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.     

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     

Ischemic Postconditioning and Nitric Oxide Administration Failed to Confer Protective Effects in a Porcine Model of Extracorporeal Cardiopulmonary Resuscitation

The protective effects of ischemic postconditioning (IPC) and nitric oxide (NO) administration have been demonstrated in several ischemic scenarios. However, current evidence regarding the effect of IPC and NO in extracorporeal cardiopulmonary resuscitation remains lacking. Fifteen female swine (body weight 45 kg) underwent veno‐arterial extracorporeal membrane oxygenation (ECMO) implantation; cardiac arrest‐ventricular fibrillation was induced by rapid ventricular pacing. After 20 min of cardiac arrest, blood flow was restored by increasing the ECMO flow rate to 4.5 L/min. The animals (five per group) were then randomly assigned to receive IPC (three cycles of 3 min ischemia and reperfusion), NO (80 ppm via oxygenator), or mild hypothermia (HT; 33.0°C). Cerebral oximetry and aortic blood pressure were monitored continuously. After 90 min of reperfusion, blood samples were drawn for the measurement of troponin I, myoglobin, creatine‐phosphokinase, alanine aminotransferase, neuron‐specific enolase, cystatin C, and reactive oxygen metabolite (ROM) levels. Significantly higher blood pressure and cerebral oxygen saturation values were observed in the HT group compared with the IPC and NO groups (P < 0.05). The levels of troponin I, myoglobin, creatine phosphokinase, and alanine aminotransferase were significantly lower in the HT group (P < 0.05); levels of neuron‐specific enolase, cystatin C, and ROM were not significantly different. IPC and NO were comparable in all monitored parameters. The results of the present study indicate that IPC and NO administration are not superior interventions to HT for the maintenance of blood pressure, cerebral oxygenation, organ protection, and suppression of oxidative stress following extracorporeal cardiopulmonary resuscitation.     

Ischemic preconditioning of a rat adipomusculocutaneous flap: which duration of ischemia/reperfusion works best?

In this study a comparison was made to the efficacy of three cycles of various duration of ischemic preconditioning (IPC) on flap survival after the critical ischemic time. Method: In the first study, 20 Sprague-Dawley rats were used (6 groups of rats; flap ischemia ranging from 6 to 14 h) to determine the critical ischemia time for the rat groin flap (6×3 cm). In the second study the rats (n=31 were divided into nine groups; ischemia and reperfusion times being 5, 10 or 15 min) a groin flap was elevated. Thereafter IPC was induced to the flap by clamping its pedicle prior to the critical ischemic time (14 h). Three cycles of IPC were applied. After seven days, the flap was retraced for viability assessment and the surviving flap area was calculated with planimetry. Results: Without pretreatment with IPC flap survival after 8, 10, 12, 13, and 14 h global ischemia was respectively 100%, 77±13%, 78±15%, 42±16%, and 2±4%. When the flaps were treated with three cycles of IPC before 14 h global ischemia the significant flap survival was noted in the 10/5 min and 15/10 min IPC groups (p<0.05). Overall, the 5 min reperfusion groups increased flap survival (38±26%; p<0.05), and had fewer total flap losses (2/9) than the 10 or 15 min reperfusion groups. Only if ischemia time was longer than reperfusion time the flaps had increased survival area (40±22%, p<0.05). Conclusion: This study demonstrated the efficacy of IPC to improve the viability of a composite flap even after a long critical ischemic time. The optimal IPC duration was 10/5 min combination, yet the 15/10 schedule was also acceptable. Received: 20 May 1998 / Accepted: 20 July 1998     

Effects of ischemic liver preconditioning on hepatic ischemia/reperfusion injury in the rat

To minimize bleeding during major liver resections or liver transplantation, surgical measures have been adopted that induce ischemia-reperfusion injury (I/R) which may significantly contribute to morbidity and mortality of partial liver resections. Several methods have sought to minimize I/R hepatic lesions. The present project assessed the protective role of ischemic preconditioning (IPC) in rat livers. The IPC was accomplished by clamping the hepatic pedicle for 5 minutes, followed by a 5-minute reperfusion (R) period before a 2-hour ischemia. Thereafter, reperfusions of 1, 3, and 24 hours were compared among IPC and control groups without IPC. Liver biopsy and blood samples were measured for mitochondrial respiratory control ratio (RCR), serum aspartate aminotransferase (AST), and alanine aminotransferase (ALT). IPC protected liver mitochondrial function. Serum aminotransferase levels were significantly lower among animals undergoing IPC compared with groups without IPC. Thus, we verified the effects of IPC for hepatocellular protection against I/R lesions.     

缺血预处理对肝硬化肝癌患者入肝血流阻断肝切除的保护作用及其机理探讨

目的　探讨缺血预处理 (IPC)对肝硬化肝癌患者入肝血流阻断肝切除的保护作用及其机理。　方法　将近期手术切除的 2 9例原发性肝癌 (HCC)随机分为 2组 :IPC组 (n =14 ) :肝门阻断切肝前先给予缺血 5min ,灌注 5min的缺血预处理 ;对照组 (n =15 ) :单纯肝门阻断切肝。比较 2组术前后肝功能的变化和肝灌注 1h时肝组织caspase 3活性和细胞凋亡的情况。　结果　术后 1、3、7d ,IPC组的天门冬氨酸氨基转移酶 (AST)、丙氨酸氨基转移酶 (ALT)明显低于对照组 (t =4 2 38,P <0 0 5 ) ;术后 3、7d ,IPC组的总胆红素 (TBIL)明显低于对照组 (t=2 2 96 ,P <0 0 5 ) ;术后 1d ,IPC组的ALB高于对照组 ,但无统计学差异 (t=2 0 2 9,P >0 0 5 )。术后 1h ,IPC组肝组织caspase 3活性和凋亡的内皮细胞均明显低于对照组 (t=2 349,P <0 0 5 )。　结论　IPC对肝硬化肝癌患者入肝血流阻断肝切除术后肝功能有良好的保护作用 ,其保护机理是通过抑制caspase 3的活性 ,从而抑制肝窦内皮细胞来实现的。     

内质网应激分子伴侣GRP78在大鼠缺血再灌注损伤肝脏中的表达

目的:观察内质网应激相关分子葡萄糖调节蛋白78(GRP78)在大鼠缺血再灌注损伤肝脏组织中的表达水平.方法:将24只健康雄性SD大鼠随机均分为假手术组,单纯肝缺血组(肝缺血30 min+再灌注0h),再灌注6h组(肝缺血30 min+再灌注6h)和再灌注12h组(肝缺血30 min+再灌注12h).分别检测各组血清丙氨酸转氨酶(ALT)和门冬氨酸转氨酶(AST)水平;肝组织病理学、凋亡情况及GRP78 mRNA表达水平.结果:与对照组比较,各实验组大鼠肝缺血后出现明显的肝组织损伤,且随着再灌注时间的延长损伤加重,表现为血清ALT和AST水平升高,明显的肝组织病理学改变,肝细胞凋亡率增加,各组间计量指标的差异均有统计学意义(均P＜0.05).大鼠肝组织GRP78 mRNA变化趋势与上述指标一致,缺血后表达明显上调,且随着再灌注时间延长而逐渐升高,各组间差异均有统计学意义(均P＜0.05).结论:缺血再灌注损伤肝脏组织中GRP78表达上调,但其具体作用还有待于探明.     

Ischemic preconditioning fails to improve microcirculation but increases apoptotic cell death in experimental pancreas transplantation

Brief periods of warm ischemia and subsequent short reperfusion before either long-term cold or warm ischemic insult (ischemic preconditioning, IPC) have proven to ameliorate ischemia/reperfusion (I/R) injury in various organs, such as the liver and lung. The aim of this study was to examine the effect of IPC on pancreatic cell apoptosis and microcirculatory impairments in experimental pancreas transplantation. Male Lewis rats served as donors and recipients of heterotopic syngeneic pancreaticoduodenal transplantation. Recipient animals were divided into two experimental groups: group Tx (n=7) received grafts without IPC, group Tx&IPC received grafts with IPC. Animals that had not undergone transplantation but whose pancreata had been exteriorized served as controls (n=5). All pancreatic grafts were preserved in University of Wisconsin solution for 6 h at 4°C. IPC was induced by interruption of the arterial blood flow for 10 min followed by 10 min of reperfusion. One and two hours after reperfusion, graft microcirculation was assessed by means of intravital microscopy (IVM). Rats were immediately killed after the second measurement and DNA breaks of acinar cells were detected by in situ terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate digoxigenin nick end-labelling (TUNEL) assay and gel electrophoresis (laddering). The apoptotic index (AI) was defined as the number of apoptotic cells per high-power field. Analysis of both groups of transplanted grafts showed a significant decrease in functional capillary density (FCD) and a significant increase in leukocyte sticking to postcapillary venules (LAV) at 1 h and 2 h of reperfusion, compared with animals that had not undergone transplantation (P<0.01). In parallel, AI was significantly increased in transplanted grafts compared to the controls (P<0.01). Grafts subjected to IPC showed no significant differences, neither for FCD nor LAV, at both time points if compared with grafts of group Tx. However, IPC resulted in a significant increase in AI (P<0.05). We can conclude that IPC has no effect on pancreatic microcirculation but enhances acinar cell apoptosis in experimental pancreas transplantation. These results indicate that IPC might increase I/R injury after pancreatic cold ischemia.     

Silibinin Improves TNF-α and M30 Expression and Histological Parameters in Rat Kidneys After Hepatic Ischemia/Reperfusion

Background: Remote kidney damage is a sequel of hepatic ischemia–reperfusion (I/R) injury. Silibinin is the main ingredient of the milk thistle plant seed extract with known antioxidant and hepatoprotective activity. Our study investigates the nephroprotective potential of intravenously administered silibinin, as a lyophilized SLB-hydoxypropyl-beta-cyclodextrin product, in hepatic I/R injury. Material and methods: 63 Wistar rats were divided into three groups: Sham (virtual intervention); Control (45 min ischemia and reperfusion); and Silibinin (200 μL intravenous silibinin administration after 45 min of ischemia). Kidney tissues were collected to determine TNF-α, M30 and histopathological changes at predetermined time intervals. Results: Comparing Sham vs. Control groups, proved that hepatic I/R injury increased renal TNF-α and M30 expression. Deterioration was observed in hyperemia/filtration of renal parenchyma and tubules, cortical filtration, tubular necrosis and edema (tissue swelling index). Intravenous silibinin administration and comparison of the Control vs. Silibinin groups showed a statistically significant decrease in TNF-α levels at 240 min following I/R (p < 0.0001), and in M30 at 180 min (p = 0.03) and 240 min (p < 0.0001). Renal parameters have significantly decreased in: hyperemia/filtration of renal parenchyma at 120 min (p = 0.003), 180 min (p = 0.0001) and 240 min (p = 0.0002); hyperemia/filtration of renal tubules at 120 min (p = 0.02), 180 min (p = 0.0001) and 240 min (p = 0.0005); cortical filtration (240 min - p = 0.005); tubular necrosis (240 min - p = 0.021); and edema (240 min - p = 0.001). Conclusion: Our study confirms that hepatic I/R injury causes remote renal damage while the intravenous administration of silibinin leads to statistically significant nephroprotective action.     

Free Radical Scavengers Improve Liver Function but Not Morphological Changes Induced by Reperfusion Injury

Objective: Reperfusion injury (RI) is associated with high generation of reactive oxygen species (ROS), but the extent of involvement of these agents in the injury remains controversial. The present study aimed to examine the effectiveness of ROS scavengers against hepatic reperfusion injury in the rat. Methods: The RI was induced in the liver using an isolated slow-flow, reflow perfused rat liver in both anterograde and retrograde perfusion. The effects of gentisic acid, N-acetyl cysteine, and trolox C on the superoxide production, liver function, and morphological changes were examined using different biochemical and histological assays. Results: The hepatic RI caused a significant (p < 0.05) increase in superoxide production and enzyme releases and a decrease in bile flow in both directions. Histological changes induced by RI include apoptosis, necrosis, pale cytoplasm, cell vacuolation, and attenuation of cell cords. Although the production of superoxide in retrograde direction was significantly less than the anterograde, the extent of the injury in the retrograde was greater than the anterograde direction. Pretreatment of the livers with each of the test compounds significantly reduced the release of lactate dehydrogenase and aspartate aminotransferase and improved bile flow in the liver exposed to hypoxia/reperfusion. However, they failed to protect the liver against the structural alterations induced by RI. Conclusion: ROS scavengers can reduce superoxide-induced damage and improve the liver function, but they are not able to prevent the structural changes. It shows that ROS are not the sole causative mechanism of hepatic RI and other mechanisms and mediators may be involved.     

Carnitine as a preventive agent in experimental renal ischemia-reperfusion injury

Reactive oxygen species generated during the reperfusion of ischemic kidney, as well as any other tissue, cause lipid peroxidation damaging the cell membrane. The aim of this study was to investigate the effect of carnitine in reperfusion injury of the kidney. Male albino rabbits were subjected to unilateral renal 1-h warm ischemia followed by 15 min of reperfusion. Group I (n=9): control group received 3 cc of isotonic saline solution and group II (n=9): carnitine group received 100 mg/kg of carnitine. Blood samples were collected at the 15th min of reperfusion from the left renal vein selectively. Preischemic and post-reperfusion serum and renal tissue MDA levels were measured by thiobarbituric acid reactive substances (TBARS) spectrophotometric analysis. The preischemic serum and tissue MDA values (sham values) for groups I and II were statistically comparable (P > 0.01). Serum and tissue MDA levels were markedly elevated after 15 min of reperfusion in group I (P < 0.01), while the values remained in the baseline levels following reperfusion in group II (P > 0.01). In group I, the major histological differences observed in the reperfused kidneys were marked edema and congestion whereas glomerular and tubular cellular integrity were well preserved in group II. Pre-treatment with carnitine in solid organ transplantations, preschock states, surgical procedures that require temporary vascular clamping etc. may be helpful to minimize the reperfusion injury in the involved tissue, reducing morbidity and mortality. Received: 22 May 2000 / Accepted: 1 February 2001     

氨溴索对大鼠肝脏缺血再灌注损伤的保护作用

目的：探讨氨溴索对大鼠肝脏缺血再灌注损伤（I/RI）的保护作用及其机制。方法：18只雄性Wistar大鼠被随机均分为假手术组、肝I/RI模型组（模型组）、氨溴索预处理+肝I/RI模型组（氨溴索预处理组）。肝I/RI模型采用阻断入肝血流30 min后再灌注方法诱导,氨溴索预处理组于缺血前20 min尾静脉注射氨溴索（100 mg/kg）,而模型组给予等体积生理盐水。术后6 h处死大鼠,检测血清丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）水平和肝组织病理学改变,同时检测肝组织超氧化物岐化酶（SOD）,谷胱甘肽（GSH）,丙二醛（MDA）含量,caspase-3的活化水平。结果：与假手术组比较,模型组与氨溴索预处理组血清ALT和AST水平明显升高（均P<0.05）;肝组织出现明显的病理学改变;肝组织SOD和GSH含量明显下降,而MDA水平明显升高（均P<0.05）;肝组织caspase-3活化水平明显升高（均P<0.05）。与模型组比较,氨溴索预处理组以上各项指标的变化均明显减弱（均P<0.05）。结论：氨溴索预处理能减轻大鼠肝脏I/RI,机制可能与其调控抗氧化和抗凋亡信号通路有关。     

钙预处理对未成熟心肌的保护作用

目的 观察钙预处理对未成熟心肌的影响.方法 采用Langendorff离体灌注模型,分为3组,缺血再灌组(I/R):离体心脏灌注10 win、工作心15 min后停灌45 min恢复灌注15 min,转为工作心模型30 min;心脏缺血预处理组(IPC):离体灌注10 min转为工作心15 min,反复2次缺血5min/再灌5min,停灌45min后恢复灌注15min,转为工作心模型30min;钙预处理组(CP):离体心脏灌注10 min、工作心15 min后,反复3次45 s无钙KH液灌流/5 min KH液灌流,停灌45 min后恢复灌注15 min,转为工作心模型30 min.以血流动力学指标、生化指标和心肌超微结构作为观察指标.结果 IPC与CP组比较,血流动力学指标、生化指标和心肌超微结构等方面均无明显差异;CP、IPC与I/R组比较,左心室功能恢复、三磷酸腺苷含量(ATP)(11.53±1.85、13.40±1.96比4.27±0.83,P＜0.01)、超氧化物歧化酶(SOD)活性(230.47±11.72、236.28±12.69比124.17±6.20,P＜0.01)、心肌线粒体Ca2+ATP酶活性(17.86±1.39、16.38±1.27比6.78 ±0.64,P＜0.01)和心肌线粒体合成ATP的能力(104.29±9.60、102.43±9.53比50.83±4.75,P＜0.01)明显增强,在心肌含水量(75.32±1.25、73.29±1.26比84.23±2.03,P＜0.01)、丙二醛含量(1.32±0.12、1.23±0.11比2.61±0.37,P＜0.01)、肌酸激酶(53.17±5.32、57.47±5.62比123.65±9.63,P＜0.01)和乳酸脱氢酶漏出率(32.16±3.23、34.48±3.43比85.43±5.93,P＜0.01)、心肌细胞内(2.54 ±0.32、2.17±0.22比4.48±0.74,P＜0.01)和心肌线粒体Ca2+含量(35.91±4.01、36.85±3.97比68.29±6.90,P＜0.01)明显减少;CP、IPC组心肌超微结构损伤较I/R组明显减轻.结论 钙预处理对未成熟心肌具有明显保护作用

Abstract：

Objective To investigate the protective effects of Ca2+ preconditioning on isolated immature myocardium.Methods Isolated working rabbit heart model was used,and 18 rabbits were randomly divided into 3 groups:ischemic/reperfusion (I/R) group receiving 45 min ischemia followed by 45 min reperfusion;myocardial ischemic preconditioning (IPC) group receiving 5 min ischemia and 5 min reperfusion 2 times before 45 min ischemia followed by 45 min reperfusion;Ca2 + preconditioning (CP)group receiving no-Ca2 + preconditioning before 45 min ischemia followed by 45 min reperfusion.The hemodynamics,biochemistry and myocardial ultrastructure were tested.Results The hemodynamics,biochemistry and myocardial ultrastructure had no significant diferrence between CP group and IPC group.As compared with I/R group,in CP and IPC groups,the left ventricular function recovery,adenosine triphosphate content (ATP) (11.53 ± 1.85,13.40 ± 1.96 vs 4.27 ±0.83,P＜0.01),superoxide dismutase (SOD)activity (230.47± 11.72,236.28 ± 12.69 vs 124.17 ±6.20,P＜0.01),Ca2+-ATPase activity of mitothondia ( 104.29 ± 9.60,102.43 ± 9.53 vs 50.83 ± 4.75,P＜0.01 ) and synthesized ATP activity of mitochondria ( 104.29 ±9.60,102.43 ±9.53 vs 50.83 ±4.75 ,P ＜0.01 ) were improved,and myocardial water content ( 75.32 ± 1.25,73.29 ± 1.26 vs 84.23 ± 2.03 ,P＜0.01 ),malondialdehyde content ( 1.32 ± 0.12,1.23 ± 0.11 vs 2.61 ± 0.37 ,P＜0.01 ),the dehydrogenase (32.16 ± 3.23,34.48 ± 3.43 vs 85.43 ± 5.93,P ＜0.01 ) and creatine kinase leakage (53.17 ±5.32,57.47±5.62 vs 123.65 ±9.63 ,P ＜0.01 ),myocardial cell Ca2+ content (2.54 ±0.32,2.17 ±0.22 vs 4.48 ±0.74 ,P ＜0.01 ) and mitochondrial Ca2+ content(35.91 ±4.01,36.85 ±3.97 vs 68.29 ±6.90,P ＜0.01 ) were reduced.The ultra.structure injury was milder in CP group and ICP group than in I/R group.Conclusion CP has signifcantly protective effects on immature myocardium.     

Hepatocellular Glycogen in Alleviation of Liver Ischemia-Reperfusion Injury During Partial Hepatectomy

Background Temporary occlusion of liver blood supply for complex liver operation is common in liver surgery. However, hepatic vascular occlusion will undoubtedly impair liver function. This study was designed to elucidate the effect of hepatocellular glycogen in alleviation of liver ischemia-reperfusion injury during hepatic vascular occlusion for partial hepatectomy. Methods Fifty-seven patients were randomly divided into an experimental group (n = 29) and a control group (n = 28). In the experimental group, patients were given high-concentration glucose intravenously during 24 h before the operation. The hepatic lesion was resected after portal triad clamping in the two groups. Noncancer liver tissue was biopsied to measure hepatic tissue ATP content and change of malondialdehyde (MDA) and superoxide dismutase (SOD). Liver function of all patients was assessed by using an automatic biochemical analysis apparatus before the operation and the first and fifth days after operation. Results The mean hepatic vascular occlusion time in the experimental group was 19.21 ± 4.54 min and in the control group it was 21.04 ± 5.11 min. Hepatic tissue ATP content of the experimental group was significantly higher than that of the control group at the end of hepatic vascular occlusion (2.15 ± 0.39 μmol/g wet tissue vs. 1.33 ± 0.44, p < 0.01) and at the point of 1-h reperfusion (2.19 ± 0.29 μmol/g wet tissue vs. 1.57 ± 0.35, p < 0.01). There was significant difference in SOD activity between the two groups at the end of hepatic vascular occlusion (130.69 ± 30.49 NU/mg pr vs. 97.83 ± 26.23, p < 0.01) and at the point of 1-h reperfusion (139.55 ± 39.88 NU/mg pr vs. 114.74 ± 25.93, p < 0.01). Significant difference was shown in MDA content between the two groups at the end of hepatic vascular occlusion (3.02 ± 0.30 nmol/mg pr vs. 3.99 ± 0.49, p < 0.01) and at the point of 1-h reperfusion (3.81 ± 0.69 nmol/mg pr vs. 5.75 ± 1.17, p < 0.01). In addition, the liver function of the experimental group was significantly better than that of the control group the first and fifth days after the operation (p < 0.01). Conclusions Abundant intracellular glycogen may reduce liver ischemia-reperfusion injury caused by hepatic vascular occlusion. It is beneficial to give a large amount of glucose before a complex liver operation during which temporary occlusion of hepatic blood flow is necessary.     

缺血预处理对大鼠肝脏缺血/再灌注损伤的延迟保护作用及机制

目的 研究缺血预处理(IPC)对大鼠肝脏缺血/再灌注损伤的延迟保护作用,并探讨线柱体ATP敏感性钾通道(mitoKATP通道)在这种保护机制中的作用. 方法 SD大鼠随机分为5组(每组8只).IPC组以肝缺血5 min作预处理;DE组以静脉注射mitoKATP通道选择性开放剂二氮嗪(DE)作为预处理;IPC+5-HD组是在IPC组基础上再予静注mitoKATP通道特异性阻滞剂5-hydroxydecanoate(5-HD)进行预处理;对照组(C组)仅以静注等量生理盐水作为预处理;上述4组均在预处理24 h后行肝缺血1 h再灌注3 h,缺血方式均为70%肝脏热缺血.假手术组(S组)仅行两次开腹手术,不作其它处理.完成预定实验操作后取血用于血清谷丙转氨酶(ALT)与乳酸脱氢酶(LDH)检测,切取肝组织用于测定超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量、湿重/干重(W/D)及观察显微及超微结构变化. 结果 C组ALT,LDH,MDA及W/D值明显高于S组(P<0.01),而SOD活性明显低于S组(P<0.01),肝脏的显微及超微结构损伤明显;IPC组与DE组的各项肝损伤指标均明显好于C组(P<0.05及P<0.01);IPC+5-HD组的肝损伤指标均差于IPC组(P<0.05及P<0.01). 结论 缺血预处理对正常大鼠肝脏I/R损伤具有延迟保护作用,肝细胞mitoKATP通道的开放在其中发挥了重要作用,作用途径可能与诱导肝脏SOD活性增加,改善肝组织微循环,减轻肝脏水肿有关.     

S-腺苷蛋氨酸对大鼠缺血再灌注肝脏能量代谢的影响

目的研究S-腺苷蛋氨酸(SAM)预处理对大鼠肝脏缺血再灌注损伤线粒体功能的影响。方法54只大鼠按随机数字表法随机均分为对照组、缺血再灌注组(I/R组)和SAM组,SAM组大鼠肝脏在缺血前2h行腹腔注射SAM预处理。3组动物在阻断肝门30min后(对照组仅做分离,不阻断肝门)复流,并于再灌注后0、1和6h抽取下腔静脉血检测ALT及AST,切取肝组织检测线粒体SOD、MDA、ATP及EC,并制备病理切片在电镜下观察线粒体的超微结构。结果再灌注后0、1及6h,I/R组ALT、AST和MDA明显高于对照组(P<0.01),SOD(除0h外)、ATP及EC明显低于对照组(P<0.01);SAM组ALT、AST及MDA(除0h外)明显低于I/R组(P<0.01),SOD(除0h外)、ATP及EC明显高于I/R组(P<0.05,P<0.01)。超微结构观察,I/R组线粒体较对照组有明显的损伤,线粒体数量减少,肿胀明显,嵴模糊不清,基质密度低;而SAM组与I/R组相比损伤程度明显减轻。结论SAM能抑制线粒体脂质过氧化反应,提高ATP的产生,最终提高线粒体的能量代谢水平,有效地减轻肝脏的缺血再灌注损伤。     

Effects of Intravenous Anesthetics on Renal Ischemia/Reperfusion Injury

Background. Renal ischemia/reperfusion (I/R)-induced tubular epithelial cell injury, called ischemic acute renal failure, is associated with high mortality in humans. Protecting the kidney against I/R injury is very important during complicated renal operations, transplantation surgery, and anesthesia. Aim. The purpose of this study was to investigate and compare the efficiency of ketamine, thiopental, propofol, etomidate, and intralipid in reducing the injury induced by free radicals in a rat model of renal I/R. Method. Forty-two Wistar rats were divided into seven groups in our study. Rats in the sham group underwent laparotomy and waited for 120 minutes (min) without ischemia. Rats in the control group were given nothing with ischemia-reperfusion. Rats in the I/R groups were given ketamine (20 mg/kg), thiopental (20 mg/kg) propofol (25 mg/kg), etomidate (10 mg/kg) and 10% intralipid (250 mg/kg) intraperitoneally 15 min prior to the ischemia for 60 min, followed by reperfusion for 60 min. The blood samples and kidney tissues of the rats were obtained under anesthesia at the end of the reperfusion period. Biochemical malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), blood urea nitrogen (BUN), creatine (Cr), aspartate aminotransferase (AST) were determined, and histopathological analysis was performed with these samples. Results. MDA level was increased significantly in the control group (p < 0.05). Histopathological findings of the control group confirmed that there was renal impairment by tubular cell swelling, interstitial edema, medullary congestion, and tubular dilatation. MDA levels were lower in the ketamine, thiopental, and propofol groups compared to the control group (p < 0.05). In the thiopental and propofol groups, the levels of histopathological scores were significantly lower than control and etomidate groups in ischemia-reperfusion. Conclusion. Our results demonstrated that I/R injury was significantly reduced in the presence of propofol and thiopental. The protective effects of these drugs may belong to their antioxidant properties. These results may indicate that propofol and thiopental anesthesia protects against functional, biochemical, and morphological damage better than control in renal I/R injury.     

Ischemic Preconditioning versus Intermittent Vascular Inflow Control during Major Liver Resection in Pigs

Ischemic preconditioning (IPC) and intermittent vascular control (IVC) have been shown to reduce the number of ischemia/reperfusion injuries during liver resections with the Pringle maneuver. Our study aimed to compare the beneficial effect of these two modalities in relation to the duration of normothermic liver ischemia. A group of 24 Landrace pigs with a mean body weight of 25 to 30 kg were subjected to extended liver resection of more than 65%. Although, 12 animals underwent IPC (10 minutes of ischemia and 10 minutes of reperfusion), and subsequently the Pringle maneuver was applied for 90 minutes (n= 6) or 120 minutes (n= 6). Another 12 animals underwent liver resection by IVC (20 minutes of ischemia alternated with 5 minutes of reperfusion) for 60 minutes (n = 6) or 120 minutes (n = 6) of inflow vascular control. At 90 minutes of liver ischemia, the IPC group demonstrated lower levels of asportate aminotransferase (AST) (173 ± 53 vs. 265 ± 106 IU; p =0.089) and malondialdehyde (MDA) (2.60 ± 1.03 vs. 5.33 ± 2.25 μmol/L; p =0.022) and higher liver tissue cAMP (200 ± 42 vs. 146 ± 40 pmol/g wet wt, p = 0.04) compared to the IVC group. However, no pathologic differences were observed between the two groups. By contrast, at 120 minutes of liver ischemia, IVC proved to be more beneficial, reflected by lower levels of AST (448 ± 135 vs. 857 ± 268 IU; p = 0.006) and MDA (8.33 ± 1.75 vs. 12.7 ± 4.31 μmol/L; (p = 0.045), a higher cAMP level (127 ± 10 vs. 97 ± 31 pmol/g wet wt p = 0.045), and eventually less cellular necrosis (necrosis score 1.66 ± 0.51 vs. 2.85 ± 1.16; p = 0.04) compared to the IPC group. It appears that IPC should be employed when liver ischemia is anticipated to last less than 90 minutes, followed by IVC when the liver ischemia is expected to last 120 minutes.     

Effect of ischemic preconditioning on the intestinal microvascular perfusion and oxygenation in reperfusion injury of the intestine

Background. Ischemia reperfusion (IR) injury of the intestine is inevitable in small bowel transplantation. Ischemic preconditioning (IPC) provides a way of protecting the intestine from damage inflicted by IR injury. This study was designed to evaluate the effect of IPC on the intestinal microvascular perfusion (IMP) and the intestinal tissue oxygenation. Methods. Rats were allocated into (a) sham group, (b) IR group consisting of 30 min of intestinal ischemia followed by 2 h of reperfusion, and (c) IPC group, as in IR group, but preceded by 10 min of ischemia and 10 min of reperfusion. Heart rate (HR), arterial oxygen saturation (SaO₂), and mean arterial pressure (MAP) were monitored continuously. IMP was continuously monitored by laser Doppler flowmetry (LDF). Near Infrared reflectance spectroscopy (NIRS) was used to monitor intestinal oxygenation, namely oxyhemoglobin (HbO₂) and cytochrome oxidase Cu_A redox state (Cyt Ox) continuously. At the end of the reperfusion, blood samples for lactate dehydrogenase (LDH) levels and biopsies of ileum for histopathological examination were obtained. Data were analyzed using one-way analysis of variance (ANOVA) and Bonferroni adjustment for multiple comparisons were used with Prism® software. Results. HR and SaO₂ did not change statistically throughout the experiment (P > 0.05). During ischemia there was an increase of MAP from the baseline in both IR (87 ± 3.6 to 111.5 ± 4.8 mmHg, P < 0.01) and IPC groups (87.7 ± 6.5 to 111.7 ± 4.6 mmHg, P < 0.01).There was a significant improvement in IMP with IPC (P < 0.01 versus IR). IPC also improved the HbO₂and Cyt Ox significantly (P < 0.001 versus IR) at the end of the reperfusion. Serum LDH levels were significantly reduced (P < 0.001) in IPC (667.1 ± 86.8) as compared to Group IR (1973.8 ± 306.5). Histological evaluation demonstrated that IPC lowered the intestinal mucosal injury grade (P < 0.01 versus IR). Conclusion. This study demonstrates that IPC improves the intestinal microvascular perfusion and tissue oxygenation in a model of warm IR injury of the intestine. These findings suggest that IPC may be applied during small bowel transplantation to reduce IR injury.