ABSTRACT
INTRODUCTION AND AIM: Hepatic encephalopathy (HE), caused by hyperammonemia resulting from liver disease, is a spectrum of neuropsychiatric and motor disorders that can lead to death. Existing therapies are deficient and alternative treatments are needed. We have shown that gene therapy with a baculovirus vector containing the glutamine synthetase (Bac-GS) gene is efficient for reducing ammonia levels in an acute hyperammonemia rat model. However, the most common condition resulting from liver disease is chronic hyperammonemia. In this work, Bac-GS was evaluated in bile-duct ligated rats, a chronic liver disease model with hyperammonemia and some characteristics of Type C HE. MATERIAL AND METHODS: Bac-GS was tested for mediating GS overexpression in HeLa cells and H9C2 myotubes. For determining the utility of Bac-GS for the reduction of ammonia levels in a chronic hyperammonemia animal model, four groups of rats were treated: control, sham, ligated with Bac-GS and ligated with Bac-GFP. Baculoviruses were injected i.m. 18 days post-surgery. Blood was drawn 2, 3 and 4 weeks post-surgery and plasma ammonia concentrations were quantified. RESULTS: In protein lysates of cells and myotubes transduced with Bac-GS, a 44 kDa band corresponding to GS was detected. Significant results were obtained in the hyperammonemic bile-duct ligated rat model, as plasma ammonia was reduced to normal levels 3 days after treatment with Bac-GS. Furthermore, a transitory effect of Bac-GS was observed. CONCLUSION: Our results show that gene therapy by delivering GS is a promising alternative for treatment of hyperammonemia in acute-on-chronic liver failure patients with HE.
Subject(s)
Baculoviridae/genetics , Genetic Therapy/methods , Hepatic Encephalopathy/etiology , Hepatic Encephalopathy/therapy , Hyperammonemia/complications , Analysis of Variance , Animals , Blotting, Western , Cells, Cultured , Chronic Disease , Disease Models, Animal , Genetic Vectors , Glutamate-Ammonia Ligase/administration & dosage , HeLa Cells/cytology , HeLa Cells/pathology , Hepatic Encephalopathy/pathology , Humans , Hyperammonemia/physiopathology , Random Allocation , Rats , Risk Factors , Sensitivity and SpecificityABSTRACT
Particulate matter may promote cardiovascular disease, possibly as a consequence of its oxidative potential. Studies using susceptible animals indicate that particulate matter aggravates atherosclerosis by increasing lipid/macrophage content in plaques. Macrophage lipid uptake requires oxidized low-density lipoprotein and scavenger receptors; same receptors are involved in particulate matter uptake. We studied in vitro particulate matter potential to oxidize low-density lipoproteins and subsequent cell uptake through scavenger receptors. Particulate matter-induced low-density lipoproteins oxidation was evaluated by the thiobarbituric acid assay. Binding/internalization was tested in wild type and scavenger receptor-transfected Chinese hamster ovary cells, and in RAW264.7 cells using fluorescently labeled low-density lipoproteins. Dose-dependent binding/internalization only occurred in scavenger receptor-transfected Chinese hamster ovary cells and RAW264.7 cells. Competition binding/internalization using particles showed that particulate matter induced decreased binding (â¼50%) and internalization (â¼70%) of particle-oxidized low-density lipoproteins and native low-density lipoproteins. Results indicate that particulate matter was capable of oxidizing low-density lipoproteins, favoring macrophage internalization, and also altered scavenger and low-density lipoproteins receptor function.
