糖尿病大鼠离体心脏对缺血/再灌注的耐受性

目的评价糖尿病大鼠离体心脏缺血／再灌注后心脏功能的改变。方法链佐星(60mg／kg)诱导的糖尿病大鼠16只(D组),年龄匹配的健康雄性SD大鼠10只(C组),戊巴比妥钠(60mg／kg)麻醉后快速取出心脏,接上主动脉插管置于Langendorg装置上,Krebs-Henseleit缓冲液逆行灌注。平衡灌注20min,待心率(HR)及冠脉流量平稳后夹闭灌注道,进行全心缺血30min,复灌40min。持续监测心肌心电活动、左心室压峰值(LVPSP)、左室舒张末压(LVEDP)和左室压力最大上升／下降速率(±dp／dtmax),计算左室发展压(LVDP=LVPSP-LVEDP),用LVDP×HR(RDPP)表示左室作功。结果与C组相比,基础状态下,D组大鼠心脏HR减慢,LVDP、RDPP和±dp／dtmax降低,LVEDP升高(P<0.05或0.01);再灌注后HR、LVDP、RDPP、冠脉流出液、±dp／dtmax等心功能指标恢复百分率升高,肌酸激酶活性降低(P<0.05或0.01);心脏缺血-停搏时间延长。结论糖尿病心脏基础心功能损伤严重,但对缺血／再灌注的耐受性增强。     

一氧化氮在异丙酚减轻大鼠离体心脏缺血再灌注损伤中的作用

目的 探讨一氧化氮在异丙酚减轻大鼠离体心脏缺血再灌注损伤中的作用.方法 成年雄性SD大鼠,体重220～330 g,采用Langendorff装置建立大鼠离体心脏灌注模型,选取符合实验标准的心脏模型18个,随机分为3组(n=6):K-H液灌注组(A组)、异丙酚灌注组(B组)和异丙酚+L-NAME灌注组(C组).各组用相应的K-H液灌注15 min,常温全心停灌20 min,然后用相应的K-H液复灌60 min.采用电化学微传感器法测定心肌一氧化氮(NO)含量,免疫组化法测定心肌一氧化氮合酶(NOS)含量,分光光度计测定心肌NOS活性,测定心率、左心室舒张末压、左心室发展压、左心室压变化速率最大值和左心室压变化速率最小值,定时收集右心室流出液以测定冠脉流量.结果 与A组相比,B组和C组复灌后各时点心功能改善(P＜0.05);与C组相比,B组复灌后各时点心功能改善(P＜0.05);B组心肌NO含量和NOS活性较A组升高(P＜0.05),但2组心肌NOS含量差异无统计学意义(P＞0.05).结论 缺血再灌注导致心肌内源性NO的含量降低.异丙酚减轻大鼠离体心脏缺血再灌注损伤可能是通过增加心肌内源性NO生成实现的.     

Temperature dependency of calcium-induced reperfusion injury in the isolated rat heart

The temperature dependence of Ca-induced reperfusion injury was studied in an isolated rat heart preparation. Hearts were subjected to 90 minutes of hypothermic arrest (20 degrees C) followed by 15 minutes of reperfusion at 20, 28, or 37 degrees C with a reperfusate containing various concentrations of Ca (0.1-2.55 mM). When reperfusion was started at 37 degrees C, the Ca concentration in the reperfusate significantly affected both postischemic functional recovery and creatine kinase leakage. Bell-shaped dose-response curves were observed. The optimal Ca concentration for 37 degrees C reperfusion was between 0.3 and 0.7 mM. When reperfusion was started at 20 degrees C, neither functional recovery nor creatine kinase leakage was dependent on the Ca concentration in the reperfusate. At 28 degrees C, functional recovery was not dependent on the Ca concentration, however, creatine kinase leakage was. These results indicate that Ca-induced reperfusion injury depends on the temperature of the reperfusate and that the boundary temperature of the reperfusate at which Ca-induced reperfusion injury becomes manifest seems to be near 28 degrees C.     

Na~ /H~ 交换抑制剂对离体鼠心肌缺血/再灌注损伤的保护作用

目的　探讨Na H 交换抑制剂二甲基阿米洛利 (DMA)对心肌缺血 再灌注损伤(MIRI)的影响。方法　离体鼠心脏置于Langendorff装置上 ,采用主动脉逆灌法 ,停灌 30min后复灌30min ,造成MIRI模型。离体鼠心脏稳定灌流 2 0min ,随机分成 4组 :对照组、DMA 5 μmol L组、10 μmol L组和 2 0 μmol L组 ,每组各 8例。记录心电图和心肌收缩力的变化 ,测定缺血前及再灌注30min时冠脉流出量以及心肌组织含钙量 ,观察心肌细胞超微结构变化。结果　与对照组相比 ,DMA10 μmol L组和 2 0 μmol L组心率、心肌收缩力和冠脉流出量的恢复程度明显提高 (P <0 0 5或0 0 1) ,心律失常发生率及心肌组织钙含量明显降低 (P <0 0 5或 0 0 1) ,超微结构改变轻微 ,而DMA5 μmol L组仅在再灌注 10min和 15min时心肌收缩力恢复程度高于对照组 (P <0 0 5 ) ,其他指标均无显著性变化。结论　DMA对心肌缺血 再灌注损伤有一定的保护作用 ,且与剂量有关     

Amelioration of reperfusion injury following hypothermic, ischemic cardioplegia in isolated, infarcted rat hearts

The left coronary artery was ligated and myocardial infarction developed in 28 rats. Three weeks later, the hearts were excised and mounted in an apparatus for perfusion of non-working isolated hearts (Langendorff). Hypothermic (15 degrees C), ischemic cardioplegia was induced for either 2 or 3 1/2 h followed by reperfusion for 45 min. Half of the hearts were reperfused with an initially gradual rise in temperature and pressure of the perfusion fluid, whereas the other half was reperfused directly with the perfusate at 37 degrees C and 100 cm H2O pressure. The hearts were examined by transmission electron microscopy and randomized for stereological analysis based on point counting on electron micrographs. Cardioplegia of 2 h duration was tolerated better than cardioplegia for 3 1/2 h (interstitial edema; P = 0.03, fraction of altered mitochondria; P = 0.001). Particularly in the hearts undergoing the longest cardioplegia, myocardial injury was less severe following a gentle reperfusion as compared with those exposed to the clinically common abrupt technique (fraction of mitochondria in the myocyte; P = 0.03, fraction of altered mitochondria; P = 0.008). In the interstitium, the luminal area of capillaries was significantly increased and the endothelial swelling less pronounced in the groups undergoing the gentle reperfusion technique, (luminal/endothelial fraction; P = 0.01). The study shows that previously infarcted hearts are susceptible to ischemic damage even after 2 h of regular hypothermic, ischemic cardioplegia and that a gentle reperfusion technique significantly ameliorates reperfusion injury.     

Toll-like receptor 4 mediates ischemia/reperfusion injury of the heart

BACKGROUND: Restoration of blood flow to the ischemic heart may paradoxically exacerbate tissue injury (ischemia/reperfusion injury). Toll-like receptor 4, expressed on several cell types, including cardiomyocytes, is a mediator of the host inflammatory response to infection. Because ischemia/reperfusion injury is characterized by an acute inflammatory reaction, we investigated toll-like receptor 4 activation in a murine model of regional myocardial ischemia/reperfusion injury. We used C3H/HeJ mice, which express a nonfunctional toll-like receptor 4, to assess the pertinence of this receptor to tissue injury after reperfusion of ischemic myocardium. METHODS: Wild-type mice (C3H/HeN) or toll-like receptor 4 mutant mice (C3H/HeJ) were subjected to 60 minutes of regional myocardial ischemia followed by 2 hours of reperfusion. At the end of reperfusion, the area at risk and the myocardial infarct size were measured as the end point of myocardial ischemia/reperfusion injury. Myocardial mitogen-activated protein kinase activation was measured by Western blotting, and nuclear translocation of nuclear factor-kappaB and activator protein-1 was determined by electrophoretic mobility shift assay. Ischemia/reperfusion-injured myocardium was also assessed by ribonuclease protection assay for expression of inflammatory mediators (tumor necrosis factor-alpha, interleukin-1beta, monocyte chemotactic factor-1, and interleukin-6). RESULTS: The area at risk was similar for all groups after myocardial ischemia/reperfusion injury. There was a 40% reduction in infarct size (as a percentage of the area at risk) in C3H/HeJ mice compared with C3H/HeN mice (P =.001). Within the myocardium, significant activation of c-Jun N-terminal kinase, p38, and extracellular signal-regulated kinase was observed in both strains after ischemia and during reperfusion as compared with an absence of mitogen-activated protein kinase activation during sham operations; however, c-Jun N-terminal kinase activity, but not p38 or extracellular signal-regulated kinase activity, was significantly reduced in C3H/HeJ mice (P <.05). In both groups, nuclear factor-kappaB and activator protein-1 nuclear translocation occurred in the myocardium during myocardial ischemia/reperfusion injury, but, by densitometric analysis, nuclear translocation of nuclear factor-kappaB and activator protein-1 was significantly decreased in C3H/HeJ mice compared with C3H/HeN mice. Interleukin-1beta, monocyte chemotactic factor-1, and interleukin-6 were detectable in reperfused ischemic myocardium but were not detected in sham-operated myocardium; the expression of each of these mediators was significantly decreased in the myocardial tissue of C3H/HeJ mice when compared with expression in the control C3H/HeN mouse strain. CONCLUSIONS: Our data suggest that toll-like receptor 4 may mediate, at least in part, myocardial ischemia/reperfusion injury. Inhibition of toll-like receptor 4 activation may be a potential therapeutic target to attenuate ischemia/reperfusion-induced tissue damage in the clinical setting.     

Ozone administration reduces reperfusion injury in an isolated rat heart model

BACKGROUND: Accumulating clinical experience with ozone administration for conditions associated with ischemia has been encouraging. The aim of our study was to determine the effect of ozone on reperfusion injury in an isolated rat heart model. METHODS: Isolated rat hearts were perfused with modified Krebs-Henseleit buffer solution via ascending aorta cannulation. After 15 minutes, perfusion was stopped and global ischemia was maintained for 30 minutes, following which perfusion was restarted, and continued for 40 minutes. Baseline hemodynamic measurements (heart rate, left ventricular developed pressure (LVDP), dP/dt, and coronary flow) were taken prior to ischemia, and every 10 minutes after reperfusion was started. Eleven hearts were treated with ozone during reperfusion and eight hearts served as controls. In the treatment group, after 5 minutes of reperfusion, ozone was administered in distilled water via a side arm for 5 minutes. RESULTS: Preischemic baseline hemodynamic measurements and coronary flow were similar in the two groups. Hearts treated with ozone during reperfusion exhibited better recovery than did controls. Mean (+/-SE) percent recovery for treatment and control groups, respectively, was: LVDP 69 +/- 2% vs 51 +/- 6% (p = 0.04); dP/dt 68.9 +/- 13.3% vs 53.7 +/- 20.4% (p = 0.05); and LVDPxHR 61.4 +/- 3.3% vs 44.4 +/- 3.5% (p = 0.02). CONCLUSION: In the isolated rat heart model, treatment with ozone during reperfusion enables better recovery than in controls. Although the mechanism by which ozone exerts its beneficial effect is not identified, it is possibly due to reduction in reperfusion injury.     

Lidocaine reduces ischaemic but not reperfusion injury in isolated rat heart

The local anaesthetic lidocaine protects the myocardium in ischaemia–reperfusionsituations. It is not known if this is the consequence of ananti-ischaemic effect or an effect on reperfusion injury. Therefore,we investigated the effect of two concentrations of lidocaineon myocardial ischaemia–reperfusion injury and on reperfusioninjury alone. We used an isolated rat heart model where heartrate, ventricular volume and coronary flow were kept constant.Hearts underwent 45 min of low-flow ischaemia followed by 90min reperfusion. Two groups received lidocaine 1.7 or 17 µgml^–1 starting 5 min before the onset of reperfusion. Intwo additional groups, lidocaine infusion started 5 min beforelow-flow ischaemia. In all groups, lidocaine administrationwas stopped after 15 min of reperfusion. One group served asan untreated control (n=11 in each group). Left ventriculardeveloped pressure (LVDP) and total creatine kinase release(CKR) were measured. Lidocaine administration during ischaemiaand reperfusion led to an improved recovery of LVDP during reperfusion(1.7 µg ml^–1, 54 (SEM 10) mm Hg; 17 µg ml^–1,71 (9) mm Hg at 30 min of reperfusion; both significantly differentfrom control (21 (4) mm Hg) (P<0.05)) and a reduced CKR (1.7µg ml^–1, 79 (13) IU; 17 µg ml^–1, 52(8) IU at 30 min of reperfusion; both significantly differentfrom control (130 (8) IU (P<0.05)). Lidocaine given duringearly reperfusion only, affected neither LVDP during reperfusion(1.7 µg ml^–1, 19 (6) mm Hg (P=1.0); 17 µgml^–1, 36 (8) mm Hg (P=0.46)) nor CKR (156 (21) IU (P=0.50)and 106 (14) IU (P=0.57)). We conclude that lidocaine protectsthe myocardium against ischaemic but not against reperfusioninjury in the isolated rat heart. Br J Anaesth 2001; 86: 846–52     

Biliverdin protects rat livers from ischemia/reperfusion injury

OBJECTIVE: To explore putative cytoprotective functions of biliverdin during hepatic ischemia/reperfusion (I/R) injury in rat models. MATERIAL AND METHODS: Male Sprague Dawley (SD) rat livers were harvested and stored for 24 hours at 4 degrees C in University of Wisconsin (UW) solution (n=18), and then perfused with blood for 2 hours on an isolated rat liver perfusion apparatus equipped for temperature (37 degrees C), pressure (13 cm H2O), and pH (7.3) maintenance. Biliverdin was added to the blood at concentrations of 10 and 50 micromol in two groups of six animals. Portal vein blood flow, bile production, and GOT/GPT levels were assessed serially. At the conclusion of the experiment, liver samples were collected for histologic evaluation using Suzuki criteria. RESULTS: BV exerted protective effects against liver I/R injury. Adjunctive biliverdin improved portal venous blood flow (mL/min/g) from the beginning of reperfusion (1.33+/-0.17 versus 0.98+/-0.15; P<.001) and increased bile production (mL/g) as compared with the control group (3.40 versus 1.88; P<.003). I/R-induced hepatocellular damage as measured by GOT/GPT release (IU/L) was diminished in the biliverdin group (91 versus 171 and 46 versus 144, respectively; P<.0001). Improved liver function by biliverdin was accompanied by preservation of the histologic structure as assessed by Suzuki criteria (3.7+/-1.4 versus 6.8+/-0.8 in untreated controls; P<.005). CONCLUSIONS: Biliverdin attenuates the ischemia/early reperfusion injury of rat liver grafts as assessed by hemodynamics, function, enzyme analysis, and histology. This study provides the rationale for novel therapeutic approaches using biliverdin to maximize the organ donor pool through the safer use of liver transplants despite prolonged periods of cold ischemia.     

Curcumin protects against ischemia/reperfusion injury in rat kidneys

OBJECTIVES: Renal ischemia followed by reperfusion leads to acute renal failure in both native kidneys and renal allograft. We investigated the effect of curcumin on ischemia-reperfusion (I/R) injury and the antioxidant effects of curcumin in rats. METHODS: Thirty rats were randomly divided into five experimental groups (control, sham, curcumin, I/R and I/R+curcumin, n=6 each). Curcumin was administered (200 mg kg(-1)) orally to curcumin and I/R+curcumin groups for 7 days. Then, the rats were subjected to bilateral renal ischemia for 45 min and followed by reperfusion for 24 h. All rats were killed and kidney function tests, serum and tissue nitric oxide (NO), protein carbonyl (PC), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels were determined. Histopathological examinations were also performed. RESULTS: Curcumin significantly improved the urea and cystatin C levels in I/R+curcumin group compared to I/R group (p<0.05). Reduction of serum GSH-Px was significantly improved by curcumin (p<0.001), but SOD enzyme activity did not alter (p>0.05). Treatment with curcumin also resulted in significant reduction in serum and tissue MDA, NO and PC and for tissue that were increased by renal I/R injury (p<0.001 for serum and p<0.05 for tissue, respectively). In histological examination, the rats treated with curcumin had nearly normal morphology of the kidney. CONCLUSIONS: Based on our results, it can be concluded that curcumin protects the kidneys against I/R injury via its antioxidant effects.     

心麻液预处理的心肌保护效果

目的 研究Ｓｔ．Ｔｈｏｍａｓ晶体心麻液预处理和缺血预处理对大鼠缺血／再灌注心肌收缩功能的影响。方法 应用离体Ｌａｎｇｅｎｄｏｒｆｆ逆行灌注模型,观察晶体心麻液预处理的缺血预处理时心肌缺血及再灌注前后ＬＶＥＤＰ,ＤＰ,＋ｄｐ／ｄｔ－,ＣＦ冠脉流出液中蛋白含量和ＬＤＨ活性以及再灌注后心肌ＳＯＤ活性,ＭＤＡ和ＡＴＰ含量的变化。结果 在缺血期间,晶体心麻液预处理显著延迟了心肌缺血性挛缩的发生时间。     

依达拉奉对大鼠小肠缺血-再灌注所致肺损伤的保护作用

目的探讨依达拉奉对大鼠小肠缺血-再灌注所致肺损伤的保护作用。方法雄性SD大鼠18只,随机均分为假手术组(Sham组),缺血-再灌注组(IR组)和依达拉奉组(E组)。Sham组只分离肠系膜上动脉,不做其他处理;IR组分离肠系膜上动脉,从大鼠尾静脉注射与E组等量的生理盐水后,用无创动脉夹夹闭120min后移去动脉夹,再灌注120min;E组在缺血-再灌注前静脉注射依达拉奉6mg/kg。再灌注120min后采集标本。肺组织HE染色后病理学检测,采集腹主动脉血液检测大鼠血清中TNF-α和IL-6浓度,取肺组织检测髓过氧化物酶(MPO)活性和丙二醛(MDA)浓度。结果与Sham组比较,IR组肺泡上皮细胞广泛水肿、炎性细胞浸润、肺泡肺萎陷、肺毛细血管扩张出血;E组肺组织病理改变较IR组明显改善,肺泡炎性渗出减少;E组病理评分为(2.1±0.7)分,明显低于IR组的(5.7±1.1)分,IR组病理评分明显高于Sham组的(1.5±0.2)分(P0.01);血清中TNF-α和IL-6的浓度明显少于IR组,肺组织中MPO活性和MDA浓度明显低于IR组(P0.01)。结论依达拉奉能够明显改善小肠缺血-再灌注性肺损伤。     

miRNA在缺血预处理保护心肌缺血/再灌注损伤中的研究进展

背景microRNAs(miRNAs)是一类由动物、 植物和病毒基因组编码的约由22个核苷酸组成的小分子单链RNA.近年来发现它在缺血预处理(ischemic preconditioning,IPC)中发挥着重要作用,有望成为研究治疗心肌缺血/再灌注损伤(ischemia/reperfusion injury,I/RI)的新靶点.目的 阐述不同miRNA在IPC保护心肌I/RI中的作用及其可能机制.内容IPC是经典的保护心肌I/RI的方法,IPC后心肌中miRNA表达谱发生改变,其中miR-1、miR-21、miR-133b-5p、miR-199a、miR-144/451可能通过不同的基因调节机制影响IPC保护心肌I/RI.趋向将miRNA与靶基因、 信号调节通路相结合将是未来研究miRNA调节IPC保护心肌I/RI的重要趋势.     

舒芬太尼后处理和七氟醚后处理对大鼠离体心脏缺血再灌注损伤的影响

目的 评价舒芬太尼后处理和七氟醚后处理对大鼠离体心脏缺血再灌注损伤的影响.方法 雄性SD大鼠,体重230～250 g,成功制备Langendorff离体灌注模型的40个心脏随机分为4组(n=10):缺血再灌注组(Ⅰ组)、七氟醚后处理组(Ⅱ组)、舒芬太尼后处理组(Ⅲ组)和七氟醚联合舒芬太尼后处理组(Ⅳ组).采用K-H液平衡灌注(灌注压10 kPa)30 min,全心缺血40 min再灌注120 min.再灌注即刻时Ⅱ组、Ⅲ组和Ⅳ组进行药物后处理15 min:Ⅱ组K-H液中通入3.0%七氟醚,Ⅲ组K-H液中加入100 nmol/L舒芬太尼,Ⅳ组同时进行七氟醚后处理和舒芬太尼后处理.分别于平衡灌注末(基础状态)、再灌注15 min、30 min、60 min、90 min、120 min时记录左室收缩压(LVSP)、左室舒张末压(LVEDP)、左室发展压(LVDP)、左室内压上升最大速率(+dp/dtmax)、左室内压下降最大速率(-dp/dtmax)、心率(HR)和灌脉流量(CF).再灌注5 min时,收集冠脉流出液,测定肌酸激酶(CK)和乳酸脱氢酶(LDH)的活性.再灌注120 min时取心肌组织,测定心肌梗死体积、Bcl-2和Bax的表达水平,并计算Bcl-2和Bax表达的比值(Bcl-2/Bax).结果 与Ⅰ组比较,Ⅱ组、Ⅲ组和Ⅳ组LVSP、LVDP、+dp/dtmax、-dp/dtmax和CF升高,LVEDP和LDH、CK的活性降低,心肌梗死体积缩小,Bcl-2表达上调,Bax表达下调,Bcl-2/Bax升高(P＜0.05或0.01);Ⅱ组、Ⅲ组和Ⅳ组间上述指标差异无统计学意义(P＞0.05).结论 舒芬太尼后处理可减轻大鼠心肌缺血再灌注损伤,联合七氟醚后处理时心肌保护作用并未增加,其心肌保护的机制与上调Bcl-2表达、下调Bax表达从而抑制细胞凋亡有关.     

Calpain inhibitor-1 reduces renal ischemia/reperfusion injury in the rat

BACKGROUND: Activation of the cysteine protease calpain has been implicated in renal ischemia/reperfusion (I/R) injury. The aim of this study was to investigate the effects of calpain inhibitor-1 (Cal I-1) in an in vivo model of renal I/R injury. METHODS: Male Wistar rats were administered Cal I-1 (10 mg/kg, IP) 30 minutes before undergoing bilateral renal ischemia (45 minutes) followed by reperfusion (6 hours). Plasma concentrations of urea, creatinine, Na(+), gamma-glutamyl transferase (gamma GT), aspartate aminotransferase (AST) and urinary Na(+), glutathione S-transferase (GST), and N-acetyl-beta-D-glucosaminidase (NAG) were measured for the assessment of renal dysfunction and I/R injury. Creatinine clearance (C(Cr)) and fractional excretion of Na(+) (FE(Na)) were used as indicators of glomerular and tubular function, respectively. Kidney myeloperoxidase (MPO) activity and malondialdehyde (MDA) levels were measured for assessment of neutrophil infiltration and lipid peroxidation, respectively. Renal sections were used for histologic grading of renal injury and for immunohistochemical localization of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). RESULTS: Cal I-1 significantly reduced I/R-mediated increases in urea, creatinine, gamma GT, AST, NAG, and FE(Na) and significantly improved C(Cr). Cal I-1 also significantly reduced kidney MPO activity and MDA levels. Cal I-1 also reduced histologic evidence of I/R-mediated renal damage and caused a substantial reduction in the expression of iNOS and COX-2, both of which involve activation of nuclear factor-kappa B (NF-kappa B). CONCLUSIONS:: These results suggest that Cal I-1 reduces the renal dysfunction and injury associated with I/R of the kidney. We suggest that the mechanism could involve the inhibition of I/R-mediated activation of NF-kappa B.     

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.     

Attenuation of ischemia/reperfusion injury by N-acetylcysteine in a rat hind limb model

BACKGROUND: Ischemia/reperfusion is a complex set of events with severe pathologic consequences. Reperfusion initiates both the local and systemic damage in part through rapid oxygen generation. N-acetylcysteine (NAC) is a scavenger of free radical species, inhibits neutrophil accumulation, acts as a vasodilator and also improves microcirculation. In present study, we examined the protective effect of NAC in a rat hind limb ischemia/ reperfusion model. Dimethyl-sulfoxide (DMSO), a well-known antioxidant was also tested for comparison. MATERIALS AND METHODS: Ischemia was induced for 4 h by vascular clamping and followed by 1 h of reperfusion. Muscle injury was evaluated in 3 groups as a saline group (control), DMSO group, and NAC group. Plasma levels of creatine kinase, lactate dehydrogenase, thiobarbituric acid reactive substances (TBARS), and blood HCO(3), as well as muscle tissue TBARS, were measured at the end of reperfusion. Muscle tissue samples were taken for histological evaluation. RESULTS: DMSO and NAC group showed significant amelioration of plasma CPK (P < 0.05, P < 0.05), plasma TBARS (P < 0.05, P < 0.05), and muscle tissue TBARS (P < 0.05, P < 0.05) compared with the control group. Similarly, neutrophil infiltration in DMSO and NAC groups were significantly less prominent than the control group (P < 0.01, P < 0.01). CONCLUSIONS: These results show that NAC improved effectively ischemia reperfusion injury in a rat hind limb model.