• Proceedings of the KSAR Conference
• /
• 2001.03a
• /
• pp.60-60
• /
• 2001

Human thrombopoietin (hTPO) is a cytokine that plays a central role in megakaryopoiesis by influencing on the development and maturation of megakaryocyte and platelet production. To induce hTPO production in the mammary gland, expression vector was constructed by combining the promoter of bovine beta-casein gene, cDNA of hTPO and neomycine resistance gene for transfection into fibroblasts. Bovine fibroblast cells derived from female ear skin were transfercted with the expression vector using Lipofectamine (Life Technology, NY). Transected cells resistant to G4l8 treatment (600 $\mu\textrm{g}$/$m\ell$) were recovered and colony formation was initiated at 13 days. The colonies with about 1 cm diameter were picked and analysed by PCR. Single transfected cells were individually transferred to enucleated oocytes. After electrofusion, the reconstructed embryos were exposed to calcium ionophore (5uM) for 5 min followed by treatment with 6-DMAP (2.5 mM) for 4h. The nuclear transfer embryos were cultured in CRlaa medium at 38.5C, 5% $CO_2$ for 7 days. Twenty three of 29 (79.3%) colonies were proved to be hTPO transfectants by PCR. The colonies were further passaged and used to produce transgenic embryos using nuclear transfer. Cleavage and developmental rates of reconstructed embryos to the blastocyst stage were 65.1% and 39.4%, respectively Of 22 blastocysts that developed from reconstructed embryos with the transfected cell, 20 embryos (90.9%) were positive for hTPO by using PCR analysis. The results suggest that somatic cell nuclear transfer is efficient for production of transgenic embryos.

• Journal of the Korean Society of Food Culture
• /
• v.38 no.2
• /
• pp.112-118
• /
• 2023

Seaweed-derived foods have long been popular in Korea because of their high content of nutrients that are beneficial to the human body. Recently, Korean seaweeds have been used as raw materials to produce new natural products with health benefits. Herein, we compared the antioxidant activity of 16 Korean seaweed extracts to explore their potential utility as health foods. The total phenolic content (TPC) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity of seaweed extracts were determined. We also investigated their ability to protect human diploid fibroblast (HDF) cells against hydrogen peroxide. The results showed that seaweed extracts at a concentration of 100 ㎍/mL did not cause any cell toxicity. Sargassum thunbergii (Jichung-i) had the highest TPC and radical scavenging effects, followed by Porphyra tenera (Gim), Silvetia siliquosa (Tteumbugi), and Sargassum fusiforme (Tot). Hydrogen peroxide increased the production of intracellular reactive oxygen species, while P. tenera (Gim), Saccharina japonica (Dasima), and S. thunbergii (Jichung-i) extracts significantly decreased it. The effect was highest in the S. thunbergii (Jichung-i)-treated HDF cells. These findings indicate that S. thunbergii (Jichung-i) shows promise as a potential antioxidant raw material.

• Restorative Dentistry and Endodontics
• /
• v.48 no.2
• /
• pp.18.1-18.10
• /
• 2023

Objectives: This study aimed to determine whether collagen triple helix repeat containing-1 (CTHRC1), which is involved in vascular remodeling and bone formation, can stimulate odontogenic differentiation and angiogenesis when administered to human dental pulp stem cells (hDPSCs). Materials and Methods: The viability of hDPSCs upon exposure to CTHRC1 was assessed with the WST-1 assay. CTHRC1 doses of 5, 10, and 20 ㎍/mL were administered to hDPSCs. Reverse-transcription polymerase reaction was used to detect dentin sialophosphoprotein, dentin matrix protein 1, vascular endothelial growth factor, and fibroblast growth factor 2. The formation of mineralization nodules was evaluated using Alizarin red. A scratch wound assay was conducted to evaluate the effect of CTHRC1 on cell migration. Data were analyzed using 1-way analysis of variance followed by the Tukey post hoc test. The threshold for statistical significance was set at p < 0.05. Results: CTHRC1 doses of 5, 10, and 20 ㎍/mL had no significant effect on the viability of hDPSCs. Mineralized nodules were formed and odontogenic markers were upregulated, indicating that CTHRC1 promoted odontogenic differentiation. Scratch wound assays demonstrated that CTHRC1 significantly enhanced the migration of hDPSCs. Conclusions: CTHRC1 promoted odontogenic differentiation and mineralization in hDPSCs.

• Development and Reproduction
• /
• v.14 no.4
• /
• pp.233-241
• /
• 2010

Human adipose stem cells are an abundant, readily available population of multipotent progenitor cells that reside in adipose tissue and these cells have characteristics very similar to bone marrow mesenchymal stromal cells (BMMSCs). However, liposuction procedure, donor age, body mass index, and harvesting sites might generate differences in the initial cell population and the preparations are a heterogeneous mixture of precursors with different subsets. Therefore, in this study, we investigated the characteristics of human thigh adipose stem cells and the differentiation potential into mesodermal and endodermal lineage. Thigh adipose stem cells maintained fibroblast-like morphology similar to BM-MSCs and they underwent average 56.5 doublings and produced $5{\times}10^{22}$ cells. These cells expressed SCF, Oct4, nanog, vimentin, CK18, FGF5, NCAM, Pax6, BMP4, HNF4a, nestin, GATA4, HLA-ABC, and HLA-DR genes at p3 and they also expressed Oct4, Thy-1, FSP, vWF, vimentin, desmin, CK18, CD54, CD4, CD106, CD31, a-SMA, HLA-ABC proteins. Moreover, they could differentiate into mesodermal lineage cells such as adipocyte, osteoblast and chondrocyte. In addition, they also differentiated into insulin secreting cells in our culture condition. In conclusion, human thigh adipose stem cells retain proliferative potential and expression patterns similar to BM-MSCs and they also differentiate into various cell types. Thus, human thigh adipose stem cells might be useful alternative cell source for clinical application.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.33 no.3
• /
• pp.181-187
• /
• 2007

In this study, we measured the anti-oxidative activity of Cirsium setidens and investigated its effect on UVA-induced MMP-1 expression in human dermal fibroblats. And then we examined possible improvement in skin elasticity by topical treatment with fomular including Cirsium setidens extract. The ethanol extract of C. setidens showed free anion radical scavenging effect(87.47 % at 1 mg/mL) and superoxide anion radical scavenging effect(61.71 % at 1 mg/mL) in the xanthine/xanthine oxidase system, respectively. At the concentration of 100 ${\mu}g/mL$, C. setidens extract showed 95.54% inhibition on lipid peroxidation of linoleic acid. UVA-induced MMP-1 expression in human dermal fibroblasts was reduced to 54.69 % by treatment with 100 ${\mu}g/mL$ of C. setidens extract. A human clinical study, in which oil-in-water emulsion with C. setidens extract was topically applied, showed significant increase in skin elasticity. These results suggest that the C. setidens extract can be effective anti-aging ingredient for cosmetics applications.

• The Journal of Korean Academy of Prosthodontics
• /
• v.45 no.3
• /
• pp.382-388
• /
• 2007

Statement of problem. The role of calcium sulfate in stimulating the growth of gingival soft tissue has been reported in few studies. Such a unique property of calcium sulfate could serve as a trouble-solving broker in compensating for the lack of soft tissues in various oral surgeries. Purpose. The purpose of this study was to compare the proliferating activities of human gingival fibroblasts seeded on various bone graft barrier materials of calcium sulfate, collagen, and polytetrafluorethylene (PTFE). Material and methods. Two calcium sulfates ($CAPSET^{(R)}$. and $CalForma^{(R)}$, Lifecore Biomedical Inc., St. Paul, Minnesota, USA), a resorbable natural collagen ($Bio-Gide^{(R)}$, Geistlich Pharma Ag., Wolhusen, Switzerland), and a non-resorbable PTFE ($TefGen-FD^{(R)}$, Lifecore Biomedical Inc., St. Paul, Minnesota, USA) served as the human gingival fibroblasts' substrates and comprised the four experimental groups, whereas the untreated floors of culture plastics were used in the control group, in this study. Cells were trypsinized, seeded, and incubated for 48 h. The proliferating activities of fibroblasts were determined by XTT and SRB assay and absorbance (optical density, OD) was measured. One-way ANOVA was used to analyze the differences in the mean OD values between the groups of CAPSET, CalForma, Bio-Gide, TefGen, and the control (p<0.05). Results. From the XTT assay, the mean OD value of the control group, the highest, was significantly greater than that of any of the four experimental groups followed by CalForma, CAPSET, TefGen, and Bio-Gide. Further, the mean OD value of CalForma, was significantly greater compared to that of Bio-Gide. From the SRB assay, Calforma showed the highest mean OD value, which was significantly greater than that of any other groups, followed by the control, CAPSET, Bio-Gide, and TefGen. The mean OD values of both the control and CAPSET were significantly greater compared to that of TefGen (p<0.05). Conclusion. Assessment of the viability and proliferation of cultured fibroblasts seeded and incubated for 48 h on various barrier-material substrates using XTT and SRB assay showed that calcium sulfate $CalForma^{(R)}$ promotes the proliferating activity of human gingival fibroblasts.

• Asian-Australasian Journal of Animal Sciences
• /
• v.22 no.1
• /
• pp.26-34
• /
• 2009

Porcine embryonic stem (ES) cells have a great potential as tools for transgenic animal production and studies of regulation of differentiation genes. Although several studies showed successful derivation of porcine ES-like cells, these cells were not maintained long-term in culture. Therefore, this study was conducted to establish porcine pluripotent ES-like cells using in vivo fertilized embryos and to maintain these cells in long term culture. Porcine ES-like cells from in vivo embryos obtained by immunosurgery or whole explant culture were successfully cultured for over 56 passages. Morphology of porcine ES-like cells was flat-shaped with a monolayer type colony. These cells stained for alkaline phosphatase throughout the culture. Furthermore, porcine ES-like cells reacted with antibodies against Oct-4, SSEA-1, SSEA-4, Tra-1-60, and Tra-1-81, which are typical markers of undifferentiated stem cells. To characterize the ability of porcine ES-like cells to differentiate into three germ layers, embryoid body formation was induced. After plating of these cells, porcine ES-like cells were spontaneously differentiated into various cell types of all three germ layers. In addition, porcine ES-like cells were successfully derived from IVF blastocysts in media containing human recombinant basic fibroblast growth factor.

• Asian-Australasian Journal of Animal Sciences
• /
• v.27 no.11
• /
• pp.1644-1651
• /
• 2014

Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because of their random integration in the host genome. In this study, we investigated the homologous recombination on the porcine ${\beta}$-casein gene locus using a knock-in vector for the ${\beta}$-casein gene locus. We developed the knock-in vector on the porcine ${\beta}$-casein gene locus and isolated knock-in fibroblast for nuclear transfer. The knock-in vector consisted of the neomycin resistance gene (neo) as a positive selectable marker gene, diphtheria toxin-A gene as negative selection marker, and 5' arm and 3' arm from the porcine ${\beta}$-casein gene. The secretion of enhanced green fluorescent protein (EGFP) was more easily detected in the cell culture media than it was by western blot analysis of cell extract of the HC11 mouse mammary epithelial cells transfected with EGFP knock-in vector. These results indicated that a knock-in system using ${\beta}$-casein gene induced high expression of transgene by the gene regulatory sequence of endogenous ${\beta}$-casein gene. These fibroblasts may be used to produce transgenic pigs for the production of therapeutic proteins via the mammary glands.

• Environmental Analysis Health and Toxicology
• /
• v.18 no.2
• /
• pp.111-120
• /
• 2003

The validation of many synthetic chemicals that may pose a genetic hazard in our environment is of great concern at present. Since these substances are not limited to the original products, and enter the environment, they have become widespread environmental pollutants, thus leading to a variety of chemicals that possibly threaten the public health. In this respect, the regulation and evaluation of the chemical hazard playa very important role to environment and human health. The clastogenicity of 17 synthetic chemicals was evaluated in Chinese hamster lung fibroblast cells in vitro. 2-Nitroaniline (CAS No. 88-74-4) induced chromosomal aberrations with statistical significance at the concentration of 86.3 ${\mu}$g/ml in the absence of metabolic activation system. 1-Chloroanthraquinone (CAS No. 82-44-0) which is one of the most cytotoxic chemical among 17 chemicals tested revealed no clastogenicity in the range of 0.8 ∼ 3.0 ${\mu}$g/ml both in the presence and absence of S-9 metabolic activation system. From the results of chromosomal aberration assay with 17 synthetic chemicals in Chinese hamster lung cells in vitro, 2-Nitroaniline (CAS No. 88-74-4) revealed weak positive clastogenic results in this study.

• Environmental Mutagens and Carcinogens
• /
• v.24 no.1
• /
• pp.33-39
• /
• 2004

To validate and to estimate the chemical hazard playa very important role to environment and human health. The detection of many synthetic chemicals including agrochemicals that may pose a genetic hazard in our environment is of great concern at present. Since these substances are not limited to the original products, and enter the environment, they have become widespread environmental pollutants, thus leading to a variety of chemicals that possibly threaten the public health. Pyrazosulfuron-ethyl [Ethyl-5-(4,6-dimethoxypyrimidin-2-ylcarbamoylsulfamoyl)-1-methylpyrazole-4-carboxylate, $C_{14}H_{18}N{6}O_{7}S,$ M.W. =414.39, CAS No. 93697-74-6], is one of well known rice herbicide belong in the sulfonyl urea group. To clarify the genotoxicity of this agrochemical, Ames bacterial reversion assay, in vitro chromosomal aberration assay with Chinese hamster lung (CHL) fibroblast and bone marrow micronucleus assay in mice were subjected. In Ames assay, although pyrazosulfuron-ethyl revealed cytotoxic at 5,000-140 $\mug/plate$ in Salmonella typhimurium TA100, no dose-dependent mutagenic potential in 4.4～70 $\mug/plate$ of S. typhimurium TA 98, TA 100, TA1535 and TA 1537 both in the absence and presence of S-9 metabolic activation system was observed. Using CHL fibroblasts, the 50% cell growth inhibition concentration $(IC_{50})$ of pyrazosulfuron-ethyl was determined as 1,243 $\mug/mL,$ and no chromosomal aberration was observed both in the absence and presence of S-9 mixture in the concentration range of 311-1,243 $\mug/mL.$ And also, in vivo micronucleus assay using mouse bone marrow, pyrazosulfuron-ethyl revealed no remarkable induction of MNPCE (micronucleated polychromatic erythrocytes/1000 polychromatic erythrocytes) in the dose range of 625-2,500 mg/kg body weight when administered orally. Consequently, Ames bacterial gene mutation with Salmonella typhimurium, in vitro chromosome aberration with mammalian cells and in vivo bone marrow micronucleus assay revealed no clastogenic potential of pyrazosulfuron-ethyl in this study.