肝细胞内糖原含量与肝脏缺血再灌注损伤关系的实验研究

目的　探讨细胞内糖原含量与肝脏缺血再灌注损伤之间的关系及其相关机理。方法　对3组糖原含量显著不同的兔肝脏缺血再灌注过程中的组织形态学、肝脏酶学、组织ATP含量及细胞膜Na K ATP酶,Ca2 ATP酶活性进行观察。结果　糖原含量高的肝脏,其细胞能量代谢旺盛、细胞膜Na K ATP酶及Ca2 ATP酶活性强、组织结构及功能损伤轻。结论　缺血前增加肝糖原负荷可显著减轻肝脏缺血再灌注损伤程度。     

肝细胞糖原拮抗肝脏缺血再灌注损伤及其相关机理的研究

为了探讨增加细胞内糖原含量能否改善肝脏缺血再灌注损伤程度,本实验对３组糖原含量显著不同的兔肝脏缺血再灌注过程中的组织形态学,肝脏酶学,组织ＡＴＰ含量,细胞膜Ｃａ＾２＋－ＡＴＰ酶活性及胞浆内游离Ｃａ＾２＋浓度进行了观察。结果：糖原含量高的肝脏,其细胞能量代谢旺盛,细胞膜Ｃａ＾２＋－ＡＴＰ酶活性强,胞浆内游离Ｃａ＾２＋浓度相对稳定,组织结构及功能损作轻,本实验结果提示：缺血前增加肝脏糖含量可显著了拮抗     

BQ123与L-arginine在肝脏缺血再灌注损伤中的作用研究

目的：探讨肝脏缺血再灌注损伤的机制及BQ123或（和）L－arginine能否有效改善肝脏缺血再灌注损伤。方法：在大鼠肝脏缺血再灌注损伤模型基础上，对组织形态学、肝脏酶学、透明质酸、血浆内皮素及免疫组织化学染色情况进行观测。结果：肝脏缺血再灌注损伤时，血肝酶、透明质酶、血浆内皮素水平均显著增高，再灌注前使用BQ123或（和）L-arginine的肝脏，其组织结构及功能损伤均显著减轻。结论：肝脏缺血再灌注损伤与肝脏微循环改善有关，若能在肝脏再灌注前使用BQ123或（和）L－arginine，可有效改善肝脏微循环，从而减轻肝缺血再灌注损伤。     

内皮素-1与肝脏缺血再灌注损伤的实验研究

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

羟丁酸钠对大鼠肝脏热缺血再灌注损伤的保护作用

目的探讨羟丁酸钠（γ-hydroxybutyrate,GHB）对肝脏热缺血再灌注损伤的保护作用.方法采用大鼠肝脏热缺血再灌注模型,观察肝脏组织形态学、酶学、脂质过氧化、细胞凋亡及血浆内皮素的变化.结果肝脏缺血再灌注损伤时,血清ALT、AST、LDH、血浆内皮素、TNF-α及肝组织MDA水平均显著增高,细胞凋亡增加,肝组织SOD明显减少.再灌注前使用GHB可以明显减轻上述改变,钠络酮可以部分阻断GHB的作用.结论GHB可能通过抗脂质过氧化,下调肝脏内皮素的表达,改善微循环,减少肝细胞凋亡,从而减轻热缺血再灌注对肝脏的损伤作用.     

山莨菪碱对大鼠肝脏缺血再灌注损伤保护作用的实验研究

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

兔后肢缺血再灌注后足背肌腱表面微循环变化

应用活体显微镜技术,观察了兔后肢常温止血带缺血2h(n=8)及5h(n=8)再灌注后最初1h 期间足背肌腱表面微循环动态变化,尤其是白细胞内皮粘附及微血管灌注状况的变化,旨在探讨缺血再灌注损伤的发生机制,从而指导临床治疗。结果表明:①肢体缺血再灌注后缺血组织微静脉内皮上粘附的白细胞数显著增加,而且缺血时间越长,增加越显著。②肢体缺血5h 再灌注后,缺血组织的微循环并不能均匀恢复,部分区域发生“无复流现象”,包括原发性无复流和继发性毛细血管灌注衰竭两种形式。提示:①白细胞内皮粘附参与了缺血再灌注损伤的病理生理过程。②肢体缺血5h 再灌注后发生的局部组织损伤并非皆属缺血再灌注损伤,部分区域可能系单纯缺血性损伤,部分区域则可能属缺血再灌注—继发性缺血损伤。     

一氧化氮,血栓素／前列环素及ATP与肝硬变大鼠肝脏缺血再灌注损伤

四氯化碳复制大鼠肝硬变模型，通过大鼠肝脏缺血再灌注损伤模型，比较硬变肝与正常肝在缺血再灌注损伤时的差异和意义。结果显示：肝硬变时对缺血再灌注损伤反应与正常大鼠不同，再灌注时肝脏一氧化氮合成释放显著增加，肝组织ＡＴＰ含量明显降低且长时间难以恢复等，可能是肝硬变时对缺血再灌注损伤更敏感、更易发生肝功能衰竭的重要原因。     

血小板激活因子及其拮抗剂在肝脏缺血再灌注损伤中的作用

通过对肝脏缺血再灌注损伤过程中PAF的变化进行初步研究,观察PAF拮抗剂海风藤酮对大鼠肝脏局部缺血再灌注损伤的保护作用。结果表明:肝脏缺血再灌注损伤时肝组织PAF含量增高,且随再灌注时间的延长而加重,海风藤酮可以拮抗损伤过程中PAF增高所致的药理作用,减轻肝脏脂质过氧化程度,改善肝功能,减轻肝脏病变程度。结果提示PAF拮抗剂可能为肝脏缺血再灌注损伤的防治提供一条新的途径。     

兔后肢缺血再灌注后足背肌腱表面微循环变化

应用活体显微镜技术，观察了兔后肢常温止血带缺血２ｈ（ｎ＝８）及５（ｎ＝８）再灌注后最初１ｈ期间足背肌腱表面微循环动态变化，尤其是白细胞内皮粘附及微血管灌注状况的变化，旨在探讨缺血再灌注损伤的发生机制，从而指导临床治疗。结果表明：①肢体缺血再灌注后缺血组织微静脉内皮上粘附的白细胞数显著增加，而且缺血时间越长，增加越显著。②肢体缺血５ｈ再灌注后，缺血组织的微循环并不能均匀恢复，部分区域发生＂无复流现象＂，包括原发性无复流和继发性毛细血管灌注衰竭两种形式。提示：①白细胞内皮粘附参与了缺血再灌注损伤的病理生理过程。②肢体缺血５ｈ再灌注后发生的局部组织损伤并非皆属缺血再灌注损伤，部分区域可能系单纯缺血性损伤，部分区域则可能属缺血再灌注一继发性缺血损伤。     

Hepatic microcirculatory failure

Liver ischemia has been considered a frequent problem in medical practice, and can be associated to a number of surgical and clinical situations, such as massive hepatic resections, sepsis, liver trauma, circulatory shock and liver transplantation. After restoring blood flow, the liver is further subjected to an additional injury more severe than that induced by ischemia. On account of the complexity of mechanisms related to pathophysiology of ischemia and reperfusion (I/R) injury, this review deals with I/R effects on sinusoidal microcirculation, especially when steatosis is present. Alterations in hepatic microcirculation are pointed as a main factor to explain lower tolerance of fatty liver to ischemia-reperfusion insult. The employment of therapeutic strategies that interfere directly with vasoactive mediators (nitric oxide and endothelins) acting on the sinusoidal perfusion seem to be determinant for the protection of the liver parenchyma against I/R. These approaches could be very suitable to take advantage of marginal specimens as fatty livers, in which the microcirculatory disarrangements hamper its employment in liver transplantation.     

The role of nitric oxide after a short period of liver ischemia-reperfusion

BACKGROUND: Liver ischemia-reperfusion injury is a serious problem during liver resection and transplantation. Nitric oxide (NO) has been suggested to have a cytoprotective effect for microcirculation, while the interaction of active oxygen species and NO produces peroxynitrite anion. The present study attempts to clarify the role of NO in liver ischemia-reperfusion injury. METHODS: Wistar male rats were subjected to 30 min of hepatic ischemia followed by reperfusion. The model rats were divided into the three following groups: a control group that was not administered NO synthase inhibitors, and two experimental groups that were administered either N(omega)-nitro-L-arginine methyl ester (L-NAME) or aminoguanidine. In each group, we examined active oxygen species and nitric oxide production, and investigated liver function by measuring serum transaminase levels. In addition, we conducted histopathologic examinations and microcirculation examinations using intravital videomicroscopy. RESULTS: In the control group, NO concentrations in the plasma increased with time after reperfusion. A decrease in NO production was detected in the groups administered NO synthase inhibitors. Elevated serum transaminase levels became more prominent after L-NAME administration, while aminoguanidine administration reduced its level. The degree of microcirculation failure was found to be more prominent in the L-NAME-administered group over both the control group and the aminoguanidine-administered group. A significantly lower survival rate was observed at 6 h after reperfusion in the L-NAME-administered group over that of the other groups. CONCLUSIONS: A reduction of the ischemia-reperfusion injury is important in inhibiting the production of high-output NO and peroxynitrite, and in maintaining NO levels necessary for maintenance of microcirculation.     

麦角新碱预处理对减轻大鼠移植肝缺血再灌注损伤的作用

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

Hepatic ischemia-reperfusion injury.

BACKGROUND: The morbidity associated with liver transplantation and major hepatic resections is partly a result of ischemia-reperfusion injury. DATA SOURCES: The entire world literature on the subject was searched via Medline. Keywords included reperfusion injury, transplantation, liver resection, nitric oxide, endothelin, cytokines, Kupffer cells, ischemic/ischaemic preconditioning, and nuclear factor-kappa B. CONCLUSIONS: An imbalance between endothelin and nitric oxide levels results in failure of the hepatic microcirculation at the onset of reperfusion. Activation of nuclear factor-kappa B in the liver promotes proinflammatory cytokine and adhesion molecule synthesis. These result in oxygen-derived free radical production and neutrophil recruitment, further contributing to cellular injury. Various therapeutic modalities acting on the above mediators have been successfully used to attenuate reperfusion injury in animal models of hepatic transplantation and resection. Application of the knowledge gained from animal models of hepatic ischemia-reperfusion to the clinical setting will improve the outcome of hepatic surgery.     

潘生丁对大鼠移植肝缺血再灌注损伤保护作用的实验研究

目的探讨潘生丁对大鼠移植肝缺血再灌注损伤的保护作用及其相关机制。方法采用改良Kam ada“二袖套法”制作肝移植模型,48只雄性SD大鼠随机分成3组:假手术组(A组),同基因大鼠肝移植组(B组)和潘生丁预处理 同基因大鼠肝移植组(C组)。于供肝再灌注2 h后测定各组PAGT、ALT、AST、SOD、MDA含量,比较肝细胞凋亡和组织形态学改变。结果移植肝再灌注2 h后C组较B组肝组织损害轻,SOD、MDA、ALT、AST、PAGT变化及肝细胞凋亡差异有显著性(P<0.05)。结论潘生丁可通过改善肝脏微循环状况,改善缺血肝脏组织的能量代谢,提高肝组织抗氧化能力,抑制细胞凋亡和减少肝脏细胞的变性坏死程度而对肝脏热缺血再灌注起保护作用。     

Analysis of ischemia-reperfusion injury in a microcirculatory model of pressure ulcers

The aim of this study was to establish a pressure ulcer model that visualizes the microcirculation, and to examine the participation of ischemia-reperfusion injury in the pathophysiology of pressure ulcers. An original system composed of a new skin fold chamber and compression device allowed loading quantitative vertical stress to the skin. An intravital microscopic technique enabled direct visualization of the microcirculation in the physiological condition and in response to pressure application. To estimate the effect of ischemia-reperfusion injury, animals were divided into two groups: the compression-release group (n = 8), in which the animals received four cycles of compression-release which consisted of 2 hours of compression followed by 1 hour of pressure release; and the compression alone group (n = 8) in which the animals underwent continuous compression for 8 hours. Functional capillary density was quantified before the compression procedure and on day 1 (35 hours) after the first evaluation. The cyclic compression-release procedure significantly decreased functional capillary density as compared to continuous compression, indicating that in our experimental setting repetition of ischemia-reperfusion cycle more severely damaged the microcirculation than single prolonged ischemic insult. This finding supports the significant contribution of ischemia-reperfusion injury to the pathophysiology of pressure ulcers at the level of dynamic in vivo microcirculation.     

Warm hepatic ischemia in pigs: effects of L-arginine and oligotide treatment.

Reperfusion injury represents a key event leading to graft nonfunction. Maintaining adequate nitric oxide levels and stimulating vasodilator synthesis can probably minimize endothelial damage. The aim of this study was to investigate the effect of L-arginine, a substrate of nitric oxide synthesis, and oligotide, a promoter of prostacyclin synthesis, on liver function and morphology after warm ischemia-reperfusion injury. After constructing a side-to-side portacaval shunt, ischemia was induced by clamping the hepatic hilum for 2 h above the shunt, in 19 female pigs divided into a control group (n = 7), an L-arginine treatment group (n = 6), and an oligotide treatment group (n = 6). Liver function tests and measurements of serum and red blood cell malondialdehyde and plasma nitric oxide levels were performed before reperfusion and at 1, 10, 60, and 120 min after reperfusion. Liver biopsies, taken before reperfusion and at 30 min and 7 days after reperfusion, were analyzed for tissue malondialdehyde, histological-ultrastructural features, and apoptosis evaluation. Thirty minutes after reperfusion, liver malondialdehyde, sinusoidal congestion, necrosis, and apoptosis were significantly lower in the L-arginine group than in the controls (p < .05). On postoperative day 7, tissue malondialdehyde decreased, while plasma nitric oxide and hepatocyte glycogen content were increased in the L-arginine group compared to controls (p < .05). This study demonstrates the protective effect of L-arginine on hepatic lipoperoxidation and liver morphology in a pig model of warm ischemia-reperfusion injury. The increased plasma levels of nitric oxide a week after ischemia-reperfusion injury support the hypothesis that it has a role in preventing liver damage. The same beneficial effect was not confirmed for oligotide.     

Orthogonal polarization spectral imaging as a tool for the assessment of hepatic microcirculation: a validation study

BACKGROUND: Quantitative analysis of liver microcirculation using intravital fluorescence microscopy in animals has increased our knowledge about ischemia-reperfusion injury. However, because of the size of the instrumentation and the necessity of fluochromes for contrast enhancement, human liver microcirculation cannot be observed. Orthogonal Polarization Spectral (OPS) imaging is a recently introduced technique that can be used to visualize the microcirculation without the need for fluorescent dyes. It is a small, hand-held device and could potentially be used to study the microcirculation of the human liver in a clinical setting. However, before implementation into clinical use its ability to quantitatively measure microcirculatory parameters must be validated. METHODS: The livers of Spraque-Dawley rats (n=9) were exteriorized, and images were obtained using OPS imaging and intravital fluorescence microscopy of the identical microvascular regions before and after the induction of a 20-min warm lobar ischemia. Images were videotaped for later computer-assisted off-line analysis. RESULTS: OPS imaging can be used to accurately quantify the sinusoidal perfusion rate, vessel diameter, and venular red blood cell velocity. Correlation parameters were significant and Bland-Altman analyses showed good agreement for data obtained from the two methods at baseline as well as during reperfusion. CONCLUSION: OPS imaging can be used to quantitatively measure microcirculatory parameters in the rat liver under both physiological and pathophysiological conditions. Thus, OPS imaging has the potential to be used to make quantitative measurements of the microcirculation in the human liver.