• 대한한방부인과학회지
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• 제19권1호
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• pp.166-177
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• 2006

Purpose : This investigation was undertaken to evaluate the antiproliferation, atoptosis of Sam-nueng(Sparganii Rhizoma) extract using MCF-7 human breast cancer cells. Methods : MCF-7 cells were cultured in Dulbecco's modified Eagle's medium/F12(DMEM/F12) supplemented with 10 % fetal bovine serum(FBS; Gibco) and antibiotics. At varying times after extract treatment, cells were harvested with scraper and processed for analysis of protein expression, proliferation, cytotoxicity and apoptosis. Results : Our results show that the extract of Sam-nueng(Sparganii Rhizoma) strongly inhibits the proliferation of MCF-7 cells in a dose and time-dependent manner. Sulforhodamine B showed that the addition of Sam-nueng(Sparganii Rhizoma) extract reduced the viability of MCF-7 cells in a dose-dependent manner. Poly[ADP(ribose)] polymerase(PARP) which serves as a marker of cells undergoing apoptosis, a major substrate for caspase-3 was extensively cleaved in the Sam-nueng(Sparganii Rhizoma)-treated cells. Conclusion : So, we can conclude that Sam-nueng(Sparganii Rhizoma) can have an inhibitive effect on MCF-7 human breast cancer cells by variable mechanisms.

• Clinical and Experimental Reproductive Medicine
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• 제27권1호
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• pp.1-7
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• 2000

Objective: Mammalian embryos undergo changes of energy environment for transfer from oviduct to uterus. Also, the human reproductive organ (oviduct, uterus) contains energy sources of different concentration (oviduct - glucose: 0.5 mM, pyruvate: 0.32 mM, lactate: 10.5 mM; uterus - goucose: 3.15 mM, pyruvate: 0.1mM, lactate: 5.87 mM, respectively). This study was conducted to examine the effect of these energy sources added in DMEM with glutamine on the mouse embryo development. Methods: There was used ICR female mouse. Two cell embryos of mouse are collected by method of 'flushing'. Flushing fluid was used Ham's F-10 added to 20% FBS. The collected 2 cell embryos were cultured in media such as Control (only DMEM), group A and B (DMEM supplemented with 0.5 mM and 3.15 mM glucose), and group C and D (DMEM supplemented with 0.1 mM and 0.32 mM pyruvate), and group E and F (DMEM supplemented with 5.87 mM and 10.5 mM lactate). All experimental media supplemented with 20% hFF, respectively. Pattern of embryo development was observed to interval at 24hr during 96hr. Results : The media with glutamine added glucose (group A: 51.0%; group B: 48.4%) was significantly (p<0.05) higher than other experimental group in development into the morula stage after 24 hr in culture, but not significantly different compared with control and the rate of development into the blastocyst was significantly (p<0.05) low in the both of pyruvate (group C: 7.9% group D: 6.8%) and lactate (group E: 7.1%, group F: 7.1%) treatment group after 48 hr in culture. Development into the blastocyst and hatched balstocyst after 72 hr in culture revealed similarly in control (81.9%) and glucose treatment group (group A: 83.3%, group B: 82.8%). However, development into the hatched and attached blastocyst after 96hr in culture revealed significantly (p<0.05) development in the glucose treatment group (group A: 82.3%, group B: 78.5%) than control (63.2%), and its of pyruvate (group C: 34.1%, group D: 34.1%) and lactate (group E: 25.9%, group F: 33.3%) treatment group were significantly (p<0.05) lower than control similar to previous observations. Conclusion : The glucose added to the DMEM with only glutamine, as energy source, was highly to the rate of development compared with control, but the other energy sources were not, synthetically. Above refer to, the human reproductive organ (oviduct, uterus) contains energy sources of different concentration. Thus, further studies are will examine continuously to effects by interaction of different energy sources in the mouse embryo development, and these results will provide to foundation on the human embryo culture.

• Journal of Animal Science and Technology
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• 제64권3호
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• pp.481-499
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• 2022

This study aims to determine the effects of D-methionine (D-Met) isomer and the methionine precursor 2-hydroxy-4-methylthiobutanoic acid i (HMBi) supplementation on milk protein synthesis on immortalized bovine mammary epithelial cell (MAC-T). MAC-T cells were seeded using 10-cm dishes and cultured in Dulbecco's modified Eagle's medium/F12 (DMEM/F12) basic medium. The basic medium of DMEM/F12 was replaced with the lactogenic DMEM/ F12 differentiation medium when 90% of MAC-T cells reached confluency. The best dosage at 0.6 mM of D-Met and HMBi and incubation time at 72 h were used uniformly for all treatments. Each treatment was replicated six times wherein treatments were randomly assigned in a 6-well plate. Cell, medium, and total protein were determined using a bicinchoninic acid protein assay kit. Genes, proteomics and metabolomics analyses were also done to determine the mechanism of the milk protein synthesis pathway. Data were analyzed by two-way analysis of variance (ANOVA) with supplement type and plate as fixed effects. The least significant difference test was used to evaluate the differences among treatments. The HMBi treatment group had the highest beta-casein and S6 kinase beta-1 (S6K1) mRNA gene expression levels. HMBi and D-Met treatments have higher gene expressions compared to the control group. In terms of medium protein content, HMBi had a higher medium protein quantity than the control although not significantly different from the D-Met group. HMBi supplementation stimulated the production of eukaryotic translation initiation factor 3 subunit protein essential for protein translation initiation resulting in higher medium protein synthesis in the HMBi group than in the control group. The protein pathway analysis results showed that the D-Met group stimulated fructose-galactose metabolism, glycolysis pathway, phosphoinositide 3 kinase, and pyruvate metabolism. The HMBi group stimulated the pentose phosphate and glycolysis pathways. Metabolite analysis revealed that the D-Met treatment group increased seven metabolites and decreased uridine monophosphate (UMP) production. HMBi supplementation increased the production of three metabolites and decreased UMP and N-acetyl-L-glutamate production. Taken together, D-Met and HMBi supplementation are effective in stimulating milk protein synthesis in MAC-T cells by genes, proteins, and metabolites stimulation linked to milk protein synthesis.

• Journal of Periodontal and Implant Science
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• 제25권2호
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• pp.210-221
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• 1995

The purpose of this study was performed to investigate the mineralization and differentiation of osteobalsts for bone regeneration in vitro and the effect of rate of the composition in periodontal cells on mineralization. For this study, healthy gingival tissues were surgically obtained from the patients during 1st premolar extraction for the purposes of orthodontic treament. Gingival tissue was washed several time with Phosphate buffered saline contained high concentration of antibiotics and antifungal agent, and cultured in Dulbecco's Modified Eagle's Medium(DMEM, Gibco, U.S.A.). Every cell were cultured in state at $37^{\circ}C$, 100% of humidity, 5% of $CO_2$ incubator. Bone marrow stromal cells were isolated from 5-clay-old rat femur with using medium irrigation mathod by syringe. Cell suspension medium were centrifuged at 1500 rpm for 5 min and then cultured in the petri dish. Two kinds of cell were freezed and stocked in the liquid nitrogen tank until experiment. Cell were incubated into the 24 multi-well plate with $5{\times}10^4$cell/well of medium at $37^{\circ}C$, 100% of humidity 5% $CO_2$ incubator for 24 hours. After discarded of the supernatent of medium, O.5ml of medium were reapplied and incubated. And counted the number of cell using the hemocytometer and inverted light microscope. We have measured the number of mineralized nodule with using Alizarin red S. staining in microscope. Furthermore every cell were observed the morphological change between every rate of co-culture of the two kinds of cell. The results were as follows; The rate of proliferation of co-culture cell revealed high rate tendency compared the bone marrow stromal cell only and low growth rate to compared with gingival fibroblast only. The tendency of formation of the mineralized nodule were observed dose-depend pattern of bone marrow stromal cell. It is concluded that the gingival fibroblast may inhibit the formation of mineralized nodule in the culture of the bone marrow stromal cell.

• Journal of Ginseng Research
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• 제10권2호
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• pp.141-150
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• 1986

This study was attempted to screen the cytotoxic activity of petroleum ether ex- tract from panax ginseng root against human colon cancer cells. Two extracts of panax ginseng root, crude and partially purified, were used for this experiment. The crude extract was prepared by extraction with petroleum ether using Soxhlet aparatus for 12 to 15 hours from panax ginseng and the extract was partially purified by silicic acid column with mixture of petroleum ether: ethyl ether (70 : 30, v/v). Three species of human colon cancer cells, HRT-18, HCT-48 and HT-29, were maintained in DMEM (Dulbecco's modified Eagle medium), and the cells were cultured in DMEM containing serial concentration of the crude or partially purified fraction to observe the cytotoxic activity of the both extracts. The effects of incubation time and concentration of the both extracts in culture medium against the growth of the each cancer cell were determined. The results obtained are summarized as follows: 1. The doubling times of the HRT-18, HCT-48 and HT-29 cells were about 20, 24 and 22 hours, respectively. 2, The inhibitory effects of the crude extract on the growth of cancer cells were increased according to the rise of concentration of the extract and incubation time. 3. The inhibitory effect of partially purified fraction on the growth of HRT-18 cell was about 4 times stronger than that of the crude extract under same experimental condition. 4 The inhibitory effects of the crude and purified fraction on the growth of each cancer cell were shown difference by the kind of the cancer cell. In view of the above results, it could be said that the petroleum ether extract of panax ginseng root inhibited the division of the human colon cancer cell, in vitro.

• 사상체질의학회지
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• 제13권1호
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• pp.51-60
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• 2001

Effects of Taeumjowetang on Lipid Peroxidation by Free Radicals and Oxidative Damage of Hepatocytes by tert-Butyl Hydroperoxide. 1. Purpose The present study was carried out to evaluate the antioxidant effects of Taeumjowetang in vitro. 2. Methods In this study, antioxidant effects of TJT on lipid peroxidation were determined according to the method of TBA. (Abbreviation) TJT : Taeumjowetang, TBA : 2-thiobarbituric acid. 3. Results : 1) TJT inhibited markedly peroxidation of linoleic acid during the autoxidation. 2) TJT inhibited lipid peroxidation induced by hydroxyl radical derived from H2O2-Fe2+ in rat liver homogenate. 3) TJT showed 66% scavenging effect on DPPH radical. 4) TJT exhibited a 25% inhibitory effect on superoxide generation from xanthine-xan thine oxidase system. 5) To investigate the antioxidative effects of TJT on the hepatocytes, cultured normal rat liver cells(Ac2F) were prepared and incubated with or without TJT. After 16~18hr, cells placed in DMEM medium without serum, and then incubated with 1mM t-BHP for 2hr. Viable cells were detected by MTT assay. In this test, TJT protected the cell death induced by t-BHP and significantly increased cell viability in the normal rat liver cell. (Abbreviation) DPPH : ${\alpha},{\alpha}$-diphenyl-${\beta}$-picryl hydrazyl, DMEM : Dulbecco's Modified Eagle Medium, t-BHP : terr-butyl hydroperoxide, 4. Conclusion These results suggested that TJT might play a protective role in lipid peroxidation by free radicals.

• 대한의용생체공학회:의공학회지
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• 제41권3호
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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.

• Restorative Dentistry and Endodontics
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• 제38권4호
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• pp.204-209
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• 2013

Objectives: The objective of this in vitro study was to evaluate and compare the cytotoxicity of four different root canal sealers i.e. Apexit Plus (Ivoclar Vivadent), Endomethasone N (Septodont), AH-26 (Dentsply) and Pulpdent Root Canal Sealer (Pulpdent), on a mouse fibroblast cell line (L929). Materials and Methods: Thirty two discs for each sealer (5 mm in diameter and 2 mm in height) were fabricated in Teflon mould. The sealer extraction was made in cell culture medium (Dulbecco's Modified Eagle's Medium, DMEM) using the ratio 1.25 $cm^2/mL$ between the surface of the sealer samples and the volume of medium in a shaker incubator. Extraction of each sealer was obtained at 24 hr, 7th day, 14th day, and one month of interval. These extracts were incubated with L929 cell line and 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) assay was done. Two-way ANOVA for interaction effects between sealer and time and Post-hoc multiple comparison using Tukey's test across all the 16 different groups were used for statistical analysis. Results: Apexit Plus root canal sealer was significantly less toxic than other sealers (p < 0.05) and showed higher cellular growth than control. Endomethasone N showed mild cytotoxicity. AH-26 showed severe toxicity which became mild after one month while Pulpdent Root Canal Sealer showed severe to moderate toxicity. Conclusions: Apexit Plus was relatively biocompatible sealer as compared to other three sealers which were cytotoxic at their initial stages, however, they became biocompatible with time.

• 대한본초학회지
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• 제20권4호
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• pp.27-31
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• 2005

Objectives : The anticancer response of three different types of water extracts of Zingiberaceae Curcuma longa L. tested for sarcoma 180. Only few studies carried out to investigate the effects of other contents of Curcuma longa L. in anticancer activities, therefore, in this study we have investigated the effects of other component then curcumin in Curcuma longa L. for anticancer a activities. Methods : Three different types of water extracts of Curcuma longa L. were prepared as follows. The sarcoma cells (S180) were maintained in Dulbecco's modified Eagle's medium (DMEM) and were seeded on 24-well cell culture cluster flat bottom with lid tissue culture treated non-pyrogenic polystyrene. The growth of sarcoma 180 was monitored for 1, 2 and 5 days. The sarcoma cells were pictured using inverted microscope and cell density was counted using hemocytometry. Results : After 5 days in the culture medium the results showed high growth of sarcoma 180 for control condition and the surface of CCP plates were fully covered with the cells. In case of medium in which the 10% of filtered water extract of Curcuma longa L. was added a very limited growth of sarcoma 180 was observed. The results were showed only small difference in cell density for two different concentrations of unfiltered water extracts of Curcuma longa L. whereasin case of filtered water extracts the control of sarcoma growth shows better result. Conclusion : The filtered water extracts showed the best result relatively to the unfiltered water extracts for two different concentrations. This indicates that the water extracts of Curcuma longa L. can have anticancer activities possibly without curcumin.

• 대한화장품학회지
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• 제27권1호
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• pp.17-27
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• 2001

A cell-based assay system for monitoring NF-$textsc{k}$B activity was developed to determine the influence of activated NF-$textsc{k}$B in human HaCaT cells. The pNF-$textsc{k}$B-SEAP-NPT plasmid that permits expression of the secreted alkaline phosphatase (SEAP) reported gene in response to the NF-$textsc{k}$B activity and contains neomycin phosphotransferase (NPT) gene for the geneticin resistance in host cells was constructed and transfected into human keratinocyte cell line HaCaT. Human HaCaT transfectant cells secreted the SEAP enzyme into the culture medium in a time-dependent manner until 72h. NF-$textsc{k}$B activities were measured in the SEAP reporter gene assay using a fluorescent detection method. The treatment of HaCaT cell transfectants with known antioxidants [e.g., N-acetyl-L-cysteine and vitamin C] showed inhibition of NF-$textsc{k}$B activity in a time-and concentration-dependent manner. The phorbol 12-myristate 13-acetate (PMA) known as a stimulator of NF-$textsc{k}$B expression demonstrated that it increased NF-$textsc{k}$B activity in a time- and concentration-dependent manner. This assay system could be used to determine the quantitative measurement of NF-$textsc{k}$B activity in the human skin and allow the screening of anti-inflammatory agents from various synthetic chemicals and natural products for dermatological purpose. Abbrevitions used: NF-$textsc{k}$B, nuclear factor kappa B; I-$textsc{k}$B, Inhibitory kappa B; SEAP, secreted alkaline phosphatase; NPT, neomycin phosphotransferease; PCR, polymerase chain reaction: dNTP, deoxynucleoside triphosphates; DMEM, dulbecco’s modified eagle medium; FBS, fetal bovine serum; PBs, phosphate-buffered saline; MUP, 4-methylumbellifery phosphate; NAC, N-acetyl-L-cysteine; DMSO, dimethyl sulfoxide; PMA, phorbol 12-myristate 13-acetate.