Inhibition of ischemia and reflow-induced liver injury by an SOD derivative that circulates bound to albumin

Ischemia followed by reflow often results in tissue injury. Although reactive oxygens seem to play an important role in the pathogenesis of postischemic reflow-induced tissue injury, the mechanism and an efficient way to inhibit oxidative injury are not known. We studied the mechanism by which hepatic transport function was inhibited by a transient occlusion followed by reflow of the portal vein and hepatic artery by using a superoxide dismutase (SOD) derivative (SM-SOD) which circulates bound to albumin with a half-life of 6 h. Occlusion of the hepatic vessels for 20 min followed by reflow for 60 min significantly inhibited transhepatic transport of cholephilic ligands, such as bromosulfophthalein (BSP) and taurocholic acid. Intravenous administration of SM-SOD markedly inhibited the reflow-induced decrease in transhepatic transport of these ligands. Thiobarbituric acid - reactive metabolites (TBAR) in the liver and plasma remained unchanged during occlusion and reflow, while TBAR in the bile increased significantly. Intravenous injection of SM-SOD inhibited the reflow-induced increase in biliary TBAR. Xanthine oxidase activity in plasma also increased during occlusion and reflow by an SM-SOD-inhibitable mechanism. Polymorphonuclear leukocyte-dependent chemiluminescence of the peripheral blood remained unchanged during occlusion, but increased markedly with time after reflow. SM-SOD also inhibited the increase in chemiluminescence almost completely. These and other results suggested that the superoxide radical and/or its metabolite(s) might play an important role in the pathogenesis of the reflow-induced liver injury and that SM-SOD might be useful for studying the mechanism for tissue injury caused by oxygen toxicity.     

Inhibition of post-ischemic reperfusion injury of the small intestine by diamine oxidase.

To elucidate the role of diamines in the pathogenesis of post-ischemic reperfusion-induced tissue injury, the effect of diamine oxidase was studied in the rat whose superior mesenteric artery was occluded for 15 min followed by 30 min reperfusion. Kinetic analysis using radiolabeled albumin revealed that the mucosal permeability of the reperfused small intestine increased significantly. Histological examination of the reperfused intestine revealed a marked degeneration of its mucosal layer. Intravenous administration of diamine oxidase inhibited the reperfusion-induced increase in mucosal permeability of the intestine almost completely and preserved the structure of the small intestine. H1-antagonist chlorphenilamine and H2-antagonist famotidine also inhibited the reperfusion injury of the small intestine. These and other results suggested that extracellular diamines might play critical roles in post-ischemic reperfusion-induced injury of the small intestine.     

Inhibition of carrageenin-induced paw edema by a superoxide dismutase derivative that circulates bound to albumin.

Although the possible involvement of superoxide radical and its metabolite(s) in the pathogenesis of various types of edema have been suggested, direct evidence supporting this concept is lacking. Since intravenously administered Cu2+Zn2(+)-type superoxide dismutase (SOD) rapidly disappeared from the circulation with a half-life of 4 min, the enzyme could not be used to test whether superoxide radicals played a critical role in the modulation of vascular permeability. We previously synthesized a SOD derivative (SM-SOD) by linking poly(styrene co-maleic acid butyl ester) (SM) to the enzyme (Ogino, T., Inoue, M., Ando, Y., Awai, M., Maeda, H. and Morino Y. (1988) Int. J. Pept. Protein Res. 32, 1583-1588); SM-SOD circulates bound to albumin with a half-life of 6 h. To test whether superoxide radicals play an important role in the regulation of vascular permeability, the effect of SM-SOD on experimental paw edema was studied in the rat. Subcutaneous injections of carrageenin to the paw rapidly induced local edema by increasing vascular permeability. Intravenous administration of SM-SOD markedly inhibited the carrageenin-induced increase in vascular permeability and suppressed the development of paw edema. In contrast, the same dose of SOD showed no such inhibitory effect. These results suggest that superoxide radical and/or its metabolite(s) might play a critical role in the pathogenesis of carrageenin-induced vasogenic edema.     

Synthesis of a superoxide dismutase derivative that circulates bound to albumin and accumulates in tissues whose pH is decreased

Protection of tissues from oxidative stress is one of the major prerequisites for aerobic life. Since intravenously injected Cu2+/Zn2+-type superoxide dismutase (SOD) disappears from the circulation with a short half-life of 5 min, its clinical use as a scavenger for superoxide radical is limited. We synthesized a human erythrocyte type SOD derivative (SM-SOD) by linking 2 mol of hydrophobic organic anion, alpha-4-[( 6-(N-maleimido)hexanoyloxymethyl]cumyl]half-butyl-esterified poly(styrene-co-maleic acid) (SM), to the cysteinyl residues of the dimeric enzyme without decreasing enzymic activity. SM-SOD, but not SOD, bound to an albumin-Sepharose column; the bound SM-SOD was eluted by a buffer solution containing 0.5% sodium dodecyl sulfate or 10 mM warfarin, suggesting that SM-SOD reversibly binds to the warfarin site on albumin. Due to the amphipathic nature of the SMI moiety, SM-SOD bound also to cell membranes particularly when the pH was decreased. In vivo analysis in the rat revealed that intravenously injected SM-SOD circulated bound to albumin with a half-life of 6 h. Postischemic reperfusion arrhythmias were almost completely prevented by a single dose of SM-SOD, but not SOD. Thus, the prolonged half-life of SM-SOD in the circulation and its preferential accumulation in an injured site with decreased pH appeared to be responsible for preventing myocardial injury. These results suggest that superoxide radical and/or its metabolite(s) would play an important role in the pathogenesis of postischemic reperfusion arrhythmias and that SM-SOD may be useful for decreasing tissue injury in ischemic heart disease.     

Cerebroprotective activity of Wedelia calendulacea on global cerebral ischemia in rats

The present study was to investigate the effect of W. calendulacea on ischemia and reperfusion-induced cerebral injury. Cerebral ischemia was induced by occluding right and left common carotid arteries (global cerebral ischemia) for 30 min followed by reperfusion for 1 h and 4 h individually. Various biochemical alterations, produced subsequent to the application of bilateral carotid artery occlusion (BCAO) followed by reperfusion viz. increase in lipid peroxidation (LPO), hydrogen peroxide (H2O2), and decrease in reduced glutathione (GSH), catalase (CAT) and superoxide dismutase (SOD), level in the brain tissue, Western blot analysis (Cu-Zn-SOD and CAT) and assessment of cerebral infarct size were measured. All those enzymes are markedly reversed and restored to near normal level in the groups pretreated with W. calendulacea (250 and 500 mg/kg given orally in single and double dose/day for 10 days) in dose-dependent way. The effect of W. calendulacea had increased significantly the protein expression of copper/zinc superoxide dismutase (Cu-Zn-SOD) and CAT in cerebral ischemia. W. claendulacea was markedly decrease cerebral infarct damages but results are not statistically significant. It can be concluded that W. calendulacea possesses a neuroprotective activity against cerebral ischemia in rat.     

Therapeutic effects of superoxide dismutase derivatives modified with mono- or polysaccharides on hepatic injury induced by ischemia/reperfusion.

Therapeutic effects of four types of recombinant superoxide dismutase (SOD) derivatives, conjugates with polysaccharides, carboxymethyl (SOD-CMD) and diethylaminoethyl (SOD-DEAED) dextrans and galactosylated (Gal-SOD) and mannosylated (Man-SOD) derivatives, on hepatic ischemia/reperfusion injury were studied in rats. Hepatic injury induced by transient occlusion and subsequent reflow of hepatic blood was evaluated by the analysis of biliary excretion of bromosulfophthalein (BSP) injected intravenously. At a dose of 10000 units/kg, native SOD and SOD-DEAE did not show any significant effect and SOD-CMD showed slight effect. On the other hand, Gal-SOD and Man-SOD, targeted to the liver parenchymal and nonparenchymal cells, respectively, by a receptor-mediated endocytosis, exhibited superior inhibitory effects. These results demonstrated that these glycosylated SOD derivatives were useful for the prevention of hepatic ischemia/reperfusion injury.     

Targeting Sod by Gene and Protein Engineering and Inhibition of Free Radical Injury

Although oxygen toxicity of tissues can be decreased by a variety of antioxidants and some enzymes, such as SOD and catalase, their protective effect on tissue injury in various diseases are fairly small predominantly because of their unfavorable in vivo behavior. To minimize oxidative stress in various diseases. such as ischemic myocardial injury, circulatory disturbance and corneal inflammation, we synthesized three types of SOD derivatives by gene and protein engineering technique. One type of SOD (SM-SOD covalently linked with hydrophobic anions) circulates bound to albumin with a half life of 6 h and accumulates in tissues whose local pH is decreased. The other type of SOD (AC-SOD covalently linked with long chain fatty acids via the ?-amino group of lysyl residues) anchors onto membranc/lipid bilaycrs of various cells. The last type of SOD (HB-SOD synthesized by constructing a fusion gene coding human CuZn-type SOD and a C-terminal heparin-binding domain) binds to heparin-like proteoglycans on vascular cndothelial cell surface. Intravenous administration of either SM-SOD or HB-SOD markedly inhibited postischcmic reflow arrhythmias in the rat. When the left anterior descending artery was occluded permanently. about 65 % of animals died within 30 min predominantly due to irreversible ventricular fibrillation; the motality of animals decreased to 15 % by administering SM-SOD either before or after occlusion. Topically administered AC-SOD bound to the corneal epithelial cell surface and polyrnorp%onuclear leukocytes and efficiently dismutated superoxide radicals at their cell surface. Thus,' endotoxin-induced kcratitis was inhibited markedly by topical instillation of AC-SOD. Unmodified SOD itself failed to inhibit the pathologic events occurring in these disease models. Thus, these SOD derivatives permit in vivo studies on the mechanism and the site for oxygen toxicity in various diseases and provide a new strategy for targeting enzymes and bioactive peptides for medical use to appropriate site(s) of their action.     

Effect of adenosine A2A receptor agonist (CGS) on ischemia/reperfusion injury in isolated rat liver

Ischemia/reperfusion injury during liver transplantation is a major cause of primary nonfunctioning graft for which there is no effective treatment other than retransplantation. Adenosine prevents ischemia-reperfusion-induced hepatic injury via its A2A receptors. The aim of this study was to investigate the role of A2A receptor agonist on apoptotic ischemia/reperfusion-induced hepatic injury in rats. Isolated rat livers within University of Wisconsin solution were randomly divided into four groups: (1) continuous perfusion of Krebs-Henseleit solution through the portal vein for 165 minutes (control); (2) 30-minute perfusion followed by 120 minutes of ischemia and 15 minutes of reperfusion; (3) like group 2, but with the administration of CGS 21680, an A2A receptor agonist, 30 microg/100 ml, for 1 minute before ischemia; (4) like group 3, but with administration of SCH 58261, an A2A receptor antagonist. Serum liver enzyme levels were measured by biochemical analysis, and intrahepatic caspase-3 activity was measured by fluorometric assay; apoptotic cells were identified by morphological criteria, the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) fluorometric assay, and immunohistochemistry for caspase-3. Results showed that at 1 minute of reperfusion, there was a statistically significant reduction in liver enzyme levels in the animals pretreated with CGS (p < 0.05). On fluorometric assay, caspase-3 activity was significantly decreased in group 3 compared to group 2 (p < 0.0002). The reduction in postischemic apoptotic hepatic injury in the CGS-treated group was confirmed morphologically, by the significantly fewer apoptotic hepatocyte cells detected (p < 0.05); immunohistochemically, by the significantly weaker activation of caspase-3 compared to the ischemic group (p < 0.05); and by the TUNEL assay (p < 0.05). In conclusion, the administration of A2A receptor agonist before induction of ischemia can attenuate postischemic apoptotic hepatic injury and thereby minimize liver injury. Apoptotic hepatic injury seems to be mediated through caspase-3 activity.     

Electron spin resonance and spin trapping technique provide direct evidence that edaravone prevents acute ischemia-reperfusion injury of the liver by limiting free radical-mediated tissue damage

A novel free radical scavenger, 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone), is used for the treatment of acute ischemic stroke and is protective in several animal models of organ injury. We tested whether edaravone is protective against acute liver warm ischemia/reperfusion injury in the rat by acting as a radical scavenger. When edaravone was administered prior to ischemia and at the time of initiation of the reperfusion, liver injury was markedly reduced. Production of oxidants in the liver in this model was assessed in vivo by spin-trapping/electron spin resonance (ESR) spectroscopy. Ischemia/reperfusion caused an increase in free radical adducts rapidly, an effect markedly blocked by edaravone. Furthermore, edaravone treatment blunted ischemia/reperfusion-induced elevation in pro-inflammatory cytokines, infiltration of leukocytes and lipid peroxidation in the liver. These results demonstrate that edaravone is an effective blocker of free radicals in vivo in the liver after ischemia/reperfusion, leading to prevention of organ injury by limiting the deleterious effects of free radicals.     

Electron spin resonance and spin trapping technique provide direct evidence that edaravone prevents acute ischemia–reperfusion injury of the liver by limiting free radical-mediated tissue damage

A novel free radical scavenger, 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone), is used for the treatment of acute ischemic stroke and is protective in several animal models of organ injury. We tested whether edaravone is protective against acute liver warm ischemia/reperfusion injury in the rat by acting as a radical scavenger. When edaravone was administered prior to ischemia and at the time of initiation of the reperfusion, liver injury was markedly reduced. Production of oxidants in the liver in this model was assessed in vivo by spin-trapping/electron spin resonance (ESR) spectroscopy. Ischemia/reperfusion caused an increase in free radical adducts rapidly, an effect markedly blocked by edaravone. Furthermore, edaravone treatment blunted ischemia/reperfusion-induced elevation in pro-inflammatory cytokines, infiltration of leukocytes and lipid peroxidation in the liver. These results demonstrate that edaravone is an effective blocker of free radicals in vivo in the liver after ischemia/reperfusion, leading to prevention of organ injury by limiting the deleterious effects of free radicals.     

Complement inhibitor, complement receptor 1-related gene/protein y-Ig attenuates intestinal damage after the onset of mesenteric ischemia/reperfusion injury in mice.

Complement receptor 1-related gene/protein y (Crry) is a murine membrane protein that regulates the activity of both classical and alternative complement pathways. We used a recombinant soluble form of Crry fused to the hinge, CH2, and CH3 domains of mouse IgG1 (Crry-Ig) to determine whether inhibition of complement activation prevents and/or reverses mesenteric ischemia/reperfusion-induced injury in mice. Mice were subjected to 30 min of ischemia, followed by 2 h of reperfusion. Crry-Ig was administered either 5 min before or 30 min after initiation of the reperfusion phase. Pretreatment with Crry-Ig reduced local intestinal mucosal injury and decreased generation of leukotriene B(4) (LTB(4)). When given 30 min after the beginning of the reperfusion phase, Crry-Ig resulted in a decrease in ischemia/reperfusion-induced intestinal mucosal injury comparable to that occurring when it was given 5 min before initiation of the reperfusion phase. The beneficial effect of Crry-Ig administered 30 min after the initiation of reperfusion coincided with a decrease in PGE(2) generation despite the fact that it did not prevent local infiltration of neutrophils and did not have a significant effect on LTB(4) production. These data suggest that complement inhibition protects animals from reperfusion-induced intestinal damage even if administered as late as 30 min into reperfusion and that the mechanism of protection is independent of neutrophil infiltration or LTB(4) inhibition.     

Neutrophil elastase contributes to the development of ischemia-reperfusion-induced liver injury by decreasing endothelial production of prostacyclin in rats

We previously reported that nitric oxide (NO) derived from endothelial NO synthase (NOS) increased endothelial prostacyclin (PGI(2)) production in rats subjected to hepatic ischemia-reperfusion (I/R). The present study was undertaken to determine whether neutrophil elastase (NE) decreases endothelial production of PGI(2), thereby contributing to the development of I/R-induced liver injury by decreasing hepatic tissue blood flow in rats. Hepatic tissue levels of 6-keto-PGF(1alpha), a stable metabolite of PGI(2), were transiently increased and peaked at 1 h after reperfusion, followed by a gradual decrease until 3 h after reperfusion. Sivelestat sodium hydrochloride and L-658,758, two NE inhibitors, reduced I/R-induced liver injury. These substances inhibited the decreases in hepatic tissue levels of 6-keto-PGF(1alpha) at 2 and 3 h after reperfusion but did not affect the levels at 1 h after reperfusion. These NE inhibitors significantly increased hepatic tissue blood flow from 1 to 3 h after reperfusion. Both hepatic I/R-induced increases in the accumulation of neutrophils and the microvascular permeability were inhibited by these two NE inhibitors. Protective effects induced by the two NE inhibitors were completely reversed by pretreatment with nitro-l-arginine methyl ester, an inhibitor of NOS, or indomethacin. Administration of iloprost, a stable derivative of PGI(2), produced effects similar to those induced by NE inhibitors. These observations strongly suggest that NE might play a critical role in the development of I/R-induced liver injury by decreasing endothelial production of NO and PGI(2), leading to a decrease in hepatic tissue blood flow resulting from inhibition of vasodilation and induction of activated neutrophil-induced microvascular injury.     

Nebivolol regulates eNOS and iNOS expressions and alleviates oxidative stress in cerebral ischemia/reperfusion injury in rats

AimsOxidative stress-induced cell damage is reported to contribute to the pathogenesis of cerebral ischemia/reperfusion injury. This study investigated the neuroprotective effect of nebivolol against cerebral ischemia/reperfusion insult in rats.Main methodsThe model adopted was that of surgically-induced forebrain ischemia, performed by means of bilateral common carotid artery occlusion for 1 h, followed by reperfusion for 24 h. The effects of 5 and 10 mg/kg nebivolol, treated for 7 days prior to ischemia/reperfusion insult, were investigated by estimating endothelial and inducible nitric oxide synthases (eNOS and iNOS) protein expressions and assessing oxidative stress-related biochemical parameters in the rat forebrain. Also, infarct volume measurement and histopathological study of the forebrain were examined.Key findingsAdministration of nebivolol increased eNOS expression with simultaneous decrease in iNOS expression in a dose dependent manner. Moreover, nebivolol inhibited ischemia/reperfusion-induced depletion of reduced glutathione level and decreased the elevated total nitric oxide end production and malondialdehyde levels, superoxide dismutase and lactate dehydrogenase activities. A notable finding is that catalase activity was not changed in response to either ischemia/reperfusion insult or nebivolol treatment. However, the results confirmed that nebivolol significantly reduced infarct volume and alleviated ischemia/reperfusion-induced histopathological changes.SignificanceThe present study demonstrates the neuroprotective effect of nebivolol against cerebral ischemia/reperfusion insult. Neuroprotection observed with nebivolol may possibly be explained by regulating eNOS and iNOS expressions and by inhibition of oxidative stress-induced injury. Thus, nebivolol may be considered as a potential candidate for treatment in patients who are prone to stroke.     

Flurbiprofen,a Cyclooxygenase Inhibitor,Protects Mice from Hepatic Ischemia/Reperfusion Injury by Inhibiting GSK-3β Signaling and Mitochondrial Permeability Transition

Flurbiprofen acts as a nonselective inhibitor for cyclooxygenases (COX-1 and COX-2), but its impact on hepatic ischemia/reperfusion (I/R) injury remains unclear. Mice were randomized into sham, I/R and flurbiprofen (Flurb) groups. The hepatic artery and portal vein to the left and median liver lobes were occluded for 90 min and unclamped for reperfusion to establish a model of segmental (70%) warm hepatic ischemia. Pretreatment of animals with flurbiprofen prior to I/R insult significantly decreased serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH), and prevented hepatocytes from I/R-induced apoptosis/necrosis. Moreover, flurbiprofen dramatically inhibited mitochondrial permeability transition (MPT) pore opening, and thus prevented mitochondrial-related cell death and apoptosis. Mechanistic studies revealed that flurbiprofen markedly inhibited glycogen synthase kinase (GSK)-3β activity and increased phosphorylation of GSK-3β at Ser9, which, consequently, could modulate the adenine nucleotide translocase (ANT)–cyclophilin D (CyP-D) complex and the susceptibility to MPT induction. Therefore, administration of flurbiprofen prior to hepatic I/R ameliorates mitochondrial and hepatocellular damage through inhibition of MPT and inactivation of GSK-3β, and provides experimental evidence for clinical use of flurbiprofen to protect liver function in surgical settings in addition to its conventional use for pain relief.     

Aggravation of ischemia/reperfusion-induced gastric lesions in streptozotocin-diabetic rats

We examined the influence of diabetes on ischemia/reperfusion-induced gastric damage in rats, in relation to the antioxidative system. Animals were injected with streptozotocin (STZ: 70 mg/kg, i.p.) and used after 5 weeks of diabetes with blood glucose levels of >350 mg/dl. Gastric mucosal blood flow (GMBF) was measured before, during and after 20 min of ischemia (1.5 ml bleeding per 100 g body weight from the carotid artery) followed by a 15-min reperfusion in the presence of acid (100 mM HCI). At the end of each experiment, gastric damage was observed macroscopically. GMBF was reduced by ischemia in all groups of rats, followed by a gradual return after reperfusion. Ischemia/reperfusion produced hemorrhagic lesions in normal rat stomachs in the presence of 100 mM HCl. These lesions were significantly aggravated when the animals were pretreated with diethyldithiocarbamate, an inhibitor of superoxide dismutase (SOD). Ischemia/reperfusion-induced damage was also markedly exacerbated in STZ-diabetic rats, but this aggravation was significantly suppressed by pretreatment with exogenous SOD or glutathione (GSH). Diabetic rat stomachs showed significantly less SOD activity as well as GSH content than normal rat stomachs. In addition, the deleterious influence of diabetes on the gastric ulcerogenic response to ischemia/reperfusion was significantly mitigated by decreasing the blood glucose levels by daily insulin treatment. These results suggest that the gastric mucosa of diabetic rats is more vulnerable to ischemia/reperfusion-induced injury, and the mechanism may be partly accounted for by impairment of the antioxidative system associated with a reduced SOD activity and GSH content.     

Fetuin-A exerts a protective effect against experimentally induced intestinal ischemia/reperfusion by suppressing autophagic cell death

Intestinal tissue is highly susceptible to ischemia/reperfusion injury in many hazardous health conditions. The anti-inflammatory and antioxidant glycoprotein fetuin-A showed efficacy in cerebral ischemic injury; however, its protective role against intestinal ischemia/reperfusion remains elusive. Therefore, this study investigated the protective role of fetuin-A supplementation against intestinal structural changes and dysfunction in a rat model of intestinal ischemia/reperfusion. We equally divided 72 male rats into control, sham, ischemia/reperfusion, and fetuin-A-pretreated ischemia/reperfusion (100 mg/kg/day fetuin-A intraperitoneally for three days prior to surgery and a third dose 1 h prior to the experiment) groups. After 2 h of reperfusion, the jejunum was dissected and examined for spontaneous contractility. A jejunal homogenate was used to assess inflammatory and oxidative stress enzymes. Staining of histological sections was carried out with hematoxylin, eosin and Masson’s trichrome stain for evaluation. Immunohistochemistry was performed to detect autophagy proteins beclin-1, LC3, and p62. This study found that fetuin-A significantly improved ischemia/reperfusion-induced mucosal injury by reducing the percentage of areas of collagen deposition, increasing the amplitude of spontaneous contraction, decreasing inflammation and oxidative stress, and upregulating p62 expression, which was accompanied by beclin-1 and LC3 downregulation. Our findings suggest that fetuin-A treatment can prevent ischemia/reperfusion-induced jejunal structural and functional changes by increasing antioxidant activity and regulating autophagy disturbances observed in the ischemia/reperfusion rat model. Furthermore, fetuin-A may provide a protective influence against intestinal ischemia/reperfusion complications.     

Proteins released from liver after ischaemia induced an elevation of heart resistance against ischaemia-reperfusion injury: 2. Beneficial effect of liver ischaemia in situ

We have shown earlier that proteins released from the heart during preconditioning may protect non-preconditioned heart during sustained ischaemia, similarly as preconditioning itself. In other our experiments we have documented that also proteins released from isolated rat liver during reperfusion after global ischaemia performed a protective effect on isolated rat heart against ischaemia-reperfusion injury. In the current study we examined the effect of liver ischaemia in situ on resistance of rat heart to ischaemia and reperfusion injury. Wistar rats (male) were subjected to liver ischaemia maintained by occlusion of portal vein and hepatic artery for 20 min, followed with 30-min reperfusion after reopening of both vessels. Then the hearts were isolated and perfused according to Langendorf. Hearts, after initial stabilisation (15 min), were subjected to 20-min ischaemia and 30-min reperfusion. During reperfusion, the haemodynamic parameters of hearts were measured. The protein pattern of high soluble fraction (HS fraction) isolated from rat blood by precipitation with ammonium sulphate was detected by SDS-PAGE. Our results showed improved parameters of pressure and contractility in the group after liver ischaemia (ischaemic group), presented by decreased diastolic pressure and increased LVDP((S-D)) in comparison with levels of these parameters in the control group. We also observed improved heart contraction-relaxation cycles parameters (dP/dt)(max) and (dP/dt)(min) in ischaemic group as compared with the control group. On the other hand, there were no significant differences in heart rate and coronary flow between both experimental groups. SDS-PAGE showed changed protein pattern in HS fraction, particularly the levels of several low molecular weight proteins increased. We conclude that liver ischaemia induced a higher resistance of heart against ischaemia-reperfusion injury. We propose that release of some cardioprotective proteins present in HS fraction can also contribute to this cardioprotection.     

Evaluation of serum uric acid changes in different forms of hepatic vascular inflow occlusion in human liver surgeries

Uric acid values in serum have been analyzed as one of the markers to predict cellular damage due to ischemia reperfusion injury in the field of organ transplantation. The present study was conducted to confirm that uric acid values in serum could be an efficient marker of ischemic injury of liver parenchyma following hepatic vascular occlusion in human liver surgery. The changes in serum uric acid values were analyzed at fixed intervals during different liver surgeries. Significant increases in serum uric acid values were observed in patients who received the Pringle's maneuver in which hepatic vascular inflow was manipulated with a repetition of 15 min occlusion and 5 min perfusion, whereas almost no changes in uric acid values were found in both groups of patients who received the hemilobal occlusion of the Glisson's triad in which the right or left vessels were manipulated with a repetition of 30 min occlusion and 5 min perfusion and the "control method" in which the hepatic vessels of the lesion side were previously cut before liver resection. Uric acid values in serum increased in patients of Pringle's maneuver compared to those of the hemilobal occlusion of the Glisson's triad and the control method though these procedures were used in larger hepatectomies rather than Pringle's maneuver. The results indicated that serum uric acid values do not always reflect the severity of ischemia of the liver parenchyma but reflect intestinal congestion because marked intestinal congestion was observed in patients of Pringle's maneuver but not in patients of the hemilobal occlusion of the Glisson's triad and the control method. The evaluation of the severity of the ischemic injury of the liver should be done with caution when uric acid is used as a marker in human liver surgery.