Carmen Peralta

Endoplasmic reticulum stress inhibition protects steatotic and non-steatotic livers in partial hepatectomy under ischemia-reperfusion

During partial hepatectomy, ischemia–reperfusion (I/R) is commonly applied in clinical practice to reduce blood flow. Steatotic livers show impaired regenerative response and reduced tolerance to hepatic injury. We examined the effects of tauroursodeoxycholic acid (TUDCA) and 4-phenyl butyric acid (PBA) in steatotic and non-steatotic livers during partial hepatectomy under I/R (PH+I/R). Their effects on the induction of unfolded protein response (UPR) and endoplasmic reticulum (ER) stress were also evaluated. We report that PBA, and especially TUDCA, reduced inflammation, apoptosis and necrosis, and improved liver regeneration in both liver types. Both compounds, especially TUDCA, protected both liver types against ER damage, as they reduced the activation of two of the three pathways of UPR (namely inositol-requiring enzyme and PKR-like ER kinase) and their target molecules caspase 12, c-Jun N-terminal kinase and C/EBP homologous protein-10. Only TUDCA, possibly mediated by extracellular signal-regulated kinase upregulation, inactivated glycogen synthase kinase-3β. This is turn, inactivated mitochondrial voltage-dependent anion channel, reduced cytochrome c release from the mitochondria and caspase 9 activation and protected both liver types against mitochondrial damage. These findings indicate that chemical chaperones, especially TUDCA, could protect steatotic and non-steatotic livers against injury and regeneration failure after PH+I/R.

Activation of peroxisome proliferator‐activated receptor‐α inhibits the injurious effects of adiponectin in rat steatotic liver undergoing ischemia–reperfusion

Hepatic steatosis is a major risk factor in ischemia–reperfusion (I/R). Adiponectin acts as an antiobesity and anti‐inflammatory hormone. Adiponectin activates peroxisome proliferator‐activated receptor‐α (PPAR‐α), a transcription factor that regulates inflammation in liver disease. Ischemic preconditioning (PC) based on brief periods of I/R protects steatotic livers against subsequent sustained I/R injury, but just how this is achieved is poorly understood. This study explains the role of PPAR‐α and adiponectin in the vulnerability shown by steatotic livers to I/R and the benefits of PC in this situation. PPAR‐α and adiponectin levels in nonsteatotic livers undergoing I/R were similar to those found in the sham group. However, reduced PPAR‐α and increased adiponectin levels, particularly the high molecular weight isoform, were observed in steatotic livers as a consequence of I/R. Our results suggest that mitogen‐activated protein kinases (MAPKs) may be positive regulators of adiponectin accumulation in steatotic livers. The addition of adiponectin small interfering RNA (siRNA) before I/R protected steatotic livers against oxidative stress and hepatic injury. The induction of PC before I/R increased PPAR‐α and reduced adiponectin levels in steatotic livers. PC, which increased PPAR‐α, as well as PPAR‐α agonist pretreatment reduced MAPK expression, adiponectin, oxidative stress, and hepatic injury that follows I/R. In addition, the administration of a PPAR‐α antagonist in preconditioned steatotic livers eliminated the beneficial effects of PC on MAPKs, adiponectin, oxidative stress, and hepatic injury. Conclusion: Steatotic livers are more predisposed to down‐regulate PPAR‐α and overexpress adiponectin when subjected to I/R. PPAR‐α agonists and adiponectin siRNA are promising candidates to protect steatotic livers. PPAR‐α agonists as well as PC, through PPAR‐α, inhibited MAPK expression following I/R. This in turn inhibited adiponectin accumulation in steatotic livers and adiponectin‐worsening effects on oxidative stress and hepatic injury. (HEPATOLOGY 2007.)

Addition of carvedilol to University Wisconsin solution improves rat steatotic and nonsteatotic liver preservation

Here we examine the effect of adding carvedilol (CVD) to University of Wisconsin (UW) solution on the preservation of steatotic and nonsteatotic livers during cold ischemia and after normothermic reperfusion. We used an isolated perfused rat liver model. The following protocols were evaluated. Protocol 1 concerned the effect of CVD after cold ischemia. Steatotic and nonsteatotic livers were preserved for 24 hours in UW solution alone or with CVD. Livers without cold ischemia were used as controls. Transaminases were evaluated in the flushing effluent. Protocol 2 involved the effect of CVD after reperfusion. Both liver types were preserved for 24 hours in UW solution alone or with CVD and then perfused ex vivo for 2 hours at 37°C. Livers flushed and perfused without ischemia were used as controls. Hepatic injury and functionality [transaminases, bile production, and hepatic clearance of sulfobromophthalein (BSP)] were evaluated after reperfusion. In addition, factors potentially involved in hepatic ischemia‐reperfusion injury, including oxidative stress (malondialdehyde and superoxide anion levels), mitochondrial damage (glutamate dehydrogenase activity), microcirculatory disorders (flow rate and vascular resistance), and adenosine triphosphate (ATP) depletion, were evaluated after reperfusion. After cold ischemia, steatotic livers preserved in UW solution showed higher transaminase levels than nonsteatotic livers. After reperfusion, steatotic livers preserved in UW solution showed higher transaminase levels and lower bile production and BSP clearance than nonsteatotic livers. Alterations in the perfusion flow rate and vascular resistance, mitochondrial damage, and reduced ATP content were more evident in steatotic livers preserved in UW solution. The addition of CVD to UW solution reduced hepatic injury, obstructed its mechanisms, and improved hepatic functionality in both liver types. We conclude that CVD is a useful additive for UW solution that improves the preservation of steatotic and nonsteatotic livers subjected to prolonged cold ischemia. Liver Transpl 16:163–171, 2010.

Therapeutic targets in liver transplantation: angiotensin II in nonsteatotic grafts and angiotensin-(1-7) in steatotic grafts.

Numerous steatotic livers are discarded as unsuitable for transplantation because of their poor tolerance of ischemia‐reperfusion(I/R). The injurious effects of angiotensin (Ang)‐II and the benefits of Ang‐(1–7) in various pathologies are well documented. We examined the generation of Ang II and Ang‐(1–7) in steatotic and nonsteatotic liver grafts from Zucker rats following transplantation. We also studied in both liver grafts the effects of Ang‐II receptors antagonists and Ang‐(1–7) receptor antagonists on hepatic I/R damage associated with transplantation. Nonsteatotic grafts showed higher Ang II levels than steatotic grafts, whereas steatotic grafts showed higher Ang‐(1–7) levels than nonsteatotic grafts. Ang II receptor antagonists protected only nonsteatotic grafts against damage, whereas Ang‐(1–7) receptor antagonists were effective only in steatotic grafts. The protection conferred by Ang II receptor antagonists in nonsteatotic grafts was associated with ERK 1/2 overexpression, whereas the beneficial effects of Ang‐(1–7) receptor antagonists in steatotic grafts may be mediated by NO inhibition. Our results show that Ang II receptor antagonists are effective only in nonsteatotic liver transplantation and point to a novel therapeutic target in liver transplantation based on Ang‐(1–7), which is specific for steatotic liver grafts.

Insulin-like growth factor and epidermal growth factor treatment: new approaches to protecting steatotic livers against ischemia-reperfusion injury.

Hepatic steatosis is a major risk factor in ischemia-reperfusion (I/R). IGF-binding proteins (IGFBPs) modulate IGF-I action by transporting circulating IGF-I to its sites of action. Epidermal growth factor (EGF) stimulates IGF-I synthesis in vitro. We examined the effect of IGF-I and EGF treatment, separately or in combination, on the vulnerability of steatotic livers to I/R. Our results indicated that I/R impaired IGF-I synthesis only in steatotic livers. Only when a high dose of IGF-I (400 microg/kg) was given to obese animals did they show high circulating IGF-I:IGFBP levels, increased hepatic IGF-I levels, and protection against damage. In lean animals, a dose of 100 microg/kg IGF-I protected nonsteatotic livers. Our results indicated that the combined administration of IGF-I and EGF resulted in hepatic injury parameters in both liver types similar to that obtained by IGF-I and EGF separately. IGF-I increased egf expression in both liver types. The beneficial role of EGF on hepatic I/R injury may be attributable to p38 inhibition in nonsteatotic livers and to PPAR gamma overexpression in steatotic livers. In conclusion, IGF-I and EGF may constitute new pharmacological strategies to reduce the inherent susceptibility of steatotic livers to I/R injury.

Matrix metalloproteinase 2 in reduced-size liver transplantation: beyond the matrix.

We studied the contribution of matrix metalloproteinase 2 (MMP2) and matrix metalloproteinase 9 (MMP9) to the beneficial effects of preconditioning (PC) in reduced‐size orthotopic liver transplantation (ROLT). We also examined the role of c‐Jun N‐terminal kinase (JNK) and whether it regulates MMP2 in these conditions. Animals were subjected to ROLT with or without PC and pharmacological modulation, and liver tissue samples were then analyzed. We found that MMP2, but notMMP9, is involved in the beneficial effects of PC in ROLT. MMP2 reduced hepatic injury and enhanced liver regeneration. Moreover, inhibition of MMP2 in PC reduced animal survival after transplantation. JNK inhibition in the PC group decreased hepatic injury and enhanced liver regeneration. Furthermore, JNK upregulated MMP2 in PC. In addition, we showed that Tissue inhibitors of matrix metalloproteinases 2 (TIMP2) was also upregulated in PC and that JNK modulation also altered its levels in ROLT and PC. Our results open up new possibilities for therapeutic treatments to reduce I/R injury and increase liver regeneration after ROLT, which are the main limitations in living‐donor transplantation.

Effect of Angiotensin II and Bradykinin Inhibition in Rat Reduced-Size Liver Transplantation

This study examined whether angiotensin II (Ang II) blockers [Ang II type I receptor antagonist, Ang II type II receptor antagonist, and angiotensin converting enzyme (ACE) inhibitor] could reduce hepatic injury and improve regeneration in reduced‐size orthotopic liver transplantation (ROLT) and whether the beneficial effects of ischemic preconditioning (PC) in ROLT could be explained by changes in Ang II. We show that small liver grafts generated Ang II after ROLT and that this was associated with increased angiotensinogen and ACE messenger RNA expression. Furthermore, inhibition of Ang II did not contribute to PC‐induced protection in ROLT. All Ang II blockers reduced hepatic injury, but none of them promoted liver regeneration. Bradykinin (BK) receptor antagonist improved liver regeneration but did not reduce hepatic injury in ROLT. Finally, the combination of Ang II blockers and BK receptor antagonists in ROLT reduced hepatic injury and improved liver regeneration. In conclusion, treatments with either Ang II blockers or BK receptor antagonists cannot, on their own, improve the outcome of ROLT. Although Ang II blockers can reduce hepatic ischemia‐reperfusion injury and BK receptor antagonists can promote liver regeneration, neither confers both benefits at the same time. Consequently, it may be of clinical interest to apply both treatments simultaneously. Liver Transpl 15:313–320, 2009.

New Insights Into Fatty Liver Preservation Using Institute Georges Lopez Preservation Solution

Institute Georges Lopez preservation solution (IGL-1) has been demonstrated to be useful for fatty liver preservation. The mechanisms responsible for this effective graft protection against ischemia-reperfusion injury are pivotal actions on generation of nitric oxide a diffusible molecule with vasodilator properties, that facilitates the up-regulation of other well-known cytoprotective genes, such as hypoxia-inducible factor-1 alpha (HIF-1alpha) and heme-oxygenase 1 (HO-1). During normoxic reperfusion, the presence of nitric oxide permits HIF-1alpha accumulation to inhibit prolyl-hydoxylases, thus promoting an additional overexpression of the HO-1 in steatotic and nonsteatotic graft livers preserved in IGL-1.

564 TLR4 INDUCTION BY TUDCA IN EXPERIMENTAL MODELS OF SYNGENEIC AND ALLOGENEIC STEATOTIC LIVER TRANSPLANTATION

3. Antioxidant capability of rMnSOD as a supplement of a commercially available preservation solution was evaluated in hepatic biopsies cold stored for transplantation. Results: 1. Cold storage induced a marked increase in O−2 levels and a decrease in NO bioavailability in SEC, those detrimental effects were abolished in cells preserved with rMnSOD. 2. In rats, administration of rMnSOD ameliorated hepatic injury and endothelial dysfunction derived from cold storage and warm reperfusion injuries. The beneficial effects of rMnSOD were associated with a reduction in hepatic O−2, ONOO − and inflammation together with an improved antioxidant activity and nitric oxide bioavailability. 3. rMnSOD added to a conventional preservation solution maintains its marked antioxidant activity avoiding oxidative stress formation in hepatic tissue preserved for transplantation. Conclusions: This study demonstrates that rMnSOD markedly improves liver viability and endothelial function after cold ischemia and warm reperfusion. rMnSOD represents a new therapeutic strategy to protect liver grafts undergoing transplantation.