Article
Early changes of gene expression in the perifocal brain tissue after traumatic injury
Untersuchungen zur Genexpression im perifokalen Gewebe nach kortikalem Schädel-Hirn-Trauma
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Published: | May 8, 2006 |
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Objective: To study the alterations of gene expression in a model of cortical traumatic brain injury. We focussed on the viable perifocal tissue to gain further insight into the mechanisms involved in the development of secondary brain damage.
Methods: A cortical brain lesion of moderate size was produced by applying a cold probe (4mm Ų, -68°C, 1 min) onto the brain of male Sprague-Dawley rats. The extent of cell death was observed by injection of propidium iodide (PI; 0.5%, before and 6h after lesion) and the spread of edema by injection of fluorescein (5%, 2h before sacrifice). In-situ perfusion was performed 12, 24, or 48 h after trauma followed by brain isolation and serial slicing (thickness, 300 µm). These slices were used for microdissection of the viable perifocal tissue (PI negative, fluorescein positive). Total RNA was extracted and subjected to reverse transcription and preparation of cRNA using biotinylated nucleotides (cytosine and uracil) for gene chip analysis (Affymetrix RAE-230A containing 13115 genes).
Results: The level of significance was set at p<0.05 corresponding to a cutoff value of 10-6,571 with Bonferroni adjustment for multiple testing. The numbers of genes significantly regulated were 1237 (12h; 875 named, thereof 537 upregulated), 1208 (24h; 825 named, thereof 521 upregulated), and 1429 (48h; 961 named, thereof 619 upregulated). The dendrogram analysis revealed the largest degree of similarity between 12 and 24 h after trauma while the control tissue differed markedly from trauma at all time points. Pathway analysis using KEGG pathways showed most significant upregulation of genes to be related to cell cycling and nucleotide synthesis via MAP kinase cascades, to steroid formation, and at later time points to extracellular matrix proteins and associated membrane proteins.
Conclusions: A moderate cortical trauma leads to marked alterations of gene expression in the perifocal viable tissue which were initially related to the regulation of intracellular signalling, cell cycling, and nucleotide metabolism. Stimulation of the steroid pathway suggests inhibition of local pro-inflammatory responses. These alterations may well relate to limit the spread of secondary brain damage.