RESUMO
Spirochetes from the Borrelia genus are known to cause diseases in humans, namely Lyme disease and relapsing fever. These organisms are commonly transmitted to humans by arthropod vectors including ticks, mite, and lice. Here, we report the molecular detection of a Borrelia sp. from a Haemaphysalis hystricis Supino tick collected from wildlife in an Orang Asli settlement in Selangor, Malaysia. Phylogenetic analyses of partial 16s rRNA and flaB gene sequences revealed that the Borrelia sp. is closely related to the relapsing fever group borreliae, Borrelia lonestari, Borrelia miyamotoi, and Borrelia theileri, as well as a number of uncharacterized Borrelia sp. from ticks in Portugal and Japan. To our knowledge, this is the first report of a Borrelia sp. detected in H. hystricis, and in Malaysia. The zoonotic potential of this Borrelia sp. merits further investigation.
Assuntos
Borrelia/classificação , Borrelia/isolamento & purificação , Ixodidae/microbiologia , Animais , Borrelia/genética , Flagelina/genética , Malásia , Filogenia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Análise de Sequência de RNA , Sus scrofa/parasitologiaRESUMO
Resting-state fMRI studies have become very important in cognitive neuroscience because they are able to identify BOLD fluctuations in brain circuits involved in motor, cognitive, or perceptual processes without the use of an explicit task. Such approaches have been fruitful when applied to various disordered populations, or to children or the elderly. However, insufficient attention has been paid to the consequences of the loud acoustic scanner noise associated with conventional fMRI acquisition, which could be an important confounding factor affecting auditory and/or cognitive networks in resting-state fMRI. Several approaches have been developed to mitigate the effects of acoustic noise on fMRI signals, including sparse sampling protocols and interleaved silent steady state (ISSS) acquisition methods, the latter being used only for task-based fMRI. Here, we developed an ISSS protocol for resting-state fMRI (rs-ISSS) consisting of rapid acquisition of a set of echo planar imaging volumes following each silent period, during which the steady state longitudinal magnetization was maintained with a train of relatively silent slice-selective excitation pulses. We evaluated the test-retest reliability of intensity and spatial extent of connectivity networks of fMRI BOLD signal across three different days for rs-ISSS and compared it with a standard resting-state fMRI (rs-STD). We also compared the strength and distribution of connectivity networks between rs-ISSS and rs-STD. We found that both rs-ISSS and rs-STD showed high reproducibility of fMRI signal across days. In addition, rs-ISSS showed a more robust pattern of functional connectivity within the somatosensory and motor networks, as well as an auditory network compared with rs-STD. An increased connectivity between the default mode network and the language network and with the anterior cingulate cortex (ACC) network was also found for rs-ISSS compared with rs-STD. Finally, region of interest analysis showed higher interhemispheric connectivity in Heschl's gyri in rs-ISSS compared with rs-STD, with lower variability across days. The present findings suggest that rs-ISSS may be advantageous for detecting network connectivity in a less noisy environment, and that resting-state studies carried out with standard scanning protocols should consider the potential effects of loud noise on the measured networks.
Assuntos
Percepção Auditiva/fisiologia , Mapeamento Encefálico/métodos , Córtex Cerebral/fisiologia , Imageamento por Ressonância Magnética/métodos , Rede Nervosa/fisiologia , Adolescente , Adulto , Mapeamento Encefálico/normas , Córtex Cerebral/diagnóstico por imagem , Imagem Ecoplanar/métodos , Imagem Ecoplanar/normas , Feminino , Humanos , Imageamento por Ressonância Magnética/normas , Masculino , Rede Nervosa/diagnóstico por imagem , Ruído , Reprodutibilidade dos Testes , Descanso , Adulto JovemRESUMO
The current and potential uses of neuroimaging in healthcare and beyond have spurred discussion about the ethical issues related to neuroimaging and neuroimaging research. This study examined the perspectives of neuroimagers on ethical issues in their research and on the ethics review process. One hundred neuroimagers from 13 Canadian neuroscience centers completed an online survey and 35 semi-structured interviews were conducted. Neuroimagers felt that most ethical and social issues identified in the literature were dealt with adequately, well, and even very well by research ethics boards (REBs), but some issues such as incidental findings and transfer of knowledge were problematic. Neuroimagers reported a range of practical problems in the ethics review process. We aimed to gather perspectives from REB on the ethics review process, but insufficient participation by REBs prevented us from reporting their perspectives. Given shortcomings identified by neuroimagers as well as longstanding issues in Canadian ethics governance, we believe that substantial challenges exist in Canadian research ethics governance that jeopardize trust, communication, and the overall soundness of research ethics governance. Neuroimagers and REBs should consider their shared responsibilities in developing guidance to handle issues such as incidental findings, risk assessment, and knowledge transfer.
Assuntos
Atitude do Pessoal de Saúde , Diagnóstico por Imagem/ética , Técnicas de Diagnóstico Neurológico/ética , Revisão da Pesquisa por Pares , Canadá , Feminino , Humanos , Achados Incidentais , Disseminação de Informação , Imageamento por Ressonância Magnética/ética , MasculinoRESUMO
Chromatin is nonrandomly distributed in nuclear space, yet the functional significance of this remains unclear. Here, we make use of transgenes carrying developmentally regulated promoters to study subnuclear gene positioning during the development of Caenorhabditis elegans. We found that small transgenes (copy number ≤50) are randomly distributed in early embryonic nuclei, independent of promoter activity. However, in differentiated tissues, these same transgenes occupied specific subnuclear positions: When promoters are repressed, transgenes are found at the nuclear periphery, whereas active, developmentally regulated promoters are enriched in the nuclear core. The absence of specific transgene positioning in embryonic nuclei does not reflect an absence of proteins that mediate perinuclear sequestration: Embryonic nuclei are able to sequester much larger transgene arrays (copy number 300-500) at the periphery. This size-dependent peripheral positioning of gene arrays in early embryos correlates with the accumulation of heterochromatic marks (H3K9me3 and H3K27me3) on large arrays. Interestingly, depletion of nuclear lamina components caused release of arrays from the nuclear envelope and interfered with their efficient silencing. Our results suggest that developmentally silenced chromatin binds the nuclear lamina in a manner correlated with the deposition of heterochromatic marks. Peripheral sequestration of chromatin may, in turn, support the maintenance of silencing.
Assuntos
Caenorhabditis elegans/genética , Dosagem de Genes/genética , Heterocromatina/metabolismo , Laminas/metabolismo , Membrana Nuclear/metabolismo , Sequências Repetitivas de Ácido Nucleico/genética , Transgenes/genética , Animais , Caenorhabditis elegans/embriologia , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Diferenciação Celular/genética , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/genética , Genes de Helmintos/genética , Modelos Genéticos , Mutação/genética , Análise de Sequência com Séries de Oligonucleotídeos , Regiões Promotoras Genéticas , Processos EstocásticosAssuntos
Corpo Estriado/patologia , Efeitos Tardios da Exposição Pré-Natal/genética , Efeitos Tardios da Exposição Pré-Natal/patologia , Receptores Nicotínicos/genética , Fumar/efeitos adversos , Regiões 3' não Traduzidas/genética , Adolescente , Distribuição de Qui-Quadrado , Feminino , Humanos , Masculino , GravidezRESUMO
In an initial epigenetic characterization of diffuse large B-cell lymphoma (DLBCL), we evaluated the DNA methylation levels of over 500 CpG islands. Twelve CpG islands (AR, CDKN1C, DLC1, DRD2, GATA4, GDNF, GRIN2B, MTHFR, MYOD1, NEUROD1, ONECUT2 and TFAP2A) showed significant methylation in over 85% of tumors. Interestingly, the methylation levels of a CpG island proximal to FLJ21062 differed between the activated B-cell-like (ABC-DLBCL) and germinal center B-cell-like (GCB-DLBCL) subtypes. In addition, we compared the methylation and expression status of 67 genes proximal (within 500 bp) to the methylation assays. We frequently observed that hypermethylated CpG islands are proximal to genes that are expressed at low or undetectable levels in tumors. However, many of these same genes were also poorly expressed in DLBCL tumors where their cognate CpG islands were hypomethylated. Nevertheless, the proportional reductions in BNIP3, MGMT, RBP1, GATA4, IGSF4, CRABP1 and FLJ21062 expression with increasing methylation suggest that epigenetic processes strongly influence these genes. Lastly, the moderate expression of several genes proximal to hypermethylated CpG tracts suggests that DNA methylation assays are not always accurate predictors of gene silencing. Overall, further investigation of the highlighted CpG islands as potential clinical biomarkers is warranted.
Assuntos
Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Linfoma Difuso de Grandes Células B/genética , Pesquisa Biomédica/normas , Ilhas de CpG/genética , Inativação Gênica , Humanos , Proteínas de Neoplasias/genéticaRESUMO
Previous studies in humans have shown the presence of an age-related reduction of hippocampal (HC) volume, as well as the presence of reduced HC volume in psychiatric populations suffering from schizophrenia, depression or post-traumatic stress disorder. Altogether, these data suggested that aging or psychiatric disease can have neurotoxic effects on the hippocampus, and lead to HC atrophy. However, these two sets of findings imply that HC volume in young healthy adults should present less variability than HC volume in older adults and psychiatric populations. In the present study, we assessed HC volume in 177 healthy men and women aged from 18 to 85 years of age. We show that the dispersion around the mean of HC volume is not different in young and older adults, so that 25% of young healthy adults present HC volume as small as the average participants aged 60 to 75 years. This shows that HC volume is as variable in young as in older adults and suggests that smaller HC volume attributed to the aging process in previous studies could in fact represent HC volume determined early in life. We also report that within similar age groups, the percentage of difference in HC volume between the individuals with the smallest HC volume (smallest quartile) and the group average is greater than the percentage of difference reported to exist between psychiatric populations and normal control in recent meta-analyses. Taken together, these results confront the notion of hippocampal atrophy in humans and raise the possibility that pre-determined inter-individual differences in HC volume in humans may determine the vulnerability for age-related cognitive impairments or psychopathology throughout the lifetime.
Assuntos
Envelhecimento , Hipocampo/anatomia & histologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Atrofia/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Despite a preponderance of studies demonstrating gene expression and/or enzymatic activation of calpain and caspase proteases after traumatic brain injury (TBI), no studies have examined the effects of injury magnitude on expression levels of these cell death effectors after TBI. Determination of the degree to which injury severity affects specific expression profiles is critical to understanding the relevant pathways contributing to post-trauma pathology and for developing targeted therapeutics. This investigation tested the hypothesis that different injury magnitudes (1.0, 1.2, and 1.6 mm) cause alterations in the regional and temporal patterns of mRNA expression of calpain-related (calpain-1 and -2, calpastatin) and caspase-related (caspases -3, -8, -9, BID) gene products after cortical impact in rats. Quantitative RT-PCR was used to compare effects of injury severity on mRNA levels in ipsilateral (injured) cortex and hippocampus, 6 h to 5 days post-injury. TBI caused increases in mRNA expression of all proteins examined, with the highest expression detected in the cortex. Generally, injury magnitude and levels of gene expression were positively correlated. High levels of gene induction were observed with BID, caspase-3, and -8, while caspase-9 mRNA had the lowest level of induction. Interestingly, although calpains are activated within minutes of TBI, calpain mRNA expression was highest 72 h to 5 days post-TBI. This study is the first analysis of the regional and temporal expression of calpains and caspases after TBI. These data provide insight into the inter-relationship of these two protease families and on the distinct but overlapping cascades of cell death after TBI.
Assuntos
Lesões Encefálicas/genética , Calpaína/genética , Caspases/genética , Regulação da Expressão Gênica/fisiologia , RNA Mensageiro/biossíntese , Animais , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3 , Lesões Encefálicas/metabolismo , Lesões Encefálicas/fisiopatologia , Calpaína/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Caspases/metabolismo , Masculino , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ativação TranscricionalRESUMO
Increases in intracellular calcium and subsequent activation of calcium-activated proteases (e.g., calpains) may play a critical role in central nervous system injury. Several studies have implicated calpain activation following subarachnoid hemorrhage (SAH). This study evaluated the effect of a calpain inhibitor administration following SAH in the rat on behavioral deficits (postinjury days 1-5, employing a battery of well-characterized assessment tasks), and blood-brain barrier permeability changes (48 h post-SAH, quantifying the microvascular alterations according to the extravasation of protein-bound Evans Blue using a spectrophotofluorimetric technique). Rats were injected with 400 microl of autologous blood into the cisterna magna to induce SAH. Within 5 min after the surgical procedure, Calpain Inhibitor II or vehicle was continuously administered intravenously for 2 days. Results indicated that Calpain Inhibitor II treatment after SAH significantly improved (a) beam balance time (day 1, p < 0.05), but not beam balance score, (b) latency to traverse the beam on days 1-4 (day 1-3, p < 0.001; day 4, p < 0.01), and (c) loss in body weight on days 4-5 (p < 0.05). Evans Blue dye extravasation was significantly less in SAH Calpain Inhibitor II-treated rats compared to SAH vehicle-treated rats in seven out of the eight brain regions studied (p < 0.001, 0.01, and 0.05). These results suggest that pharmacological inhibition of a relatively selective, membrane-permeant calpain inhibitor can significantly reduce some pathophysiological SAH consequences, and indicate that the inhibition of calpain may be a beneficial therapeutic approach to reduce post-SAH global brain dysfunction.
Assuntos
Sintomas Comportamentais/tratamento farmacológico , Barreira Hematoencefálica/efeitos dos fármacos , Calpaína/antagonistas & inibidores , Oligopeptídeos/administração & dosagem , Hemorragia Subaracnóidea/tratamento farmacológico , Animais , Sintomas Comportamentais/enzimologia , Sintomas Comportamentais/fisiopatologia , Barreira Hematoencefálica/fisiologia , Peso Corporal/efeitos dos fármacos , Peso Corporal/fisiologia , Masculino , Destreza Motora/efeitos dos fármacos , Destreza Motora/fisiologia , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Hemorragia Subaracnóidea/enzimologia , Hemorragia Subaracnóidea/fisiopatologia , Hemorragia Subaracnóidea/psicologiaRESUMO
The contributions of calpain and caspase-3 to apoptosis and necrosis after central nervous system (CNS) trauma are relatively unexplored. No study has examined concurrent activation of calpain and caspase-3 in necrotic or apoptotic cell death after any CNS insult. Experiments used a model of oxygen-glucose deprivation (OGD) in primary septo-hippocampal cultures and assessed cell viability, occurrence of apoptotic and necrotic cell death phenotypes, and protease activation. Immunoblots using an antibody detecting calpain and caspase-3 proteolysis of alpha-spectrin showed greater accumulation of calpain-mediated breakdown products (BDPs) compared with caspase-3-mediated BDPs. Administration of calpain and caspase-3 inhibitors confirmed that activation of these proteases contributed to cell death, as inferred by lactate dehydrogenase release. Oxygen-glucose deprivation resulted in expression of apoptotic and necrotic cell death phenotypes, especially in neurons. Immunocytochemical studies of calpain and caspase-3 activation in apoptotic cells indicated that these proteases are almost always concurrently activated during apoptosis. These data demonstrate that calpain and caspase-3 activation is associated with expression of apoptotic cell death phenotypes after OGD, and that calpain activation, in combination with caspase-3 activation, could contribute to the expression of apoptotic cell death by assisting in the degradation of important cellular proteins.
Assuntos
Calpaína/metabolismo , Caspases/metabolismo , Glucose/farmacologia , Neurônios/enzimologia , Oxigênio/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Western Blotting , Calpaína/antagonistas & inibidores , Caspase 3 , Inibidores de Caspase , Células Cultivadas , Ativação Enzimática/fisiologia , Inibidores Enzimáticos/farmacologia , Infecções por Fusobacterium , Hipocampo/citologia , Neuroglia/citologia , Neuroglia/efeitos dos fármacos , Neuroglia/enzimologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Fenótipo , Ratos , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/patologia , Formação de Roseta , Septo do Cérebro/citologia , Espectrina/metabolismo , Acidente Vascular Cerebral/enzimologia , Acidente Vascular Cerebral/patologiaRESUMO
Although a number of increased CSF proteins have been correlated with brain damage and outcome after traumatic brain injury (TBI), a major limitation of currently tested biomarkers is a lack of specificity for defining neuropathological cascades. Identification of surrogate biomarkers that are elevated in CSF in response to brain injury and that offer insight into one or more pathological neurochemical events will provide critical information for appropriate administration of therapeutic compounds for treatment of TBI patients. Non-erythroid alpha II-spectrin is a cytoskeletal protein that is a substrate of both calpain and caspase-3 cysteine proteases. As we have previously demonstrated, cleavage of alpha II-spectrin by calpain and caspase-3 results in accumulation of protease-specific spectrin breakdown products (SBDPs) that can be used to monitor the magnitude and temporal duration of protease activation. However, accumulation of alpha II-spectrin and alpha II-SBDPs in CSF after TBI has never been examined. Following a moderate level (2.0 mm) of controlled cortical impact TBI in rodents, native alpha II-spectrin protein was decreased in brain tissue and increased in CSF from 24 h to 72 h after injury. In addition, calpain-specific SBDPs were observed to increase in both brain and CSF after injury. Increases in the calpain-specific 145 kDa SBDP in CSF were 244%, 530% and 665% of sham-injured control animals at 24 h, 48 h and 72 h after TBI, respectively. The caspase-3-specific SBDP was observed to increase in CSF in some animals but to a lesser degree. Importantly, levels of these proteins were undetectable in CSF of uninjured control rats. These results indicate that detection of alpha II-spectrin and alpha II-SBDPs is a powerful discriminator of outcome and protease activation after TBI. In accord with our previous studies, results also indicate that calpain may be a more important effector of cell death after moderate TBI than caspase-3.
Assuntos
Lesões Encefálicas/líquido cefalorraquidiano , Calpaína/fisiologia , Espectrina/líquido cefalorraquidiano , Ferimentos não Penetrantes/líquido cefalorraquidiano , Animais , Calpaína/metabolismo , Caspase 3 , Caspases/metabolismo , Córtex Cerebral/metabolismo , Eritrócitos/metabolismo , Masculino , Peptídeo Hidrolases/metabolismo , Isoformas de Proteínas/sangue , Isoformas de Proteínas/líquido cefalorraquidiano , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Ratos , Ratos Sprague-Dawley , Espectrina/química , Espectrina/metabolismoRESUMO
Motivated by the vast amount of information that is rapidly accumulating about the human brain in digital form, we embarked upon a program in 1992 to develop a four-dimensional probabilistic atlas and reference system for the human brain. Through an International Consortium for Brain Mapping (ICBM) a dataset is being collected that includes 7000 subjects between the ages of eighteen and ninety years and including 342 mono- and dizygotic twins. Data on each subject includes detailed demographic, clinical, behavioural and imaging information. DNA has been collected for genotyping from 5800 subjects. A component of the programme uses post-mortem tissue to determine the probabilistic distribution of microscopic cyto- and chemoarchitectural regions in the human brain. This, combined with macroscopic information about structure and function derived from subjects in vivo, provides the first large scale opportunity to gain meaningful insights into the concordance or discordance in micro- and macroscopic structure and function. The philosophy, strategy, algorithm development, data acquisition techniques and validation methods are described in this report along with database structures. Examples of results are described for the normal adult human brain as well as examples in patients with Alzheimer's disease and multiple sclerosis. The ability to quantify the variance of the human brain as a function of age in a large population of subjects for whom data is also available about their genetic composition and behaviour will allow for the first assessment of cerebral genotype-phenotype-behavioural correlations in humans to take place in a population this large. This approach and its application should provide new insights and opportunities for investigators interested in basic neuroscience, clinical diagnostics and the evaluation of neuropsychiatric disorders in patients.
Assuntos
Mapeamento Encefálico/métodos , Encéfalo/anatomia & histologia , Bases de Dados Factuais , Neuroanatomia/métodos , Adulto , Mapeamento Encefálico/instrumentação , Humanos , Imageamento por Ressonância Magnética , Modelos Estatísticos , Neuroanatomia/instrumentaçãoRESUMO
The authors describe the development of a four-dimensional atlas and reference system that includes both macroscopic and microscopic information on structure and function of the human brain in persons between the ages of 18 and 90 years. Given the presumed large but previously unquantified degree of structural and functional variance among normal persons in the human population, the basis for this atlas and reference system is probabilistic. Through the efforts of the International Consortium for Brain Mapping (ICBM), 7,000 subjects will be included in the initial phase of database and atlas development. For each subject, detailed demographic, clinical, behavioral, and imaging information is being collected. In addition, 5,800 subjects will contribute DNA for the purpose of determining genotype- phenotype-behavioral correlations. The process of developing the strategies, algorithms, data collection methods, validation approaches, database structures, and distribution of results is described in this report. Examples of applications of the approach are described for the normal brain in both adults and children as well as in patients with schizophrenia. This project should provide new insights into the relationship between microscopic and macroscopic structure and function in the human brain and should have important implications in basic neuroscience, clinical diagnostics, and cerebral disorders.
Assuntos
Anatomia Artística , Anatomia Transversal , Encéfalo/anatomia & histologia , Bases de Dados Factuais , Ilustração Médica , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Algoritmos , Humanos , Aumento da Imagem/métodos , Imageamento Tridimensional , Imageamento por Ressonância Magnética , Pessoa de Meia-Idade , Neuroanatomia/métodos , Probabilidade , Esquizofrenia/patologiaRESUMO
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.
Assuntos
Apoptose/fisiologia , Lesões Encefálicas/metabolismo , Caspases/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Animais , Caspase 3 , Caspase 8 , Caspase 9 , Córtex Cerebral/química , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Modelos Animais de Doenças , Immunoblotting , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Masculino , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
This review focuses on the maturation of brain white-matter, as revealed by magnetic resonance (MR) imaging carried out in healthy subjects. The review begins with a brief description of the nature of the MR signal and its possible biological underpinnings, and proceeds with a description of MR findings obtained in newborns, infants, children and adolescents. On MR images, a significant decrease in water content leads to a decrease of longitudinal relaxation times (T1) and transverse relaxation times (T2) and consequent "adult-like" appearance of T1-weighted and T2-weighted images becomes evident towards the end of the first year of life. Owing to the onset of myelination and the related increase of lipid content, MR images gradually acquire an exquisite grey-white matter contrast in a temporal sequence reflecting the time course of myelination. Albeit less pronounced, age-related changes in white matter continue during childhood and adolescence; white matter increases its overall volume and becomes more myelinated in a region-specific fashion. Detection of more subtle changes during this "late" phase of brain development is greatly aided by computational analyses of MR images. The review also briefly outlines future directions, including the use of novel MR techniques such as diffusion tensor imaging and magnetization transfer, as well as the suggestion for the concurrent use of experimental behavioral test-batteries, with structural MR imaging, to study developmental changes in structure-function relationships.
Assuntos
Encéfalo/anatomia & histologia , Encéfalo/crescimento & desenvolvimento , Imageamento por Ressonância Magnética/métodos , Fibras Nervosas Mielinizadas/ultraestrutura , Adolescente , Fatores Etários , Encéfalo/metabolismo , Córtex Cerebral/citologia , Córtex Cerebral/crescimento & desenvolvimento , Córtex Cerebral/metabolismo , Criança , Pré-Escolar , Humanos , Lactente , Recém-Nascido , Imageamento por Ressonância Magnética/tendências , Fibras Nervosas Mielinizadas/metabolismo , Testes Neuropsicológicos , Desempenho Psicomotor/fisiologiaRESUMO
Primary septo-hippocampal cell cultures were incubated in varying concentrations of tumor necrosis factor (TNF-alpha; 0.3-500 ng/ml) to examine proteolysis of the cytoskeletal protein alpha-spectrin (240 kDa) to a signature 145 kDa fragment by calpain and to the apoptotic-linked 120-kDa fragment by caspase-3. The effects of TNF-alpha incubation on morphology and cell viability were assayed by fluorescein diacetate-propidium iodide (FDA-PI) staining, assays of lactate dehydrogenase (LDH) release, nuclear chromatin alterations (Hoechst 33258), and internucleosomal DNA fragmentation. Incubation with varying concentrations of TNF-alpha produced rapid increases in LDH release and nuclear PI uptake that were sustained over 48 hr. Incubation with 30 ng/ml TNF-alpha yielded maximal, 3-fold, increase in LDH release and was associated with caspase-specific 120-kDa fragment but not calpain-specific 145-kDa fragment as early as 3.5 hr after injury. Incubation with the pan-caspase inhibitor, carbobenzosy- Asp-CH(2)-OC (O)-2-6-dichlorobenzene (Z-D-DCB, 50-140 microM) significantly reduced LDH release produced by TNF-alpha. Apoptotic-associated oligonucleosomal-sized DNA fragmentation on agarose gels was detected from 6 to 72 hr after exposure to TNF-alpha. Histochemical changes included chromatin condensation, nuclear fragmentation, and formation of apoptotic bodies. Results of this study suggest TNF-alpha may induce caspase-3 activation but not calpain activation in septo-hippocampal cultures and that this activation of caspase-3 at least partially contributes to TNF-alpha-induced apoptosis.
Assuntos
Apoptose/efeitos dos fármacos , Caspases/metabolismo , Hipocampo/efeitos dos fármacos , Proteínas do Tecido Nervoso/metabolismo , Neurônios/efeitos dos fármacos , Septo Pelúcido/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Animais , Ácido Aspártico/análogos & derivados , Ácido Aspártico/farmacologia , Calpaína/análise , Caspase 3 , Inibidores de Caspase , Células Cultivadas/efeitos dos fármacos , Cicloeximida/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Fragmentação do DNA , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Hipocampo/citologia , Proteínas do Tecido Nervoso/antagonistas & inibidores , Neurônios/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Ratos , Septo Pelúcido/citologia , Espectrina/metabolismoRESUMO
Forkhead-associated (FHA) domains are multifunctional phosphopeptide-binding modules and are the hallmark of the conserved family of Rad53-like checkpoint protein kinases. Rad53-like kinases, including the human tumor suppressor protein Chk2, play crucial roles in cell cycle arrest and activation of repair processes following DNA damage and replication blocks. Here we show that ectopic expression of the N-terminal FHA domain (FHA1) of the yeast Rad53 kinase causes a growth defect by arresting the cell cycle in G(1). This phenotype was highly specific for the Rad53-FHA1 domain and not observed with the similar Rad53-FHA2, Dun1-FHA, and Chk2-FHA domains, and it was abrogated by mutations that abolished binding to a phosphothreonine-containing peptide in vitro. Furthermore, replacement of the RAD53 gene with alleles containing amino acid substitutions in the FHA1 domain resulted in an increased DNA damage sensitivity in vivo. Taken together, these data demonstrate that the FHA1 domain contributes to the checkpoint function of Rad53, possibly by associating with a phosphorylated target protein in response to DNA damage in G(1).
Assuntos
Proteínas de Ciclo Celular , Ciclo Celular , Proteínas Nucleares/química , Proteínas Quinases/química , Proteínas Quinases/fisiologia , Proteínas Serina-Treonina Quinases , Proteínas de Saccharomyces cerevisiae , Fatores de Transcrição/química , Alelos , Proteínas de Bactérias/metabolismo , Divisão Celular , Quinase do Ponto de Checagem 2 , Dano ao DNA/genética , Citometria de Fluxo , Fatores de Transcrição Forkhead , Immunoblotting , Modelos Moleculares , Mutagênese Sítio-Dirigida , Mutação , Proteínas Nucleares/genética , Peptídeos/química , Fenótipo , Fosforilação , Proteínas Quinases/genética , Estrutura Terciária de Proteína , Proteínas Recombinantes/metabolismo , Fatores de Tempo , Fatores de Transcrição/genética , Leveduras/metabolismoRESUMO
In unselected patients with intractable temporal lobe epilepsy (TLE), approximately 15% do not have detectable hippocampal atrophy on MRI. The purpose of this study was to evaluate whether T2 relaxometry can identify hippocampal pathology and lateralize the epileptic focus in patients with intractable TLE, who do not demonstrate hippocampal atrophy on volumetric MRI (MRIV). We selected 14 patients with unilateral TLE who had unilateral atrophy and 11 patients with unilateral TLE who had no evidence of atrophy on MRIV. Images were acquired on a 1.5 T MR scan using a dual echo sequence with 23 contiguous oblique coronal slices in all patients and in 14 healthy subjects. Fitting a single exponential decay equation to the imaging data generated T2 maps. Averages of six slices containing the head, body, and tail of the hippocampus were used to calculate hippocampal T2 relaxation times (HT2). The epileptic focus was defined by history, video-EEG, and surgical response. All TLE patients with hippocampal atrophy and 9/11 (82%) patients with normal MRI had abnormally high HT2 ipsilateral to the epileptic focus. Bilateral abnormal HT2 were found in 6/14 (43%) of patients with unilateral hippocampal atrophy and 2/11 (18%) of patients with normal MRI. However, this increase was always greater ipsilateral to the epileptic focus. Qualitative hippocampal pathology showed gliosis and neuronal loss in 10/14 operated patients with hippocampal atrophy on MRIV and in 5/7 operated patients with normal MRI. In conclusion, hippocampal T2 mapping provides evidence of hippocampal damage in the majority of patients with intractable TLE who have no evidence of atrophy on MRI and can correctly lateralize the epileptic focus in most patients.
Assuntos
Dominância Cerebral/fisiologia , Epilepsia do Lobo Temporal/diagnóstico , Hipocampo/patologia , Imageamento por Ressonância Magnética , Adulto , Atrofia , Mapeamento Encefálico , Epilepsia do Lobo Temporal/fisiopatologia , Feminino , Gliose/diagnóstico , Gliose/fisiopatologia , Hipocampo/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Degeneração Neural/diagnóstico , Degeneração Neural/fisiopatologia , Valores de Referência , Lobo Temporal/patologia , Lobo Temporal/fisiopatologiaRESUMO
This study investigated the temporal expression and cell subtype distribution of activated caspase-3 following cortical impact-induced traumatic brain injury in rats. The animals were killed and examined for protein expression of the proteolytically active subunit of caspase-3, p18, at intervals from 6 h to 14 days after injury. In addition, we also investigated the effect of caspase-3 activation on proteolysis of the cytoskeletal protein alpha-spectrin. Increased protein levels of p18 and the caspase-3-specific 120-kDa breakdown product to alpha-spectrin were seen in the cortex ipsilateral to the injury site from 6 to 72 h after the trauma. Immunohistological examinations revealed increased expression of p18 in neurons, astrocytes, and oligodendrocytes from 6 to 72 h following impact injury. In contrast, no evidence of caspase-3 activation was seen in microglia at all time points investigated. Quantitative analysis of caspase-3-positive cells revealed that the number of caspase-3-positive neurons exceeded the number of caspase-3-positive glia cells from 6 to 72 h after injury. Moreover, concurrent assessment of nuclear histopathology using hematoxylin identified p18-immunopositive cells exhibiting apoptotic-like morphological profiles in the cortex ipsilateral to the injury site. In contrast, no evidence of increased p18 expression or alpha-spectrin proteolysis was seen in the ipsilateral hippocampus, contralateral cortex, or hippocampus up to 14 days after the impact. Our results are the first to demonstrate the concurrent expression of activated caspase-3 in different CNS cells after traumatic brain injury in the rat. Our findings also suggest a contributory role of activated caspase-3 in neuronal and glial apoptotic degeneration after experimental TBI in vivo.