• THE JOURNAL OF KOREAN ORIENTAL ONCOLOGY
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• v.6 no.1
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• pp.81-97
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• 2000

Objectives: This experimental study was carried out to evaluate the effects of aqueous and methanol extracts of Hedyotis diffusa which has long been used for cancer treatment in oriental medicines on the induction of apoptotic cell death in human lymphoid leukemia cell line, HL-60. Methods: Cells were treated with various concentrations (200 to $0.4{\mu}g$) and periods (6 to 30 hr) of $H_2O$ and methanol extracts of Hedyotis diffusa. Then, cells were tested for viability by MTT assay. Cells wrere treated with $200{\mu}g/ml$ of methanol extract fork various periods. Genomic DNA was isolated, separated, on 1.5% agarose gels, stained with ethidium bromide and visualized under UV light. Cells were treated with $200{\mu}g/ml$ of each extract for 16 hr. Then, cells were treated with Hoechst dye 33342 and observed by fluorescence microscopy. Cells were treated with various doses of each for 12 hr and $100{\mu}g/ml$ of methanol extract for various periods. Lysate from the cells used to measure the activity of Caspase-1 and-3 proteases by using fluorogenic peptide substrates including acetyl-YVAD-AMC and acetyl-DEVD-AMC, respectively. Cells were treated with $200{\mu}g/ml$ of each extract for various periods. Cell lysates were immunoprecipated with anti-JNKl antibodies. The immune complex was reacted with $32^p-ATP$ and c-Jun as a substrate. The phosphotransferase activity of JNKI was measured by using PhosphoImage analyzer (Fuji Co., Japan). Nuclear extracts were isolated and incubated with oligonucleotide probe of $NF-{\kappa}B$. Transcriptional activation of ${\kappa}B$ was measured by using EMSA and visualized by PhosphoImage analyzer (Fuji Co, Japan). Cell lysates were prepared and analyzed by Western blotting with anti-Bc12 antibodies and anti-Bax antibodies. Cells were pretreated with various doses of methanol extract for 2 hr. Then, the extract was removed by centrifugation. Cells were resuspended with RPMI-1640 media containing 0.3% agarose, 10% FBS, overlayred onto bottom layer agarose and incubated at $CO_2$ incubator for 6 days. The number of colony was counted under light microscopy ($\time100$). Results: The death of HL-60 cells was markedly induced by the addition of methanol extract of Hedyotis diffusa in a dose and time-dependent manners. The apoptotic characteristic ladder pattern of DNA strand break was observed in death of HL-60 cells. In addition, it was shown nucleus chromatin condensation and fragmentation under Hoechst staining. Therefore, Hedyotis diffusa extract-induced death of HL-60 cells is mediated by apoptotic signaling processes. The activity of Caspase 3-like proteases remained in a basal level in HL-60 cells treated with aqueous extract of Hedyotis diffusa. However, it was markedly increased in HL-60 cells treated with methanol extract of Hedyotis diffusa. In addition, the phosphotransferase activity of JNKl was increased in HL-60 cells treated with methanol extract of Hedyotis diffusa. Furthermore, the activation of transcriptional activator, $NF-{\kappa}B$ was markedly induced by methanol extract of Hedyotis diffusa. Anti-apoptotic Bc12 was cleaved into 23Kda fragment by treatment of methanol extract of Hedyotis diffusa. However, expression of proapoptotic Bax protein was increased by treatment of methanol extract of Hedyotis diffusa in a time-dependent manner. Furthermore, methanol extract markedly inhibited the colony forming efficiency of HL-60 cells in semisolid agar culture. Conclusions: Above results suggest that methanol extract of Hedyotis diffusa induces the apoptotic death of human leukemic HL-60 cells via activations of Caspase-3 proteases, JNKI, transcriptional activator $NF-{\kappa}B$, In addition, our results also suggest that methanol extract of Hedyotis diffusa reduces the malignant potential of HL-60 cells via down regulation of colony forming effciency through cleavage of Bc12 as well as induction of Bax.

• Clinical and Experimental Reproductive Medicine
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• v.29 no.1
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• pp.1-12
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• 2002

Objective: In order to acquire the technique for the establishment of human embryonic stem cells (ESe) derived from the human frozen-thawed embryos produced in IVF-ET program, this study was performed to establish mouse ESC derived from the in vitro fertilized embryos. Materials and Methods: After Fl hybrid (C57BL female $\times$ CBA mael) female mice were superovulated with PMSG and hCG treatment, their oocytes were retrieved and inseminated, and the fertilized embryos were cultured for 96-120 hours until the expected stages of blastocysts were obtained. To isolate the inner cell mass (ICM), either the blastocysts were treated with immunosurgery, or the whole embryos were cultured for 4 days. Isolated ICMs were then cultured onto STO feeder cell layer, and the resultant ICM colonies were subcultured with trypsin-EDTA treatment. During the subculture process, ESC-like cell colonies were observed with phase contrast microscopy. To identify ESC in the subcultured ESC-like cell colonies, alkaline phosphatase activity and Oct-4 (octamer-binding transcription factor-4) expression were examined by immunohistochemistry and RT-PCR, respectively. To examine the spontaneous differentiation, ESC-like cell colonies were cultured without STO feeder cell layer and leukemia inhibitory factor (LIF). Results: Seven ESC-like cell lines were established from ICMs isolated from the in vitro fertilized embryos. According to the developmental stage, the growth of ICMs isolated from the expanded blastocysts was significantly better than that of ICMs isolated from the hatched blastocysts (80.3% vs. 58.7%, p<0.05). ESC-like cell colonies were only obtained from ICMs of expanded blastocysts. However, the ICMs isolated from the embryos treated with immunosurgery were poorly grown and frequently differentiated during the culture process. The established ESC-like cell colonies were positively stained with alkaline phosphatase and expressed Oct-4, and their morphology resembled that observed in the previously reported mouse ESC. In addition, following the extended in vitro culture process, they maintained their expression of cell surface markers characteristic of the pluripotent stem cells such as alkaline phosphatase and Oct-4. When cultured without STO feeder cell layer and LIF, they were spontaneously differentiated into the various types of cells. Conclusion: The findings of this study suggest that the establishment of mouse ESC can be successfully derived from the in vitro fertilized embryos. The established ESC-like cells expressed the cell surface markers characteristic of the pluripotent stem cells and spontaneously differentiated into the various types of cells.

• Journal of Animal Environmental Science
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• v.18 no.3
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• pp.257-266
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• 2012

Near infrared reflectance spectroscopy (NIRS) has become increasingly used as a rapid, accurate method of evaluating some chemical constituents in cereal and dired animal forages. Analysis of forage quality by NIRS usually involves dry grinding samples. Costs might be reduced if samples could be analyzed without drying or grinding. The objective of this study was to investigate effect of sample preparations on prediction ability of chemical composition and fermentation parameter for Italian ryegrass silages by NIRS. A population of 147 Italian ryegrass silages representing a wide range in chemical parameters were used in this investigation. Samples were scanned at 1nm intervals over the wavelength range 680-2500 nm and the optical data recorded as log 1/Reflectance (log 1/R) and scanned in oven-dried grinding and fresh ungrinding condition. The spectral data were regressed against a range of chemical parameters using partial least squares (PLS) multivariate analysis in conjunction with four spectral math treatments to reduced the effect of extraneous noise. The optimum calibrations were selected on the basis of minimizing the standard error of cross validation (SECV) and maximizing the correlation coefficient of cross validation (${R^2}_{CV}$). The results of this study show that NIRS predicted the chemical parameters with high degree of accuracy in oven-dried grinding treatment except for moisture contents. Prediction accuracy of the moisture contents was better for fresh ungrinding treatment (SECV 1.37%, $R^2$ 0.96) than for oven-dried grinding treatments (SECV 4.31%, $R^2$ 0.68). Although the statistical indexes for accuracy of the prediction were the lower in fresh ungrinding treatment, fresh treatment may be acceptable when processing is costly or when some changes in component due to the processing are expected. Results of this experiment showed the possibility of NIRS method to predict the chemical composition and fermentation parameter of Italian ryegrass silages as routine analysis method in feeding value evaluation and for farmer advice.

• Korean Journal of Environmental Biology
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• v.37 no.2
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• pp.155-163
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• 2019

The Paralichthys olivaceus (mean weight 34.3±3.5 g) was used in the conduction of density experiment in this study to establish the optimal density determination for a period of 13 weeks. The density consisted of four groups. These were group 1 (500 fish, 40 fish m^-2), group 2 (750 fish, 60 fish m^-2), group 3 (1,000 fish, 80 fish m^-2) and group 4 (1,250 fish, 100 fish m^-2), each in 12.56 ㎡ circular water tanks. In the high density groups, such as groups 1 and 2, the nitrite was increased to over 21 mg L^-1 (group 3) and 25 mg L^-1 (group 4). The experiment of the groups was terminated by skin ulcers and mortality after 49 days for group 3 and 43 days for group 4. The water quality environment, such as the water temperature, dissolved oxygen, salinity and pH, remained constant. The ammonia and nitrite in groups 1 and 2 remained stable after ammonia and nitrite peak. However, the experiments in groups 3 and 4 were terminated due to the mortality induced by high nitrite. Hematological parameters, such as hemoglobin and hematocrit, were significantly decreased in the high density groups. The plasma components were significantly changed in the high density groups, such as groups 3 and 4. The mean weight of groups 1 and 2 after 13 weeks were 91.3 g and 83.7 g, respectively. The survival rates were 99.4% and 98.2%, respectively. The final FCR was 0.6 in both groups. The results of this study show that the density of 80 fish m^-2 or more induce mortality due to high nitrite with hematological changes. Additionally, they indicate that the 60 fish m^-2 indicate proper density in bio-floc environment in olive flounder weighing less than 100 g.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.7
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• pp.3349-3355
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• 2012

Background and objectives: Chromosomal abnormalities play an important role in genesis of acute lymphoblastic leukemia (ALL) and have prognostic implications. Five major risk stratifying fusion genes in ALL are BCR-ABL, MLL-AF4, ETV6-RUNX11, E2A-PBX1 and SIL-TAL1. This work aimed to detect common chromosomal translocations and associated fusion oncogenes in adult ALL patients and study their relationship with clinical features and treatment outcome. Methods: We studied fusion oncogenes in 104 adult ALL patients using RT-PCR and interphase-FISH at diagnosis and their association with clinical characteristics and treatment outcome. Results: Five most common fusion genes i.e. BCR-ABL (t 9; 22), TCF3-PBX1 (t 1; 19), ETV6-RUNX1 (t 12; 21), MLL-AF4 (t 4; 11) and SIL-TAL1 (Del 1p32) were found in 82/104 (79%) patients. TCF3-PBX1 fusion gene was associated with lymphadenopathy, SIL-TAL1 positive patients had frequent organomegaly and usually presented with a platelets count of less than $50{\times}10^9/l$. Survival of patients with fusion gene ETV6-RUNX1 was better when compared to patients harboring other genes. MLL-AF4 and BCR-ABL positivity characterized a subset of adult ALL patients with aggressive clinical behaviour and a poor outcome. Conclusions: This is the first study from Pakistan which investigated the frequency of5 fusion oncogenes in adult ALL patients, and their association with clinical features, treatment response and outcome. Frequencies of some of the oncogenes were different from those reported elsewhere and they appear to be associated with distinct clinical characteristics and treatment outcome. This information will help in the prognostic stratification and risk adapted management of adult ALL patients.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.28 no.6
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• pp.812-813
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• 1995

Lactococcal cells are nutritionally fastidious and thus, generally cultured either in milk or M17 medium (Terzaghi and Sandine, 1975). In this study, Lactococcus cremoris wild-type (KH) and its less­proteolytic mutant (KHA1) cells were grown on the M17 medium or with modified M17 medium by replicated parallel experiments. The modified M17 medium had the same composition as M17 medium, except that lactose was replaced by glucose. Analyses of culture-broth samples, in which the M17 and the modified M17 media were used, were conducted by high-performance liquid chromatography (HPLC). But, working with these media created noisy problems in analyses of samples. Therefore, a new semi-synthetic medium was developed on the basis of nutritional requirements (Morishita et al., 1981). The composition of the semi-synthetic medium determined on the basis of the nutritional requirements and the composition of milk, is presented in Table 1. The composition of M17 medium is also presented and compared in the table. L. cremoris KH and KHA1 cells were grown again on the new synthetic medium containing glucose or lactose. The broth samples were then drawn and analyzed by HPLC. Clearer separations of fermented products were achieved from the new medium than those with the M17 and the modified M17 media. In comparison with the M17 or the modified M17 media, growth on the new medium was good (Kim et al, 1993). Additional fermentations were also carried out at a controlled pH of 7.0, where enhanced growth of lactococcal cells was obtained. In the fermentations, samples were also analyzed for the concentrations of sugar and lactic acid. The results showed that the new synthetic medium was as good as or better than the M 17 and the modified M 17 media. This is because casein hydrolysate in the synthetic medium provided a ready supply of amino acids and peptides for L. cremoris KH and KHA1 cells. Lactic acid bacteria (LAB) including Lactococcal cells have been known to be an effective means of preserving foods, at the same time as giving particular tastes in fields of dairy products. LAB also have always occupied an important place in the technology of sea products, and marine LAB have known to be present in traditional fermented products (Ohhira et al, 1988). To apply the new synthetic medium to marine LAB, two different LAB were isolated from pickled anchovy and pollacks caviar and were grown on the new media in which various concentrations of NaCl $(3, 5, 7 and 10\%)$ added. They were also grown on the medium solution in natural seawater $(35\%o\;salinity)$ and on the solution of natural seawater itself, too. As seen in Fig. 1, Marine LAB were grown best on the synthetic medium solution in natural seawater and the higher concentrations of NaCl were added to the medium, the longer lag-phase of growth profile appeared. Marine LAB in natural seawater were not grown well. From these results, the synthetic medium seems good to cultivate cells which are essential to get salted fish aged. In this study, it showed that the new synthetic medium provided adequate nutrition for L. cremoris KH and KHA1 cells, which have been used as cheese starters (Stadhouders et al, 1988). Using this new medium, the acid production capability of starter cultures could be also measured quantitatively. Thus, this new medium was inferior to the M17 or the modified M17 medium in culturing the cheese starters and in measuring fermentation characteristics of the starter cells. Moreover, this new medium found to be good for selected and well-identified marine LAB which are used in rapid fermentations of low-salted fish.

• Journal of Korean Society of Forest Science
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• v.83 no.3
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• pp.286-298
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• 1994

The object of this study was to examine the user's impacts on the environmental deteriorations of trail at Ticket Office - Paekryunsa (Temple) Hyangch$\hat{o}$kpong - Dongyupryung - Chilyun Fall area in T$\hat{o}$kyusan National Park. Four trails were sampled in the study area according to the amount of users. Then the user's impacts on trail were measured at each trail. The Ticket Office-Paekryunsa trail was the most used district and followed at Paekryunsa-Hyangch$\hat{o}$kpong trail, Hyangch$\hat{o}$kpong-Dongyupryung trail in descending order. Dongyupryung-Chilyun Fall trail is not used by people because of rest rotation system. The entire width of trail was greater at the more heavily used trail. Maximum depth, cross-sectional area loss, and surface texture and roughness of trail were the highest at Paekryunsa-Hyangch$\hat{o}$kpong trail. Soil hardness, soil acidity, soil moisture content, organic matter content, and exchange canon were influenced by trampling. Soil hardness, soil acidity and exchange canon increased in tramples soil, but content of soil moisture and organic matter decreased therein. Environmental deteriorations of trail were significantly influenced by the amount of users and the slope of trail. Bared lands about $2.000m^2$ were appeared by trampling and camping around Hyangch$\hat{o}$kpong. Effects of carrying of rest rotation system for National Park were partly recognized at Dongyupryung-Chilyun Fall trail.

• KSBB Journal
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• v.29 no.4
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• pp.285-296
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• 2014

Although the problems of the algal blooms have been world-widely observed in freshwater, estuary, and marine throughout the year, it is not yet certain what are the basic causes of such blooms. Consequently, it is very difficult to predict when and where algal blooms occur. The constituents of the Asian dust are in a good agreement with the elements required for the algal growth, which suggests some possible relationship between the algal blooms and the Asian dust. There have been frequently algal blooms in drinking water from rivers or lakes. However, there is no any algal blooms in upwelling waters where the Asian dust cannot penetrate into the soil due to its relatively weak settling velocity (size of particles, $4.5{\pm}1.5{\mu}m$), which implies the possible close relationship of the Asian dust with algal blooms. The present initiative study is thus intended firstly in Korea to illustrate such a relationship by reviewing typical previous studies along with 12 years of weekly iron profiles (2001~2012) and two slant culture experiments with the dissolved Asian dust. The result showed bacterial suspected colonies in the slant culture experiment that are qualitatively in a good agreement with the recent Japanese studies. Since the diatoms require cheap energy (8%) compared to other phytoplankton (100%) to synthesize their cell walls by silicate, the present results can be used to predict algal blooms by diatoms if the concentrations of iron and silicate are available during spring and fall. It can be postulated that the algal blooms occur only if the environmental factors such as light, nutrients, calm water surface layer, temperature, and pH are simultaneously satisfied with the requirements of the micronutrients of mineral ions supplied by the Asian dust as enzymatic cofactors for the rapid bio-synthesis of the macromolecules during algal blooms. Simple eco-friendly methods to regulate the algal blooms are suggested for the initial stage of blooming with limited area: 1) to cover up the water surface with black curtain and inhibit photosynthesis during the day time, 2) to blow air (20.9%) or pure oxygen into the bottom of the water and inhibit rubisco for carbon uptake and nitrate reductase for nitrogen uptake activities in algal growth during the night, 3) to eliminate the resting spores or cysts by suction of bottom sediments as deep as 5 cm to prevent the next year germinations.

• Journal of the Korea Organic Resources Recycling Association
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• v.9 no.1
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• pp.56-64
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• 2001

Composting of livestock feces is economic and safe process to decrease the possibility of direct leakage of organic pollutants to ecosystem from commercial and environmental point of view. This study was conducted with three different experiments related to composting of livestock feces. The purpose of experiment 1 was to investigate changes of characteristic of compost pile during composting period by low temperature in cold season. To compare composting effect of experimental compost pile and control pile exposed in cold air, experimental compost piles were warmed up by hot air until their temperatures were reached at $35^{\circ}C$. Sawdust, Ricehull and Ricestraw were mixed with livestock feces as bulking agent. The highest temperatures of compost pile during composting period were in sawdust, rice hull, rice straw, and control were $75^{\circ}C$, $76^{\circ}C$, $68^{\circ}C$, $45^{\circ}C$ respectively. Moisture content, pH, C/N and volume of compost were decreased during composting period. Experiment 2 was carried out to study utilization effect of compost by plant. A corn was cultivated for 3 years on fertilized land with compost and chemical fertilizer. The amount of harvest and nutrition value of corn were analyzed. In first year of trial, the amount of harvest of corn on land treated with compost was lower by 20% than that of land treated with chemical fertilizer. In second year, there was no difference in yield of com between compost and chemical fertilizer. In third year, the yield of com on land fertilized with compost was much more than that of land fertilized with chemical fertilizer. The purpose of experiment 3 was to estimate the decrease of malodorous gas originating from livestock feces by bio-filter. Four types of bio-filters filled with saw dust, night soil, fermented compost and leaf mold were manufactured and tested. Each bio-filter achieved 87-95% $NH_3$ removal efficiency. This performance was maintained for 10 days. The highest $NH_3$ removal efficiency was achieved by leaf mold on the first day of operation period. It reduced the concentration of $NH_3$ by about 95%. Night soil and fermented compost showed nearly equal performance of 93 to 94% for 10 days from the beginning of operation. The concentration of hydrogen sulfide and methyl mercaptan originating for compost were equal to or less than $3mg/{\ell}$ and $2mg/{\ell}$, respectively. After passing throughout the bio-filter, hydrogen sulfide and methyl mercaptan were not detected.

• Korean Journal of Environmental Biology
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• v.25 no.4
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• pp.303-312
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• 2007

In order to investigate the dynamics of phytoplankton standing crops affecting by environmental factors, biological and environmental factors, this study was examined in the marine ranching ground of Tongyeong coastal waters from 2000 to 2007. During the study, mean water temperature and salinity were 16.7$^{\circ}C$ and 32.9 psu, respectively. pH, DO and SS varied from 7.81$\sim$8.09, 3.02$\sim$8.97 mg $L^{-1}$ and 2.7$\sim$32.2 mg $L^{-1}$, respectively. Mean concentrations of dissolved inorganic nitrogen, phosphate and silicate were 21.75 ${\mu}M$, 0.90 ${\mu}M$ and 14.38 ${\mu}M$, respectively. Chlorophyll a concentrations varied from 0.02 ${\mu}g$ $L^{-1}$ to 25.29 ${\mu}g$ $L^{-1}$ with mean a value of 2.0 ${\mu}g$ $L^{-1}$. These factors did show significant differences on each layer and season, while did not show on the sampling stations. Phytoplankton standing crops varied from $4.21\times10^3$ cells $L^{-1}$ to $1.44\times10^6$ cells $L^{-1}$ with a mean value of $1.92\times10^5$ cells $L^{-1}$. Especially, variations of phytoplankton standing crops had an unimodal pattern as only bloomed in autumn rather than a bimodal pattern as generally bloomed in spring and autumn. In results of stepwise multiple regression analysis, the coefficient of determination $(R^2)$ for total standing crops was 0.35 and the standing crops were affected by water temperature, salinity, phosphate and silicate. The factors affected were different seasonally; water temperature in spring, salinity in summer, water temperature, salinity and silicate in autumn and water temperature, salinity and suspended solids in winter. Therefore, the results from the statistical analysis showed that the environmental factors influencing on the variations of the phytoplankton standing crops were predominantly water temperature and salinity.