• Clinical and Experimental Reproductive Medicine
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• v.27 no.4
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• pp.345-351
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• 2000

Objective: Hatching of the blastocyst from the zona pellucida (ZP) is a key event in mammalian implantation. In vivo, two factors have been identified as possible mediators of hatching: lysis of the ZP by substances elaborated either from the embryo or female reproductive tract and pressure exerted on the zona by expansion of the blastocyst. Two methods of zona manipulation were already in use to enhance the ability of embryos to hatch: mechanical PZD and chemical ZD by acidic Tyrode's solution. But several controversies of each method have been reported. The purpose of this study was to investigate the effect of pronase for mouse embryo hatching. Methods: Mouse embryos were obtained following ovulation induction of $F_1$ animals. Fresh and cryo-thawed morula embryos were exposed to 0.5, 1.0, 2.0, 5.0 ${\mu}g/ml$ pronase in Ham's F10 for 72 hrs. Main outcome measures were the rates of partial hatching and completely hatched blastocysts, and cell number of it. Results: In fresh and cryo-thawed group, the rates of completely hatched blastocyst were significantly higher in 5 ${\mu}g/ml$ pronase treatment group than control group. There was no difference in completely hatched blastocyst total cell number between pronase treatment group and control group. This suggest that pronase treatment did not harmful in mouse embryo development. In pronase treatment group, zona pellucida were thinner than control group. Conclusion: The addition of pronase to culture media may accelerate the hatching of embryo. So, enzymatic treatment of the zona may provide a valuable and effective assisted hatching technique for human in-vitro fertilization-embryo transfer.

• Microbiology and Biotechnology Letters
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• v.28 no.2
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• pp.117-123
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• 2000

The object of this work is to improve the maintenance of bioluminescence from stored Photobacterium phosphoreum in a view of developing continuous monitoring system for pollutants. The long-term experiments were performed to determine the effect of storage temperature and immobilization on the maintenance of bioluminescence and viability of P. phosphoreum. A naturally luminescent bacterium, P. phosphoreum was starved in 2.5% Nael solution at $20^{\circ}C$, $4^{\circ}C$, -$20^{\circ}C$ and -$70^{\circ}C$ for 30 days. In vivo luminescence was measured by luminometry, and total cell concentrations and concentrations of culturable and viable cells were determined by acridine orange staining, dilution plate counting, and direct viable counting, respectively. The bioluminescence emission from cells stored at 4De was maintained up to 10 days while those with starved cells at other temperature ranges decreased to background level within 3 days. In terms of viability of cells, concentrations of cells stored at $20^{\circ}C$ were rapidly decreased as a result of cell lysis, leading to a drop in culturable and viable counts while cells stored at $4^{\circ}C$ was shown viable but nonculturable state during starvation. With immobilized cells on strontium alginate, the bioluminescence showed higher maintenance than free cells and decreased with count number of nonculturable cells.

• Journal of the korean academy of Pediatric Dentistry
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• v.30 no.1
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• pp.80-91
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• 2003

Mutans streptococci, especially S. mutans and S. sobrinus strongly implicated in pathogenesis of dental caries, the major cause of tooth loss in children. Use of an antibacterial agent controlling dental caries has been rationalized. The present study was performed to observe the antibacterial effect of inorganic polyphosphates (polyP) on S. mutans and S. sobrinus. S. mutans GS5 and S. sobrinus 6715 were grown in brain-heart infusion broth with or without polyP. Minimal inhibitory concentration (MIC) of polyP for S. mutans GS5 was determined to be 0.08% and that for S. sobrius 6715 was 0.17%. PolyP 15 added to the growing culture of S. mutans GS5 and S. sobrinus 6715 at their exponential phase was as effective in inhibiting the growth of S. mutans GS5 and S. sobrinus 6715 as polyP added at the very beginning of the culture. More than 85% of the cells lost their viability determined by viable cell count when polyP 15 was added to the culture of growing S. mutans GS5 at MIC, suggesting that polyP 15 has bacterial effect on the bacterium. And more than 99.9% of the cells lost their viability determined by viable cell count when polyP 15 was added to the culture of growing S. sobrinus 6715 at MIC, suggesting that polyP 15 has bacterial effect on the bacterium. Intracellular nucleotide release from S. mutans CS5 and S. sobrinus 6715 was increased in the presence of polyP 15 for 5h but was not really reversed by the addition of divalent cations like $Ca^{++}\;and\;Mg^{++}$. The majority of the cells appeared to be atypical in their shape, demonstrating accumulation of highly electron-dense granules and ghost cells. The overall results suggest that polyP have a strong bactericidal activity against S. mutans and S. sobrinus in which lysis in relation to chelation may not play the major role but unknown mechanism that possibly affects the viability of the bacterium may be involved. PolyP may be used as an agent for prevention of dental caries.

• Journal of Dental Rehabilitation and Applied Science
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• v.19 no.2
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• pp.87-96
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• 2003

The purposes of this study were to find out the optimum intensity of magnetic field where magnetism could promote the activity of osteoblast, and to discover the possibility of clinical application in the areas of dental implants and bone grafts by confirming the effect of clinically increasing bone formation. In this experiment, we used the Neodymium magnet, which had magnetic power six times as strong as the current ones and enabled the resistances against the demagnetization up to 20 to 50 times to be minimized with the size of 1mm in sight. In order to culture cells, a specially designed device was used. It was made to adjust the distance and accordingly to control the intensity of the magnetic field, by placing the cell culture plate in the center with a magnet of 1mm long and thick installed on the both ends. Using MC3T3-E1 cell, a kind of osteoblast-like cell, we cultured, for 24 hours, not only the test group which had been cultured under the magnetic fields with different intensity of 5, 10, 50, 100, 500, and 1000 Gauss, but also the control group excluding the influences of the magnetic field. After observing the cell's form and the density of the culture medium through an inverted microscope, we made a series of proceedings needed for the immunofluoroscence staining, such as fixation, normal serum reaction, primary antibody reaction, and secondary antibody reaction. And with a fluorescence microscope, we observed those-above and compared the frequency of expression of IFG-1 receptor. To make a Western immunoblotting analysis, the cells cultured under the same condition as the above had the procedure of the lysis buffer and the acrylamide gel electrophoresis was carried out. Protein transferred into the nitrocellulose membrane and tested on the primary and the secondary antibody reactions was observed and compared. The results were as follows: When observed through an inverted microscope, the nuclear divisions of the cells under the magnetic field of 10 Gauss were the most active, and the density of the cells could be observed the most enormously. As the result of an immunofluoroscence staining of IGF-1 receptor, the expression of IFG-1 was the most frequently observed under the magnetic field of 10 Gauss. On the other hand, few differences of consideration were made between the test group cultured under the magnetic fields of 5, 500, and 1000 Gauss and the control group. In respect of the expression of IFG-1 receptor, the test group cultured under the magnetic fields of 50 and 100 Gauss were higher than the control group, and lower than that cultured under the magnetic field of 10 Gauss.(p<0.05) According to the Western immunoblotting analysis, the band of IFG-1 receptor which had 85KDa of molecular weight was the darkest. Judging from the above-mentioned results, the growth factor receptor of an osteoblast cell which was an important criterion for the bone formation was increased in maximum under the magnetic field of 10 Gauss. Moreover it was observed that the optimum intensity of magnetic field in which magnetism made the activity of the osteoblast cell increase was about 10 Gauss.

• Journal of Pharmacopuncture
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• v.11 no.1
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• pp.71-82
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• 2008

Objectives : The purpose of this study is to investigate the effects of Spirodelae Herba pharmacopuncture(SHP) on the adipogenesis in 3T3 cells, lipolysis in rat epididymal adipocytes and histological changes in porcine adipose tissue. Methods : Inhibition of preadipocyte differentiation and/or stimulation of lipolysis play important roles in reducing obesity. 3T3 preadipocytes were differentiated with adipogenic reagents by incubating for 3 days in the absence or presence of SHP ranging from 0.01 to $1.0mg/m{\ell}$. The effect of SHP on adipogenesis was examined by measuring glycerol-3-phosphate dehydrogenase(GPDH) activity and by Oil Red O staining. Mature adipocytes from rat epididymal fat pad was incubated with SHP ranging from 0.01 to $1.0mg/m{\ell}$ for 3 days. The effect of SHP on lipolysis was examined by measuring free glycerol released. Fat tissue from porcine skin was injected with SHP ranging from 0.1 to $10.0mg/m{\ell}$ to examine the effect of SHP onhistological changes under light microscopy. Results : Following results were obtained from the preadipocyte proliferation and lipolysis adipocyte and histologic investigation of fat tissue 1. SHP showed the effect of decreased preadipocyte proliferation on the high dosage($1mg/m{\ell}$). 2. SHP showed the effect of decreased the activity of glycerol-3-phosphate dehydrogenase (GPDH) on the high dosage($1mg/m{\ell}$). 3. Investigated the changes in lipolysis of differentiated adipocyte after treated SHP, we knew that these pharmacopuncture showed increasing the effect of lipolysis in all concentration significantly. 4. Investigated the histological changes in porcine fat tissue after treated SHP, we knew that these pharmacopuncture showed significant activity to the lysis of extensive cell membranes on high dosage($10.0mg/m{\ell}$). Conclusions : These results suggest that SHP efficiently induces diminishing proliferation of preadipocyte and lipolysis in adipose tissue.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.4031-4036
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• 2012

Background: The negative signaling provided by interactions of the co-inhibitory molecule, programmed death-1 (PD-1), and its ligands, B7-H1 (PD-L1) and B7-DC (PD-L2), is a critical mechanism contributing to tumor evasion; blockade of this pathway has been proven to enhance cytotoxic activity and mediate antitumor therapy. Here we evaluated the anti-tumor efficacy of AAV-mediated delivery of the extracellular domain of murine PD-1 (sPD-1) to a tumor site. Material and Methods: An rAAV vector was constructed in which the expression of sPD-1, a known negative regulator of TCR signals, is driven by human cytomegalovirus immediate early promoter (CMV-P), using a triple plasmid transfection system. Tumor-bearing mice were then treated with the AAV/sPD1 construct and expression of sPD-1 in tumor tissues was determined by semi quantitative RT-PCR, and tumor weights and cytotoxic activity of splenocytes were measured. Results: Analysis of tumor homogenates revealed sPD-1 mRNA to be significantly overexpressed in rAAV/sPD-1 treated mice as compared with control levels. Its use for local gene therapy at the inoculation site of H22 hepatoma cells could inhibit tumor growth, also enhancing lysis of tumor cells by lymphocytes stimulated specifically with an antigen. In addition, PD-1 was also found expressed on the surfaces of activated CD8+ T cells. Conclusion: This study confirmed that expression of the soluble extracellular domain of PD-1 molecule could reduce tumor microenvironment inhibitory effects on T cells and enhance cytotoxicity. This suggests that it might be a potential target for development of therapies to augment T-cell responses in patients with malignancies.

• Microbiology and Biotechnology Letters
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• v.13 no.4
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• pp.397-402
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• 1985

The fermentation of various sugars by C. thermosaccharolyticum was examined under pH controlled, anaerobic condition. The kinetic model for Product formation at various sugars was the combination of growth and non-growth associated mode. In the utilization of a single sugar, glucose was the best carbon source for growth. The specific growth rate of glucose, xylose and cellobiose were 0.363 h$^{-1}$, 0.242 h$^{-1}$ and 0.144 h$^{-1}$ respectively. The production of ethanol from glucose showed a negatively growth associated mode, so the higher growth rate decreased the productivity of ethanol. The maximum concentrations of the produced ethanol were 2.42 g/l, 3.76 g/l, and 3.4 g/l on glucose, xylose, and cellobiose. No glucose was detected during cellobiose fermentation. Sequential utilization of sugars was observed in the mixtures of glucose, xylose and cellobiose. It preferred glucose, followed by xylose and then cellobiose. The presence of other sugars had little or no effect on the rate of another sugar utilization. Cell lysis at the end of fermentation occured more slowly in the mixtures of sugars than a single sugar.

• The korean journal of orthodontics
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• v.26 no.5 s.58
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• pp.591-599
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• 1996

This study was undertaken to investigate the cytotoxicity of orthodontic wires after doing various treatments to the wires. 018x025 inch Stainless steel(A) and Co-Cr(B) wires were used and each of them were divided into 4 groups. A-1 and B-1 groups were as received state, and A-2 and B-2 groups were heat treated. A-3 and B-3 groups were electropolished after heat treatment, and A-4 and B-4 groups were soldered with Ag-solder. Each group had 3 wires and these were sterilized with Ethylene Oxide gas. We used human gingival fibroblast cell culture and agar overlay technique to investigate the cytotoxicity of each group of wires. The cytotoxicity of wire was assessed using reaction index (zone index/lysis index). The findings of this study were as follows. 1. Both of the stainless steel wire and Co-Cr wire showed no cytotoxicity in as received state. 2. Heat treatment or electropolishing of the wires had no effect on the cytotoxicity of the wires 3. Soldered stainless steel wires showed a little wider zone of discoloration than soldered Co-Cr wires, but the zone index and cytotoxicity(reaction index) was not different. 4. Soldered wires showed moderate cytotoxicity in both of the wires.

• Korean Journal of Environmental Biology
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• v.28 no.3
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• pp.172-178
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• 2010

The bloom of the genus Stephanodiscus was gradually extinguished after 18 April. Counts of bacterial population were increased as the diatom bloom was disappeared. Numbers of the heterotrophic nanoflagellates and ciliates were also increased during the disappearance of the bloom. The densities of the mesozooplankton, the major predator of the diatoms, started to increase in April. However, their growth was suppressed during the bloom period of the diatoms (from January to March). During the bloom period of the diatoms, the monthly average value of the basic productivity amounted up to 11,765.7 mgC $m^{-2}day^{-1}$, which is relatively high value considering the low temperature and light during that period. The growth rate of phytoplankton in March, when the bloom was beginning to be supressed was 0.007. The growth rate of phytoplankton was negative value in April when the decreasing of the algal density was started.

• Korean Journal of Food Science and Technology
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• v.22 no.6
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• pp.611-617
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• 1990

This study was conducted to breed a high vitamin $B_{12}$ producer by the fusion of protoplasts between Bacillus natto and Bacillus megaterium. Auxotrophic mutants of Bacillus natto SH-34 ($thr^-try^-rif^r$) and Bacillus megaterium BK-13 ($arg^-ade^-lys^-str^r$) which showed high protease activity and production of vitamin $B_{12}$, respectively, were isolated for the fusion experiment. Protoplasts were induced by incubating the cells with lysis solution containing $500{\mu}/ml$ lysozyme, and the ratio of protoplast and regeneration formation were ranged from 99% and 67%, respectively. Fusion frequencies of fusants between Bacillus natto SH-34 and Bacillus megaterium BK-13 were appeared in the ranges of $1.0{\times}10^{-5}$ under the treatment of 30% PEG 6000 containing 3% PVP. The fusant, MNF-72 showed the highest product yield of $7.85{\mu}g/g-cell\;vitamin\;B_{12}$ in production medium. For the improvement of productivity, the immobilization of fusants with sodium alginate was carried out. In batch and continuous fermentation systems, the productivity were determined to be $0.58{\mu}g/ml.hr\;and\;0.80{\mu}g/ml.hr\;vitamin\;B_{12}$ under optimum condition, respectivity.