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Repetitive Model of Mild Traumatic Brain Injury Produces Cortical Abnormalities Detectable by Magnetic Resonance Diffusion Imaging, Histopathology, and Behavior

View Article: PubMed Central - PubMed

ABSTRACT

Noninvasive detection of mild traumatic brain injury (mTBI) is important for evaluating acute through chronic effects of head injuries, particularly after repetitive impacts. To better detect abnormalities from mTBI, we performed longitudinal studies (baseline, 3, 6, and 42 days) using magnetic resonance diffusion tensor imaging (DTI) and diffusion kurtosis imaging (DKI) in adult mice after repetitive mTBI (r-mTBI; daily × 5) or sham procedure. This r-mTBI produced righting reflex delay and was first characterized in the corpus callosum to demonstrate low levels of axon damage, astrogliosis, and microglial activation, without microhemorrhages. High-resolution DTI-DKI was then combined with post-imaging pathological validation along with behavioral assessments targeted for the impact regions. In the corpus callosum, only DTI fractional anisotropy at 42 days showed significant change post-injury. Conversely, cortical regions under the impact site (M1–M2, anterior cingulate) had reduced axial diffusivity (AD) at all time points with a corresponding increase in axial kurtosis (Ka) at 6 days. Post-imaging neuropathology showed microglial activation in both the corpus callosum and cortex at 42 days after r-mTBI. Increased cortical microglial activation correlated with decreased cortical AD after r-mTBI (r = −0.853; n = 5). Using Thy1-YFP-16 mice to fluorescently label neuronal cell bodies and processes revealed low levels of axon damage in the cortex after r-mTBI. Finally, r-mTBI produced social deficits consistent with the function of this anterior cingulate region of cortex. Overall, vulnerability of cortical regions is demonstrated after mild repetitive injury, with underlying differences of DTI and DKI, microglial activation, and behavioral deficits.

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Post-imaging neuroinflammation in the corpus callosum and cortex after r-mTBI. Immunohistochemistry was used to detect microglia/macrophage cells with CD11b and astrocytes with GFAP in mice perfused after the final imaging scan, that is, 6 weeks post-injury or sham procedure. (A and B) Slight astrogliosis and microglia/macrophage activation were evident in r-mTBI mice (B) compared to r-sham mice (A). (C–H) Quantitative analysis shows a significant increase in microglial activation in the corpus callosum of r-mTBI mice relative to sham (C and D). Astrogliosis in the corpus callosum was not significantly increased after r-mTBI (E). In the cortex, the r-mTBI mice had a significant increase in the area of CD11b immunolabeling and in the number of activated microglia (F and G). Astrogliosis in the medial cortex was not significantly increased after r-mTBI (H). Dashed lines outline the corpus callosum. Values are mean ± standard error of the mean; n = 5; scale bar = 50 μm; *p < 0.05. DAPI, 4′,6-diamidino-2-phenylindole; GFAP, glial fibrillary acidic protein; r-mTBI, repetitive mild traumatic brain injury; r-sham, repetitive sham.
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f5: Post-imaging neuroinflammation in the corpus callosum and cortex after r-mTBI. Immunohistochemistry was used to detect microglia/macrophage cells with CD11b and astrocytes with GFAP in mice perfused after the final imaging scan, that is, 6 weeks post-injury or sham procedure. (A and B) Slight astrogliosis and microglia/macrophage activation were evident in r-mTBI mice (B) compared to r-sham mice (A). (C–H) Quantitative analysis shows a significant increase in microglial activation in the corpus callosum of r-mTBI mice relative to sham (C and D). Astrogliosis in the corpus callosum was not significantly increased after r-mTBI (E). In the cortex, the r-mTBI mice had a significant increase in the area of CD11b immunolabeling and in the number of activated microglia (F and G). Astrogliosis in the medial cortex was not significantly increased after r-mTBI (H). Dashed lines outline the corpus callosum. Values are mean ± standard error of the mean; n = 5; scale bar = 50 μm; *p < 0.05. DAPI, 4′,6-diamidino-2-phenylindole; GFAP, glial fibrillary acidic protein; r-mTBI, repetitive mild traumatic brain injury; r-sham, repetitive sham.

Mentions: After completing the final MRI scan at the 6-week time point, mice were perfused for post-imaging analysis of the underlying pathology in the corpus callosum and cortical ROIs from the DTI and DKI analysis. We first examined neuroinflammation using CD11b and GFAP for detection of microglia and astrocytes, respectively. The r-mTBI tissues from our longitudinal MRI analysis showed mild, but persistent, neuroinflammation in the corpus callosum (Fig. 5A,B). Specifically, in r-mTBI mice compared to r-sham mice, the percent area immunolabeled was increased for CD11b (Fig. 5C). Morphological classification of individual CD11b-labeled cells also showed that the number of activated microglia increased after r-mTBI (Fig. 5D). Amoeboid CD11b-labeled cells were not found in any condition or time point (data not shown). After r-mTBI, astrogliosis in the corpus callosum was not significantly different from the r-sham condition (Fig. 5E). Overall, in the r-mTBI mice, neuroinflammation in the corpus callosum was much less robust relative to the s-mTBI model, which was used as a positive control comparison for corpus callosum pathology detectable by DTI (Supplementary Figs. 2 and 3) (see online supplementary material at http://www.liebertpub.com). Neuroinflammation in medial cortical regions, that is, directly under the impact site, in r-mTBI mice, showed a significant increase in microglial activation, but not astrogliosis (Fig. 5F–H). In contrast, the single-injury model did not exhibit significant neuroinflammation in the cortex with matched analysis of immunoreactivity for either CD11b (Supplementary Fig. 3F,G) (see online supplementary material at http://www.liebertpub.com) or GFAP (Supplementary Fig. 3H) (see online supplementary material at http://www.liebertpub.com) and did not have cortical abnormalities for DTI or DKI analysis (Supplementary Fig. 2) (see online supplementary material at http://www.liebertpub.com).


Repetitive Model of Mild Traumatic Brain Injury Produces Cortical Abnormalities Detectable by Magnetic Resonance Diffusion Imaging, Histopathology, and Behavior
Post-imaging neuroinflammation in the corpus callosum and cortex after r-mTBI. Immunohistochemistry was used to detect microglia/macrophage cells with CD11b and astrocytes with GFAP in mice perfused after the final imaging scan, that is, 6 weeks post-injury or sham procedure. (A and B) Slight astrogliosis and microglia/macrophage activation were evident in r-mTBI mice (B) compared to r-sham mice (A). (C–H) Quantitative analysis shows a significant increase in microglial activation in the corpus callosum of r-mTBI mice relative to sham (C and D). Astrogliosis in the corpus callosum was not significantly increased after r-mTBI (E). In the cortex, the r-mTBI mice had a significant increase in the area of CD11b immunolabeling and in the number of activated microglia (F and G). Astrogliosis in the medial cortex was not significantly increased after r-mTBI (H). Dashed lines outline the corpus callosum. Values are mean ± standard error of the mean; n = 5; scale bar = 50 μm; *p < 0.05. DAPI, 4′,6-diamidino-2-phenylindole; GFAP, glial fibrillary acidic protein; r-mTBI, repetitive mild traumatic brain injury; r-sham, repetitive sham.
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f5: Post-imaging neuroinflammation in the corpus callosum and cortex after r-mTBI. Immunohistochemistry was used to detect microglia/macrophage cells with CD11b and astrocytes with GFAP in mice perfused after the final imaging scan, that is, 6 weeks post-injury or sham procedure. (A and B) Slight astrogliosis and microglia/macrophage activation were evident in r-mTBI mice (B) compared to r-sham mice (A). (C–H) Quantitative analysis shows a significant increase in microglial activation in the corpus callosum of r-mTBI mice relative to sham (C and D). Astrogliosis in the corpus callosum was not significantly increased after r-mTBI (E). In the cortex, the r-mTBI mice had a significant increase in the area of CD11b immunolabeling and in the number of activated microglia (F and G). Astrogliosis in the medial cortex was not significantly increased after r-mTBI (H). Dashed lines outline the corpus callosum. Values are mean ± standard error of the mean; n = 5; scale bar = 50 μm; *p < 0.05. DAPI, 4′,6-diamidino-2-phenylindole; GFAP, glial fibrillary acidic protein; r-mTBI, repetitive mild traumatic brain injury; r-sham, repetitive sham.
Mentions: After completing the final MRI scan at the 6-week time point, mice were perfused for post-imaging analysis of the underlying pathology in the corpus callosum and cortical ROIs from the DTI and DKI analysis. We first examined neuroinflammation using CD11b and GFAP for detection of microglia and astrocytes, respectively. The r-mTBI tissues from our longitudinal MRI analysis showed mild, but persistent, neuroinflammation in the corpus callosum (Fig. 5A,B). Specifically, in r-mTBI mice compared to r-sham mice, the percent area immunolabeled was increased for CD11b (Fig. 5C). Morphological classification of individual CD11b-labeled cells also showed that the number of activated microglia increased after r-mTBI (Fig. 5D). Amoeboid CD11b-labeled cells were not found in any condition or time point (data not shown). After r-mTBI, astrogliosis in the corpus callosum was not significantly different from the r-sham condition (Fig. 5E). Overall, in the r-mTBI mice, neuroinflammation in the corpus callosum was much less robust relative to the s-mTBI model, which was used as a positive control comparison for corpus callosum pathology detectable by DTI (Supplementary Figs. 2 and 3) (see online supplementary material at http://www.liebertpub.com). Neuroinflammation in medial cortical regions, that is, directly under the impact site, in r-mTBI mice, showed a significant increase in microglial activation, but not astrogliosis (Fig. 5F–H). In contrast, the single-injury model did not exhibit significant neuroinflammation in the cortex with matched analysis of immunoreactivity for either CD11b (Supplementary Fig. 3F,G) (see online supplementary material at http://www.liebertpub.com) or GFAP (Supplementary Fig. 3H) (see online supplementary material at http://www.liebertpub.com) and did not have cortical abnormalities for DTI or DKI analysis (Supplementary Fig. 2) (see online supplementary material at http://www.liebertpub.com).

View Article: PubMed Central - PubMed

ABSTRACT

Noninvasive detection of mild traumatic brain injury (mTBI) is important for evaluating acute through chronic effects of head injuries, particularly after repetitive impacts. To better detect abnormalities from mTBI, we performed longitudinal studies (baseline, 3, 6, and 42 days) using magnetic resonance diffusion tensor imaging (DTI) and diffusion kurtosis imaging (DKI) in adult mice after repetitive mTBI (r-mTBI; daily&thinsp;&times;&thinsp;5) or sham procedure. This r-mTBI produced righting reflex delay and was first characterized in the corpus callosum to demonstrate low levels of axon damage, astrogliosis, and microglial activation, without microhemorrhages. High-resolution DTI-DKI was then combined with post-imaging pathological validation along with behavioral assessments targeted for the impact regions. In the corpus callosum, only DTI fractional anisotropy at 42 days showed significant change post-injury. Conversely, cortical regions under the impact site (M1&ndash;M2, anterior cingulate) had reduced axial diffusivity (AD) at all time points with a corresponding increase in axial kurtosis (Ka) at 6 days. Post-imaging neuropathology showed microglial activation in both the corpus callosum and cortex at 42 days after r-mTBI. Increased cortical microglial activation correlated with decreased cortical AD after r-mTBI (r&thinsp;=&thinsp;&minus;0.853; n&thinsp;=&thinsp;5). Using Thy1-YFP-16 mice to fluorescently label neuronal cell bodies and processes revealed low levels of axon damage in the cortex after r-mTBI. Finally, r-mTBI produced social deficits consistent with the function of this anterior cingulate region of cortex. Overall, vulnerability of cortical regions is demonstrated after mild repetitive injury, with underlying differences of DTI and DKI, microglial activation, and behavioral deficits.

No MeSH data available.


Related in: MedlinePlus