• The Journal of Korean Medicine
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• v.21 no.4
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• pp.64-72
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• 2000

Objectives : This study was designed to investigate the neuroprotective effect of Hwansodan(Huanshao-dan) on the apoptosis induced by withdrawal of neurotrophic support. Methods : PCl2 pheochromocytoma cells have been used extensively as a model for studying the cellular and molecular effects of neuronal cells. The viability of cells was measured by MTT assay. We used DNA fragmentation and caspase 3-like protease activation assay. Results : The water extract of Hwansodan(Huanshao-dan) significantly showed protective effects on serum and glucose deprivation-induced apoptotic death. Hwansodan(Huanshao-dan) also prevents DNA fragmentation and caspase 3-like protease activation, representing typical neuronal apoptotic phenomena in PCl2 pheochromocytoma cells and induces tyrosine phosphorylation of proteins around 44 kDa, which was identified as ERK1 with electrophoretic gel mobility shift by Western blot. In addition, MAPK kinase(MEK) inhibitor PD98059 and Ras inactivator, ${\alpha}-hydroxyfarnesylphosphonic$ acid attenuated the neuroprotective effects of Hwansodan(Huanshao-dan) in serum-deprived PCl2 cells. Conclusions : These results indicate that Ras/MEK/ERK signaling pathway plays a key role in neuroprotective effects of Hwansodan(Huanshao-dan) in serum-deprived PCl2 cells. Taken together, we suggest the possibility that Hwansodan(Huanshao-dan) might provide a neurotrophic-like activity in PCl2 cells.

• KSBB Journal
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• v.28 no.4
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• pp.230-237
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• 2013

Amyloid ${\beta}$ peptide (A${\beta}$) still best known as a molecule to cause Alzheimer's disease (AD). AD is characterized by the accumulation and deposition of A${\beta}$ within the brain, leading to neuronal cell loss and perturbation of synaptic function by causing free radical formation, inflammation and apoptosis. We investigated the inflammatory action of A${\beta}$ on two types of brain cells, neuronal cells (SH-SY5Y) and neuroglia cells (C6), and its mechanism. We measured the production of NO-iNOS, TNF-${\alpha}$, and ICAM-1 using RT-PCR and Western blot analysis less than the concentration of cytotoxic effects (> 70% survivability). A${\beta}$ had no effect on the production of NO and TNF-${\alpha}$, but significantly increases of iNOS and ICAM-1. Based on this, we suggest that the inflammatory effect of A${\beta}$ results from the action of ICAM-1 in neuronal cells, rather than the release of inflammatory mediators such as NO and TNF-${\alpha}$ in neuroglia cells. In addition, we confirmed whether p53 was related to the action of A${\beta}$ by using SH-SY5Y ($p53^{-/-}$) dominant cells. Neither the expression of p53 nor the cytotoxicity of SH-SY5Y ($p53^{-/-}$) cells were directly affected by A${\beta}$. However, ICAM-1 was not expressed in SH-SY5Y ($p53^{-/-}$) cells. This means that p53- independent pathway exists in the expression of ICAM-1 by A${\beta}$ while p53 plays a role as an on-and-off switch.

• The Korean Journal of Physiology and Pharmacology
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• v.11 no.6
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• pp.239-246
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• 2007

Expressions of endoplasmic reticulum stress response (ERSR) genes were examined during the neuronal differentiation of rat fetal cortical precursor cells (rCPC) and rat pheochromocytoma PC12 cells. When rCPC were differentiated into neuronal cells for 7 days, early stem cell marker, nest in, expression was decreased from day 4, and neuronal markers such as neurofilament-L, -M and Tuj1 were increased after day 4. In this condition, expressions of BIP, ATF6, and phosphorylated PERK as well as their down stream signaling molecules such as CHOP, ATF4, XBP1, GADD34, Nrf2 and $p58^{IPK}$ were significantly increased, suggesting the induction of ERSR during neuronal differentiation of rCPC. ERSR was also induced during the differentiation of PC12 cells for 9 days with NGF. Neurofilament-L transcript was time-dependently increased. Both mRNA and protein levels of Tuj1 were increased after the induction, and the significant increase in NeuN was observed at day 9. Similar to the expression patterns of neuronal markers, BIP/GRP78 and CHOP mRNAs were highly increased at day 9, and ATF4 mRNA was also increased from day 7. These results strongly suggest the induction and possible role of ERSR in neuronal differentiation process. Further study to identify targets responsible for neuronal induction will be necessary.

• The Journal of Internal Korean Medicine
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• v.24 no.2
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• pp.348-357
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• 2003

Objectives : The purpose of this investigation is to evaluate the effects of Aucklandiae Radix Moschus(木香 麝香)and to study the mechanism for neuronal death protection in hypoxia with Embryonic day 20 (E20) cortical cells of a rat (Sprague Dawley). Methods : E20 cortical cells used in this investigation were dissociated in Neurobasal media and grown for 14 days in vitro (DIV). On 14 DIV, Aucklandiae Radix Moschus(木香 麝香) was added to the culture media for 72 hrs. On 17 DIV, cells were given a hypoxic shock and further incubated in normoxia for another three days. On 20 DIV, Moschus(麝香)'s effects for neuronal death protection were evaluated by LDH assay and the mechanisms were studied by Bcl-2, Bak, Bax, caspase family. Results : This study indicate that Aucklandiae Radix(木香)'s effects for neuronal death protection in normoxia and Scutellariae Radix(麝香)'s effects for neuronal death protection in hypoxia were confirmed by LDH assay in culture method of Embryonic day 20(E20) cortical neuroblast. Moschus(麝香)'s mechanism for neuronal death protection in hypoxia is to increase the anti-apoptosis protein Bcl-2. Conclusions : It may be reasonable to propose that Moschus(麝香) protects delayed neuronal death in hypoxia by increasing Bcl-2, thereby reducing mitochondrial permeability transition(PT) pores, the cytochrome c channels.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.38 no.6
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• pp.343-353
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• 2012

Objectives: This aim of this study was to effectively isolate mesenchymal stem cells (hSMSCs) from human submandibular skin tissues (termed hSMSCs) and evaluate their characteristics. These hSMSCs were then chemically induced to the neuronal lineage and analyzed for their neurogenic characteristics in vitro. Materials and Methods: Submandibular skin tissues were harvested from four adult patients and cultured in stem cell media. Isolated hSMSCs were evaluated for their multipotency and other stem cell characteristics. These cells were differentiated into neuronal cells with a chemical induction protocol. During the neuronal induction of hSMSCs, morphological changes and the expression of neuron-specific proteins (by fluorescence-activated cell sorting [FACS]) were evaluated. Results: The hSMSCs showed plate-adherence, fibroblast-like growth, expression of the stem-cell transcription factors Oct 4 and Nanog, and positive staining for mesenchymal stem cell (MSC) marker proteins (CD29, CD44, CD90, CD105, and vimentin) and a neural precursor marker (nestin). Moreover, the hSMSCs in this study were successfully differentiated into multiple mesenchymal lineages, including osteocytes, adipocytes, and chondrocytes. Neuron-like cell morphology and various neural markers were highly visible six hours after the neuronal induction of hSMSCs, but their neuron-like characteristics disappeared over time (24-48 hrs). Interestingly, when the chemical induction medium was changed to Dulbecco's Modified Eagle Medium (DMEM) supplemented with fetal bovine serum (FBS), the differentiated cells returned to their hSMSC morphology, and their cell number increased. These results indicate that chemically induced neuron-like cells should not be considered true nerve cells. Conclusion: Isolated hSMSCs have MSC characteristics and express a neural precursor marker, suggesting that human skin is a source of stem cells. However, the in vitro chemical neuronal induction of hSMSC does not produce long-lasting nerve cells and more studies are required before their use in nerve-tissue transplants.

• Molecules and Cells
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• v.38 no.8
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• pp.729-733
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• 2015

C. elegans has two functional peroxidasins (PXN), PXN-1 and PXN-2. PXN-2 is essential to consolidate the extracellular matrix during development and is suggested to interact with PXN-1 antagonistically. pxn-1 is involved in neuronal development and possibly maintenance; therefore, we investigated the relationship between pxn-1 and pxn-2 in neuronal development and in aging. During neuronal development, defects caused by pxn-1 overexpression were suppressed by overexpression of both pxn-1 and pxn-2. In neuronal aging process, pxn-1 mutants showed less age-related neuronal defects, such as neuronal outgrowth, neuronal wavy processes, and enhanced short-term memory performance. In addition, pxn-2 overexpressing animals retained an intact neuronal morphology when compared with age-matched controls. Consistent with these results, overexpression of both pxn-1 and pxn-2 restored the severe neuronal defects present with pxn-1 overexpression. These results implied that there is a negative relationship between pxn-1 and pxn-2 via pxn-1 regulating pxn-2. Therefore, pxn-1 may function in neuronal development and age-related neuronal maintenance through pxn-2.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.4
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• pp.771-777
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• 2008

Chungpaesagan-tang which is used for treating patients of brain in cerebrovascular disease frequently from clinical doctor has not reported about the effect of neuronal aptosis caused of brain ischemia. The aim of this study is to investigate effect of Chungpaesagan-tang protecting neuronal cells from being damaged by brain ischemia through using organotypic hippocampal slice cultures. We caused ischemic damage to organotypic hippocampal slice cultures by oxygen and glucose deprivation. And added Chungpaesagan-tang extract to cultures. thereafter we measured area percentage of propidium iodide (PI)-stained neuronal cell, lactate dehydrogenase (LDH) levels in culture media and Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells. Area percentage of PI-stained neuronal cells and count of TUNEL-positive cells in CA1 and DG area of organotypic hippocampal slice culture were significantly decreased in pertinent density level of Chungpaesagan-tang extract. LDH levels in culture media of organotypic hippocampal slice culture were significantly decreased in pertinent density level of Chungpaesagan-tang extract. Within pertinent density level, Chungpaesagan-tang has cell protection effect that prevents brain ischemia damaging neuronal cells and apoptosis increasing.

• Journal of Microbiology and Biotechnology
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• v.20 no.4
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• pp.700-707
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• 2010

A novel antioxidative peptide (APVPH I, antioxidative peptides from venison protein hydrolysates I) was purified from venison by enzymatic hydrolysis, column chromatography of DEAE-Sephacel, and high-performance liquid chromatography. The molecular mass of the purified peptide was found to be 9,853 Da and the amino acid sequences of the purified peptide was Met-Gln-Ile-Phe-Val-Lys-Thr-Leu-Thr-Gly. The purpose of this study was to evaluate the effects of APVPH I against $H_2O_2$-induced neuronal cells damage in PC-12 cells. Antioxidative enzyme levels in cultured neuronal cells were increased in the presence of the peptide. In addition, APVPH I inhibited productions of nitric oxide (NO), reactive oxygen species (ROS), malondialdehyde (MDA), and cell death against $H_2O_2$-induced neuronal cell damage in PC-12 cells. It was presumed to be APVPH I involved in regulating the apoptosis-related gene expression in the cell environment. The present results indicate that APVPH I substantially contributes to antioxidative properties in neuronal cells.

• Journal of Korean Neurosurgical Society
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• v.59 no.3
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• pp.259-268
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• 2016

Objective : Accumulation of advanced glycation end-products (AGE) and mitochondrial glycation is importantly implicated in the pathological changes of the brain associated with diabetic complications, Alzheimer disease, and aging. The present study was undertaken to determine whether sildenafil, a type 5 phosphodiesterase type (PDE-5) inhibitor, has beneficial effect on neuronal cells challenged with AGE-induced oxidative stress to preserve their mitochondrial functional integrity. Methods : HT-22 hippocampal neuronal cells were exposed to AGE and changes in the mitochondrial functional parameters were determined. Pretreatment of cells with sildenafil effectively ameliorated these AGE-induced deterioration of mitochondrial functional integrity. Results : AGE-treated cells lost their mitochondrial functional integrity which was estimated by their MTT reduction ability and intracellular ATP concentration. These cells exhibited stimulated generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential, induction of mitochondrial permeability transition, and release of the cytochrome C, activation of the caspase-3 accompanied by apoptosis. Western blot analyses and qRT-PCR demonstrated that sildenafil increased the expression level of the heme oxygenase-1 (HO-1). CoPP and bilirubin, an inducer of HO-1 and a metabolic product of HO-1, respectively, provided a similar protective effects. On the contrary, the HO-1 inhibitor ZnPP IX blocked the effect of sildenafil. Transfection with HO-1 siRNA significantly reduced the protective effect of sildenafil on the loss of MTT reduction ability and MPT induction in AGE-treated cells. Conclusion : Taken together, our results suggested that sildenafil provides beneficial effect to protect the HT-22 hippocampal neuronal cells against AGE-induced deterioration of mitochondrial integrity, and upregulation of HO-1 is involved in the underlying mechanism.

• Advances in Traditional Medicine
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• v.1 no.1
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• pp.55-63
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• 2000

Soeumin Sibjeundaibo-tang (SJDBT) and Soyangin Sibimijihwang-tang (SMJHT) have been used traditionally to improve the systemic blood circulation and biological energy production in the patients with circulatory and neuronal diseases. The object of this study is to determine the protective effects of SJDBT and SMJHT extracts on the spontaneous and glutamate-induced neuronal damages in cultured cells derived from mice cerebral cortex. At 14 days after beginning the cultures, the activity of lactate dehydrogenase released into the culture media was significantly decreased by treatment of cerebroneuronal cells with SJDBT and SMJHT (0.1 mg/ml) for 7 days. By comparison with the normal cells, cerebroneuronal morphology was dramatically changed by treatment of glutamate (1 mM) for 12 hrs, and this was conspicuously recovered by pretreatment of cerebroneural cells with SJDBT and SMJHT (0.1-1.0 mg/ml) for 2 days. Moreover, glutamated-induced DNA fragmentation was also protected by pretreatment of cerebroneuronal cells with those extracts. These results suggest that naturally occurring and glutamate-induced degeneration of cultured cerebrocortical cells may be related, in part, to the process of apoptotic cell death. The pharmacological properties of SJDBT and SMJHT extracts to improve cerebroneuronal degeneration may be considered as one of useful medicines that can prevent cerebrocortical impairments resulted from age-dependent and excitotoxicity-induced neuronal degeneration in human brain.