Proteomics of the Injured Rat Sciatic Nerve Reveals Protein Expression Dynamics During Regeneration

Ka Wan Li,Fred De Winter,Joost Verhaagen,Martin P. Hornshaw,Yvonne Gouwenberg,Floor J. Stam,August B. Smit,Connie R. Jiménez

doi:10.1074/mcp.m400076-mcp200

Abstract

Using proteomics, we investigated the temporal expression profiles of proteins in rat sciatic nerve after experimental crush. Extracts of sciatic nerves collected at 5, 10, and 35 days after injury were analyzed by two-dimensional gel electrophoresis and quantitative image analysis. Of the approximately 1,500 protein spots resolved on each gel, 121 showed significant regulation during at least one time point. Using cluster analysis, these proteins were grouped into two expression profiles of down-regulation and four of up-regulation. These profiles mainly reflected differences in cellular origins in addition to different functional roles. Mass spectrometric analysis identified 82 proteins pertaining to several functional classes, i.e. acute-phase proteins, antioxidant proteins, and proteins involved in protein synthesis/maturation/degradation, cytoskeletal (re)organization, and in lipid metabolism. Several proteins not previously implicated in nerve regeneration were identified, e.g. translationally controlled tumor protein, annexin A9/31, vitamin D-binding protein, alpha-crystallin B, alpha-synuclein, dimethylargininases, and reticulocalbin. Real-time PCR analysis of selected genes showed which were expressed in the nerve versus the dorsal root ganglion neurons. In conclusion, this study highlights the complexity and temporal aspect of the molecular process underlying nerve regeneration and points to the importance of glial and inflammatory determinants.

Highlights

Using proteomics, we investigated the temporal expression profiles of proteins in rat sciatic nerve after experimental crush
The injury-induced processes include Wallerian degeneration/demyelination in the first 2 weeks [30], Schwann cell proliferation peaking at day 4 after crush [17], macrophage infiltration starting at 3– 4 days and peaking at 7 days after crush [17, 30], axonal regeneration, and remyelination starting 1–2 weeks after nerve crush [17]
We investigated the effect of peripheral nerve injury on the global protein expression patterns in sciatic nerve at 5, 10, and 35 days after injury

Summary

Introduction

We investigated the temporal expression profiles of proteins in rat sciatic nerve after experimental crush. Of the ϳ1,500 protein spots resolved on each gel, 121 showed significant regulation during at least one time point Using cluster analysis, these proteins were grouped into two expression profiles of down-regulation and four of up-regulation. To study the molecular mechanisms that govern regeneration of the sciatic nerve, global gene expression profiling of dorsal root ganglia (DRG) and sciatic nerve has been employed recently These studies revealed that the regenerationassociated genes can be categorized into several groups, including cytoskeletal proteins, neurotransmitter metabolizing enzymes, neuropeptides, growth factors, and signal transduction molecules in the DRG [11,12,13,14], as well as genes encoding proteins involved in inflammation, proliferation, and myelination in the nerve [15,16,17]

Methods

Results

Conclusion