A density-based proteomics sample fractionation technology: folate deficiency induced oxidative stress response in...

Folate deficiency (FD) alters hepatic methionine metabolism and is associated with increased hepatocellular apoptosis. Additionally, mice deprived of folate showed increased oxidative damage in brain tissue, leading to cognitive impairment. Most previous studies have focused independently on either liver, the main tissue of folate storage and metabolism, or brain, where folate regulates neurogenesis and programs cell death. The aim of this study was to apply a powerful, rapid proteomics approach to understand potential subcellular correlations of folate deficiency in both brain and liver of the same rat. This approach combined a new density-based sample fractionation technology (enhanced density gradient extraction = Edge technology) with other conventional proteomics techniques, such as western blot analysis, 2DE, and mass spectrometry. The brain and the liver from individual rats, fed normal or FD diets for 6 wks, were homogenized and then fractionated using the Edge 200 Separation System. Subsequently, all fractions from brain and liver, from control and treated rats, were analyzed by western blot using two markers of oxidative stress: glutathione peroxidase 1 (GPx1) and glucose-regulated protein 75 (GRP75). certain fractions were selected based on western blot analysis and were further analyzed by 2DE. protein spots of interest were identified by MALDI-TOF/TOF. The results demonstrated that edge technology provides a powerful density based separation and enrichment method for rapid screening of potential FD markers and their possible correlations to both liver and brain diseases.