• Asian Pacific Journal of Cancer Prevention
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• v.14 no.8
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• pp.4533-4537
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• 2013

Objective: To use microarray chip technology for screening of stem cell radiation related miRNAs in laryngeal squamous cell carcinoma; study and explore the relationship of miRNAs with radiosensitivity of laryngeal squamous cells. Method: After conventional culture and amplification of the laryngeal squamous carcinoma cell line Hep-2, CD 133+ cells were screened out with combination of isolated culture of stem cell microspheres and FACS for preparation of laryngeal cancer stem cells. After radiation treatment, miRNAs of laryngeal squamous carcinoma stem cells before and after radiation were enriched and purified. After microarray hybridization with mammalian miRNA and scanning of fluorescence signal, the miRNAs of laryngeal squamous carcinoma stem cells before and after radiation was subject to differential screening and clustering analysis. Real-time quantitative RT-PCR was used to verify part of the differentially expressed miRNAs. Results: 70 miRNAs related to laryngeal cancer stem cell radiation with 2-fold difference in expression were screened out, in which 62 were down-regulated and 8 were up-regulated. Fluorescent quantitative RT-PCR results were consistent with miRNAs chip results. Conclusion: Some miRNAs may be involved in self-regulation with laryngeal squamous carcinoma stem cell radiation.

• Biomedical Science Letters
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• v.13 no.4
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• pp.273-278
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• 2007

Stem cells have been the subject of increasing scientific interest because of their utility in numerous biomedical applications. Stem cells are capable of renewing themselves; that is, they can be continuously cultured in an undifferentiated state, giving rise to more specialized cells of the human body. Therefore, stem cells are an important new tools for developing unique, in vitro model systems to test drugs and chemicals and a potential to predict or anticipate toxicity in humans. In the present study, in vitro cultured F3 immortalized human neural stem cell line and in vivo adult Sprague Dawley rats was used to evaluate the cytotoxicity of anticancer drug paclitaxel. In vitro apoptotic activity of paclitaxel was evaluated in F3 cell line by a MTT assay and DAPI test. The cell death was induced with the treatment of 20 nM paclitaxel and chromatin degradation was detected by DAPI staining, which was analyzed by fluorescent microscope. In vivo studies, we also observed nestin immunoreactivity on subventricular zone, which is stem cell rich region in the adult brain of the SD rat. Immunofluorescent staining result shows that pixel intensities of nestin were decreased in a dose dependent manner. These results suggest that paclitaxel is able to induce cytotoxic activity both in F3 neural stem cell line and neural stem cell in SD rat brain.

• Molecules and Cells
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• v.37 no.1
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• pp.31-35
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• 2014

Induced pluripotent stem cells (iPSCs) are capable of unlimited self-renewal and can give rise to all three germ layers, thereby providing a new platform with which to study mammalian development and epigenetic reprogramming. However, iPSC generation may result in subtle epigenetic variations, such as the aberrant methylation of the Dlk1-Dio3 locus, among the clones, and this heterogeneity constitutes a major drawback to harnessing the full potential of iPSCs. Vitamin C has recently emerged as a safeguard to ensure the normal imprinting of the Dlk1-Dio3 locus during reprogramming. Here, we show that vitamin C exerts its effect in a manner that is independent of the reprogramming kinetics. Moreover, we demonstrate that reprogramming cells under 2i conditions leads to the early upregulation of Prdm14, which in turn results in a highly homogeneous population of authentic pluripotent colonies and prevents the abnormal silencing of the Dlk1-Dio3 locus.

• BMB Reports
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• v.51 no.2
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• pp.79-84
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• 2018

Niemann-Pick type C disease (NP-C) is a fatal neurodegenerative disorder caused by a deficiency of NPC1 gene function, which leads to severe neuroinflammation such as astrogliosis. While reports demonstrating neuroinflammation are prevalent in NP-C, information about the onset and progression of cerebellar astrogliosis in this disorder is lacking. Using gene targeting, we generated vascular endothelial growth factor (VEGF) conditional null mutant mice. Deletion of VEGF in cerebellar Purkinje neurons (PNs) led to a significant increase of astrogliosis in the brain of NP-C mice in addition to the loss of PNs, suggesting PN-derived VEGF as an important factor in NP-C pathology. Moreover, replenishment of VEGF in neurons improved brain pathology in NP-C mice. Overall, our data provide a new pathological perspective on cerebellar astrogliosis in NP-C and suggest the importance of VEGF as a therapeutic target for this disease.

• Development and Reproduction
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• v.13 no.4
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• pp.227-237
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• 2009

Recently, adipose mesenchymal stem cells (AdMSC) that are similar to bone marrow MSC and blood derived MSC are thought to be another source for stem cell therapy. However, the diseases that can be applied for stem cells therapy are age-dependent degenerative diseases. Accordingly, the present study investigated the growth and differentiation potential to neural cells of human AdMSC (hAdMSC) obtained from aged thirty, forty and fifty. The growth of cells and cell viability were measured by passage and neural differentiation of hAdMSC was induced in neural differentiation condition for 10 days. Our results demonstrated that cell number, viability and morphology were not different from hAdMSC by age and passage. Immunofluorescence analysis of neural cell marker (TuJ1, NSE, Sox2, GFAP or MAP2) demonstrated no significant differences in neural cell differentiation by age and passage. As the number of passage was increased, the mRNA level of MAP2 and Sox2 was decreased in hAdMSC from age of 50 compared to hAdMSC from age of 30. In conclusion, the present study demonstrated that ability of neural cell differentiation of hAdMSC was maintained with ages, suggesting that autologous stem cells from aged people can be applied for stem cell therapy with age-dependent neural disease with the same stem cell quality and ability as stem cell derived from young age.

• Korean Journal of Clinical Laboratory Science
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• v.51 no.4
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• pp.475-483
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• 2019

The use of stem cells in cell-based therapy has attracted extensive interest in the field of regenerative medicine, and it has been applied to numerous incurable diseases due to the inherent abilities of self-renewal and differentiation. However, there still exist some severe obstacles, such as requirement of cell expansion before the treatment, and low survival at the treated site. To overcome these disadvantages of stem cells, we used the carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM 6) gene, which functions to increase cell-cell interaction as well as anti-apoptosis. We first confirmed whether CEACAM 6 is expressed in various cell lines at the protein level (including in stem cells), followed by evaluating and selecting the optimal transfection conditions into stem cells. The CEACAM 6 gene was transfected into stem cells to prolong cell survival and preserve from damage by oxidative stress. After confirming the CEACAM 6 expression in transfected stem cells, the cell survival was assessed under oxidative condition by exposing to hydrogen peroxide (H₂O₂) to mimic the chronic environment-induced cellular damage. CEACAM 6 expressing stem cells show increased cell viability compared to the non-CEACAM 6 expressing cells. We propose that the application of the CEACAM 6 gene is a potential option, capable of expanding and enhancing the therapeutic effects of stem cells.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.20 no.6
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• pp.95-103
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• 2019

Periodontal inflammation, a major kind of periodontal diseases, is characterized to bleed, pain, and teeth loss, and it is resulted from oxidative stress. Membrane-free stem cell extract could avoid the immunogencity rejection by removal of cell membrane. In the present study, we investigated the protective effect of membrane-free stem cell extract from oxidative stress-induced periodontal inflammation in human periodontal ligament fibroblasts (HPLF). In the cell viability measurement, membrane-free stem cell extract showed significant increase of cell viability, compared with the $H_2O_2$-treated control group. To further investigation of molecular mechanisms, we measured inflammation and apoptosis related protein expressions. Membrane-free stem cell extract attenuated inflammation-related protein expressions such as nuclear factor kappa light chain enhancer of activated B cells, inducible nitric oxide synthase, and interleukin-6. In addition, the treatment of membrane-free stem cell extract decreased apoptotic protein expressions such as cleaved caspase-9, -3, poly (ADP-ribose) polymerase, and B-cell lymphoma 2 (Bcl-2)-associated X protein/Bcl-2 ratio in the $H_2O_2$-treated HPLF cells. In conclusion, membrane-free stem cell extract exhibited anti-oxidative stress effects by regulation of inflammation and apoptosis in HPLF, suggesting that it could be used as the treatment agents for periodontal inflammatory disease.

• International Journal of Stem Cells
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• v.16 no.3
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• pp.356-362
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• 2023

Glutathione (GSH) is a chief cellular antioxidant, affecting stem cell functions. The cellular GSH level is dynamically altered by the redox buffering system and transcription factors, including NRF2. Additionally, GSH is differentially regulated in each organelle. We previously reported a protocol for monitoring the real-time GSH levels in live stem cells using the reversible GSH sensor FreSHtracer. However, GSH-based stem cell analysis needs be comprehensive and organelle-specific. Hence, in this study, we demonstrate a detailed protocol to measure the GSH regeneration capacity (GRC) in living stem cells by measuring the intensities of the FreSHtracer and the mitochondrial GSH sensor MitoFreSHtracer using a high-content screening confocal microscope. This protocol typically analyses the GRC in approximately 4 h following the seeding of the cells onto plates. This protocol is simple and quantitative. With some minor modifications, it can be employed flexibly to measure the GRC for the whole-cell area or just the mitochondria in all adherent mammalian stem cells.

• Korean Journal of Plant Tissue Culture
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• v.27 no.6
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• pp.423-428
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• 2000

In 1997 when cloned sheep Dolly and soon after Polly were born, it had become head-line news because in the former the nucleus that gave rise to the lamb came from cells of six-year-old adult sheep and in the latter case a foreign gene was inserted into the donor nucleus to make the cloned sheep produce human protein, factor IX, in e milk. In the last few years, once the realm of science fiction, cloned mammals especially in livestock have become almost commonplace. What the press accounts often fail to convey, however, is that behind every success lie hundreds of failures. Many of the nuclear-transferred egg cells fail to undergo normal cell divisions. Even when an embryo does successfully implant in the womb, pregnancy often ends in miscarriage. A significant fraction of the animals that are born die shortly after birth and some of those that survived have serious developmental abnormalities. Efficiency remains at less than one % out of some hundred attempts to clone an animal. These facts show that something is fundamentally wrong and enormous hurdles must be overcome before cloning becomes practical. Cloning researchers now tent to put aside their effort to create live animals in order to probe the fundamental questions on cell biology including stem cells, the questions of whether the hereditary material in the nucleus of each cell remains intact throughout development, and how transferred nucleus is reprogrammed exactly like the zygotic nucleus. Stem cells are defined as those cells which can divide to produce a daughter cell like themselves (self-renewal) as well as a daughter cell that will give rise to specific differentiated cells (cell-differentiation). Multicellular organisms are formed from a single totipotent stem cell commonly called fertilized egg or zygote. As this cell and its progeny undergo cell divisions the potency of the stem cells in each tissue and organ become gradually restricted in the order of totipotent, pluripotent, and multipotent. The differentiation potential of multipotent stem cells in each tissue has been thought to be limited to cell lineages present in the organ from which they were derived. Recent studies, however, revealed that multipotent stem cells derived from adult tissues have much wider differentiation potential than was previously thought. These cells can differentiate into developmentally unrelated cell types, such as nerve stem cell into blood cells or muscle stem cell into brain cells. Neural stem cells isolated from the adult forebrain were recently shown to be capable of repopulating the hematopoietic system and produce blood cells in irradiated condition. In plants although the term$\boxDr$ stem cell$\boxUl$is not used, some cells in the second layer of tunica at the apical meristem of shoot, some nucellar cells surrounding the embryo sac, and initial cells of adventive buds are considered to be equivalent to the totipotent stem cells of mammals. The telomere ends of linear eukaryotic chromosomes cannot be replicated because the RNA primer at the end of a completed lagging strand cannot be replaced with DNA, causing 5' end gap. A chromosome would be shortened by the length of RNA primer with every cycle of DNA replication and cell division. Essential genes located near the ends of chromosomes would inevitably be deleted by end-shortening, thereby killing the descendants of the original cells. Telomeric DNA has an unusual sequence consisting of up to 1,000 or more tandem repeat of a simple sequence. For example, chromosome of mammal including human has the repeating telomeric sequence of TTAGGG and that of higher plant is TTTAGGG. This non-genic tandem repeat prevents the death of cell despite the continued shortening of chromosome length. In contrast with the somatic cells germ line cells have the mechanism to fill-up the 5' end gap of telomere, thus maintaining the original length of chromosome. Cem line cells exhibit active enzyme telomerase which functions to maintain the stable length of telomere. Some of the cloned animals are reported prematurely getting old. It has to be ascertained whether the multipotent stem cells in the tissues of adult mammals have the original telomeres or shortened telomeres.

• Molecules and Cells
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• v.41 no.3
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• pp.207-213
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• 2018

Hypoxic culture is widely recognized as a method to efficiently expand human mesenchymal stem cells (MSCs) without loss of stem cell properties. However, the molecular basis of how hypoxia priming benefits MSC expansion remains unclear. In this report, our systemic quantitative proteomic and RT-PCR analyses revealed the involvement of hypoxic conditioning activated genes in the signaling process of the mitotic cell cycle. Introduction of screened two mitotic cyclins, CCNA2 and CCNB1, significantly extended the proliferation lifespan of MSCs in normoxic condition. Our results provide important molecular evidence that multipotency of human MSCs by hypoxic conditioning is determined by the mitotic cell cycle duration. Thus, the activation of mitotic cyclins could be a potential strategy to the application of stem cell therapy.