The failure of recent clinical trials in traumatic brain injury (TBI) demonstrates the critical need for more comprehensive preclinical studies. To determine the molecular mechanisms of selective vulnerability of the rat hippocampus, our recent studies have focused on the molecular determinants of neuronal injury and survival. The goal of the present proposal is to test the feasibility of using RNA interference (RNAi) in vivo to effectively knockdown TBI-induced gene expression in rat hippocampal neurons. We will use short interfering RNA (siRNA) in vivo to address the hypothesis that silencing of neuronal nitric oxide synthase (nNOS) expression will reduce the numbers of injured hippocampal neurons after TBI and, conversely, that silencing of glutathione peroxidase-1 (GPx-1) expression will increase the numbers of injured hippocampal neurons after TBI.
Specific Aim 1. To examine the role of nNOS in the pathophysiology of TBI, we will knockdown the expression of nNOS in rat hippocampus before TBI and quantify the influence on downstream gene expression and on numbers of injured neurons after TBI.
Specific Aim 2. To evaluate the consequences of silencing a gene that is implicated in endogenous neuroprotective responses, we will knockdown expression of GPx-1 in the rat hippocampus and quantify the influence on downstream gene expression and on numbers of injured neurons after TBI. We have previously shown that siRNA for the GPx-1 and nNOS genes reduces gene expression in rat hippocampal progenitor (HC2S2) cells. Although RNAi represents a powerful therapeutic possibility for human diseases, the therapeutic potential of RNAi has not been studied in animal models of TBI. Successful completion of these exploratory studies will help in guiding the design of future studies aimed at understanding the role of injury-induced genes after TBI and in guiding the therapeutic application of future studies of RNAi in brain injury and perhaps silencing of disease genes in other neurodegenerative disorders. ? ?
