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Effects of antioxidant treatment on blast-induced brain injury.

Du X, Ewert DL, Cheng W, West MB, Lu J, Li W, Floyd RA, Kopke RD - PLoS ONE (2013)

Bottom Line: The biomarkers examined included an oxidative stress marker (4-hydroxy-2-nonenal, 4-HNE), an immediate early gene (c-fos), a neural injury marker (glial fibrillary acidic protein, GFAP) and two axonal injury markers [amyloid beta (A4) precursor protein, APP, and 68 kDa neurofilament, NF-68].Antioxidant treatment reduced the following: 4-HNE in the hippocampus and the forceps major corpus callosum, c-fos expression in the retrosplenial cortex, GFAP expression in the dorsal cochlear nucleus (DCN), and APP and NF-68 expression in the hippocampus, auditory cortex, and MGN.This preliminary study indicates that antioxidant treatment may provide therapeutic protection to the central auditory pathway (the DCN and MGN) and the non-auditory central nervous system (hippocampus and retrosplenial cortex), suggesting that these compounds have the potential to simultaneously treat blast-induced injuries in the brain and auditory system.

View Article: PubMed Central - PubMed

Affiliation: Hough Ear Institute, Oklahoma City, Oklahoma, United States of America.

ABSTRACT
Blast-induced traumatic brain injury has dramatically increased in combat troops in today's military operations. We previously reported that antioxidant treatment can provide protection to the peripheral auditory end organ, the cochlea. In the present study, we examined biomarker expression in the brains of rats at different time points (3 hours to 21 days) after three successive 14 psi blast overpressure exposures to evaluate antioxidant treatment effects on blast-induced brain injury. Rats in the treatment groups received a combination of antioxidants (2,4-disulfonyl α-phenyl tertiary butyl nitrone and N-acetylcysteine) one hour after blast exposure and then twice a day for the following two days. The biomarkers examined included an oxidative stress marker (4-hydroxy-2-nonenal, 4-HNE), an immediate early gene (c-fos), a neural injury marker (glial fibrillary acidic protein, GFAP) and two axonal injury markers [amyloid beta (A4) precursor protein, APP, and 68 kDa neurofilament, NF-68]. The results demonstrate that blast exposure induced or up-regulated the following: 4-HNE production in the dorsal hippocampus commissure and the forceps major corpus callosum near the lateral ventricle; c-fos and GFAP expression in most regions of the brain, including the retrosplenial cortex, the hippocampus, the cochlear nucleus, and the inferior colliculus; and NF-68 and APP expression in the hippocampus, the auditory cortex, and the medial geniculate nucleus (MGN). Antioxidant treatment reduced the following: 4-HNE in the hippocampus and the forceps major corpus callosum, c-fos expression in the retrosplenial cortex, GFAP expression in the dorsal cochlear nucleus (DCN), and APP and NF-68 expression in the hippocampus, auditory cortex, and MGN. This preliminary study indicates that antioxidant treatment may provide therapeutic protection to the central auditory pathway (the DCN and MGN) and the non-auditory central nervous system (hippocampus and retrosplenial cortex), suggesting that these compounds have the potential to simultaneously treat blast-induced injuries in the brain and auditory system.

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Examples of c-Fos immunostaining images obtained from the granular RC of the NC (A), 3H-B, (B), and 3H-B/T (C) groups by light microscopy.Few c-fos-positive cells were observed in layer three in the cortex of the NC group (A). Numerous c-fos-positive stained cells were seen in layer three in the cortex of the 3H-B group (B), and decreased numbers of c-fos-positive cells were seen in the cortex of the 3H-B/T group relative to the 3H-B group (C). C-fos-positive cells in the RC were quantified and statistically analyzed (D). Four to six sections (8-12 images) from each rat brain (6 rats in each group) were used in this analysis. Significantly increased numbers of c-fos-positive cells were found in the 3H-B and 7D-B groups compared to the NC group (all p < 0.001). Significant differences were also found between the 3H-B and 3H-B/T groups (p < 0.001), suggesting that antioxidant treatment suppressed c-fos upregulation in the cortex at this time point after blast exposure. However, no significant difference was found between the 7D-B and 7D-B/T groups (p > 0.05). C-fos-positive cells in the DCN were quantified and statistically analyzed (E). Significantly increased numbers of c-fos-positive cells were found in the lateral region of the DCN in the 3H-B and 3H-B/T groups compared to the NC group (all p < 0.05), however no antioxidant treatment effect was observed in the DCN at this time point (all p > 0.05). ### indicates p < 0.001 compared to normal controls. *** indicate p < 0.001 compared to the blast only group. Error bars represent standard error of the means. Scale bar in C = 500 µm for A-C.
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pone-0080138-g002: Examples of c-Fos immunostaining images obtained from the granular RC of the NC (A), 3H-B, (B), and 3H-B/T (C) groups by light microscopy.Few c-fos-positive cells were observed in layer three in the cortex of the NC group (A). Numerous c-fos-positive stained cells were seen in layer three in the cortex of the 3H-B group (B), and decreased numbers of c-fos-positive cells were seen in the cortex of the 3H-B/T group relative to the 3H-B group (C). C-fos-positive cells in the RC were quantified and statistically analyzed (D). Four to six sections (8-12 images) from each rat brain (6 rats in each group) were used in this analysis. Significantly increased numbers of c-fos-positive cells were found in the 3H-B and 7D-B groups compared to the NC group (all p < 0.001). Significant differences were also found between the 3H-B and 3H-B/T groups (p < 0.001), suggesting that antioxidant treatment suppressed c-fos upregulation in the cortex at this time point after blast exposure. However, no significant difference was found between the 7D-B and 7D-B/T groups (p > 0.05). C-fos-positive cells in the DCN were quantified and statistically analyzed (E). Significantly increased numbers of c-fos-positive cells were found in the lateral region of the DCN in the 3H-B and 3H-B/T groups compared to the NC group (all p < 0.05), however no antioxidant treatment effect was observed in the DCN at this time point (all p > 0.05). ### indicates p < 0.001 compared to normal controls. *** indicate p < 0.001 compared to the blast only group. Error bars represent standard error of the means. Scale bar in C = 500 µm for A-C.

Mentions: C-fos is an immediate-early gene that is widely used as a marker for neuronal activity. Exposure to impulse noise has been shown to induce prolonged c-fos expression in the auditory cortex (AC) and the DCN [65], as well as in the cerebral cortex, the thalamus, and the hippocampus [66]. To determine whether our bTBI model resulted in induced c-fos expression, brain sections from normal controls and blast-exposed animals were immunostained with a c-fos antibody and subjected to comparative histological analyses. Few c-fos-positive cells were observed in the cortex (the RC and AC, Figure 2A), the DCN, the IC, and the hippocampus of normal control brains. In comparison, the degree of c-fos-positive immunostaining was strikingly higher in the RC of the brain in blast-exposed animals at three hours post-bTBI compared to normal controls (Figure 2B, and 2D, p < 0.05 or 0.01). We found that antioxidant treatment significantly attenuated c-fos expression in the RC in blast-exposed animals over this same time course (p < 0.001, F (8, 584) = 27.93, Figure 2C and 2D). Second peaks of RC c-fos expression were observed 7 days after blast exposure in both blast-exposed (p < 0.001) and blast-exposed plus antioxidant treatment (p < 0.01) groups when compared to the NC group (Figure 2D). However, no treatment effect was observed at this time point (p > 0.05). In contrast to the bTBI effects observed in the RC on c-fos expression, the observed density of c-fos-positive cells in the AC did not reflect significant induction of this immediate-early biomarker in this region of the brain 3 hours following blast-exposure (p > 0.05, Table S1).


Effects of antioxidant treatment on blast-induced brain injury.

Du X, Ewert DL, Cheng W, West MB, Lu J, Li W, Floyd RA, Kopke RD - PLoS ONE (2013)

Examples of c-Fos immunostaining images obtained from the granular RC of the NC (A), 3H-B, (B), and 3H-B/T (C) groups by light microscopy.Few c-fos-positive cells were observed in layer three in the cortex of the NC group (A). Numerous c-fos-positive stained cells were seen in layer three in the cortex of the 3H-B group (B), and decreased numbers of c-fos-positive cells were seen in the cortex of the 3H-B/T group relative to the 3H-B group (C). C-fos-positive cells in the RC were quantified and statistically analyzed (D). Four to six sections (8-12 images) from each rat brain (6 rats in each group) were used in this analysis. Significantly increased numbers of c-fos-positive cells were found in the 3H-B and 7D-B groups compared to the NC group (all p < 0.001). Significant differences were also found between the 3H-B and 3H-B/T groups (p < 0.001), suggesting that antioxidant treatment suppressed c-fos upregulation in the cortex at this time point after blast exposure. However, no significant difference was found between the 7D-B and 7D-B/T groups (p > 0.05). C-fos-positive cells in the DCN were quantified and statistically analyzed (E). Significantly increased numbers of c-fos-positive cells were found in the lateral region of the DCN in the 3H-B and 3H-B/T groups compared to the NC group (all p < 0.05), however no antioxidant treatment effect was observed in the DCN at this time point (all p > 0.05). ### indicates p < 0.001 compared to normal controls. *** indicate p < 0.001 compared to the blast only group. Error bars represent standard error of the means. Scale bar in C = 500 µm for A-C.
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Related In: Results  -  Collection

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pone-0080138-g002: Examples of c-Fos immunostaining images obtained from the granular RC of the NC (A), 3H-B, (B), and 3H-B/T (C) groups by light microscopy.Few c-fos-positive cells were observed in layer three in the cortex of the NC group (A). Numerous c-fos-positive stained cells were seen in layer three in the cortex of the 3H-B group (B), and decreased numbers of c-fos-positive cells were seen in the cortex of the 3H-B/T group relative to the 3H-B group (C). C-fos-positive cells in the RC were quantified and statistically analyzed (D). Four to six sections (8-12 images) from each rat brain (6 rats in each group) were used in this analysis. Significantly increased numbers of c-fos-positive cells were found in the 3H-B and 7D-B groups compared to the NC group (all p < 0.001). Significant differences were also found between the 3H-B and 3H-B/T groups (p < 0.001), suggesting that antioxidant treatment suppressed c-fos upregulation in the cortex at this time point after blast exposure. However, no significant difference was found between the 7D-B and 7D-B/T groups (p > 0.05). C-fos-positive cells in the DCN were quantified and statistically analyzed (E). Significantly increased numbers of c-fos-positive cells were found in the lateral region of the DCN in the 3H-B and 3H-B/T groups compared to the NC group (all p < 0.05), however no antioxidant treatment effect was observed in the DCN at this time point (all p > 0.05). ### indicates p < 0.001 compared to normal controls. *** indicate p < 0.001 compared to the blast only group. Error bars represent standard error of the means. Scale bar in C = 500 µm for A-C.
Mentions: C-fos is an immediate-early gene that is widely used as a marker for neuronal activity. Exposure to impulse noise has been shown to induce prolonged c-fos expression in the auditory cortex (AC) and the DCN [65], as well as in the cerebral cortex, the thalamus, and the hippocampus [66]. To determine whether our bTBI model resulted in induced c-fos expression, brain sections from normal controls and blast-exposed animals were immunostained with a c-fos antibody and subjected to comparative histological analyses. Few c-fos-positive cells were observed in the cortex (the RC and AC, Figure 2A), the DCN, the IC, and the hippocampus of normal control brains. In comparison, the degree of c-fos-positive immunostaining was strikingly higher in the RC of the brain in blast-exposed animals at three hours post-bTBI compared to normal controls (Figure 2B, and 2D, p < 0.05 or 0.01). We found that antioxidant treatment significantly attenuated c-fos expression in the RC in blast-exposed animals over this same time course (p < 0.001, F (8, 584) = 27.93, Figure 2C and 2D). Second peaks of RC c-fos expression were observed 7 days after blast exposure in both blast-exposed (p < 0.001) and blast-exposed plus antioxidant treatment (p < 0.01) groups when compared to the NC group (Figure 2D). However, no treatment effect was observed at this time point (p > 0.05). In contrast to the bTBI effects observed in the RC on c-fos expression, the observed density of c-fos-positive cells in the AC did not reflect significant induction of this immediate-early biomarker in this region of the brain 3 hours following blast-exposure (p > 0.05, Table S1).

Bottom Line: The biomarkers examined included an oxidative stress marker (4-hydroxy-2-nonenal, 4-HNE), an immediate early gene (c-fos), a neural injury marker (glial fibrillary acidic protein, GFAP) and two axonal injury markers [amyloid beta (A4) precursor protein, APP, and 68 kDa neurofilament, NF-68].Antioxidant treatment reduced the following: 4-HNE in the hippocampus and the forceps major corpus callosum, c-fos expression in the retrosplenial cortex, GFAP expression in the dorsal cochlear nucleus (DCN), and APP and NF-68 expression in the hippocampus, auditory cortex, and MGN.This preliminary study indicates that antioxidant treatment may provide therapeutic protection to the central auditory pathway (the DCN and MGN) and the non-auditory central nervous system (hippocampus and retrosplenial cortex), suggesting that these compounds have the potential to simultaneously treat blast-induced injuries in the brain and auditory system.

View Article: PubMed Central - PubMed

Affiliation: Hough Ear Institute, Oklahoma City, Oklahoma, United States of America.

ABSTRACT
Blast-induced traumatic brain injury has dramatically increased in combat troops in today's military operations. We previously reported that antioxidant treatment can provide protection to the peripheral auditory end organ, the cochlea. In the present study, we examined biomarker expression in the brains of rats at different time points (3 hours to 21 days) after three successive 14 psi blast overpressure exposures to evaluate antioxidant treatment effects on blast-induced brain injury. Rats in the treatment groups received a combination of antioxidants (2,4-disulfonyl α-phenyl tertiary butyl nitrone and N-acetylcysteine) one hour after blast exposure and then twice a day for the following two days. The biomarkers examined included an oxidative stress marker (4-hydroxy-2-nonenal, 4-HNE), an immediate early gene (c-fos), a neural injury marker (glial fibrillary acidic protein, GFAP) and two axonal injury markers [amyloid beta (A4) precursor protein, APP, and 68 kDa neurofilament, NF-68]. The results demonstrate that blast exposure induced or up-regulated the following: 4-HNE production in the dorsal hippocampus commissure and the forceps major corpus callosum near the lateral ventricle; c-fos and GFAP expression in most regions of the brain, including the retrosplenial cortex, the hippocampus, the cochlear nucleus, and the inferior colliculus; and NF-68 and APP expression in the hippocampus, the auditory cortex, and the medial geniculate nucleus (MGN). Antioxidant treatment reduced the following: 4-HNE in the hippocampus and the forceps major corpus callosum, c-fos expression in the retrosplenial cortex, GFAP expression in the dorsal cochlear nucleus (DCN), and APP and NF-68 expression in the hippocampus, auditory cortex, and MGN. This preliminary study indicates that antioxidant treatment may provide therapeutic protection to the central auditory pathway (the DCN and MGN) and the non-auditory central nervous system (hippocampus and retrosplenial cortex), suggesting that these compounds have the potential to simultaneously treat blast-induced injuries in the brain and auditory system.

Show MeSH
Related in: MedlinePlus