Comparison of ischemic preconditioning and intermittent and continuous inflow occlusion in the murine liver

OBJECTIVE: To compare protection of the liver by ischemic preconditioning and intermittent inflow occlusion in a mouse model of prolonged periods of ischemia. SUMMARY BACKGROUND DATA: Preconditioning (short ischemic stress prior to a prolonged period of ischemia) and intermittent inflow occlusion protect the liver against reperfusion injury. This is the first study comparing these two modalities with continuous inflow occlusion (control). METHODS: Mice were subjected to 75 or 120 minutes of 70% hepatic ischemia and 3 hours of reperfusion. Each ischemic period was evaluated using three different protocols: continuous ischemia (control), preconditioning (10 minutes ischemia and 15 minutes reperfusion) prior to the prolonged ischemic insult, and intermittent clamping (cycles of 15 minutes ischemia and 5 minutes reperfusion). Organ injury was evaluated using serum levels of aspartate aminotransferase (AST), hematoxylin and eosin staining, and specific markers of apoptosis (cytochrome C release, caspase 3 activity, and TUNEL staining). Animal survival was determined using a model of total hepatic ischemia. RESULTS: Intermittent inflow occlusion and ischemic preconditioning were both protective against ischemic insults of 75 and 120 minutes compared with controls (continuous ischemia only). Protection against 75 minutes of ischemia was comparable in the intermittent clamping and the ischemic preconditioning group, whereas intermittent clamping was superior at 120 minutes of ischemia. One hundred percent animal survival was observed after 75 minutes of total hepatic ischemia using both protective protocols, whereas all animals subjected to continuous ischemia died after surgery. After 120 minutes of ischemia, intermittent inflow occlusion was associated with better animal survival (71%) compared with preconditioning (14%). CONCLUSIONS: Preconditioning and intermittent clamping are both protective against prolonged periods of ischemia. In the clinical setting, preconditioning is superior for ischemic periods of up to 75 minutes because it is not associated with blood loss during transection of the liver. However, for prolonged ischemic insults exceeding 75 minutes, intermittent clamping is superior to preconditioning.     

Increased ischemic injury in old mouse liver: an ATP-dependent mechanism.

Although livers exhibit only minimal morphologic changes with age, how older livers tolerate pathologic conditions such as normothermic ischemia is unknown. Young 6-week-old mice and old 60-week-old mice underwent 60 minutes of hepatic ischemia and various periods of reperfusion. Markers of hepatocyte injury, hepatic energy content, and mitochondrial function were determined. Ischemic preconditioning and glucose injection were evaluated as protective strategies against reperfusion injury in old mice. Reperfusion injury was far worse in old mice compared with mice in the young control group. Ischemic preconditioning was highly protective against reperfusion injury in young but not in old mice. Older livers had dramatically reduced adenosine triphosphate (ATP) levels and glycogen contents. The low intrahepatic energy level in old mice was associated with a reduced mitochondrial ATP production. Preoperative injection of glucose restored the intrahepatic ATP content and protected against reperfusion injury. Furthermore, glucose injection restored the protective effect of ischemic preconditioning, resulting in additive protection when both strategies were combined. Aging of the liver is associated with mitochondrial dysfunction and decreased intrahepatic energy content, resulting in poorer tolerance against ischemic injury. Improving intrahepatic ATP levels in old livers by glucose injection protects the old liver against ischemic injury and restores the protective effects of ischemic preconditioning.     

Hepatic ischemia/reperfusion injury in P-selectin and intercellular adhesion molecule-1 double-mutant mice

Neutrophil adhesion and recruitment represents one of the early cellular events that occur during hepatic ischemia/reperfusion (IR) injury and plays a critical role in determining the extent of tissue damage. The adhesion molecules, such as selectins and intercellular adhesion molecules (ICAM), are important in mediating neutrophil-endothelial cell interactions and neutrophil emigration. The goal of this study was to evaluate the role of P-selectin and ICAM-1 in hepatic IR injury. Male wild-type and P-selectin/ICAM-1-deficient (P/I null) mice underwent 90 minutes of partial hepatic ischemia followed by reperfusion at various time points (0, 1.5, 3, and 6 hours). Reperfusion caused a time-dependent hepatocellular injury in both wild-type and P/I null mice as judged by plasma alanine aminotransferase (ALT) levels and liver histopathology examination. Although ALT levels were slightly lower in the P/I null mice compared with the wild-type mice the differences were not statistically significant. Neutrophil infiltration to the ischemic liver was observed in both mouse groups after 6 hours of reperfusion; however, the infiltration to the midzonal region of the ischemic liver was more pronounced in the wild-type group. This study suggests that hepatocellular injury induced after hepatic IR was independent of P-selectin and ICAM-1 in this model of acute inflammatory tissue injury.     

缺血预处理和Caspase抑制剂对缺血再灌注损伤保护作用的比较

目的　比较缺血预处理和Caspase抑制剂治疗对大鼠肝缺血再灌注的保护作用及其机制。方法　SD大鼠随机分为 6组 :( 1)缺血再灌注1 (IR1 )组 ;( 2 )IR2 组 ;( 3 )缺血预处理1 (IP1 )组 ;( 4 )IP2 组 ;( 5 )Caspase抑制剂治疗1 (T1 )组 ;( 6)T2 组。比较各组大鼠 70 %肝脏 60min或 12 0min缺血 ,在再灌注 3h时的肝组织Caspase 3活性 ,肝细胞凋亡率和血清AST和ALT水平 ,及实验动物 7d存活率。结果　在 60min缺血及 3h再灌注时间 ,IP1 组和T1 组的保护作用相同 ,在 12 0min缺血及 3h再灌注时 ,T2 组对Caspase活性和肝细胞凋亡的抑制优于IP2 组 (P <0 .0 1) ,但两者的AST和ALT水平及动物 7d存活率均无显著性差异。结论　缺血预处理和Caspase抑制剂治疗对鼠肝缺血再灌注损伤都有保护作用 ,两者的保护效果无显著性差异。缺血预处理对缺血再灌注损伤的保护更简便、经济、安全 ,临床应用前景十分广阔。     

缺血预处理减轻大鼠肝缺血再灌注损伤的免疫机制研究

目的探讨大鼠肝缺血再灌注损伤(HIRI)的免疫机制和缺血预处理(IPC)的保护作用。 方法80只大鼠被随机分为假手术组(A组)、肝门阻断20 min组(B组)、30 min组(C组)、40 min组(D组)以及肝门阻断30 min前预处理组(E组),每组再分为再灌注2 h亚组和24 h亚组,各8只。检测再灌注后2 h、24 h的血丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、白细胞介素10、12(IL-10、IL-12)以及外周血T淋巴细胞亚群的水平,观察再灌注后2 h、24 h时的存活率及肝脏病理情况。 结果随着肝门阻断时间的延长,ALT、AST显著升高,肝内炎症细胞浸润增加,24 h存活率逐渐降低。D组再灌注2 h时,CD8+ T淋巴细胞显著升高,CD4+/CD8+比值下降,调节性T淋巴细胞显著减少,血清IL-10显著降低,而IL-12水平显著升高。再灌注后24 h,B组大鼠各项指标逐步恢复至假手术组水平,而D组大鼠CD4+T淋巴细胞、CD4+/CD8+比值尤其是Treg显著升高,且IL-10水平显著升高,IL-12水平明显降低。与C组相比,E组阻断30 min后无一例死亡,再灌注2 h的ALT、AST水平显著降低,Treg和IL-10水平显著升高,IL-12水平明显降低,再灌注24 h后各项指标恢复并接近假手术组水平,肝脏病理损伤较轻。 结论肝缺血再灌注引起肝脏损伤甚至死亡,可能与诱导T淋巴细胞尤其是Treg和细胞因子的紊乱有关。缺血预处理可以增加再灌注早期的Treg细胞,有效纠正免疫紊乱,减轻损伤。     

The role of HMGB-1 on the development of necrosis during hepatic ischemia and hepatic ischemia/reperfusion injury in mice

BACKGROUND: High-mobility group 1 (HMGB-1) is a late mediator of endotoxin lethality in mice. The release of HMGB-1 is delayed compared to other proinflammatory cytokines that mediate shock and tissue injury. The purpose of this study was to investigate the role of HMGB-1 levels in response to hepatic ischemia, hepatic I/R injury, and the relationship between changes in HMGB-1 and other cytokines. MATERIALS AND METHODS: Three murine models were employed: our robust model of segmental hepatic warm ischemia (SHWI), a model of partial hepatic ischemia/reperfusion injury (PHIRI), and a model of total hepatic ischemia/reperfusion injury (THIRI). Over a 48-h period following ischemic insult and reperfusion using these models, serum HMGB-1 concentrations, concentrations of HMGB-1 in ischemic and nonischemic lobes, and serum concentrations of TNF-alpha and IL-6 levels were determined in mice. An anti-HMGB-1 antibody treatment was used in SHWI and THIRI to evaluate what aspects of response to ischemia and reperfusion were potentially mediated by HMGB-1. RESULTS: Hepatic HMGB-1 tissue concentrations exhibited biphasic changes in SHWI mice, which were increased in the ischemic lobes relative to nonischemic lobes at 12 h but decreased relative to nonischemic lobes at 24 h after ischemic insult. These results suggested that HMGB-1 was released into the systemic circulation by necrotic cells over the first 12 h but this process may be complete by 24 h postischemia. By 6 to 12 h after SHWI, serum TNF-alpha began to increase significantly and continued to increase for 18 h, followed by a sudden decline. Similarly, serum IL-6 increased over 1-3 h after SHWI and then decreased over the next 6 h. Treatment with an anti-HMGB-1 antibody significantly prolonged survival time in SHWI and THIRI. CONCLUSIONS: HMGB-1 plays a significant role in the response to hepatic ischemia and hepatic ischemia/reperfusion injury. The present study demonstrated a time-dependent production of HMGB-1 following hepatic warm ischemia in mice. The inherent HMGB-1 in ischemic areas was exhausted and HMGB-1 may be released by necrotic cells. HMGB-1 activation is involved in immediate proinflammatory stress response to I/R and anti-HMGB-1 antibody treatment remarkably improved survival. We demonstrated that systemic HMGB-1 accumulation was measured at an earlier phase of the hepatic ischemia and ischemia/reperfusion injury model than LPS-induced endotoxemia.     

Ischemic preconditioning and intermittent clamping increase the tolerance of fatty liver to hepatic ischemia-reperfusion injury in the rat

INTRODUCTION: Liver ischemia-reperfusion (I/R) injury is a well-known cause of morbidity and mortality following liver surgery and transplantation. Hepatic steatosis increases the extent of cellular injury incurred during I/R injury. We sought to identify measures that reduced the untoward sequelae of liver I/R injury. METHODS: Male Zucker rats were subjected to 75 minutes of 70% hepatic ischemia, and 3 hours of reperfusion. The ischemic periods were based on the following protocols: continuous clamping (CC) for 75 minutes; intermittent clamping (IC) with five cycles of 15 minutes clamp on and 5 minutes clamp off; or ischemic preconditioning (IP) with 10 minutes clamp on, 15 minutes off, and 60 minutes on (n=7 in each group). Warm I/R injury was evaluated using serum levels of aspartate aminotransferase (AST), serum interleukin (IL)-6, as well as hematoxylin and eosin staining. RESULTS: Hepatocellular injury was significantly reduced with IP or IC compared with CC (AST: 3285+/-122.3 and 2875+/-285.4 compared with 5436.3+/--984.7 units/L, respectively; P<.01). Serum IL-6 level was also significantly reduced with IP and IC compared with CC (70+/-8.8 and 76+/-6.2 compared with 147+/-8.5 ng/l, respectively (p<.01). Histological analysis also revealed that IC and IP provided significant protection compared with the CC group. CONCLUSION: IC and IP increased the tolerance of a fatty liver to hepatic I/R injury.     

Maladaptive role of IL-6 in ischemic acute renal failure

The role of IL-6 was investigated in murine ischemic acute renal failure. The renal pedicles were clamped for 17 min, and the mice were studied at various times after reperfusion. We found that serum IL-6 increased after murine ischemic renal injury. This increase was associated with increased IL-6 mRNA in the ischemic kidney but not in the contralateral kidney or the liver. Maximal IL-6 production occurred at 4 to 8 h and decreased to baseline by 24 h. Reperfusion of the kidney was required for IL-6 production. In situ hybridization and immunohistochemistry showed that macrophages infiltrated areas adjacent to the vascular bundles in the outer medulla within hours of reperfusion and showed that these macrophages produced IL-6 mRNA. For understanding how macrophages were stimulated to produce IL-6, an in vitro model in which S3 proximal tubular cells were injured by reactive oxygen species was set up. These injured cells released molecules that activated macrophages to produce IL-6 in vitro. IL-6 that was produced in response to renal ischemia was maladaptive because transgenic knockout of IL-6 ameliorated renal injury as measured by serum creatinine and histology. IL-6 transgenic knockout mice were lethally irradiated, and their bone marrow was reconstituted with wild-type IL-6 cells. Such bone marrow transfers abolished the protective effects of transgenic IL-6 knockout. It is concluded that macrophages infiltrate the area of the vascular bundles of the outer medulla, these macrophages produce IL-6, and this IL-6 exacerbates ischemic murine acute renal failure.     

Preconditioning protects against ischemia/reperfusion injury of the liver

Ischemic preconditioning (IPC) of an organ may induce protection against the injury caused by longer duration of ischemia and subsequent reperfusion. In a standardized model of such injury in the rat liver, we used the following protocol to investigate whether adenosine played a role in IPC by preventing its enzymatic degradation by dipyridamole pretreatment according to the following protocol: group 1, non-ischemic control rats; group 2, ischemic control rats subjected to 60 minutes of ischemia by clamping of the common hepatic artery followed by 60 minutes of reperfusion; group 3, IPC with 10 minutes of ischemia followed by 15 minutes of reperfusion, prior to the ischemia/reperfusion period as in group 2; group 4, pharmacologic preconditioning with administration of dipyridamole prior to the ischemia/reperfusion period as in group 2. Peripheral liver blood flow was significantly reduced during clamping (groups 2 to 4). After unclamping, blood flow was still reduced in the ischemic rats (group 2) but had returned to preclamp values in the animals that had been subjected to ischemic (group 3) or pharmacologic (group 4) preconditioning. Liver cell injury was significantly increased in the ischemia group (group 2) only. In our experimental model of ischemia/reperfusion injury in the rat liver, we found an equally beneficial effect with ischemic and pharmacologic preconditioning. Adenosine appears to be a crucial factor in IPC.     

缺血预处理对鼠后肢缺血再灌注下脊髓腰骶膨大运动神经元的保护作用

[目的]对大鼠一侧后肢的缺血再灌注损伤行不同条件缺血预处理,观察脊髓腰骶膨大处运动神经元超微结构的变化,探讨其保护作用.[方法]通过血管夹暂时阻断大鼠左侧髂总、髂内和髂外动脉,建立大鼠后肢缺血模型.以缺血6h再灌注2h和12 h分为A、B两组,预处理方式为缺血10 min血液复流10 min,按无预处理、预处理1次、无时间间隔预处理2、3次,分成A0、A1、A2、A3;B0、B1、B2、B3组.取材腰骶膨大处脊髓灰质前角,光镜及透射电镜下观察不同条件预处理对缺血再灌注损伤中脊髓前角超微结构变化.[结果]组织形态观察结果显示A0组缺血再灌注损伤后神经元细胞减少,核溶解消失,损伤明显,预处理后神经元细胞增多,可见部分细胞核存在;B0组缺血再灌注损伤后神经元细胞大部分坏死溶解,预处理后坏死溶解现象减轻.超微结构观察显示A0和B0组脊髓前角神经兀细胞不同程度核周器扩张损伤,出现内质网扩张、大量线粒体空泡以及核膜溶解消失.预处理后损伤程度有所减轻,叠加预处理后损伤进一步减轻.[结论]缺血预处理能减轻大鼠肢体缺血再灌注下脊髓腰骶膨大运动神经元的损伤,对脊髓具有保护作用.反复3次预处理产生的保护作用优于2次预处理及1次预处理.     

安氟醚和异氟醚对肝脏缺血/再灌注损害的影响

目的 研究安氟醚和异氟醚预处理对肝脏缺血／再灌注损害的影响。方法 １８只家兔随机分３组,对照组阻断肝劝脉和门静脉血流形成肝缺血４５ｍｉｎ,开放再灌注１２０ｍｉｎ;安氟醚和异氟醚预处理组,于缺血再灌注前分别吸入１．６８％安氟醚和１．１５％异氟醚２０ｍｉｎ和药物清除１０ｍｉｎ。结果 缺血前,三组血清谷丙转氨酶（ＡＬＴ）,谷草转氨酶（ＡＳＴ）、乳酸脱氢酶（ＬＤＨ）、碱性磷酸酶（ＡＬＰ）和谷氨酰酶（ＧＣＴ     

Effects of ischemia and omeprazole preconditioning on functional recovery of isolated rat heart

Objective

The aim of this study was to compare protective effects of ischemic and potential protective effects of pharmacological preconditioning with omeprazole on isolated rat heart subjected to ischemia/reperfusion.

Methods

The hearts of male Wistar albino rats were excised and perfused on a Langendorff apparatus. In control group (CG) after stabilization period, hearts were subjected to global ischemia (perfusion was totally stopped) for 20 minutes and 30 minutes of reperfusion. Hearts of group II (IPC) were submitted to ischemic preconditioning lasting 5 minutes before 20 minutes of ischemia and 30 minutes of reperfusion. In third group (OPC) hearts first underwent preconditioning lasting 5 minutes with 100μM omeprazole, and then submitted 20 minutes of ischemia and 30 minutes of reperfusion.

Results

Administration of omeprazole before ischemia induction had protective effect on myocardium function recovery especially regarding to values of systolic left ventricular pressure and dp/dt max. Also our findings are that values of coronary flow did not change between OPC and IPC groups in last point of reperfusion.

Conclusion

Based on our results it seems that ischemic preconditioning could be used as first window of protection after ischemic injury especially because all investigated parameters showed continuous trend of recovery of myocardial function. On the other hand, preconditioning with omeprazole induced sudden trend of recovery with positive myocardium protection, although less effective than results obtained with ischemic preconditioning not withstand, we must consider that omeprazole may be used in many clinical circumstances where direct coronary clamping for ischemic preconditioning is not possible.     

IL-10 is not protective in intestinal ischemia reperfusion injury

BACKGROUND: Ischemia/reperfusion of the small intestine disrupts gut barrier function, increases bacterial translocation, and activates systemic pro-inflammatory responses. Pharmacological treatment with the anti-inflammatory cytokine interleukin-10 (IL-10) following ischemia to muscle reduces the severity of local and systemic inflammation. While endogenous IL-10 is protective in murine models of acute endotoxemia, its physiological role during direct gut injury is unknown. PATIENTS AND MATERIALS: Mice genetically deficient in IL-10 (IL-10(-/-)) and their normal littermates (IL-10(+/+)) underwent 20 to 50 min of gut ischemia by occlusion of the superior mesenteric artery. RESULTS: Both short- and long-term (>16 h) survival after reperfusion of IL-10(-/-) mice was identical to that of the wild-type littermates, with 50% mortality observed at 35 min of occlusion. The small bowel demonstrated discrete gross areas of hemorrhage and ischemia localized to the jejunum. No significant difference in the extent or time for occurrence of macroscopic or microscopic intestinal damage to the small bowel was observed in IL-10(-/-) or IL-10(+/+) mice, despite the marked elevation in serum IL-6. CONCLUSIONS: The absolute serum concentration of IL-6 in the presence or the absence of IL-10 does not affect local or systemic response to ischemic intestinal injury. These results also demonstrate that the anti-inflammatory cytokine IL-10 does not play a significant local or systemic protective role in this model of ischemia/reperfusion.     

Ischemic Preconditioning Protects the Pig Liver by Preserving the Mitochondrial Structure and Downregulating Caspase-3 Activity

Summary Background Data: The beneficial effects of ischemic preconditioning (IPC) on hepatic ischemia-reperfusion injury (I/RI) have been described. However, the way in which IPC causes the changes in mitochondrial ultrastructure seen in hepatic I/RI is not well understood. Objective: The objective of the present study was to determine whether IPC protects the liver from changes in mitochondrial structure and caspase 3 activity in the early phase of post-ischemic injury. Methods: A pig model consisting of 90 min of hepatic ischemia and 180 min of reperfusion was employed. Eighteen female pigs were randomly divided into three groups: sham-operated, non-preconditioned, and ischemic preconditioned (10 min ischemia followed by 10 min reperfusion). Serum concentrations of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and thiobarbituric acid reactive substances (TBARS), as well as bile flow, were measured. Liver biopsies were taken after reperfusion for histological, immunohistochemical (anti-caspase 3), and ultrastructural examinations. Results: The IPC procedure increased bile flow (p < 0.01), reduced serum AST level (p < 0.01), and reduced serum concentration of TBARS at 180 min of reperfusion (p = 0.05). Ischemic-preconditioned liver cells had less caspase 3 activity than the non-preconditioning group (p < 0.01), and changes in mitochondrial ultrastructure were reduced (p < 0.01). Conclusion: IPC exerts a powerful protective effect against hepatic I/RI in the early phase of reperfusion, which may be mediated by preservation of mitochondrial structure and inhibition of caspase-3 activity.     

阿片受体在缺血预处理中的作用

目的　研究阿片受体激动在缺血预处理限制心肌梗死(心梗)范围作用中的地位。方法　建立在体家兔心脏缺血再灌注模型,观察阿片受体拮抗剂纳洛酮对缺血预处理心肌保护作用的影响。在离体兔心模型上,观察阿片受体激动剂吗啡代替短暂缺血刺激,对随后发生的急性心梗范围的影响,并用纳洛酮及蛋白激酶C阻断剂Chelerythrine分别进行干预。电镜观察吗啡预处理对离体心脏缺血心肌超微结构改变的影响。结果　在体心脏模型上,5min缺血及10min再灌注的预处理可显著缩小随后30min缺血引起的心梗范围(P＜0.01);在预处理前10min或30min、缺血前5min给予纳洛酮均可使预处理所产生的限制心梗范围的作用消失。离体心脏经历5min全心缺血及10min再灌注的预处理后,可缩小心梗范围(P＜0.05);在缺血前如给予吗啡预灌注15min,亦可缩小心梗范围(P＜0.05),其作用可被纳络酮或Chelerythrine分别阻断。30min缺血后心肌线粒体等超微结构严重受损,如给吗啡预灌流,心肌损伤明显减轻。结论　阿片受体激动参与了缺血预处理的心肌保护;吗啡预处理可以减轻缺血心肌损伤及产生限制心梗范围的心肌保护作用;吗啡是通过心脏局部的阿片受体介导,激活蛋白激酶C从而产生心肌保护作用。     

Protective effects of ischemic preconditioning for liver resection performed under inflow occlusion in humans

OBJECTIVE: To determine whether ischemic preconditioning protects the human liver against a subsequent period of ischemia in patients undergoing hemihepatectomy, and to identify possible underlying protective mechanisms of ischemic preconditioning, such as inhibition of hepatocellular apoptosis. SUMMARY BACKGROUND DATA: Ischemic preconditioning is a short period of ischemia followed by a brief period of reperfusion before a sustained ischemic insult. Recent studies in rodents suggest that ischemic preconditioning is a simple and powerful protective modality against ischemic injury of the liver. The underlying mechanisms are thought to be related to downregulation of the apoptotic pathway. METHODS: Twenty-four patients undergoing hemihepatectomy for various reasons alternatively received ischemic preconditioning (10 minutes of ischemia and 10 minutes of reperfusion) before transection of the liver performed under inflow occlusion for exactly 30 minutes. Liver wedge and Tru-cut biopsy samples were obtained at the opening of the abdomen and 30 minutes after the end of the hepatectomy. Serum levels of aspartate transferase, alanine transferase, bilirubin and prothrombin time were determined daily until discharge. Hepatocellular apoptosis was evaluated by in situ terminal deoxynucleotidyl transferase mediated d-UTP nick end-labeling (TUNEL) assay and electron microscopy. Caspase 3 and 8 activities were measured in tissue using specific fluorometric assays. RESULTS: Serum levels of aspartate transferase and alanine transferase were reduced by more than twofold in patients subjected to ischemic preconditioning versus controls. The analysis of a subgroup of patients with mild to moderate steatosis indicated possible increased protective effects of ischemic preconditioning. In situ TUNEL staining demonstrated a dramatic reduction in the number of apoptotic sinusoidal lining cells in the ischemic preconditioning group. Electron microscopy confirmed features of apoptosis present in control but not in ischemic preconditioning patients. There was no significant difference in caspase 3 and 8 activity when patients with ischemic preconditioning were compared with controls. CONCLUSIONS: Ischemic preconditioning is a simple and effective modality protecting the liver against subsequent prolonged periods of ischemia. This strategy may be a more attractive technique than intermittent inflow occlusion, which is associated with increased blood loss during each period of reperfusion.     

Effect of ischemic preconditioning on hepatic microcirculation and function in a rat model of ischemia reperfusion injury

Ischemic preconditioning (IPC) may protect the liver from ischemia reperfusion injury by nitric oxide formation. This study has investigated the effect of ischemic preconditioning on hepatic microcirculation (HM), and the relationship between nitric oxide metabolism and HM in preconditioning. Rats were allocated to 5 groups: 1. sham laparotomy; 2. 45 minutes lobar ischemia followed by 2-hour reperfusion (IR); 3. IPC with 5 minutes ischemia and 10 minutes reperfusion before IR; 4. L-arginine before IR; and 5. L-NAME + IPC before IR. HM was monitored by laser Doppler flowmeter. Liver transaminases, adenosine triphosphate, nitrites + nitrates, and guanosine 3'5'-cyclic monophosphate (cGMP) were measured. Nitric oxide synthase (NOS) distribution was studied using nicotinamide adeninine dinucleotide phosphate (NADPH) diaphorase histochemistry. At the end of reperfusion phase, in the IR group, flow in the HM recovered partially to 25.8% of baseline (P < .05 versus sham), whereas IPC improved HM to 49.5% of baseline (P < .01 versus IR). With L-arginine treatment, HM was 31.6% of baseline (NS versus IR), showing no attenuation of liver injury. In the preconditioned group treated with L-NAME, HM declined to 10.2% of baseline, suggesting not only a blockade of the preconditioning effect, but also an exacerbated liver injury. Hepatocellular injury was reduced by IPC, and L-arginine and was increased by NO inhibition with L-NAME. IPC also increased nitrate + nitrate (NOx) and cGMP concentrations. NOS detected by NADPH diaphorase staining was associated with hepatocytes and vascular endothelium, and was induced by IPC. IPC induced NOS and attenuated HM impairment and hepatocellular injury. These data strongly suggest a role for nitric oxide in IPC. (Liver Transpl 2002;8:1182-1191.)     

Ischemic preconditioning and liver tolerance to warm or cold ischemia: experimental studies in large animals

BACKGROUND: In the rodent, ischemic preconditioning (IPC) has been shown to improve the tolerance of the liver to ischemia-reperfusion under normothermic or hypothermic conditions. The aim of the present study was to test this hypothesis in a dog model, which may be more relevant to the human. METHODS: Beagle dogs were used in two distinct animal models of hepatic warm ischemia and orthotopic liver transplantation (hypothermic ischemia). IPC consisted of 10 minutes of ischemia followed by 10 minutes of reperfusion. In the first model, livers were exposed to 55 minutes prolonged warm ischemia and reperfused for 3 days (n = 6). In the second model, livers were retrieved and preserved for 48 hours at 4 degrees C in University of Wisconsin solution, transplanted, and reperfused without immunosuppression for 7 days (n = 5). In each model, nonpreconditioned animals served as controls (n = 5 in each group). Also, isolated dog hepatocytes were subjected to warm and cold storage ischemia-reperfusion to model the animal transplant studies using IPC. RESULTS: In the first model (warm ischemia), IPC significantly decreased serum aminotransferase activity at 6 and 24 hours post-reperfusion. After 1 hour of reperfusion, preconditioned livers contained more adenosine triphosphate and produced more bile and less myeloperoxidase activity (neutrophils) relative to controls. In the second model (hypothermic preservation), IPC was not protective. Finally, IPC significantly attenuated hepatocyte cell death after cold storage and warm reperfusion in vitro. CONCLUSIONS: IPC is effective in large animals for protecting the liver against warm ischemia-reperfusion injury but not injury associated with cold ischemia and reperfusion (preservation injury). However, the IPC effect observed in isolated hepatocytes suggests that preconditioning for preservation is theoretically possible.     

肿瘤坏死因子-α在心肌缺血预处理中的变化及其意义

目的 探讨缺血预处理(IP)对心肌细胞的保护机制.方法 在我院行瓣膜置换手术的患者36例,根据是否采取缺血预处理分为预处理组(20例)和对照组(16例),比较两组炎症因子肿瘤坏死因子(TNF)-α和心肌细胞bcl-2、Caspase-3的变化,观察IP对机体炎症反应的影响.结果 两组患者细胞因子TNF-α均在主动脉开放6 h时达到高峰,术后5 d恢复到术前水平.预处理组开放后6 h、术后1 d、术后2 d TNF-α水平明显低于对照组(P＜0.05).复灌后对照组心肌细胞bcl-2蛋白表达轻度提高,与阻断前比较差异无统计学意义;而复灌后IP组心肌细胞bcl-2蛋白表达明显提高,与阻断前和对照组比较,差异有统计学意义(P＜0.05).对照组复灌后心肌细胞Caspase-3、TNF-α蛋白表达显著提高,IP组心肌细胞中Caspase-3、TNF-α蛋白表达也增高,但比对照组降低,两组间差异有统计学意义(P＜0.05).术后2 d血清TNF-α与bcl-2呈负相关;术后2 d血清及开放后心肌TNF-α与Caspase-3有相关性.结论 缺血预处理可能通过降低机体的炎症反应途径达到心肌细胞保护的效果.