A highly annotated whole-genome sequence of a Korean individual.

Recent advances in sequencing technologies have initiated an era of personal genome sequences. To date, human genome sequences have been reported for individuals with ancestry in three distinct geographical regions: a Yoruba African, two individuals of northwest European origin, and a person from China. Here we provide a highly annotated, whole-genome sequence for a Korean individual, known as AK1. The genome of AK1 was determined by an exacting, combined approach that included whole-genome shotgun sequencing (27.8x coverage), targeted bacterial artificial chromosome sequencing, and high-resolution comparative genomic hybridization using custom microarrays featuring more than 24 million probes. Alignment to the NCBI reference, a composite of several ethnic clades, disclosed nearly 3.45 million single nucleotide polymorphisms (SNPs), including 10,162 non-synonymous SNPs, and 170,202 deletion or insertion polymorphisms (indels). SNP and indel densities were strongly correlated genome-wide. Applying very conservative criteria yielded highly reliable copy number variants for clinical considerations. Potential medical phenotypes were annotated for non-synonymous SNPs, coding domain indels, and structural variants. The integration of several human whole-genome sequences derived from several ethnic groups will assist in understanding genetic ancestry, migration patterns and population bottlenecks.

Mechanisms of anti-inflammatory and neuroprotective actions of PPAR-gamma agonists.

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors of the nuclear hormone receptor superfamily. The 3 PPAR isoforms (alpha, delta/beta and gamma) are known to control many physiological functions including glucose absorption, lipid balance, and cell growth and differentiation. Of interest, PPAR-gamma activation was recently shown to mitigate the inflammation associated with chronic and acute neurological insults. Particular attention was paid to test the therapeutic potential of PPAR agonists in acute conditions like stroke, spinal cord injury (SCI) and traumatic brain injury (TBI), in which massive inflammation plays a detrimental role. While 15d-prostaglandin J2 (15d PGJ2) is the natural ligand of PPAR-gamma, the thiazolidinediones (TZDs) are potent exogenous agonists. Due to their insulin-sensitizing properties, 2 TZDs rosiglitazone and pioglitazone are currently FDA-approved for type-2 diabetes treatment. Recent studies from our laboratory and other groups have shown that TZDs induce significant neuroprotection in animal models of focal ischemia and SCI by multiple mechanisms. The beneficial actions of TZDs were observed to be both PPAR-gamma-dependent as well as -independent. The major mechanism of TZD-induced neuroprotection seems to be prevention of microglial activation and inflammatory cytokine and chemokine expression. TZDs were also shown to prevent the activation of pro-inflammatory transcription factors at the same time promoting the anti-oxidant mechanisms in the injured CNS. This review article discusses the multiple mechanisms of TZD-induced neuroprotection in various animal models of CNS injury with an emphasis on stroke.

Role of transcription factors in mediating post-ischemic cerebral inflammation and brain damage.

Inflammation is a known precipitator of neuronal death after cerebral ischemia. The mechanisms that promote or curtail the start and spread of inflammation in brain are still being debated. By virtue of their capability to modulate gene expression, several transcription factors induced in the ischemic brain can modulate the post-ischemic inflammation. While the induction of transcription factors such as IRF1, NF-kappaB, ATF-2, STAT3, Egr1 and C/EBPbeta is thought to promote post-ischemic inflammation, activation of transcription factors such as HIF-1, CREB, c-fos, PPARalpha, PPARgamma and p53 is thought to prevent post-ischemic inflammation and neuronal damage. Of these, PPARgamma which is a ligand-activated transcription factor was recently shown to prevent inflammatory gene expression in several animal models CNS disorders. This review article discusses some of the molecular mechanisms of PPARgamma induction by its agonists following focal cerebral ischemia.

Glutamate transporter EAAT4 is increased in hippocampal astrocytes following lateral fluid-percussion injury in the rat.

Functional impairment of glutamate transporters contributes to excitotoxic damage and exacerbation of injury in certain neurodegenerative disorders. Several high-affinity sodium-dependent glutamate transporters have been cloned thus far. Of these, EAAT4 is abundantly expressed in Purkinje cells of the cerebellum in rats. However, little is currently known regarding levels of EAAT4 following traumatic brain injury (TBI). In this study, EAAT4 changes were examined for up to 7 days after moderate fluid-percussion by immunoblotting and immunohistochemistry. TBI caused a 20% and 25% increase in EAAT4 levels in the injured hippocampus at day 3 and day 7 following the insult. Immunohistochemical analysis revealed this increase to be localized in cells exhibiting morphological characteristics of astrocytes. In addition, increased EAAT4 immunoreactivity was observed in astrocytes in the ipsilateral cortex and cerebellum at day 3 post-injury that persisted up to 7 days after the insult. Given the reported novel characteristics of chloride conductance displayed by this transporter, our findings of increased EAAT4 levels suggest this protein may play an important role in the pathophysiology of TBI.

Early, transient increase in complexin I and complexin II in the cerebral cortex following traumatic brain injury is attenuated by N-acetylcysteine.

Alteration of excitatory neurotransmission is a key feature of traumatic brain injury (TBI) in which extracellular glutamate levels rise. Although increased synaptic release of glutamate occurs at the injury site, the precise mechanism is unclear. Complexin I and complexin II constitute a family of cytosolic proteins involved in the regulation of neurotransmitter release, competing with the chaperone protein alpha-SNAP (soluble N-ethylmaleimide-sensitive factor-attachment protein) for binding to the synaptic vesicle protein synaptobrevin as well as the synaptic membrane proteins SNAP-25 and syntaxin, which together form the SNAP receptor (SNARE) complex. Complexin I is predominantly a marker of axosomatic (inhibitory) synapses, whereas complexin II mainly labels axodendritic and axospinous synapses, the majority of which are excitatory. In order to examine the role of these proteins in TBI, we have studied levels of both complexins in the injured hemisphere by immunoblotting over a time period ranging from 6 h to 7 days following lateral fluid-percussion brain injury in the rat. Transient increases in the levels of complexin I and complexin II proteins were detected in the injured cerebral cortex 6 h following TBI. This increase was followed by a decrease of complexin I in the injured cortex and hippocampus, and a decrease in both complexins in the injured thalamus region at day 3 and day 7 post-injury. The early, transient increase in the injured cortex was completely blocked by N-acetylcysteine (NAC) administered 5 min following trauma, suggesting an involvement of oxidative stress. Neuronal loss was also reduced in the injured hemisphere with post-TBI NAC treatment. Our findings suggest a dysregulation of both inhibitory and excitatory neurotransmission following traumatic injury that is responsive to antioxidant treatment. These alterations in complexin levels may also play an important role in neuronal cell loss following TBI, and thus contribute to the pathophysiology of cerebral damage following brain injury.

Excitotoxic mechanisms and the role of astrocytic glutamate transporters in traumatic brain injury.

Glutamate excitotoxicity plays an important role in the development of secondary injuries that occur following traumatic brain injury (TBI), and contributes significantly to expansion of the total volume of injury. Acute increases in extracellular glutamate levels have been detected in both experimental brain trauma models and in human patients, and can lead to over-stimulation of glutamate receptors, resulting in a cascade of excitotoxic-related mechanisms culminating in neuronal damage. These elevated levels of glutamate can be effectively controlled by the astrocytic glutamate transporters GLAST (EAAT1) and GLT-1 (EAAT2). However, evidence indicate these transporters and splice variant are downregulated shortly following the insult, which then precipitates glutamate-mediated excitotoxic conditions. Lack of success with glutamate receptor antagonists as a potential source of clinical intervention treatment following TBI has resulted in the necessity for a better understanding of the mechanisms that underlie the process of excitotoxicity, including the function and regulation of glutamate transporters. Such new insight should improve the likelihood of development of novel avenues for therapeutic intervention following TBI.

Alzheimer type II astrocytic changes following sub-acute exposure to manganese and its prevention by antioxidant treatment.

Exposure to manganese in an industrial or clinical setting can lead to manganism, a neurological disorder with similarities to Parkinson's disease. Although the pathogenetic basis of this disorder is unclear, studies indicate this metal is highly accumulated in astrocytes, suggesting an involvement of these glial cells. To investigate this issue, we have used a recently characterized, sub-acute model of manganese neurotoxicity. Treatment of rats with manganese (II) chloride (50 mg/kg body weight, i.p.) once daily for 1 or 4 days led to increases in manganese levels of up to 232, 523, and 427% in the cerebral cortex, globus pallidus, and cerebellum, respectively, by instrumental neutron activation analysis. These changes were accompanied by development of pathological changes in glial morphology identified as Alzheimer type II astrocytosis in both cortical and sub-cortical structures. Co-treatment with either the antioxidant N-acetylcysteine or the manganese chelator 1,2-cyclohexylenedinitrilotetraacetic acid completely blocked this pathology, indicating the cellular transformation may be mediated by oxidative stress associated with the presence of this metal. These findings represent, to our knowledge, the first report of early induction of this pathological hallmark of manganese neurotoxicity, an event previously considered a consequence of chronic exposure to manganese in primates and in human cases of manganism. Our results also indicate that use of this rodent model may provide a novel opportunity to examine the nature and role of the Alzheimer type II astrocyte in the pathophysiology of this disorder as well as in other disease processes in which cerebral accumulation of manganese occurs.

N-acetylcysteine attenuates early induction of heme oxygenase-1 following traumatic brain injury.

Traumatic brain injury (TBI) results in a cascade of events that includes the production of reactive oxygen species. Heme oxygenase-1 (HO-1) is induced in glial cells following head trauma, suggestive of oxidative stress. We have studied the temporal and spatial effects of the antioxidant N-acetylcysteine (NAC) on HO-1 levels following lateral fluid-percussion injury by immunoblotting and immunohistochemistry. In the injured cerebral cortex, maximal HO-1 induction was seen 6 h post-TBI and was maintained for up to 24 h following the insult, while the ipsilateral hippocampus and thalamus showed marked induction at 24 h postinjury. In all three brain regions, little or no HO-1 immunoreactivity was observed on the contralateral side. Astrocytes exhibited positive immunoreactivity for HO-1 in the injured cerebral cortex, hippocampus, and thalamus, while some neurons and microglia were also immunoreactive in the injured cortex. The administration of NAC 5 min following TBI resulted in a marked reduction in this widespread induction of HO-1, concomitant with a decrease in the volume of injury in all three brain regions. Together, these findings indicate that HO-1 induction is related to both oxidative and injury characteristics of the affected tissue, suggesting that protein expression of this gene is a credible marker of oxidative damage in this model of TBI.

Therapeutic effect of anti CEACAM6 monoclonal antibody against lung adenocarcinoma by enhancing anoikis sensitivity.

Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) plays a crucial role in tumorigenesis of lung cancer. However, the therapeutic potential for anti CEACAM6 monoclonal antibody (mAb) has only been limitedly explored. Here, we evaluate the therapeutic potential of naked anti CEACAM6 mAb against lung adenocarcinoma. Clone 8F5, recognizing B domain of CEACAM6, is established by immunizing A549 cells and screening for clones double positive for A549 and CEACAM6-Fc recombinant protein. We found that 85.7% of 70 resected lung adenocarcinoma tissue sections were positive for CEACAM6, whereas all squamous cell carcinoma examined were negative. A549 cells with high levels of CEACAM6 demonstrated more aggressive growth nature and showed increased paclitaxel chemosensitivity upon 8F5 binding. Treatment with 8F5 to A549 decreased cellular CEACAM6 expression and reversed anoikis resistance. 8F5 also decreased cellular status of Akt phosphorylation and increased apoptosis via caspase activation. In a mouse model of lung adenocarcinoma with xenotransplanted A549 cells, 8F5 treatment alone demonstrated 40% tumor growth inhibition. When combined with paclitaxel treatment, 8F5 markedly enhanced tumor growth inhibition, up to 80%. In summary, we demonstrate that anti CEACAM6 mAb is an effective therapeutic treatment for lung adenocarcinoma whose effect is further enhanced by combined treatment with paclitaxel.

Temporal patterns of cortical proliferation of glial cell populations after traumatic brain injury in mice.

TBI (traumatic brain injury) triggers an inflammatory cascade, gliosis and cell proliferation following cell death in the pericontusional area and surrounding the site of injury. In order to better understand the proliferative response following CCI (controlled cortical impact) injury, we systematically analyzed the phenotype of dividing cells at several time points post-lesion. C57BL/6 mice were subjected to mild to moderate CCI over the left sensory motor cortex. At different time points following injury, mice were injected with BrdU (bromodeoxyuridine) four times at 3-h intervals and then killed. The greatest number of proliferating cells in the pericontusional region was detected at 3 dpi (days post-injury). At 1 dpi, NG2+ cells were the most proliferative population, and at 3 and 7 dpi the Iba-1+ microglial cells were proliferating more. A smaller, but significant number of GFAP+ (glial fibrillary acidic protein) astrocytes proliferated at all three time points. Interestingly, at 3 dpi we found a small number of proliferating neuroblasts [DCX+ (doublecortin)] in the injured cortex. To determine the cell fate of proliferative cells, mice were injected four times with BrdU at 3 dpi and killed at 28 dpi. Approximately 70% of proliferative cells observed at 28 dpi were GFAP+ astrocytes. In conclusion, our data suggest that the specific glial cell types respond differentially to injury, suggesting that each cell type responds to a specific pattern of growth factor stimulation at each time point after injury.

Receptor protein tyrosine phosphatase σ binds to neurons in the adult mouse brain.

The role of type IIA receptor protein tyrosine phosphatases (RPTPs), which includes LAR, RPTPσ and RPTPδ, in the nervous system is becoming increasingly recognized. Evidence supports a significant role for these RPTPs during the development of the nervous system as well as after injury, and mutations in RPTPs are associated with human disease. However, a major open question is the nature of the ligands that interact with type IIA RPTPs in the adult brain. Candidates include several different proteins as well as the glycosaminoglycan chains of proteoglycans. In order to investigate this problem, we used a receptor affinity probe assay with RPTPσ-AP fusion proteins on sections of adult mouse brain and to cultured neurons. Our results demonstrate that the major binding sites for RPTPσ in adult mouse brain are on neurons and are not proteoglycan GAG chains, as RPTPσ binding overlaps with the neuronal marker NeuN and was not significantly altered by treatments which eliminate chondroitin sulfate, heparan sulfate, or both. We also demonstrate no overlap of binding of RPTPσ with perineuronal nets, and a unique modulation of RPTPσ binding to brain by divalent cations. Our data therefore point to neuronal proteins, rather than CSPGs, as being the ligands for RPTPσ in the adult, uninjured brain.

Characterization of Two Novel mAbs Recognizing Different Epitopes on CD43.

JL1, a specific epitope on CD43, is a potential biomarker for the diagnosis of acute leukemia. Although qualitative assays for detecting leukemia-specific CD43 exist, there is a need to develop quantitative assays for the same. Here, we developed two novel monoclonal antibodies (mAbs), 2C8 and 8E10, recognizing different epitopes on CD43. These clones are capable of pairing with YG5, another mAb against JL1 epitope, because they were selectively obtained using sandwich ELISA. Antigens recognized by 2C8 and 8E10 were confirmed as CD43 by western blotting using the CD43-hFC recombinant protein. When expression on various leukemic cell lines was investigated, 2C8 and 8E10 displayed a disparity in the distribution of the epitope. Enzyme assays revealed that these mAbs recognized a sialic acid-dependent epitope on CD43. Using normal thymus and lymph node paraffin-embedded tissues, we confirmed a difference in the epitopes recognized by the two mAbs that was predicted based on the maturity of the cells in the tissue. In summary, we developed and characterized two mAbs, 2C8 and 8E10, which can be used with YG5 in a sandwich ELISA for detecting leukemia-specific CD43.

An epigenomic roadmap to induced pluripotency reveals DNA methylation as a reprogramming modulator.

Reprogramming of somatic cells to induced pluripotent stem cells involves a dynamic rearrangement of the epigenetic landscape. To characterize this epigenomic roadmap, we have performed MethylC-seq, ChIP-seq (H3K4/K27/K36me3) and RNA-Seq on samples taken at several time points during murine secondary reprogramming as part of Project Grandiose. We find that DNA methylation gain during reprogramming occurs gradually, while loss is achieved only at the ESC-like state. Binding sites of activated factors exhibit focal demethylation during reprogramming, while ESC-like pluripotent cells are distinguished by extension of demethylation to the wider neighbourhood. We observed that genes with CpG-rich promoters demonstrate stable low methylation and strong engagement of histone marks, whereas genes with CpG-poor promoters are safeguarded by methylation. Such DNA methylation-driven control is the key to the regulation of ESC-pluripotency genes, including Dppa4, Dppa5a and Esrrb. These results reveal the crucial role that DNA methylation plays as an epigenetic switch driving somatic cells to pluripotency.

Heritabilities of facial measurements and their latent factors in korean families.

Genetic studies on facial morphology targeting healthy populations are fundamental in understanding the specific genetic influences involved; yet, most studies to date, if not all, have been focused on congenital diseases accompanied by facial anomalies. To study the specific genetic cues determining facial morphology, we estimated familial correlations and heritabilities of 14 facial measurements and 3 latent factors inferred from a factor analysis in a subset of the Korean population. The study included a total of 229 individuals from 38 families. We evaluated a total of 14 facial measurements using 2D digital photographs. We performed factor analysis to infer common latent variables. The heritabilities of 13 facial measurements were statistically significant (p < 0.05) and ranged from 0.25 to 0.61. Of these, the heritability of intercanthal width in the orbital region was found to be the highest (h (2) = 0.61, SE = 0.14). Three factors (lower face portion, orbital region, and vertical length) were obtained through factor analysis, where the heritability values ranged from 0.45 to 0.55. The heritability values for each factor were higher than the mean heritability value of individual original measurements. We have confirmed the genetic influence on facial anthropometric traits and suggest a potential way to categorize and analyze the facial portions into different groups.

Alterations in sulfated chondroitin glycosaminoglycans following controlled cortical impact injury in mice.

Chondroitin sulfate proteoglycans (CSPGs) play a pivotal role in many neuronal growth mechanisms including axon guidance and the modulation of repair processes following injury to the spinal cord or brain. Many actions of CSPGs in the central nervous system (CNS) are governed by the specific sulfation pattern on the glycosaminoglycan (GAG) chains attached to CSPG core proteins. To elucidate the role of CSPGs and sulfated GAG chains following traumatic brain injury (TBI), controlled cortical impact injury of mild to moderate severity was performed over the left sensory motor cortex in mice. Using immunoblotting and immunostaining, we found that TBI resulted in an increase in the CSPGs neurocan and NG2 expression in a tight band surrounding the injury core, which overlapped with the presence of 4-sulfated CS GAGs but not with 6-sulfated GAGs. This increase was observed as early as 7 days post injury (dpi), and persisted for up to 28 dpi. Labeling with markers against microglia/macrophages, NG2+ cells, fibroblasts, and astrocytes showed that these cells were all localized in the area, suggesting multiple origins of chondroitin-4-sulfate increase. TBI also caused a decrease in the expression of aggrecan and phosphacan in the pericontusional cortex with a concomitant reduction in the number of perineuronal nets. In summary, we describe a dual response in CSPGs whereby they may be actively involved in complex repair processes following TBI.

Genome-wide linkage analysis for ocular and nasal anthropometric traits in a Mongolian population.

Anthropometric traits for eyes and nose are complex quantitative traits influenced by genetic and environmental factors. To date, there have been few reports on the contribution of genetic influence to these traits in Asian populations. The aim of this study was to determine the genetic effect and quantitative trait locus (QTL) of seven traits eyes- and nose-related anthropometric measurements in an isolated Mongolian population. Frontal and lateral photographs were obtained from 1,014 individuals (434 males and 580 females) of Mongolian origin. A total of 349 short tandem repeat markers on 22 autosomes were genotyped for each individual. Heritability estimates of the seven ocular and nasal traits, adjusted for significant covariates, ranged from 0.48 to 0.90, providing evidence for a genetic influence. Variance-component linkage analyses revealed 10 suggestive linkage signals on 5q34 (LOD=3.2), 18q12.2 (LOD=2.7), 5q15 (LOD=2.0), 9q34.2 (LOD=1.9), 5q34 (LOD=1.9), 17q22 (LOD=1.9), 13q33.3 (LOD=2.7), 1q36.22 (LOD=1.9), 4q32.1 (LOD=2.1) and 15q22.31 (LOD=2.9). Our study provides the first evidence that genetics influences nasal and ocular traits in a Mongolian population. Additional collaborative efforts will further extend our understanding of the link between genetic factors and human anthropometric traits.

PPARgamma agonist rosiglitazone is neuroprotective after traumatic brain injury via anti-inflammatory and anti-oxidative mechanisms.

Peroxisome proliferator-activated receptor (PPAR)-gamma is a ligand-activated transcription factor of nuclear hormone receptor superfamily. Thiazolidinedione rosiglitazone is a potent agonist of PPARgamma which was shown to induce neuroprotection in animal models of focal ischemia and spinal cord injury. We currently evaluated the therapeutic potential of rosiglitazone (6 mg/kg at 5 min, 6 h and 24 h; i.p.) following controlled cortical impact (CCI)-induced traumatic brain injury (TBI) in adult mice. CCI injury increased the cortical PPARgamma mRNA levels which were further elevated by rosiglitazone treatment. In addition, rosiglitazone treatment significantly decreased the cortical lesion volume measured at 7 days compared to vehicle treatment (by 56+/-7%; p<0.05; n=6/group). Following TBI, the spared cortex of the rosiglitazone group showed significantly less numbers of GSI-B4(+) activated microglia/macrophages and ICAM1(+) capillaries, and curtailed induction of pro-inflammatory genes IL6, MCP1 and ICAM1 compared to vehicle group. Rosiglitazone-treated mice also showed significantly less number of TUNEL(+) apoptotic neurons and curtailed induction of caspase-3 and Bax, compared to vehicle control. In addition, rosiglitazone significantly enhanced the post-TBI expression of the neuroprotective chaperones HSP27, HSP70 and HSP32/HO1, and the anti-oxidant enzymes catalase, Cu/Zn-SOD and Mn-SOD, compared to vehicle. Treatment with GW9662 (a specific PPARgamma antagonist) prevented all the above PPARgamma-mediated actions. Thus, PPARgamma activation confers neuroprotection after TBI by anti-inflammatory, anti-apoptotic and anti-oxidative mechanisms.

Thiazolidinedione class of peroxisome proliferator-activated receptor gamma agonists prevents neuronal damage, motor dysfunction, myelin loss, neuropathic pain, and inflammation after spinal cord injury in adult rats.

Thiazolidinediones (TZDs) are potent synthetic agonists of the ligand-activated transcription factor peroxisome proliferator-activated receptor-gamma (PPARgamma). TZDs were shown to induce neuroprotection after cerebral ischemia by blocking inflammation. As spinal cord injury (SCI) induces massive inflammation that precipitates secondary neuronal death, we currently analyzed the therapeutic efficacy of TZDs pioglitazone and rosiglitazone after SCI in adult rats. Both pioglitazone and rosiglitazone (1.5 mg/kg i.p.; four doses at 5 min and 12, 24, and 48 h) significantly decreased the lesion size (by 57 to 68%, p < 0.05), motor neuron loss (by 3- to 10-fold, p < 0.05), myelin loss (by 66 to 75%, p < 0.05), astrogliosis (by 46 to 61%, p < 0.05), and microglial activation (by 59 to 78%, p < 0.05) after SCI. TZDs significantly enhanced the motor function recovery (at 7 days after SCI, the motor scores were 37 to 45% higher in the TZD groups over the vehicle group; p < 0.05), but the treatment was effective only when the first injection was given by 2 h after SCI. At 28 days after SCI, chronic thermal hyperalgesia was decreased significantly (by 31 to 39%; p < 0.05) in the pioglitazone group compared with the vehicle group. At 6 h after SCI, the pioglitazone group showed significantly less induction of inflammatory genes [interleukin (IL)-6 by 83%, IL-1beta by 87%, monocyte chemoattractant protein-1 by 75%, intracellular adhesion molecule-1 by 84%, and early growth response-1 by 67%] compared with the vehicle group (p < 0.05 in all cases). Pioglitazone also significantly enhanced the post-SCI induction of neuroprotective heat shock proteins and antioxidant enzymes. Pretreatment with a PPARgamma antagonist, 2-chloro-5-nitro-N-phenyl-benzamide (GW9662), prevented the neuroprotection induced by pioglitazone.

Gene expression changes in thalamus and inferior colliculus associated with inflammation, cellular stress, metabolism and structural damage in thiamine deficiency.

Identification of gene expression changes that promote focal neuronal death and neurological dysfunction can further our understanding of the pathophysiology of these disease states and could lead to new pharmacological and molecular therapies. Impairment of oxidative metabolism is a pathogenetic mechanism underlying neuronal death in many chronic neurodegenerative diseases as well as in Wernicke's encephalopathy (WE), a disorder induced by thiamine deficiency (TD). To identify functional pathways that lead to neuronal damage in this disorder, we have examined gene expression changes in the vulnerable thalamus and inferior colliculus of TD rats using Affymetrix Rat Genome GeneChip analysis in combination with gene ontology and functional categorization assessment utilizing the NetAffx GO Mining Tool. Of the 15 927 transcripts analysed, 125 in thalamus and 141 in inferior colliculus were more abundantly expressed in TD rats compared with control animals. In both regions, the major functional categories of transcripts that were increased in abundance after TD were those associated with inflammation (approximately 33%), stress (approximately 20%), cell death and repair ( approximately 26%), and metabolic perturbation (approximately 19%), together constituting approximately 98% of all transcripts up-regulated. These changes occurred against a background of neuronal cell loss and reactive astro- and microgliosis in both structures. Our results indicate that (i) TD produces changes in gene expression that are consistent with the observed dysfunction and pathology, and (ii) similar alterations in expression occur in thalamus and inferior colliculus, brain regions previously considered to differ in pathology. These findings provide important new insight into processes responsible for lesion development in TD, and possibly WE.

Early loss of the glutamate transporter splice-variant GLT-1v in rat cerebral cortex following lateral fluid-percussion injury.

Glutamate transporter proteins are essential for the control of interstitial glutamate levels, with an impairment of their function or levels being a major potential contributor to excitotoxicity. We have investigated the effects of lateral fluid percussion on the levels of the glutamate transporter proteins GLT-1alpha, its splice variant GLT-1v, GLAST, and EAAC1 in the rat in order to evaluate their pathogenetic role in this model of traumatic brain injury (TBI). Immunoblot analysis revealed neuronal loss in the cerebral cortex was accompanied by a 54% decrease in GLT-1v 6 h following the insult which progressed to an 83% loss of the transporter after 24 h. No changes in GLT-1alpha, GLAST, or EAAC1 were observed in this brain region at either time point. GLT-1v content was also decreased by 55% and 68% in the hippocampus and thalamus, respectively, at 6 h post-injury, but recovered fully after 24 h in both brain regions. In contrast, levels of GLT-1alpha were increased in the hippocampus at 6 h and 24 h post-TBI. These alterations in transporter protein content were also confirmed using immunohistochemical methods. Our results show for the first time a pattern of early, dynamic changes in the levels of GLT-1 transporter splice variants in different brain regions in this trauma model. In addition, correlation of GLT-1v levels with both neuronal cell loss and alpha-internexin content in the injured cortex suggests that loss of this novel glutamate transporter may be a key factor in determining cerebral vulnerability following this type of brain injury.