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At least 307 records · Page 17Linked to original sources
PubMed2003

mRNA expression patterns of the cGMP-hydrolyzing phosphodiesterases types 2, 5, and 9 during development of the rat brain.

Recent evidence indicates that cGMP plays an important role in neural development and neurotransmission. Since cGMP levels depend critically on the activities of phosphodiesterase (PDE) enzymes, mRNA expression patterns were examined for several key cGMP-hydrolyzing PDEs (type 2 [PDE2], 5 [PDE5], and 9 [PDE9]) in rat brain at defined developmental stages. Riboprobes were used for nonradioactive in situ hybridization on sections derived from embryonic animals at 15 days gestation (E15) and several postnatal stages (P0, P5, P10, P21) until adulthood (3 months). At all stages PDE9 mRNA was present throughout the whole central nervous system, with highest levels observed in cerebellar Purkinje cells, whereas PDE2 and PDE5 mRNA expression was more restricted. Like PDE9, PDE5 mRNA was abundant in cerebellar Purkinje cells, although it was observed only on and after postnatal day 10 in these cells. In other brain regions, PDE5 mRNA expression was minimal, detected in olfactory bulb, cortical layers, and in hippocampus. PDE2 mRNA was distributed more widely, with highest levels in medial habenula, and abundant expression in olfactory bulb, olfactory tubercle, cortex, amygdala, striatum, and hippocampus. Double immunostaining of PDE2, PDE5, or PDE9 mRNAs with the neuronal marker NeuN and the glial cell marker glial fibrillary acidic protein revealed that these mRNAs were predominantly expressed in neuronal cell bodies. Our data indicate that three cGMP-hydrolyzing PDE families have distinct expression patterns, although specific cell types coexpress mRNAs for all three enzymes. Thus, it appears that differential expression of PDE isoforms may provide a mechanism to match cGMP hydrolysis to the functional demands of individual brain regions.

Animals↗
PubMed2007

Low doses of ethanol and hypoxia administered together act synergistically to promote the death of cortical neurons.

The majority of clinically relevant injuries to the developing brain are increasingly being recognized as the result of a combination of least two insults. Evidence from epidemiologic and clinical studies suggests that many of these insults do not cause injury in and of themselves. Rather, they act as predisposing factors (pre-conditioning cytotoxic factors, such as chronic ethanol consumption), increasing the susceptibility to a second, injury-producing (precipitating), unfavorable event (such as hypoxia-ischemia). In this study we make use of primary neuronal cultures to test, in a reproducible system, this two-hit hypothesis. We show that, when administered in combination, low levels of two clinically important environmental insults, hypoxia and ethanol, despite having minimal effects on their own, can act synergistically produce a significant degree of neuronal injury. We further show that one indicator of apoptotic cell death, activated caspase 3, undetectable in neurons subjected to low levels of either insult alone, is detectable when they are administered together.

Animals↗
PubMed2004

Functional restoration of acoustic units and adult-generated neurons after hypothalamic lesion.

The hypothalamus of the adult ring dove contains acoustic units that respond to species-specific coo vocalization. Loss of nest coo leads to unsuccessful breeding. However, the recovery of nest coo in some doves suggests that these units are capable of self-renewal. We have previously shown that lesioning the hypothalamus generates the addition of new neurons at the lesioned area. In this study, we sought to determine whether lesion-induced new neurons are involved in the recovery of coo-responsive units. We systematically recorded electrical activity in the ventromedial nucleus (VMN) of the hypothalamus, before and after lesion, for varying periods up to 3 months. Recordings were made when the birds were at rest (spontaneous discharge) and when the birds were exposed to acoustic stimulations (evoked discharge). Concurrently, the lesioned area was monitored for changes in cell types by using bromodeoxyuridine (BrdU) to label newly divided cells and NeuN to identify mature neurons. For 1 month after lesion, there was no sign of electrical activity, and only BrdU-labeled cells were present. When the first electrical activity occurred, it displayed abnormal spontaneous bursting patterns. The mature discharge patterns (both spontaneous and evoked) occurred after detection of BrdU+/NeuN+ double-labeled cells 2-3 months postlesion and were similar to those found in intact and sham-lesioned birds. Double-labeled cells bore morphologic characteristics of a neuron and were confirmed with z-stack analysis using confocal laser scanning microscopy. Moreover, double-labeled cells were not stained for glial fibrillary acidic protein (GFAP), suggesting that they were neurons. The number of coo-responsive units was significantly correlated with that of BrdU+/NeuN+ cells. Furthermore, the marker for recording sites revealed that coo-responsive units were colocalized with BrdU+/NeuN+ cells. Taken together, the evidence strongly suggests that lesion-induced addition of new neurons promotes the functional recovery of the adult hypothalamus.

Acoustic Stimulation↗
PubMed2005

Differential properties of dentate gyrus and CA1 neural precursors.

In the present article we investigated the properties of CA1 and dentate gyrus cell precursors in adult rodents both in vivo and in vitro. Cell proliferation in situ was investigated by rating the number of cells incorporating BrdU after kainate-induced seizures. CA1 precursors displayed a greater proliferation capacity than dentate gyrus precursors. The majority of BrdU-labeled cells in CA1 expressed Nestin and Mash-1, two markers of neural precursors. BrdU-positive cells in the dentate gyrus expressed Nestin, but only a few expressed Mash-1. In animals pretreated with the antimitotic azacytidine, the capacity of kainate to enhance the proliferation was higher in CA1 than in the dentate gyrus. Differences in intrinsic progenitor cell activity could underlie these different expansion capacities. Thus, we compared the renewal- expansion and multipotency of dentate gyrus and CA1 precursors isolated in vitro. We found that the dissected CA1 region, including the periventricular zone, is enriched in neurosphere-forming cells (presumed stem cells), which respond to either EGF or FGF-2. Dentate gyrus contains fewer neurosphere-forming cells and none that respond to FGF-2 alone. Neurospheres generated from CA1 were multipotent and produced neurons, astrocytes, and oligodendrocytes, while dentate gyrus neurospheres mostly produced glial cells. The analysis of the effects of EGF on organotypic cultures of hippocampal slices depicted similar features: BrdU and Nestin immunoreactivities increased after EGF treatment in CA1 but not in the dentate gyrus. These results suggest that CA1 precursors are more stem-cell-like than granule cell precursors, which may represent a more restricted precursor cell.

Analysis of Variance↗
PubMed1993

Cloning and functional expression of a Schistosoma japonicum cDNA homologous to the enolase gene family.

A cDNA encoding the complete open reading frame of Schistosoma japonicum enolase has been cloned. The 1494bp cDNA (C30) was isolated from a S. japonicum cDNA expression library immunoscreened with hyperimmune rabbit sera raised against soluble adult S. japonicum proteins. The ORF encodes a protein of 434 amino acids exhibiting 72% identity to human, murine, and rat enolases, and 62% identity to Saccharomyces cerevisiae enolase. The inferred molecular mass of the protein is 47,251 Daltons, similar to that reported for the enolases of other species. In vitro translation of C30 also generated a protein of 47kDa. After subcloning and expression, the recombinant protein was purified by affinity chromatography under non-denaturing conditions and shown to exhibit functional enolase enzymatic activity.

Amino Acid Sequence↗
PubMed1994

Time-course curves of enzyme reactions with dilute substrates can be determined from the reverse reactions.

We present a novel and useful principle that can be applied to enzyme reactions with dilute substrates and enables to obtain the desired time-course curve from the reverse reaction when the latter can be measured more easily. Let us imagine an enzyme mixture that can catalyze the interconversion between substances A, B, C,.... If one substance (sufficiently dilute) is added to the mixture as the substrate and the time-course curve of the concentration of another substance is followed, then, "the time-course curve of A-->B" (which is the time-course curve of the concentration of substance B when substance A is added as the substrate) and that of the reverse reaction B-->A will coincide when plotted at appropriate scales of magnitude. Similar relation holds on any pair of substances even if they are widely separated in any type of reaction scheme. The principle was proved mathematically and holds on any first-order reaction network and can be applied on an enzyme reaction with a dilute substrate because it is reduced to a first-order reaction network. An example of experimental application of this principle is also given.

Catalysis↗
PubMed1987

Medulloblastomas in childhood: histological factors influencing patients' outcome.

Sixty-five medulloblastomas in infancy and childhood treated from 1965 through 1981 were reviewed, and the correlation between histological findings of medulloblastomas and clinical course of the patients was studied. Thirty-five patients died but the remaining 30 are alive and without clinical evidence of recurrence 5 years or more after surgery. Certain histological features on light microscopic examinations (e.g., pleomorphism of tumor cells, nuclear-cytoplasmic ratio, mitotic index, degree of vascularity and endothelial proliferation) do influence patient outcome with statistical significance (P less than 0.05). Thirty out of 65 medulloblastomas were examined further, using immunohistochemical methods with glial fibrillary acidic protein (GFAP), neuron specific enolase (NSE), and factor VIII/vW factor (F VIII/vWF). GFAP stain was negative in 20%, NSE stain in 13.3% and F VIII/vWF stain in 16.7% of the medulloblastomas studied. "Desmoplastic" medulloblastomas showed a strong tendency toward positive NSE and GFAP staining in the glomerular portion. There was no correlation between patient outcome and the results of applied immunohistochemical studies. Our data indicate that certain histological features may influence patient outcome, but the degree and pattern of cellular differentiation do not predict outcome.

Adolescent↗
PubMed1984

Immunohistochemistry of folliculo-stellate cells in normal human adenohypophyses and in pituitary adenomas.

Presence and distribution of S-100 protein (S-100), neuron-specific enolase (NSE), cytokeratin polypeptides, glial fibrillary acidic protein (GFAP), vimentin, actin, lysozyme and pituitary hormones (prolactin, hGH, ACTH, beta-FSH, beta-LH, beta-TSH, alpha subunit) in folliculo-stellate cells (FSC) were studied in seven normal human pituitary glands and 28 pituitary adenomas using peroxidase-antiperoxidase and the avidin-biotin immunohistochemical techniques. Approximately 5% of the cells of the adenohypophysis were agranular, non-hormone-producing FSC most of which showed a conspicuous and strong reaction with S-100 antibodies but some were, in addition, GFAP- and vimentin-positive. In contrast to endocrine cells (EC), FSC were not decorated by antibodies to NSE or cytokeratins. In addition to supportive functions, these cells, due to their close special relationship to EC, seem to have transport and other metabolic functions yet to be elucidated. By their S-100 reactivity and their distribution FSC are comparable to glial cells of the central and Schwann and satellite cells of the peripheral nervous system (PNS) as well as to supportive cells in neuroendocrine organs and related tumors (e.g., pheochromocytomas, paragangliomas, carcinoids). With one exception, S-100 reactive FSC were not found in pituitary adenomas. The immunohistochemical demonstration of S-100 protein in pituitary tissue is, therefore, a reliable aid in the discrimination between adenomas and normal pituitary tissue, particularly in small and poorly preserved specimens.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenoma↗
PubMed1987

Selective localization of gamma-enolase in stromal cells of cerebellar hemangioblastomas.

Three cases of cerebellar hemangioblastoma were studied using the immunoperoxidase technique to localize gamma-enolase, also known as neuron-specific enolase. The stromal cells demonstrated positive staining for gamma-enolase, while endothelial cells and pericytes showed no reactivity. Two vascular lesions, an angiosarcoma and a cutaneous angioma, were studied and found to be nonreactive for gamma-enolase. All tumors were also tested for factor VIII/von Willebrand factor, glial fibrillary acidic protein, and the S-100 protein. The lack of expression of gamma-enolase in endothelial cells of hemangioblastomas demonstrates a clear antigenic distinction from neighboring gamma-enolase-positive stromal cells. The significance of this finding and its implications for stromal cell histogenesis are discussed.

Adult↗
PubMed1993

Biochemical correlates to cortical dysplasia, gliosis, and astrocytoma infiltration in human epileptogenic cortex.

The study provides detailed biochemical correlates to the common histopathological diagnoses in epilepsy. A dot immunobinding procedure was used for quantification of NSE, GFA, S-100, NCAM, NF 68 and NF 200. The material consisted of samples from 48 patients either selected for surgical treatment of partial epilepsy or for disorders not related to epilepsy. The histopathological diagnosis of the epileptic cases was: MCD (mild cortical dysplasia, microdysgenesis), gliosis, astrocytoma, ganglioglioma, oligodendroglioma and single cases. The concentration in non-epileptic white matter, in per cent of that in grey matter was: NSE, 85; GFA, 175; S-100, 117; NCAM, 43; NF 68,227 and NF 200, 173. The concentration of NSE as well as of GFA was close to normal in the specimens of the MCD and gliosis groups and of one subgroup of the astrocytomas. There was a striking inverse relationship of the GFA vs the NSE concentrations in the whole material. The concentrations of S-100 showed no such inverse relationship to NSE levels. In all the epileptic groups, total NCAM was lower than 50% of that of the non-epileptic group. The mean NF 68 and NF 200 concentration in the gliosis and astrocytoma groups was 75% of that of the non-epileptic group while the corresponding value for the MCD group was 50%. There was a positive correlation of immunochemically determined GFA and the histopathological gliosis score in the samples of epileptogenic cortex. There was no correlation between the concentration of GFA in the samples and the duration of epilepsy.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1991

A primary encephalocele culture yields a pure population of human astrocytes.

In order to develop a reliable source of human astrocytes for in vitro studies, we established a primary explant culture of a human encephalocele. This culture yielded a population of cells which were karyotypically normal, morphologically resembled astrocytes, expressed glial fibrillary acid protein, and responded mitogenically to exogenous growth factors. We conclude that white matter derived from human encephaloceles can be used to generate pure populations of normal astrocytes.

Astrocytes↗
PubMed1992

Presence of typical neuronal markers in serially cultured cells from adult human brain.

Typical markers for neurons but not for astroglia have been identified in cells cultured from a sample of normal adult human temporal lobe, which was removed to gain access to a glioma. Cells were grown in medium containing growth factors, including fibroblast growth factor and nerve growth factor. The cells grew slowly (doubling time, 18 days) and have been carried as far as passage 8 over 10 months. Both immunoblotting and immunocytochemistry with redundant antibodies demonstrated the presence of neurofilaments (NF-H, NF-M, NF-L), but not glial fibrillary acidic protein (GFAP). Neuron-specific enolase (NSE) was also found. Morphologically, the cultures consisted of a pleimorphic population of cells with frequent long processes. Cells demonstrating neuronal rather than astroglial markers can be cultured from normal adult human brain.

Biomarkers↗
PubMed1988

Third ventricular primary cerebral neuroblastoma. Electron-microscopic and immunohistochemical study.

A case of third ventricular primary cerebral neuroblastoma with secondary hydrocephalus is reported. Light microscopy showed a cell pattern that resembled either ependymoma or oligodendroglioma. The tumor was confirmed to be neuroblastoma by electron microscopy and immunohistochemistry. Immunoperoxidase staining was positive for neuron-specific enolase and negative for glial fibrillary acidic protein.

Adolescent↗
PubMed1992

Periventricular neurocytoma: a pathological entity.

Three cases of periventricular neurocytomas are presented. All patients had a large but well-circumscribed, hyperdense tumor with insignificant contrast enhancement in the lateral and third ventricle, causing hydrocephalus. Calcification was present in one patient. Angiography revealed a blush tumor enhancement in two cases. Surgical removal was complete in two patients and incomplete in one. Light microscopy showed a cell pattern that resembled either ependymoma or oligodendroglioma. However, in all cases the tumor was confirmed to be a neurocytoma by immunocytochemical analysis that showed reactivity for synaptophysin and/or neuron-specific enolase in a high percentage of neoplastic cells. With respect to the literature it is concluded that neurocytomas represent an individual pathological entity of supratentorial midline tumors. Complete surgical removal without irradiation is the recommended treatment.

Adult↗
PubMed1983

Characterization of alpha alpha, beta beta, gamma gamma and alpha gamma human enolase isozymes, and preparation of hybrid enolases (alpha gamma, beta gamma and alpha beta) from homodimeric forms.

Four forms (alpha alpha, beta beta, gamma gamma and alpha gamma) of enolase (2-phospho-D-glycerate hydrolyase, EC 4.2.1.11) were purified from human brain and skeletal muscle. Each purified preparation showed a single band on SDS-polyacrylamide gel electrophoresis, with a relative mobility corresponding to a molecular weight of about 46 000 for alpha and gamma subunits, or 44 000 for the beta subunit. The four enzymes showed similar values of specific activity (about 80 units/mg), optimal pH (pH 6.8-6.9), and Km for 2-phosphoglycerate (about 3 X 10(-5) M). Amino acid analysis, peptide mapping analysis with a limited proteolysis, and immunochemical studies of the purified enzymes revealed that the three subunits, alpha, beta and gamma, are distinct proteins. Three hybrid enolases (alpha gamma, beta gamma and alpha beta) could be prepared from a mixture of two homodimeric enolases by dissociation and reassociation procedures, followed by isolation with ion-exchange column chromatography.

Amino Acids↗
PubMed1992

Characterization of the interaction of yeast enolase with polynucleotides.

Yeast enolase is inhibited under certain conditions by DNA. The enzyme binds to single-stranded DNA-cellulose. Inhibition was used for routine characterization of the interaction. The presence of the substrate 2-phospho-D-glycerate reduces inhibition and binding. Both yeast enolase isozymes behave similarly. Impure yeast enolase was purified by adsorption onto a single-stranded DNA-cellulose column followed by elution with substrate. Interaction with RNA, double-stranded DNA, or degraded DNA results in less inhibition, suggesting that yeast enolase preferentially binds single-stranded DNA. However, yeast enolase is not a DNA-unwinding protein. The enzyme is inhibited by the short synthetic oligodeoxynucleotides G6, G8 and G10 but not T8 or T6, suggesting some base specificity in the interaction. The interaction is stronger at more acid pH values, with an apparent pK of 5.6. The interaction is prevented by 0.3 M KCl, suggesting that electrostatic factors are important. Histidine or lysine reverse the inhibition at lower concentrations, while phosphate is still more effective. Binding of single-stranded DNA to enolase reduces the reaction of protein histidyl residues with diethylpyrocarbonate. The inhibition of yeast enolase by single-stranded DNA is not total, and suggests the active site is not directly involved in the interaction. Binding of substrate may induce a conformational change in the enzyme that interferes with DNA binding and vice versa.

Animals↗
PubMed1990

Selective adherence of neurons and glial cells from dissociated cerebral and spinal cord microcarrier cultures.

In stationary cultures of dissociated brain and spinal cord grown on microcarriers (MCs), the neuronal and ependymal cells attached to the MCs forming floating aggregates in which they grow in a three-dimensional pattern. The glial and meningeal elements on the contrary, tend to dissociate from the aggregates and adhere to the plastic dish where they divide to form a monolayer. This different behavior of CNS components is not observed in rotating cultures in which all CNS cells remain attached to the MCs and develop into mature floating structures. This cell separation in stationary MC-cultures which is documented here by SEM and immunocytochemistry, may be useful for analysis and evaluation of the metabolic biochemical events of each of the cellular components derived from the same culture.

Animals↗