PEG35 and Glutathione Improve Mitochondrial Function and Reduce Oxidative Stress in Cold Fatty Liver Graft Preservation

Abstract

The need to meet the demand for transplants entails the use of steatotic livers, more vulnerable to ischemia-reperfusion (IR) injury. Therefore, finding the optimal composition of static cold storage (SCS) preservation solutions is crucial. Given that ROS regulation is a therapeutic strategy for liver IR injury, we have added increasing concentrations of PEG35 and glutathione (GSH) to the preservation solutions (IGL-1 and IGL-2) and evaluated the possible protection against energy depletion and oxidative stress. Fatty livers from obese Zücker rats were isolated and randomly distributed in the control (Sham) preserved (24 h at 4 °C) in IGL-0 (without PEG35 and 3 mmol/L GSH), IGL-1 (1 g/L PEG35, and 3 mmol/L GSH), and IGL-2 (5 g/L PEG35 and 9 mmol/L GSH). Energy metabolites (ATP and succinate) and the expression of mitochondrial oxidative phosphorylation complexes (OXPHOS) were determined. Mitochondrial carrier uncoupling protein 2 (UCP2), PTEN-induced kinase 1 (PINK1), nuclear factor-erythroid 2 related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and the inflammasome (NLRP3) expressions were analyzed. As biomarkers of oxidative stress, protein oxidation (AOPP) and carbonylation (DNP derivatives), and lipid peroxidation (malondialdehyde (MDA)–thiobarbituric acid (TBA) adducts) were measured. In addition, the reduced and oxidized glutathione (GSH and GSSG) and enzymatic (Cu–Zn superoxide dismutase (SOD), CAT, GSH S-T, GSH-Px, and GSH-R) antioxidant capacities were determined. Our results showed that the cold preservation of fatty liver graft depleted ATP, accumulated succinate and increased oxidative stress. In contrast, the preservation with IGL-2 solution maintained ATP production, decreased succinate levels and increased OXPHOS complexes I and II, UCP2, and PINK-1 expression, therefore maintaining mitochondrial integrity. IGL-2 also protected against oxidative stress by increasing Nrf2 and HO-1 expression and GSH levels. Therefore, the presence of PEG35 in storage solutions may be a valuable option as an antioxidant agent for organ preservation in clinical transplantation.

Liver Graft Hypothermic Static and Oxygenated Perfusion (HOPE) Strategies: A Mitochondrial Crossroads

“…Mitochondrial aldehyde dehydrogenase-2 (ALDH2) is best known for its critical detoxifying role in liver alcohol metabolism, but there is growing evidence that it also plays a role in IRI through the prevention of oxidative stress processes (oxygen free radicals and toxic 4-hydroxy-nonenal generation) [ 20 , 43 ]. PEG35 in rinse [ 34 ] and IGL-1/IGL-2 solutions [ 33 , 44 ] was a determinant for increasing mitochondrial protection and function, as well as preventing lipid peroxidation when compared to identical solutions with or without PEG35 [ 44 ]. The high antioxidant capacity of IGL-2 was corroborated by MDA and AOPP when compared to livers preserved in UW and HTK ( Figure 4 ).…”

“…Although oxygen levels of the media embedding the organ (preservation solution) are very low, there is still an impairment of mitochondrial oxidative phosphorylation, leading to the generation of reactive oxygen species (ROS). In this sense, NO generation in IGL-1/IGL-2 solutions [ 33 , 38 ] could act, among many other functions, as an ROS scavenger in the lipoperoxidation process [ 44 ]. Interestingly, these involved cytoprotective mechanisms observed for PEG35 in IGL-1 and IGL-2 solutions were similar to those we proposed for liver ischemic preconditioning strategies in a rat model, where AMPK and NO are also well-known liver preconditioning cytoprotective factors [ 36 , 37 ].…”

“…[ 53 ]. This was one reason why we proposed increasing the content of glutathione in IGL-2 solution [ 44 ] vs. UW and IGL-1 [ 33 ]…”

“…In this line, the use of PEG35 would be “killing two birds with one stone”, avoiding red blood cell hyperaggregation and bringing the solution to more adequate parameters of viscosity in order not to damage the endothelial cells and their associated structures, such as glycocalyx, its application in the narrowed sinusoids of steatotic liver being especially interesting [ 16 , 45 ]. Furthermore, these benefits are not only applied at the physical level, as the use of IGL-2 confers higher mitochondrial protection and assures better conditions for the ALDH2 enzymatic function to prevent oxidative stress in hypothermic graft preservation [ 43 , 44 ].…”

Antifibrotic preventive effect of polyethylene glycol (PEG) 3350 in methotrexateinduced hepatoxicity model

“…Nowadays, many antioxidant agents are used due to their protective and therapeutic properties, and they can largely eliminate the risks of oxidative damage 7 . Polyethylene glycol (PEG) is a safe, affordable, widely available, and highly effective osmotic laxative that acts as a faecal cleaner to remove faecal nitrogen 8 . In addition to its laxative activity, PEG also has antioxidant activity 9 .…”