• IMMUNE NETWORK
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• 제11권6호
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• pp.342-347
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• 2011

Background: Glial cells are involved in immune and inflammatory responses in the central nervous system (CNS). Glial cells such as microglia and astrocytes also provide structural and functional support for neurons. Migration and morphological changes of CNS cells are associated with their physiological as well as pathological functions. The secreted protein lipocalin-2 (LCN2) has been previously implicated in regulation of diverse cellular processes of glia and neurons, including cell migration and morphology. Methods: Here, we employed a zebrafish model to analyze the role of LCN2 in CNS cell migration and morphology in vivo. In the first part of this study, we examined the indirect effect of LCN2 on cell migration and morphology of microglia, astrocytes, and neurons cultured in vitro. Results: Conditioned media collected from LCN2-treated astrocytes augmented migration of glia and neurons in the Boyden chamber assay. The conditioned media also increased the number of neuronal processes. Next, in order to further understand the role of LCN2 in the CNS in vivo, LCN2 was ectopically expressed in the zebrafish spinal cord. Expression of exogenous LCN2 modulated neuronal cell migration in the spinal cord of zebrafish embryos, supporting the role of LCN2 as a cell migration regulator in the CNS. Conclusion: Thus, LCN2 proteins secreted under diverse conditions may play an important role in CNS immune and inflammatory responses by controlling cell migration and morphology.

• Toxicological Research
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• 제12권1호
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• pp.69-79
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• 1996

Primary culture of cerebellar neuronal cells derived from 8-day old Long-Evans rats was used. Pure granule cells, astrocytes or mixed cells culture systems were prepared. These cells were differentiated and developed synaptic connections. And the astrocytes were identified by immunostaining with glial fibrillary acidic protein (GFAP). Methamphetamine (MAP), which acts on dopaminergic system and cadmium (Cd), a toxic heavy metal, were applied and biochemical assays and electrophysiological studies were performed. $LC_50$ values estimated by MTT assay of MAP and Cd were 3 mM and 2$\mu M$ respectively. Cells were treated with 1 mM or 2 mM MAP and 1$\mu M$ $CdCl_2$ for 48 hour, and the incubation media were analyzed for the content of released LDH. MAP (2 mM) and Cd significantly increased the LDH release. Cell viability was decreased in both groups and some cytopathological changes like cell swelling or vacuolization were seen. The cerebellar granule cells were used for measuring membrane currents using whole-cell clamp technique. Sodium and potassium currents were not affected by MAP neither Cd, but calcium current was significantly reduced by Cd but not affected by MAP. Therefore, in vitro neurotoxicity test system using neuronaI cells and astrocytes cultures were established and can be used in screening of potential neurotoxic chemicals.

• Biomolecules & Therapeutics
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• 제18권1호
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• pp.48-55
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• 2010

Neural progenitor cells (NPCs) differentiate into astrocytes, neurons and oligodendrocytes, which is controlled by various factors in brain. Recent evidences suggest that small molecules modulating the proliferation and differentiation of NPCs may have therapeutic value as well as the potential use as chemical probes. Phylligenin is a lignan with anti-inflammatory activity that is isolated from the fruits of Forsythia koreana. We investigated effects of phylligenin on proliferation and differentiation of NPCs. Treatment of phylligenin decreased the number of proliferating NPCs in culture without effects on the differentiation and survival of neural cells such as neurons and astrocytes. To examine the mechanism of the decreased NPCs number, we performed cell cycle analysis. Proliferation of NPCs was decreased via G1-S transition block by phylligenin treatment, and it was mediated by the increase of p21 level. However, phylligenin did not induce apoptosis of NPCs as determined by TUNEL assay and PARP cleavage. We also found that viability of glioma cell lines such as C6 and U87MG glioma cells, but not that of primary neuron and astrocyte, was inhibited by phylligenin. These results suggest that phylligenin selectively inhibits proliferation of rapidly growing cells such as neural stem cells and glioma cells. Given that the possible role of brain tumor stem cells in the pathology of brain cancers, the inhibitory effects of phylligenin might be useful in the development of new therapeutic agents against brain cancers.

• 대한한방내과학회지
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• 제19권2호
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• pp.381-391
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• 1998

천축황(天竺黃)은 한의학에서 청풍열(淸風熱)과 치담(治痰)하는 효능으로 중풍과 불어증(不語症)을 치료하는데 널리 사용되고 있다. 본 연구에서는 천축황(天竺黃)이 실험적인 치매(Alzheimer Disease; AD)를 유발시키는 물질로 알려진 $amyloid-{\beta}\;(A{\beta}\;peptide)$를 흰쥐의 신경세포의 일종인 astrocyte에 처리하여 그 세포독성과 보호효과 및 세포막 지질의 과산화에 미치는 영향을 검토하였다. 천축황(天竺黃)은 $A{\beta}$에 의한 신경세포에 대한 손상을 억제하여 세포증식을 촉진하여 예방 및 보호효과를 나타내었다. 또한, 세포막 지질의 과산화의 지표인 malondialdehyde (MDA)생성이 $A{\beta}$처리로 크게 증가하였으나 세포막 파괴에 의한 뇌세포 파괴의 전형적인 현상이 천축황(天竺黃)의 전(前)처리와 후(後)처리로 크게 감소되었다. 그리고, 이러한 결과들은 천축황(天竺黃)이 신경세포의 하나인 astrocyte에 대한 보호효과와 세포막지질의 과산화 저해 및 $A{\beta}$처리와 같은 치매유발 독성에 대한 적응능력 향상을 통한 뇌신경의 보호효과를 주는 것으로 노인성 치매 등 임상적 응용에 그 효과가 기대된다.

• 한국수정란이식학회지
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• 제32권2호
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• pp.39-45
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• 2017

Mesenchymal stem cells (MSCs) have been considered an alternative source of neuronal lineage cells, which are difficult to isolate from brain and expand in vitro. Previous studies have reported that MSCs expressing Nestin ($Nestin^+$ MSCs), a neuronal stem/progenitor cell marker, exhibit increased transcriptional levels of neural development-related genes, indicating that $Nestin^+$ MSCs may exert potential with neurogenic differentiation. Accordingly, we investigated the effects of the presence of $Nestin^+$ MSCs in bone-marrow-derived primary cells (BMPCs) on enhanced neurogenic differentiation of BMPCs by identifying the presence of $Nestin^+$ MSCs in uncultured and cultured BMPCs. The percentage of $Nestin^+$ MSCs in BMPCs was measured per passage by double staining with Nestin and CD90, an MSC marker. The efficiency of neurogenic differentiation was compared among passages, revealing the highest and lowest yields of $Nestin^+$ MSCs. The presence of $Nestin^+$ MSCs was identified in BMPCs before in vitro culture, and the highest and lowest percentages of $Nestin^+$ MSCs in BMPCs was observed at the third (P3) and fifth passages (P5). Moreover, significantly the higher efficiency of differentiation into neurons, oligodendrocyte precursor cells and astrocytes was detected in BMPCs at P3, compared with P5. In conclusion, these results demonstrate that neurogenic differentiation can be enhanced by increasing the proportion of $Nestin^+$ MSCs in cultured BMPCs.

• Molecular & Cellular Toxicology
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• 제4권2호
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• pp.106-112
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• 2008

Parkinson's disease (PD) progresses severely by a gradual loss of dopaminergic neurons in the substantia nigra (SN). Epidemiological studies showed that the incidences of PD were reduced by smoking of which the major component, nicotine might be neuroprotective. But the function of nicotine, which might suppress the incidences of PD, is still unknown. Fortunately, recently it was reported that a glial reaction and inflammatory processes might participate in a selective loss of dopaminergic neurons in the SN. The levels of tumour necrosis factor (TNF)-${\alpha}$ synthesised by astrocytes and microglia are elevated in striatum and cerebrospinal fluid (CSF) in PD. TNF-${\alpha}$ kills the cultured dopaminergic neurons through the apoptosis mechanism. TNF-${\alpha}$ release from glial cells may mediate progression of nigral degeneration in PD. Nicotine pretreatment considerably decreases microglial activation with significant reduction of TNF-${\alpha}$ mRNA expression and TNF-${\alpha}$ release induced by lipopholysaccharide (LPS) stimulation. Thus, this study was intended to explore the role of nicotine pretreatment to inhibit the expressions of TNF-${\alpha}$ mRNA in human fetal astrocytes (HFA) stimulated with IL-$1{\beta}$. The results are as follows: HFA were pretreated with 0.1, 1, and $10{\mu}g/mL$ of nicotine and then stimulated with IL-$1{\beta}$ (100 pg/mL) for 2h. The inhibitory effect of nicotine on expressions of TNF-${\alpha}$ mRNA in HFA with pretreated $0.1{\mu}g/mL$ of nicotine was first noted at 8hr, and the inhibitory effect was maximal at 12 h. The inhibitory effect at $1{\mu}g/mL$ of nicotine was inhibited maximal at 24 h. Cytotoxic effects of nicotine were noted above $10{\mu}g/mL$ of nicotine. Moreover, Nicotine at 0.1, 1 and $10{\mu}g/mL$concentrations significantly inhibited IL-$1{\beta}$-induced TF-${\kappa}B$ activation. Collectively, these results indicate that in activated HFA, nicotine may inhibit the expression of TNF-${\alpha}$ mRNA through the pathway which suppresses the NF-${\kappa}B$ activation. This study suggests that nicotine might be neuroprotective to dopaminergic neurons in the SN and reduce the incidences of PD.

• Reproductive and Developmental Biology
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• 제28권4호
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• pp.247-252
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• 2004

Human embryonic stem (ES) cells are derived from the inner cell mass of the preimplantation embryo. Human ES cells have the capacity to differentiate into various types of cells in the body. Human ES cells are indefinite source of cells for cell therapy in various degenerative disorders including neuronal disorders. Directed differentiation of human ES cells is a prerequisite for their clinical application. The objective of this study is to develop the culture condition for the derivation of neural precursor cells from human ES cells. Neural precursor cells were derived from human ES cells in a stepwise culture condition. Neural precursor cells in the form of neural rosette structures developed into neurospheres when cultured in suspension. Suspension culture of neurospheres has been maintained over 4 months. Expressions of nestin, soxl, sox2, pax3 and pax6 transcripts were upregulated during differentiation into neural precursor cells by RT-PCR analysis. In contrast, expression of oct4 was dramatically downregulated in neural precursor cells. Immunocytochemical analyses of neural precursor cells demonstrated expression of nestin and SOX1. When induced to differentiate on an adhesive substrate, neuro-spheres were able to differentiate into three lineages of neural systems, including neurons, astrocytes and oligo-dendrocytes. Transcripts of sox1 and pax6 were downregulated during differentiation of neural precursor cells into neurons. In contrast, expression of map2ab was elevated in the differentiated cells, relative to those in neural precursor cells. Neurons derived from neural precursor cells expressed NCAM, Tuj1, MAP2ab, NeuN and NF200 in immunocytochemical analyses. Presence of astrocytes was confirmed by expression of GFAP immuno-cytochemically. Oligodendrocytes were also observed by positive immuno-reactivities against oligodendrocyte marker O1. Results of this study demonstrate that a stepwise culture condition is developed for the derivation of neural precursor cells from human ES cells.

• 생명과학회지
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• 제17권5호
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• pp.706-713
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• 2007

Scutellaria baicalensis GEORGI(SB) is used in oriental medicine for the treatment of incipient strokes. Although it has been reported that SB is neuroprotective in a hypoxia model, its mechanism is poorly understood. Here, we investigated the effect of SB on the modulation of heme oxygenase-1(HO-1), which has important biological roles in regulating mitochondrial heme protein turnover and in protecting against conditions such as hypoxia, neurodegenerative diseases, or sepsis. Rat cerebrocortical day In vitro(DIV)12 cells were grown in neurobasal medium. On DIV12 cells were treated with SB($20{\mu}g/ml$) and given a hypoxic shock ($2%\;O_2/5%\;CO_2,\;3\;hr$) on DIV14. In situ hybridization results revealed that SB upregulated HO-1 mRNA in neuronal dendrites in both normoxia and hypoxia(38.5% and 59.2%, respectively). At the protein level, SB upregulated HO-1 in the neuronal soma in both normoxia and hypoxia(22.4% and 15.7%, respectively). Interestingly, most significant increase was associated with astrocytes, which increased HO-1 protein by 77.5% compared to SB-untreated culture. These results indicate that SB upregulates both neuronal and glial HO-1 expression, which contributes to the neuroprotection efficacy in hypoxia).

• Anatomy and Cell Biology
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• 제56권2호
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• pp.219-227
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• 2023

Adult neurogenesis has been reported in the hypothalamus, subventricular zone and subgranular zone in the hippocamp. Recent studies indicated that new cells in the hypothalamus are affected by diet. We previously showed beneficial effects of safflower seed oil (SSO), a rich source of linoleic acid (LA; 74%), on proliferation and differentiation of neural stem cells (NSCs) in vitro. In this study, the effect of SSO on hypothalamic neurogenesis was investigated in vivo, in comparison to synthetic LA. Adult mice were treated with SSO (400 mg/kg) and pure synthetic LA (300 mg/kg), at similar concentrations of LA, for 8 weeks and then hypothalamic NSCs were cultured and subsequently used for Neurosphere-forming assay. In addition, serum levels of brain-derived neurotrophic factor (BNDF) were measured using enzyme-linked immunosorbent assay. Administration of SSO for 8 weeks in adult mice promoted the proliferation of NSCs isolated from SSO-treated mice. Immunofluorescence staining of the hypothalamus showed that the frequency of astrocytes (glial fibrillary acidic protein⁺ cells) are not affected by LA or SSO. However, the frequency of immature (doublecortin⁺ cells) and mature (neuronal nuclei⁺ cells) neurons significantly increased in LA- and SSO-treated mice, compared to vehicle. Furthermore, both LA and SSO caused a significant increase in the serum levels of BDNF. Importantly, SSO acted more potently than LA in all experiments. The presence of other fatty acids in SSO, such as oleic acid and palmitic acid, suggests that they could be responsible for SSO positive effect on hypothalamic proliferation and neurogenesis, compared to synthetic LA at similar concentrations.

• 생명과학회지
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• 제27권6호
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• pp.623-631
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• 2017

뇌실하 영역과 subgranular zone은 뇌에서 평생 새로운 신경 세포를 만들어 내는 곳이다. 이 부위에 있는 신경줄기세포는 세포 분열을 통해서 줄기 세포군을 계속 유지할 뿐만 아니라, 신경 세포와 신경 교세포로 분화한다. 세포 분열을 측정하기 위해 thymidine 유사체인 5-ethynyl-2'-deoxyuridine (EdU)가 사용되어 왔다. 몇몇의 경우에서는 새롭게 만들어지는 신경 세포를 표지하려는 목적으로 사용되었다. 이번 연구에서는, EdU가 쥐의 뇌실 하영역에서 분리해낸 신경줄기세포의 분열과 분화에 어떠한 영향을 미치는 지를 보여주었다. 첫째, 신경줄기세포가 EdU를 포함하는 세포 증식 배양액에서 24시간 동안 배양되었을 때, 추후에 분화를 유도하여도 신경세포로 분화가 전혀 일어나지 않았다. EdU를 1시간 동안 처리했을 때도 신경세포로의 분화가 상당부분 저해되었다. 둘째, EdU는 농도가 높을수록, 처리시간이 많을수록 신경줄기세포의 증식을 더욱 많이 저해하였다. 끝으로, EdU는 신경 교세포 중에서 oligodendrocyte으로의 분화는 억제하였지만, astrocyte로의 분화는 오히려 증가시켰다. 본 연구결과는 뇌실하 영역 신경줄기세포의 분화에 EdU가 어떠한 영향을 미치는 지를 처음으로 보여주었고, 이러한 결과들은 신경 세포와 oligodendrocyte로의 분화에 세포 분열이 반드시 필요하다는 것을 제안하고 있다.