Optimization of Quantitative Proteomics Using 2-Dimensional Difference Gel Electrophoresis to Characterize Molecular Mechanisms of Chemical Warfare Nerve Agent Exposure in the Rat Brain

摘要

This report describes the optimization and use of 2-dimensional difference gel electrophoresis (2-D DIGE) to investigate the proteomic changes induced by chemical warfare nerve agents (CWNAs) in whole cell homogenates and isolated mitochondria from rat brain regions. Rats were pretreated with the oxime HI-6, exposed to a seizurogenic dose of the CWNA soman, and then administered atropine methyl nitrate (AMN). Control animals were given HI-6, an equivalent volume of agent vehicle (saline), and AMN. Brain tissue was dissected into anatomical regions and either homogenized or processed for mitochondrial isolation. Proteins were fluorescently labeled, separated in two dimensions, and analyzed to quantify changes in protein abundance. Increases and decreases in protein abundance, as well as changes in protein isomer form, were observed in response to soman exposure, HI-6/AMN treatment, and/or soman exposure in the presence of HI- 6/AMN treatment. The proteomics approach described in this report in combination with mass spectrometry has recently been used to identify proteins and pathways altered by exposure to CWNA.