• Proceedings of the KSAR Conference
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• 2003.06a
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• pp.17-17
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• 2003

The milk of transgenic animals may provide an attractive vehicle for large-scale production of hEPO. Since glycosylation is cell type specific, recombinant human EPO (rhEPO) produced in different host cells contain different patterns of oligosaccharides, which could affect the biological functions. However, there have been no reports on the characteristics of rhEPO derived from milk of transgenic animals. To address this objective, several transgenic mice by using pWAPhEPO and/or pBC1hEPO expression vector were produced. However, 2 lines of pWAPhEPO founder female mouse died during late gestational day (day 18) before offspring could be obtained. They showed a severe splenomegaly, Unlike those of pWAPhEPO, mammary gland epithelial cells from biopsies of lactating pBC1hEPO transgenic mice had marked immunoreactivity to EPO and any activity was not detected in other tissues. The expression level of rhEPO is about 0.7% of mammary gland cellular total soluble proteins and an amount of 300~500 mg/L rhEPO is secreted into milk. Furthermore, the pBC1hEPO transgenic mice transmitted this character to their progeny in mendelian manner. In order to determine the extent of glycosylation variation, N-linked oligosaccharide structures present in the milk-derived rhEPO were characterized. Most of milk-derived rhEPO is fully glycosylated. the biological activity of milk-derived rhEPO was comparable to that of purified CHO-derived rhEPO, and milk-derived rhEPO showed relatively stable after freezing and thawing. Taken together, the results illustrate the potential of transgenic animals in the large-scale production of biopharmaceuticals.

• Bulletin of the Korean Chemical Society
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• v.34 no.4
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• pp.1089-1095
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• 2013

The tumor necrosis factor-receptor 1 (TNFR1)-associated death domain protein (TRADD) contains an N-terminal TRAF binding domain and a C-terminal death domain. TRADD is known to interact directly with TNF receptor 2 (TNFR2) and the Fas-associated death domain protein (FADD), which are signal transducers that activate NF-${\kappa}B$ and induce apoptosis, respectively. To date, there has been no structural information on the TRADD and FADD death domain (DDs) complex. In this study, the death domains of TRADD and FADD were co-expressed and purified from Escherichia coli for structural characterization. We found that human TRADD (hTRADD) interacted strongly with mouse FADD (mFADD) via their DDs and interacted weakly with human FADD (hFADD)-DD. Moreover, the structures of the TRADD-DD:FADD-DD complexes were separately modeled from predicted structures in the protein data bank (PDB). The results of this study will have important applications in human diseases such as cancer, AIDS, degenerative and autoimmune diseases, and infectious diseases.

• Journal of the Korean Society of Food Science and Nutrition
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• v.29 no.2
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• pp.322-328
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• 2000

This study was performed to determine the antimutagenic and cytotoxic effect of the Phellinus linteus methanol extract on Salmonella typhimurium TA98, TA100 and human cancer cell lines. In the Ames test, methanol extract of P. linteus alone did not exhibit any mutagenicity but showed substantial inhibitory effects against mutation induced by N-methyl-N'-nitro-N-nitrosoguanidine(MNNG), 4-nitroquinoline-nitroquinoline-1-oxide(4NQO), 3-amino-1,4-dimethyl-5H-pyrdo[4,3-blindol(Trp-P-1) and benzo(α)pyrene(B(α)P). The methanol extracts of P. linteus(200㎍/plate) showed approximately 78.3%, 78.7% and 88.1% inhibitory effect on the mutagenesis induced by 4NQO, Trp-P-1 and B(α)P. The anticancer effects of P. linteus extract against human breast adenocarcinoma(MCF7), human lung carcinoma (A549), human fibrosarcoma (HT1080), human hepatocelular carcinoma (Hep3B) and human epitheloid carcinoma (HeLa) were investigated. The treatment of 1mg/mL P. linteus extracts had the highest cytotoxicity against MCF7 (92.0%), followed by Hep3B (84.9%), A549 (84.2%) and HT1080 (82.9%). In contrast 1mg/mL treatment of P. linteus extracts had only 10∼40% cytotoxicity on normal human liver cell (WRL68).

• Journal of Microbiology
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• v.40 no.3
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• pp.224-229
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• 2002

The effect of human cytomegalovirus (HCMV) on three human monocyte cell lines at different stages of differentiation was investigated. While the viability of HL-60 cells or U-937 cells was not significantly affected by HCMV infection, the viability of THP-1 cells was reduced. Acridine orange/ethidiurn bromide staining revealed that the reduction of THP-1 cell viability was due to increased apoptotic death following HCMV infection. Apoptosis in HL-60 cells was not affected by HCMV infection, and induction of apoptosis of U-937 cells by HCMV was intermediate between HL-60 and THP-1 cells. Since HL-60 cells are the least differentiated and THP-1 cells are the most differentiated, the induction of apoptosis of human monocytes appears to be related to the degree of cell differentiation. Flow cytometric and confocal microscopic studies using fluorescent calcium indicator Fluo-3 suggested a significant increase in intracellular free calcium concentration (［Ca$\^$2+/］i) in THP-1 cells undergoing apoptosis by HCMV infection. Again ［Ca$\^$2+/］i in HCMV-infected HL-60 cells was not critically altered, and that in HCMV-infected U-937 cells was intermediate between THP-1 cells and HL-60 cells. Calcium influx blockers such as verapamil and nifedipine partially reversed HCMV-induced apoptosis in THP-1 cells.

• Journal of the Korean Society of Food Science and Nutrition
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• v.24 no.5
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• pp.651-657
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• 1995

The antioxidative activity of each fraction in human milk was examined using H2O2 and FeSO4-induced lipid peroxidation of mouse liver homogenate in order to elucidate the antioxidative substances of human milk. High molecular weight(~>20KD) fraction had more antioxidative effect on lipid peroxidation than low molecular weight(~ <20KD) fraction. Furthermore, the changes of antioxidative enzyme activities were estimated during lactation to study the roles of human milk. The human milk showed high activities of catalase, glutathione(GSH) peroxidase and GSH S-transferase. These results suggest that the antioxidative activities may mostly be attributed to high molecular weight fraction containing catalase, GSH peroxidase and GSH S-transferase.

• Journal of the Korean Society of Food Science and Nutrition
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• v.44 no.9
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• pp.1279-1285
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• 2015

The current study aimed to investigate effects of ethanol extract of white sesame seed (WSE) as well as a major constituent of white sesame seed, ${\beta}-sitosterol$, on the growth, migration, and adhesion of H1299 human lung cancer cells. Treatment with WSE at concentrations of 150, 300, and $600{\mu}g/mL$ dose-dependently inhibited cell growth (to 51.5~82.6% of control). Treatment with ${\beta}-sitosterol$ at concentrations of 3.125, 6.25, 12.5, and $25{\mu}M$ inhibited cell growth to a greater extent (to 27.5~49.0% of control) than that with WSE (P<0.05). Treatment with WSE (at concentration of $600{\mu}g/mL$) or ${\beta}-sitosterol$ (at concentration of $25{\mu}M$) resulted in increased sub-G1 cell population, indicating their apoptosis-inducing activities. ${\beta}-sitosterol$ was effective in inhibiting both cell migration (to 80.8~86.2% of control at a concentration range of $3.125{\sim}25{\mu}M$) and adhesion (to 21.5~37.4% of control at a concentration range of $6.25{\sim}25{\mu}M$), whereas WSE at a concentration range of $150{\sim}600{\mu}g/mL$ was ineffective. These results indicate that ${\beta}-sitosterol$ is more active than WSE in inhibiting growth, migration, and adhesion of H1299 human lung cancer cells. Further studies are needed to determine if similar effects are reproduced in vivo.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.1
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• pp.19-27
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• 2004

Objective: This study was to examine the in vitro neural cell differentiation patterns of human embryonic stem (hES) cells following treatment of various neurotrophic factors [basic fibroblast growth factor (bFGF), retinoic acid (RA), brain derived neurotrophic factor (BDNF) and transforming growth factor (TGF)-$\alpha$], particulary in dopaminergic neuron formation. Methods: The hES cells were induced to differentiate by bFGF and RA. Group I) In bFGF induction method, embryoid bodies (EBs, for 4 days) derived from hES were plated onto gelatin dish, selected for 8 days in ITSFn medium and expanded at the presence of bFGF (10 ng/ml) for another 6 days followed by a final differentiation in N2 medium for 7, 14 and 21 days. Group II) For RA induction, EBs were exposed of RA ($10^{-6}M$) for 4 days and allowed to differentiate in N2 medium for 7, 14 and 21 days. Group III) To examine the effects of additional neurotrophic factors, bFGF or RA induced cells were exposed to either BDNF (10 ng/ml) or TGF-$\alpha$ (10 ng/ml) during the 21 days of final differentiation. Neuron differentiation and dopamine secretion were examined by indirect immunocytochemistry and HPLC, respectively. Results: The bFGF or RA treated hES cells were resulted in similar neural cell differentiation patterns at the terminal differentiation stage, specifically, 75% neurons and 11% glial cells. Additionally, treatment of hES cells with BDNF or TGF-$\alpha$ during the terminal differentiation stage led to significantly increased tyrosine hydroxylase (TH) expression of a dopaminergic neuron marker, compared to control (p<0.05). In contrast, no effect was observed on the rate of mature neuron (NF-200) or glutamic acid decarboxylase-positive neurons. Immunocytochemistry and HPLC analyses revealed the higher levels of TH expression (20.3%) and dopamine secretion (265.5 $\pm$ 62.8 pmol/mg) in bFGF and TGF-sequentially treated hES cells than those in $\alpha$ RA or BDNF treated hES cells. Conclusion: These results indicate that the generation of dopamine secretory neurons from in vitro differentiated hES cells can be improved by TGF-$\alpha$ addition in the bFGF induction protocol.

• Microbiology and Biotechnology Letters
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• v.18 no.4
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• pp.401-405
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• 1990

Maximum specific pro-UK production rate was achieved at 15 mllmin of perfusion rate as $5.7 \times 10^{-8}$ $\mu g$/h/cell by cytostatic cultivation of human cell line 579 with DMEM and 590 FBS. As perfusion rates were increased, glutamine uptake rates were also increased but ammonium production rates remained relatively constant, which resulted in low ratio of ammonium to glutamine at high perfusion rate. Partially it quantitatively explains why the productivity is increased in perfusion cultivations. At maintenance period of 15 days by controlling metabolic process, such as 5 mM of glucose, 2 mM of gtutamine, 10% of air saturation and pH 6.2, high speecific product production rate and product yield on substrate were obtained as $12\times 10^{-8}$$\mu g$/h/cell abd 0.226 mglg of glucose, respetively. This product yield corresponds to 0.223 mg/day of productivity at 10 mllmin of perfusion rate.

• Archives of Pharmacal Research
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• v.28 no.4
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• pp.463-468
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• 2005

The purpose of the present study was to develop a standard protocol for imidapril hydrochloride bioequivalence testing. For this reason, a specific LC-MS method was developed and validated for the determination of imidapril in human plasma. A solid-phase extraction cartridge, $Sep-pak^{R}$ C18, was used to extract imidapril and ramipril (an internal standard) from deproteinized plasma. The compounds were separated using a XTerra $MS^{R}$?C18 column ($3.5 {\mu}m, 2.1\times150 mm$) and $acetonitrile-0.1\%$ formic acid (67:33, v/v) adjusted to pH 2.4 by 2 mmol/L ammonium formic acid, as mobile phase at 0.3 mL/min. Imidapril was detected as m/z 406 at a retention time of ca. 2.3 min, and ramipril as m/z 417 at ca. 3.6 min. The described method showed acceptable specificity, linearity from 0.5 to 100 ng/mL, precision (expressed as a relative standard deviation of less than $15\%$), accuracy, and stability. The plasma concentration-versus-time curves of eight healthy male volunteers administered a single dose of imidapril (10 mg), gave an $AUC_{12hr}$ of imidapril of $121.48\pm35.81 ng mL^{-1} h$, and $C_{max} and T_{max}$ values of $32.59\pm9.76 ng/mL and 1.75\pm0.27 h$. The developed method should be useful for the determination of imidapril in plasma with sufficient sensitivity and specificity in bioequivalence study.

• YAKHAK HOEJI
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• v.47 no.6
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• pp.422-431
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• 2003

The use of recombinant human erythropoietin (rhEPO), a stimulator of erythropoiesis, banned in sports because of the medical risk associated with thrombosis. Due to analytical difficulties to differentiate between natural human EPO (hEPO) and rhEPO, blood parameters of erythropoiesis such as contents of hemoglobin (cut-off value <17.5 g/d l for man, and < 16.0 g/dl for women), hematocrit and reticulocytes (cut-off value <2.0%) were measured to focus the misuse of rhEPO. We conducted anti-doping test for 122 blood samples of the World Cup athletes. The mean values of key parameters are as follows; 14.5$\pm$1.0 g/dl for hemoglobin, 41.7$\pm$2.8% for hematocrit, and 1.3$\pm$0.4% for reticulocyte. Blood sample was found to be stable up to 8 hours for the reticulocyte measurement. In addition, the soluble transferrin receptor and ferritin levels were measured by immunoassay methods using plasma samples (n=28) in which the mean value was 0.8$\pm$0.5 $\mu\textrm{g}$/$m\ell$ and 54.6$\pm$33.7 ng/$m\ell$, respectively. The results indicate that all samples tested were negative for the blood parameters of indirect anti-doping test for hEPO misuse. The statistical evaluation suggest that several other parameters such as red blood cell, mean corpuscular hemoglobin concentration, mean corpuscular volume, mean corpuscular hemoglobin and white blood cell could be considered as factors influencing hEPO function in addition to five parameters mentioned.