Failure of regeneration of the steatotic rat liver: disruption at two different levels in the regeneration pathway

Hepatic resection or transplantation in patients with fatty liver is associated with increased morbidity and mortality. The regenerative capacity of fatty livers after major tissue loss is unknown. Interleukin 6 (IL-6) is a potent inducer of hepatic regeneration in normal and ischemic livers. Therefore, we studied hepatic regeneration at day 1, day 2, and day 4 in a model of 70% hepatectomy in obese and lean Zucker rats, and obese Zucker rats pretreated with recombinant interleukin 6 (rIL-6). The mitotic cycle in hepatocytes was investigated by 4 different markers of regeneration representing distinct phases of mitosis (proliferating cell nuclear antigen [PCNA] = G(1) phase, bromodeoxy uridine [BrdU] = S phase, mitotic index, and regenerated liver weight = M phase). Obese Zucker rats had significantly decreased regenerative capacity compared with lean Zucker rats (PCNA, BrdU, mitotic index, regenerated liver weight) at days 1 and 2 after surgery. Four days after resection fatty animals showed an increase in the mitotic index indicating a delay of regeneration in steatotic livers. Animal survival after 70% hepatectomy was significantly decreased in obese rats compared with lean animals. Pretreatment of obese animals with rIL-6 normalized PCNA expression (G(1) phase) in steatotic hepatocytes but failed to increase DNA synthesis (BrdU, S phase), mitosis (mitotic index and regenerated liver weight, M phase), and animal survival. These results indicate major impairment of hepatic regeneration in steatotic livers. Two different blockages of regeneration must be present, one rIL-6 sensitive, at the level of IL-6 or upstream, and a second, rIL-6 resistant, at the level of G(1)/S-phase transition.     

ICAM-1 triggers liver regeneration through leukocyte recruitment and Kupffer cell-dependent release of TNF-alpha/IL-6 in mice

BACKGROUND & AIMS: Tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 mediate hepatocyte proliferation in vivo, suggesting that local and systemic inflammatory reactions may trigger hepatic regeneration after major tissue loss. METHODS: Wild-type, intercellular adhesion molecule (ICAM)-1-/-, and neutropenic-induced mice were subjected to 70% hepatectomy. Three different approaches to block and/or deplete liver macrophages (Kupffer cells) were used. RESULTS: We found that liver from ICAM-1-deficient mice exhibited impaired regeneration after partial hepatectomy. This finding is associated with dramatic decrease in leukocyte recruitment and tissue TNF-alpha and IL-6 levels. All markers of hepatocyte proliferation were restored in ICAM-/- mice by injections of recombinant IL-6. Neutropenic animals and liver macrophage (Kupffer cell) depletion resulted in similar failure of regeneration with low levels of TNF-alpha and IL-6. CONCLUSIONS: The data suggest a novel pathway in which ICAM-1 binds to leukocytes after hepatectomy, triggering hepatocyte proliferation through Kupffer cell-dependent release of TNF-alpha and IL-6.     

Endogenous IL-13 protects hepatocytes and vascular endothelial cells during ischemia/reperfusion injury

Hepatic ischemia/reperfusion injury involves a complex inflammatory cascade resulting in neutrophil-mediated injury of hepatocytes. Previous studies from our laboratory have established that exogenous administration of the anti-inflammatory cytokines interleukin 10 (IL-10) and IL-13 can ameliorate the inflammatory response and significantly reduce hepatocellular injury. The purpose of the present study was to determine if IL-10 and IL-13 function as endogenous regulators of the hepatic inflammatory response to ischemia/reperfusion. Wild-type, IL-10-, and IL-13-deficient (IL-10(-/-), IL-13(-/-)) mice were exposed to 90 minutes of partial hepatic ischemia and up to 24 hours of reperfusion. In wild-type mice, expression of IL-10 and IL-13 shared similar expression profiles with maximal production after 8 hours of reperfusion. There were no significant differences between wild-type and IL-10(-/-) mice in response to hepatic ischemia and reperfusion. IL-13(-/-) mice had much greater liver injury, as assessed biochemically and histologically, than wild-type mice. There were no differences between wild-type and IL-13(-/-) mice in their production of inflammatory cytokines, but IL-13(-/-) mice displayed disrupted neutrophil accumulation, with less neutrophils present in the hepatic parenchyma and far more neutrophils adherent to the endothelium of large hepatic venules than wild-type mice. These observations were associated with increased liver endothelial cell injury in IL-13(-/-) mice, as measured by serum levels of hyaluronic acid. In vitro, IL-13 protected hepatocytes from H(2)O(2)-induced cytotoxicity. In conclusion, IL-10 is not an important endogenous regulator of the inflammatory response to hepatic ischemia/reperfusion. In contrast, endogenous IL-13 appears to be critical for the control of this response, with prominent protective effects on hepatocytes and hepatic endothelial cells.     

Requirement for interleukin-12 in the pathogenesis of warm hepatic ischemia/reperfusion injury in mice

Hepatic ischemia and reperfusion causes neutrophil-dependent liver injury. Although the mechanisms of ischemia/reperfusion-induced liver neutrophil recruitment are somewhat understood, less is known regarding the early events that initiate the inflammatory injury. Using a murine model of partial hepatic ischemia and reperfusion, we evaluated the role of endogenous interleukin (IL)-12 in this inflammatory response. Hepatic ischemia for 90 minutes and reperfusion for up to 4 hours resulted in hepatocyte expression of IL-12. By 8 hours of reperfusion there were large increases in serum levels of interferon-gamma (IFNgamma) and tumor necrosis factor-alpha (TNFalpha). In addition, hepatic ischemia/reperfusion caused significant increases in liver neutrophil recruitment, hepatocellular injury, and liver edema, as defined by liver myeloperoxidase content, serum alanine aminotransferase, and liver wet to dry weight ratios, respectively. In mice treated with neutralizing antibody to IL-12 and in mice deficient in the IL-12 p40 gene, ischemia/reperfusion-induced increases in IFNgamma and TNFalpha were greatly diminished. These conditions also caused significant reductions in liver myeloperoxidase content and attenuated the parameters of liver injury. The data suggest that IL-12 is required for the full induction of injury after hepatic ischemia and reperfusion.     

Platelets and platelet-derived serotonin promote tissue repair after normothermic hepatic ischemia in mice

Hepatic ischemia and reperfusion (I/R) leads to the formation of leukocyte-platelet aggregates. Upon activation, platelets generate reactive oxygen species and release proapoptotic and proinflammatory mediators as well as growth factors. In cold hepatic ischemia, adhesion of platelets to endothelial cells mediates sinusoidal endothelial cell apoptosis. Furthermore, platelet-derived serotonin mediates liver regeneration. We hypothesized that platelets may contribute to reperfusion injury and repair after normothermic hepatic ischemia. The aim of this study was to assess the impact of platelets in normothermic hepatic I/R injury using models of impaired platelet function and immune thrombocytopenia. Inhibition of platelet function in mice was achieved via clopidogrel feeding. Immune thrombocytopenia was induced via intraperitoneal injection of anti-CD41 antibody. Platelet-derived serotonin was investigated using mice lacking tryptophan hydroxylase 1. Mice were subjected to 60 minutes of partial hepatic ischemia and various time points of reperfusion. Hepatic injury was determined via AST and histological analysis of the necrotic area as well as leukocyte infiltration. Liver regeneration was determined via proliferating cell nuclear antigen and Ki67 immunohistochemistry. Neither inhibition of platelet function nor platelet depletion led to a reduction of I/R injury. Liver regeneration and repair were significantly impaired in platelet-depleted animals. Mice lacking peripheral serotonin were deficient in hepatocyte proliferation, but otherwise displayed normal tissue remodeling. Conclusion: Platelets have no direct impact on the pathogenesis of normothermic I/R injury. However, they mediate tissue repair and liver regeneration. Furthermore, platelet-derived serotonin is a mediator of hepatocyte proliferation in the postischemic liver, but has no impact on tissue remodeling.     

Exogenous melatonin protects small‐for‐size liver grafts by promoting monocyte infiltration and releases interleukin‐6

Defective regeneration of small‐for‐size (SFS ) liver remnants and partial grafts remains a key limiting factor in the application of liver surgery and transplantation. Exogenous melatonin (MLT ) has protective effects on hepatic ischemia‐reperfusion injury (IRI ), but its influence on graft regeneration is unknown. The aim of the study is to investigate the role of MLT in IRI and graft regeneration in settings of partial liver transplantation. We established three mouse models to study hepatic IRI and regeneration associated with partial liver transplantation: (I) IR +PH group: 60 minutes liver ischemia (IR ) plus 2/3 hepatectomy (PH ); (II ) IR +exPH group: 60 minutes liver IR plus extended hepatectomy (exPH ) associated with the SFS syndrome; (III ) SFS ‐LT group: Arterialized 30% SFS liver transplant. Each group was divided into MLT or vehicle‐treated subgroups. Hepatic injury, inflammatory signatures, liver regeneration, and animal survival rates were assessed. MLT reduced liver injury, enhanced liver regeneration, and promoted interleukin (IL ) 6, IL 10, and tumor necrosis factor‐α release by infiltrating, inflammatory Ly6C+ F4/80+ monocytes in the IR +PH group. MLT ‐induced IL 6 significantly improved hepatic microcirculation and survival in the IR +exPH model. In the SFS ‐LT group, MLT promoted graft regeneration and increased recipient survival along with increased IL 6/GP 130‐STAT 3 signaling. In IL 6 ^?/? mice, MLT failed to promote liver recovery, which could be restored through recombinant IL 6. In the IR +exPH and SFS ‐LT groups, inhibition of the IL 6 co‐receptor GP 130 through SC 144 abolished the beneficial effects of MLT . MLT ameliorates SFS liver graft IRI and restores regeneration through monocyte‐released IL 6 and downstream IL 6/GP 130‐STAT 3 signaling.     

Paneth cell-derived interleukin-17A causes multiorgan dysfunction after hepatic ischemia and reperfusion injury

Hepatic ischemia and reperfusion (IR) injury is a major clinical problem that leads to frequent extrahepatic complications including intestinal dysfunction and acute kidney injury (AKI). In this study we aimed to determine the mechanisms of hepatic IR-induced extrahepatic organ dysfunction. Mice subjected to 60 minutes of hepatic IR not only developed severe hepatic injury but also developed significant AKI and small intestinal injury. Hepatic IR induced small intestinal Paneth cell degranulation and increased interleukin-17A (IL-17A) levels in portal vein plasma and small intestine. We also detected increased levels of IL-17A messenger RNA (mRNA) and protein in Paneth cells after hepatic IR with laser capture dissection. IL-17A-neutralizing antibody treatment or genetic deletion of either IL-17A or IL-17A receptors significantly protected against hepatic IR-induced acute liver, kidney, and intestinal injury. Leukocyte IL-17A does not contribute to organ injury, as infusion of wildtype splenocytes failed to exacerbate liver and kidney injury in IL-17A-deficient mice after hepatic IR. Depletion of Paneth cell numbers by pharmacological (with dithizone) or genetic intervention (SOX9 flox/flox Villin cre+/- mice) significantly attenuated intestinal, hepatic, and renal injury following liver IR. Finally, depletion of Paneth cell numbers significantly decreased small intestinal IL-17A release and plasma IL-17A levels after liver IR. CONCLUSION: Taken together, the results show that Paneth cell-derived IL-17A plays a critical role in hepatic IR injury and extrahepatic organ dysfunction. Modulation of Paneth cell dysregulation may have therapeutic implications by reducing systemic complications arising from hepatic IR.     

Intestinal ischemia-reperfusion impairs liver regeneration after partial hepatectomy in rats

BACKGROUND/AIMS: The deleterious effects of intestinal ischemia-reperfusion on liver are realized, but its effect on the regenerative capacity of the liver has not been studied. Our aim in this study was to determine the effect of intestinal ischemia-reperfusion on liver regeneration. METHODOLOGY: Sprague-Dawley rats were randomly divided into six groups; two sham-operated, two hepatectomy, and two hepatectomy with intestinal ischemia-reperfusion groups. To create intestinal ischemia-reperfusion, the superior mesenteric artery and collateral arteries supplying the small intestine were occluded for 20 minutes. Partial hepatectomy was performed during the period of ischemia. Ischemia-reperfusion injury in the mucosal layer of the small intestine was scored in light microscopy. Liver regeneration parameters (proliferating cell nuclear antigen labeling index for hepatocytes and liver regeneration rate), and serum levels of aspartate aminotransferase and alanine aminotransferase were studied on day 1 or 4 after operation. RESULTS: Mucosal injury score was high in the hepatectomy with intestinal ischemia-reperfusion groups. Liver regeneration rate and proliferating cell nuclear antigen labeling index were less in these groups than the hepatectomy groups on day 1 and 4. There were no differences in the serum levels of aspartate aminotransferase and alanine aminotransferase between hepatectomy and hepatectomy with intestinal ischemia-reperfusion groups. The mortality rate was higher in the hepatectomy with intestinal ischemia-reperfusion groups than the other groups. CONCLUSIONS: Ischemia and reperfusion of the small intestine impaired liver regeneration with high mortality after partial hepatectomy in the rats.     

Amelioration of hepatic ischemia/reperfusion injury in the remnant liver after partial hepatectomy in rats

BACKGROUND AND AIMS: Reactive oxygen species have been implicated in the development of hepatic ischemia/reperfusion (I/R) injury. I/R injury remains an important problem in massive hepatectomy and organ transplantation. The aim of this study was to examine the effect of edaravone, a newly synthesized free radical scavenger, on I/R injury in the remnant liver after partial hepatectomy in rats. METHODS: Partial (70%) hepatic ischemia was induced in rats by occluding the hepatic artery, portal vein, and bile duct to left and median lobes of liver. Total hepatic ischemia (Pringle maneuver) was induced by occluding the hepatoduodenal ligament. Edaravone was intravenously administered to rats just before reperfusion and partial (70%) hepatectomy was performed just after reperfusion. RESULTS: Edaravone significantly reduced the increases in the levels of serum alanine aminotransferase and aspartate aminotransferase in rats with liver injury induced by 90-min of partial ischemia followed by 120-min of reperfusion. Histopathological analysis showed that edaravone prevented inflammatory changes in the livers with I/R injury. Edaravone also decreased the levels of myeloperoxidase activity, which is an index of neutrophil infiltration, and interleukin-6 mRNA, which is a proinflammatory cytokine. Additionally, edaravone improved the survival rate in partial hepatectomy rats with I/R injury induced by the Pringle maneuver. CONCLUSIONS: Edaravone administration prior to reperfusion protected the liver against I/R injury. Edaravone also improved the function of the remnant liver with I/R injury after partial hepatectomy. Therefore, edaravone may have applicability for major hepatectomy and liver transplantation in the clinical setting.     

Neutrophil-mediated liver injury during hepatic ischemia-reperfusion in rats

BACKGROUND: Neutrophil plays an important role in hepatic ischemia-reperfusion injury. We investigated neutrophil infiltration in liver tissue, Kupffer cells' role in neutrophil accumulation, and apoptosis and regeneration of hepatocytes in liver ischemia-reperfusion injury. METHODS: Vascular microclamps were placed across the pedicles of the median and left lateral lobes for 90 minutes after 30% hepatectomy with the resection of caudate, right lateral and quadrate lobes and papillary process. Gadolinium chloride (GdCl3) was used to destroy Kupffer cells. Neutrophil activity was inhibited with Urge-8, a monoclonal antibody against neutrophil produced in our laboratory. GdCl3 (10 mg/kg) and Urge-8 (50 mg/kg) were given intravenously in respective groups. Ischemia control, GdCl3 and Urge-8 groups were compared. RESULTS: Following hepatic reperfusion, serum interleukin-8 (IL-8) levels and hepatic neutrophil counts peaked at 3 hours, and peak concentrations of alanine aminotransferase (ALT) occurred at 6 hours. Animals of the control group showed increases in neutrophil infiltration in liver tissue, liver enzyme levels, and apoptosis index of hepatocytes and decreases in overall survival rate and proliferating cell nuclear antigen (PCNA) expression of hepatocytes. The survival rates and PCNA proportion of hepatocytes were higher and the levels of hepatic neutrophil infiltration, liver enzymes, and hepatocyte apoptosis after reperfusion were lower in the GdCl3 and Urge-8 groups than those in the ischemia control group. CONCLUSIONS: Blockades of Kupffer cells' activity and neutrophil infiltration by GdCl3 and Urge-8 eliminate neutrophil-mediated hepatic injury and enhance subsequent hepatic regeneration during liver ischemiareperfusion.     

Peroxisome proliferator-activated receptor-gamma protects against hepatic ischemia/reperfusion injury in mice

The function of peroxisome proliferator-activated receptor-gamma (PPARgamma) in hepatic inflammation and injury is unclear. In this study, we sought to determine the role of PPARgamma in hepatic ischemia/reperfusion injury in mice. Male mice were subjected to 90 minutes of partial hepatic ischemia followed by up to 8 hours of reperfusion. PPARgamma was found to be constitutively activated in hepatocytes but not in nonparenchymal cells. Upon induction of ischemia, hepatic PPARgamma activation rapidly decreased and remained suppressed throughout the 8-hour reperfusion period. This reduced activation was not a result of decreased protein availability as hepatic nuclear PPARgamma, retinoid X receptor-alpha (RXRalpha), and PPARgamma/RXRalpha heterodimer expression was maintained. Accompanying the decrease in PPARgamma activation was a decrease in the expression of the natural ligand 15-deoxy-Delta(12,14)-prostaglandin J(2). This was associated with reduced interaction of PPARgamma and the coactivator, p300. To determine whether PPARgamma activation is hepatoprotective during hepatic ischemia/reperfusion injury, mice were treated with the PPARgamma agonists, rosiglitazone and connecting peptide. These treatments increased PPARgamma activation and reduced liver injury compared to untreated mice. Furthermore, PPARgamma-deficient mice had more liver injury after ischemia/reperfusion than their wild-type counterparts. Conclusion: These data suggest that PPARgamma is an important endogenous regulator of, and potential therapeutic target for, ischemic liver injury.     

Interleukin-6 is a key mediator of the hepatoprotective and pro-proliferative effects of ischaemic preconditioning in mice

BACKGROUND/AIMS: The biological effects of ischaemic preconditioning include NF-kappaB activation, increased TNF synthesis, stimulation of cell cycle entry and hepatoprotection against ischaemia-reperfusion (IR) injury. Low dose TNF initiates the priming phase of liver regeneration via NF-kappaB and IL-6. To determine whether (1) IL-6 is released during preconditioning and confers protection against hepatic IR injury, and (2) IL-6 could mediate the biological effects of preconditioning. METHODS: Wildtype (wt) and TNF-/- C57BL6 mice were subjected to 90 min partial hepatic ischaemia and 2-44 h reperfusion with or without prior 10 min ischaemic preconditioning. To restitute liver injury, TNF-/- mice were administered murine TNF 5 microg/kg iv 1 min prior to IR. Murine recombinant IL-6 (500 ng/kg iv) was administered 30 min prior to IR, either to wt mice or to TNF-/--repleted mice; in the latter case, 1 min before preconditioning. RESULTS: In wt mice, IL-6 attenuated hepatic IR injury and stimulated cell cycle entry. IR injury in TNF-repleted TNF-/- mice was not ameliorated by preconditioning. However, prior IL-6 administration conferred hepatoprotection (IL-6/preconditioned: 349+/-169 U/L vs vehicle/preconditioned: 1250+/-608 U/L, P<0.01). CONCLUSIONS: IL-6 is one likely mediator of the hepatoprotective and pro-proliferative effects of ischaemic preconditioning.     

Liver regeneration is impaired in lipodystrophic fatty liver dystrophy mice

We previously reported that mice subjected to partial hepatectomy exhibit rapid development of hypoglycemia followed by transient accumulation of fat in the early regenerating liver. We also showed that disrupting these metabolic alterations results in impaired liver regeneration. The studies reported here were undertaken to further characterize and investigate the functional importance of changes in systemic adipose metabolism during normal liver regeneration. The results showed that a systemic catabolic response is induced in each of two distinct, commonly used experimental models of liver regeneration (partial hepatectomy and carbon tetrachloride treatment), and that this response occurs in proportion to the degree of induced hepatic insufficiency. Together, these observations suggest that catabolism of systemic adipose stores may be essential for normal liver regeneration. To test this possibility, we investigated the hepatic regenerative response in fatty liver dystrophy (fld) mice, which exhibit partial lipodystrophy and have diminished peripheral adipose stores. The results showed that the development of hypoglycemia and hepatic accumulation of fat was attenuated and liver regeneration was impaired following partial hepatectomy in these animals. The fld mice also exhibited increased hepatic p21 expression and diminished plasma levels of the adipose-derived hormones adiponectin and leptin, which have each been implicated as regulators of liver regeneration. Conclusion: These data suggest that the hypoglycemia that develops after partial hepatectomy induces systemic lipolysis followed by accumulation of fat derived from peripheral stores in the early regenerating liver, and that these events may be essential for initiation of normal liver regeneration.     

Attenuation of microcirculatory disturbance after liver ischemia by newly synthesized inflammatory cytokine suppressor,FR167653

BACKGROUND/AIMS: Inflammatory cytokines, such as interleukin-1 beta and tumor necrosis factor-alpha, which activate neutrophils, contribute to hepatic warm ischemia-reperfusion injury. However, the role of the cytokines in hepatic microcirculation immediately after reperfusion is still unclear. This study was carried out to investigate whether FR167653, a dual inhibitor of interleukin-1 beta and tumor necrosis factor-alpha, attenuates hepatic microcirculatory disturbance at the initial phase of reperfusion following liver ischemia. METHODOLOGY: Adult mongrel dogs were subjected to 90 minutes of liver ischemia by a Pringle's maneuver under portosystemic bypass. The animals were divided into two groups: a control group (n = 10), subjected to hepatic warm ischemia only, and a FR167653 administered group (n = 5), which received 1 mg/kg/h FR167653 for 4 hours since 30 minutes before the ischemia to 2 hours after the reperfusion continuously. Seven days animal survival, hepatic tissue blood flow, liver function test, hepatic venous blood concentration of endothelin-1 and plasminogen activator inhibitor-1, liver tissue biochemistry, and histopathology were analyzed. RESULTS: The treatment with FR167653 attenuated microcirculatory disturbance, lessened liver injury, enhanced adenine nucleotides resynthesis, and improved animal survival after liver ischemia. In addition, FR167653 significantly inhibited release of both endothelin-1 and plasminogen activator inhibitor-1 from the liver cells. CONCLUSIONS: These results suggest that the inflammatory cytokines induce microcirculatory disturbance in the initial phase of reperfusion following liver ischemia.     

Interleukin 6 alleviates hepatic steatosis and ischemia/reperfusion injury in mice with fatty liver disease

Fatty liver, formerly associated predominantly with excessive alcohol intake, is now also recognized as a complication of obesity and an important precursor state to more severe forms of liver pathology including ischemia/reperfusion injury. No standard protocol for treating fatty liver exists at this time. We therefore examined the effects of 10 days of interleukin 6 (IL-6) injection in 3 murine models of fatty liver: leptin deficient ob/ob mice, ethanol-fed mice, and mice fed a high-fat diet. In all 3 models, IL-6 injection decreased steatosis and normalized serum aminotransferase. The beneficial effects of IL-6 treatment in vivo resulted in part from an increase in mitochondrial beta oxidation of fatty acid and an increase in hepatic export of triglyceride and cholesterol. However, administration of IL-6 to isolated cultured steatotic hepatocytes failed to decrease lipid contents, suggesting that the beneficial effects of IL-6 in vivo do not result from its effects on hepatocytes alone. IL-6 treatment increased hepatic peroxisome proliferator-activated receptor (PPAR) alpha and decreased liver and serum tumor necrosis factor (TNF) alpha. Finally, 10 days of treatment with IL-6 prevented the susceptibility of fatty livers to warm ischemia/reperfusion injury. In conclusion, long-term IL-6 administration ameliorates fatty livers and protects against warm ischemia/reperfusion fatty liver injury, suggesting the therapeutic potential of IL-6 in treating human fatty liver disease.     

Erratum

Liver resections are frequently associated with significant ischemia-reperfusion (I-R) injury of the liver remnant. The aim of this study was to investigate whether deferoxamine (DFO) can ameliorate I-R injury during major hepatectomies performed under vascular exclusion of the liver in a porcine model. Twelve female domestic pigs were divided into control (n = 6) and DFO treatment (n = 6) groups and subjected to 150 min. liver ischemia followed by 70% hepatectomy and 24 hours reperfusion. Pigs in the DFO group received a continuous intravenous infusion of 100 mg/kg DFO. Liver remnant injury was evaluated by liver function tests, hepatic histology as well as serum and liver tissue malondialdehyde (MDA) concentrations. Deferoxamine-treated animals had reduced total bilirubin, γ-glutamyl transferase and ammonia levels as well as hepatocyte necrosis and oxidative injury. In a subsequent randomized clinical trial using DFO for I-R protection during major liver surgery, preliminary results revealed amelioration of hepatocellular damage, oxidative and inflammatory serum markers and apoptotic response in liver remnant biopsies.