• Journal of Microbiology and Biotechnology
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• v.18 no.5
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• pp.852-858
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• 2008

An extracellular protease (Mc1) was isolated from the nematode-trapping fungus Monacrosporium cystosporium by gel filtration, anion-exchange, and hydrophobic interaction chromatographies. This protease had a molecular mass of approximately 38 kDa and displayed an optimal activity at pH 7-9 and $56^{\circ}C$ (over 30 min). Its proteolytic activity was highly sensitive to the serine protease inhibitor PMSF (phenylmethylsulfonylfluoride, 0.1 mM), indicating that it belonged to the serine-type peptidase group. The Michaelis constant ($K_m$) and $V_max$ for substrate N-Suc-Ala-Ala-Pro-Phe-pNA were $1.67{\times}10^{-4}\;M$ and 0.6071 $OD_{410}$ per 30 s, respectively. This protease could degrade a broad range of substrates including casein, gelatin, BSA (bovine serum albumin), and nematode cuticle. Moreover, the enzyme could immobilize the free-living nematode Panagrellus redivivus and the pine wood nematode Bursaphelenchus xylophilus, suggesting that it might playa role in infection against nematodes. The encoding gene of Mc1 was composed of one intron and two exons, coding for a polypeptide of 405 amino acid residues. The deduced amino acid sequence of Mcl showed 61.4-91.9% identity to serine proteases from other nematode-trapping fungi. Our results identified that Mcl possessed biochemical properties including optimal reaction condition and substrate preference that are different from previously identified serine proteases.

• Animal Bioscience
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• v.36 no.8
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• pp.1167-1179
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• 2023

Objective: Matrix metalloproteinases (MMPs) are a family of endoproteases produced by various tissues and cells and play important roles in angiogenesis, tissue repair, immune response, and endometrial remodeling. However, the expression and function of MMPs in the pig endometrium during the estrous cycle and pregnancy have not been fully elucidated. Thus, we determined the expression, localization, and regulation of MMP2, MMP8, MMP9, MMP12, and MMP13 in the endometrium throughout the estrous cycle and at the maternal-conceptus interface during pregnancy in pigs. Methods: Endometrial tissues during the estrous cycle and pregnancy and conceptus and chorioallantoic tissues during pregnancy were obtained and the expression of MMPs was analyzed. The effects of steroid hormones and cytokines on the expression of MMPs were determined in endometrial explant cultures. Results: Expression levels of MMP12 and MMP13 changed during the estrous cycle, while expression of MMP2, MMP9, MMP12, and MMP13 changed during pregnancy. Expression of MMP2, MMP8, and MMP13 mRNAs was cell type-specific at the maternal-conceptus interface. Gelatin zymography showed that enzymatically active MMP2 was present in endometrial tissues. In endometrial explant cultures, estradiol-17β induced the expression of MMP8 and MMP12, progesterone decreased the expression of MMP12, interleukin-1β increased the expression of MMP2, MMP8, MMP9, and MMP13, and interferon-γ increased the expression of MMP2. Conclusion: These results suggest that MMPs expressed in response to steroids and cytokines play an important role in the establishment and maintenance of pregnancy by regulating endometrial remodeling and processing bioactive molecules in pigs.

• Korean Journal of Food Science and Technology
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• v.43 no.2
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• pp.213-219
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• 2011

The metastatic effect of Eupatorium japonicum extract (EJE) on MDA-MB-231 human breast cancer cells was investigated. MDA-MB-231 cells were treated with various concentrations of EJE (0, 5, 10 and $20{\mu}g/mL$). EJE inhibited cell migration, invasion and adhesion of MDA-MB-231 cells in dose-dependent manners. Gelatin zymography exhibited that EJE significantly down regulated secretion of matrix metalloproteinase (MMP)-9 and MMP-2. EJE decreased the protein levels of tissue inhibitor of metalloproteinase (TIMP)-1 but increased TIMP-2 levels. Additionally, EJE reduced the protein and mRNA levels of urokinase-type plasminogen activator (uPA), vascular endothelial growth factor (VEGF) and intercellular adhesion molecule (ICAM). In several solvent fractions of EJE, the hexane fraction markedly decreased MDAMB-231 cell migration. Thus, these finding suggest that EJE may be a potential antimetastatic agent, which can considerably inhibit the metastatic and invasive capacity of breast cancer cells.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.12 no.1
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• pp.19-26
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• 1979

To evaluate the sanitary conditions of four species of commercial shellfish (Crassostrea gigas, Tapes japonica, Mytilus coruscus and Anadara granosa), the psychrotrophic coliforms and the bacterial flora were monthly examined from January to December, 1978. Viable cell counts of the samples submitted were $1.3\times10^3\~2.5\times10^6/g$ and the numbers of coliform and E. coli were above $0\~5.5\times10^2/g\;and\;2\~9.2\times10^2/100g$ respectively. Among the 373 coliform bacteria isolated from the samples submitted, 298 strains were identified as coliform by IMViC test, $44^{\circ}C$ culture and gelatin liquefaction test. E. coli type I and K. aerogenes type I were the most predominant ones among the identified coliform as 138 strains and 71 strains respectively. The psychrotrophic coliform growing at $5^{\circ}C$ were 186 $strains(50\%)$ among 373 strains of isolated coliform. Vibrio and Pseudomonas were the most predominant groups. Among 453 strains of isolated bacteria, Vibrio was 240 $strains(53\%)$ and Pseudomonas was 91 $strains(20\%)$. Moraxella, Acinetobacter, Enterobacteriaceae and Flavobacterium-Cytophage were scantly detected. In seasonal variation of general bacterial flora, a lot of Vibrio appeared from May to October, did Moraxella during the winter season and the other genuses showed no seasonal change.

• Journal of Periodontal and Implant Science
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• v.33 no.2
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• pp.277-288
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• 2003

It has been suggested that increased number and activity of phagocytes in periodontitis lesion results in a high degree of reactive oxygen species (ROS) such as superoxide anion, hydrogen peroxide, nitric oxide and peroxynitrite. There are few reports on the relationship between ROS and MMPs expressions in gingival fibroblast. We studied to elucidate whether and how ROS, especially nitric oxide affects the MMP expression. Human gingival fibroblasts and HTl080 cells (human fibrosarcoma sell line as reference) were grown in DMEM supplemented with 10 mM HEPES, 50 mg/L gentamicin, and 10% heat inactivated fetal bovine serum with addition of various reactive oxygen species (ROS). Culture media conditioned by cells were examined by gelatin zymography. HT1080 cells expressed proMMP-2 and proMMP-9, but human gingival fibroblasts (HGF) produced only proMMP-2. Hydrogen peroxide upregulated MMP-9 expression in HT1080 cells, whereas in human gingival fibroblast SNP treatment showed marked increase in MMP-2 level compared to other ROS. These results suggest that the effects of ROS on MMPs expressions are cell-type specific. RT-PCR for MMP-2 and TIMP-2 m-RNA were performed using total RNA from cultured cells under the influence various kinase inhibitors. In HT1080 cells, treatment with FPTI III (Ras processing inhibitor) and LY294002 (PI3-kinase inhibitor) resulted in inhibition of MMP-2 and MMP-9 expressions, suggesting that Ras/P13-kinase pathway is important for MMPs expression in HT1080 cells. In gingival fibroblasts, treatment with FPTI III and PDTC (NF-kB inhibitor) showed marked decrease in MMP-2 regardless of the of SNP , suggesting that Ras/NF-kB could be the key pathway for NO-induced MMP-2 expression in gingival fibroblasts. This study showed that ROS, especially nitric oxide, could be the critical mediator of periodontal disease progression through control of MMP-2 expression in gingival fibroblasts possibly via Ras/NF-kB pathway.

• Journal of the Korean Crystal Growth and Crystal Technology
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• v.1 no.2
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• pp.79-93
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• 1991

With the advent of ULSI, many problems in previous metallization techniques and interconnection materials have become more serious. In this work, selective deposition of copper to fill the submicron contact has been tried. After forming electro-deposited copper films on p-type (100) silicon wafer using 0.75M $CuSO_4{\cdot}$5H_2O$ as an electrolyte, the effect of deposition time, current density and concentration of an additive on film properties were investigated. Film thickness, particle size and resistivity were analyzed by Alpha Step, SEM and 4 - point probe measurement respectively. The deposition rate was about $0.5-0.6\mu\textrm{m}$/min at $2A/dm^2$ and the particle size increased with increasing current density. The resistivities of electro-deposited copper films were about $3-6{\mu}{\Omega}{\cdot}$cm for the particle size above $4000{\AA}$. By the addition of 0.2 g/l gelatin, the particle size was reduced to less than $0.1{\mu}m $ and selective plugging of copper on submicron contacts could be successfully achieved.

• Korean Journal of Microbiology
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• v.30 no.1
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• pp.15-29
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• 1992

Vibrio vuln$cus has been recognized as a pathogen of septicemia and wound infection, when the organism attacks high-risk persons with a history of hepatic disease. alcohol abuse. diabetes or any debilitative disease. Forty six strains of K vulnzjicus. isolated from 1025 marine specimens from May to Novemver for three years. from 1985 to 1987. were studied for their biochemical properties. growth requirements, serotype and drug susceptibilities. The isolates were different in their various biochemical reactions. Ninety-five percent of isolated strains were able to ferment lactose, while most strains didn't utilize sucrose in their biochemical test, for example ornithine, gelatin and mannitol were quite dit'ferent composition than those described in other reports. It was found that the biochemical test wasn't useful for identifying strain. The type of somatic 0 antiserum was determined in isolates from marine sources and in patients with Vibrio septicemia. In patient isolates. 1-2 group were 24% and 1-4 group were 42%. However. 02 group(33%) were more abundant in isolates from marine sources. Minimal inhibitory concentrations(M1Cs) of chloramphenicol, tetracycline. erythromycin and ampicillin were determinef for V vuln~ficus by broth dilution method. MIC90 was I , 0.25, :! and 4,ug/ml in patient isolates. 1, 0.25, 2 and 2 ,ug/ml in marine isolates. The divalent chelating agent, IDTA. inhibited the growth of V. vuln!'ficus at 6.25 mMlml of MIC90.

• Safety and Health at Work
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• v.1 no.2
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• pp.183-191
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• 2010

Objectives: The objective of this study is to investigate the distribution patterns and exposure concentrations of bioaerosols in industries suspected to have high levels of bioaerosol exposure. Methods: We selected 11 plants including 3 livestock feed plants (LF industry), 3 metal working fluids handling plants (MWFs industry), and 5 sawmills and measured total airborne bacteria, fungi, endotoxins, as well as dust. Airborne bacteria and fungi were measured with one stage impactor, six stage cascade impactor, and gelatin filters. Endotoxins were measured with polycarbonate filters. Results: The geometric means (GM) of the airborne concentrations of bacteria, fungi, and endotoxins were 1,864, $2,252\;CFU/m^3$, and $31.5\;EU/m^3$, respectively at the sawmills, followed by the LF industry (535, $585\;CFU/m^3$, and $22.0\;EU/m^3$) and MWFs industry (258, $331\;CFU/m^3$, and $8.7\;EU/m^3$). These concentrations by industry type were significantly statistically different (p < 0.01). The ratio of indoor to outdoor concentration was 6.2, 1.9, 3.2, and 3.2 for bacteria, fungi, endotoxins, and dust in the LF industry, 5.0, 0.9, 2.3, and 12.5 in the MWFs industry, and 3.7, 4.1, 3.3, and 9.7 in sawmills. The respiratory fractions of bioaerosols were differentiated by bioaerosol types and industry types: the respiratory fraction of bacteria in the LF industry, MWF industry, and sawmills was 59.4%, 72.0%, and 57.7%, respectively, and that of fungi was 77.3%, 89.5%, and 83.7% in the same order. Conclusion: We found that bioaerosol concentration was the highest in sawmills, followed by LF industry facilities and MWFs industry facilities. The indoor/outdoor ratio of microorganisms was larger than 1 and respiratory fraction of microorganisms was more than 50% of the total microorganism concentrations which might penetrate respiratory tract easily. All these findings suggest that bioaerosol in the surveyed industries should be controlled to prevent worker respiratory diseases.

• Journal of Periodontal and Implant Science
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• v.49 no.5
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• pp.270-286
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• 2019

Purpose: Despite the well-known anti-inflammatory effects of vitamin D in periodontal health, its mechanism has not been fully elucidated. In the present study, the effect of vitamin D on strengthening E-cadherin junctions (ECJs) was explored in human gingival keratinocytes (HGKs). ECJs are the major type of intercellular junction within the junctional epithelium, where loose intercellular junctions develop and microbial invasion primarily occurs. Methods: HOK-16B cells, an immortalized normal human gingival cell line, were used for the study. To mimic the inflammatory environment, cells were treated with tumor necrosis factor-alpha ($TNF-{\alpha}$). Matrix metalloproteinases (MMPs) in the culture medium were assessed by an MMP antibody microarray and gelatin zymography. The expression of various molecules was investigated using western blotting. The extent of ECJ development was evaluated by comparing the average relative extent of the ECJs around the periphery of each cell after immunocytochemical E-cadherin staining. Vitamin D receptor (VDR) expression was examined via immunohistochemical analysis. Results: $TNF-{\alpha}$ downregulated the development of the ECJs of the HGKs. Dissociation of the ECJs by $TNF-{\alpha}$ was accompanied by the upregulation of MMP-9 production and suppressed by a specific MMP-9 inhibitor, Bay 11-7082. Exogenous MMP-9 decreased the development of ECJs. Vitamin D reduced the production of MMP-9 and attenuated the breakdown of ECJs in the HGKs treated with $TNF-{\alpha}$. In addition, vitamin D downregulated $TNF-{\alpha}$-induced nuclear factor kappa B ($NF-{\kappa}B$) signaling in the HGKs. VDR was expressed in the gingival epithelium, including the junctional epithelium. Conclusions: These results suggest that vitamin D may avert $TNF-{\alpha}$-induced downregulation of the development of ECJs in HGKs by decreasing the production of MMP-9, which was upregulated by $TNF-{\alpha}$. Vitamin D may reinforce ECJs by downregulating $NF-{\kappa}B$ signaling, which is upregulated by $TNF-{\alpha}$. Strengthening the epithelial barrier may be a way for vitamin D to protect the periodontium from bacterial invasion.

• Journal of Chest Surgery
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• v.38 no.1 s.246
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• pp.38-43
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• 2005

Matrix metalloproteinase-2 (MMP-2) is a class of proteolytic enzymes that digest collagen type IV and other components of the basement membrane. It plays a key role in the local invasion and the formation of distant metastases by various malignant tumors. The aim of this study was to evaluate the activity of MMP-2 and its significance as a prognostic marker in resected stage I non-small cell lung cancer (NSCLC). Material and Method: In this study we obtained fresh-frozen samples of tumor and non-tumor tissues from 34 patients with stage I NSCLC who underwent resection without preoperative radiotherapy or chemotherapy. After the extraction of total protein from tissue samples, MMP-2 activities were assessed by gelatin-substrate-zymography. The activities were divided into the higher or lower groups. Result: The MMP-2 activities were higher in tumor tissues than in non-tumor tissues. The MMP-2 activity of non-tumor tissues in recurrent group was higher than in non-recurrent group (p<0.01). Also the patients with higher MMP-2 activity of non-tumor tissues showed poor 5 year survival (p<0.01). Conclusion: This result indicates that the higher level of MMP-2 activity in the non-tumor tissue is associated with the recurrence and survival after the resection of stage I NSCLC. Therefore, MMP-2 activity in the non-tumor tissue could be used as a potential prognostic marker for the resected stage I-NSCLC.