Traumatic brain injury coupled with secondary hypoxia leads to a mild endoplasmic reticulum stress response in the rat

Introduction

Traumatic brain injury (TBI) initiates a series of secondary injury cascades that participate in the pathogenesis of traumatic cell injury. These include electrophysiological, biochemical, and molecular events associated with excitotoxicity, inflammation and apoptosis. An immediate reaction by vulnerable and nonvulnerable brain regions to trauma is a cellular stress response. Genes involved in producing a variety of stress response proteins, including cytosolic, mitochondrial, and endoplasmic reticulum (ER) associated, may be rapidly induced or upregulated after injury. These genetic responses may also be exacerbated by secondary hypoxia, which represents a common secondary insult in TBI patients. In this study, specific stress genes were assessed by in situ hybridization after TBI both with and without a 30 min secondary hypoxic insult.

Methods

Male Sprague-Dawley rats were traumatized by moderate (1.8–2.2 atm) fluid-percussion brain injury. Experimental groups included sham and moderate TBI, with or without secondary hypoxia. Four and 24 h survival times were assessed. Brains were removed, cryosectioned, and hybridized to 35S labeled riboprobes for the following genes: HSP70, HSP60, GRP78, calnexin, and PDI.

Results

The cytosolic chaperone HSP70 was strongly induced by TBI in the ipsilateral cerebral cortex at 4 hours. Secondary hypoxia increased this expression to include areas of the ipsilateral hippocampus and thalamus, and the bilateral striatum. The mitochondrial membrane bound chaperone HSP60 was induced by TBI at a lower level in the ipsilateral outer cortical layers at 4–24 hours and was not affected by secondary hypoxia. GRP78, an endoplasmic reticulum stress response gene, was induced by TBI in the ipsilateral outer cortical layers, as well as in the dentate gyrus. Secondary hypoxia did not alter this expression. Calnexin and PDI, both constitutive ER protein folding mediators, were unaffected by TBI or hypoxia. Hypoxia alone failed to induce any of these stress genes in sham operated control rats.

Conclusions

The cytosolic response to stress caused by TBI is very strong and is augmented with the addition of secondary hypoxia. In contrast, the ER and mitochondrial stress response to TBI, even in the presence of secondary hypoxia, is comparatively mild. These findings indicate that distinct parts of the cell may respond to metabolic stresses and cerebral insults in unique ways and possibly by different mechanisms. Induction of either cytoplasmic stress genes or ER luminal stress genes reflects in which cellular compartments non-native proteins may be overproduced. This study suggests non-native proteins are overproduced mainly in the cytoplasm, and to a lesser degree, in the mitochondria and ER lumen after TBI. These new findings may help to clarify therapeutic targets to reduce the detrimental consequences of secondary insults after TBI.