Change and role of heme oxygenase-1 in injured lungs following limb ischemia／reperfusion in rats

Lowerlimbischemiafollowedbyreperfusionisanimportantandcommonclinicalevent .Bothclinicalobservationandanimalexperimentindicatethatrestorationofbloodflowcansavethelimbsbutresultsinmultisystemorgandysfunctionevendeath .1Althoughthesystemicinflammationoflimbischemia/reperfusion (I/R )candamageanyorgan ,theonsetofthesyndromeisusuallyheraldedbythedevelopmentof pulmonarydysfunction .2 ,3Thiskindofpulmonarydysfunctionischaracterizedbyincreasedlungvascular permeabilityandpulmonaryhypertension ,whichis…     

Cytokines contribute to early hepatic parenchymal injury and microvascular dysfunction after bilateral hindlimb ischemia.

PURPOSE: Hepatic dysfunction may contribute to death from multiple organ dysfunction after abdominal aortic surgery. Several factors are likely responsible, and the purpose of this study was to determine whether the cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin 1 (IL-1) are involved in initiating this remote hepatic injury. METHODS: In a normotensive rat model of 4-hour bilateral hindlimb ischemia/reperfusion (I/R), we measured systemic TNF-alpha and IL-1 levels throughout the I/R period. Rats were randomly assigned to either the 3-hour control group, the 3-hour I/R group, or the I/R group with administration of a polyclonal antibody (PAb) to TNF-alpha (I/R + TNF-alpha PAb). Direct evidence of lethal hepatocyte injury through the labeling of nuclei by propidium iodide (per 10(-1)mm(3)) and altered microvascular perfusion were assessed by using intravital microscopy. RESULTS: Systemic TNF-alpha peaked at 83.97 pg/mL (P <.05, n = 5) at 30 minutes of reperfusion and returned to baseline in 60 to 90 minutes. No significant change in systemic IL-1 was detected (P <.05, n = 4). Alanine aminotransferase increased 2.5-fold in the I/R group through 3 hours of reperfusion (P <.05, n = 4), and TNF-alpha PAb did not attenuate this alanine aminotransferase increase (P <.05, n = 6). Lethal hepatocyte injury increased by 8-fold in the I/R group compared with the control group (P <.05, n = 5), whereas TNF-alpha PAb significantly reduced this injury (P <.05, n = 4). No regional differences in injury were noted within the acinus. Total perfusion within the microvascular unit did not drop; however, significant flow heterogeneity was observed. The proportion of continuously perfused sinusoids declined in the I/R group after 3 hours of reperfusion in both periportal (62.0 +/- 2.2, P <.05) and, to a lesser, although significant, degree, in the pericentral regions (73. 2 +/- 1.73, P <.05). CONCLUSION: By scavenging extracellular TNF-alpha with a PAb, we provide direct evidence that TNF-alpha contributes to, but is not solely responsible for, early remote hepatocellular injury and microvascular dysfunction. The administration of TNF-alpha PAb reduced lethal hepatocyte injury in both regions of the acinus and also improved perfusion in the periportal region (76.8 +/- 5.41, P <.05), but not in the pericentral region. This suggests that TNF-alpha released during reperfusion mediates early remote hepatocellular injury and microvascular dysfunction after a remote ischemic insult.     

Decreased microvascular perfusion in the rabbit ear after six hours of ischemia

The cellular injury produced by reperfusion of ischemic tissue with oxygen-rich blood has been studied in numerous tissues but has not been investigated extensively in thermoregulatory tissue. This study was designed (a) to compare 4 and 6 hours of ischemia to document the evidence of impaired capillary perfusion after resumption of blood flow (reperfusion injury) in a thermoregulatory end organ (the rabbit ear), and (b) to examine, with use of vital capillaroscopy (VC) and laser Doppler flowmetry (LDF), the altered microvascular blood flow in the rabbit ear after ischemia and reperfusion. One ear from each of five rabbits underwent warm ischemia for 4 hours. VC showed no deficits of capillary perfusion in these ears after reperfusion; LDF measurements in both ears also demonstrated no significant difference between control and reperfusion blood flow. One ear from each of eight additional rabbits underwent 6 hours of warm ischemia. LDF values were significantly reduced in the ischemic ear after reperfusion as compared with baseline measurements for that ear and as compared with the control ear. VC showed arrested perfusion and static plasma gaps within three to five capillaries per high-power field (an area of 300 × 500 μm) in the ischemic ear and good perfusion of all vessels in the contralateral control ear. This evidence of reperfusion injury in a thermoregulatory end organ may help to explain the poor functional result that often occurs after replantation of an amputated digit.     

大鼠骨骼肌缺血再灌注时血红素氧合酶-1的表达

目的：观察大鼠缺血再灌注时骨骼肌中血红素氧合酶－1（HO－1）表达的变化，方法：夹闭大鼠股动脉造成下肢缺血模型，分别采集假手术（S）组，单纯缺血（I）4小时组及缺血4小时再灌注（R）2、4、8、16和24小时组的比目鱼肌，检测其组织学和丙二醛（MDA）含量变化，通过Northern印迹、Western印迹及免疫组织化学分析，观察HO－1表达的变化。结果：S组和I组未见HO－1mRNA表达；R2组可见其表达信号，且随着再灌注时间延长表达信号逐渐增强，到R8组时达到高峰，之后渐减弱，R24组时已消失。蛋白表达的变化与mRNA变化基本一致，免疫组织化学显示，R组HO－1阳性信号主要出现在骨骼肌细胞浆内，S组和I组均未见阳性信号。与S组相比，R各组MDA含量显著增高（P＜0．05），但R8组较R4组显著减低（P＜0．05）。结论：肢缺血再灌注可诱发骨骼肌HO－1的表达，其表达可能具有保护作用。     

Inhalation of carbon monoxide reduces skeletal muscle injury after hind limb ischemia-reperfusion injury in mice

BackgroundThe purpose of this study was to determine if inhaled carbon monoxide (CO) can ameliorate skeletal muscle injury, modulate endogenous heme oxygenase-1 expression, and improve indexes of tissue integrity and inflammation after hind limb ischemia reperfusion.MethodsC57BL6 mice inhaling CO (250 ppm) or room air were subjected to 1.5 hours of ischemia followed by limb reperfusion for either 3 or 6 hours (total treatment time, 4.5 or 7.5 h). After the initial period of reperfusion, all mice breathed only room air until 24 hours after the onset of ischemia. Mice were killed at either the end of CO treatment or at 24 hours' reperfusion. Skeletal muscle was subjected to histologic and biochemical analysis.ResultsCO treatment for 7.5 hours protected skeletal muscle from histologic and structural evidence of skeletal muscle injury. Serum and tissue cytokines were reduced significantly (P < .05) in mice treated with CO for 7.5 hours. Tubulin, heme oxygenase, and adenosine triphosphate levels were higher in CO-treated mice.ConclusionsInhaled CO protected muscle from structural injury and energy depletion after ischemia reperfusion.     

肝糖原贮备对热缺血再灌注大鼠肝细胞凋亡的影响

目的探讨肝糖原贮备对热缺血再灌注肝细胞凋亡及肝损伤的影响。方法建立大鼠肝热缺血模型。实验分组:术前24h静脉注射25%葡萄糖组,2ml/只,每6h1次,高糖饮食(H组);术前禁食24h,饮水不限(L组);正常饮食对照组(N组)和假手术组(S组)。缺血45min,再灌注2、24h取材。流式细胞术检测细胞凋亡及Bcl-2、Bax蛋白。同时进行肝酶学检测、肝组织形态学观察。结果1.细胞凋亡及蛋白表达:(1)24h细胞凋亡百分率。S组N组(P<0.01)。2.肝酶学指标:各时相点4组间比较差异有统计学意义(P<0.05),S组相似文献   

肝细胞凋亡在肝硬化大鼠肝缺血再灌注损伤中的意义

目的 研究肝硬化大鼠肝缺血再灌注（I／R）损伤和硬化肝比正常肝更容易损伤的机制是否与肝细胞凋亡有关？方法 建立原位肝I／R模型，将肝硬化大鼠随机分为2组：A组：缺血时间（I）＝20min;B组：I＝30min;C组：正常大鼠，I＝30min，比较灌注前后各组血清AST、ALT的变化和肝细胞凋亡的百分数。结果 肝硬化大鼠肝I／R后，AST、ALT明显升高，以灌注后6h为高峰，灌注24、72h后逐渐下降。灌注6h后，B组的血清转氨酶为3组中最高（P＜0．05），说明B组肝损伤最严重。肝细胞凋亡在I／R后明显增多，以灌注后6h为高峰，随后逐渐下降，变化与转氨酶一致。灌注后6h，B、A、C组肝细胞凋亡的百分数分别为20．9％、13．5％和10．7％，B组明显高于A、C两组（P＜0．01）。再灌注72h内未见明显肝细胞坏死。结论 肝细胞凋亡是肝硬化大鼠I／R损伤肝细胞死亡的主要形式，肝细胞凋亡与肝缺血时间密切相关，肝硬化肝细胞比正常肝细胞容易发生凋亡是硬化肝对缺血敏感的重要原因。     

Preconditioning with tin-protoporphyrin IX attenuates ischemia/reperfusion injury in the rat kidney

BACKGROUND: Heme oxygenase (HO)-1 is induced as a unique stress response and leads to a transient resistance against oxidative damage, including ischemia and reperfusion (I/R) injury. In the present study, we examined whether HO-1 induction may confer a protection against I/R injury in the rat kidney. METHODS: Lewis rats were divided into four groups as follows: (1) vehicle group; (2) group treated with ferri-protoporphyrin IX (hemin), an inducer of HO; (3) group treated with low-dose tin-protoporphyrin IX (SnPP), an inhibitor of HO; and (4) group treated with high-dose SnPP. Renal warm ischemia for 60 minutes was performed 24 hours after each treatment. RESULTS: At 24 hours after treatment, hemin induced a significant increase in renal HO activity, but failed to induce HO-1 protein synthesis. Although both low- and high-dose SnPP reduced HO activity, a marked HO-1 expression was observed only in the high-dose SnPP-treated kidney. Hemin exacerbated the renal function after reperfusion, while high-dose SnPP significantly suppressed the intercellular adhesion molecule (ICAM)-1 expression, the infiltration of ED-1-positive macrophages and the expression of activated caspase-3, which resulted in attenuation of apoptotic cell death and ameliorated I/R injury. CONCLUSION: These results suggest that prior induction of HO-1 protein by high-dose SnPP may lead to anti-inflammatory and antiapoptotic effects on warm renal I/R injury independently of its enzyme activity, and that HO enzyme activation may not always act as an antioxidant, especially under I/R-induced oxidative stress.     

VEGF mitigates histone‐induced pyroptosis in the remote liver injury associated with renal allograft ischemia–reperfusion injury in rats

Clinical evidence has indicated a possible link between renal injury and remote liver injury. We investigated whether extracellular histone mediates remote hepatic damage after renal graft ischemia–reperfusion injury, while vascular endothelial growth factor (VEGF) is protective against remote hepatic injury. In vitro, hepatocyte HepG2 cultures were treated with histone. In vivo, the Brown‐Norway renal graft was stored in 4°C preservation solution for 24 hours and then transplanted into a Lewis rat recipient; blood samples and livers from recipients were harvested 24 hours after surgery. Prolonged cold ischemia in renal grafts enhanced liver injury 24 hours after engraftment. Caspase‐1, ASC, NLRP3, and AIM2 expressions in hepatocyte, CD68⁺‐infiltrating macrophages, tissue, and serum interleukin‐1β and ‐18 were greatly elevated, indicating that pyroptosis occurred in the liver and resulted in acute liver functional impairment. Blocking the caspase‐1 pathway decreased the number of necrotic hepatocytes. VEGF treatment suppressed the hepatocyte pyroptosis and liver function was partially restored. Our data suggested that renal allograft ischemia–reperfusion injury is likely associated with acute liver damage due to hepatocyte pyroptosis induced by histone and such injury may be protected by VEGF administration. VEGF, therefore, may serve as a new strategy against other remote organ injuries related to renal transplantation.     

Heme oxygenase-1 induction by the clinically used anesthetic isoflurane protects rat livers from ischemia/reperfusion injury

OBJECTIVE: It was the aim of this study to characterize the influence of isoflurane-induced heme oxygenase-1 (HO-1) expression on hepatocellular integrity after ischemia and reperfusion. SUMMARY BACKGROUND DATA: Abundant experimental data characterize HO-1 as one of the most powerful inducible enzymes that contribute to the protection of the liver and other organs after harmful stimuli. Therapeutic strategies aimed at utilizing the protective effects of HO-1 are hampered by the fact that most pharmacological inducers of this enzyme perturb organ function by themselves and are not available for use in patients because of their toxicity and undesirable or unknown side effects. METHODS: Rats were pretreated with isoflurane before induction of partial hepatic ischemia (1 hour) and reperfusion (1 hour). At the end of each experiment, blood and liver tissue were obtained for molecular biologic, histologic, and immunohistochemical analyses. RESULTS: Isoflurane pretreatment increased hepatic HO-1 mRNA, HO-1 protein, HO enzyme activity, and decreased plasma levels of AST, ALT, and alpha-GST. Histologic analysis of livers obtained from isoflurane-pretreated rats showed a reduction of necrotic areas, particularly in the perivenular region, the predominant site of isoflurane-induced HO-1 expression. In addition, sinusoidal congestion that could otherwise be observed after ischemia/reperfusion was inhibited by the anesthetic. Furthermore, isoflurane augmented hepatic microvascular blood flow and lowered the malondialdehyde content within the liver compared with control animals. Administration of tin protoporphyrin IX inhibited HO activity and abolished the isoflurane-induced protective effects. CONCLUSIONS: This study provides first evidence that pretreatment with the nontoxic and clinically approved anesthetic isoflurane induces hepatic HO-1 expression, and thereby protects rat livers from ischemia/reperfusion injury.     

The effect of mannitol versus dimethyl thiourea at attenuating ischemia/reperfusion-induced injury to skeletal muscle

OBJECTIVE: Mannitol is used as a treatment for skeletal muscle ischemia/reperfusion (I/R) injury in humans, despite the fact that its effectiveness in vivo is still disputed. The purpose of this study was to determine the efficacy of mannitol in attenuating I/R injury at the microcirculatory level. METHODS: The study was designed as an experimental study with male Wistar rats. The main outcome measures were intravital microscopy, which was used to measure capillary perfusion, capillary and venular red blood cell velocity (VRBC), and leukocyte-endothelial interactions in the extensor digitorum longus muscle of the rat hind limb before and after ischemia. In addition, tissue injury was assessed during reperfusion with the fluorescent vital dyes bisbenzimide and ethidium bromide. Dimethyl thiourea (DMTU), a highly effective therapeutic agent of experimental I/R injury, was used as a positive control. RESULTS: No-flow ischemia (2 hour) resulted in a 40% drop in capillary perfusion, a decline in capillary and venular VRBC, and increased leukocyte venular adherence and tissue infiltration. Tissue injury increased to a constant level during reperfusion. Mannitol attenuated capillary malperfusion during the first 60 minutes of reperfusion and prevented a decline in capillary VRBC. However, mannitol did not reduce tissue injury or leukocyte adherence and infiltration during reperfusion. By comparison, DMTU not only prevented the perfusion deficits and the increases in leukocyte venular adherence and tissue infiltration but significantly reduced the magnitude of tissue injury. CONCLUSION: Our findings suggest that mannitol may be of limited value for the prevention of early reperfusion-induced injury after no-flow ischemia in skeletal muscle. By comparison, DMTU was highly efficacious by not only reducing microvascular perfusion deficits but by also reducing leukocyte-endothelial cell interactions and the incidence of cellular injury.     

A noninvasive murine model of hind limb ischemia-reperfusion injury

BACKGROUND: This study describes a novel murine method of the Controlled Tension Tourniquet (CTT). The CTT applies a measured circumferential tension to hind limbs using a tourniquet attached to digital strain gauges, and is useful for investigating hind limb ischemia reperfusion (IR). MATERIALS AND METHODS: Mice were subjected to 1, 3, or 6 h of unilateral hind limb ischemia followed by either 4 or 24 h of reperfusion. Blood flow in the ischemic, reperfused, and contralateral limbs was monitored using a Laser Doppler Imager. Edema in the IR limbs was documented by changes in the wet weight to dry weight ratio. Myeloperoxidase and tetrazolium based mitochondrial activity assays indicated neutrophil infiltration and tissue viability, respectively. RESULTS: During reperfusion following 1, 4, or 6 h, flow stabilized at 100%, 53%, and 23% of baseline levels, respectively. Edema was present all in IR limbs after 4 h of reperfusion, but increased with the duration of ischemia. After 24 h of reperfusion neutrophil infiltration was equivalent in all IR limbs after all intervals of ischemia. After 24 h of reperfusion, tissue viability after 1 h of ischemia was equivalent to sham or contralateral limbs. At 3 or 6 h of ischemia and 24 h reperfusion decreased tissue viability to 40% of sham and contralateral limbs. CONCLUSIONS: The CTT provides a reproducible, noninvasive model of acute limb ischemia, which reflects the biochemical indices of microvascular injury, inflammation and flow characteristic of reperfusion injury.     

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.     

三种灌注液对断指再植组织再灌注损伤保护作用的研究

目的 研究三种不同灌注液对断肢再植后缺血再灌注损伤的保护作用。方法 建立大鼠后肢的断肢再植模型，应用3种不同灌注液（A组肝素钠组，B组肝素钠＋利多卡因组，C组肝素钠＋地塞米松组）对离断肢体进行灌注，然后进行再植。分别于缺血前、缺血6h、通血60min取材，测定断肢皮肤中丙二醛（MDA）、透射电镜观察缺血6h后再通血60min血管内膜的改变。结果 3组再植肢体成活率相当，B组灌注前后MDA差值明显低于A组，差异有统计学意义。B组、C组血管内膜改变较A组轻微。结论 断肢再植前应用含利多卡因、地塞米松灌注液对离断指进行灌注，可以减轻再植肢体的缺血再灌注损伤，且对血管内膜有保护作用。     

Evidence for reperfusion injury in cortical bone as a function of crush injury ischemia duration: a rabbit bone chamber study

A model for critical limb ischemia was produced by occluding femoral vessels in 24 rabbits with a pneumatic cuff for 0, 2, 4, or 6 hours. Immediate sequelae and subsequent creeping substitution of cortical bone were observed in vivo using an implanted tibial window, the optical bone chamber implant (with intravital microscopy), and then by light and fluorescence microscopy of fluorochrome-labeled and surface-stained ground sections of retrieved implants. Six rabbits were used as controls (0 h) for each ischemia treatment, and the animals were monitored for 5 weeks postocclusion. A subpopulation of 13 implants was retrieved after euthanization and then histologically assessed for bone necrosis and remodeling. The hypothesis tested was that reperfusion injury during the 24 h after occluder release (reperfusion phase), and vessel perfusion/caliber, angiogenesis, and net bone resorption during the 5 subsequent weeks (creeping substitution phase), would exhibit ischemia duration-dependent effects. All animals could bear weight on the affected limb to ambulate by 1 week posttreatment. Two-way analysis of variance (ANOVA) comparison of the resulting data confirmed a significant difference between control and ischemia-treated rabbits for: (1) vessel perfusion/reperfusion; (2) vessel caliber; and (3) net bone resorption. Vascular responses to 4 vs. 6 h of ischemia were not significantly different, but net bone resorption was strictly ischemia duration-dependent. The conclusion that reperfusion injury was the mechanism spreading ischemia to more vessels was supported by a decrease in reperfusion and caliber of vessels, and an increase in vascular permeability and leukocyte adherence during the reperfusion phase. It is postulated that reperfusion injury produces a secondary ischemia that amplifies the occlusion-created primary ischemia and, in the present work, may have been succeeded by progressive episodes of ischemia, similar to the infarction pattern of ischemic hearts.     

Liver viability after ischemia-reperfusion

Lack of a reproducible model to quantitatively assess hepatocellular injury following ischemia has made it difficult to assess new strategies for minimizing hepatic injury. We studied the progression of hepatocellular injury after ischemia and ischemia with reperfusion in rats. Irreversible injury was quantitated using a triphenyltetrazolium chloride assay that was shown to correlate with ultrastructural changes. Adenosine triphosphate decreased to 36% of basal values after 30 minutes, but returned to normal with reperfusion with no decrease in viability. In contrast, viability fell by 30% after 60 minutes of ischemia, and by 64% when 60 minutes of ischemia was followed by reperfusion. We conclude that reperfusion of ischemic liver increases the degree of irreversible damage. The model employed here seems to be useful for studying ischemic and reperfusion injury in the liver.     

Acute lung injury following reperfusion after ischemia in the hind limbs of rats.

In this study, we proposed that oxygen free radicals participate in the acute pulmonary injury that follows limb ischemia/reperfusion. Using an established model of hind limb ischemia, reproducible lung injury occurred after reperfusion. Lung microvascular permeability was measured with 125I-BSA and increased two-fold after 30 minutes of reperfusion. Pulmonary injury was blocked with DMSO, DMTU, allopurinol, indomethacin, and SOD plus catalase. The degree of pulmonary neutrophil sequestration as assessed by tissue myeloperoxidase activity was significantly diminished in animals pretreated with antioxidants. Pretreatment with indomethacin did not attenuate the neutrophil sequestration within the pulmonary parenchyma. These data suggest that increased lung microvascular permeability and neutrophil accumulation occur following hind limb ischemia/reperfusion. Therapeutic interventions with oxygen radical inhibitors blocked this process, while the prostaglandin inhibitor, indomethacin, only reduced lung permeability.     

应用256层螺旋CT灌注技术研究兔肝I/R后血流动力学变化

目的：应用256层螺旋CT灌注成像分析兔部分肝脏缺血再灌注损伤（I/R）后阶段血流动力学的变化规律及其价值。
方法：新西兰大白兔阻断肝左叶血供60min后,恢复血供,按缺血再灌注的时间分为6,12,24 h和假手术（S）组,每组6只。各组分别采用256层螺旋CT行全肝灌注成像和病理学分析。在灌注图上测量肝动脉灌注量（HAP）,门静脉灌注量（HPP）,总灌注量（TLP）以及肝动脉灌注指数（HPI）。
结果：(1)在I/R 6,12,24 h组肝脏血流出现差异性分布（低灌注的肝组织,即梗死区）。(2)在梗死的肝组织中,与S组相比,I/R 6 h 组中HPP,TLP外,I/R12 h和24 h组HAP,HPP,TLP均低于对照组,而HPI则高于对照组.在灌注相对正常的肝组织中各灌注参数均呈下降趋势。(3)I/R组梗死与未梗死肝组织CT灌注参数之间差异有显著性。
结论：CT灌注参数能够客观准确地反映肝I/R病理过程中血流动力学的变化。     

In vivo microscopy of microcirculatory injury in skeletal muscle following ischemia/reperfusion

While the sequence of biochemical and cellular events in the pathogenesis of ischemia/reperfusion injury is increasingly well understood, the way that these processes interact at the level of microcirculation to promote a distinctive reperfusion injury is less well defined. It is becoming clear, however, that these processes are initiated at the level of microcirculation, and that microcirculatory damage may precede actual tissue injury. Such damage causes microvascular no-reflow, which in turn effectively prolongs the time of tissue ischemia and extends tissue injury. Recently, microcirculatory models have been adapted for study of the microvascular effects of ischemia/reperfusion. We have used a new in vivo mouse cremaster muscle model to study, by direct and quantitative measurement, the acute microvascular changes involved in ischemia/reperfusion. Previously described changes in capillary perfusion and venular leukocyte adhesion were observed in this model following reperfusion after prolonged ischemia (4–6 hours). We have further characterized an intense reactive vasoconstriction or vasospasm that occurs after prolonged ischemia; this vessel reaction may represent an important overlooked cause of no-reflow following ischemia/reperfusion. This article summarizes our work in the context of other available methods that have been used to define the microvascular changes of ischemia/reperfusion. © 1994 Wiley-Liss, Inc.     

Influence of ischemic preconditioning and nitric oxide on microcirculation and the degree of rat liver injury in the model of ischemia and reperfusion

The aim of this study was to assess the influence of ischemic preconditioning (IPC) on parenchymal liver blood flow during the early phase of reperfusion after 60 minutes of ischemia, additionally modified by adding N-nitro-L-arginine methyl ester (L-NAME). Our research involved 4 groups of rats (10 animals in each group), which underwent liver ischemia and 24 hours of reperfusion. Group I, ischemia/reperfusion (IR) was performed; group II, IPC, 10 minutes of ischemia and 10 minutes of reperfusion, and IR after that; group III, L-NAME (10 mg/kg intravenous [iv]), 10 minutes before IR; and group IV, L-NAME before IPC + IR. Activity of APAT, ALAT, GGTP, and FA was marked in serum in 90 minutes and 24 hours of reperfusion. In the liver biopsies at 24 hours of reperfusion, we analyzed reaction on adenosine-3-phosphatase stimulated by Mg++ and performed histological examination. The parenchymal perfusion was measured using a laser-doppler blood flowmeter (model PeriFlux System5000, Perimed Inc., United Kingdom). IPC during reperfusion led to minor injuries of the organ, with statistically significant normalization of enzymes compared with group 1, and a better reaction to the adenosine-3-phosphatase IPC produced faster and full return of perfusion to the 68.3 value at 24 hours (59.1 in the 60 minutes). In groups III and IV at 60 minutes, the perfusion was not statistically different from that in group 1. IPC causes full and faster blood return in the early phase of reperfusion and minor injury of liver parenchyma and liver sinus. The protective effect observed, especially in the first 60 minutes of reperfusion, was limited by L-NAME and was influenced by the action of nitric oxide.