基质金属蛋白酶在心肌缺血再灌注损伤中的作用

肌缺血再灌注损伤是指缺血心肌组织在恢复血流供给后,其细胞代谢功能障碍及结构破坏反而加重的现象,主要表现在心肌收缩与舒张功能障碍、血管内皮功能障碍、微循环血流紊乱、细胞代谢失调、电解质平衡紊乱、细胞凋亡与坏死等,并伴随着氧自由基的大量产生和毒性损伤以及炎症反应的激活,是一个极其复杂的病理过程。基质金属蛋白酶(MMPs)及其组织抑制物(TIMPs)是心肌组织中多种细胞分泌的内源性细胞因子,其作用涵盖了细胞外基质降解、炎症反应激活、调节血管功能、影响细胞凋亡与存活等众多病理生理过程,而这些过程均在心肌缺血再灌注损伤中发挥着重要的作用。     

基质金属蛋白酶在心肌缺血再灌注损伤中的作用

肌缺血再灌注损伤是指缺血心肌组织在恢复血流供给后,其细胞代谢功能障碍及结构破坏反而加重的现象,主要表现在心肌收缩与舒张功能障碍、血管内皮功能障碍、微循环血流紊乱、细胞代谢失调、电解质平衡紊乱、细胞凋亡与坏死等,并伴随着氧自由基的大量产生和毒性损伤以及炎症反应的激活,是一个极其复杂的病理过程。基质金属蛋白酶（MMPs）及其组织抑制物（TIMPs）是心肌组织中多种细胞分泌的内源性细胞因子,其作用涵盖了细胞外基质降解、炎症反应激活、调节血管功能、影响细胞凋亡与存活等众多病理生理过程,而这些过程均在心肌缺血再灌注损伤中发挥着重要的作用。     

Inflammation in Traumatic Brain Injury: Role of Cytokines and Chemokines

A traumatic injury to the adult mammalian central nervous system (CNS), such as a stab wound lesion, results in reactive astrogliosis and the migration of hematogenous cells into the damaged neural tissue. The roles of cytokines and growth factors released locally by the damaged endogenous cells are recognized in controlling the cellular changes that occur following CNS injury. However, the role of chemokines, a novel class of chemoattractant cytokines, is only recently being studied in regulating inflammatory cell invasion in the injured/diseased CNS (1). The mRNAs for several chemokines have been shown to be upregulated in experimental allergic encephalomyelitis (EAE), an inflammatory demyelinating disease of the CNS, but chemokine expression in traumatic brain injury has not been studied in detail. Astrocytes have been demonstrated to participate in numerous processes that occur following injury to the CNS. In particular, astrocytic expression of cytokines and growth factors in the injured CNS has been well reviewed (2). Recently a few studies have detected the presence of chemokines in astrocytes following traumatic brain injury (3,4). These studies have suggested that chemokines may represent a promising target for future therapy of inflammatory conditions. This review summarizes the events that occur in traumatic brain injury and discusses the roles of resident and non-resident cells in the expression of growth factors, cytokines and chemokines in the injured CNS.     

Role of the Prostaglandin E2 EP1 Receptor in Traumatic Brain Injury

Brain injuries promote upregulation of so-called proinflammatory prostaglandins, notably prostaglandin E₂ (PGE₂), leading to overactivation of a class of its cognate G-protein-coupled receptors, including EP1, which is considered a promising target for treatment of ischemic stroke. However, the role of the EP1 receptor is complex and depends on the type of brain injury. This study is focused on the investigation of the role of the EP1 receptor in a controlled cortical impact (CCI) model, a preclinical model of traumatic brain injury (TBI). The therapeutic effects of post-treatments with a widely studied EP1 receptor antagonist, SC-51089, were examined in wildtype and EP1 receptor knockout C57BL/6 mice. Neurological deficit scores (NDS) were assessed 24 and 48 h following CCI or sham surgery, and brain immunohistochemical pathology was assessed 48 h after surgery. In wildtype mice, CCI resulted in an obvious cortical lesion and localized hippocampal edema with an associated significant increase in NDS compared to sham-operated animals. Post-treatments with the selective EP1 receptor antagonist SC-51089 or genetic knockout of EP1 receptor had no significant effects on cortical lesions and hippocampal swelling or on the NDS 24 and 48 h after CCI. Immunohistochemistry studies revealed CCI-induced gliosis and microglial activation in selected ipsilateral brain regions that were not affected by SC-51089 or in the EP1 receptor-deleted mice. This study provides further clarification on the respective contribution of the EP1 receptor in TBI and suggests that, under this experimental paradigm, the EP1 receptor would have limited effects in modulating acute neurological and anatomical pathologies following contusive brain trauma. Findings from this protocol, in combination with previous studies demonstrating differential roles of EP1 receptor in ischemic, neurotoxic, and hemorrhagic conditions, provide scientific background and further clarification of potential therapeutic application of prospective prostaglandin G-protein-coupled receptor drugs in the clinic for treatment of TBI and other acute brain injuries.     

Endocannabinoids and Traumatic Brain Injury

In response to traumatic brain injury, there is local and transient accumulation of 2-AG at the site of injury, peaking at 4 h and sustained up to at least 24 h. Neuroprotection exerted by exogenous 2-AG suggests that the formation of 2-AG may serve as a molecular regulator of pathophysiological events, attenuating the brain damage. Inhibition of this protective effect by SR-141716A, a CB₁ cannabinoid receptor antagonist, and the lack of effect of 2-AG in CB₁ knockout mice suggest that 2-AG and the CB₁ receptor may be important in the pathophysiology of traumatic brain injury. 2-AG exerts its neuroprotective effect after traumatic brain injury, at least in part, by inhibition of NF-κB transactivation. 2-AG also inhibits, at an early stage (2–4 h), the expression of the main proinflammatory cytokines, TNF-α, IL-6, and IL-1β, and is accompanied by reduction of BBB permeability. Moreover, the CB₁, CB₂, and TRVP1 receptors are expressed on microvascular endothelial cells, and their activation by 2-AG counteracts endothelin (ET-1)-induced cerebral microvascular responses (namely, Ca²⁺ mobilization and cytoskeleton rearrangement). This suggests that the functional interaction between 2-AG and ET-1 may provide a potential alternative pathway for abrogating ET-1-inducible vasoconstriction after brain injury and play a role in the neuroprotective effects exerted by 2-AG, as a potent vasodilator.     

冷诱导RNA结合蛋白在急性脑外伤后的表达变化及其促炎作用研究

目的:观察小鼠急性脑外伤后,冷诱导RNA结合蛋白(CIRP)的表达变化情况及其对炎症因子的影响,旨在探讨CIRP在神经系统损伤性疾病中的作用,为该疾病的临床治疗提供新的思路。方法:成年小鼠随机分为正常组(n=10)和手术组(n=36),用液压打击法建立小鼠急性脑外伤模型,利用HE染色法观察模型建立后小鼠大脑的损伤情况;损伤后24、48 h时间点,采用实时PCR检测大脑损伤部位CIRP和炎症因子肿瘤坏死因子α(TNFα)m RNA的表达,免疫组织化学法检测CIRP蛋白水平的表达;分析CIRP表达变化与炎症因子TNFα相关性。结果:急性脑外伤后小鼠脑损伤部位的CIRP m RNA及蛋白表达水平呈上升趋势,CIRP表达与炎症因子TNFα表达正相关。结论:CIRP可能参与了小鼠脑外伤的炎性反应过程。     

Expression of Dixdc1 and its Role in Astrocyte Proliferation after Traumatic Brain Injury

DIX domain containing 1 (Dixdc1), a positive regulator of Wnt signaling pathway, is recently reported to play a role in the neurogenesis. However, the distribution and function of Dixdc1 in the central nervous system (CNS) after brain injury are still unclear. We used an acute traumatic brain injury (TBI) model in adult rats to investigate whether Dixdc1 is involved in CNS injury and repair. Western blot analysis and immunohistochemistry showed a time-dependent up-regulation of Dixdc1 expression in ipsilateral cortex after TBI. Double immunofluorescent staining indicated a colocalization of Dixdc1 with astrocytes and neurons. Moreover, we detected a colocalization of Ki-67, a cell proliferation marker with GFAP and Dixdc1 after TBI. In primary cultured astrocytes stimulated with lipopolysaccharide, we found enhanced expression of Dixdc1 in parallel with up-regulation of Ki-67 and cyclin A, another cell proliferation marker. In addition, knockdown of Dixdc1 expression in primary astrocytes with Dixdc1-specific siRNA transfection induced G0/G1 arrest of cell cycle and significantly decreased cell proliferation. In conclusion, all these data suggest that up-regulation of Dixdc1 protein expression is potentially involved in astrocyte proliferation after traumatic brain injury in the rat.     

Delayed Phospholipid Degradation in Rat Brain After Traumatic Brain Injury

Abstract: Lipid second messengers such as arachidonic acid and its metabolites and diacylglycerols (DAGs) are affected in brain injury. Therefore, changes in the pool size and the fatty acid composition of free fatty acids (FFAs) and DAGs were analyzed in different rat brain areas 4 and 35 days after traumatic injury. Cortical impact injury of low-grade severity was applied in the right frontal somatosensory cortex. Four days after injury, FFAs and DAGs were increased by three- and twofold, respectively, in the injured cortex and to a lesser extent in the contralateral cortex compared with sham-operated animals. Docosahexaenoic acid followed by stearic acid, and arachidonic acid, displayed the greatest changes in both FFAs and DAGs. By day 35, free stearic, oleic, and arachidonic acids remained elevated in the damaged cortex (1.5-fold each). DAGs showed the greatest change, reaching values 2.7-fold higher than sham in all frontal and occipital cortical areas, including brainstem. Oleoyl- and arachidonoyl-DAGs (four- and threefold increase, respectively) followed by docosahexaenoyl-DAGs (twofold) contributed to the DAG accumulation. These results reveal that traumatic brain injury triggers a sustained and time-dependent activation of phospholipase-mediated signaling pathways leading to membrane phospholipid degradation and targeting, early on, docosahexaenoyl phospholipid-enriched excitable membranes.     

Quantification of Axonal Damage in Traumatic Brain Injury

Abstract : Diffuse axonal injury is a primary feature of head trauma and is one of the most frequent causes of mortality and morbidity. Diffuse axonal injury is microscopic in nature and difficult or impossible to detect with imaging techniques. The objective of the present study was to determine whether axonal injury in head trauma patients could be quantified by measuring levels of CSF tau proteins. Tau proteins are structural microtubule binding proteins primarily localized in the axonal compartment of neurons. Monoclonal antibodies recognizing the form of tau found in the CSF of head trauma patients were developed by differential CSF hybridoma screening using CSF from head trauma and control patients. Clones positive for head trauma CSF tau proteins were used to characterize this form of tau and for ELISA development. Using the developed ELISA, CSF tau levels were elevated >1,000-fold in head trauma patients (mean, 1,519 ng/ml of CSF) when compared with patients with multiple sclerosis (mean, 0.014 ng/ml of CSF ; p < 0.001), normal pressure hydrocephalus (nondetectable CSF tau), neurologic controls (mean, 0.031 ng/ml of CSF ; p < 0.001), or nonneurologic controls (nondetectable CSF tau ; p < 0.001). In head trauma, a relationship between clinical improvement and decreased CSF tau levels was observed. These data suggest that CSF tau levels may prove a clinically useful assay for quantifying the axonal injury associated with head trauma and monitoring efficacy of neuroprotective agents. Affinity purification of CSF tau from head trauma patients indicated a uniform cleavage of ~ 18 kDa from all six tau isoforms, reducing their apparent molecular sizes to 30-50 kDa. These cleaved forms of CSF tau consisted of the interior portion of the tau sequence, including the microtubule binding domain, as judged by cyanogen bromide digestion. Consistent with these data, CSF cleaved tau bound taxolpolymerized microtubules, indicating a functionally intact microtubule binding domain. Furthermore, epitope mapping studies suggested that CSF cleaved tau proteins consist of the interior portion of the tau sequence with cleavage at both N and C terminals.     

Efficacy of N-Acetyl Cysteine in Traumatic Brain Injury

In this study, using two different injury models in two different species, we found that early post-injury treatment with N-Acetyl Cysteine (NAC) reversed the behavioral deficits associated with the TBI. These data suggest generalization of a protocol similar to our recent clinical trial with NAC in blast-induced mTBI in a battlefield setting [1], to mild concussion from blunt trauma. This study used both weight drop in mice and fluid percussion injury in rats. These were chosen to simulate either mild or moderate traumatic brain injury (TBI). For mice, we used novel object recognition and the Y maze. For rats, we used the Morris water maze. NAC was administered beginning 30–60 minutes after injury. Behavioral deficits due to injury in both species were significantly reversed by NAC treatment. We thus conclude NAC produces significant behavioral recovery after injury. Future preclinical studies are needed to define the mechanism of action, perhaps leading to more effective therapies in man.     

Selective Inhibition of Matrix Metalloproteinase-9 Attenuates Secondary Damage Resulting from Severe Traumatic Brain Injury

Traumatic brain injury (TBI) is a leading cause of death and long-term disability. Following the initial insult, severe TBI progresses to a secondary injury phase associated with biochemical and cellular changes. The secondary injury is thought to be responsible for the development of many of the neurological deficits observed after TBI and also provides a window of opportunity for therapeutic intervention. Matrix metalloproteinase-9 (MMP-9 or gelatinase B) expression is elevated in neurological diseases and its activation is an important factor in detrimental outcomes including excitotoxicity, mitochondrial dysfunction and apoptosis, and increases in inflammatory responses and astrogliosis. In this study, we used an experimental mouse model of TBI to examine the role of MMP-9 and the therapeutic potential of SB-3CT, a mechanism-based gelatinase selective inhibitor, in ameliorating the secondary injury. We observed that activation of MMP-9 occurred within one day following TBI, and remained elevated for 7 days after the initial insult. SB-3CT effectively attenuated MMP-9 activity, reduced brain lesion volumes and prevented neuronal loss and dendritic degeneration. Pharmacokinetic studies revealed that SB-3CT and its active metabolite, p-OH SB-3CT, were rapidly absorbed and distributed to the brain. Moreover, SB-3CT treatment mitigated microglial activation and astrogliosis after TBI. Importantly, SB-3CT treatment improved long-term neurobehavioral outcomes, including sensorimotor function, and hippocampus-associated spatial learning and memory. These results demonstrate that MMP-9 is a key target for therapy to attenuate secondary injury cascades and that this class of mechanism-based gelatinase inhibitor–with such desirable pharmacokinetic properties–holds considerable promise as a potential pharmacological treatment of TBI.     

Cerebrospinal Fluid Taurine after Traumatic Brain Injury

In the experimental setting, taurine is known to be released from swollen cells to reestablish their normal volume. However, its clinical relevance has not been fully understood. This study was undertaken to reveal changes in cerebrospinal fluid (CSF) amino acids concentration in patients with severe traumatic brain injury (TBI). The study included eight patients, in whom a ventricular catheter was inserted to measure intracranial pressure and obtain CSF samples for 5 days. CSF obtained from patients with normal pressure hydrocephalus served as a control. CSF taurine concentration increased 1.8 times control (P < 0.05) after TBI and returned to control value approximately 67 h after injury. Taurine decreased further and remained lower than control thereafter. Phosphoethanolamine showed similar increase, whereas glutamine decreased transiently and arginine remained close to control value. The present data support the period of astrocytic swelling observed after TBI in other morphological studies. The mechanism and consequences of CSF taurine decrease in the subacute stage of TBI need to be elucidated.     

Depletion of Brain Docosahexaenoic Acid Impairs Recovery from Traumatic Brain Injury

Omega-3 fatty acids are crucial for proper development and function of the brain where docosahexaenoic acid (DHA), the primary omega-3 fatty acid in the brain, is retained avidly by the neuronal membranes. We investigated the effect of DHA depletion in the brain on the outcome of traumatic brain injury (TBI). Pregnant mice were put on an omega-3 fatty acid adequate or deficient diet from gestation day 14 and the pups were raised on the respective diets. Continuation of this dietary regime for three generations resulted in approximately 70% loss of DHA in the brain. Controlled cortical impact was delivered to both groups of mice to produce severe TBI and the functional recovery was compared. Compared to the omega-3 adequate mice, the DHA depleted mice exhibited significantly slower recovery from motor deficits evaluated by the rotarod and the beam walk tests. Furthermore, the DHA deficient mice showed greater anxiety-like behavior tested in the open field test as well as cognitive deficits evaluated by the novel object recognition test. The level of alpha spectrin II breakdown products, the markers of TBI, was significantly elevated in the deficient mouse cortices, indicating that the injury is greater in the deficient brains. This observation was further supported by the reduction of NeuN positive cells around the site of injury in the deficient mice, indicating exacerbated neuronal death after injury. These results suggest an important influence of the brain DHA status on TBI outcome.     

创伤性脑损伤中神经炎症相关细胞的研究进展

创伤性脑损伤(traumatic brain injury, TBI), 亦称颅脑损伤或头部外伤, 专指由外伤引起的脑组织损害。然而,从轻度到重度的TBI,改善TBI患者预后的治疗方法都十分匮乏。神经炎症可引起脑外伤后急性继发性损伤,并与慢性神经退行性疾病有关,因此,系统了解参与TBI后神经炎性反应的细胞显得尤为重要。主要对TBI中参与炎症反应的细胞（如小胶质细胞、星形胶质细胞、少突细胞、中性粒细胞和淋巴细胞）的启动以及相互作用的最新研究进展进行了综述,以期为临床研究提供新的策略。     

Changes in Body Temperature Rhythm after Traumatic Brain Injury

We performed this study to determine whether in head injured patients body temperature rhythmicity exists outside the usual spectrum. Temperature data of in total 22 patients with head injury were analyzed using the Regressive and Iterative Cosinor methods. We found that circadian rhythm often remained, and usually was combined with rhythms in ultradian or infradian ranges. Tau shifts over consecutive days were observed in three severely head injured patients (Glasgow Coma Scale score ≤ 8). To validate the results we used surrogate data. Detection of temperature rhythms in this study may serve to estimate the clinical importance of biological rhythms in head injury.     

Changes in Body Temperature Rhythm after Traumatic Brain Injury

We performed this study to determine whether in head injured patients body temperature rhythmicity exists outside the usual spectrum. Temperature data of in total 22 patients with head injury were analyzed using the Regressive and Iterative Cosinor methods. We found that circadian rhythm often remained, and usually was combined with rhythms in ultradian or infradian ranges. Tau shifts over consecutive days were observed in three severely head injured patients (Glasgow Coma Scale score ≤ 8). To validate the results we used surrogate data. Detection of temperature rhythms in this study may serve to estimate the clinical importance of biological rhythms in head injury.     

Controlled Cortical Impact Model for Traumatic Brain Injury

Every year over a million Americans suffer a traumatic brain injury (TBI). Combined with the incidence of TBIs worldwide, the physical, emotional, social, and economical effects are staggering. Therefore, further research into the effects of TBI and effective treatments is necessary. The controlled cortical impact (CCI) model induces traumatic brain injuries ranging from mild to severe. This method uses a rigid impactor to deliver mechanical energy to an intact dura exposed following a craniectomy. Impact is made under precise parameters at a set velocity to achieve a pre-determined deformation depth. Although other TBI models, such as weight drop and fluid percussion, exist, CCI is more accurate, easier to control, and most importantly, produces traumatic brain injuries similar to those seen in humans. However, no TBI model is currently able to reproduce pathological changes identical to those seen in human patients. The CCI model allows investigation into the short-term and long-term effects of TBI, such as neuronal death, memory deficits, and cerebral edema, as well as potential therapeutic treatments for TBI.