Hydrogen Protects Against Liver Damage In Rats With Ischemia-Reperfusion

Background and Aims:Liver ischemia/reperfusion (I/R) injury is a frequently observed condition caused by restoring blood supply of liver after hepatic ischemia which involved a series of pathophysiological processes, such as radical generation, neutrophil infiltration and release of inflammatory mediators.Liver surgery often requires clamping of the portal triad to reduce intraoperative blood loss, and is inevitable to cause liver I/R injury which can increase postoperative liver dysfunction and even liver failure. Experimental and clinical studies have demonstrated the pivotal role of oxidative stress associated with over-production of ROS, which may cause lipid peroxidation and inflammatory injury in the promotion of hepatic injury after ischemia/reperfusion.Molecular hydrogen (H2) possesses antioxidative effects through selective reduction of the levels of hydroxyl radical. Previous studies have demonstrated that H2in the form of gas or dissolved in water can suppress tissue injuries in brain, liver, heart and intestine caused by oxidative stress following ischemia-reperfusion. Hydrogen also suppresses the inflammation induced in tissue-destructive diseases such as colitis, hepatitis, and transplantation-induced intestinal graft injury.Hydrogen-rich saline provides a high concentration of hydrogen that can be easily and safely applied. We here aim to investigate the effects of hydrogen-rich saline on the prevention of liver and mitochondria injury induced by liver ischemia/reperfusion in rats.Methods:1. We established a rat model of liver ischemia-reperfusion, and collected blood and liver samples180min after reperfusion.2. Liver function was measured using an autoanalyzer.3. HE staining was used to observe the change of liver morphology and neutrophils accumulation.4. Hepatic MDA, SOD, and GSH levels were assessed spectrophotometrically to investigate the degree of oxidizing reaction in liver. 5. Levels of TNF-α, IL-1β, IL-6, and HMGB1were measured with a commercial ELISA kit.6. We separated mitochondria and cytosol by differential centrifugation7. We observed hepatocyte mitochondrial injury by transmission electron microscope.8. Mitochondrial MDA, GSH, and GSSG levels were assessed by spectrophotometry.9. ATP content was measured with Sigma ATP bioluminescent assay kit based on the luciferin-luciferase method by a chemiluminescence analyzer. Oxygen consumption of isolated mitochondria was determined polarographically with a Clark oxygen electrode.Results:1. HS treatment significantly reduced I/R-induced liver damage.1) Administration of5ml/kg of HS considerably suppressed the release of ALT and AST from the liver, compared with NS-treated rats.2) Moderate hepatic cell edema, necrosis, and neutrophil cell infiltration were seen in HS treated groups and the increased ductular proliferation observed after I/R wassignificantly diminished after hydrogen treatment.2. HS ameliorated I/R-induced liver damage by reducing oxidative stress and inflammatory reaction in liver tissue.1) HS treatment dramatically suppressed the production of MDA in rats with OJ. Treatment with HS markedly increased the hepatic SOD and GSH activities in I/R rats2) When liver ischemia was followed by HS treatment, hepatic TNF-a, IL-1β and IL-6concentrations were significantly reduced compared with NS group. Treatment with HS significantly attenuated HMGB1levels in the liver of I/R rats.3. HS reduced I/R-induced mitochondrial damage and oxidizing reaction1) HS dramatically decreased the level of mitochondrial MDA and GSSG in rats with liver I/R and markedly increased the mitochondrial GSH content in I/R rats2) ATP content was significantly prevented in the rats treated with HS. HS promoted a significant increase of RCR and P/O compared to rats treated by NS. Conclusions:1. Hydrogen-rich saline attenuates I/R-induced liver damage.2. Hydrogen-rich saline inhibits I/R-induced liver oxidative stress.3. Hydrogen-rich saline attenuates I/R-induced liver inflammation.4. Hydrogen-rich saline attenuates I/R-induced liver mitochondria oxidative stress5. Hydrogen-rich saline can inhibit hepatocyte mitochondrial ATP consumption, breath control and p/o ratio to protect liver.Hydrogen-rich saline attenuates I/R-induced liver damage, possibly by reduction of inflammation and oxidative stress. Hydrogen-rich saline can also reduce I/R-induced liver mitochondria injury.