ornithine-phenylacetate and Disease-Models--Animal

Hepatic encephalopathy (HE) is a debilitating neurological complication of cirrhosis. By definition, HE is considered a reversible disorder, and therefore HE should resolve following liver transplantation (LT). However, persisting neurological complications are observed in as many as 47% of LT recipients. LT is an invasive surgical procedure accompanied by various perioperative factors such as blood loss and hypotension which could influence outcomes post-LT. We hypothesize that minimal HE (MHE) renders the brain frail and susceptible to hypotension-induced neuronal cell death. Six-week bile duct-ligated (BDL) rats with MHE and respective SHAM-controls were used. Several degrees of hypotension (mean arterial pressure of 30, 60 and 90 mm Hg) were induced via blood withdrawal from the femoral artery and maintained for 120 min. Brains were collected for neuronal cell count and apoptotic analysis. In a separate group, BDL rats were treated for MHE with the ammonia-lowering strategy ornithine phenylacetate (OP; MNK-6105), administered orally (1 g/kg) for 3 weeks before induction of hypotension. Hypotension 30 and 60 mm Hg (not 90 mm Hg) significantly decreased neuronal marker expression (NeuN) and cresyl violet staining in the frontal cortex compared to respective hypotensive SHAM-operated controls as well as non-hypotensive BDL rats. Neuronal degeneration was associated with an increase in cleaved caspase-3, suggesting the mechanism of cell death was apoptotic. OP treatment attenuated hyperammonaemia, improved anxiety and activity, and protected the brain against hypotension-induced neuronal cell death. Our findings demonstrate that rats with chronic liver disease and MHE are more susceptible to hypotension-induced neuronal cell degeneration. This highlights MHE at the time of LT is a risk factor for poor neurological outcome post-transplant and that treating for MHE pre-LT might reduce this risk.

Topics: Ammonia; Animals; Antigens, Nuclear; Anxiety; Apoptosis; Behavior, Animal; Bile Ducts; Caspase 3; Cerebrovascular Circulation; Disease Models, Animal; Hepatic Encephalopathy; Hyperammonemia; Hypotension; Ligation; Male; Nerve Tissue Proteins; Neurodegenerative Diseases; Neurons; Ornithine; Rats; Rats, Sprague-Dawley

In liver failure, ammonia homeostasis is dependent upon the function of the ammonia metabolising enzymes, glutamine synthetase (GS) and glutaminase (GA) but data about their protein expression and activity are lacking. The aims of this study were to determine the protein expression and activity of GS and GA in individual organs in a rat model of chronic liver disease and to test whether the treatment with the ammonia-lowering agent ornithine phenylacetate (OP) modulates their activities.. 49 SD rats were studied 35 days after sham-operation or bile duct ligation (BDL). The BDL group received: L-ornithine (0.6 mg/kg/day), Phenylacetate (0.6 mg/kg/day), OP (0.6 mg/kg/day) or placebo (saline) for 5 days prior to sacrifice. Arterial ammonia, amino acids and liver biochemistry were measured. Expressions of GS and GA were determined by Western-blotting and activities by end-point methods in liver, muscle, gut, kidney, lung, and frontal cortex.. In BDL rats, hepatic GS enzyme activity was reduced by more than 80% compared to sham rats. Further, in BDL rats GA activity was reduced in liver but increased in the gut, muscle and frontal cortex compared to sham rats. OP treatment resulted in a reduction in hyperammonemia in BDL rats, associated with increased GS activity in the muscle and reduced gut GA activity.. In a rat model of chronic liver failure, hyperammonemia is associated with inadequate compensation by liver and muscle GS activity and increased gut GA activity. OP reduces plasma ammonia by increasing GS in the muscle and reducing GA activity in the gut providing additional insights into its mechanism of its action. GS and GA may serve as important future therapeutic targets for hyperammonemia in liver failure.

Topics: Ammonia; Animals; Bile Ducts; Disease Models, Animal; End Stage Liver Disease; Glutamate-Ammonia Ligase; Glutaminase; Ligation; Liver; Male; Ornithine; Rats; Rats, Sprague-Dawley

Glycine is an important ammoniagenic amino acid, which is increased in acute liver failure (ALF). We have previously shown that L-ornithine phenylacetate (OP) attenuates ammonia rise and intracranial pressure in pigs suffering from ALF but failed to demonstrate a stoichiometric relationship between change in plasma ammonia levels and excretion of phenylacetylglutamine in urine. The aim was to investigate the impact of OP treatment on the phenylacetylglycine pathway as an alternative and additional ammonia-lowering pathway. A well-validated and -characterized large porcine model of ALF (portacaval anastomosis, followed by hepatic artery ligation), which recapitulates the cardinal features of human ALF, was used. Twenty-four female pigs were randomized into three groups: (1) sham operated + vehicle, (2) ALF + vehicle, and (3) ALF + OP. There was a significant increase in arterial glycine concentration in ALF (P < 0.001 compared with sham), with a three-fold increase in glycine release into the systemic circulation from the kidney compared with the sham group. This increase was attenuated in both the blood and brain of the OP-treated animals (P < 0.001 and P < 0.05, respectively), and the attenuation was associated with renal removal of glycine through excretion of the conjugation product phenylacetylglycine in urine (ALF + vehicle: 1,060 ± 106 μmol/l; ALF + OP: 27,625 ± 2,670 μmol/l; P < 0.003). Data from this study provide solid evidence for the existence of a novel, additional pathway for ammonia removal in ALF, involving glycine production and removal, which is targeted by OP.

Topics: Ammonia; Animals; Biomarkers; Brain; Disease Models, Animal; Female; Glycine; Hyperammonemia; Kidney; Liver Failure, Acute; Ornithine; Random Allocation; Swine; Time Factors

Ornithine phenylacetate (OP) is a new drug that has been proposed for the treatment of hepatic encephalopathy (HE) because it decreases plasma ammonia. We performed a study to assess if OP would impact on neuronal function.. Motor-evoked potentials (MEP), a surrogate of hepatic encephalopathy, were assessed (without anesthesia) in rats with portacaval anastomosis (PCA) that received gastrointestinal blood (GIB). Rats were pre-treated with OP prior to GIB. Ammonia and related metabolites (plasma, urine, and brain microdialysis) were assessed by HPLC and mass spectroscopy.. OP (one dose or 3 days) prevented disturbances in MEP induced by GIB in PCA rats. In rats treated with OP for 3 days, the amplitude and latency of MEP remained stable (-1% and +1%), while in the control group the amplitude decreased -21% and the latency increased +12% (p<0.01). OP attenuated the rise of ammonia in plasma by 45%, ammonia in brain microdialysate by 48%, induced a faster glutamine rise and the appearance of phenylacetylglutamine in plasma and urine. In addition, OP was associated with a lower concentration of ammonia and glutamate in brain microdialysate (approx. 50%).. OP prevents abnormalities in MEP precipitated by GIB in a model of HE. This is probably due to the enhancement of glutamine synthesis and metabolism, which results in a lower rise of plasma ammonia and the prevention of changes in glutamate in microdialysate. Thus, OP may be a good drug to prevent HE precipitated by gastrointestinal bleeding.

Topics: Amino Acids; Ammonia; Animals; Brain; Disease Models, Animal; Evoked Potentials, Motor; Glutamine; Hepatic Encephalopathy; Male; Ornithine; Phenylacetates; Portacaval Shunt, Surgical; Rats; Rats, Sprague-Dawley