Inhibiting endogenous tissue plasminogen activator enhanced neuronal apoptosis and axonal injury after traumatic brain injury.

Tissue plasminogen activator is usually used for the treatment of acute ischemic stroke, but the role of endogenous tissue plasminogen activator in traumatic brain injury has been rarely reported. A rat model of traumatic brain injury was established by weight-drop method. The tissue plasminogen activator inhibitor neuroserpin (5 µL, 0.25 mg/mL) was injected into the lateral ventricle. Neurological function was assessed by neurological severity score. Neuronal and axonal injuries were assessed by hematoxylin-eosin staining and Bielschowsky silver staining. Protein level of endogenous tissue plasminogen activator was analyzed by western blot assay. Apoptotic marker cleaved caspase-3, neuronal marker neurofilament light chain, astrocyte marker glial fibrillary acidic protein and microglial marker Iba-1 were analyzed by immunohistochemical staining. Apoptotic cell types were detected by immunofluorescence double labeling. Apoptotic cells in the damaged cortex were detected by terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP-biotin nick-end labeling staining. Degenerating neurons in the damaged cortex were detected by Fluoro-Jade B staining. Expression of tissue plasminogen activator was increased at 6 hours, and peaked at 3 days after traumatic brain injury. Neuronal apoptosis and axonal injury were detected after traumatic brain injury. Moreover, neuroserpin enhanced neuronal apoptosis, neuronal injury and axonal injury, and activated microglia and astrocytes. Neuroserpin further deteriorated neurobehavioral function in rats with traumatic brain injury. Our findings confirm that inhibition of endogenous tissue plasminogen activator aggravates neuronal apoptosis and axonal injury after traumatic brain injury, and activates microglia and astrocytes. This study was approved by the Biomedical Ethics Committee of Animal Experiments of Shaanxi Province of China in June 2015.

Vascular endothelial growth factor A promotes platelet adhesion to collagen IV and causes early brain injury after subarachnoid hemorrhage.

The role of vascular endothelial growth factor A in platelet adhesion in cerebral microvessels in the early stage of subarachnoid hemorrhage remains unclear. In this study, the endovascular puncture method was used to produce a rat model of subarachnoid hemorrhage. Then, 30 minutes later, vascular endothelial growth factor A antagonist anti-vascular endothelial growth factor receptor 2 antibody, 10 µg, was injected into the right ventricle. Immunohistochemistry and western blot assay were used to assess expression of vascular endothelial growth factor A, occludin and claudin-5. Immunohistochemical double labeling was conducted to examine co-expression of GP Ia-II integrin and type IV collagen. TUNEL was used to detect apoptosis in the hippocampus. Neurological score was used to assess behavioral performance. After subarachnoid hemorrhage, the expression of vascular endothelial growth factor A increased in the hippocampus, while occludin and claudin-5 expression levels decreased. Co-expression of GP Ia-II integrin and type IV collagen and the number of apoptotic cells increased, whereas behavioral performance was markedly impaired. After treatment with anti-vascular endothelial growth factor receptor 2 antibody, occludin and claudin-5 expression recovered, while co-expression of GP Ia-II integrin and type IV collagen and the number of apoptotic cells decreased. Furthermore, behavioral performance improved notably. Our findings suggest that increased vascular endothelial growth factor A levels promote platelet adhesion and contribute to early brain injury after subarachnoid hemorrhage. This study was approved by the Biomedical Ethics Committee, Medical College of Xi'an Jiaotong University, China in December 2015.

Dynamic expression of nerve growth factor and its receptor TrkA after subarachnoid hemorrhage in rat brain.

Delayed ischemic neurologic deficit after subarachnoid hemorrhage results from loss of neural cells. Nerve growth factor and its receptor TrkA may promote regeneration of neural cells, but their expression after subarachnoid hemorrhage remains unclear. In the present study, a rat model of subarachnoid hemorrhage was established using two injections of autologous blood into the cistern magna. Immunohisto-chemical staining suggested that the expression of nerve growth factor and TrkA in the cerebral cortex and brainstem increased at 6 hours, peaked at 12 hours and decreased 1 day after induction of subarachnoid hemorrhage, whereas the expression in the hippocampus increased at 6 hours, peaked on day 1, and decreased 3 days later. Compared with those for the rats in the sham and saline groups, neurobehavioral scores decreased significantly 12 hours and 3 days after subarachnoid hemorrhage (P < 0.05). These results suggest that the expression of nerve growth factor and its receptor TrkA is dynamically changed in the rat brain and may thus participate in neuronal survival and nerve regeneration after subarachnoid hemorrhage.

Rapidly increased vasopressin promotes acute platelet aggregation and early brain injury after experimental subarachnoid hemorrhage in a rat model.

OBJECTIVE: To investigate the dynamic expression of vasopressin and its potential role in rat brain tissue after experimental subarachnoid hemorrhage (SAH). METHODS: Male Sprague-Dawley rats were divided into 10min, 1h, 6h, 24h, 48h and 72h groups. The SAH model was established by endovascular puncture. ELISA and immunohistochemistry were performed to evaluate dynamic expression of vasopressin. Immunohistochemistry of GPIIb/IIIa integrin was used to assess platelet aggregation. Double immunofluorescence labeling was carried out to observe the reaction between vasopressin and platelet. Early brain injury was evaluated by apoptotic cells counting. Neurobehavioral score was performed to assess neuroprotective role of SR 49059 (a selective antagonists of vasopressin receptor). RESULTS: In peripheral blood and hypothalamus, vasopressin increased rapidly at 6h and 24h. Expression of GPIIb/IIIa integrin peaked at 24h in cortex and hippocampus. Immunofluorescence showed that vasopressin and GPIIb/IIIa integrin located at the same site. Administration of SR 49059 significantly decreased platelet aggregation and number of apoptotic cells. The neurobehavioral score was promoted significantly after the intervention. CONCLUSION: The results indicate that rapidly increased vasopressin could induce platelet aggregation and contribute to early brain injury after SAH.

Meta-analysis of the efficacy and safety of therapeutic hypothermia in children with acute traumatic brain injury.

OBJECTIVE: To evaluate the efficacy and safety of therapeutic hypothermia in children with acute traumatic brain injury (TBI). METHODS: A systematic literature review using PubMed, Embase, Cochrane Library, Chinese National Knowledge Infrastructure, Wanfang, VIP, and Chinese Biomedical Database was performed to retrieve studies of randomized controlled trials (RCTs) on therapeutic hypothermia for children with TBI published before March 2014. Data extraction and quality evaluation of RCTs were performed by 2 investigators independently. A meta-analysis was performed by RevMan 5.2.7. RESULTS: There were 7 RCTs comprising 442 children (218 in hypothermia group and 224 in normothermia group). Meta-analysis showed therapeutic hypothermia could increase mortality compared with the normothermia group (relative risk [RR] = 1.84, 95% confidence interval [CI] = 1.15-2.93, P = 0.01). On the Glasgow Outcome Scale (GOS), the following scores did not differ between the hypothermia group and normothermia group: 3-month GOS 4-5 (RR = 0.89, 95% CI = 0.68-1.16, P = 0.39), 3-month GOS 1-3 (RR = 1.19, 95% CI = 0.80-1.76, P = 0.39), 6-month GOS 4-5 (RR = 0.91, 95% CI = 0.78-1.07, P = 0.26), and 6-month GOS 1-3 (RR = 1.18, 95% CI = 0.88-1.59, P = 0.27). Hypothermia did not increase the rate of pneumonia (RR = 0.84, 95% CI = 0.63-1.12, P = 0.23) or bleeding (RR = 0.94, 95% CI = 0.39-2.26, P = 0.89), but the incidence of arrhythmias was higher in the hypothermia group (RR = 2.60, 95% CI = 1.06-6.41, P = 0.04). CONCLUSIONS: No benefit of therapeutic hypothermia in children with TBI is shown in this study; therapeutic hypothermia may increase the risk of mortality and arrhythmia. There is no evidence that therapeutic hypothermia improves prognosis of children with TBI; there is also no evidence that therapeutic hypothermia increases the risk of pneumonia and coagulation dysfunction. These results are limited by the quality of the included studies and need to be considered with caution. Further large-scale, well-designed RCTs on this topic are needed.

Etanercept alleviates early brain injury following experimental subarachnoid hemorrhage and the possible role of tumor necrosis factor-α and c-Jun N-terminal kinase pathway.

Cerebral inflammation plays a crucial role in early brain injury (EBI) after subarachnoid hemorrhage (SAH). This study investigated the effects of c-Jun N-terminal kinase (JNK) inhibitor SP600125, acetylcholine (Ach), etanercept, and anti-TNF-α on cellular apoptosis in the cerebral cortex and the hippocampus, in order to establish the role of JNK and TNF-α in EBI. The SAH model was established using an endovascular puncture protocol. The reliability of the EBI model was determined by phosphorylated-Bad (pBad) immunohistochemistry. Neurological scores were recorded and western blot was used to detect the expression of JNK and TNF-α, and TUNEL assay was used to mark apoptotic cells. The results showed that pBad positive cells were evenly distributed in the cerebral cortex at different time points. The highest expression of pBad was reached 1 day after SAH, and pJNK and TNF-α reached their peak expression at 2 days after SAH. SP600125, Ach, and etanercept significantly decreased the level of pJNK and TNF-α in the cerebral cortex and the hippocampus. In addition, SP600125 and etanercept reduced cellular apoptosis in the cerebral cortex and the hippocampus and significantly improved neurological scores at 2 days after SAH potentially via inhibition of the JNK-TNF-α pathway. Ach reduced cellular apoptosis only in the cerebral cortex. It is possible that JNK induces TNF-α expression, which in turn enhances JNK expression in EBI after SAH, leading to increased apoptosis in the cerebral cortex and the hippocampus. Thus, our results indicate that that etanercept may be a potential therapeutic agent to alleviate EBI.