Biomedical subjects

Yong-Sung Lee

Publications and source records attributed to Yong-Sung Lee.

At least 19 recordsLinked to original sources

PubMed2004

Dexamethasone inhibits proliferation of adult hippocampal neurogenesis in vivo and in vitro.

Activation of glucocorticoid receptor (GR) induces a reduction of adult hippocampal neurogenesis found in dentate gyrus (DG). However, the nature of specific effects by glucocorticoid in hippocampal neurogenesis is not known. In this report, we show differential effects of dexamethasone (DEX), a glucocorticoid receptor agonist, on proliferation and functional differentiation of adult hippocampal progenitor cells in DG. Two-month-old adult rats received daily injections of DEX for 9 days and were sacrificed 12 h and 28 days after the ninth injection. Proliferation assays showed that DEX inhibited proliferation of neural progenitor cells and the inhibitory effects of DEX was not detected 28 days after recovery. Functional differentiation studies using B-cell lymphoma protein-2 (Bcl-2), brain-derived neurotrophic factor (BDNF), p-ERK, and neuronal nuclear protein (NeuN) antibodies revealed that the expressions of Bcl-2 and BDNF were not significantly different between control and DEX-treated rats. In contrast, however, the activation of extracellular signal-regulated kinase (ERK) was downregulated 12 h, but not 28 days, after the DEX treatment. When adult hippocampal progenitor cell cultures were treated with subchronic DEX, proliferation of the progenitor cells was suppressed. Taken these in vitro and in vivo results together, it is concluded that glucocorticoid receptor activation blocks only proliferation, but not differentiation, in hippocampal neurogenesis.

Animals↗

PubMed2004

Temporal expression changes during differentiation of neural stem cells derived from mouse embryonic stem cell.

Temporal analysis in gene expression during differentiation of neural stem cells (NSCs) was performed by using in-house microarrays composed of 10,368 genes. The changes in mRNA level were measured during differentiation day 1, 2, 3, 6, 12, and 15. Out of 10,368 genes analyzed, 259 genes were up-regulated or down-regulated by 2-fold or more at least at one time-point during differentiation, and were classified into six clusters based on their expression patterns by K-means clustering. Clusters characterized by gradual increase have large numbers of genes involved in transport and cell adhesion; those which showed gradual decrease have much of genes in nucleic acid metabolism, cell cycle, transcription factor, and RNA processing. In situ hybridization (ISH) validated microarray data and it also showed that Fox M1, cyclin D2, and CDK4 were highly expressed in CNS germinal zones and ectonucleotide pyrophosphatase/phosphodiesterase 2 (Enpp2) was highly expressed in choroid plexus where stem/progenitor cells are possibly located. Together, this clustering analysis of expression patterns of functionally classified genes may give insight into understanding of CNS development and mechanisms of NSCs proliferation and differentiation.

Animals↗

PubMed2004

Changes of gene expression profiles during neuronal differentiation of central nervous system precursors treated with ascorbic acid.

Ascorbic acid (AA) has been shown to increase the yield of dopaminergic (DA) neurons derived from basic fibroblast growth factor (bFGF)-expanded mesencephalic precursors. To understand the molecular mechanisms underlying this phenomenon, we used cDNA microarray analysis to examine differential expression of neuronal genes following AA treatment. The putative precursor cells were isolated from E13 rat ventral mesencephalons and expanded in the presence of bFGF. Cells were incubated in mitogen-free media supplemented with 200 microM AA or were left untreated as a control, and total RNA was isolated at different time points (expansion stage and 1, 3, and 6 days after induction of differentiation) and subjected to cDNA microarray analysis. Differentiation was evaluated by Western blot analysis and immunocytochemistry of neuron-specific markers. AA treatment of the mesencephalic precursors increased the expression of neuronal (MAP2) and astrocytic (glial fibrillary acidic protein) markers and the percentage of tyrosine hydroxylase (TH)-positive cells. The microarray analysis revealed that 12 known genes were up-regulated and 20 known genes were down-regulated in expansion-stage AA-treated cells. Six days after the induction of differentiation, AA-treated cells showed up-regulation of 48 known genes and down-regulation of 5 known genes. Our results identified several proteins, such as transferrin, S-100, and somatostatin, as being differentially regulated in AA-treated mesencephalic precursors. This novel result may lead to a better understanding of the molecular mechanisms underlying the AA-induced differentiation of mesencephalic precursors into DA neurons and may form the basis for improved DA neuronal production for treatment of Parkinson's disease patients.

Animals↗

PubMed2004

Ascorbic acid responsive genes during neuronal differentiation of embryonic stem cells.

Ascorbic acid has been reported to enhance differentiation of embryonic stem (ES) cells into neurons, however, the specific functions of ascorbic acid have not been defined yet. To address this issue, gene expression profiling was performed using cDNA microarray. Ascorbic acid increased the expressions of genes involved in neurogenesis, maturation, and neurotransmission. Furthermore, statistical analysis using Fisher's exact test revealed ascorbic acid significantly modulated the genes involved in cell adhesion and development category. These results provide information on the role for ascorbic acid during neuronal differentiation of ES cells and might contribute to large-scale generation of neurons for future clinical treatment.

Animals↗

PubMed2004

The complete mitochondrial genome of the javeline goby Acanthogobius hasta (Perciformes, Gobiidae) and phylogenetic considerations.

We isolated Acanthogobius hasta mitochondrial DNA by long-polymerase chain reaction (long-PCR) with conserved primers, and sequenced this mitogenome with primer walking. The resultant A. hasta mitochondrial DNA sequence was found to consist of 16,663 bp with a structural organization conserved relative to that of other fish. In this paper, we report the basic characteristics of the A. hasta mitochondrial genome including structural organization, base composition of rRNAs and the tRNAs and protein-encoding genes, and characteristics of mitochondrial tRNAs. These findings are applicable to molecular phylogenetics in the suborder Gobioidei.

Animals↗

PubMed2004

Differential effects of corticosterone and dexamethasone on hippocampal neurogenesis in vitro.

Prenatal stress during fetal development results in the blockade of neurogenesis in the dentate gyrus in adulthood. Present study was undertaken to investigate the dominant role of the glucocorticoid receptors in corticosterone actions on the neurogenesis of fetal hippocampal progenitor cells. For that purpose, expressions of key molecules affected by corticosterone and dexamethasone were compared during proliferation and differentiation of the hippocampal progenitor cells. Corticosterone (2 microM) significantly decreased the number of bromodeoxyuridine-labeled cells (about 50%) and caused the dendritic atrophy in microtubule-associated protein 2-labeled cells. The expressions of NeuroD, BDNF, and NR1 mRNA levels and protein levels of p-ERK and p-CREB were remarkably decreased by corticosterone in a dose-dependent manner. In contrast, dexamethasone, a glucocorticoid receptor (GR) specific agonist, had an inhibitory effect on proliferation, but not differentiation. It is concluded that corticosterone elicits its effects on neurogenesis including proliferation and differentiation whereas stimulation of the glucocorticoid receptor is sufficient to decrease only proliferation.

Animals↗

PubMed2004

Enhanced in vitro midbrain dopamine neuron differentiation, dopaminergic function, neurite outgrowth, and 1-methyl-4-phenylpyridium resistance in mouse embryonic stem cells overexpressing Bcl-XL.

Embryonic stem (ES) cells provide a potentially unlimited source of specialized cells for regenerative medicine. The ease of inducing stable genetic modifications in ES cells allows for in vitro manipulations to enhance differentiation into specific cell types and to optimize in vivo function of differentiated progeny in animal models of disease. We have generated mouse ES cells that constitutively express Bcl-XL, an antiapoptotic protein of Bcl-2 family. In vitro differentiation of Bcl-XL overexpressing ES (Bcl-ES) cells resulted in higher expression of genes related to midbrain dopamine (DA) neuron development and increased the number of ES-derived neurons expressing midbrain DA markers compared with differentiation of wild-type ES cells. Moreover, DA neurons derived from Bcl-ES cells were less susceptible to 1-methyl-4-phenylpyridium, a neurotoxin for DA neurons. On transplantation into parkinsonian rats, the Bcl-ES-derived DA neurons exhibited more extensive fiber outgrowth and led to a more pronounced reversal of behavioral symptoms than wild-type ES-derived DA neurons. These data suggest a role for Bcl-XL during in vitro midbrain DA neuron differentiation and provide an improved system for cell transplantation in a preclinical animal model of Parkinson's disease.

1-Methyl-4-phenylpyridinium↗

PubMed2004

Human zinc finger protein 161, a novel transcriptional activator of the dopamine transporter.

The dopamine transporter (DAT) terminates dopaminergic neurotransmission via reuptake of released dopamine into presynaptic neurons. We have cloned 2.5 kb of the regulatory region upstream of human DAT (hDAT) and constructed a series of deletion mutants to test promoter activity. A comparison of promoter activity between non-neural and neuronal cell lines reveals an interesting difference in pattern. In the PC12 cell line, activity of the proximal promoter is strongly silenced by one or more unidentified elements spanning positions -395 to -2465 of the hDAT gene. Our studies focus on identifying and characterizing the activating factor for hDAT transcription in the sequence between -2511 and -2492 (5(')-CTA CCT GCA CAG TTC ACG GA-3('), termed HY1). In this investigation, we cloned the zinc finger protein 161 (ZFP161) gene as a HY1-binding factor, using the yeast one-hybrid screen. Recombinant ZFP161 was produced to evaluate the DNA-binding properties of the protein. The ability of ZFP161 to directly bind HY1 was examined in an electrophoretic mobility shift assay. RT-PCR analyses revealed that transfection of ZFP161 induced hDAT mRNA expression in HEK293 cells. We additionally confirmed the expression and localization of the DAT protein, using a specific antibody. Both the HY1 sequence and the downstream region were necessary for activation of the hDAT promoter by ZFP161. This finding suggests that the site of cofactor interaction with ZFP161 may exist downstream of HY1.

Animals↗

PubMed2004

Comprehensive transcriptome analysis of differentiation of embryonic stem cells into midbrain and hindbrain neurons.

Neurogenesis is one of the most complex events in embryonic development. However, little information is available regarding the molecular events that occur during neurogenesis. To identify regulatory genes and underlying mechanisms involved in the differentiation of embryonic stem (ES) cells to neurons, gene expression profiling was performed using cDNA microarrays. In mouse ES cells, we compared the gene expression of each differentiated cell stage using a five-stage lineage selection method. Of 10,368 genes, 1633 (16%) known regulatory genes were differentially expressed at least 2-fold or greater at one or more stages. At stage 3, during which ES cells differentiate into neural stem cells, modulation of nearly 1000 genes was observed. Most of transcription factors (Otx2, Ebf-3, Ptx3, Sox4, 13, 18, engrailed, Irx2, Pax8, and Lim3), signaling molecules (Wnt, TGF, and Shh family members), and extracellular matrix/adhesion molecules (collagens, MAPs, and NCAM) were up-regulated. However, some genes which may play important roles in maintaining the pluripotency of ES cells (Kruppel-like factor 2, 4, 5, 9, myeloblast oncogene like2, ZFP 57, and Esg-1) were down-regulated. The many genes identified with this approach that are modulated during neurogenesis will facilitate studies of the mechanisms underlying ES cell differentiation, neural induction, and neurogenesis.

Animals↗

PubMed2004

Syntaxin 1A and receptor for activated C kinase interact with the N-terminal region of human dopamine transporter.

The dopamine transporter (DAT) regulates the extent and duration of dopamine receptor activation through sodium-dependant reuptake of dopamine into presynaptic neurons, resulting in termination of dopaminergic neurotransmission. Using the yeast two-hybrid system, we have identified novel interactions between DAT, the SNARE protein syntaxin 1A, and the receptor for activated C kinases (RACK1). This association involves the intracellular N-terminal domain of human DAT (hDAT). Our data suggest that hDAT may exist as dimers or oligomers and that its protein-protein interactions with syntaxin 1A and RACK1 form functional regulatory complexes that may mediate DAT trafficking through modulation of hDAT phosphorylation by PKC.

Antigens, Surface↗

PubMed2004

Lithium selectively increases neuronal differentiation of hippocampal neural progenitor cells both in vitro and in vivo.

Lithium has been demonstrated to increase neurogenesis in the dentate gyrus of rodent hippocampus. The present study was undertaken to investigate the effects of lithium on the proliferation and differentiation of rat neural progenitor cells in hippocampus both in vitro and in vivo. Lithium chloride (1-3 mM) produced a significant increase in the number of bromodeoxyuridine (BrdU)-positive cells in high-density cultures, but did not increase clonal size in low-density cultures. Lithium chloride at 1 mM (within the therapeutic range) also increased the number of cells double-labeled with BrdU antibody and TuJ1 (a class III beta-tubulin antibody) in high-density cultures and the number of TuJ1-positive cells in a clone of low-density cultures, whereas it decreased the number of glial fibrillary acidic protein-positive cells in both cultures. These results suggest that lithium selectively increased differentiation of neuronal progenitors. These actions of lithium appeared to enhance a neuronal subtype, calbindin(D28k)-positive cells, and involved a phosphorylated extracellular signal-regulated kinase and phosphorylated cyclic AMP response element-binding protein-dependent pathway both in vitro and in vivo. These findings suggest that lithium in therapeutic amounts may elicit its beneficial effects via facilitation of neural progenitor differentiation toward a calbindin(D28k)-positive neuronal cell type.

Animals↗

PubMed2003

Evaluation of normalization methods for microarray data.

BACKGROUND: Microarray technology allows the monitoring of expression levels for thousands of genes simultaneously. This novel technique helps us to understand gene regulation as well as gene by gene interactions more systematically. In the microarray experiment, however, many undesirable systematic variations are observed. Even in replicated experiment, some variations are commonly observed. Normalization is the process of removing some sources of variation which affect the measured gene expression levels. Although a number of normalization methods have been proposed, it has been difficult to decide which methods perform best. Normalization plays an important role in the earlier stage of microarray data analysis. The subsequent analysis results are highly dependent on normalization. RESULTS: In this paper, we use the variability among the replicated slides to compare performance of normalization methods. We also compare normalization methods with regard to bias and mean square error using simulated data. CONCLUSIONS: Our results show that intensity-dependent normalization often performs better than global normalization methods, and that linear and nonlinear normalization methods perform similarly. These conclusions are based on analysis of 36 cDNA microarrays of 3,840 genes obtained in an experiment to search for changes in gene expression profiles during neuronal differentiation of cortical stem cells. Simulation studies confirm our findings.

Analysis of Variance↗

PubMed2003

Erythropoietin and bone morphogenetic protein 7 mediate ascorbate-induced dopaminergic differentiation from embryonic mesencephalic precursors.

Mesencephalic precursors derived from early (embryonic day 12; E12) rat embryos were grown in vitro using mitogen basic fibroblast growth factor (bFGF) and these cells efficiently differentiated into dopaminergic (DA) neurons. However, this in vitro DA differentiation was poor in mesencephalic precursors isolated from later embryos (E13-15). Ascorbate (AA) treatment enhanced yields of DA neurons from E12 precursors, and increased the number of DA neurons generated from E13 precursors to levels attained when using E12 precursors. AA markedly up-regulated expression of bone morphogenetic protein 7 (BMP7) and erythropoietin (Epo) in precursors, but did not affect expression of a number of genes known to regulate midbrain DA development. The addition of these recombinant proteins or blockers revealed that both BMP7 and Epo mediate AA-induced DA neuron differentiation.

Animals↗

PubMed2003

Ascorbate-induced differentiation of embryonic cortical precursors into neurons and astrocytes.

A specific role for ascorbate (AA) in brain development has been postulated based on a rise of AA levels in fetal brain (Kratzing et al., 1985). To evaluate the role of AA during CNS development, we analyzed the survival, proliferation, and differentiation of AA-treated CNS precursor cells isolated from rat embryonic cortex. Immunocytochemical analyses revealed that AA promoted the in vitro differentiation of CNS precursor cells into neurons and astrocytes in a cell density-dependent manner. Additionally, AA increased the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) of postmitotic neurons in primary neuronal cultures. Differential expression analysis of genes specific to neuronal or glial differentiation revealed an AA-dependent increase in the expression of genes that could potentially compound the effects of AA on cell differentiation. These data suggest that AA may act in the developing brain to stimulate the generation of CNS neurons and glia, thereby assisting in the formation of neural circuits by promoting the acquisition of neuronal synaptic functions.

Animals↗

PubMed2003

Chronic lithium enhances hippocampal long-term potentiation, but not neurogenesis, in the aged rat dentate gyrus.

We investigated the hippocampal long-term potentiation (LTP), neurogenesis, and the activation of signaling molecules in the 20-month-old aged rats following chronic lithium treatment. Chronic lithium treatment produced a significant 79% increase in the numbers of BrdU(+) cells after treatment completion in the dentate gyrus (DG). Both LTP obtained from slices perfused with artificial cerebrospinal fluid (ACSF-LTP), and LTP recorded in the presence of bicuculline (bicuculline-LTP) were significantly greater in the lithium group than in the saline controls. Our results show that as with young rats, chronic lithium can substantially increase LTP and the number of BrdU(+) cells in the aged rats. However, neurogenesis, assessed by colocalization of NeuN and BrdU, was not detected in the aged rat DG subjected to chronic lithium treatment. Therefore, it is concluded that the increase in LTP and the number of BrdU(+) cells might not be associated with increases in neurogenesis in the granule cell layer of the DG. Lithium might has a beneficial effects through other signaling pathways in the aged brain.

Aging↗

PubMed2003

Statistical tests for identifying differentially expressed genes in time-course microarray experiments.

MOTIVATION: Microarray technology allows the monitoring of expression levels for thousands of genes simultaneously. In time-course experiments in which gene expression is monitored over time, we are interested in testing gene expression profiles for different experimental groups. However, no sophisticated analytic methods have yet been proposed to handle time-course experiment data. RESULTS: We propose a statistical test procedure based on the ANOVA model to identify genes that have different gene expression profiles among experimental groups in time-course experiments. Especially, we propose a permutation test which does not require the normality assumption. For this test, we use residuals from the ANOVA model only with time-effects. Using this test, we detect genes that have different gene expression profiles among experimental groups. The proposed model is illustrated using cDNA microarrays of 3840 genes obtained in an experiment to search for changes in gene expression profiles during neuronal differentiation of cortical stem cells.

Algorithms↗

PubMed2003

Neurons and astrocytes secrete factors that cause stem cells to differentiate into neurons and astrocytes, respectively.

We examined the role of soluble factors secreted by neurons and astrocytes in the differentiation of CNS stem cells. We showed that the soluble factors from neurons strongly induced multipotent cortical stem cells to acquire neuronal identity, while the factors from astrocytes promoted astrocytic differentiation. Neurons secreted the brain-derived neurotrophic factor and neurotrophin-3 to induce neuron differentiation, while astrocytes secreted ciliary neurotrophic factor for astrocyte differentiation. Both neurons and astrocytes secrete bone morphogenetic proteins (BMPs). Using BMP antagonists it was shown that BMPs were responsible for the neuron-induced neuronal differentiation, as well as the astrocyte-induced astrocytic differentiation. These findings demonstrate the importance of soluble signals in lineage-specific differentiation and provide evidence for the roles of neurons and astrocytes in stem cell differentiation.

Animals↗

PubMed2003

Lithium enhances long-term potentiation independently of hippocampal neurogenesis in the rat dentate gyrus.

We measured the temporal and spatial profiles of neural precursor cells, hippocampal long-term potentiation (LTP), and signaling molecules in neurogenesis-induced adult rats. Chronic lithium treatment produced a significant 54% and 40% increase in the numbers of bromodeoxyuridine [BrdU(+)] cells after 12 h and 28 days, respectively, after treatment completion in the dentate gyrus (DG). Both LTP obtained from slices perfused with artificial cerebrospinal fluid (ACSF-LTP) and LTP recorded in the presence of bicuculline (bicuculline-LTP) were significantly greater in the lithium group than in the saline controls. Although the number of BrdU(+) cells, approximately 90% of which were double-labeled with a neural marker neuronal nuclear protein, were markedly increased in the granule cell layer (GCL) 28 days after the completion of the 28-day lithium treatment, the magnitude of LTP observed at this time was similar to that observed 12 h after completing the 28-day lithium treatment. However, protein levels of calcium and calmodulin-dependent protein kinase II, p-Elk and TrkB were highly elevated until 28 days after the 28-day lithium treatment. Acute lithium treatment for 2 days also enhanced LTP, which was accompanied by the elevated expression of p-CREB, but not by neurogenesis. Our results suggest that the enhancement of LTP is independent of the increased number of neurons per se and it is more closely associated with key molecules, which are probably involved in neurogenesis.

Animals↗