Joan Roselló-Catafau

Ischemic Preconditioning Increases the Tolerance of Fatty Liver to Hepatic Ischemia-Reperfusion Injury in the Rat

Hepatic steatosis is a major risk factor in ischemia-reperfusion. The present study evaluates whether preconditioning, demonstrated to be effective in normal livers, could also confer protection in the presence of steatosis and investigates the potential underlying protective mechanisms. Fatty rats had increased hepatic injury and decreased survival after 60 minutes of ischemia compared with lean rats. Fatty livers showed a degree of neutrophil accumulation and microcirculatory alterations similar to that of normal livers. However, in presence of steatosis, an increased lipid peroxidation that could be reduced with glutathione-ester pretreatment was observed after hepatic reperfusion. Ischemic preconditioning reduced hepatic injury and increased animal survival. Both in normal and fatty livers, this endogenous protective mechanism was found to control lipid peroxidation, hepatic microcirculation failure, and neutrophil accumulation, reducing the subsequent hepatic injury. These beneficial effects could be mediated by nitric oxide, because the inhibition of nitric oxide synthesis and nitric oxide donor pretreatment abolished and simulated, respectively, the benefits of preconditioning. Thus, ischemic preconditioning could be an effective surgical strategy to reduce the hepatic ischemia-reperfusion injury in normal and fatty livers under normothermic conditions, including hepatic resections, and liver transplantation.

Ischemic preconditioning: A defense mechanism against the reactive oxygen species generated after hepatic ischemia reperfusion

Background. Preconditioning protects against both liver and lung damage after hepatic ischemia-reperfusion (I/R). Xanthine and xanthine oxidase (XOD) may contribute to the development of hepatic I/R. Objective. To evaluate whether preconditioning could modulate the injurious effects of xanthine/XOD on the liver and lung after hepatic I/R. Methods. Hepatic I/R or preconditioning previous to I/R was induced in rats. Xanthine and xanthine dehydrogenase/xanthine oxidase (XDH/XOD) in liver and plasma were measured. Hepatic injury and inflammatory response in the lung was evaluated. Results. Preconditioning reduced xanthine accumulation and conversion of XDH to XOD in liver during sustained ischemia. This could reduce the generation of reactive oxygen species (ROS) from XOD, and therefore, attenuate hepatic I/R injury. Inhibition of XOD prevented postischemic ROS generation and hepatic injury. Administration of xanthine and XOD to preconditioned rats led to hepatic MDA and transaminase levels similar to those found after hepatic I/R. Preconditioning, resulting in low circulating levels of xanthine and XOD activity, reduced neutrophil accumulation, oxidative stress, and microvascular disorders seen in lung after hepatic I/R. Inhibition of XOD attenuated the inflammatory damage in lung after hepatic I/R. Administration of xanthine and XOD abolished the benefits of preconditioning on lung damage. Conclusions. Preconditioning, by blocking the xanthine/XOD pathway for ROS generation, would confer protection against the liver and lung injuries induced by hepatic I/R.

Ischemic preconditioning affects interleukin release in fatty livers of rats undergoing ischemia/reperfusion

The present study evaluates the effect of ischemic preconditioning on interleukin‐1 (IL‐1) and interleukin‐10 (IL‐10) generation following hepatic ischemia/reperfusion (I/R) in normal and steatotic livers as well as the role of nitric oxide (NO) in this process. Increased IL‐1β and IL‐10 levels were observed in normal livers after I/R. Steatotic livers showed higher IL‐1β levels than normal livers, and IL‐10 at control levels. The injurious role of IL‐1β and the benefits of IL‐10 on hepatic I/R injury was shown with the use of IL‐1 receptor antagonist (IL‐1ra), anti‐IL‐10 polyclonal antibody against IL‐10 (anti‐IL‐10) and exogenous IL‐10. The effective dose of these treatments was different in both types of livers. Preconditioning prevented IL‐1β release and increased IL‐10 generation after I/R in normal and steatotic livers. IL‐1β or anti‐IL‐10 pretreatments reversed the benefits of preconditioning. IL‐1β action inhibition in a preconditioned group that was pretreated with anti‐IL‐10 did not modify the benefits of preconditioning. In addition, anti‐IL‐10 pretreatment in the preconditioned group resulted in IL‐1β levels comparable to those observed after I/R. NO inhibition eliminated the benefits of preconditioning on IL‐10 release, IL‐1β levels, and hepatic injury. In conclusion, preconditioning, through IL‐10 overproduction, inhibits IL‐1β release and the ensuing hepatic I/R injury in normal and steatotic livers. IL‐10 generation induced by preconditioning could be mediated by NO. (HEPATOLOGY 2004;39:688–698.)

Activation of alveolar macrophages in lung injury associated with experimental acute pancreatitis is mediated by the liver.

OBJECTIVE To evaluate (1) whether alveolar macrophages are activated as a consequence of acute pancreatitis (AP), (2) the implication of inflammatory factors released by these macrophages in the process of neutrophil migration into the lungs observed in lung injury induced by AP, and (3) the role of the liver in the activation of alveolar macrophages. SUMMARY BACKGROUND DATA Acute lung injury is the extrapancreatic complication most frequently associated with death and complications in severe AP. Neutrophil infiltration into the lungs seems to be related to the release of systemic and local mediators. The liver and alveolar macrophages are sources of mediators that have been suggested to participate in the lung damage associated with AP. METHODS Pancreatitis was induced in rats by intraductal administration of 5% sodium taurocholate. The inflammatory process in the lung and the activation of alveolar macrophages were investigated in animals with and without portocaval shunting 3 hours after AP induction. Alveolar macrophages were obtained by bronchoalveolar lavage. The generation of nitric oxide, leukotriene B4, tumor necrosis factor-alpha, and MIP-2 by alveolar macrophages and the chemotactic activity of supernatants of cultured macrophages were evaluated. RESULTS Pancreatitis was associated with increased infiltration of neutrophils into the lungs 3 hours after induction. This effect was prevented by the portocaval shunt. Alveolar macrophages obtained after induction of pancreatitis generated increased levels of nitric oxide, tumor necrosis factor-alpha, and MIP-2, but not leukotriene B4. In addition, supernatants of these macrophages exhibited a chemotactic activity for neutrophils when instilled into the lungs of unmanipulated animals. All these effects were abolished when portocaval shunting was carried out before induction of pancreatitis. CONCLUSION Lung damage induced by experimental AP is associated with alveolar macrophage activation. The liver mediates the alveolar macrophage activation in this experimental model.

Endogenous nitric oxide and exogenous nitric oxide supplementation in hepatic ischemia-reperfusion injury in the rat.

BACKGROUND Although nitric oxide (NO) is thought to be beneficial in hepatic ischemia-reperfusion (I/R), the mechanisms for this effect are not well established. METHODS To investigate the effects of endogenous NO and exogenous NO supplementation on hepatic I/R injury and their pathogenic mechanisms, serum ALT and hyaluronic acid (endothelial cell damage), and hepatic malondialdehyde and H2O2 (oxidative stress), myeloperoxidase activity (leukocyte accumulation), and endothelin (vasoconstrictor peptide opposite to NO) were determined at different reperfusion periods in untreated rats and rats receiving L-NAME, L-NAME+L-arginine, and spermine NONOate (exogenous NO donor). RESULTS After reperfusion every parameter increased in untreated animals. Endogenous NO synthesis inhibition by L-NAME increased hepatocyte and endothelial damage as compared to untreated rats, which was reverted and even improved by the addition of L-arginine. Spermine NONOate also improved this damage. However, different mechanisms account for the beneficial effect of endogenous and exogenous NO. Oxidative stress decreased by both L-NAME and L-NAME+L-arginine, but remained unmodified by spermine NONOate. Myeloperoxidase increased by L-NAME and this effect was reverted by the addition of L-arginine, whereas no change was observed with spermine NONOate. Endothelin levels were not modified by L-NAME and L-NAME+L-arginine, but decreased with spermine NONOate. CONCLUSIONS These results suggest that, although both endogenous and exogenous NO exert a protective role in experimental hepatic I/R injury, the mechanisms of the beneficial effect of the two sources of NO are different.

Preservation of Steatotic Livers in IGL-1 Solution

A new Institut Georges Lopez (IGL‐1) solution was used to preserve steatotic livers. Steatotic (obese [Ob]) and nonsteatotic (lean [Ln]) livers from Zücker rats (n = 16, 8 Ln and 8 Ob) were preserved for 24 hours at 4°C in University of Wisconsin (UW) or IGL‐1 solution, respectively, and then perfused ex vivo for 2 hours at 37°C. Additionally, Ob and Ln livers (n = 16, 8 Ln and 8 Ob) were preserved in IGL‐1 plus Nω‐nitro‐L‐arginine methyl ester hydrochloride (L‐NAME). Hepatic injury and function (aminotransferases, bile production, bromosulfophthalein clearance), and factors potentially involved in the susceptibility of steatotic livers to ischemia‐reperfusion injury, such as oxidative stress, mitochondrial damage, and vascular resistance, were studied. Nitric oxide (NO) production and constitutive and inducible NO synthase were also measured. Steatotic and nonsteatotic livers preserved in IGL‐1 solution showed lower transaminases, malondialdehyde, glutamate dehydrogenase levels, and higher bile production than UW‐solution‐preserved livers. IGL‐1 solution protected against oxidative stress, mitochondrial damage and the alterations in vascular resistance associated with cold ischemia‐reperfusion. Thus, at the end of reperfusion period, aspartate aminotransferase levels in steatotic livers were 281 ± 6 U/L in UW vs. 202 ± 10 U/L in IGL‐1 solution. Glutamate dehydrogenase was 463 ± 75 U/L in UW vs. 111 ± 4 U/L in IGL‐1 solution, and oxidative stress was 3.0 ± 0.1 nmol/mg prot in UW vs. 2.0 ± 0.1 nmol/mg prot in IGL‐1 solution. These beneficial effects of IGL‐1 solution were abolished by the addition of L‐NAME, which implicates NO in the benefits of IGL‐1. In conclusion, IGL‐1 solution provided steatotic livers with better protection against the deleterious effects of cold ischemia‐reperfusion injury than did UW solution. Liver Transpl 12:1215‐1223, 2006.

Role of P-Selectin and ICAM-1 in Pancreatitis-Induced Lung Inflammation in Rats: Significance of Oxidative Stress

OBJECTIVE To investigate the role of P-selectin and intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of lung injury associated with pancreatitis, and the relation between xanthine oxidase-derived oxidants and expression of these adhesion molecules. SUMMARY BACKGROUND DATA In acute pancreatitis, acute respiratory distress syndrome occurs in the early stages of disease. This process is mediated by neutrophil infiltration. METHODS Pancreatitis was induced in rats by intraductal administration of 5% sodium taurocholate. ICAM-1 and P-selectin expression was measured using radiolabeled monoclonal antibodies. Neutrophil infiltration and plasma levels of xanthine oxidase were also evaluated. RESULTS Pancreatitis induces increases in P-selectin expression in lung, whereas ICAM-1 is unchanged from baseline levels. Immunoneutralization of either P-selectin or ICAM-1 prevents the infiltration of neutrophils into the lung. Xanthine and xanthine oxidase activity were increased after induction of pancreatitis. Xanthine oxidase inhibition prevents the upregulation of P-selectin in lung and neutrophil infiltration. CONCLUSIONS During acute pancreatitis, P-selectin is upregulated in the pulmonary endothelium and is a key determinant of leukocyte recruitment. Constitutive ICAM-1 is also involved in the process of cell infiltration into the lung. The increased expression of P-selectin appears to be triggered by a mechanism dependent on free radicals generated by xanthine oxidase released by the damaged pancreas.

Protective Effect of Ischemic Preconditioning on Cold Preservation and Reperfusion Injury Associated With Rat Intestinal Transplantation

ObjectiveTo define the protective effect of ischemic preconditioning on cold ischemia and reperfusion injury associated with intestinal transplantation, and the role of nitric oxide in this process. Summary Background DataIschemia/reperfusion injury continues to be a significant obstacle in small bowel transplantation. Preconditioning is a mechanism that protects against this injury. MethodsTo study the capacity of preconditioning to prevent cold ischemia-associated injury and the inflammatory response associated with intestinal transplantation, the authors studied a control group of animals, cold ischemia groups with or without previous preconditioning and with or without previous administration of L-NAME or NONOS, and intestinal transplantation groups with or without previous preconditioning and with or without previous administration of L-NAME or NONOS. ResultsHistologic findings and the release of lactate dehydrogenase into the preservation solution showed that preconditioning protects against cold ischemic preservation-associated injury. Preconditioning also prevented the inflammatory response associated with intestinal transplantation, measured by the above parameters and by neutrophil recruitment in the intestine. Inhibition of nitric oxide eliminates the protective effect. ConclusionsPreconditioning protects the intestinal grafts from cold preservation and reperfusion injury in the rat intestinal transplantation model. Nitric oxide is involved in this protection.

Is Ischemic Preconditioning a Useful Strategy in Steatotic Liver Transplantation

This study examined the effect of preconditioning on steatotic livers for transplantation and attempted to identify the underlying protective mechanisms. Blood flow alterations, neutrophil accumulation, tumor necrosis factor α release and lipid peroxidation were observed in nonsteatotic livers after transplantation. Steatotic and nonsteatotic liver grafts were similar in their blood flow, neutrophil accumulation, and TNF release after transplantation. However, in the presence of steatosis, lipid peroxidation and hepatic injury increased. In addition, recipients of steatotic liver grafts were more vulnerable to lung damage associated with transplantation. The conversion of xanthine dehydrogenase to xanthine oxidase and the accumulation of xanthine during cold ischemia was greater in steatotic than in nonsteatotic liver grafts. The results obtained with xanthine oxidase inhibitors indicated that xanthine/xanthine oxidase could be responsible for the increased lipid peroxidation as well as the exacerbated liver and lung damage associated with transplantation of steatotic livers. Preconditioning reduced the xanthine accumulation and percentage of xanthine oxidase seen in steatotic liver grafts during cold ischemia, and conferred protection against liver and lung damage following transplantation. The benefits of preconditioning could be mediated by nitric oxide. These findings suggest that preconditioning could be a relevant new strategy to protect against the inherent risk of steatotic liver failure following transplantation.

Modification of oxidative stress in response to intestinal preconditioning

Previous studies have demonstrated that intestinal preconditioning protects the organ from ischemia reperfusion damage. Xanthine oxidase mediating free radical generation contributes to the development of injury associated to ischemia reperfusion. Thus, any process able to modulate the oxygen free radical generation system could attenuate the injury. Also, it is known that nitric oxide is implicated in the preconditioning response. The aim of this work is to determine: (1) the effect of intestinal preconditioning on the xanthine oxidase system, (2) the relevance of this system in the development of injury, and (3) its relationship with nitric oxide. For this purpose, we have determined the activity of the xanthine dehydrogenase/xanthine oxidase system, the levels of its substrate (xanthine), and end-product (uric acid) and oxidant stress status in rat small intestine subjected to ischemic pre-conditioning. The effects of nitric oxide inhibition have also been evaluated. Results show that the percentage of xanthine dehydrogenase to xanthine oxidase conversion, xanthine, uric acid concentration, lipoperoxides, and reduced glutathione were significantly reduced in preconditioned rats irrespectively of nitric oxide inhibition. In summary, this work shows that oxidative stress in intestinal preconditioning is reduced as consequence of the diminished conversion of xanthine dehydrogenase to xanthine oxidase, and also as a consequence of the reduced availability of xanthine.