Pharmacological inhibition of lipid peroxidation attenuates calpain-mediated cytoskeletal degradation after traumatic brain injury

Free radical-induced lipid peroxidation (LP) is critical in the evolution of secondary injury following traumatic brain injury (TBI). Previous studies in our laboratory demonstrated that U-83836E, a potent LP inhibitor, can reduce post-TBI LP along with an improved maintenance of mouse cortical mitochondrial bioenergetics and calcium (Ca(2+)) buffering following severe (1.0 mm; 3.5 m/s) controlled cortical impact TBI (CCI-TBI). Based upon this preservation of a major Ca(2+) homeostatic mechanism, we have now performed dose-response and therapeutic window analyses of the ability of U-83836E to reduce post-traumatic calpain-mediated cytoskeletal (α-spectrin) proteolysis in ipsilateral cortical homogenates at its 24 h post-TBI peak. In the dose-response analysis, mice were treated with a single i.v. dose of vehicle or U-83836E (0.1, 0.3, 1.3, 3.0, 10.0 or 30.0 mg/kg) at 15 min after injury. U-83836E produced a dose-related attenuation of α-spectrin degradation with the maximal decrease being achieved at 3.0 mg/kg. Next, the therapeutic window was tested by delaying the single 3 mg/kg i.v. dose from 15 min post-injury out to 1, 3, 6 or 12 h. No reduction in α-spectrin degradation was observed when the treatment delay was 1 h or longer. However, in a third experiment, we re-examined the window with repeated U-83836E dosing (3.0 mg/kg i.v. followed by 10 mg/kg i.p. maintenance doses at 1 and 3 h after the initial i.v. dose) which significantly reduced 24 h α-α-spectrin degradation even when treatment initiation was withheld until 12 h post-TBI. These results demonstrate the relationship between post-TBI LP, disruptions in neuronal Ca(2+) homeostasis and calpain-mediated cytoskeletal damage.     

Modulation of autophagy in traumatic brain injury

Traumatic brain injury (TBI) is defined as a traumatically induced structural injury or physiological disruption of brain function as a result of external forces, leading to adult disability and death. A growing body of evidence reveals that alterations in autophagy-related proteins exist extensively in both experimentally and clinically after TBI. Of note, the autophagy pathway plays an essential role in pathophysiological processes, such as oxidative stress, inflammatory response, and apoptosis, thus contributing to neurological properties of TBI. With this in mind, this review summarizes a comprehensive overview on the beneficial and detrimental effects of autophagy in pathophysiological conditions and how these activities are linked to the pathogenesis of TBI. Moreover, the relationship between oxidative stress, inflammation, apoptosis, and autophagy occur TBI. Ultimately, multiple compounds and various drugs targeting the autophagy pathway are well described in TBI. Therefore, autophagy flux represents a potential clinical therapeutic value for the treatment of TBI and its complications.     

Dimethyl fumarate treatment after traumatic brain injury prevents depletion of antioxidative brain glutathione and confers neuroprotection

Dimethyl fumarate (DMF) is an immunomodulatory compound to treat multiple sclerosis and psoriasis with neuroprotective potential. Its mechanism of action involves activation of the antioxidant pathway regulator Nuclear factor erythroid 2‐related factor 2 thereby increasing synthesis of the cellular antioxidant glutathione (GSH). The objective of this study was to investigate whether post‐traumatic DMF treatment is beneficial after experimental traumatic brain injury (TBI). Adult C57Bl/6 mice were subjected to controlled cortical impact followed by oral administration of DMF (80 mg/kg body weight) or vehicle at 3, 24, 48, and 72 h after the inflicted TBI. At 4 days after lesion (dal), DMF‐treated mice displayed less neurological deficits than vehicle‐treated mice and reduced histopathological brain damage. At the same time, the TBI‐evoked depletion of brain GSH was prevented by DMF treatment. However, nuclear factor erythroid 2‐related factor 2 target gene mRNA expression involved in antioxidant and detoxifying pathways was increased in both treatment groups at 4 dal. Blood brain barrier leakage, as assessed by immunoglobulin G extravasation, inflammatory marker mRNA expression, and CD45⁺ leukocyte infiltration into the perilesional brain tissue was induced by TBI but not significantly altered by DMF treatment. Collectively, our data demonstrate that post‐traumatic DMF treatment improves neurological outcome and reduces brain tissue loss in a clinically relevant model of TBI. Our findings suggest that DMF treatment confers neuroprotection after TBI via preservation of brain GSH levels rather than by modulating neuroinflammation.

     

Effect of age and partial water deprivation on lipid peroxidation in the brain of male garden lizard

Lipid peroxidation estimated as thiobarbituric acid reactive substance increased with advancing age in the brain of male garden lizard. The degree of increase in the parameter was almost double in senescent phase (middle-aged to old) than in maturation phase (young to middle-aged). Partial water deprivation led to an increase in lipid peroxidation of the brain of young and middle-aged lizards. On the other hand, a similar treatment to old counterparts caused a decrease in the parameter.     

Methamphetamine and lipid peroxidation in the rat retina

BACKGROUND: The use of psychoactive drugs during adolescence and early adult life has increased in the last few decades. It is known that developmental exposure to psychostimulants affects the sensory systems, and the retina has been shown to be a target tissue. This work was conducted to evaluate the pattern of lipid peroxidation in the rat retina following prenatal exposure to methamphetamine (MA). METHODS: Pregnant female Wistar rats were given MA (5 mg/kg of body weight/day; SC, in 0.9% saline) from GD 8 to 22. Offspring were sacrificed at postnatal days (PNDs) 7, 14, and 21. The retinas were homogenized, and both the total antioxidant and superoxide dismutase (SOD) activities were measured by enzymatic-colorimetric methods. The lipid peroxidation byproducts (malondialdehyde [MDA] and MDA-like metabolites) were measured by the thiobarbituric acid test. RESULTS: Total antioxidant levels were lower in the MA group at PND 21 in both males and females. The activity of SOD was higher in PND 7 females from the MA group. MDA levels were higher in the MA group at PND 21 in both genders. CONCLUSIONS: These findings suggest that prenatal-induced MA toxicity in the retina may be related to lipid peroxidation processes and oxidative stress.     

颅脑创伤后大鼠脑组织脑红蛋白表达变化及其与神经元凋亡的关系研究

目的：研究大鼠弥漫性颅脑创伤后脑组织中脑红蛋白的表达变化情况,探究创伤后脑红蛋白表达变化及其与神经元凋亡的关系。方法：采用雄性SD大鼠50只,随机分为10组（n=5）空白对照组、伤后30min、1h、2h、6h、12h、24h、48h、72h和5d组。以Marmarou’s自由落体打击装置复制颅脑创伤模型,采用免疫组化技术检测伤后不同时间脑组织中脑红蛋白的表达情况及神经元凋亡相关基因Bax、Bcl-2表达情况,并对所得数据进行统计学分析。结果：致伤区皮层神经元脑红蛋白表达分别于伤后2h、72h呈现出两次高峰表达;伤后30min～1h、48～72h期间大脑皮层区脑红蛋白表达的上调均伴随着Bax/Bcl-2比值上升趋势减缓甚至呈现下降趋势。结论：弥漫性颅脑创伤后脑组织中脑红蛋白的高表达在一定程度上可以拮抗创伤应激及伤后继发缺血、缺氧性损伤所导致的神经元凋亡,在颅脑创伤的超早期（〈3h）、急性期（〈72h）可能具有一定的神经保护作用。     

Temporal and spatial profile of caspase 8 expression and proteolysis after experimental traumatic brain injury

Recent studies have demonstrated that the downstream caspases, such as caspase 3, act as executors of the apoptotic cascade after traumatic brain injury (TBI) in vivo. However, little is known about the involvement of caspases in the initiation phase of apoptosis, and the interaction between these initiator caspases (e.g. caspase 8) and executor caspases after experimental brain injuries in vitro and in vivo. This study investigated the temporal expression and cell subtype distribution of procaspase 8 and cleaved caspase 8 p20 from 1 h to 14 days after cortical impact-induced TBI in rats. Caspase 8 messenger RNA levels, estimated by semiquantitaive RT-PCR, were elevated from 1 h to 72 h in the traumatized cortex. Western blotting revealed increased immunoreactivity for procaspase 8 and the proteolytically active subunit of caspase 8, p20, in the ipsilateral cortex from 6 to 72 h after injury, with a peak at 24 h after TBI. Similar to our previous studies, immunoreactivity for the p18 fragment of activated caspase 3 also increased in the current study from 6 to 72 h after TBI, but peaked at a later timepoint (48 h) as compared with proteolyzed caspase 8 p20. Immunohistologic examinations revealed increased expression of caspase 8 in neurons, astrocytes and oligodendrocytes. Assessment of DNA damage using TUNEL identified caspase 8- and caspase 3-immunopositive cells with apoptotic-like morphology in the cortex ipsilateral to the injury site, and immunohistochemical investigations of caspase 8 and activated caspase 3 revealed expression of both proteases in cortical layers 2-5 after TBI. Quantitative analysis revealed that the number of caspase 8 positive cells exceeds the number of caspase 3 expressing cells up to 24 h after impact injury. In contrast, no evidence of caspase 8 and caspase 3 activation was seen in the ipsilateral hippocampus, contralateral cortex and hippocampus up to 14 days after the impact. Our results provide the first evidence of caspase 8 activation after experimental TBI and suggest that this may occur in neurons, astrocytes and oligodendrocytes. Our findings also suggest a contributory role of caspase 8 activation to caspase 3 mediated apoptotic cell death after experimental TBI in vivo.     

Extracellular N-acetylaspartate depletion in traumatic brain injury

N-Acetylaspartate (NAA) is almost exclusively localized in neurons in the adult brain and is present in high concentration in the CNS. It can be measured by proton magnetic resonance spectroscopy and is seen as a marker of neuronal damage and death. NMR spectroscopy and animal models have shown NAA depletion to occur in various types of chronic and acute brain injury. We investigated 19 patients with traumatic brain injury (TBI). Microdialysis was utilized to recover NAA, lactate, pyruvate, glycerol and glutamate, at 12-h intervals. These markers were correlated with survival and a 6-month Glasgow Outcome Score. Eleven patients died and eight survived. A linear mixed model analysis showed a significant effect of outcome and of the interaction between time of injury and outcome on NAA levels (p = 0.009 and p = 0.004, respectively). Overall, extracellular NAA was 34% lower in non-survivors. A significant non-recoverable fall was observed in this group from day 4 onwards, with a concomitant rise in lactate-pyruvate ratio and glycerol. These results suggest that mitochondrial dysfunction is a significant contributor to poor outcome following TBI and propose extracellular NAA as a potential marker for monitoring interventions aimed at preserving mitochondrial function.     

Region specific increase in the antioxidant enzymes and lipid peroxidation products in the brain of rats exposed to lead

The objective of this study is to determine the effect of lead (pb) on antioxidant enzymes and lipid peroxidation products in different regions of rat brain. Wistar male rats were treated with lead acetate (500 ppm) through drinking water for a period of 8 weeks. Control animals were maintained on sodium acetate. Treated and control rats were sacrificed at intervals of 1st, 4th and 8th week and the whole brains were dissected on ice into four regions namely the cerebellum, the hippocampus, the frontal cortex and the brain stem. Antioxidant enzymes namely catalase and superoxide dismutase in all the four regions of brain were determined. In addition, lipid peroxidation products were also estimated. The results indicated a gradual increase in the activity of antioxidant enzymes in different regions of the brain and this response was time-dependent. However, the increase was more in the cerebellum and the hippocampus compared to other regions of the brain. The lipid peroxidation products also showed a similar trend suggesting increased effect of lead in these two regions of the brain. The data indicated a region-specific oxidative stress in the brain exposed to lead.     

Clinical characteristics and pathophysiological mechanisms of focal and diffuse traumatic brain injury

Traumatic brain injury (TBI) is a frequent and clinically highly heterogeneous neurological disorder with large socioeconomic consequences. TBI severity classification, based on the hospital admission Glasgow Coma Scale (GCS) score, ranges from mild (GCS 13–15) and moderate (GCS 9–12) to severe (GCS ≤ 8). The GCS reflects the risk of dying from TBI, which is low after mild (∼1%), intermediate after moderate (up to 15%) and high (up to 40%) after severe TBI. Intracranial damage can be focal, such as epidural and subdural haematomas and parenchymal contusions, or diffuse, for example traumatic axonal injury and diffuse cerebral oedema, although this distinction is somewhat arbitrary. Study of the cellular and molecular post-traumatic processes is essential for the understanding of TBI pathophysiology but even more to find therapeutic targets for the development of neuroprotective drugs to be eventually used in human beings. To date, studies in vitro and in vivo, mainly in animals but also in human beings, are unravelling the pathological TBI mechanisms at high pace. Nevertheless, TBI pathophysiology is all but completely elucidated. Neuroprotective treatment studies in human beings have been disappointing thus far and have not resulted in commonly accepted drugs. This review presents an overview on the clinical aspects and the pathophysiology of focal and diffuse TBI, and it highlights several acknowledged important events that occur on molecular and cellular level after TBI.     

牛磺酸对脑创伤大鼠的脑保护作用

目的:探讨牛磺酸(Tau)预处理对弥漫性脑创伤(TBI)大鼠脑皮层超氧化物歧化酶(SOD)活力、丙二醛(MDA)含量、脑含水量(BWC)和脑皮层水孔通道蛋白4(AQP4)表达的影响。方法:复制大鼠TBI模型,分为假手术组(S组)、TBI组(T组)、低剂量Tau组(L组)和高剂量Tau组(H组),用比色法测定脑皮层匀浆液中SOD活力和MDA含量;干/湿法测定BWC;免疫组织化学检测脑皮层AQP4的表达。结果:T组大鼠脑皮层SOD活力显著低于S组,T组MDA含量、BWC和脑皮层AQP4的表达显著高于S组;H、L组脑皮层SOD活力显著高于T组,H、L组MDA含量、BWC和脑皮层AQP4的表达显著低于T组;H、L组之间差异无显著性。结论:Tau可能通过清除TBI后产生的的氧自由基、下调TBI大鼠脑皮层AQP4的表达减轻脑水肿,发挥其脑保护作用。     

Sirtuin 1 alleviates neuroinflammation-induced apoptosis after traumatic brain injury

Sirtuin 1 (SIRT1) plays a very important role in a wide range of biological responses, such as metabolism, inflammation and cell apoptosis. Changes in the levels of SIRT1 have been detected in the brain after traumatic brain injury (TBI). Further, SIRT1 has shown a neuroprotective effect in some models of neuronal death; however, its role and working mechanisms are not well understood in the model of TBI. This study aimed to address this issue. SIRT1-specific inhibitor (sirtinol) and activator (A3) were introduced to explore the role of SIRT1 in cell apoptosis. Results of the study suggest that SIRT1 plays an important role in neuronal apoptosis after TBI by inhibiting NF-κB, IL-6 and TNF-α deacetylation and the apoptotic pathway sequentially, possibly by alleviating neuroinflammation.     

A model of aluminum exposure associated with lipid peroxidation in rat brain

We have developed a rat model to investigate the relationship between aluminum exposure and aluminum accumulation, and with oxidative damage in brain tissues. Intraperitoneal injections of aluminum lactate for 7 wk (the total aluminum dosage per rat was approx 100 mg) significantly increased aluminum levels in the brain. The concentration of lipid peroxidation products (thiobarbituric acid-reactive substances [TBARS]) also increased in the brain following aluminum lactate injections. No significant correlations between the concentrations of aluminum and of TBARS were found in the whole brain. Subcellular analysis revealed that aluminum lactate injections led to a significant increase in the concentration of aluminum in the mitochondrial fraction but had no significant effect on the concentration of peroxides in any subcellular fraction. These results suggest that aluminum accumulation induced by the aluminum lactate administration associates with the acceleration of lipid peroxidation in rat brain. Furthermore, these data indicate that the pro-oxidant effect of aluminum may be indirect and concentration independent. The experimental conditions used here provide an animal model of aluminum accumulation in the brain that should prove useful for further investigations of the mechanisms of aluminum neurotoxicity.     

Caspase 7: increased expression and activation after traumatic brain injury in rats

Caspases, a cysteine proteinase family, are required for the initiation and execution phases of apoptosis. It has been suggested that caspase 7, an apoptosis executioner implicated in cell death proteolysis, is redundant to the main executioner caspase 3 and it is generally believed that it is not present in the brain or present in only minute amounts with highly restricted activity. Here we report evidence that caspase 7 is up-regulated and activated after traumatic brain injury (TBI) in rats. TBI disrupts homeostasis resulting in pathological apoptotic activation. After controlled cortical impact TBI of adult male rats we observed, by semiquantitative real-time PCR, increased mRNA levels within the traumatized cortex and hippocampus peaking in the former about 5 days post-injury and in the latter within 6-24 h of trauma. The activation of caspase 7 protein after TBI, demonstrated by immunoblot by the increase of the active form of caspase 7 peaking 5 days post-injury in the cortex and hippocampus, was found to be up-regulated in both neurons and astrocytes by immunohistochemistry. These findings, the first to document the up-regulation of caspase 7 in the brain after acute brain injury in rats, suggest that caspase 7 activation could contribute to neuronal cell death on a scale not previously recognized.