Functional analysis of protein complex involved in neuropathic pain by proteomics

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Since human genome project is accomplished, proteomics that enables global analysis of proteins expressed in cells and tissues has been paid much attention at present. Primary afferent fibers are originated from pseudounipolar sensory neurons in dorsal root ganglia (DRG) and innervate polymodal receptors in the periphery, which are most effectively excited by noxious heat, chemical and mechanical stimuli, and transmit these impulses in the superficial dorsal horn of the spinal cord, an important site of pain processing. Nerve injury often causes intractable and chronic pain. Since neuronal plasticity has been shown to be an important component in the generation of neuropathic pain, changes in gene expression, protein expression and post-translational protein modification have been extensively studied in the DRG as well as the spinal cord. In order to elucidate the mechanisms of generation of neuropathic pain, here we attempted to identify functional molecules involved in 1) neural pola … More rity of axonal transport of proteins produced in the DRG through primary afferent fibers and 2) generation of neuropathic pain, especially NMDA receptor complex in the postsynaptic density (PSD) by use of proteomic technologies and obtained following results.1)We established large-sized two-dimensional gel electrophoresis (20 X 70 cm) and subsequent identification by MALDI-TOF-MS. We also established differential analysis of protein expression of two samples in a single gel by Ettan DIGE system.2)We found that 69 and 61 spots were at least 2-fold abundantly expressed in lumbar spinal nerve segments peripheral (P) and central (C) to the DRG respectively, among more than 800 protein spots visualized by silver staining. One of the unique spots in the P fraction was identified as an isoform of collapsin response mediator protein-2 (CRMP-2), which was decreased after nerve injury.3)We found that phosphorylation of NMDA receptor subtype NR2B increased in the PSD of spinal dorsal horn associated with neuropathic pain. At present, we have examined changes of components of NMDA receptor complex in spinal PSD of inflammatory and neuropathic pain model by proteomics. Less