Preconditioning methods in the management of hepatic ischemia reperfusion‐ induced injury: Update on molecular and future perspectives

Hepatic IR (ischemia reperfusion) injury is a commonly encountered obstacle in the post‐operative management of hepatic surgery. Hepatic IR occurs during ‘Pringle maneuver’ for reduction of blood loss or during a brief period of cold storage followed by reperfusion of liver grafts. The stress induced during hepatic IR, triggers a spectrum of cellular responses leading to the varying degrees of hepatic complications which in turn affect the post operative care. Different preconditioning methods either activate or subdue different sets of molecular signals, resulting in varied levels of protection against hepatic IR injury. Yet, there is a serious lacuna in the knowledge regarding the choice of preconditioning methods and the resulting molecular changes in order to assess the efficiency and choice of these methods correctly. This review provides an update on the various preconditioning approaches such as surgical/ischemic, antioxidant, pharmaceutical and genetic preconditioning strategies published during last six years (2009–2015). Further, we discuss the attenuation or inhibition of specific inflammatory, apoptotic and necrotic markers in the various experimental models of liver IR subjected to different preconditioning strategies. While enlisting the controversies in the ischemic preconditioning strategy, we bring out the uncertainties in the existing molecular targets and their reliability in the attenuation of hepatic IR injury. Future research studies would include the novel preconditioning strategies employ i) the targeted gene silencing of key molecular targets inducing IR, ii) hyper expression of beneficial molecular signals against IR via gene transfer techniques. The above studies would see the combination of these latest techniques with the established preconditioning strategies for better post‐operative hepatic management.     

心肌缺血/再灌注损伤后的肺组织损伤

目的初步探讨心肌缺血／再灌注损伤对肺组织损伤的可能机制。方法选取雄性成年SD大鼠（4～6月龄）,体重130～160g,建立成年大鼠缺血／再灌注模型。运用CK和MPO试剂盒检测肌酸激酶（CK）和髓过氧化物酶（MPO）的含量,运用双抗体夹心ABC—ELISA法检测细胞间黏附分子-1（ICAM-1）的含量。结果与伪手术组相比,缺血／再灌注大鼠的AN／AAR比值明显增高（P〈0．05）,CK在心肌缺血／再灌注大鼠血清中的含量明显升高（P〈0．05）,MPO与ICAM-1在心肌缺血／再灌注组大鼠血清和肺组织中含量明显升高（P〈0．05）。结论大鼠心肌缺血再灌注损伤后肺组织受到一定的损伤,可能与体循环中炎性介质的作用及肺组织的炎性应激有关。     

Amelioration of hepatic ischemia/reperfusion injury in the remnant liver after partial hepatectomy in rats

BACKGROUND AND AIMS: Reactive oxygen species have been implicated in the development of hepatic ischemia/reperfusion (I/R) injury. I/R injury remains an important problem in massive hepatectomy and organ transplantation. The aim of this study was to examine the effect of edaravone, a newly synthesized free radical scavenger, on I/R injury in the remnant liver after partial hepatectomy in rats. METHODS: Partial (70%) hepatic ischemia was induced in rats by occluding the hepatic artery, portal vein, and bile duct to left and median lobes of liver. Total hepatic ischemia (Pringle maneuver) was induced by occluding the hepatoduodenal ligament. Edaravone was intravenously administered to rats just before reperfusion and partial (70%) hepatectomy was performed just after reperfusion. RESULTS: Edaravone significantly reduced the increases in the levels of serum alanine aminotransferase and aspartate aminotransferase in rats with liver injury induced by 90-min of partial ischemia followed by 120-min of reperfusion. Histopathological analysis showed that edaravone prevented inflammatory changes in the livers with I/R injury. Edaravone also decreased the levels of myeloperoxidase activity, which is an index of neutrophil infiltration, and interleukin-6 mRNA, which is a proinflammatory cytokine. Additionally, edaravone improved the survival rate in partial hepatectomy rats with I/R injury induced by the Pringle maneuver. CONCLUSIONS: Edaravone administration prior to reperfusion protected the liver against I/R injury. Edaravone also improved the function of the remnant liver with I/R injury after partial hepatectomy. Therefore, edaravone may have applicability for major hepatectomy and liver transplantation in the clinical setting.     

Splenic artery ligation improves remnant liver function in partially hepatectomized rats with ischemia/reperfusion injury

Background: In liver resection, the temporary occlusion of the hepatoduodenal ligament (Pringle maneuver) is often used. However, the maneuver causes severe ischemia/reperfusion injury in the remnant liver. Our aim was to investigate the effects of splenic artery ligation on the liver function in partially hepatectomized rat with the Pringle maneuver. Methods: The Pringle maneuver was conducted for 30 min just before a two‐thirds partial hepatectomy. Splenic artery ligation was performed before the Pringle maneuver. The efficacy of splenic artery ligation was assessed by survival, serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), recovery of remnant liver weight, and portal pressure. Results: On day 3, animal survival was four rats of 12 in partially hepatectomized rats with the Pringle maneuver and 10 rats of 12 in the splenic artery ligation‐treated partially hepatectomized rats with the Pringle maneuver. A two‐thirds partial hepatectomy alone or splenic artery ligation itself did not show any effects on the survival. Compared with partially hepatectomized rats with the Pringle maneuver, splenic artery‐ligated animals had lower serum AST and ALT levels, and higher recovery of remnant liver weight. Splenic artery ligation significantly reduced the portal pressure and also decreased the fatality in excessively hepatectomized rats. Conclusions: Splenic artery ligation ameliorated the remnant liver function in partially hepatectomized rats with the Pringle maneuver and excessively hepatectomized rats. The amelioration may be mediated at least by decreasing portal pressure.     

Effect of MG132 on proteasome activity and prooxidant/antioxidant status of rat liver subjected to ischemia/reperfusion injury.

Aim: Previous studies have shown that proteasome inhibitors exerted protective effects against ischemia/reperfusion injury (IRI) of brain, heart, kidney and intestine. The aim of the present study was to investigate: (i) whether the proteasome inhibitor MG132 protects rat liver against IRI; and (ii) whether MG132 modulates prooxidant/antioxidant status of rat liver subjected to warm IRI. Methods: The left lateral and medial lobes (approximately 70% of the total liver volume) of livers of male Wistar rats were subjected to 30-min ischemia followed by 60-min reperfusion. Lactate dehydrogenase (LDH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels were measured in the plasma. Proteasome chymotryptic-like (ChT-L) activity, levels of thiobarbituric acid-reactive substances (TBARS), protein carbonyls (PC) and glutathione (GSH), as well as superoxidase dismutase (SOD), catalase (CAT), glutathionine peroxidase and glutathionine reductase activities were measured in liver fractions. Results: Thirty-min ischemia followed by 60-min reperfusion increased liver TBARS and PC, CAT and SOD activities, but decreased GSH level. Ischemia/reperfusion-induced oxidative stress was exacerbated in mitochondria, indicating that these organelles are the preferential target of IRI. Plasma LDH and AST levels were decreased by MG132 during both ischemia and reperfusion, while ALT values were decreased only after 30 min of reperfusion. MG132 did not significantly affect liver TBARS and GSH levels, but it increased PC and decreased ChT-L activity; the activities of CAT and SOD were also decreased. Conclusions: MG132 exerts a protective effect during the early phase of reperfusion and it modulates prooxidant/antioxidant status of rat liver subjected to warm IRI.     

Splenectomy-reduced hepatic injury induced by ischemia/reperfusion in the rat

Abstract: In the present study, we investigated the role of the spleen in experimental hepatic ischemia/reperfusion in the rat. After a 90-min period of ischemia in the left and middle hepatic lobes, the ischemia was released and the liver was reperfused for up to 24 h. Plasma alanine aminotransferase reached a peak 3 h after the onset of reperfusion, and gradually decreased thereafter. A histological examination revealed evidence of hepatocellular necrosis and degeneration, especially 24 h after the onset of reperfusion. In addition, there was a noticeable accumulation of polymorphonuclear cells in the liver following ischemia/reperfusion. A splenectomy performed just prior to ischemia/reperfusion reduced both biochemical and histological hepatocellular injury. The number of polymorphonuclear cells in the liver following ischemia/reperfusion was significantly reduced in rats subjected to splenectomy, suggesting that the increase in polymorphonuclear cells may contribute to liver injury. The number of mononuclear cells also increased in the marginal zones of the spleen following ischemia/reperfusion, and appeared to be derived from the splenic monocyte/macrophage population, based on immunohistochemical studies. The spleen plays an important role in the pathogenesis of hepatic ischemia/reperfusion injury and the splenic monocyte/macrophage population contributes to liver damage.     

抗氧化剂mTHC对大鼠脏器缺血再灌注损伤的拮抗作用

目的探讨抗氧化剂mTHC对缺血再灌注（I/R）大鼠脏器损伤的拮抗作用。方法取24只Wistar大鼠建立I/R损伤模型。灌注前15min和缺血前期分别予mTHC 10mg/kg给药2次。再灌注末期将大鼠断头,取肝、回肠和肺组织样本做生化分析;以肺湿重/干重计算肺损伤程度。结果应用mTHC后,肝脏、小肠、肺丙二醛（MDA）水平分别为（49.2±1.5）、（21.2±2.7）、（79.4±3.7）μmol/g,P〈0.01;髓过氧化物酶（MPO）水平分别为（14.7±1.1）、（17.8±1.1）、（36.7±2.1）U/g,P〈0.05;还原型谷胱甘肽（GSH）水平分别为（1.34±0.1）、（1.72±0.1）、（1.20±0.1）μmol/g,P〈0.05,AST、ALT分别为（36.2±2.4）、（50.1±2.2）mg/dl,肺湿重/干重为6.9±0.5。结论mTHC可能对I/R诱发的器官损伤有保护作用。     

大鼠脑缺血/再灌注损伤后血管内皮细胞生长因子的表达

目的　探讨大鼠局灶性脑缺血 /再灌注后脑组织内血管内皮细胞生长因子 (VEGF)的表达及意义。方法　制备大鼠局灶性脑缺血再灌注模型 ,应用免疫组化 S- P法检测 VEGF蛋白的表达。结果　缺血再灌注损伤后 VEGF表达增加 ,随再灌注时间的延长 ,在缺血灶周边区 ,阳性表达的小血管数量明显增多。结论　脑缺血再灌注损伤可以诱导 VEGF表达增强 ,VEGF可促进毛细血管增生。     

Melatonin abates liver ischemia/reperfusion injury by improving the balance between nitric oxide and endothelin

Introduction Melatonin (5-methoxy-N-acetyltryptamine) is a naturally occurring hormone derived from the amino acid tryptophan andproduced mainly by the pineal gland (pinealocytes) in the brain as well as in the retina and gastrointestinal tract. Although the major role of melatonin is in the sleep-wake cycle through its circadian fluctuation, a large body of literature has recently demonstrated that melatonin also exerts complex physiological and pharmacological effects on multiple systems and…     

JAK2/STAT3 activation by melatonin attenuates the mitochondrial oxidative damage induced by myocardial ischemia/reperfusion injury

Ischemia/reperfusion injury (IRI) is harmful to the cardiovascular system and causes mitochondrial oxidative stress. Numerous data indicate that the JAK2/STAT3 signaling pathway is specifically involved in preventing myocardial IRI. Melatonin has potent activity against IRI and may regulate JAK2/STAT3 signaling. This study investigated the protective effect of melatonin pretreatment on myocardial IRI and elucidated its potential mechanism. Perfused isolated rat hearts and cultured neonatal rat cardiomyocytes were exposed to melatonin in the absence or presence of the JAK2/STAT3 inhibitor AG490 or JAK2 siRNA and then subjected to IR. Melatonin conferred a cardio‐protective effect, as shown by improved postischemic cardiac function, decreased infarct size, reduced apoptotic index, diminished lactate dehydrogenase release, up‐regulation of the anti‐apoptotic protein Bcl2, and down‐regulation of the pro‐apoptotic protein Bax. AG490 or JAK2 siRNA blocked melatonin‐mediated cardio‐protection by inhibiting JAK2/STAT3 signaling. Melatonin exposure also resulted in a well‐preserved mitochondrial redox potential, significantly elevated mitochondrial superoxide dismutase (SOD) activity, and decreased formation of mitochondrial hydrogen peroxide (H₂O₂) and malondialdehyde (MDA), which indicates that the IR‐induced mitochondrial oxidative damage was significantly attenuated. However, this melatonin‐induced effect on mitochondrial function was reversed by AG490 or JAK2 siRNA treatment. In summary, our results demonstrate that melatonin pretreatment can attenuate IRI by reducing IR‐induced mitochondrial oxidative damage via the activation of the JAK2/STAT3 signaling pathway.     

Resveratrol attenuates oxidative stress and histological alterations induced by liver ischemia/reperfusion in rats

AIM： To investigate the effects of resveratrol on liver ischemia/reperfusion （I/R） injury in rats. METHODS： A total of 40 male Sprague-Dawley rats weighing 240-290 g were randomized into four groups of ten： （1） controls： data from unmanipulated animals; （2） sham group： rats subjected to the surgical procedure, except for liver I/R, and given saline; （3） I/R group： rats underwent liver ischemia for 45 rain followed by reperfusion for 45 rain; （4） I-R/Resveratrol group： rats pretreated with resveratrol （10 umol/L, iv）. Liver tissues were obtained to determine antioxidant enzyme levels and for biochemical and histological evaluation. RESULTS： Plasma aminotransferase activities were higher in the I/R group than in the I-R/Resveratrol group. Malondialdehyde levels and the hepatic injury score decreased, while superoxide dismutase, catalase, and glutathione peroxidase levels increased in group 4 compared to group 3. In group 4, histopathological changes were significantly attenuated in resveratroltreated livers. CONCLUSION： These results suggest that resveratrol has protective effects against hepatic I/R injury, and is a potential therapeutic drug for ischemia reperfusionrelated liver injury.     

丙酮酸对缺血/再灌注大鼠心肌的保护作用

目的:探讨丙酮酸(Pyr)对缺血/再灌注(I/R)大鼠心肌的影响及其可能的机制。方法:30只SD成年大鼠随机分为假手术(Sham)组、I/R组及I/R+Pyr组,每组10只。I/R+Pyr组大鼠于再灌前5 min,开始持续性静脉灌注Pyr 2 h[25 mg/(kg·h)]。再灌注2 h后,利用多道生理记录仪检测大鼠在体血流动力学指标:平均动脉压(MABP)、左室收缩压(LVSP)及左室最大收缩、舒张末压微分(±LV dP/dt max)。采用Western blot方法检测磷酸化-JNK(p-JNK)和总的JNK(t-JNK)表达。用原位缺口末端标记法(TUNEL)评价心肌细胞的凋亡。结果:I/R组的MABP、LVSP、±LV dP/dt max显著低于Sham组(P0.01),Pyr干预可增加I/R后MABP、LVSP及±LV dP/dt max的水平(P0.05)。与Sham组相比,I/R组大鼠左心室p-JNK的水平明显增高(P0.01);而Pyr可降低大鼠左心室p-JNK的水平(P0.01),并抑制心肌细胞凋亡(P0.05)。结论:Pyr可改善I/R大鼠心肌的功能,抑制心肌细胞凋亡,其机制可能与抑制JNK信号的激活有关。     

Fish oil alleviates liver injury induced by intestinal ischemia/reperfusion via AMPK/SIRT-1/autophagy pathway

AIM To evaluate whether fish oil(FO) can protect liver injury induced by intestinal ischemia/reperfusion(I/R) via the AMPK/SIRT-1/autophagy pathway.METHODS Ischemia in wistar rats was induced by superior mesenteric artery occlusion for 60 min and reperfusion for 240 min. One milliliter per day of FO emulsion or normal saline was administered by intraperitoneal injection for 5 consecutive days to each animal. Animals were sacrificed at the end of reperfusion. Blood andtissue samples were collected for analyses. AMPK, SIRT-1, and Beclin-1 expression was determined in lipopolysaccharide(LPS)-stimulated HepG2 cells with or without FO emulsion treatment.RESULTS Intestinal I/R induced significant liver morphological changes and increased serum alanine aminotransferase and aspartate aminotransferase levels. Expression of p-AMPK/AMPK, SIRT-1, and autophagy markers was decreased whereas tumor necrosis factor-α(TNF-α) and malonaldehyde(MDA) were increased. FO emulsion blocked the changes of the above indicators effectively. Besides, in LPS-stimulated HepG2 cells, small interfering RNA(siRNA) targeting AMPK impaired the FO induced increase of p-AMPK, SIRT-1, and Beclin-1 and decrease of TNF-α and MDA. SIRT-1 siRNA impaired the increase of SIRT-1 and Beclin-1 and the decrease of TNF-α and MDA.CONCLUSION Our study indicates that FO may protect the liver against intestinal I/R induced injury through the AMPK/SIRT-1/autophagy pathway.