• Journal of the Korea Organic Resources Recycling Association
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• v.8 no.4
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• pp.147-152
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• 2000

For the probiotic feed production from residual food waste by using the yeast Kluyveromyces marxianus, aerobic liquid fermentation was attempted at $35^{\circ}C$. After grinding finely, optimal fermentation conditions of the substrate was investigated in shaking incubator. By controlling water content yeast growth was studied at each different solid content of 5, 10 and 15% respectively. The most active growth of the yeast was shown at 10%. For the stimulation of the cell growth, mixed culture with Aspersillus oryzae was conducted in a 2 litre-jar fermenter. As the results, the yeast growth rate was increased, but the maximum viable cell count amounted was slightly higher as $3.5{\times}10^9/ml$ than single culture.

• Journal of Biomedical Engineering Research
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• v.41 no.3
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• pp.138-145
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• 2020

Adipocytes are the main constituent of adipose tissue. Understanding the molecular basis of adipogenesis is pivotal to finding the therapeutic targets for treatment of obesity. Melatonin is associated with obesity and its mechanism is currently under intensive investigation. The objective of this study was to investigate the effect of melatonin on adipogenesis in differentiating preadipocytes. 3T3-L1 preadipocytes were cultured in Dulbecco's modified Eagle's medium (DMEM) containing 5% calf serum at 37℃ with 5% CO₂ in a humidified incubator. Differentiation was induced using DMEM with 10% fetal bovine serum supplemented with MDI two days after cell confluence (day 0). Cells were treated with 0, 10 and 100 μM melatonin on either day 0 or day 5. 72 hours after each treatment, lipid accumulation was measured by oil red O staining. Proteins were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to membranes. As a result, lipid accumulation decreased with melatonin treatment. ERK pathway, activated when differentiation is induced, also decreased with an increase in melatonin concentration. Furthermore, the expression of key adipogenic factors, C/EBPα, C/EBPβ, and PPARγ, were reduced by melatonin treatment. These results imply that melatonin may inhibit the process of adipogenesis and may have a role as a new anti-obesity agent.

• The Journal of Korean Obstetrics and Gynecology
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• v.20 no.1
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• pp.50-60
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• 2007

Purpose : This investigation was undertaken to evaluate the anti-proliferation, in hexane, chloroform, ethyl acetate, butanol and water fraction from extract of Gyulyupsanbyonbang(GYSB) using MCF-7 human breast cancer cells. Methods : GYSB was added to distilled water(1500ml) and was boiled then filtered. The residue was suspended in distilled water and extracted with hexane, chloroform, ethyl acetate, butanol and water. MCF-7 cells were cultured in RPMI1640 complex badge, NIH3T3 was cultered in 37$^{\circ}$C, 5% moisture incubator of carbon dioxide with Dulbecco's Modified Eagle Medium(DMEM) supplemented with 10% fetal bovine serum and antibiotics. Cell cytotoxicity test about cancer cell was measured used MTT assay. Results: When it synthesizes a result, hexane and butanol fraction had shown anti-proliferation effect and safety together, and those anti-proliferation effect operating selectively appeared. Ethyl acetate fraction had anti-proliferation effect however, it was not selective. The Chloroform and water soluble fraction did not almost appear anti-proliferation effect. Conclusion : I can conclude that GYSB have anti-proliferation effect and safety together on MCF-7 cells. It suggest that GYSB may be useful for brest cancer patients.

• Journal of Embryo Transfer
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• v.9 no.1
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• pp.23-29
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• 1994

The long term goal of this research is to develop an efficient procedure for large scale production of genetically identical or cloned animals. To improve nuclear transpalntation efficiency in the rabbit, this study evaluated the age of nuclear recipient oocytes on the different steps of nuclear transplantation. The ovulated oocytes in different ages were collected from the superovulated does by flushing oviducts with Dulbecco's phosphate buffered saline(D-PBS) supplemented with 10% fetal calf serum(FCS) from 13 to 15, 17 to 20 and 23 to 26 hours after hCG injection. The denuded oocytes were used as nuclear recipient cytoplasm following enucleation by micromanipulation. The blastomeres separated from the 8-cell embryos were used as nuclear donor. The enucleated oocytes receiving a blastomere in the perivitteline space were fused in the 0.28 M mannitol solution at 1.5 kV/cm, 60 sec for three times. The fused oocytes were co-cultured with the monolayered rabbit oviductal epithelial cells in TGM-199 solution with 10% FCS for 72 hours at 37$^{\circ}C$ in a 5% $CO_2$ incubator. The cultured nuclear transplant embryos and in vivo developed embryos collected at 72 hours after hCG injection were stained with Hoechst 33342 dye. Their cell numbers were counted under a fluorescent microscope. The results obtained were summarized as follows ; 1. The aged oocytes(20 hrs. post hCG) showed significantly(P<0.05) higher fusionrates(70 ~ 90%) than the recently ovulated oocytes(30.8%) 2. The aged oocytes which were electrically activated and fused at 20 hours developed to blastocyst at significantly(P<0.05) high rate, while none of the recently ovulated oocytes developed to blastocyst. 3. Even though the aged oocytes at 23~26 hours showed higher fusion rate(85.7%), not only they were inadequate to manipulate but also their developmental potential to blastocyst was highly impaired. 4. The developmental potential in vitro of nuclear transplant embryos was significantly retarded than in vivo deveolped embryos.

• Korean Journal of Agricultural Science
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• v.48 no.3
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• pp.557-565
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• 2021

Interest in research on various medicinal plants has increased globally over the last few decades, possibly due to their possible antibacterial and antioxidant activities. The present study was conducted to verify the antioxidant effects, antibacterial activity, and collagen synthesis and cell viability outcomes of adipocytes upon exposure to Abeliophyllum distichum Nakai (AdN). Antibacterial activity was measured through the Disc diffusion method to compare the growth ability of pathogenic microorganisms (E.coli, Salmonella). The absorbance was measured at 560 nm to calculate the active oxygen scavenging ability. Fibroblasts were dispensed in a 96-well plate at a density of 1 × 10⁵ cells·well^-1. The amount of procollagen was measured in each case using a procollagen type 1 C-peptide EIA KIT. The cytotoxicity of the Abeliophyllum distichum Nakai extract against animal adipocytes (Hanwoo backfat cells) was determined using a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, a method that measures the conversion of MTS to Formazan by means of mitochondrial dehydrogenases. The concentrations of the samples were made to be 0.0125, 0025, 0.05, 0.1, and 0.5% and all were -completely absorbed into the disc in an incubator at 37℃ for 24 to 36 hours. For the 0.125 mg·disc^-1, effects of Abeliophyllum distichum Nakai on the antioxidant effect, antibacterial activity, and cell viability of adipocytes were found. However, Abeliophyllum distichum Nakai had no effect on collagen synthesis, thus suggesting that AdN extracts may be useful for the prevention and/or treatment of obesity.

• Journal of Periodontal and Implant Science
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• v.29 no.1
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• pp.173-192
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• 1999

The purpose of this study was to evaluate the effects of bioactive glass and natural coral on the human periodontal ligament fibroblast(HPLF) behaviors during the regeneration process of peridontium. To determine the cellular events occuring in the presence of the particles of bioactive glass and natural coral, HPLF were isolated from healthy premolar teeth extracted for orthodontic treatment. Cells were cultured in ${\alpha}$MEM at 37$^{\circ}C$, 5% $CO_2$, 95% humidity incubator. Bioactive glass and natural coral were powdered, and each particles(<40${\mu}$m) were placed on the cultured cells at the concentration of 0.3mg/ml, and 1,0mg/ml for experimental group. In control group no particles were added. And each group was evaluated by examining the cell morphology under phase-contrast micrograph at 4 day and transmission electron micrograph(TEM) and scanning electron micrograph(SEM) at 14 day, alkaline phosphatase activity at 5 and 9 day, protain synthesis at 4 day, DNA synthesis at 1, 2, 3 and 4 day, cell proliferation at 1, 3, 5,7 and 9 day and the formation of bone nodule at 30 day after culturing all groups in mineralizing supplemented mediun, No significant changes in cell morphology by adding these two matirials were found under phase contrast microscopy and TEM. HPLF phagocytocized each particles suggesting that HPLF is involved in the process of resorbing each particles and that bioactive glass were more biocompatible than natural coral. The ALPase activity of bioactive glass 0.3 mg/ml was similar with control groups and all the rests of control groups were significantly low(P<0.01) indicating a transient dedifferentiation of HPLF in the presence of bioactive glass and natural coral particles. There were no significant differences of protein synthesis between all groups. The DNA synthesis in experimental groups were significantly lower than control groups at 1, 2 and 3 day (P<0.01) but became similar to control groups at 4 day. Between control groups, the DNA synthesis in bioactive glass O.3mglml group was significantly higher than other groups(P<0.01). Cell proliferation in natural coral 1.0mg/ml and bioactive glass 1.0mglml groups were significantly lower than control group at 3 day(P<0.05) and there were no differences at 5, 7, 9 day. There were more bone nodule formation in experimental groups than in control groups. In conclusion, these results indicated that bioactive glass and natural coral have some effects of a transient dedifferentiation on HPLF and regeneration of periodontal tissues, however any significant cytotoxic effect on HPLF by these two particles were not found.

• Parasites, Hosts and Diseases
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• v.28 no.2
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• pp.91-100
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• 1990

Paragonimus westermani is a tissue migrating parasite in the early stage until arriving at lung, and most of the parasites spend their life spans there. Considerable immune responses including activation of macrophages are taken place during the residence of parasites in the host. However, concerning the immunologic defense mechanisms of the host against this parasite, only a few document is available so far. In this study, the cytotoxic effect of peritoneal macrophages under the presence of antibody and/or complement against metacercariae of F. westermani was investigated in vitro. Metacercarlae were collected from the crayfish, Cambaroides similis and hatched out in Tyrode solution (pH 7.4). Plastic adherent cells from normal or infected rat (Wistar) peritoneal exudates were used as experimental macrophages. Polyclonal antibodies were obtained from infected rats and a cat. Cat IgG was fractioned with ion exchange chromatography. Fresh rabbit complement was used according to experimental scheme. Various combinations of peritoneal macrophages, normal or infected rat serum, complement and cat IgG were incubated at $36^{\circ}C$ in 5% $CO_2$ incubator for 6, 14, 24 and 48 hours. The results obtained were as follows: 1. P. westermani infection activated peritoneal macrophages non-specifically and this activation induced increases of cell adherence and cytotoxicity on metacercariae. 2. In the presence of infected rat serum the antibody.dependent cell-mediated cytotoxicity of peritoneal macrophages on metacercariae was significantly increased and showed a peak at 6-hour incubation. But the cytotoxic effect was markedly reduced after inactivation of complement and heat.labile IgE antibody by the heating of infected serum at 56$^{\circ}C$ for 30 minutes. 3. The highest cytotoxic effect (100%) of concomitant incubation with IgG and complement showed 24 hours after incubation, although cell adherence was relatively low at 6-hour incubation and 0% at 24-hour incubation. 4. Coordinative functions of complement with serum and IgG were effective in cell adherence and in cytotoxicity, but it is not clear the independent role of complement on the macrophage- mediated cytotoxicity in this study- With these results it is assumed that P. westermani infection can induce the non-specific activation of peritoneal macrophages, and strum antibodies including IgE antibody might enhance the cytotoxicity by macrophages,

• Journal of Periodontal and Implant Science
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• v.30 no.2
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• pp.389-405
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• 2000

The purpose of this study is to evaluate the effect of IGF-I for DNA synthetic activity and the mRNA expression of bone matrix protein, type I collagen and osteopontin in prolifetation and differentiation of MC3T3-E1 cells. To evaluate DNA synthetic activity, cells were seeded at $2{\times}10^4cells/ml$ in 24 well plates and to evaluate mRNA of type I collagen and osteopontin cells were seeded at $5{\times}10^5cells/ml$ in 100mm culture dishes. These cells were cultured in alpha-minimum essential medium(${\alpha}-MEM$) containing 10% fetal bovine serum at $37^{\circ}C$, 5% $CO_2$ incubator. For DNA synthetic activity test 1, 10, 100ng/ml IGF-I were added to the cells which had been cultured for 3 days before 24 hours. For type I collagen mRNA expression 1, 10ng/ml IGF-I were added to the cells which had been cultured for 5, 10 days and for osteopontin mRNA expression 0.1, 1, 10ng/ml IGF-I were added to the cells which had been cultured for 5, 15, 20 days. Cell proliferaton was measured by the incorporation of [$^3H$]-thymidine into DNA and expression for type I collagen and osteopontin were measured by northern blot analysis. The results were as follows : DNA synthetic activity were generally higher in experimental group than control group. Expressions of type I collagen mRNA were higher at 5 day group and much lower at 10 day group in the control groups. In the experimental groups, mRNA expressions were slightly increased when 1 ng/ml IGF-I were added to 5 day group and decreased in all experimental 10 day groups. Expressions of osteopontin mRNA were higher at 20 day groups and lower at 15 day groups than the control groups. In the experimental groups, mRNA expressions were incereased when 0.1, 1 ng/ml IGF-I were added to 5 day group and in all the 15 day groups, but decreased when 0.1, 1, 10 ng/ml IGF-I were added to 20 day groups. IGF-I stimulated DNA synthetic activity of MC3T3-E1 cells during proliferation stage significantly, did not greatly changed effects on type I collagen mRNA expression and stimulated osteopontin mRNA expression at 15 day especially. In conclusion, we suggests that IGF-I have a tendency of stimulation effect of DNA synthetic activity but do not stimulate type I collagen mRNA in proliferation stage of MC3T3-E1 cell cultures, and stimulate osteopontin mRNA in differentiation stage of MC3T3-E1 cell cultures.

• Journal of Embryo Transfer
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• v.13 no.1
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• pp.11-19
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• 1998

This study was to determine the effect of ionomycin and 6-dimethylaminopurine (6-DMAP) and/or elcetrical stimulation on the oocyte activation and production of rabbit nuclear transplant embryos. The oocytes were collected from the oviduct of superovulated rabbits at 14 h post hCG injection and cultured in TCM-199 containing 10% FBS until 19 h post hCG injection. To determine the optimum concentration and exposure time of 6-DMAP, some oocytes were activated with 5 $\mu$M ionomycin for 5 min and then in 2.0 mM 6-DMAP for 0.5 to 3.0 h, or in 1.0 to 3.0 mM 6-DMAP for 2.0 h. Other control oocytes were stimulated electrically(3X, 1.25 kV/cm, 60 $\mu$sec) in 0.3 M mannitol solution supplemented with 100 $\mu$M CaCl$_2$ and MgCl$_2$. The nuclear donor embryos of 8-cell stage were synchronized to G$_1$ phase of 16-cell stage, and the recipient cytoplasms were obtained from removal of the first polar body and a portion of membrane bound cytoplasm of the oocytes collected at 15 h post hCG injection. A separated blastomere was injected into the perivitelline space of the enucleated oocytes. The oocytes injected with nucleus were cultured until 19 h post hCG and then electrofused and activated by electrical stimulation with or without ionomycin and 6-DMAP. These nuclear transplant embryos were cultured in TCM-199 containing 10% FBS in 39˚C, 5% CO2 incubator for 120 h. For the oncytes activated parthenogenetically with electrical stimulation with or with-out ionomycin and the various concentration of exposure time of 6-DMAP, the highest cleavage(92.3%) and development to blastocyst stage(41.0%) were resulted from the oocytes activated by ionomycin and 2.0 mM 6-DMAP for 2.0 h, which were found to be significantly(P<0.05) higher than the cleavage(45.2%) and developement to blastocyst stage(14.3%) from the oocytes activated with electrical stimulation. The significantly(P<0.05) more oocytes(71.4%) developed to 4 cell stage at 24 h post activation by ionomycin and 6-DMAP than those by electrical stimulation(18.9%). For the nuclear transplant embryos, the cleavage rate was similarly high in oocyte activation by electrical stimulation with(79.4%) or without ionomycin and 6-DMAP(70.5%). However, the embryo development to blastocyst stage was significantly(P<0.05) higher in oocyte activation by electrical stimulation with ionomycin and 6-DMAP(44.4%) than by electrical stimulation only(25.0%). The significantly(P<0.05) more nuclear transplant embryos(45.6%) developed to 4 cell stage at 18 h post activation by electrical stimulation with ionomycin and 6-DMAP than those by electrical stimulation only(10.6%). These results indicated that the supplemental oocyte activation by ionomycin and 6-DMAP with electrical stimulation enhanced and accelerated the preimplanted in vitro development of the rabbit nuclear transplant embryos.

• Journal of Periodontal and Implant Science
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• v.28 no.1
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• pp.133-143
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• 1998

Diabetes mellitus is a systemic disease with profound effects on oral health and periodontal wound healing. Uncontrolled diabetes adversely affects surgical wound healing and is often associated with abnormal proliferation of fibroblasts. Human gingival fibroblasts and PDL cells were chosen because they are intimately involved in periodontal therapy and are important for the success of surgical procedure such as guided tissue regeneration. The aim of the present study was to elucidate whether cellular activity and collagen synthesis by glucose pre-treated human gingival fibroblasts and PDL cells are influenced by insulin, and whether healthy cells differ from glucose treated cells. Cells were cultured with DMEM at $37^{\circ}C$, 5% $CO_2$, 100% humidified incubator. To evaluate the effect of glucose on gingival fibroblasts and periodontal ligament cells, the cells were seeded at a cell density of $1{\times}10^4\;cells/well$ culture plates and treated with 20 and 50mM of glucose for 5 days. Then MTT assay was carried out. To evaluate the effect of insulin on glucose-pretreated cells, the cells were seeded at a cell density of $1{\times}10^4\;cells/well$ culture plates and treated with 20 and 50mM of glucose for 5 days. After incubation, $10^3$, $10^4$ and $10^5mU/l$ of insulin were also added to the each well and incubated for 2 days, respectively. Then, MTT assay and collagen synthesis assay were carried out. The results indicate that cellular activity of gingival fibroblasts significantly increased by glucose while periodontal ligament cells were unaffected and cellular activity of gingival fibroblasts and periodontal ligament cells were unaffected by insulin. Collagen synthesis of gingival fibroblast with 20mM glucose and insulin unaffected, but 50mM glucose and insulin increased than control. Collagen synthesis of periodontal ligament cell with 20mM glucose and $10^5mU/l$ insulin significantly increased than other groups and 50mM glucose pretreated PDL cells significantly increased at $10^3mU/l$ insulin but decreased at $10^4mU/l$ insulin. Our findings indicated that these cell types differed in their growth response to glucose, and the increase in collagen synthesis was significantly raised at insulin level of $10^3mU/l$ in gingival fibroblasts and periodontal ligament cells except 20mM glucose pretreated periodontal ligament cells.