• International Journal of Oral Biology
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• v.35 no.4
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• pp.209-214
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• 2010

Cells that have endogenous multipotent properties can be used as a starting source for the generation of induced pluripotent cells (iPSC). In addition, small molecules associated with epigenetic reprogramming are also widely used to enhance the multi- or pluripotency of such cells. Skinderived precursor cells (SKPs) are multipotent, sphereforming and embryonic neural crest-related precursor cells. These cells can be isolated from a juvenile or adult mammalian dermis. SKPs are also an efficient starting cell source for reprogramming and the generation of iPSCs because of the high expression levels of Sox2 and Klf4 in these cells as well as their endogenous multipotency. In this study, valproic acid (VPA), a histone deacetylase (HDAC) inhibitor, was tested in the generation of iPSCs as a potential enhancer of the reprogramming potential of SKPs. SKPs were isolated from the back skins of 5-6 week old C57BL/6 X DBA/2 F1 mice. After passage 3, the SKPs was treated with 2 mM of VPA and the quantitative real time RT-PCR was performed to quantify the expression of Oct4 and Klf4 (pluripotency specific genes), and Snai2 and Ngfr (neural crest specific genes). The results show that Oct4 and Klf4 expression was decreased by VPA treatment. However, there were no significant changes in neural crest specific gene expression following VPA treatment. Hence, although VPA is one of the most potent of the HDAC inhibitors, it does not enhance the reprogramming of multipotent skin precursor cells in mice.

• Journal of Genetic Medicine
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• v.5 no.2
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• pp.100-104
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• 2008

DNA methylation is one of many epigenetic mechanisms that regulate gene expression in the human body. From the view of epigenetics, there are two phases of development, one for germ cell development and another for embryo development. This review will discuss the basic mechanism of methylation, its role in gene expression, and the role of methylation in embryonic reprogramming. Methylation of genes is very critical to embryo development and should be explored further in order to increase our understanding of development.

• Proceedings of the KSAR Conference
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• 2005.06a
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• pp.3-8
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• 2005

• The Korean Journal of Physiology and Pharmacology
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• v.24 no.6
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• pp.463-472
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• 2020

Direct reprogramming, also known as a trans-differentiation, is a technique to allow mature cells to be converted into other types of cells without inducing a pluripotent stage. It has been suggested as a major strategy to acquire the desired type of cells in cell-based therapies to repair damaged tissues. Studies related to switching the fate of cells through epigenetic modification have been progressing and they can bypass safety issues raised by the virus-based transfection methods. In this study, a protocol was established to directly convert fully differentiated fibroblasts into diverse mesenchymal-lineage cells, such as osteoblasts, adipocytes, chondrocytes, and ectodermal cells, including neurons, by means of DNA demethylation, immediately followed by culturing in various differentiating media. First, 24 h exposure of 5-azacytidine (5-aza-CN), a well-characterized DNA methyl transferase inhibitor, to NIH-3T3 murine fibroblast cells induced the expression of stem-cell markers, that is, increasing cell plasticity. Next, 5-aza-CN treated fibroblasts were cultured in osteogenic, adipogenic, chondrogenic, and neurogenic media with or without bone morphogenetic protein 2 for a designated period. Differentiation of each desired type of cell was verified by quantitative reverse transcriptase-polymerase chain reaction/western blot assays for appropriate marker expression and by various staining methods, such as alkaline phosphatase/alizarin red S/oil red O/alcian blue. These proposed procedures allowed easier acquisition of the desired cells without any transgenic modification, using direct reprogramming technology, and thus may help make it more available in the clinical fields of regenerative medicine.

• Molecules and Cells
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• v.46 no.4
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• pp.209-218
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• 2023

In induced pluripotent stem cells (iPSCs), pluripotency is induced artificially by introducing the transcription factors Oct4, Sox2, Klf4, and c-Myc. When a transgene is introduced using a viral vector, the transgene may be integrated into the host genome and cause a mutation and cancer. No integration occurs when an episomal vector is used, but this method has a limitation in that remnants of the virus or vector remain in the cell, which limits the use of such iPSCs in therapeutic applications. Chemical reprogramming, which relies on treatment with small-molecule compounds to induce pluripotency, can overcome this problem. In this method, reprogramming is induced according to the gene expression pattern of extra-embryonic endoderm (XEN) cells, which are used as an intermediate stage in pluripotency induction. Therefore, iPSCs can be induced only from established XEN cells. We induced XEN cells using small molecules that modulate a signaling pathway and affect epigenetic modifications, and devised a culture method which can produce homogeneous XEN cells. At least 4 passages were required to establish morphologically homogeneous chemically induced XEN (CiXEN) cells, whose properties were similar to those of XEN cells, as revealed through cellular and molecular characterization. Chemically iPSCs derived from CiXEN cells showed characteristics similar to those of mouse embryonic stem cells. Our results show that the homogeneity of CiXEN cells is critical for the efficient induction of pluripotency by chemicals.

• BMB Reports
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• v.47 no.11
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• pp.609-618
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• 2014

DNA methylation at cytosines (5mC) is a major epigenetic modification involved in the regulation of multiple biological processes in mammals. How methylation is reversed was until recently poorly understood. The family of dioxygenases commonly known as Ten-eleven translocation (Tet) proteins are responsible for the oxidation of 5mC into three new forms, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). Current models link Tet-mediated 5mC oxidation with active DNA demethylation. The higher oxidation products (5fC and 5caC) are recognized and excised by the DNA glycosylase TDG via the base excision repair pathway. Like DNA methyltransferases, Tet enzymes are important for embryonic development. We will examine the mechanism and biological significance of Tet-mediated 5mC oxidation in the context of pronuclear DNA demethylation in mouse early embryos. In contrast to its role in active demethylation in the germ cells and early embryo, a number of lines of evidence suggest that the intragenic 5hmC present in brain may act as a stable mark instead. This short review explores mechanistic aspects of TET oxidation activity, the impact Tet enzymes have on epigenome organization and their contribution to the regulation of early embryonic and neuronal development.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.2
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• pp.266-277
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• 2014

Somatic cell nuclear transfer (SCNT) has generally demonstrated that a differentiated cell can convert into a undifferentiated or pluripotent state. In the SCNT experiment, nuclear reprogramming is induced by exposure of introduced donor nuclei to the recipient cytoplasm of matured oocytes. However, because the efficiency of SCNT still remains low, a combination of SCNT technique with the ex-ovo method may improve the normal development of SCNT embryos. Here we hypothesized that treatment of somatic cells with extracts prepared from the germinal vesicle (GV) stage Siberian sturgeon oocytes prior to their use as nuclear donor for SCNT would improve in vitro development. A reversible permeability protocol with $4{\mu}g/mL$ of digitonin for 2 min at $4^{\circ}C$ in order to deliver Siberian sturgeon oocyte extract (SOE) to porcine fetal fibroblasts (PFFs) was carried out. As results, the intensity of H3K9ac staining in PFFs following treatment of SOE for 7 h at $18^{\circ}C$ was significantly increased but the intensity of H3K9me3 staining in PFFs was significantly decreased as compared with the control (p<0.05). Additionally, the level of histone acetylation in SCNT embryos at the zygote stage was significantly increased when reconstructed using SOE-treated cells (p<0.05), similar to that of IVF embryos at the zygote stage. The number of apoptotic cells was significantly decreased and pluripotency markers (Nanog, Oct4 and Sox2) were highly expressed in the blastocyst stage of SCNT embryos reconstructed using SOE-treated cells as nuclear donor (p<0.05). And there was observed a better development to the blastocyst stage in the SOE-treated group (p<0.05). Our results suggested that pre-treatment of cells with SOE could improve epigenetic reprogramming and the quality of porcine SCNT embryos.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.10
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• pp.4271-4274
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• 2014

The epithelial to mesenchymal transition (EMT) is a key step during embryonic morphogenesis and plays an important role in drug resistance and metastasis in diverse solid tumors. We previously reported that 48 h treatment of anti-cancer drug doxorubicin could induce EMT in human gastric cancer BGC-823 cells. However, the long term effects of this transient drug treatment were unknown. In this study we found that after 48 h treatment with $0.1{\mu}g/ml$ doxorubicin, most cells died during next week, while a minor population of cells survived and formed colonies. We propagated the surviving cells in drug free medium and found that these long term cultured drug survival cells (abbreviated as ltDSCs) retained a mesenchymal-like cell morphology, and expressed high levels of EMT-related molecules such as vimentin, twist and ${\beta}$-catenin. The expression of chromatin reprogramming factors, Oct4 and c-myc, were also higher in ltDSCs than parental cells. We further demonstrated that the protein level of p300 was upregulated in ltDSCs, and inhibition of p300 by siRNA suppressed the expression of vimentin. Moreover, the ltDSCs had higher colony forming ability and were more drug resistant when compared to parental cells. Our results suggested that an epigenetic mechanism is involved in the EMT of ltDSCs.

• Clinical and Experimental Pediatrics
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• v.53 no.8
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• pp.786-789
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• 2010

Induced pluripotent stem (iPS) cells are generated by epigenetic reprogramming of somatic cells through the exogenous expression of transcription factors. Recently, the generation of iPS cells from patients with a variety of genetic diseases was found to likely have a major impact on regenerative medicine, because these cells self-renew indefinitely in culture while retaining the capacity to differentiate into any cell type in the body, thereby enabling disease investigation and drug development. This review focuses on the current state of iPS cell technology and discusses the potential applications of these cells for disease modeling; drug discovery; and eventually, cell replacement therapy.

• Journal of Korean Neurosurgical Society
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• v.64 no.3
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• pp.406-413
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• 2021

Sacrococcygeal teratoma (SCT) is an extragonadal germ cell tumor (GCT) that develops in the fetal and neonatal periods. SCT is a type I GCT in which only teratoma and yolk sac tumors arise from extragonadal sites. SCT is the most common type I GCT and is believed to originate through epigenetic reprogramming of early primordial germ cells migrating from the yolk sac to the gonadal ridges. Fetal SCT diagnosed in utero presents many obstetrical problems. For high-risk fetuses, fetal interventions (devascularization and debulking) are under development. Most patients with SCT are operated on after birth. Complete surgical resection is the key for tumor control, and the anatomical location of the tumor determines the surgical approaches. Incomplete resection and malignant histology are risk factors for recurrence. Approximately 10-15% of patients have a tumor recurrence, which is frequently of malignant histology. Long-term surveillance with monitoring of serum alpha fetoprotein and magnetic resonance imaging is required. Survivors of SCT may suffer anorectal, urological, and sexual sequelae later in their life, and comprehensive evaluation and care are required.