• Clinical and Experimental Reproductive Medicine
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• v.29 no.2
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• pp.139-147
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• 2002

Objective : This study was to establish a reproducible differentiation system from the parthenogenetic mouse embryonic stem (P-mES02) cells into functional cardiomyocytes like as in vitro fertilization mouse embryonic stem (mES01) cells. Materials and Methods: To induce differentiation, P-mES02 cells were dissociated and aggregated in suspension culture environment for embryoid body (EB) formation. For differentiation into cardiomyocytes, day 4 EBs were treated with 0.75% dimethyl sulfoxide (DMSO) for another 4 days (4-/4+) and then were plated onto gelatin-coated dish. Cultured cells were observed daily using an inverted light microscope to determine the day of contraction onset and total duration of continuous contractile activity for each contracting focus. This frequency was compared with the results of DMSO not treated P-mES02 group (4-/4-) and mES01 groups (4-/4+ or 4-/4-). For confirm the generation of cardiomyocytes, beating cell masses were treated with trypsin-EDTA, dispersed cells were plated onto glass coverslips and incubated for 48 h. Attached cells were fixed using 4% paraformaldehyde and incubated with specific antibodies (Abs) to detect cardiomyocytes (anti-sarcomeric ? -actinin Ab, 1 : 100; anti-cardiac troponin I Ab, 1 : 2000) for 1 h. And the cells were finally treated with FITC or TRITC labelled 2nd Abs, respectively, then they were examined under fluorescence microscopy. Results: Rhythmically contracting areas in mES01 or P-mES02 cells were firstly appeared at 9 or 10 days after EBs plating, respectively. The highest cumulative frequency of beating EBs was not different in both treatment groups (mES01 and P-mES02, 4-/4+) with the results of 61.3 % at 13 days and 69.8% at 15 days, respectively. Also, the contracting duration of individual beating EBs was different from minimal 7 days to maximal 53 days. However, DMSO not treated groups (mES01 and P-mES02, 4-/4-) also had contracting characteristics although their frequency was a few compared to those of DMSO treated groups (6.0% and 4.0%). Cells recovered from the spontaneously contracting areas within EBs in both treated groups were stained positively with muscle specific anti-sarcomeric ? -actinin Ab and cardiac specific anti-cardiac troponin I Ab. Conclusion: This study demonstrated that the P-mES02 cell-derived cardiomyocytes displayed similarly structural properties to mES01 cell-derived cardiomyocytes and that the DMSO treatment enhanced the cardiomyocytes differentiation in vitro.

• The Journal of Internal Korean Medicine
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• v.29 no.3
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• pp.554-566
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• 2008

Objectives : Modified Gamgil-tang is a prescription commonly used for respiratory diseases. This thesis was carried out to check the treatment effects and diversity of drug formulation by comparing extraction method of ethanol and water of modified Gamgil-tang. Methods : All experiments were carried out with water and 50% ethanol extraction for comparison. In vivo experiment, hyaluronidase inhibitory effects and trypsin inhibitory effects were tested to measure the anti-inflammatory effects activity. Scavenging effects of DPPH free radical, xanthine oxidase inhibitory effects and inhibition on TBA-RS formation were experimented to measure anti-oxidative effects. With the in vivo experiment, ICR group mice and SD group rats were used as experimental animals. An anti-inflammatory effects experiment were carried out to measure the action on carrageenin-induced hind paw edema: analgesic effects were measured using writhing syndrome induced by 0.7% acetic acid in mice: antipyretic effect was measured using endotoxin, and inhibitory effects of increase vascular permeability induced by 0.5% histamine were measured. Results : For extraction of glycyrrhizin contents, ethanol extract was extracted 2 times of that of water extract. Anti-inflammatory effects showed high in ethanol extract. Anti-oxidative effects measured high in ethanol extract. No significant result was found in inhibition on TBA-RS formation. Analgesic effects were found to be similar in water and ethanol extract. Antipyretic effects were found to be stronger in water extract. Inhibitory effects of increase vascular permeability induced by 0.5% histamine showed stronger in ethanol extract. Conclusion : By measuring anti-inflammatory effects, analgesic effects, antipyretic effects, anti-oxidative effects, and histamine permeation inhibition effects both in water extract and ethanol extract after adding agents such as Mentha Herba, Gardenias Fructus, and propolis to existing Gamgil-Tang, ethanol extract was found to be more effective in anti-inflammatory effects, analgesic effects, anti-oxidative effects, and histamine permeation inhibition effects. The converse was found for antipyretic effect.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.4
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• pp.534-541
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• 2009

The aim of this study was to evaluate the effects of gamma irradiation (${\gamma}$-irradiation) at doses of 15, 30 and 45 kGy on chemical composition, anti-nutritional factors, ruminal dry matter (DM) and crude protein (CP) degradibility, in vitro CP digestibility and to monitor the fate of true proteins of full-fat soybean (SB) in the rumen. Nylon bags of untreated or ${\gamma}$-irradiated SB were suspended in the rumens of three ruminally-fistulated bulls for up to 48 h and resulting data were fitted to a nonlinear degradation model to calculate degradation parameters of DM and CP. Proteins of untreated and treated SB bag residues were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Digestibility of rumen undegraded CP was estimated using the three-step in vitro procedure. The chemical composition of raw and irradiated soybeans was similar. Results showed that phytic acid in ${\gamma}$-irradiated SB at dose of 30 kGy was eliminated completely. The trypsin inhibitor activity of 15, 30 and 45 kGy ${\gamma}$-irradiated SB was decreased (p<0.01) by 18.4, 55.5 and 63.5%, respectively. From in sacco results, ${\gamma}$-irradiation decreased (p<0.05) the washout fractions of DM and CP at doses of 30 and 45 kGy, but increased (p<0.05) the potentially degradable fractions. Gamma irradiation at doses of 15, 30 and 45 kGy decreased (p<0.05) effective degradability of CP at a rumen outflow rate of 0.05 $h^{-1}$ by 4.4, 14.4 and 26.5%, respectively. On the contrary, digestibility of ruminally undegraded CP of irradiated SB at doses of 30 and 45 kGy was improved (p<0.05) by 12 and 28%, respectively. Electrophoretic analysis of untreated soybean proteins incubated in the rumen revealed that ${\beta}$-conglycinin subunits had disappeared at 2 h of incubation time, whereas the subunits of glycinin were more resistant to degradation until 16 h of incubation. From the SDS-PAGE patterns, acidic subunits of 15, 30 and 45 kGy ${\gamma}$-irradiated SB disappeared after 8, 8 and 16 h of incubation, respectively, while the basic subunits of glycinin were not degraded completely until 24, 48 and 48 h of incubation, respectively. It was concluded that ${\gamma}$-irradiated soybean proteins at doses higher than 15 kGy could be effectively protected from ruminal degradation.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.10
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• pp.1660-1669
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• 2018

Objective: This study was conducted to evaluate the effect of probiotics (Bacillus subtilis and Enterococcus faecium) and xylo-oligosaccharide (XOS) supplementation on growth performance, nutrient digestibility, serum profiles, intestinal health, fecal microbiota and noxious gas emission in weanling pigs. Methods: A total of 240 weanling pigs ([Yorkshire${\times}$Landrace]${\times}$Duroc) with an average body weight (BW) of $6.3{\pm}0.15kg$ were used in this 28-day trial. Pigs were randomly allocated in 1 of the following 4 dietary treatments in a $2{\times}2$ factorial arrangement with 2 levels of probiotics (0 and 500 mg/kg probiotics) and XOS (0 and 200 mg/kg XOS) based on the BW and sex. Results: Administration of probiotics or XOS improved average daily gain (p<0.05) during 0 to 14 d and the overall period, while pigs that were treated with XOS had a greater average daily gain and feed efficiency (p<0.05) compared with unsupplemented treatments throughout 15 to 28 d and the whole experiment. Either probiotics or XOS treatments increased the apparent total tract digestibility of nutrients (p<0.05) during 0 to 14 d. No effects on serum profiles were observed among treatments. The XOS increased villus height: crypt depth ratio in jejunum (p<0.05). The supplementation of probiotics (500 mg/kg) or XOS (200 mg/kg) alone improved the apparent total tract digestibility of dry matter, nitrogen and gross energy on d 14, the activity of trypsin and decreased fecal NH3 concentration (p<0.05). Administration of XOS decreased fecal Escherichia coli counts (p<0.05), while increased lactobacilli (p<0.05) on d 14. There was no interaction between dietary supplementation of probiotics and XOS. Conclusion: Inclusion of XOS at 200 mg/kg or probiotics (Bacillus subtilis and Enterococcus faecium) at 500 mg/kg in diets containing no antibiotics significantly improved the growth performance of weanling pigs. Once XOS is supplemented, further providing of probiotics is not needed since it exerts little additional effects.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2008.04a
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• pp.5-20
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• 2008

GLP-1-based drugs (GLP-1 analogues and DPP IV inhibitors) and incretin mimetics are currently one of the most exciting classes of agents for type II diabetes. GLP-1, a gut peptide, is an incretin that potentiates glucose-dependent insulin release from the pancreas, slows GI-transit and stimulates the proliferation of beta-cells. DPP IV inhibitors act like incretins by inhibiting DPP IV which inactivates GLP-1. LC15-0133 is a competitive, reversible DPP IV inhibitor ($IC_{50}$ = 24 nM, Ki=0.247 nM) with excellent selectivity over other critical human proteases such as DPP II, DPP 8, elastase, trypsin. and urokinase. LC15-0133 showed long half-life and good bioavailability in rats and dogs. Inhibition of plasma DPP IV activity by LC15-0133 was kept more than 50% 24 hours after oral dosing in rats and dogs at 0.1 mg/kg and 0.02 mg/kg, respectively. The Minimum effective doses of LC15-0133 were 0.01 mg/kg for lowering blood glucose excursion during oral glucose tolerance test and 0.1 mg/kg for increasing glucose-induced GLP-1 response in C57BL/6 mice. Repeat oral administration of LC15-0133 for 1 month delayed the progression to diabetes and reduced HbA1c levels in a dose-dependent manner in Zucker Diabetic Fatty rats. In conclusion, LC15-0133 is a novel, potent, selective and orally active DPP IV inhibitor and showed an excellent blood glucose lowering effects in various animal models.

• Journal of Periodontal and Implant Science
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• v.32 no.4
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• pp.865-873
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• 2002

The primary cause of tooth loss after 30 years of age is periodontal disease. Destruction of alveolar bone by periodontal disease is done by bone resorbing activity of osteoclasts. Understanding differentiation and activation mechanism of osteoclasts is essential for controling periodontal disease. The purpose of this study is to identify the possible effects of Vitamin D and cytokines affecting osteoclasts and its precursor cells. Four to six week-old mice were killed and humerus, radius, tibia and femur were removed aseptically and washed two times with Hank's solution containing penicillin-streptomycin and then soft tissue were removed. Bone marrow cells were collected by 22 gauge needle. Cells were cultured in Hank's solution containing 1 mg/ml type II collagenase, 0.05% trypsin, 41mM EDTA. Supernatant solution was removed 5 times after 15 minutes of digestion with above mentioned enzyme solution, and remained bone particles were maintained in alpha-MEM for 15 minutes and $4^{\circ}C$ temperature. Bone particles were agitated for 1 minute and supernatant solution containing osteoclast precursor cells were filtrated with cell stainer. These separated osteoclast precursor cells were dispensed with 100-mm culture dish by $1{\times}10^7$ cells unit and cultured in ${\alpha}$- MEM containing 20 ng/ml recombinant human M-CSF, 30 ng/ml recombinant human soluble osteoclast differentiation factor and 10% fetal calf serum for 2 and 7 days. Total RNA of osteoclast precursor cells were extracted using RNeasy kit. One ${\mu}g$ of total RNA was reverse transcribed in $42^{\circ}C$ for 30 minutes using SuperScriptII reverse transcriptase. Expression of transcribed receptors of each hormone and cytokine were traced with 1 ${\mu}l$ of cDNA solution by PCR amplification. Vitamin D receptor WAS found in cells cultured for 7 days. TNF-${\alpha}$ receptor was found in cells cultured for 2 days and amount of receptors were increased by 7 days. IL-1 type I receptor was not found in cells cultured 2 and 7 days. But, IL-1 receptor type II was found in cells cultured for 2 days. TGF-${\alpha},{\beta}$type I receptor was found in cells cultured 2 and 7 days, and amount of receptors were increased by 7 days of culture. These results implies Vitamin D and cytokines can affect osteoclasts directly, and affecting period in differentiation cycle of osteoclasts is different by Vitamin D and cytokines.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.41 no.3
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• pp.243-252
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• 2015

Melanosome, the pigment granule in melanocyte, determines the color of skin when it moves into the keratinocyte. Inhibition of melanosome transfer from melanocyte to keratinocyte results in skin depigmentation. Protease activated receptor-2 (PAR-2) is involved in signal transduction systems via cell membrane and increases the melasome transfer when it is activated by cleavage of their extracellular amino acid sequence by trypsin or by a peptide such as SLIGKV. Here, we showed that lobaric acid inhibited PAR-2 activation and affected the mobilization of $Ca2^+$. The uptake of fluorescent microspheres and isolated melanosomes from melan-a melanocytes to keratinocytes induced by SLIGKV were inhibited by lobaric acid. Also, confocal microscopy studies illustrated a decreased melanosome transfer to keratinocytes in melanocyte-keratinocyte co-culture system by lobaric acid. In addition, lobaric acid induced visible skin lightening effect in human skin tissue culture model, melanoderm$^{(R)}$. Our data suggest that lobaric acid could be an effective skin lightening agent that works via regulation of phagocytic activity of keratinocytes.

• Journal of Dairy Science and Biotechnology
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• v.17 no.1
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• pp.1-10
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• 1999

A bacteriocin produced by Lab. acidophilus GP4A isolated from fecal contents of pig was characterized. Lab. acidophilus GP4A produced a heat-stable and pH-resistant bacteriocin, which was hydrolyzed by trypsin and pepsin and active against various microorganisms. Lab. acidophilus GP4A produced bacteriocin at maximum rate when grown in MRS broth(pH 6.5${\sim}$7.5) at$37^{\cric}C$ or $40^{\cric}C$. The bacteriocin produced by Lab. acidophilus GP4A inhibited the growth of Lactobacillus delbrueckii subsp. lactis 4794 in early logarithmic phase. The bacteriocin was purified by ammonium sulfate precipitation and Octyl sepharose CL-4B column chromatography. The purification resulted in a final yield of 21.7% and a 13.6-fold increase in the specific activity.

• Korean Journal of Plant Tissue Culture
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• v.24 no.5
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• pp.305-311
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• 1997

Bacillus thuringiensis, a gram-positive soil bacterium, is characterized by its ability to produce crystalline inclusions during sporulation. The crystal proteins exhibit a highly specific insecticidal activity. An insecticidal crystal protein (ICP), Cry II A, is specifically toxic to both lepidopteran and dipteran insects. In this study, tobacco plants transformed by the cry II A gene have been generated. The Cry II A crystal protein was purified from E. coli JM103 harboring cry II A gene by differential solubility. The activated Cry II A was prepared by tryptic digestion. The purified protoxin (70 kDa) and the activated toxin (50 kDa) were analyzed by SDS-PAGE. To generate the transgenic tobacco having cry II A gene, the cry II A gene was subcloned to a plant expression vector, pSRL2, having two CaMV 35S promoters. The recombinant plasmid was transformed into tobacco (N. tabacum var. Petit Havana SR1) by Agrobacterium-mediated leaf disc transformation. Through the regeneration, six putative transgenic tobacco plants were obtained and three transformants were confirmed by Southern blot analysis. It has been found that one plant had single copy of cry II A gene, another had two copies of the gene, and the third had a truncated gene. After the immunochemical confirmation of cry II A expression in plants, the transgenic tobacco plants will be used to study the genetics of future generation with the insecticidal crystal protein gene cry II A.

• Korean Journal of Environmental Biology
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• v.36 no.3
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• pp.377-384
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• 2018

This study focused on the effects of feeding on postlarvae of shrimp, Litopenaeus vannamei, during the identified acclimation time to low salinity. A total of 5 different salinity groups with or without feeding (32, 24, 16, 8, and 2 psu, 1 liter, triplicates) were prepared, and 30 shrimp were settled at PL21 (postlarvae) and placed in each group. After 24 hours of the experimentation process, the survival rate of the fed and starved groups was observed to be lower in the 2 psu group compared to other salinity groups, with the rate of 86.6% and 81.1%, respectively. The condition index of glucose and triglyceride, which are important factors for osmoregulation and as energy sources, was 4.2-7.6 times and 2.7-3.4 times higher in the fed groups than the starved groups at all the levels of salinities. The creatine level increased by 1.1-1.5 times in the starved groups as compared to the fed groups. Likewise, the activity of all the digestive enzymes like, lipase, ${\alpha}$-amylase, trypsin, and alkaline protease were clearly higher in the fed groups (ANOVA, p<0.05). Apparently, it was observed that feeding is effective for the postlarvae of shrimp, which shows a characteristic fast metabolism and larval development, during the acclimation period to low salinity.