Amelioration of liver injury by ischaemic preconditioning

BACKGROUND: Ischaemic preconditioning, i.e. preparatory brief ischaemia before subsequent long ischaemia, can effectively protect the heart from ischaemia-reperfusion injury in animals. The purpose of this study was to demonstrate the same phenomenon in the liver. METHODS: Using warm ischaemia-reperfusion of 70 per cent of the liver followed by resection of the non-ischaemic portion in rats, livers with 10 min of ischaemic preconditioning, i.e. 10 min of warm ischaemia and reperfusion, were compared with those that had not been subjected to such a manoeuvre. RESULTS: At 120 min after reperfusion following 40 min of warm ischaemia, the livers with 10 min of ischaemic preconditioning had a significantly lower mean(s.d.) serum alanine aminotransferase level (492(217) versus 1236(695) units/l; P < 0005) and lactic dehydrogenase level (7905(4002) versus 15066(9201) units/l; P< 0.05), as well as a higher bile output (0.12(0.03) versus 0.09(0.04) ml per g liver; P < 0.05) and liver tissue adenosine 5'-triphosphate level (78(13) versus 61(11) per cent; P< 0.05) than the control livers. The necrosis rate, histologically defined as the percentage of necrotic area in given liver sections, was reduced significantly by this manoeuvre (mean(s.d.) 1.3(1.3) versus 5.3(1.7) per cent; P< 0.05). CONCLUSION: Ischaemic preconditioning exerts a protective effect on hepatic warm ischaemia-reperfusion injury. Such a manoeuvre may be useful for hepatic resection in the clinical setting.     

Effects of inhibitors of the activity of poly (ADP-ribose) synthetase on the liver injury caused by ischaemia-reperfusion: a comparison with radical scavengers

1. Poly (ADP-ribose) synthetase (PARS) is a nuclear enzyme activated by strand breaks in DNA which are caused by reactive oxygen species (ROS) and peroxynitrite. Excessive activation of PARS may contribute to the hepatocyte injury caused by ROS in vitro and inhibitors of PARS activity reduce the degree of reperfusion injury of the heart, skeletal muscle and brain in vivo. Here we compared the effects of various inhibitors of the activity of PARS with those of deferoxamine (an iron chelator which prevents the generation of hydroxyl radicals) and tiron (an intracellular scavenger of superoxide anion) on the degree of hepatic injury caused by ischaemia and reperfusion of the liver in the anaesthetized rat or rabbit. 2. In the rat, ischaemia (30 or 60 min) and reperfusion (120 min) of the liver resulted in significant increases in the serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) indicating the development of liver injury. Intravenous administration of the PARS inhibitors 3-aminobenzamide (3-AB, 10 mg kg(-1) or 30 mg kg(-1)), 1,5-dihydroxyisoquinoline (ISO, 1 mg kg(-1)) or 4-amino-1,8-naphthalimide (4-AN, 3 mg kg(-1)) before reperfusion did not reduce the degree of liver injury caused by ischaemia-reperfusion. 3. In contrast to the PARS inhibitors, deferoxamine (40 mg kg(-1)) or tiron (300 mg kg(-1)) significantly attenuated the rise in the serum levels of AST and ALT caused by ischaemia-reperfusion of the liver of the rat. 4. In the rabbit, the degree of liver injury caused by ischaemia (60 min) and reperfusion (120 min) was also not affected by 3-AB (10 mg kg(-1)) or ISO (1 mg kg(-1)). 5. These results support the view that the generation of oxygen-derived free radicals mediates the liver injury associated with reperfusion of the ischaemic liver by mechanism(s) which are independent of the activation of PARS.     

A comparison of the efficacy and tolerability of dorzolamide and acetazolamide as adjunctive therapy to timolol. Oral to Topical CAI Study Group

BACKGROUND: Recent observations provide evidence that complement is involved in the pathophysiology of ischemia/reperfusion injury. In this study, we assessed the impact of complement inhibition on hepatic microcirculation and graft function using a rat model of liver transplantation. METHODS: Arterialized orthotopic liver transplantation was performed in Lewis rats after cold preservation (University of Wisconsin solution, 4 degrees C, 24 h). Eight animals received the physiological complement regulator soluble complement receptor type 1 (sCR1) intravenously 1 min before reperfusion. Controls received Ringer's solution (n=8). Microvascular perfusion, leukocyte adhesion, and Kupffer cell phagocytic activity were studied 30-100 min after reperfusion by in vivo microscopy. RESULTS: Microvascular perfusion in hepatic sinusoids was improved in the sCR1 group (87+/-0.7% vs. 50+/-1%; P < 0.001). The number of adherent leukocytes was reduced in sinusoids (68.3+/-4.7 vs. 334.1+/-15.8 [adherent leukocytes per mm < or = liver surface]; P < 0.001) and in postsinusoidal venules after sCR1 treatment (306.6+/-21.8 vs. 931.6+/-55.9 [adherent leukocytes per mm < or = endothelial surface]; P < 0.001). Kupffer cell phagocytic activity was decreased in the sCR1 group compared to controls. Postischemic bile production reflecting hepatocellular function was increased by almost 200% (P = 0.004) after complement inhibition. Plasmatic liver enzyme activity was decreased significantly upon sCR1 treatment, indicating reduced parenchymal cell injury. CONCLUSIONS: Our results provide further evidence that the complement system plays a decisive role in hepatic ischemia/reperfusion injury. We conclude that complement inhibition by sCR1 represents an effective treatment to prevent reperfusion injury in liver transplantation.     

Dibutyryl cAMP reduces nonparenchymal cell damage during cold preservation of rat livers

Sinusoidal lining cells are the main target for cold preservation injury and are further damaged with reperfusion. Different agents known to increase intracellular cAMP levels have been shown beneficial. This study was designed to assess the possible protective effect of a cAMP analogue on nonparenchymal cells of rat livers, during cold storage and during reperfusion. Parameters reflecting the status of the liver microvasculature were analyzed. The initial effluent collected after preservation reflects release during the cold storage period; therefore we measured interleukin-1 (IL-1) and endothelin-1 (ET-1) levels in these samples in order to detect and quantitate the degree of activation and/or disruption of Kupffer and sinusoidal endothelial lining cells. Rat livers were harvested after in situ flush with Ringer's lactate with or without 2 mM dibutyryl cAMP, excised, and stored in the same solution at 4 degrees C. After 6 hr, livers were perfused with Krebs-Henseleit buffer for 90 min. Physiological parameters were monitored throughout the perfusion. Perfusate samples were collected every 30 min for RIA measurements of IL-1 and ET-1. Treatment resulted in a significant decrease in release of ET-1 and IL-1 during storage. Likewise, livers treated with cAMP had a significantly improved bile output and decreased portal vein resistance during reperfusion. The beneficial effect granted by the analogue during cold storage and reperfusion was evident on parenchymal and nonparenchymal cells. Levels of ET-1 and IL-1 in the caval effluent confirm and quantitate preservation damage.     

A clinical experience with a hierarchically controlled myoelectric hand prosthesis with vibro-tactile feedback

The decline of plasma fibronectin after surgery, trauma, and burn, as well as during severe sepsis after injury, appears to limit hepatic Kupffer cell phagocytic activity. Intravenous infusion of gelatin-coated particles to simulate blood-borne particulate collagenous tissue debris in the circulation after injury also depletes plasma fibronectin. We used soluble gelatin conjugated with 125I-labeled dilactitol tyramine (DLT-gelatin) as a model of soluble collagenous tissue debris. We studied its blood clearance as well as organ localization in normal and postburn rats. Fibronectin-deficient plasma harvested early after burn exhibited limited ability to support in vitro phagocytic uptake of the gelatinized microparticles by Kupffer cells in liver tissue from normal rats. However, Kupffer cells in liver tissue from normal and postburn rats phagocytized the test particles at a normal rate when incubated in normal plasma. The DLT-gelatin ligand bound to fibronectin in a dose-dependent manner as verified by its capture with anti-fibronectin coated plastic wells when coincubated with purified fibronectin. By gel filtration chromatography, the binding of fibronectin with the DLT-gelatin ligand was readily detected, resulting in the formation of a high-molecular-weight complex. In normal animals the plasma clearance and liver localization of 125I-DLT-gelatin was competitively inhibited by infusion of excess nonradioactive gelatin. The blood clearance and liver localization of the soluble gelatin ligand were also impaired after burn injury during periods of fibronectin deficiency similarly to the pattern observed with gelatin-coated microparticles. By autoradiography, the cellular site for the uptake of the 125I-DLT-gelatin was primarily but not exclusively hepatic Kupffer cells; 125I-DLT-asialofetuin and 125I-DLT-ovalbumin were removed by hepatocytes and sinusoidal endothelial cells, respectively. Thus, gelatin conjugated with 125I-DLT can be used to simulate blood-borne soluble collagenous tissue debris after burn. It rapidly binds to plasma fibronectin before its hepatic Kupffer cell removal, and its blood clearance is markedly delayed after burn injury during periods of plasma fibronectin deficiency.     

Glutathione defense in non-parenchymal cells

Toxicity to nonparenchymal cells can result in disruption of the hepatic microcirculation, altered production of cytokines, and hepatic fibrosis. Many of the relevant insults produce oxidative stress or toxic metabolites that require glutathione detoxification. This article reviews the role of sinusoidal endothelial cell glutathione (GSH) in reperfusion injury, cytomegalovirus infection, and hepatic venoocclusive disease. The effects of oxidative stress and antioxidants on Kupffer cell production of cytokines and, in particular the potential benefit of antioxidants in the setting of reperfusion injury, are discussed. Oxidative stress upregulates collagen gene expression by stellate cells, and this is modulated by antioxidants. Current thinking on intrahepatic GSH and cysteine homeostasis is discussed. Finally, I review the published data on nonparenchymal GSH levels, glutathione S-transferase activity and isoenzyme pattern, and glutathione peroxidase activity.     

Repeatability of the Petrifilm HEC test and agreement with a hydrophobic grid membrane filtration method for the enumeration of Escherichia coli O157:H7 on beef carcasses

The review highlights recent advances in our understanding of basic mechanisms of reperfusion injury after warm hepatic ischemia. Kupffer cells play a central role as the initial cytotoxic cell type and as a source of many proinflammatory mediators. Subsequently, neutrophils are activated and recruited into the liver. Factors and conditions are outlined that determine whether neutrophils undergo apoptosis without causing damage or migrate out of the sinusoids and attack parenchymal cells. In addition to the inevitable inflammatory response during reperfusion, microcirculatory perfusion failure, due to an imbalance between the actions of vasodilators and vasoconstrictors, also has a serious impact on reperfusion injury. A better understanding of the basic pathophysiology will reveal potential targets for therapeutic interventions and will show us how to avoid risk factors that may aggravate reperfusion injury.     

Is allopurinol beneficial in the prevention of renal ischaemia-reperfusion injury in the rat?: evaluation by near-infrared spectroscopy

1. The role of allopurinol in the protection of kidney function following ischaemia-reperfusion injury has been investigated using the novel technique of near-infrared spectroscopy. 2. An in vivo model of rat kidney ischaemia was used, with the expected falls in blood and tissue oxygenation seen and confirmed by near-infrared spectroscopy. 3. Allopurinol infusion increased the rate of reperfusion of oxygenated blood seen in control rats (P < 0.05). 4. Allopurinol enhanced the rate of tissue oxygenation during early reperfusion (P < 0.01). 5. This study provides further evidence for the proposed benefits of allopurinol in ischaemia-reperfusion injury. Furthermore, the potential of near-infrared spectroscopy as a technique of value in interventional studies of this nature is confirmed.     

Antiangiogenic agents protect liver sinusoidal lining cells from cold preservation injury in rat liver transplantation

BACKGROUND & AIMS: Low temperature preservation causes unique liver injuries to the sinusoidal lining cells characterized by endothelial cell detachment and rounding and Kupffer cell activation. These changes are similar to those observed during the early stages of angiogenesis. The aim of this study was to investigate if cold preservation injury is caused by the activation of angiogenic mechanisms. METHODS: Livers were obtained from rats pretreated with three well-known antiangiogenic agents (minocycline, interferon alfa-2b, and fumagillin) and were stored for various durations in cold preservation solutions. The effects of the drugs were evaluated by morphometric assessment of endothelial cell injury in H&E, trypan blue, and immunostained (TIE2/Tek) biopsy specimens. Graft functions and survival were evaluated in isolated perfused rat liver and arterialized orthotopic liver transplantation models. RESULTS: Sinusoidal lining cell integrity and viability were significantly improved in animals pretreated with the drugs. Reperfusion injury and survival were also better in pretreated animals. Interferon alfa was the most potent agent, reducing injury even in livers preserved in the current most commonly used solution (University of Wisconsin solution). CONCLUSIONS: Cold preservation injury of liver may be the results of angiogenic mechanisms. This novel observation provides a rationale for improved liver preservation using antiangiogenic agents.     

The effect of ischemia and ischemia-reperfusion on ion transport systems

Interruption of cerebral blood flow leads to dissipation of ionic gradients as the consequence of ionic channel overstimulation and ionic pump failure. The aim of this work was to study the possible effects of ischaemia and ischaemia followed by reperfusion on biochemical properties of endoplasmic calcium pump and synaptosomal sodium pump and sodium/calcium exchanger. The results presented in this study showed that 15 minute ischaemia led to the inhibition of all three ionic transport systems, however in different degrees. 60 minute reperfusion following 15 minute ischaemia led to partial recovery of calcium pump and sodium/calcium exchanger. The activity of sodium pump was still significantly depressed. Ischaemia and ischemia followed by reperfusion did not affect kinetic parameters of calcium pump. On the other side, both ischaemia and ischaemia-reperfusion led to an increase of sodium pump affinity to ATP and a decrease of the enzyme affinity to potassium. The possible causes of the changes, as the alteration of membrane structure or altered enzymes phosphorylation are discussed in the study. In addition to the inhibitory effect of ischaemia-reperfusion injury, intracellular water accumulation, as the possible consequence of altered ion homeostasis, is documented by nuclear magnetic resonance (imaging).     

Cyclosporin-A reduces leukocyte accumulation and protects against myocardial ischaemia reperfusion injury in rats

The present study was designed to evaluate the effect of cyclosporin A in a rat model of myocardial ischaemia reperfusion injury (MI/R). Anaesthetized rats were subjected to total occlusion (20 min) of the left main coronary artery followed by 5 h reperfusion (MI/R). Sham myocardial ischaemia-reperfusion rats (Sham MI/R) were used as controls. Myocardial necrosis, myocardial myeloperoxidase activity (MPO), serum creatinine phosphokinase activity (CPK), serum tumor necrosis factor (TNF-alpha), cardiac mRNA for TNF-alpha, cardiac intercellular adhesion molecule-1 (ICAM-1) immunostaining and myocardial contractility (left ventricle dP/dtmax) were evaluated. Myocardial ischaemia plus reperfusion in untreated rats produced marked myocardial necrosis, increased serum CPK activity and myeloperoxidase activity (a marker of leukocyte accumulation) both in the area-at-risk and in the necrotic area, reduced myocardial contractility and induced a marked increase in the serum levels of the TNF-alpha. Furthermore increased cardiac mRNA for TNF-alpha was measurable within 10 to 20 min of left main coronary artery occlusion in the area-at-risk and increased levels were generally sustained for 0.5 h. Finally, myocardial ischaemia-reperfusion injury increased ICAM-1 staining in the myocardium. Administration of cyclosporin A (0.25, 0.5 and 1 mg/kg as an i.v. infusion 5 min after coronary artery occlusion) lowered myocardial necrosis and myeloperoxidase activity in the area-at-risk and in the necrotic area, decreased serum CPK activity, increased myocardial contractility, reduced serum levels of TNF-alpha and the cardiac cytokine mRNA levels, and blunted ICAM-1 immunostaining in the injured myocardium. The data suggest that cyclosporin A suppresses leukocyte accumulation and protects against myocardial ischaemia-reperfusion injury.     

Role of the donor liver in the origin of platelet disorders and hyperfibrinolysis in liver transplantation

We investigated the role of the donor liver in the origin of platelet disorders and hemostatic defects in liver transplantation. Eighteen pigs received an orthotopic or a heterotopic, auxiliary liver graft. Liver biopsies were taken for electron microscopic studies 5-10 min after reperfusion in nine animals. Blood samples were taken from the first hepatic outflow and from the systemic circulation before and 5 min after graft recirculation. Electron microscopy did not show any evidence of microthrombi or platelet aggregation in the graft, either after orthotopic liver transplantation or after heterotopic liver transplantation. Most blood platelets, which were lying free in the sinusoids, showed cell processes and many seemed to have lost their granulae, suggesting a degree of platelet activation. There were also signs of phagocytosis of platelets by the Kupffer cells. In the hepatic outflow, platelet count was significantly lower (p < 0.05) and fibrinolytic activity significantly higher (p < 0.01), than systemic post-reperfusion values. There were no important changes in the coagulation parameters. No significant changes were found between the effects on hemostasis of orthotopic and auxiliary graft reperfusion. In the second part of the study evidence for platelet activation was found after graft reperfusion in human liver transplantation. Plasma levels of platelet factor-4 and beta-thromboglobulin increased significantly after graft reperfusion. These studies suggest that platelet disorders and increased fibrinolytic activity are the major components of the hemostatic defect after graft recirculation in liver transplantation. Sequestration of platelets in the graft is probably due to the accumulation of (activated and degranulated) platelets in the sinusoids and phagocytosis by Kupffer cells.     

Arthroscopic repair of full-thickness tears of the rotator cuff

OBJECTIVE: The authors' goal was to determine the effects of specific binding and blockade of P- and E-selectins by a soluble P-selectin glycoprotein ligand-1 (PSGL-1) in rat models of hepatic in vivo warm ischemia and ex vivo cold ischemia. The authors also sought to determine the effect of selectin blockade on isograft survival in a syngeneic rat orthotopic liver transplant model. SUMMARY BACKGROUND DATA: Ischemia/reperfusion (I/R) injury is a major factor in poor graft function after liver transplantation, which may profoundly influence early graft function and late changes. It is hypothesized that I/R injury leads to the upregulation of P-selectin, which is then rapidly translocated to endothelial cell surfaces within 5 minutes of reperfusion of the liver, initiating steps leading to tethering of polymorphonuclear neutrophil leukocytes to the vascular intima. Local production by leukocytes of interleukin-1, tumor necrosis factor-alpha, or both induces P-selectin expression on the endothelium and continues the cascade of events, which increases cell adherence and infiltration of the organ. METHODS: To examine directly the effects of selectins in a warm hepatic I/R injury model, 100 microg of PSGL-1 or saline was given through the portal vein at the time of total hepatic inflow occlusion. The effects of PSGL-1 in cold ischemia were assessed using an isolated perfused rat liver after 6 hours of 4 degrees C storage in University of Wisconsin (UW) solution, with or without the instillation of PSGL-1 before the storage. To evaluate the effect of selectin blockade on liver transplant survival, syngeneic orthotopic liver transplants were performed between inbred male Sprague-Dawley rats after 24 hours of cold ischemic storage in UW solution. A separate group of animals received two doses of 100 microg of PSGL-1 through the portal vein before storage and before reperfusion of the transplanted liver. Recipient survival was assessed at 7 days, and the Kaplan-Meier product limit estimate method was used for univariate calculations of time-dependent recipient survival events. RESULTS: In an in vivo warm rat liver ischemia model, perfusion with PSGL-1 afforded considerable protection from I/R injury, as demonstrated by decreased transaminase release, reduced histologic hepatocyte damage, and suppressed neutrophil infiltration, versus controls (p < 0.05). When cold stored livers were reperfused, PSGL-1 reduced the degree of hepatocyte transaminase release, reduced neutrophil infiltration, and decreased histologic hepatocyte damage (p < 0.05 vs. UW-only controls). On reperfusion, livers treated with PSGL-1 demonstrated increased portal vein blood flow and bile production (p < 0.05 vs. UW-only controls). In addition, 90% of the rats receiving liver isografts stored in UW solution supplemented with PSGL-1 survived 7 days versus 50% of those whose transplanted syngeneic livers had been stored in UW alone (p < 0.05). CONCLUSIONS: Selectins play an important role in I/R injury of the liver. Early modulation of the interaction between P-selectin and its ligand decreases hepatocyte injury, neutrophil adhesion, and subsequent migration in both warm and cold rat liver ischemia models. In addition, the use of PSGL-1 before ischemic storage and before transplantation prevents hepatic injury, as documented by a significant increase in liver isograft survival. These findings have important clinical ramifications: early inhibition of alloantigen-independent mechanisms during the I/R damage may influence both short- and long-term survival of liver allografts.     

Gentle in situ liver manipulation during organ harvest decreases survival after rat liver transplantation: role of Kupffer cells

BACKGROUND: The etiology of primary graft nonfunction and dysfunction is unknown but most likely involves Kupffer cell-dependent reperfusion injury. However, the donor operation and surgical technique may also have an effect on the outcome after transplantation. Because liver manipulation during harvest cannot be prevented completely with standard procedures, its effect on survival was assessed here. METHODS: Donor livers were harvested from female Sprague-Dawley rats (200-230 g). Briefly, after minimal dissection during the first 12 min, livers were either manipulated gently or left alone for 13 subsequent minutes. At 25 min, all livers were perfused with cold University of Wisconsin solution via the portal vein, and transplantation was performed after cold storage (1 hr). In some rats, Kupffer cells were destroyed with gadolinium chloride or inactivated with dietary glycine before harvest. Survival, proteolytic activity in the rinse effluent, serum transaminases, trypan blue distribution to index microcirculation, and histology were compared. RESULTS: In the nonmanipulated group, survival was 100% after transplantation; however, gentle manipulation decreased survival by 70%. Further, manipulation elevated transaminases fivefold and caused about 200% necrosis. At harvest, proteolytic activity and the time for trypan blue to distribute homogeneously were elevated three- to eightfold by manipulation. Gadolinium chloride and glycine prevented the effects of manipulation on all parameters studied. CONCLUSION: These data indicate for the first time that brief, gentle manipulation of the donor liver has a marked detrimental effect on survival by priming or activating Kupffer cells. This may represent an important early event in pathogenesis, because Kupffer cells play an important role in primary graft nonfunction.     

Lower limb ischaemia-reperfusion injury alters gastrointestinal structure and function

BACKGROUND: It has been suggested that bowel permeability is altered following abdominal aortic aneurysm surgery. The effect of ischaemia-reperfusion injury to the lower limb on the morphological structure, neutrophil infiltration and permeability of the bowel was investigated. METHODS: Histological assessment of the bowel was undertaken in five groups of Wistar rats: control, 3 h of bilateral hind limb ischaemia and 3 h of bilateral hind limb ischaemia followed by 1, 2 or 3 h of reperfusion. Using an everted gut sac model and 14C-labelled polyethylene glycol, the effect of ischaemia-reperfusion on small bowel permeability was studied. RESULTS: The small bowel showed a significant decrease in mucosal thickness, villus height and crypt depth in animals subjected to ischaemia followed by 2-hr reperfusion (mean(s.e.m.) 420(15), 217(9) and 163(6) microns respectively) compared with controls (481(11), 245(6) and 195(6) microns) (P < 0.05). Neutrophil count within the lamina propria was similar in the different groups. A significant increase in mean(s.e.m.) 14C-labelled polyethylene glycol translocation was detected in animals subjected to ischaemia-reperfusion compared with controls (760(40) versus 560(27) c.p.m. per ml per h) (P < 0.05). CONCLUSION: These data suggest that reperfusion of acutely ischaemic extremities produces structural and functional changes in the small intestine, although these changes are not associated with increased neutrophil infiltration within the bowel wall.     

Maitotoxin, a cationic channel activator

ICAM-1 mediates the recruitment of neutrophils through the endothelium to the site of inflammation by the ICAM-1/Mac-1 and ICAM-1/LFA-1 adhesion pathways. In extrahepatic cholestasis, recruitment of neutrophils is a main feature of the inflammatory infiltrate in areas of parenchymal damage. The aim of the present study was to describe the light and electron microscopical localization of ICAM-1 expression in the liver of cholestatic patients. The peroxidase-antiperoxidase technique was used. Increased ICAM-1 expression was detected on sinusoidal endothelial and Kupffer cells. A de novo ICAM-1 expression was described on some Ito cells and the sinusoidal hepatocyte membrane in areas of parenchymal injury. In the portal areas of livers of cholestatic patients, ICAM-1 was observed on the endothelial surface of portal veins and on hepatic arteries. Occasionally, ICAM-1 was found on the surface of bile duct epithelia. It is suggested that ICAM-1 expression is up-regulated by cytokines like TNF-alpha, IL-1 and interferons released from activated Kupffer cells. The mechanisms of ICAM-1 upregulation and neutrophil recruitment in the liver during extrahepatic cholestasis are discussed.     

Midazolam-flumazenil topical anaesthesia for microlaryngoscopy with jet ventilation

BACKGROUND: Enhanced production of endothelin-1 (ET-1), vasoconstrictive 21 amino acids produced by endothelial cells during ischemia and after reperfusion of the liver, is known to cause sinusoidal constriction and microcirculatory disturbances, which lead to severe tissue damage. Using a 2-hour hepatic vascular exclusion model in dogs, we tested our hypothesis that neutralization of ET-1 by monoclonal anti-ET-1 and anti-ET-2 antibody (AwETN40) abates vascular dysfunction and ameliorates ischemia/reperfusion injury of the liver. STUDY DESIGN: After skeletonization, the liver was made totally ischemic by cross-clamping the portal vein, the hepatic artery, and the vena cava (above and below the liver). Veno-venous bypass was used to decompress splanchnic and inferior systemic congestion. AwETN40, 5 mg/kg, was administered intravenously 10 minutes before ischemia (treatment group, n = 5). Nontreated animals were used as controls (control group, n = 10). Animal survival, hepatic tissue blood flow, liver function tests, total bile acid, high-energy phosphate, ET-1 levels, and liver histopathology were studied. RESULTS: Treatment with AwETN40 improved 2-week animal survival from 30% to 100%. Hepatic tissue blood flow after reperfusion was significantly higher in the treatment group. The treatment significantly attenuated liver enzyme release, total bile acid, and changes in adenine nucleotides. Immunoreactive ET-1 levels in the hepatic venous blood of the control group showed a significant increase and remained high for up to 24 hours after reperfusion. Histopathologic alterations were significantly lessened in the treatment group. CONCLUSIONS: These results indicate that ET-1 is involved in ischemia/reperfusion injury of the liver, which can be ameliorated by the monoclonal anti-ET-1 and anti-ET-2 antibody AwETN40.     

Kupffer cell inactivation prevents lipopolysaccharide-induced structural changes in the rat liver sinusoid: an electron-microscopic study

Scanning and transmission electron-microscopic examination of the rat liver sinusoid was performed in this study after in vivo treatment of rats with gram-negative bacterial lipopolysaccharide (LPS, 1 mg/Kg(-1) body weight), with or without pretreatment with gadolinium chloride (GdCl3 10 mg(Kg(-1) body weight). Twenty-seven and 48 hours after GdCl3 administration, to inactivate/eliminate part of the Kupffer cell population, a decrease in the number of visualized Kupffer cells was observed, without evident effects on the sinusoidal endothelial cell or on the hepatocyte. Three and 24 hours after its administration, LPS produced ultrastructural changes in the sinusoid characterized by morphological evidence of Kupffer cell activation (i.e., swelling and expanded philopodia anchoring the Kupffer cell to the luminal surface of the sinusoidal wall), and a marked decrease in the population of endothelial cell fenestration. The reduction in the number of fenestrae was associated with a change in the diameter of fenestrae and can be interpreted as a component of the "capillarization" process of the hepatic sinusoid. Such ultrastructural changes were prevented by the administration of GdCl3 24 hours before LPS injection. Hence, these findings suggest that LPS-induced structural changes in the liver sinusoid are mediated by an LPS-induced Kupffer cell activation. Coupled with previous experimental data, showing similar effects of GdCl3 on one of the hepatic sinusoidal endothelial cell (SEC) functions, i.e., hyaluronan scavenging, the data presented in this study strongly support the view that Kupffer cells modulate both the hepatic SEC's functional as well as ultrastructural properties.     

Apoptosis of neutrophils and their elimination by Kupffer cells in rat liver

The fate of neutrophils in the peripheral circulation is poorly understood. In this study, the role of Kupffer cells in eliminating aged and apoptotic neutrophils was investigated. Liver, spleen, lung, and blood samples from Wistar rats were examined by light and electron microscopy, the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) method, and immunohistochemistry after the intravenous injection of OK-432, a streptococcal preparation. Neutrophils were trapped predominantly in the periportal and midzonal regions of hepatic lobules and were in contact with endothelial cells and Kupffer cells, or were surrounded by Kupffer cells. The trapping of neutrophils peaked after 6 hours. Apoptotic neutrophils, with or without buds, were found in the lumen of hepatic sinusoids as early as 6 hours, reached maximal levels after 12 hours, and represented greater than 60% of the total number of neutrophils in the liver. The presence of apoptotic neutrophils was correlated with the degree of neutrophil phagocytosis. Double-staining showed that TUNEL-positive neutrophils were phagocytosed or encircled by ED1- or ED2-positive Kupffer cells. In contrast, apoptosis and phagocytosis of neutrophils were rare in the spleen, lung, and peripheral blood. These results suggested that the appearance of apoptotic neutrophils in the hepatic sinusoids and their rapid clearance by Kupffer cells occurs after the invasion of bacteria (i.e., bacteremia or bacteriotoxemia) or the release of inflammatory mediators into the blood stream. These findings have important implications for the regulation of neutrophil homeostasis, the limitation of inflammation and tissue injury, and provide insight into the physiological removal of circulating, senescent neutrophils.