• Journal of Korean Medicine Rehabilitation
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• v.25 no.2
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• pp.15-35
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• 2015

Objectives This study was performed to investigate the effects of Bujasasim-tang ethanol extract (BST) on oxidative stress, inflammation and osteoarthritic rat model. Methods To ensure safety of BST, heavy metal levels were measured and cytotoxicity test was done. In vitro, To evaluate antioxidative effects of BST, total phenolic contents, 1,1-diphenyl-2-picryl-hydrazyl (DPPH), 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) scavenging activity, reactive oxygen species (ROS) levels were measured. Also, to evaluate anti-inflammatory effects of BST treated group, total nitric oxide (NO) and pro-inflammatory cytokines (IL-$1{\beta}$, IL-6, TNF-${\alpha}$) levels were measured in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. In vivo, We injected MIA $50{\mu}l$ (60 mg/ml) into knee joints of rats to induce osteoarthritis. Rats were divided into total 3 groups (normal, control, BST treated group, each n=7). Normal group was not treated at all without inducing osteoarthritis and taken normal diet. Control group was induced osteoarthritis by MIA and taken with 2 ml of distilled water once a day for 4 weeks. BST treated group was induced osteoarthritis by MIA and taken BST 2 ml (200 mg/kg/mouse) once a day for 4 weeks. We evaluated dynamic weight bearing with the Incapacitance Test Meter. At the end of experiment, the rats were sacrificed to observe the functions of liver and kidney, changes of WBC, neutrophil, lymphocyte, monocyte levels in blood, to evaluate the levels of pro-inflammatory cytokines, tissue inhibitor of metallopreteinases-1 (TIMP-1), matrix metalloproteinase-9 (MMP-9), prostaglandin $E_2$ ($PGE_2$), leukotriene $B_4$ ($LTB_4$) within serum. We observed change of articular structures by Hematoxylin & Eosin (H&E), safranin-O staining method and measured amount of cartilage by micro CT-arthrography. Statistical analysis was done by unpaired student's t-test with significance level at p<0.05 in SPSS 11.0 for windows. Results 1. Safety of the BST was identified. 2. AST, ALT, BUN, creatinine levels of BST treated group were within normal limit. In vitro, 1. DPPH and ABTS free radical scavenging activities of BST showed dose-dependent increase. 2. ROS production were significantly decreased. 3. Total nitric oxide (NO) and IL-$1{\beta}$ production were decreased. 4. IL-6 and TNF-${\alpha}$ production were significantly decreased. In vivo, 1. Weight bearing ability was significantly increased. 2. WBC, neutrophil, lymphocyte, monocyte levels in blood were decreased. 3. IL-$1{\beta}$ and TNF-${\alpha}$ levels in serum were significantly decreased. and the IL-6 level was decreased. 4. TIMP-1, MMP-9, $LTB_4$, $PGE_2$ levels in serum were significantly decreased. 5. Cartilage volume of BST treated group was significantly increased. Also changes of cartilage, synovial membrane, fibrous tissue were suppressed. Conclusions The results obtained in this study Bujasasim-tang have effects of antioxidative, anti-inflammatory, relieve pain and protection of cartilage. Therefore we expect that Bujasasim-tang is effective treatment for osteoarthritis.

• Journal of Animal Science and Technology
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• v.46 no.4
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• pp.537-546
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• 2004

Adiponectin is adipocyte complement-related protein which is highly specialized to play important roles in metabolic and honnonal processes. This protein, called GBP-28, AdipoQ, and Acrp30, is encoded by the adipose most abundant gene transcript 1 (APM1) which locates on human chromosome 3q27 and mouse chromosome 16. In order to determine chromosomal localization of the porcine APM1, we carried out PCR analysis using somatic cell hybrid panel as well as porcine whole genome radiation hybrid (RH) panel. The result showed that the porcine APM1 located on chromosome 13q41 or 13q46-49. These locations were further investigated with the two point analysis of RH panel, revealed the most significant linked marker (LOD score 20.29) being SIAT1 (8 cRs away), where the fat-related QTL located. From the SSCP analysis of APM1 using 8 pig breeds, two distinct SSCP types were detected from K~ native and Korean wild pigs. The determined sequences in Korean native and Korean wild pigs showed that two nucleotide positions (T672C and C705G) were substituted. The primary sequence of the porcine APM1 has 79 to 87% identity with those of human, mouse, and bovine APM1. The domain structures of the porcine APM1 such as signal sequence, hypervariable region, collagenous region. and globular domain are also similar to those of mammalian genes.

• Korean Journal of Veterinary Research
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• v.33 no.4
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• pp.623-632
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• 1993

This study was conducted to develop a sensitized latex-antigen for serodiagnosis of toxoplasmosis in animals. Tachyzoites of T gondii(RH-strain) harvested from mouse peritoneal cavity were purified through the filtraton of polycarbonate membrane(pore size, $3.0{{\mu}m}$, Costar Co.) and disrupted by ultrasonicator. The tachyzoite suspension was ultracentrifuged for 30 min at $60,000{\times}g(4{^{\circ}C})$ and the supernatant was used as a water-lysate antigen. Polystyrene latex particles of $0.8{{\mu}m}$ in diameter(Sigma) were used for the preparation of sensitized latex-antigen suspension. The several parameters including the preparation conditions, incubation buffer. serum dilution buffer and stability of agglutination reactions were evaluated and the results obtained were summarized as follows : 1. The antigen consisting of a water-lysate of T gondii tachyzoites was adsorbed onto polystyrene latex particles of $0.8{{\mu}m}$ in diameter by adding a latex suspension to an equal volume of diluted antigen solution and by incubating the mixture at $37{^{\circ}C}$ under different conditions. 2. The optimum incubation buffer used for the antigen sensitization was 0.1M Tris-HCl buffer(pH 8.0). 3. The optimum serum dilution buffer used for the latex agglutination test was 0.1M Tris-HCl-NaCl buffer(pH 7.4) containing 300 mM NaCl. But 0.1M Tris-HCl-NaCl buffer(pH 7.4) containing 300-600 mM NaCl, 0.5% BSA and 0.01% Tween-20 improved the agglutination pattems and cleared the background of microplate well without the effects on L.A titer. 4. The time required for antigen sensitization was 40 and 60 min in incubation buffer(pH 8.0) at $37{^{\circ}C}$. But the optimun time for antigen sensitization was min at $37{^{\circ}C}$. 5. The optimun quantity of antigen absorbed on latex particles for proper agglutination was the range of 20 to $32{\mu}g$ of latex particles. 6. The optimun concentration of the latex-antigen suspension for the proper agglutination reaction was determined as 0.2%(w/v). 7. The specificity, rapidity and simplicity of the latex-particle agglutination test suggested that it might be adaptable to large scale serum screening.

• Biomolecules & Therapeutics
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• v.25 no.2
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• pp.112-121
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• 2017

Drug-induced liver injury (DILI) is the serious and fatal drug-associated adverse effect, but its incidence is very low and individual variation in severity is substantial. Acetaminophen (APAP)-induced liver injury accounts for >50% of reported DILI cases but little is known for the cause of individual variations in the severity. Intrinsic genetic variation is considered a key element but the identity of the genes was not well-established. Here, pre-biopsy method and microarray technique was applied to uncover the key genes for APAP-induced liver injury in mice, and a cause and effect experiment employing quantitative real-time PCR was conducted to confirm the correlation between the uncovered genes and APAP-induced hepatotoxicity. We identified the innately and differentially expressed genes of mice susceptible to APAP-induced hepatotoxicity in the pre-biopsied liver tissue before APAP treatment through microarray analysis of the global gene expression profiles (Affymetrix $GeneChip^{(R)}$ Mouse Gene 1.0 ST for 28,853 genes). Expression of 16 genes including Gdap10, Lpl, Gabra3 and Ccrn4l were significantly different (t-test: FDR <10%) more than 1.5 fold in the susceptible animals than resistant. To confirm the association with the susceptibility to APAP-induced hepatotoxicity, another set of animals were measured for the expression level of selected 4 genes (higher two and lower two genes) in the liver pre-biopsy and their sensitivity to APAP-induced hepatotoxicity was evaluated by post hoc. Notably, the expressions of Gabra3 and Lpl were significantly correlated with the severity of liver injury (p<0.05) demonstrating that these genes may be linked to the susceptibility to APAP-induced hepatotoxicity.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.24 no.1
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• pp.121-141
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• 2011

Objective : This Experimental study was done to investigate the Effect of Taklisodok-um and Hwangryunhaedok-tang(TH) on Atopic Dermatitis. Methods : We assessed effects of TH on the IgE, IL-4, IL-5, IL-6, IgM, IgG1, IFN-${\gamma}$ in vivo, on the IL-4, IL-5, CCR3 in the skin tissues of ear and dorsum with NC/Nga mice. And we assessed effects of TH on the COX-2, IL-$1{\beta}$, TNF-${\alpha}$, IL-6 with RAW 264.7 cell. Results and Conclusion : 1. IgE, IL-4, IL-5, IL-6, IgM, IgGl levels in the serum of TH treated NC/Nga mouse group were decreased compared to the untreated control mice. IFN-r showed a increase in the experimental group compared to the untreated control group. The spleen weight of TH treated NC/Nga mice was decreased compared to the untreated control group. 2. mRNA expression levels of IL-4, IL-5 and CCR3 in the skin tissues of TH treated NC/Nga mice were decreased, and expression levels of IL-6 in the skin tissues of TH treated NC/Nga mice were decreased compared to the untreated control group. IFN-${\gamma}$ mRNA expression levels were increased compared to the untreated control group. 3. Judged from that IL-$1{\beta}$, TNF-${\alpha}$, IL-6 express of gene, effect of inflammatory Cytokines revelation were decreased compared to the untreated control group. 4. Depend on the strength of TH, inflammatory RAW 264.7 in the serum of TH were inhibited compared to the untreated control serum that leaded a COX-2 activity model. 5. Histological observation of the ear and skin tissues showed that the extents of inflammation and infiltrated immune cells in the epidermis and dermis of TH treated NC/Nga mice were highly reduced compared to the untreated control group.

• Korean Journal of Animal Reproduction
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• v.20 no.2
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• pp.207-214
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• 1996

To examine the developmental capacity of manipulated embryos after ultrarapid refreezing and thawing, mouse embryos were biopsied at 4-cell stage, frozen twice at 4-cell and morula stages, respectively, and then transferred to rec-ipients. Single blastomeres were biopsied from 4-cell embryos by a modified aspiration method. Biopsied 4-cell embryos were equilibrated into freezing medium at room temperature for 2.5 min, loaded into 40 $\mu$I of freezing medium in 0.25 ml plastic straw and then directly immersed into liqiud nitrogen. Freezing medium for 4-cell embryos consisted of 4.0 M ethylene glycol and O.25 M sucrose in dPBS supplemented with 6 mg/lm BSA. Morulae were frozen into freezing medium containing 5.0 M glycerol instead of ethylene glycol. Thawing was conducted by agitating each straw in 3TC water for 20 sec. The c content of each straw was expelled into 0.5 ml of dilution medium, which consisted of 0.25 M sucrose and 3 mg/ml BSA in dPBS. The thawed embryos were rehydrated in dilution medium for 10 min, washed 3 times with dPBS and then cultured in M16 medium at 37$^{\circ}C$, 5% CO$_2$ in air. Blastocysts that developed from frozen or refrozne biopsied embryos were transferred to recipients on Day 3 of pseudopregnancy, respectively. In vitro and in vivo developmental rates of the biopsied and intact 4~cell embryos after freezing and thawing were 78 (10l/130) and 25% (10/40), and 91 (114/125) and 30% (12/40), respectively. Although the rates of in vitro development of biopsied and intact embryos to blastocyst stage were significantly different after freezing and thawing (P

• Parasites, Hosts and Diseases
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• v.28 no.3
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• pp.143-154
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• 1990

Observations were made on the differences of cell-mediated responses in mice of three infectiorl groups di여erently scheduled in their severity with pathogenic Acanthamoeba culbertseni. Infections were done by dropping $5{\;}{\mu}l$ saline suspension containing $3{\times}10^3,{\;}1{\times}10^4,{\;}or{\;}1{\times}10^5$ trophosoites, respectively. Amoebae were cultured anenically in CGV medium and inoculated into the right nasal cavity of CSH/HeJ mice aging around 6∼8 weeks, under the anesthesia by intraperitoneal injection of secobarbital. Delayed type hypersensitivity (DTH) responses in footpad and blastogenlc responses of mouse spleen cells using ($^3H$)-thymidine and the serum antibody titer were measured up to day 14 after infection, and natural killer cell activities were measured up to day, i after infection. The results obtained in this study were as follows: 1. The mice infected with $3{\times}10^3$ trophosoites showed mortality rate of 17%, and 345 in the mice infected with $1{\times}10^4$ trophozoites and 65% with $1{\times}10^5$ trophozoites. 2. In regard to DTH responses in all experimental groups, the level increased on day 7 and declined on day 14 after infection, but their differences could not be noted between infected and control groups. 3. The blastogenic responses of splenocytes treated with amoeba Iysates and lipopolysaccharides (LPS) showed no difference from the control group. The blastogenic responses of splenocytes treated with concanavalin A were declined significantly in the experimental group as compared with the control group, but the blastogenic responses of splenocytes treated with polyinosinic acid were not different from the control group. There was also no difference among three infected groups. 4. The cytotoxic activity of the natural killer cells was activated on day 1 after infection and declined to the level of control group on day 2 in all experimental groups. On day 5 after infection, the natural killer cell cytotoxicity was significantly suppressed as compared with the control groups. 5. The serum antibody titers of the infected mice increased after day 7, but there was no statistical difference between the three infected groups. In summary of the results, there was no difference in cell-mediated immune responses of three experimental groups scheduled with different infection intensities. But there was a significant difference in cell$.$mediated immune responses between infected and control mice. It is considered that cell-mediated immune responses should be involved in murine model infected with A. culbertsoni.

• Tuberculosis and Respiratory Diseases
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• v.53 no.6
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• pp.619-634
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• 2002

Background : Many inflammatory mediators and collagenases are involved in the pathogenesis of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). The increase of matrix metalloproteinase-9 (MMP-9, gelatinase-B) produced mainly by inflammatory cells was reported in many ALI models and connective tissue cells. In this study, the expression of MMP-9 in ventilator-induced lung injury (VILI) model and the effects of matrix metalloproteinase inhibitor (MMPI) on VILI were investigated. Methods : Eighteen Sprague-Dawley rats were divided into three groups: low tidal Volume (LVT, 7mL/Kg tidal volume, 3 $cmH_2O$ PEEP, 40/min), high tidal volume (HVT, 30mL/Kg tidal volume, no PEEP, 40/min) and high tidal volume with MMPI (HVT+MMPI) groups. Mechanical ventilation was performed in room air for 2 hours. The 20 mg/Kg of CMT-3 (chemically modified tetracycline-3, 6-demethyl 6-deoxy 4-dedimethylamino tetracycline) was gavaged as MMPI from three days before mechanical ventilation. The degree of lung injury was measured with wet-to-dry weight ratio and acute lung injury score. Expression of MMP-9 was studied by immunohistochemical stain with a mouse monoclonal anti-rat MMP-9 $IgG_1$. Results : In the LVT, HVT and HVT+MMPI groups, the wet-to-dry weight ratio was $4.70{\pm}0.14$, $6.82{\pm}1.28$ and $4.92{\pm}0.98$, respectively. In the HVT group, the ratio was significantly higher than other groups (p<0.05). Acute lung injury score measured by five-point scale was $3.25{\pm}1.17$, $12.83{\pm}1.17$ and $4.67{\pm}0.52$, respectively. The HVT group was significantly damaged by VILI and MMPI protects injuries by mechanical ventilation (p<0.05). Expression of MMP-9 measured by four-point scale was $3.33{\pm}2.07$, $12.17{\pm}2.79$ and $3.60{\pm}1.95$, respectively, which were significantly higher in the HVT group (p<0.05). Conclusion : VILI increases significantly the expression of MMP-9 and MMPI prevents lung injury induced by mechanical ventilation through the inhibition of MMP-9.

• Analytical Science and Technology
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• v.28 no.5
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• pp.331-340
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• 2015

Mutations in Fused in Sarcoma (FUS) have been identified in patients with amyotrophic lateral sclerosis (ALS) or Frontotemporal Dementia (FTD). Pathological FUS is mis-localized to cytosol and forms aggregates associated with stress granules (SG), while FUS is normally localized to nucleus. However, it is largely unknown how pathological FUS forms SG-aggregates and which domains are responsible for this process. In this study, we examined cellular localization and aggregation of ALS-linked FUS missense mutants (P525L, R521C, R521H, R521G), analyzed the domains responsible for cytosolic FUS aggregation in HEK293T cells, and confirmed this in cultured mouse neurons. To do this, we firstly generated missense mutants of FUS and then examined their cellular localization. We found that P525L was mostly mis-localized to cytosol and formed FUS-positive SG aggregates while R521C, R521H, or R521G was localized to both nucleus and cytosol. To further characterize the domains required for aggregate formation of cytosolic FUS, we generated different domain-deletion mutants using FUS-∆17 which has a deletion of nuclear localization signal. Interestingly, cytosolic FUS without SYGQ and RGG1 domain or cytosolic FUS without RGG2-ZnF-RGG3 domain did not form FUS-positive SG aggregates, while cytosolic FUS without RRM domain generated more aggregates compared to FUS-∆17. Taken together, these data suggest that SYGQ-RGG1 or RGG2-ZnF-RGG3 domain contributes to formation of cytosolic aggregate, while RRM domain might interfere with FUS aggregation. Therefore, our studies will provide important insight for understanding cellular pathogenesis of neurodegeneration associated with FUS aggregate as well as finding therapeutic targets for ALS or FTD.

• Nutrition Research and Practice
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• v.5 no.3
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• pp.198-204
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• 2011

Target herbal ingredient (THI) is an extract made from two herbs, Scutellariae Radix and Platycodi Radix. It has been developed as a treatment for metabolic diseases such as hyperlipidemia, atherosclerosis, and hypertension. One component of these two herbs has been reported to have anti-inflammatory, anti-hyperlipidemic, and anti-obesity activities. However, there have been no reports about the effects of the mixed extract of these two herbs on metabolic diseases. In this study, we investigated the metabolic effects of THI using a diet-induced obesity (DIO) mouse model. High-fat diet (HFD) mice were orally administered daily with 250 mg/kg of THI. After 10 weeks of treatment, the THI-administered HFD mice showed reduction of body weights and epididymal white adipose tissue weights as well as improved glucose tolerance. In addition, the level of total cholesterol in the serum was markedly reduced. To elucidate the molecular mechanism of the metabolic effects of THI in vitro, 3T3-L1 cells were treated with THI, after which the mRNA levels of adipogenic transcription factors, including C/$EBP{\alpha}$ and $PPAR{\gamma}$, were measured. The results show that the expression of these two transcription factors was down regulated by THI in a dose-dependent manner. We also examined the combinatorial effects of THI and swimming exercise on metabolic status. THI administration simultaneously accompanied by swimming exercise had a synergistic effect on serum cholesterol levels. These findings suggest that THI could be developed as a supplement for improving metabolic status.