Terushige Toyooka

Enhanced hippocampal neurodegeneration after traumatic or kainate excitotoxicity in GFAP-null mice

Astrocytes perform a variety of functions in the adult central nervous system. Recent evidence suggests that the upregulation of glial fibrillary acidic protein (GFAP), an astrocyte-specific intermediate filament component, is a biological marker of neurotoxicity after cerebral injury. We herein compared the response to traumatic brain injury or kainic acid (KA)-induced neurotoxicity in GFAP knockout (GFAP-KO) and wild-type (WT) mice. Seventy-two hours after injury, all GFAP-KO mice showed hippocampal CA3 neurodegeneration, whereas WT mice did not show neurodegeneration. Seventy-two hours after KA administration, GFAP-KO mice were more susceptible to KA-induced seizures and had an increased number of pyknotic damaged CA3 neurons than did WT mice. These results indicate that GFAP plays a crucial role in pyramidal neuronal survival after injury or KA-induced neurotoxicity.

Increased xCT expression correlates with tumor invasion and outcome in patients with glioblastomas.

BACKGROUND xCT is a light chain of the cystine/glutamate antiporter system xc. Glutamate that is released by system xc plays an important role in the infiltration of glioblastoma (GBM) cells. Furthermore, increased glutathione synthesis by system xc may protect tumor cells against oxidative stress induced by radiotherapy and chemotherapy. OBJECTIVE To investigate whether the levels of xCT expression correlated with infiltrative imaging phenotypes on magnetic resonance imaging and outcomes in patients with GBMs. METHODS Forty patients with histologically confirmed primary GBMs were included in the study. Patient charts were retrospectively reviewed for age, sex, Karnofsky Performance Status Scale score, Mini-Mental State Examination score, magnetic resonance imaging features, xCT expression, isocitrate dehydrogenase 1 R132H expression, O-methylguanine-DNA methyltransferase promoter methylation status, type of surgery, progression-free survival, and overall survival. RESULTS In invasive margins, xCT expression was weak in 20 patients and strong in 20 patients. A Cox regression model revealed that a Karnofsky Performance Status Scale score less than 60 (hazard ratio [HR]: 4.525; P = .01), partial removal (HR: 2.839; P = .03), and strong xCT expression (HR: 4.134; P < .001) were significantly associated with shorter progression-free survival and that partial removal (HR: 2.865; P = .03), weak isocitrate dehydrogenase 1 R132H expression (HR: 15.729; P = .01), and strong xCT expression (HR: 2.863; P = .04) were significantly associated with shorter overall survival. CONCLUSION These findings suggest that xCT is an independent predictive factor in GBMs.

Effect of Decompressive Craniectomy on Aquaporin-4 Expression after Lateral Fluid Percussion Injury in Rats

Decompressive craniectomy is one therapeutic option for severe traumatic brain injury (TBI), and it has long been used for the treatment of patients with malignant post-traumatic brain edema. A lack of definitive evidence, however, prevents physicians from drawing any conclusions about the effects of decompressive craniectomy for the treatment of TBI. Therefore, the aim of the present study was to investigate the influence of decompressive craniectomy on post-traumatic brain edema formation. The aquaporin-4 (AQP4) water channel is predominantly expressed in astrocytes, and it plays an important role in the regulation of brain water homeostasis. In the present study, we investigated the time course of AQP4 expression and the water content of traumatized cortex following decompressive craniectomy after TBI. Adult male Sprague-Dawley rats (300-400 g) were subjected to lateral fluid percussion injury using the Dragonfly device. The effect of decompressive craniectomy was studied in traumatized rats without craniectomy (closed skull, DC-), and in rats craniectomized immediately after trauma (DC+). AQP4 expression was investigated with a Western blot analysis and immunohistochemistry. Brain edema was measured using the wet weight/dry weight method. At 48 h after TBI, AQP4 expression of the DC- group was significantly increased compared with the DC+ group (p < 0.01). In addition, the cortical water content of the DC- group was significantly increased compared to the DC+ group at the same time point (p < 0.05). The present results suggest that decompressive craniectomy may affect AQP4 expression and reduce brain edema formation after TBI.

Temporal and Spatial Profile of Phosphorylated Connexin43 after Traumatic Brain Injury in Rats

Gap junctions are conductive channels formed by membrane proteins termed connexins (Cx), which permit the intercellular exchange of metabolites, ions, and small molecules. Junctional permeability is regulated by pH, membrane potential, and intracellular secondary messengers. The purpose of this study was to elucidate the expression and distribution of astrocytic gap junctions in the hippocampus and the cortex after traumatic brain injury (TBI) in vivo. Adult male Sprague-Dawley rats (300-400 g) were subjected to lateral fluid percussion injury (FPI) at moderate severity (2.6-2.8 atm, 12 msec) using a Dragonfly device model. Phosphorylated gap junction protein levels were quantified using Western blot analysis. Spatial distribution of immunoreactivity for phosphorylated Cx43 (p-Cx43) was analyzed by immunohistochemistry. Our findings showed that p-Cx43 expression in the ipsilateral hippocampus was significantly induced at 1 h after TBI, and remained at a high level until 24 h after injury. The p-Cx43 protein content reached a maximum level at 6 h after injury. In addition, the immunoreactivity for p-Cx43 was localized in the astrocytes surrounding ipsilateral CA3 pyramidal neurons. On the other hand, the protein level in the ipsilateral cortex was not significantly different at any time point after TBI. Double immunostaining using phosphorylated ERK (p-ERK) showed that p-Cx43 and p-ERK immunoreactivities were enhanced in the same astrocytes at 6 h after injury. These findings suggest that astrocytic gap junctions participate in pathophysiological processes in the hippocampus after TBI.

Alteration of gap junction proteins (connexins) following lateral fluid percussion injury in rats

Gap junctions are intercellular channels that mediate the cytoplasmic exchange of small hydrophilic molecules and are formed by a family of integral membrane proteins called connexins (Cxs). Cx43 is expressed predominantly in astrocytes, while Cx36 is expressed in neurons. In this study, we show alteration of Cx43 and Cx36 in the hippocampus after traumatic brain injury in rats. Adult male Sprague-Dawley rats were subjected to lateral fluid percussion injury of moderate severity. Brain coronal sections were used for immunohistochemistry with Cx43 and Cx36 antibodies. Cx43 immunoreactivity was increased in reactive astrocytes in the damaged hippocampus 24 hours after injury, and persisted for 72 hours. On the other hand, Cx36 immunoreactivity increased in CA3 neurons 1 hour after injury, and decreased later. These results indicate that gap junctions might participate in the pathophysiological process after traumatic brain injury.

Microvascular decompression for hemifacial spasm caused by the vertebral artery: a simple and effective transposition method using surgical glue

BACKGROUND Microvascular decompression for the treatment of trigeminal neuralgia and hemifacial spasm (HFS) has been established and has brought about excellent results. However, recurrence or lack of relief from the symptoms was experienced in some cases in which the nerve root or root exit zone were compressed by the vertebral (VA) and basilar arteries. We experienced a severe HFS case, mainly caused by a compression with the tortuous VA. A more simple transposition technique of the offending VA using surgical glue is reported. METHODS A 64-year-old male had been suffering from HFS for six years. The tortuous right VA with anterior inferior cerebellar arteries (AICA) was considered to be responsible for his vascular compression syndrome. The VA was carefully and slowly dislocated away from the pontomedullary junction toward the cranial base. Then the arteriosclerotic portion of the VA wall was chosen, where a small piece of Surgicel (ETHICON, Inc., Somerville, NJ) is placed beforehand so as to use a minimum amount of glue (Biobond; Mitsubishi Pharma Corp., Osaka, Japan). Until the glue is hardened, the VA is held away from the brain stem. After this procedure, the ordinary transposition of AICA branches was added. RESULTS The annoying HFS completely disappeared immediately after the operation. CONCLUSION The advantage of our method is the simplicity of the procedure. It requires less space and time while the other methods using tapes, strips, and clips need a greater working space and more time.

Focal brain edema and natriuretic peptides in patients with subarachnoid hemorrhage

Central salt wasting syndrome may be caused by pathological increases in serum natriuretic peptides after subarachnoid hemorrhage (SAH). However, it is unclear as to why the serum concentration of atrial natriuretic peptide (ANP) or brain natriuretic peptide (BNP) increases in the subacute phase of SAH. The present study was designed to assess the correlation between focal brain edema and serum concentration of ANP or BNP in patients with SAH. Focal brain edema was found in 8 SAH-patients and peaked between days 4 and 7 of SAH. The mean serum ANP and BNP levels in patients with focal brain edema were significantly higher than those in patients without focal brain edema between days 4 and 14 of SAH. These results suggest that focal brain edema might correlate with increased levels of ANP and BNP in the subacute phase of SAH.

High expression of L-type amino acid transporter 1 in infiltrating glioma cells

L-type amino acid transporter 1 (LAT1), a neutral amino acid transport agent, is essential for the transport of large neutral amino acids. LAT1 also corresponds to tumor-associated gene-1 (TA1), an oncofetal antigen that is expressed primarily in fetal tissues and cancer cells such as glioma cells. We have investigated the expression of the transporter in the human primary glioma tissue from 68 patients. Among these patients, we could see the border zone between tumors and normal bain tissues in 10 patients. By WHO criteria, two of the specimens were diagnosed as grade 2, three as grade 3, and five as grade 4 [glioblastoma multiforme (GBM)]. In 9 of 10 cases, we could identify the infiltrating glioma cells associated with stronger immunoreactivity for LAT1. These tumor cells aggregated around the neurons in the border zone and were often found in the perivascular space. In one GBM case, the tumors seemed to develop expansively and separated from the normal brain with a border of arachnoid membrane. The expression of LAT1 was always higher in infiltrating glioma cells than in cells located in the center of the tumor. These findings suggest that LAT1 is one of the molecular targets for glioma therapy.

Down-Regulation of Glial Fibrillary Acidic Protein and Vimentin by RNA Interference Improves Acute Urinary Dysfunction Associated with Spinal Cord Injury in Rats

In spinal cord injury, glial scarring, a result of overexpressed intermediate filament (IF) proteins including glial fibrillary acidic protein (GFAP) and vimentin, is one of the largest obstacles in axonal regeneration. We postulated that specific suppression of IF proteins in the injured CNS might inhibit the excessive reactivity of astrocytes and thereby suppress glial scarring. siRNAs targeting GFAP and vimentin were transfected separately into C6 glioma cells and rat hippocampal astrocytes. These siRNAs suppressed both biphasic elements of each IF proteins: the ordinarily expressed elements having slow turnover and the immediately inducible elements stimulated by tumor necrosis factor-a (TNF-α). Moreover, adenovirus vectors expressing GFAP or vimentin siRNAs suppressed the proliferation of C6 glioma cells on days 3-9 after infection. Finally, each siRNA mixed with atelocollagen was applied together to the contused thoracic spines of spinal cord injury (SCI) model rats. The introduction of GFAP and vimentin siRNAs prevented the overexpression of IF proteins in the injured lesion (namely, in the white matter surrounding the long tract where the lateral funiculus exists and in the gray matter near the anterior horn neurons). Furthermore, the starting date of spontaneous voiding was significantly accelerated by application of GFAP and vimentin siRNAs. The inhibition of undesirable glial activity surrounding micturition-related pathways improved acute urinary dysfunction due to neurogenic bladder. In conclusion, the down-regulation of IF proteins by RNAi suppresses the overproliferation of reactive astrocytes and thereby might be an effective treatment for spinal cord injury.

Role of the activated extracellular signal-regulated kinase pathway on histological and behavioral outcome after traumatic brain injury in rats.

The extracellular signal-regulated kinase (ERK) pathway, which modulates the activity of many transcriptional factors leading to the proliferation of various cells, is activated in lesions in regions of selective vulnerability after traumatic brain injury (TBI). In the present study, using the ERK inhibitor U0126, we investigated the role of the ERK pathway in histopathological and behavioral outcomes after TBI. Adult male Sprague-Dawley rats, weighing 300-400 g were subjected to lateral fluid percussion brain injury. The ERK inhibitor U0126 was injected intravenously before injury at 100, 200 and 400 microg/kg. The severity of CA3 neuronal damage was evaluated by the number of surviving CA3 neurons 7 days after injury. The contusional lesion volume 72 h after injury was analysed using a computer-assisted analysis system. Three different motor skill tasks were measured on days 1-5, 7, 14 and 21 after injury. Pretreatment with U0126 significantly reduced both CA3 neuronal damage and contusional lesion volume after injury. In addition, administration of U0126 ameliorated motor function recovery on days 3, 4 and 5 after injury. Therefore, inhibition of ERK phosphorylation could be a potentially effective therapeutic target after TBI.