• Biomolecules & Therapeutics
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• v.23 no.5
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• pp.391-399
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• 2015

As a major component of the epidermal tissue, a primary keratinocyte has served as an essential tool not only for the study of pathogenesis of skin-related diseases but also for the assessment of potential toxicities of various chemicals used in cosmetics. However, its short lifespan in ex vivo setting has been a great hurdle for many practical applications. Therefore, a number of immortalization attempts have been made with success to overcome this limitation. In order to understand the immortalization process of a primary keratinocyte, several key biological phenomena governing its lifespan will be reviewed first. Then, various immortalization methods for the establishment of stable keratinocyte cell lines will be explained. Finally, its application to a three-dimensional skin culture system will be described.

• Biomedical Science Letters
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• v.10 no.2
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• pp.85-92
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• 2004

Immortalization of primary corneal cells has influence on pharmacy, medical and biological fields. Especially, investigation of immortalization mechanism using viral oncoproteins is useful for medical treatments, and these cell lines will be useful materials for toxic test of medical supplies and cell biological experiments. Rabbit corneal fibroblasts in culture undergo a finite number of divisions before they reach a terminally non-proliferating state known as replicative senescence. Therefore, we attempted to induce immortalization of rabbit corneal fibroblasts with SV 40 large T antigen. As a result of experiment, expression of SV 40 large T antigen was confirmed, and expression of proteins related to cell cycle repressor was decreased in the transfection group compared with non-transfection group. According to the results of cell cycle phase distribution test, SV 40 large T antigen-transfected cells had obtained higher proliferation rate than primary cells. It was confirmed that during induction of immortalization, SV 40 large T antigen was not able to increase telomerase activity. In conclusion, we made a rabbit corneal fibroblast cell line with SV40 large T antigen. This cell line will be useful for further studies of mammalian fibroblast biology, particularly with regard to angiogenesis and malignant transformation. In addition, this cell line offers opportunity for testing potential therapeutics and can be used for toxicity tests of materials or cosmetics. In the future, our cell line can potentially be utilized in a wide range of biology related fields.

• Genomics & Informatics
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• v.4 no.4
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• pp.161-166
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• 2006

The establishment of DNA microarray technology has enabled high-throughput analysis and molecular profiling of various types of cancers. By using the gene expression data from microarray analysis we are able to investigate diagnostic applications at the molecular level. The most important step in the application of microarray technology to cancer diagnostics is the selection of specific markers from gene expression profiles. In order to select markers of Immortalization and transformation we used c-myc and $H-ras^{V12}$ oncogene-transfected NIH3T3 cells as our model system. We have identified 8751 differentially expressed genes in the immortalization/transformation model by multivariate permutation F-test (95% confidence, FDR<0.01). Using the support vector machine algorithm, we selected 13 discriminative genes which could be used to predict immortalization and transformation with perfect accuracy. We assayed $H-ras^{V12}$-transfected 'transformed' cells to validate our immortalization/transformation dassification system. The selected molecular markers generated valuable additional information for tumor diagnosis, prognosis and therapy development.

• IMMUNE NETWORK
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• v.2 no.4
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• pp.183-188
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• 2002

Gradual attrition of telomere to a critical short length elicits successive cellular response of cellular senescence and crisis. Cancer cells evade this process by maintaining functional telomeres via one of two known mechanisms of telomere maintenance. The first and most frequent mechanism involves reactivation of enzyme activity of telomerase, a ribonucleoprotein complex mainly via transcriptional up-regulation of TERT, a catalytic subunit of telomerase complex. The second mechanism utilizes telomerase-independent way termed ALT (for Alternative Lengthening of Telomere), which possibly involves recombination pathways. Thus master key for cellular immortalization is supposed to possess adequate telomere reserves. Indeed, telomerase can alone induce the immortalization under culture on feeder cell layers without generally known inactivation mechanism of tumor suppressor genes. Including this phenomena, this review will focus on telomerase and telomere-associated proteins, thereby implication of these proteins for cellular immortalization processes.

• Proceedings of the Korea Environmental Mutagen Society Conference
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• 2003.10a
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• pp.122-122
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• 2003

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.2
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• pp.92-98
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• 2000

Mesangial cell has several key roles in thee control of glomerular function: it partocipates in the regulation of glomerular filtration rate, macromolecular clearance, and as both a source and target of numerous hormones and autocrines. Many of these insights into mesangial cell function have been obtained by studying mesangial cells in culture. However, no suitble cell lines have established yet. We here reported the immortalization of rat kidney glomeruar mesangial cell by transfection of E6 and E7 genes of human papillomavirus type 16 (HPV-16) via electroporation and lipofection. The reslts showed that only electroporation could transfect the genes to mesangial cells and the transfected cells maintained the viability for longer than 6 months. Fluorescence microscopic observation showed that cellular contractility and phagocytosis, which are the two main phenotypes of mesangial cells with rat glomerular epithelial cells showed that the growth of mesangial dells was suppressed by epithelial cell, but the growth of epithelisl cells was enhanced by mesangial cells. Moreover, Such results may imply that the glomerular cell-cell interaction plays an important role in the regulation of cell proliferation and differentiation.

• Molecules and Cells
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• v.27 no.2
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• pp.143-148
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• 2009

EBV-transformed lymphoblastoid cell lines (LCLs) are used as a resource for human genetic, immunological, and pharmacogenomic studies. We investigated the biological activity of 20 LCL strains during continuous long-term subculture up to a passage number of 160. Out of 20 LCL strains, 17 proliferated up to a passage number of 160, at which point LCLs are generally considered as "immortalized". The other three LCL strains lost the ability to proliferate at an average passage number of 41, during which these LCLs may have undergone cellular crisis. These non-immortal LCL strains exhibited no telomerase activity, decreased EBV gene expression, and a lower copy number of the EBV genome and mitochondrial DNA when compared with immortal LCLs. Thus, this study suggests that sustained EBV viral activity as well as telomerase activity may be required for complete LCL immortalization.

• Journal of the Korean Magnetic Resonance Society
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• v.17 no.1
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• pp.30-39
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• 2013

Immortalization-upregulated protein-1 (IMUP-1) genes have been cloned and are known to be involved in SV40-mediated immortalization. IMUP-1 gene is highly expressed in various cancer cell lines and tumors, suggesting the possibility that they might be involved in tumorigenicity. Previously, there were several problems for overexpression of IMUP-1 in bacterial expression systems including low solubility and aggregation due to unstructured property. To investigate the structural properties, it is necessary to obtain lots of pure and soluble proteins. Accordingly, the co-expression systems of bacterial chaperone proteins, GroEL-GroES, were used to increase solubility of IMUP-1. From the analysis of NMR and CD experiment data, it is suggested that the protein adopt typical the random coil properties in solution.

• Proceedings of the Korean Society of Toxicology Conference
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• 2003.10b
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• pp.122-122
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• 2003

Endometrial tissue is an interesting model for intrinsic and extrinsic factors, ie hormones and growth factors, involved in its normal pathologic development and its cyclic growth. The endometrial cells were isolated from endometrial tissue of the proliferative phase obtained by hysterectomy and separated stromal and epithelial cells.(omitted)

• Development and Reproduction
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• v.17 no.2
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• pp.99-109
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• 2013

Trophoblasts, in the placenta, play a role for placental development as well as implantation in the early pregnancy. The characteristics and functions of trophoblast are identified by their localization and potency for proliferation, differentiation, and invasion. Thus, inadequate trophoblast cell death induces trophoblast dysfunction resulting in abnormal placental development and several gynecological diseases. Recently, it was reported that increased immortalization-upregulated protein-2 (IMUP-2) by hypoxia influences trophoblast apoptosis. However, IMUP-2 function on autophagy, which is type II programmed cell death remains unclear. In this study, we analyzed IMUP-2 expression in trophoblast cells (HTR8-SVneo) and compared IMUP-2 effects on cell death including apoptosis and autophagy in trophoblast regardless of IMUP-2 expression. Increased IMUP-2 in trophoblast by IMUP-2 gene transfection induces cell death, especially, apoptosis increases more than autophagy (p<0.05). However, the decreased IMUP-2 in trophoblasts after siRNA treatment decreased apoptosis with the decreased activities of caspase 3 and 7. The expressions of LC3 and MDC as an autophagosome makers and phosphorylated mTOR, which is a negative regulator for autophagy, increased. In addition, the S phase of cell cycle increased in trophoblasts when IMUP-2 expression decreased. Taken together, the alteration of IMUP-2 can control the balance between apoptosis and autophagy of trophoblasts resulting in functional involvement in placental development and in gynecological diseases by regulating the function of trophoblasts.