic ischemia. Pretreatment with a nonselective endothelin receptor antagonist in continuous hepatic ischemia significantly ameliorated plasma AST levels and hepatic SO(2) values with less hepatic sinusoidal congestion, resulting in an improvement in the 7-day survival rate. CONCLUSIONS: Continuous hepatic ischemia in the cirrhotic liver has disadvantages relating to microcirculatory derangement with more ET-1 production in partial hepatectomy. In liver surgery, pharmacological regulation of ET-1 production may lead to attenuation of reperfusion injuries for ischemically damaged cirrhotic liver.     

Ischemic preconditioning prevents apoptotic cell death and necrosis in early and intermediate phases of liver ischemia-reperfusion injury in rats.

Ischemic preconditioning (IPC) may be useful in attenuating the hepatic ischemia reperfusion (IR) syndrome by means of improving cell resistance to anoxia and reoxygenation and preventing cell death. Since there are insufficient data available regarding the chronology of preconditioning effects, we investigated the role of IPC, to test the hypothesis that liver protection would occur during the early and intermediate phases of the reperfusion period. Wistar rats (n = 72) were randomly assigned into six experimental groups, 12 animals each. A 40-min ischemia to the left lateral and median liver lobes was induced by selective hepatic pedicle clamping followed by 30 min or 240 min of reperfusion (IR30 and IR240). IPC groups (IPC30 and IPC240) underwent a 10 min of ischemia followed by 10 min of reperfusion preceding the definitive 40-min ischemic period. Sham-operated animals were followed for 30 and 240 min. Hepatic enzymes and histological evaluation were performed after the reperfusion period. Hepatic ischemia-reperfusion (IR30 and IR240) induced marked increases in liver enzymes levels after 30 min and particularly after 240 min. IPC effectively attenuated those enzymatic increases. Microvesicular steatosis was observed after 30 min, but not 240 min, of reperfusion in both IPC and IR livers. Necrosis was detected in 66.7% of IR240 and only in 8.3% of IPC240. Both hepatocyte and sinusoidal apoptosis were markedly attenuated by IPC. We conclude that IPC provided protection against hepatic ischemia reperfusion injury in early and intermediate phases of the reperfusion period, reducing hepatic enzymatic leakage and ameliorating hepatic apoptosis and necrosis.     

FR167653 improves survival and pulmonary injury after partial hepatectomy under ischemia/reperfusion in rats.

BACKGROUND: FR167653 is a potent suppressant of production of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1 beta, which play an important role in hepatic and pulmonary injury due to ischemia/reperfusion of the liver and in liver regeneration after hepatectomy. We examined the effects of FR167653 on hepatectomy under ischemia/reperfusion in rats. METHODS: After initial 15-min ischemia and 5-min reperfusion, 70% hepatectomy was performed during the second 15-min ischemia period in FR167653-treated (FR group) and saline-treated (saline group) rats. The survival rate, relative liver weight, TNF-alpha, IL-1 beta, DNA synthesis rate of the remnant liver, and histological change and adhesion molecule (ICAM-1) of the lung were examined. Serum glutamic pyruvic transaminase and hepatic malondialdehyde were also measured. RESULTS: Expressions of TNF-alpha and IL-1 beta in the remnant liver were significantly inhibited in the FR group compared to the saline group. The survival was significantly better and pulmonary damage was less in the FR group after hepatectomy under ischemia/reperfusion. ICAM-1 expression of the lung was not altered after hepatectomy and was not significantly different between the two groups. Liver regeneration and injury were not significantly different between the two groups. CONCLUSION: FR167653 does not affect liver injury and regeneration after hepatectomy under ischemia/reperfusion, while it ameliorates pulmonary injury and improves the survival.     

不同预处理对肝硬化兔缺血再灌注肝组织HSP70及ET-1表达的影响

目的:探讨两种预处理方式,即经典缺血预处理(IPC)与肢体缺血预处理(LIPC),对肝硬化兔肝缺血再灌注(I/R)损伤的保护作用及可能的作用机制。方法:皮下注射CCl4-橄榄油溶液制备兔肝硬化模型,随后将模型兔随机分为假手术组,肝I/R组(I/R组),IPC+肝I/R组(IPC组),LIPC+肝I/R组(LIPC组),每组7只。肝I/R模型制作方法:阻断入肝血流30 min,再灌注2 h;IPC诱导方法:在行肝I/R处理前阻断入肝血流10 min,开放10 min;LIPC诱导方法:在行肝I/R处理前24 h,采用止血带捆扎兔单侧后肢5 min,再开放5 min,重复3次。各组于再灌注2 h后切取肝组织,行组织形态学观察,用ELISA法测定内皮素1(ET-1)含量及Western blot法检测热休克蛋白(HSP70)的表达。结果:与假手术组比较,其余各组在肝硬化病变的基础上均出现不同程度的变性、水肿和炎性细胞浸润,但IPC组与LIPC组明显轻于I/R组,而LIPC组及IPC组间病变程度无明显差异;与假手术组比较,其余各组肝组织ET-1含量和HSP70的表达均明显增加(均P<0.05),但IPC组与LIPC组肝组织ET-1含量低于I/R组,HSP70的表达高于I/R组(均P<0.05),而上述2项指标在LIPC及IPC组间均无统计学差异(均P>0.05)。结论:LIPC和IPC均能对肝硬化肝I/R损伤有保护作用,且保护强度相似,其机制可能均与抑制ET-1的释放及增加HSP70的表达有关;LIPC具有无创性,可能具有更大的临床应用前景。     

Beneficial effects of gaseous hydrogen sulfide in hepatic ischemia/reperfusion injury

Hydrogen sulfide (H₂S) can induce a reversible hypometabolic state, which could protect against hypoxia. In this study we investigated whether H₂S could protect livers from ischemia/reperfusion injury (IRI). Male C57BL/6 mice were subjected to partial hepatic IRI for 60 min. Animals received 0 (IRI) or 100 ppm H₂S (IRI + H₂S) from 30 min prior to ischemia until 5 min before reperfusion. Core body temperature was maintained at 37 °C. Animals were sacrificed after 1, 6 or 24 h. Hepatic ischemia caused extensive hepatic necrosis in the IRI animals which coincided with an increase in ALT and AST serum levels. Animals treated with H₂S showed attenuated serum ALT and AST levels and reduced necrotic lesions after 24 h. IRI animals had increased Bcl‐2 mRNA expression and increased active Caspase 3 protein, which were both significantly lower in H₂S treated animals. Increased TNFα and IL‐6 mRNA in the IRI livers was significantly attenuated by H₂S treatment, as was hepatic influx of Ly‐6G positive granulocytes. Hepatic superoxide production after ischemia was attenuated by H₂S treatment. In hepatic ischemia/reperfusion injury, gaseous H₂S treatment is highly protective, substantially reducing necrosis, apoptosis and inflammation. Gaseous H₂S is therefore a very promising treatment for reducing IRI during hepatic transplantation.     

FK 506 ameliorates normothermic liver ischemia in rats by suppressing production of tumor necrosis factor

In recent years, there has been growing evidence that tumor necrosis factor-± (TNF) plays an important role in the development of hepatic injury after ischemia-reperfusion. We have previously demonstrated that the immunosuppressants, cyclosporine, azathioprine and FK 506 (FK), have a protective effect on warm ischemic injury of the rat liver. In the present study, we attempted to elucidate the mechanism for the beneficial effect of FK on liver ischemia, with special reference to the suppression of TNF production. After 60 min and 90 min of warm liver ischemia, the survival rates were significantly improved by FK pretherapy. This was associated with amelioration of hepatic injury, as assessed by histological examinations and determinations of serum AST and lipid peroxide levels in the liver. After 60 min of liver ischemia, TNF was measurable during the reperfusion period in the sera of the control animals, peaking of 6 h after reperfusion (123 ± 15.8 pg/ml, mean SEM). In contrast, pretreatment with FK significantly suppressed the elevation of serum TNF levels at the same time point (75.8 ± 13.1 pg/ml, P < 0.05). The present data showed that liver ischemia-reperfusion resulted in TNF production, and that FK could protect the liver from reperfusion injury by suppressing this production of TNF.     

Sequential periods of preconditioning decrease laparoscopy-related elevations in hepatic TNF-alpha and IL-6 levels in rats

BACKGROUND: Laparoscopy induces an ischemia/reperfusion (I/R) injury that elicits the production of inflammatory cytokines. The aim of this study was to test whether laparoscopic preconditioning (LP) could change hepatic inflammatory response. MATERIALS AND METHODS: Twenty-four male Spraque Dawley rats were assigned into one of three experimental groups: the control group (C) was subjected to sham operation; the I/R group was subjected to 60 minutes of pneumoperitoneum followed by 45 minutes of desufflation; and the ischemic preconditioning (IP) group was subjected to preconditioning prior to the induction of I/R. Preconditioning was defined as two cycles of 5 minutes of pneumoperitoneum followed immediately by 5 minutes of desufflation. Hepatic tumor necrosis factor alpha (TNF-alpha) and interleukin (IL)-6 levels were evaluated as an inflammatory response. RESULTS: Hepatic TNF-alpha levels increased in the I/R group compared with the C group (9.64 +/- 0.77 pg/mg protein vs. 6.53 +/- 0.48 pg/mg protein, P < 0.01). The decreased TNF-alpha levels in the IP group were not statistically significant when compared to the I/R group (8.09 +/- 0.74 pg/mg protein). Hepatic IL-6 levels increased in the I/R group compared to the C group (4.17 +/- 0.31 pg/mg protein vs. 2.93 +/- 0.20 pg/mg, P < 0.05). IP reduced the hepatic IL-6 levels significantly compared to the I/R group (3.31 +/- 0.22 pg/mg protein vs. 4.17 +/- 0.31 pg/mg protein, P <0.05). CONCLUSION: Laparoscopy induces cytokine response in various organs including the liver. LP could alter the production of cytokines prior to sustained laparoscopic procedures. Preconditioning may be advisable, especially for elderly or other patients with hepatic, renal, or cardiac dysfunction. Further studies are needed to adapt this concept to clinical settings.     

Amelioration of tumor necrosis factor release by cyclosporine in warm ischemia/reperfusion injury of the rat liver: with special reference to hepatic ultrastructure

Mechanisms by which an immunosuppressant (cyclosporine, CsA) ameliorates warm ischemic injury of the liver were studied. Female Sprague-Dawley rats were subjected to 60-min normothermic liver ischemia. Animals were assigned to one of two groups: group I, controls with vehicle treatment; group II, treatment with CsA (10 mg/kg). CsA was given orally for 4 consecutive days prior to the induction of hepatic ischemia. In addition to a survival study, plasma levels of endotoxin, serum activity of tumor necrosis factor-α (TNF), and serum levels of aminotransferases were measured in blood samples collected from the suprahepatic vena cava, and hepatic ultrastructural alterations were examined under an electron microscope. The 7-day survival rate was significantly higher in the CsA-treated animals. In the control group, serum TNF levels were elevated following reperfusion and peaked at 3 h. When the values at 3 h post reflow were compared, the animals given CsA had significantly lower levels of TNF (170.0 ± 30.5 pg/ml for group I, 67.6 ± 13.7 for group II, mean ± SEM; P < 0.05). The sinusoidal lining cells and hepatocytes were drastically destroyed at 6 h post reflow in the control group, although the degree of injury at 1–3 h was less severe. On the other hand, the endothelium and parenchymal liver cells in the CsA-treated group were well preserved at 6 h in comparison with those in the control group. Our data suggest that modulation of TNF production is one of the mechanisms through which CsA prevents the exacerbation of ischemia/reperfusion injury of the liver. Received for publication on Aug. 12, 1998; accepted on Feb. 2, 1999     

Prevention of I/R injury in fatty livers by ischemic preconditioning is associated with increased mitochondrial tolerance: the key role of ATPsynthase and mitochondrial permeability transition

Ischemia/reperfusion (I/R) injury is a commonly encountered clinical problem and occurs probably as a consequence of irreversible mitochondrial injury. The increased susceptibility of fatty livers to ischemic injury is associated with depletion of adenosine triphosphate (ATP) content, which is preserved by preconditioning. Mitochondria being the main ATP production source for the cell, we aimed to evaluate whether ischemic preconditioning (IPC) of fatty livers prevents the impairment in mitochondrial function induced by I/R. Lean and steatotic animals were subjected to 90 min of hepatic warm ischemia and 12 h of reperfusion. IPC effect was tested in fatty livers. After reperfusion, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured. Mitochondrial membrane potential, mitochondrial respiration and susceptibility to mitochondrial permeability transition (MPT) were evaluated, as well as ATPase activity and adenine nucleotides. IPC of fatty livers decreased serum AST and ALT levels. Fatty animals subjected to I/R exhibited decreased mitochondrial membrane potential and a delay in the repolarization after a phosphorylation cycle, associated with increased state 4 respiration. Increased tolerance to MPT induction, preservation of F₁F_o-ATPsynthase activity and mitochondrial bioenergetics were observed in ischemic preconditioned fatty livers. Thus, IPC is an endogenous protecting mechanism that preserves mitochondrial function and bioenergetics in fatty livers.     

Role of anti-tumor necrosis factor-alpha in ischemia/reperfusion injury in isolated rat liver in a blood-free environment

BACKGROUND: Warm ischemia/reperfusion injury during liver transplantation is the most important cause of primary nonfunction of liver allografts. Tumor-necrosis factor (TNF)-alpha apparently mediates tissue damage by inducing apoptosis and/or necrosis in liver transplants. The aim of the study was to determine, using an isolated rat liver model, if pretreatment with anti-TNF-alpha monoclonal antibodies can attenuate ischemia/reperfusion liver injury. Specifically, its effect on liver cell apoptosis through the modulation of caspase activity was examined in a blood-free environment. METHODS: Isolated rat livers were perfused with Krebs-Henseleit solution and randomly divided into three groups: (1) continuous perfusion for 165 min (control); (2) perfusion for 90 min, break for 60 min (ischemia), and reperfusion for 15 min; (3) as with group 2, but with administration of monoclonal mouse anti-rat TNF-alpha monoclonal antibodies before induction of ischemia. Caspase-3- and -9-like activity was measured by fluorometric assay, and apoptotic cells were identified by morphological criteria and application of the terminal deoxnucleotidyl transferase-mediated dUTP nick-end-labeling (Tunel) assay. RESULTS: Portal pressure increased significantly in group 2 (14.8+/-2.3 mm Hg) compared to group 3, which showed no change (P<0.05). Significant amounts of TNF-alpha were detected in the effluent in group 2 at 1 min of reperfusion (147+/-8.9 pg/ml) compared to group 3 (30+/-6.7 pg/ml, P<0.05). Statistically significant reductions in liver enzyme levels were also noted in the animals pretreated with TNF-alpha antibodies (P<0.02). Caspase-3 and -9 activity was significantly decreased (270 and 160%, respectively) in group 3 compared to group 2 (P<0.005 and <0.05, respectively). A significant reduction in postischemic hepatic injury was noted on Tunel assay: many apoptotic hepatocyte cells were detected in group 2 but not in livers pretreated with monoclonal mouse anti-TNF-alpha antibodies (group 3). CONCLUSIONS: Neutralization with specific monoclonal antibodies against TNF before ischemia induction can attenuate postischemic hepatic injury. Apoptotic injury seems to be ameliorated through modulation of caspase-3- and -9-like activity.     

Effect of ischemic preconditioning in whole liver transplantation from deceased donors. A pilot study.

The effect of ischemic preconditioning (IPC) in orthotopic liver transplantation (OLT) has not yet been clarified. We performed a pilot study to evaluate the effects of IPC in OLT by comparing the outcomes of recipients of grafts from deceased donors randomly assigned to receive (IPC+ group, n = 23) or not (IPC- group, n = 24) IPC (10-min ischemia + 15-min reperfusion). In 10 cases in the IPC+ group and in 12 in the IPC- group, the expression of inducible nitric oxide synthase (iNOS), neutrophil infiltration, and hepatocellular apoptosis were tested by immunohistochemistry in prereperfusion and postreperfusion biopsies. Median aspartate aminotransferase (AST) levels were lower in the IPC+ group vs. the IPC- group on postoperative days 1 and 2 (398 vs. 1,234 U/L, P = 0.002; and 283 vs. 685 U/L, P = 0.009). Alanine aminotransferases were lower in the IPC+ vs. the IPC- group on postoperative days 1, 2, and 3 (333 vs. 934 U/L, P = 0.016; 492 vs. 1,040 U/L, P = 0.008; and 386 vs. 735 U/L, P = 0.022). Bilirubin levels and prothrombin activity throughout the first 3 postoperative weeks, incidence of graft nonfunction and graft and patient survival rates were similar between groups. Prereperfusion and postreperfusion immunohistochemical parameters did not differ between groups. iNOS was higher postreperfusion vs. prereperfusion in the IPC- group (P = 0.008). Neutrophil infiltration was higher postreperfusion vs. prereperfusion in both groups (IPC+, P = 0.007; IPC-, P = 0.003). Prereperfusion and postreperfusion apoptosis was minimal in both groups. In conclusion, IPC reduced ischemia/reperfusion injury through a decrease of hepatocellular necrosis, but it showed no clinical benefits.     

Effect of ischemic preconditioning on hepatic tolerance to cold ischemia in the rat

Abstract A short warm ischemia before reperfusion has been shown to improve the tolerance of the heart and the liver to a prolonged warm ischaemia. The present experimental study was conducted to evaluate the effect of such preconditioning on hepatic tolerance to an extended cold ischemia. In a model of isolated perfused liver, livers from Wistar rats (250–350 g) were stored for 24 h in UW (4°C) immediately after harvesting and reperfused for 3 h at 37°C. Control livers subjected to a 24-h cold ischemia were compared to livers subjected to preconditioning (defined as a 5- or 10-min clamping of the hepatic pedicle followed by a 10-min reperfusion before liver harvesting) prior to the definitive 24-h cold ischemia. While there was no difference in bile production between the preconditioned groups and the controls, transaminases and LDH release was significantly increased, vascular resistance was enhanced, and preservation injury was more extensive in both preconditioned groups as compared to controls. In contrast to the beneficial effect reported on prolonged warm ischaemia, preconditioning has a deleterious effect on hepatic tolerance to an extended cold ischemia.     

缺血预适应对老年大鼠缺血-再灌注心肌的影响

目的探讨缺血预适应（ischemic preconditioning,IPC）对老年大鼠缺血-再灌注损伤（I/R）后心肌的影响。方法取Wistar大鼠56只,其中21~23月龄（老年鼠）和4~5月龄（青年鼠）各28只,建立离体心脏Langendorff灌注模型,按随机数字表法分为7组（每组8只）：青年对照组、青年I/R组、青年IPC组、老年对照组、老年I/R组、老年IPC组、老年强化IPC组。对照组采用全心灌流90 min,不做任何处理;I/R组采用心脏平衡灌流30min后,缺血30 min,再复灌30     

Akt activation protects rat liver from ischemia/reperfusion injury

BACKGROUND: Apoptosis as well as necrosis may play an important role in hepatic ischemia/reperfusion (I/R) injury. Akt, a serine-threonine protein kinase, is known to promote cell survival. We investigated whether gene transfer of constitutively active or dominant negative Akt could affect hepatic I/R injury. MATERIALS AND METHODS: Hepatic I/R injury was induced in rats by Pringle's maneuver for 20 min followed by reperfusion. Adenoviruses encoding a constitutively active form of Akt (myrAkt), a dominant negative form of Akt (dnAkt), or beta-galactosidase (LacZ) were injected through the tail vein 72 h before hepatic I/R. RESULTS: Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) staining demonstrated a significant increase in the positive cells 240 min after reperfusion. Immunoblotting with phospho-Akt antibody showed phosphorylation of Akt from 90 to 180 min after reperfusion. The expression of myrAkt reduced the number of TUNEL-positive cells and hepatic necrosis around the central veins in the liver after reperfusion. This expression also significantly inhibited the increase in serum alanine aminotransferase (297 +/- 131 IU/L, P < 0.05) 120 min after I/R, compared with increases in uninfected (1761 +/- 671 IU/L), LacZ adenovirus (1528 +/- 671 IU/L)-, and dnAkt adenovirus (1342 +/- 485 IU/L)-infected rats. MyrAkt expression phosphorylated Bad and inhibited the release of cytochrome-c after reperfusion. No difference in nuclear translocation of nuclear factor (NF)-kappaB, p65 was seen among the three groups of rats, however. CONCLUSION: Adenoviral gene transfer of myrAkt could inhibit apoptotic cell death and subsequent hepatic I/R injury in the rat, through Bad, not NF-kappaB.     

The role of ischemic preconditioning in rat liver graft

OBJECTIVE: The objective of this study was to investigate the protective effects of different modes of ischemic preconditioning (IPC) on an ischemia/reperfusion (I/R) injury in rat liver graft. METHODS: A total of 192 Wistar rats were randomly allocated into 4 groups, each including 48 rats: control group (C), experimental group 1 (E(1)), experimental group 2 (E(2)), and experimental group 3 (E(3)). IPC was not performed in group C. Among the animals in the experimental groups, IPC was performed by blocking blood flow by the portal vein and the hepatic artery followed by reperfusion by removal of the clamp before donor liver resection: Group E(1), 5-minute ischemia and 10-minute reperfusion; Group E(2), 5-minute ischemia and 5-minute reperfusion and immediately the same procedure; and Group E(3), 10-minute ischemia and 15-minute reperfusion. Liver transplantations were performed 4 hours after IPC. At 0.5 hour, 2 hours, 6 hours, and 24 hours after portal vein reperfusion recipient blood and graft samples were obtained to determine the levels of ALT, AST, TNF-alpha, and apoptosis index (AI). RESULTS: At 0.5 hour and 2 hours after portal vein reperfusion, serum tumor necrosis factor (TNF)-alpha in the experimental groups (E(1), E(2), and E(3)) was significantly lower than in the control group (P < .05). The values in group E(2) were significantly lower than those in groups E(1) and E(3) (P < .05). At 24 hours serum TNF-alpha in group E(2) was significantly lower than groups C, E(1), and E(3) (P < .05). At 2 hours and 6 hours, AI values in experimental groups (E(1), E(2), and E(3)) were significantly lower than in group C (P < .05). AI in group E(2) was significantly lower than that in groups E(1) and E(3) (P < .05). At 24 hours, AI values among experimental groups (E(1), E(2), and E(3)) were significantly lower than that in the control group (P < .05). CONCLUSION: IPC may attenuate liver graft injury by decreasing apoptosis of hepatocytes and production of TNF-alpha. The method of IPC with 5-minute ischemia and 5-minute reperfusion followed immediately by another cycle of the same procedure was a better way to protect a liver graft from I/R injury.     

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.     

缺血预处理对肝脏缺血再灌注损伤中MPO、TNF-α、ET、NO的影响

目的：探讨缺血预处理（IPC）对肝脏缺血再灌注（I/R）损伤中的中性粒细胞和某些细胞因子的影响。方法：采用大鼠部分肝脏原位缺血再灌注损伤模型,30只Wistar大鼠随机分成：①正常对照组;②缺血再灌注对照组;③预处理组。②、③组均在60min再灌注完成后取血及肝组织标本,检测血清AST、ALT、LDH、NO、ET-1、TNF-α、及肝组织中髓过氧化酶（MPO）活性和肝细胞病理改变。结果：预处理组与再灌注对照组比较,肝功明显改善,NO含量升高,ET-1、TNF-α浓度和MPO活性明显降低,两组比较差异具有显著性。结论：缺血预处理对肝脏缺血再灌注损伤有明显保护作用,可能与提高内源性NO水平、减轻中性粒细胞在肝脏中的渗出和聚集、抑制ET-1、TNF-α的合成有关。