• Proceedings of the Korean Society of Machine Tool Engineers Conference
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• 2005.05a
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• pp.3-8
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• 2005

We introduced mechanical stimuli and micropatterned substrate with micro fibers to investigate the effects of those on neurite outgrowth along with nerve growth factor (NGF) in vitro. Microfiber substrates were fabricated using an electrospinning process. And PC-12 cells cultured on substrates were simulated with nerver growth factor and laminar flow shear stress in a fluid flow system The results suggest that microfiber substrates and fluid-induced shear stress are promising for simulating neuronal regeneration in a desired direction.

• BMB Reports
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• v.45 no.9
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• pp.521-525
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• 2012

In addition to its pivotal role in neuronal survival, PI3K/Akt signaling is integral to neuronal differentiation and neurite outgrowth. However, the exact role of Akt in neuronal differentiation is still controversial. Here, we found that nuclear expression of CA-Akt resulted in unusual rapid neurite outgrowth and overexpression of KD-Akt caused multiple dendrite growth without specific axon elongation. Moreover, microarray data revealed that the expression of FOXQ1 expression was about 10-fold higher in cells with nuclear, active Akt than in control cells. Quantitative real-time PCR analysis showed that mRNA levels were upregulated in NLS-CA-Akt cells as compared to KD or EV cells. Furthermore, our FACS analysis demonstrated that overexpression of NLS-CA-Akt accumulate cells in the G1 phase within 24 h, fitting with the rapid sprouting of neuritis. Thus, our data implied that at least in this early time frame, the overexpression of nuclear, active Akt forced cells into neurite development through probably FOXQ1regulation.

• BMB Reports
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• v.33 no.6
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• pp.525-530
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• 2000

The nerve growth factor (NGF) induces neuronal differentiation and neurite outgrowth of PC12 cells, whereas epidermal growth factors (EGF) stimulate growth and proliferation of the cells. In spite of this difference, NGF-or EGF-treated PC12 cells share various properties in cellular-signaling pathways. These include the activation of the phosphoinositide (PI)-3 kinase, 70 kDa S6 kinase, and in the mitogen-activated protein (MAP) kinase pathway, following the binding of these growth factors to intrinsic receptor tyrosine kinases (RTKs). Therefore, many studies have been attempted to access the critical signaling events in determining the differentiation and proliferation of PC12 cells. In this study, we investigated the cytosolic phospholipase $A_2$ ($cPLA_2$) in neurite behavior in order to identify the differences of signaling pathways between the NGF-induced differentiation and the EGF-induced proliferation of PC12 cells. We have showed here that the $cPLA_2$ was translocated from cytosol to membrane only in NGF-treated cells. We also demonstrated that this translocation is associated with NGF-induced activation of phospholipase $C-{\gamma}(PLC-{\gamma})$, which elevates intracellular $Ca^{2+}$ concentration. These results reveal that the translocation of $cPLA_2$ may be a requisite event in the neuronal differentiation of PC12 cells. Various phospholipase inhibitors were used to confirm the importance of these enzymes in the differentiation of PC12 cells. Neomycin B, a PLC inhibitor, dramatically inhibited the neurite outgrowth, and two distinct $PLA_2$ inhibitors, 4-bromophenacyl bromide (BPB) and arachidonyltrifluoro-methyl ketone ($AACOCF_3$) also suppressed the neurite outgrowth of the cells, as well Taken together, these data indicated that $cPLA_2$ is involved in NGF-induced neuronal differentiation and neurite outgrowth of PC12 cells.

• Toxicological Research
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• v.12 no.2
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• pp.203-211
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• 1996

Methanol has been widely used as an industrial solvent and environmental exposure to methanol would be expected to be increasing. In humans, methanol causes metabolic acidosis and damage to ocular system, and can lead to death in severe and untreated case. Clinical symptoms are attributed to accumulation of forrnic acid which is a metabolic product of methanol. In humans and primates, formic acid is accumulated after methanol intake but not in rodents due to the rapid metabolism of methanol. Neverthless, the developmental and reproductive toxicity were reported in rodents. Previous reports showed that perinatal exposure to ethanol produces a variety of damage in human central nervous system by direct neurotoxicity. This suggests that the mechanism of toxic symptoms by methanol in rodents might mimic that of ethanol in human. In the present study I hypothesized that methanol can also induce toxicity in neuronal cells. For the study, primary culture of rat hippocampal neurons and glias were empolyed. Hippocampal cells were prepared from the embryonic day-17 fetuses and maintained up to 7 days. Effect of methanol (10, 100, 500 and 1000 mM) on neurite outgrowth and cell viability was investigated at 0, 18 and 24 hours following methanol treatment. To study the changes in proliferation of glial cells, protein content was measured at 7 days. Neuronal cell viability in culture was not altered during 0-24 hours after methanol treatment. 10 and 100 mM methanol treatment significantly enhanced neurite outgrowth between 18-24 hours. 7-day exposure to 10 or 100 mM methanol significantly increased protein contents but that to 1000 mM methanol decreased in culture. In conclusion, methanol may have a variety of effects on growing and differentiation of neurons and glial cells in hippocampus. Treatment with low concentration of methanol caused that neurite outgrowth was enhanced during 18-24 hours and the numbers of glial cell were increased for 7 days. High concentration of methanol brought about decreased protein contents. At present, the mechanism responsible for the methanol- induced enhancement of neurite outgrowth is not clear. Further studies are required to delineate the mechanism possibly by employing molecular biological techniques.

• The Korea Journal of Herbology
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• v.30 no.2
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• pp.43-48
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• 2015

Objectives : The aim of this study was to investigate the memory enhancing properties of extract of Hoelen Cum Radix (HCR) and its possible mechanism in mice of normal condition. Methods : We evaluated the effects of HCR on cognitive function and memory enhancement in normal mice. Male ICR mice were orally administrated with HCR 100 mg/kg for 7 days and equal volume of saline was administrated to the control group in the same condition. We conducted two behavioral tests which measure the spatial working memory (Y-maze test) and cognitive fear memory (passive avoidance test). We also investigated whether HCR affects the hippocampal neurogenesis in the brain. To assess the effects of HCR on neural progenitor cell differentiation and neurite outgrowth in the early stage of hippocampal neurogenesis, we performed doublecortin (DCX), a direct neurogenesis marker, immunohistochemical analysis in the dentate gyrus (DG) of the mouse hippocampus. Results : HCR significantly enhanced memory and cognitive function as determined by the Y-maze test (p<0.05) and passive avoidance test (p<0.001). Moreover, HCR increased DCX positive cells (p<0.01) and neurite length (p<0.01) compared to the control group. These results indicated that HCR stimulates differentiation of neural progenitor cells and promotes neurite outgrowth in hippocampal DG of the mice. Conclusion : We concluded that HCR shows memory enhancing effects through the stimulation of hippocampal neurogenesis as a consequence of accelerated neuronal differentiation and neurite outgrowth in the DG of the hippocampus after HCR treatment.

• International Journal of Oral Biology
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• v.41 no.4
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• pp.231-236
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• 2016

Inhibition of Rho-associated coiled coil-containing kinase (ROCK) has been reported to promote differentiation of neuronal cells. Here, we examined the effect of Y-27632, a ROCK inhibitor, on the outgrowth of neurites in PC12 cells. Y-27632 caused a rapid induction of neurite outgrowth in PC12 cells in a time-dependent manner. The neurite outgrowth, triggered by Y-27632, was accompanied by Rac1 activation, and was attenuated by Rac1 inhibitor NSC23766, in a concentration-dependent manner. Y-27632 also induced an increase in the production of reactive oxygen species (ROS). Pretreatment with N-acetylcysteine, an ROS scavenger, inhibited the ROS generation and neurite outgrowth in response to Y-27632. These results indicate that the activation of Rac1 and the generation of ROS contribute to the neurite outgrowth triggered by Y-27632 in PC12 cells.

• BMB Reports
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• v.40 no.6
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• pp.888-894
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• 2007

Src homology (SH) domains of phospholipase C-$\gamma1$ (PLC-$\gamma1$) impair NGF-mediated PC12 cells differentiation. However, whether the enzymatic activity is also implicated in this process remains elusive. Here, we report that the enzymatic activity of phospholipase C-$\gamma1$ (PLC-$\gamma1$) is at least partially involved to the blockage of neuronal differentiation via an abrogation of MAPK activation, as well as sustained Akt activation. By contrast, Overexpression of WT-PLC-$\gamma1$ exhibited sustained NGF-induced MAPK activation, and triggered transient Akt activation resulting in profound inhibition of neurite outgrowth. However, lipase-inactive mutant (LIM) PLC-$\gamma1$ cells fail to suppress neurite outgrowth, although it contains intact SH domains, specifically enhancing the expression of cyclin D1 and p21 proteins, which regulate the function of retinoblastoma Rb protein. These observations show that the lipase inactive mutant of PLC-$\gamma1$ does not alter NGF-induced neuronal differentiation via enzymatic inability and the modulation of cell cycle regulatory proteins independent on SH3 domain.

• 한국생물공학회:학술대회논문집
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• 2005.10a
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• pp.859-859
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• 2005

• Journal of Microbiology and Biotechnology
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• v.19 no.11
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• pp.1490-1495
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• 2009

The development of nanotechnology has penetrated the fields of biology and medicine, resulting in remarkable applications for tissue regeneration. In order to apply this technology to tissue engineering, we have developed nano-scaled 3D scaffolds consisting of growth factor-loaded heparin/poly(l-lysine) nanoparticles (NPs) attached to the surface of polymeric micro spheres via polyionic complex methods. Growth factor-loaded NPs were simply produced as polyelectrolyte complexes with diameters of 100-200 nm. They were then coated onto positively charged poly(lactic-co-glycolic acid) (PLGA) pretreated with polyethyleneimine to enable cell adhesion, proliferation, and stimulation of neurite outgrowth. Propidium iodide staining and $\beta$-tubulin analysis revealed that neuronal PC12 cells proliferated extensively, expressed significant amounts of b-tubulin, and showed well-structured neurite outgrowth on polymeric microspheres by stimulation with growth factors. These results suggest that cellular adhesion and biological functionality on prepared PLGA microspheres enabled terminal differentiation of neuronal cells.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1993.11a
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• pp.28-33
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• 1993

Regulation of nerve growth factor (NGF)-induced neuronal differentiation by GTPase activating protein(GAP) and its mechanism were investigated in rat pheochromocytoma cell line, PCl2. Overexpression of GAP caused the delay in the onset of neurite outgrowth of PCl2 eel Is in response to NGF. GAP has been known to inhibit p21$\^$ras/, the activated form of which induces neuronal differentiation. Therefore, the activity of p21$\^$ras/ was compared in control cells and cells overexpressing GAP indirectly by measuring the activities of B-Raf and MAP kinase that are known to be positively regulated by p21$\^$ras/. Surprisingly, NGF-induced activities of these two proteins were the same in control eells and GAP-overexpressing cells. Activities of Trk, PLC-r and SMC that act at a site upstream to p21$\^$ras/ in NGF signal transduction pathway were not also affected by GAP overexpression. Interestingly, however, the extent of tyrosine phosphorylation of SNT was found to be remarkably low in cells overexpressing GAP. It has been shown previously that neurotrophins and not mitogens induce SNT tyrosine phosphorylation in PCl2 cells. Thus it is possible that the timing of NGF-induced neuronal differntiation may be in part regulated by SNT and the slower onset of neurite outgrowth in cells overexpressing GAP may be through the inhibition of SNT by GAP.