Diffuse Axonal Injury Rats Research Articles

Annexin A1 conveys neuroprotective function via inhibiting oxidative stress in diffuse axonal injury of rats.

Diffuse axonal injury (DAI) is a critical pathological facet of traumatic brain injury (TBI). Oxidative stress plays a significant role in the progress of DAI. Annexin A1 (AnxA1) has been demonstrated to benefit from recovery of neurofunctional outcomes after TBI. However, whether AnxA1 exhibits neuronal protective function by modulating oxidative stress in DAI remains unknown. Expression of AnxA1 was evaluated via real-time PCR and western blotting in rat brainstem after DAI. The neurological effect of AnxA1 following DAI through quantification of modified neurologic severity score (mNSS) was compared between wild-type and AnxA1-knockout rats. Brain edema and neuronal apoptosis, as well as expression of oxidative factors and inflammatory cytokines, were analyzed between wild-type and AnxA1 deficiency rats after DAI. Furthermore, mNSS, oxidative and inflammatory cytokines were assayed after timely administration of recombinant AnxA1 for DAI rats. In the brainstem of DAI, the expression of AnxA1 remarkably increased. Ablation of AnxA1 increased the mNSS score and brain water content of rats after DAI. Neuron apoptosis in the brainstem after DAI was exaggerated by AnxA1 deficiency. In addition, AnxA1 deficiency significantly upregulated the level of oxidative and inflammatory factors in the brainstem of DAI rats. Moreover, mNSS decreased by AnxA1 treatment in rats following DAI. Expression of oxidative and inflammatory molecules in rat brainstem subjected to DAI inhibited by AnxA1 administration. AnxA1 exhibited neuronal protective function in the progression of DAI mainly dependent on suppressing oxidative stress and inflammation.

Ketogenic diet protects myelin and axons in diffuse axonal injury

ABSTRACT Background Ketogenic diet (KD) has been identified as a potential therapy to enhance recovery after traumatic brain injury (TBI). Diffuse axonal injury (DAI) is a common type of traumatic brain injury that is characterized by delayed axonal disconnection. Previous studies showed that demyelination resulting from oligodendrocyte damage contributes to axonal degeneration in DAI. Aim The present study tests a hypothesis that ketone bodies from the ketogenic diet confers protection for myelin and attenuates degeneration of demyelinated axon in DAI. Methods A modified Marmarou’s model of DAI was induced in adult rats. The DAI rats were fed with KD and analyzed with western blot, transmission electron microscope, ELISA test and immunohistochemistry. Meanwhile, a co-culture of primary oligodendrocytes and neurons was treated with ketone body β-hydroxybutryate (βHB) to test for its effects on the myelin-axon unit. Results Here we report that rats fed with KD showed an increased fatty acid metabolism and ketonemia. This dietary intervention significantly reduced demyelination and attenuated axonal damage in rats following DAI, likely through inhibition of DAI-induced excessive mitochondrial fission and promoting mitochondrial fusion. In an in vitro model of myelination, the ketone body βHB increased myelination significantly and reduced axonal degeneration induced by glucose deprivation (GD). βHB robustly increased cell viability, inhibited GD-induced collapse of mitochondrial membrane potential and attenuated death of oligodendrocytes. Conclusion Ketone bodies protect myelin-forming oligodendrocytes and reduce axonal damage. Ketogenic diet maybe a promising therapy for DAI.

Inhibition of Macrophage Migration Inhibitory Factor Protects against Inflammation through a Toll-like Receptor-Related Pathway after Diffuse Axonal Injury in Rats.

Objective We have previously demonstrated that inflammation induced by toll-like receptors (TLRs) 2/4 exert cerebral deleterious effects after diffuse axonal injury (DAI); however, the underlying mechanisms are not fully understood. Macrophage migration inhibitory factor (MIF) is a multifunctional cytokine involved in inflammatory responses. The purpose of this study was to investigate the role of MIF in inflammation induced by TLRs in the cortices of DAI rats. Methods The rat DAI model was established by head rotational acceleration and confirmed by β-APP, HE, and silver staining. MIF protein expression at 3 h, 6 h, 12 h, 1 d, and 3 d after DAI was measured by western blot. The localization of MIF was measured by immunofluorescence. MIF antagonist ISO-1 was intracerebroventricularly injected to inhibit MIF. Neuronal and axonal injury and glial responses were assessed by TUNEL, immunohistochemistry, and TEM. Expression of TLR2, TLR4, ERK, phospho-ERK, NF-κB, and phospho-NF-κB was examined by western blot. The level of IL-1β, IL-6, and TNF-α was measured by ELISA. Results MIF expression was significantly increased, peaking at 1 day after DAI, and MIF was mainly localized in microglial cells and neurons. ISO-1 suppressed neuronal apoptosis, axonal injury, and glial responses and decreased the expression of downstream signaling molecules related to TLR2/4, including ERK, phospho-ERK, NF-κB, phospho-NF-κB, IL-1β, IL-6, and TNF-α. Conclusion MIF was involved in the neuronal and axonal damage through a TLR-related pathway following DAI.

Serum levels of tau protein increase according to the severity of the injury in DAI rat model.

Traumatic brain injury (TBI) in the form of diffuse axonal injury (DAI) is difficult to diagnose in the early phase of the injury. Early diagnosis of DAI may provide opportunity for developing treatment and management strategies. Tau protein has been demonstrated to increase in the early phase of TBI with high diagnostic accuracy in patients with DAI. We tested the biological plausibility of tau protein using a rat DAI model by evaluating the association between serum tau levels and the severity of brain injury.DAI was induced in animals using the Marmarou model.After a survival of 60 minutes, rats were anesthetized and sacrificed after obtaining blood samples (5ml) from the heart. Eighteenratswereemployedin the present studyand were randomly subjected to sham-operated control (n=4), mild DAI (n=7), and severe DAI (n=7). Of seven severe DAI rats, two rats that had focal injury caused by skull fracture were excluded in the measurement of tau protein level. The serum levels of tau protein in the rat DAI model were found to increase significantly and consistently according to the severity of the injury.Rats with DAI showed significantly higher serum levels of tau protein compared to sham rats; the severe DAI rats had higher levels of tau than moderate DAI and sham rats (sham vs. mild, P=0.02; mild vs. severe, P=0.02). In conclusion, serum tau protein levels may be useful as a biomarker for diagnosing and estimating the severity of DAI in the early phase.

Effect of α7nAChR on learning and memory dysfunction in a rat model of diffuse axonal injury

Diffuse axonal injury (DAI) is the predominant effect of severe traumatic brain injury and significantly contributes to cognitive deficits. The mechanisms that underlie these cognitive deficits are often associated with complex molecular alterations. α7nAChR, one of the abundant and widespread nicotinic acetylcholine receptors (nAChRs) in the brain, plays important physiological functions in the central nervous system. However, the relationship between temporospatial alterations in the α7nAChR and DAI-related learning and memory dysfunction are not completely understood. Our study detected temporospatial alterations of α7nAChR in vulnerable areas (hippocampus, internal capsule, corpus callosum and brain stem) of DAI rats and evaluated the development and progression of learning and memory dysfunction via the Morris water maze (MWM). We determined that α7nAChR expression in vulnerable areas was mainly reduced at the recovery of DAI in rats. Moreover, the escape latency of the injured group increased significantly and the percentages of the distance travelled and time spent in the target quadrant were significantly decreased after DAI. Furthermore, α7nAChR expression in the vulnerable area was significantly positively correlated with MWM performance after DAI according to regression analysis. In addition, we determined that a selective α7nAChR agonist significantly improved learning and memory dysfunction. Rats in the α7nAChR agonist group showed better learning and memory performance than those in the antagonist group. These results demonstrate that microstructural injury-induced alterations of α7nAChR in the vulnerable area are significantly correlated with learning and memory dysfunctions after DAI and that augmentation of the α7nAChR level by its agonist contributes to the improvement of learning and memory function.

Sulforaphane administration alleviates diffuse axonal injury (DAI) via regulation signaling pathway of NRF2 and HO-1.

Nuclear factor erythroid 2-related factor 2 (Nrf2) can alleviate diffuse axonal injury (DAI)-induced apoptosis by regulating expression of heme oxygenase-1 (HO-1), while sulforaphane (SFN) was shown to reduce oxidative stress by increasing the expression of Nrf2. Therefore, we aimed to investigate therapeutic effect of SFN in the treatment of DAI and the ability of SFN to reduce oxidative stress. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to observe the effects of H2 O 2 and SFN on cell viability. Fluorometric assay, Western blot analysis, and flow cytometry were conducted to validate the protective role of SFN in an animal model of DAI. In addition, the levels of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GPx) were measured in DAI rats treated by SFN, while Western blot, immunohistochemistry assay, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay were carried out to verify the effect of SFN in different animal groups. Cell viability was reduced by H2 O 2 in a dose-dependent manner, while the treatment by SFN significantly promoted cell growth. Meanwhile the administration of SFN effectively reduced the levels of caspase-3/poly(ADP-ribose) polymerase (PARP) activity increased by the H 2 O 2 treatment, indicating that the protective effect of SFN could be mediated by its ability to suppress caspase-3 activation and PARP cleavage. In addition, the SFN treatment reduced the intracellular reactive oxygen species (ROS) generation induced by H 2 O 2 . Moreover, the MDA levels of SOD/GPx activity in various rat groups showed the protective effects of SFN in DAI rats. It is suspected that the protective effect of SFN was exerted via the activation of the Nrf2/HO-1 signaling pathway. In this study, DAI and DAI + phosphate-buffered saline (PBS) groups also showed the presence of more TUNEL-positive cells compared with the sham-operated group, while the SFN treatment reduced the extent of neuronal apoptosis. By activating the Nrf2/HO-1 signaling pathway and reducing the activity of caspase-3, SFN reduces the apoptosis of neurons in brain trauma-induced DAI.

Microstructural and Neurochemical Changes in the Rat Brain After Diffuse Axonal Injury.

Diffuse axonal injury (DAI) is one of the devastating types of traumatic brain injury, but is difficult to detect on conventional imaging in its early stages. To test the technical feasibility and diagnostic value of diffusion kurtosis imaging (DKI) and glutamate chemical exchange saturation transfer (GluCEST) imaging in the brain after DAI. Prospective. Sixty Sprague-Dawley rats. The DAI model was induced by using the impact acceleration model of Marmarou et al with modified settings. A 7.0T animal MR scanner with a fast spin-echo sequence (T2 -weighted imaging), fast spin-echo multislice sequence (DKI), echo planar imaging in the signal of the chemical exchange saturation transfer sequence (CEST), and point-resolved spectroscopy sequence (hydrogenproton magnetic resonance spectroscopy, 1 H-MRS). Brain MRI scanned before and 2 hours after injury. DKI images were processed with MatLab and MRIcro software, GluCEST images were processed using software routines written in MatLab, and spectroscopic data were postprocessed with LCModel. The parameters of these techniques were assessed using the independent sample t-test and Pearson correlation. Mean kurtosis and mean diffusivity values were significantly higher than controls in the parietal lobe, hippocampus, and thalamus (P < 0.01). However, fractional anisotropy was lower only in the parietal lobe, with no detectable changes in the hippocampus and thalamus. GluCEST values of the parietal lobe, hippocampus, and thalamus were significantly higher than controls in DAI rats (P < 0.01). This change was further validated through 1 H-MRS. A positive correlation was observed between glutamate (Glu) and glutamate compound (Glx) concentrations and GluCEST values (Glu: R2 = 0.589, Glx: R2 = 0.878). DKI and GluCEST might be acceptably sensitive for tracking microstructural and neurochemical changes in the brain following DAI. 1 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;49:1069-1077.

Neuroprotective effect of berberine against learning and memory deficits in diffuse axonal injury.

The aim of the present study was to assess the neuroprotective effect of berberine against learning and memory deficits in diffuse axonal injury (DAI). DAI rats were orally gavaged with berberine at a dose of 200 mg/kg of body weight for 4 weeks. Behavioral tests were used to analyze the neuroprotective effect of berberine against DAI-induced learning and memory deficits. In the present study, treatment with berberine significantly protected against DAI-induced inhibition of learning and memory in rats. Notably, berberine significantly suppressed the levels of tumor necrosis factor, interleukin-1β and monocyte chemoattractant protein-1, as well as reduced the protein expression levels of nuclear factor-κB, Bcl-2-associated X protein and cytochrome c in DAI rats. In addition, berberine significantly suppressed the protein expression of p38 mitogen-activated protein kinase, activating transcription factor 2 and vascular endothelial growth factor in DAI rats. These results suggested that berberine exhibited a neuroprotective effect against learning and memory deficits in severe DAI through the suppression of inflammation, angiogenesis and apoptosis in a rat model.

Involvement of Toll Like Receptor 2 Signaling in Secondary Injury during Experimental Diffuse Axonal Injury in Rats.

Treatment of diffuse axonal injury (DAI) remains challenging in clinical practice due to the unclear pathophysiological mechanism. Uncontrolled, excessive inflammation is one of the most recognized mechanisms that contribute to the secondary injury after DAI. Toll like receptor 2 (TLR2) is highlighted for the initiation of a vicious self-propagating inflammatory circle. However, the role and detailed mechanism of TLR2 in secondary injury is yet mostly unknown. In this study, we demonstrated the expression of TLR2 levels in cortex, corpus callosum, and internal capsule and the localization of TLR2 in neurons and glial cells in rat DAI models. Intracerebral knockdown of TLR2 significantly downregulated TLR2 expression, attenuated cortical apoptosis, lessened glial response, and reduced the secondary axonal and neuronal injury in the cortex by inhibiting phosphorylation of mitogen-activated protein kinases (MAPK) including Erk, JNK, and p38, translocation of NF-κB p65 from the cytoplasm to the nucleus, and decreasing levels of proinflammatory cytokines including interleukin-6, interleukin-1β, and tumor necrosis factor-α. On the contrary, administration of TLR2 agonist to DAI rats achieved an opposite effect. Collectively, we demonstrated that TLR2 was involved in mediating secondary injury after DAI by inducing inflammation via the MAPK and NF-κB pathways.

Inhibition of TLR4 Signalling-Induced Inflammation Attenuates Secondary Injury after Diffuse Axonal Injury in Rats

Increasing evidence suggests that secondary injury after diffuse axonal injury (DAI) damages more axons than the initial insult, but the underlying mechanisms of this phenomenon are not fully understood. Recent studies show that toll-like receptor 4 (TLR4) plays a critical role in promoting adaptive immune responses and have been shown to be associated with brain damage. The purpose of this study was to investigate the role of the TLR4 signalling pathway in secondary axonal injury in the cortices of DAI rats. TLR4 was mainly localized in microglial cells and neurons, and the levels of TLR4 downstream signalling molecules, including TLR4, myeloid differentiation primary response gene 88, toll/IR-1-(TIR-) domain-containing adaptor protein inducing interferon-beta, interferon regulatory factor 3, interferon β, nuclear factor κB (NF-κB) p65, and phospho-NF-κB p65, significantly increased and peaked at 1 d after DAI. Inhibition of TLR4 by TAK-242 attenuated apoptosis, neuronal and axonal injury, and glial responses. The neuroprotective effects of TLR4 inhibition were associated with decreases in the levels of TLR4 downstream signalling molecules and inflammatory factors, including interleukin-1β, interleukin-6, and tumour necrosis factor-α. These results suggest that the TLR4 signalling pathway plays an important role in secondary injury and may be an important therapeutic target following DAI.

Cognitive dysfunction and histological findings in rats with chronic-stage contusion and diffuse axonal injury

The Morris water maze (MWM) technique is well known as a prominent method of evaluating learning acquisition and memory retention impairments in rats. We previously reported on a modified fluid percussion device that is able to consistently produce experimental cortical contusion (CC) and diffuse axonal injury (DAI) in separate groups of rats. The purpose of the present protocol is to evaluate the differences in learning acquisition and memory retention impairments between these two types of injured rats in the chronic stage using the MWM technique. CC and DAI rats are respectively induced by lateral and midline fluid percussion. We also compare the histological differences between these two different types of traumatic brain injury. The results show statistically significant differences in learning acquisition impairment between the sham and CC rats and between the sham and DAI rats. However, a difference in memory retention impairment was expected to be seen only between the sham and DAI rats. Histologically, the loss of CA3 pyramidal cells in the hippocampus was observed ipsilaterally in the CC and bilaterally in DAI. Neuronal cell loss was observed in bilaterally in layer II of the entorhinal cortex in DAI, but not in CC. Themes: Neural basis of behavior Topics: Learning and memory: systems and functions

Evaluation of learning and memory dysfunction and histological findings in rats with chronic stage contusion and diffuse axonal injury

We previously reported a modified fluid percussion device capable of consistently producing experimental cortical contusion (CC) and diffuse axonal injury (DAI) in separate groups of rats by lateral and midline fluid percussion, respectively. The purpose of the present study was to compare the differences in learning acquisition and memory retention impairments between these two types of injured rats in the chronic stage using the Morris water maze technique. We also compared the histological differences between these two different types of traumatic brain injury. The results showed a statistically significant difference in learning acquisition impairment between the sham and CC rats and also between the sham and DAI rats. However, a significant difference in memory retention impairment was observed only between the sham and DAI rats. Histologically, the neuronal cell loss of CA3 pyramidal cells in the hippocampus was observed on the ipsilateral side in the CC and bilaterally in DAI. The neuronal cell loss was seen in bilateral entorhinal cortex layer II in DAI, but it was not seen in CC. From these results, we speculate that the marked cell loss in the hippocampus CA3 region in both CC and DAI rats was related to the impairment of spatial learning acquisition. The marked cell loss in entorhinal cortex layer II in DAI rats may be one of the important factors in the impairment of spatial memory retention.