Dynamic Alterations Of Six SCIRR Proteins, Arrestin, Dynein, Dynactin, Tubulin, Glypican, And Vimentin, And Their MRNA, In The Injuried Rat Spinal Cords At Different Times, Identified By Western Blot And RT-PCR

Spinal cord injury affects people of all ages and can result in severe damage, leading to paraplegia, tetraplegia and death. The long-established view that central nervous system (CNS) neurons are inherently incapable of regeneration is no longer tenable. There is growing evidence supporting that adult brain and spinal cord possess the potential ability to regeneration [1, 2]. Both intrinsic neuronal and extrinsic environmental factors contribute to the lack of regeneration that is normally observed. The changes of incalculable proteins and genes happened during the spinal cord injury and regeneration (SCIRR). Lately several interesting molecules have been discovered to inhibit axon growth, such as the chondroitin sulfate proteoglycans, semaphorins, MAG and Nogo[3]. However, quite a lot of unknown molecules involving in the progress remain to be discovered. On the basis of the model of rat spinal cord transection injured, we made a research about compared proteomics with improved proteomic technology. Due to the complexity and extensive protein-protein and protein-lipid interactions, proteins in the CNS tissues are extraordinarily stable and resistant to isolation. So we employed the sequential extraction strategy for sample preparation: spinal cord tissue was first extracted in Tris buffer solution; the pellets were resuspended in an enhanced solution. Fractionation treatment facilitates the isolation of insoluble and low abundance proteins. phenotyping was accomplished using 2-DE; and image analysis was performed with ImageMaster 2-D Elite software. And we saw at least thirty one proteins in the injured spinal cord were shown up-regulated at folds. On this foundation we select six proteins among the up-regulated proteins, and investigated the dynamic alterations of 6 SCIRR related proteins and their corresponding mRNA after spinal cord injured with Western blot and RT-PCR. Although the exact mechanisms of up-regulated proteins what we select in spinal cord injury are not clear at present, they may involve in various processes including metabolic and stress responsive changes, neural degeneration and regeneration, axonal outgrowth and synaptic plasticity. Young adult rats were performed either mere thoracic laminectomy or spinal cord transection after laminectomy. And the comparative analysis existed in control, transection injured 1day, 3days and 5days. As a result, proteins Arrestin, Dynein, Dynactin and Tubulin in the injured spinal cord were shown up-regulated. And in the level of mRNA, the changes were either equal with that of protein or not notable. The dynamics of the genes of dynein and dynactin changed line-by-line. The expression of the protein Glypican in injured 1, 3days decreased. But in injured 5 days, the increasing of its quantity was remarkable. Simultaneously the dynamic alteration of glypican's mRNA was accordance with that of its protein. Namely, the expression fell obviously in injured 3 day, and moved up in injured 5 day. We also observed that the dynamic alteration of protein Vimentin in SCI was more than it in normal spinal cord. But in injury 1 day, the expression is the lowest. At the same time, the changes of its mRNA expression were equal with that of its protein. We think these dynamic alterations were related with spinal cord injury. In the other hand, we hope this study can lay the foundation for deeper researches. In a word, based on high-throughput proteome techniques, we reported the dynamics of six proteins and mRNA revolved in spinal cord injured and...