• Title/Summary/Keyword: ITS-PCR

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Inhibitory effects of Paridis Rhizoma in the activation and proliferation of eosinophils: implications on its regulatory roles for asthma (rmIL-5로 유도된 호산구의 활성화 및 성장에서 중루의 천식반응 억제효과)

  • Sin, Mi-Kyung;Kil, Ki-Jeong;Lee, Young-Chul;Kim, Jin-Suk;Seo, Young-Bae;Roh, Seong-Soo
    • The Korea Journal of Herbology
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    • v.20 no.2
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    • pp.159-169
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    • 2005
  • Objectives : This study was carried out for the purpose of knowing the effect from anti-arthma action of the abstraction from a extract of Paridis Rhizoma(EPR). In order to know what the effect of controlling an abstraction from Paridis Rhizoma. and about the expression of B cells and Ig E cells, mast cells it was necessary for it to be activated by ovalbumin. Methods : In order to know what the effect was on the organization of cytokine gene expression from The increase and divorce of the B cells and allergic acting by EPR, we found it necessary to examine the BALF. At the same time, as we examined the histamine release by ELISA method, we also examined the effect of EPR. Results : EPR at $100\;{\mu}g/ml$, the highest concentration examined did not have any cytotoxic effects on mLFCs. In FACS analysis, number of granulocyte/lymphocyte, $CD3e^+/CCR3^+,\;CD4^+\;and\;CD23^+/B220^+$ in asthma-induced lung cells were significantly decreased by EPR treatment compared to the control group. In RT-PCR analysis, mRNA expression for CCR3, eotaxin and histamine in asthma-induced lung cells, which was induced by rIL-3 plus rmIL-5 treatments, was significantly decreased by EPR treatment. In ELISA analysis, production levels of IL-4, IL-13 and histamine in asthma-induced lung cells, which were induced by rIL-3 plus rmIL-5 co-treatment, were significantly decreased by EPR treatment. EPR treatments significantly inhibited the proliferation of eosinohils prepared from asthma-induced mouse lung tissues compared to the non-EPR treated control cells. Immunohistochemical analysis revealed that EPR treatment significantly decreased the levels of eosipnphil activation compared to non-treated cells. Conclusion : The present data suggested that Paridis Rhizoma may have an effects on the inhibition of parameters associated with asthma responses in eosinpophils, and thus implicate the possibility for the clinical application of Paridis Rhizoma.

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Comparing the anti-inflammatory effect of nanoencapsulated lycopene and lycopene on RAW 264.7 macrophage cell line (RAW 264.7 대식세포주에서 나노입자화 리코펜의 항염증 증진 효과)

  • Seo, Eun Young;Kim, Myung Hwan;Kim, Woo-Kyoung;Chang, Moon-Jeong
    • Journal of Nutrition and Health
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    • v.48 no.6
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    • pp.459-467
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    • 2015
  • Purpose: We developed a method to load lycopene into maltodextrin and cyclodextrin in an attempt to overcome the poor bioavailability and improve the anti-inflammatory effect of this polyphenol. Methods: Nanosized lycopenes were encapsulated into biodegradable amphiphillic cyclodextrin and maltodextrin molecules prepared using a high pressure homogenizer at 15,000~25,000 psi. Cell damage was induced by lipopolysaccharides (LPS) in a mouse macrophage cell line, RAW 264.7. The cells were subjected to various doses of free lycopene (FL) and nanoencapsulated lycopene (NEL). RT-PCR was used to quantify the tumor necrosis factor (TNF-${\alpha}$), interleukin-$1{\beta}$ (IL-$1{\beta}$), IL-6, inducible nitric oxide synthase (iNOS), and cyclooxigenase-2 (COX-2) mRNA levels, while ELISA was used to determine the protein levels of TNF-${\alpha}$, IL-$1{\beta}$, and IL-6. Results: NEL significantly reduced the mRNA expression of IL-6 and IL-$1{\beta}$ at the highest dose, while not in cells treated with FL. In addition, NEL treatment caused a significant reduction in IL-6 and TNF-${\alpha}$ protein levels, compared to cells treated with a similar dose of FL. In addition, mRNA expression of iNOS and COX-2 enzyme in the activated macrophages was more efficiently suppressed by NEL than by FL. Conclusion: Overall, our results suggest that lycopene is a potential inflammation reducing agent and nanoencapsulation of lycopene can further improve its anti-inflammatory effect during tissue-damaging inflammatory conditions.

Characterization of a Chitinase Gene and Screening of Cold Active Chitinase from Polar Microorganisms (극지유래 저온활성 Chitinase 생산균주의 스크리닝과 Chitinase 유전자 클로닝)

  • Park, Yu Kyung;Kim, Jung Eun;Lee, Hyoungseok;Kim, Ji Hyun;Park, Ha Ju;Kim, Dockyu;Park, Mira;Yim, Joung Han;Kim, Il-Chan
    • Korean Journal of Microbiology
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    • v.48 no.4
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    • pp.293-297
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    • 2012
  • Of the 169 strains of microorganisms stored in Polar and Alpine Microbial Collection of Korea Polar Research Institute, 27 strains were selected for their chitinase activity on ZoBell plates supplemented with 0.4% colloidal chitin. Among them, PAMC 21693 strain have shown the highest chitinolytic enzyme activity toward pNP-$(GlcNAc)_1$ at low temperature and the highest growth rate at $4^{\circ}C$. We cloned a full-length chitinase gene of 2,857 bp which contains an open reading frame of 2,169 bp encoding 872-amino acid polypeptide. Recombinant chitinase protein was expressed in E. coli and its molecular weight was confirmed 96 kDa. In this paper, we suggest the potential use of cold-active chitinase from polar microorganisms in the field of biotechnology.

The Functional Relevance of Prepro-melanin Concentrating Hormone (pMCH) to Skin Color Change, Blind-side Malpigmentation and Feeding of Oliver Flounder Paralichthys olivaceus

  • Kang, Duk-Young;Kim, Hyo-Chan;Kang, Han-Seung
    • Fisheries and Aquatic Sciences
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    • v.17 no.3
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    • pp.325-337
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    • 2014
  • To assess the functional structure of prepro-melanin-concentrating hormone (pMCH), we isolated and cloned pMCH (of-pMCH) mRNA from the brain of the olive flounder, Paralichthys olivaceus, and compared its amino acid sequence with those from other animals. In addition, to examine whether activation of the brain of-pMCH gene is influenced by background color, density, and feeding, we compared pMCH mRNA activities against different background colors (bright and dark) and at different densities (100% PCA and 200% PCA). To examine whether the pMCH gene is related with malpigmentation of blind-side skin and appetite, we compared pMCH gene expression between ordinary and hypermelanic flounders, and between feeding and fasting flounders. The of-pMCH cDNA was 405 bp in the open reading frame [ORF] and encoded a protein of 135 amino acids; MCH was 51 bp in length and encoded a protein of 17 amino acids. An obvious single band of the expected size was obtained from the brain and pituitary by RT-PCR. In addition, of-pMCH gene activity was significantly higher in the bright background only at low density (< 100% PCA) making the ocular skin of fish whitening, and in ordinary fish. However, the gene activity was significantly decreased in dark background, at high density (>200% PCA), and in hypermelano fish. These results suggest that skin whitening camouflage of the flounder is induced by high MCH gene activity, and the density disturbs the function of background color in the physiological color change. Moreover, our data suggest that a low level of MCH gene activity may be related to malpigmentation of the blind-side skin. In feeding, although pMCH gene activity was significantly increased by feeding in the white background, the pMCH gene activity in the dark background was not influenced by feeding, indicating that the MCH gene activity increased by feeding can be offset by dark background color, or is unaffected by appetite. In conclusion, this study showed that MCH gene expression is related to ocular-skin whitening camouflage and blind-skin hypermelanosis, and is influenced by background color and density.

Cloning and Characterization of a Novel Mannanase from Paenibacillus sp. BME-14

  • Fu, Xiaoyu;Huang, Xiaoluo;Liu, Pengfu;Lin, Ling;Wu, Gaobing;Li, Chanjuan;Feng, Chunfang;Hong, Yuzhi
    • Journal of Microbiology and Biotechnology
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    • v.20 no.3
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    • pp.518-524
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    • 2010
  • A mannanase gene (man26B) was obtained from a sea bacterium, Paenibacillus sp. BME-14, through the constructed genomic library and inverse PCR. The gene of man26B had an open reading frame of 1,428 bp that encoded a peptide of 475- amino acid residues with a calculated molecular mass of 53 kDa. Man26B possessed two domains, a carbohydrate binding module (CBM) belonging to family 6 and a family 26 catalytic domain (CD) of glycosyl hydrolases, which showed the highest homology to Cel44C of P. polymyxa (60% identity). The optimum pH and temperature for enzymatic activity of Man26B were 4.5 and $60^{\circ}C$, respectively. The activity of Man26B was not affected by $Mg^{2+}$ and $Co^{2+}$, but was inhibited by $Hg^{2+},\;Ca^{2+},\;Cu^{2+},\;Mn^{2+},\;K^+,\;Na^+$, and $\beta$-mercaptoethanol, and slightly enhanced by $Pb^{2+}$ and $Zn^{2+}$. EDTA did not affect the activity of Man26B, which indicates that it does not require divalent ions to function. Man26B showed a high specific activity for LBG and konjac glucomannan, with $K_m,\;V_{max}$, and $k_{cat}$ values of 3.80 mg/ml, 91.70 ${\mu}mol$/min/mg protein, and 77.08/s, respectively, being observed when LBG was the substrate. Furthermore, deletion of the CBM6 domain increased the enzyme stability while enabling it to retain 80% and 60% of its initial activity after treatment at $80^{\circ}C$ and $90^{\circ}C$ for 30 min, respectively. This finding will be useful in industrial applications of Man26B, because of the harsh circumstances associated with such processes.

Antibiotic Resistance and Bacterial Biofilm Formation by Staphylococcus aureus Strains Isolated from Various Foods (식품에서 분리한 황색포도상구균의 항생제 내성 특징 및 균막 형성)

  • Lee, Joo-Young;Wang, Hae-Jin;Shin, Dong-Bin;Cho, Yong-Sun
    • Microbiology and Biotechnology Letters
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    • v.41 no.1
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    • pp.96-104
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    • 2013
  • Staphylococcus aureus is a major human pathogen that produces a wide array of toxins, leading to a number of adverse symptoms. We examined 275 strains of Staphylococcus aureus isolated from various foods between 2006 and 2008 for antimicrobial susceptibility. At least 259 (94.2%) of the tested strains showed antibiotic resistant properties, and 106 (40.7%) of them showed multiple antibiotic resistance. Eleven of the tested strains were resistant to oxacillin and mec A-positive. Moreover, oxacillin-resistant strains were significantly more likely to be multi-drug resistant (p < 0.01). Of the 275 isolates tested, 24.4% were noted as being positive for slime production and 30.5% were positive for biofilm assay. Antibiotic resistance was not associated with a significantly higher prevalence of biofilm formation. Twenty strains were classified using the DiversiLab system. Most of the strains could be classified into 2 clusters and 4 unique types. All 10 mec A-positive strains (cluster I) were grouped together into the same sub-cluster. Cluster II (6 strains) was not found to be resistant to oxacillin in this study. Although the prevalence of methicillin-resistant S. aureus in food is currently low, the risk of its transmission through the food chain cannot be disregarded.

Microbial Floral Dynamics of Chinese Traditional Soybean Paste (Doujiang) and Commercial Soybean Paste

  • Gao, Xiuzhi;Liu, Hui;Yi, Xinxin;Liu, Yiqian;Wang, Xiaodong;Xu, Wensheng;Tong, Qigen;Cui, Zongjun
    • Journal of Microbiology and Biotechnology
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    • v.23 no.12
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    • pp.1717-1725
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    • 2013
  • Traditional soybean paste from Shandong Liangshan and Tianyuan Jiangyuan commercial soybean paste were chosen for analysis and comparison of their bacterial and fungal dynamics using denaturing gel gradient electrophoresis and 16S rRNA gene clone libraries. The bacterial diversity results showed that more than 20 types of bacteria were present in traditional Shandong soybean paste during its fermentation process, whereas only six types of bacteria were present in the commercial soybean paste. The predominant bacteria in the Shandong soybean paste were most closely related to Leuconostoc spp., an uncultured bacterium, Lactococcus lactis, Bacillus licheniformis, Bacillus spp., and Citrobacter freundii. The predominant bacteria in the Tianyuan Jiangyuan soybean paste were most closely related to an uncultured bacterium, Bacillus licheniformis, and an uncultured Leuconostoc spp. The fungal diversity results showed that 10 types of fungi were present in the Shandong soybean paste during the fermentation process, with the predominant fungi being most closely related to Geotrichum spp., an uncultured fungal clone, Aspergillus oryzae, and yeast species. The predominant fungus in the commercial soybean paste was Aspergillus oryzae.

Expression Analysis of Lily Type Lectin Isotypes in the Rock Bream, Oplegnathus fasciatus: in the Tissue, Developmental Stage and Viral Infection

  • Lee, Young Mee;Yang, In Jung;Noh, Jae Koo;Kim, Hyun Chul;Park, Choul-Ji;Park, Jong-Won;Noh, Gyeong Eon;Kim, Woo-Jin;Kim, Kyung-Kil
    • Development and Reproduction
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    • v.20 no.4
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    • pp.297-304
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    • 2016
  • Lectins belong to the pattern-recognition receptors (PRRs) class and play important roles in the recognition and elimination of pathogens via the innate immune system. Recently, it was reported that lily-type lectin-1 is involved when a pathogen attacks in the early immune response of fish. However, this study is limited to information that the lectin is involved in the innate immune response against viral infection. In the present study, the lily-type lectin-2 and -3 of Oplegnathus fasciatus (OfLTL-2 and 3) have been presented to be included B-lectin domain and two D-mannose binding sites in the amino acid sequence that an important feature for the fundamental structure. To investigate the functional properties of OfLTLs, the tissue distribution in the healthy rock bream and temporal expression during early developmental stage analysis are performed using quantitative real-time PCR. OfLTL-2 and 3 are predominantly expressed in the liver and skin, but rarely expressed in other organ. Also, the transcripts of OfLTLs are not expressed during the early developmental stage but its transcripts are increased after immune-related organs which are fully formed. In the challenge experiment with RBIV (rock bream iridovirus), the expression of OfLTLs was increased much more strongly in the late response than the early, unlike previously known. These results suggest that OfLTLs are specifically expressed in the immune-related tissues when those organs are fully formed and it can be inferred that the more intensively involved in the second half to the virus infection.

Effects of 17β-estradiol, Interleukin-1β, and Human Chorionic Gonadotropin on Activity and mRNA Expression of Plasminogen Activators in Porcine Endometrial Cells

  • Hwangbo, Yong;Cheong, Hee-Tae;Yang, Boo-Keun;Park, Choon-Keun
    • Development and Reproduction
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    • v.22 no.2
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    • pp.155-163
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    • 2018
  • This study aimed to investigate changes in the activity and mRNA expression of plasminogen activators (PAs) induced by $17{\beta}$-estradiol ($E_2$), human chorionic gonadotropin (hCG), and interleukin-$1{\beta}$ ($IL-1{\beta}$) in porcine endometrial cells. Endometrial cells were isolated from the epithelium and cultured to 80% confluence. They were then treated for 24 h with $E_2$ (0.2, 2, 20, and 200 ng/mL), $IL-1{\beta}$ (0.1, 1, 10, and 100 ng/mL), and hCG (0.5, 1, 1.5 and 2 IU/mL). mRNA expressions of urokinase-type (uPA) and tissue-type (tPA) PAs were analyzed using reverse transcription PCR, and activities were measured using a PA activity assay. mRNA expressions of uPA and tPA increased with $E_2$ treatment; however, this was not significant. Similarly, treatment with hCG did not influence the mRNA expressions of PAs. Interestingly, treatment with 0.1 ng/mL $IL-1{\beta}$ significantly reduced the mRNA expression of uPA, but did not affect that of tPA. Treatment with 2, 20, and 200 ng/mL $E_2$ increased PA activity compared with the control group; treatment with 0.1 and 1 ng/mL $IL-1{\beta}$ significantly increased PA activity compared with the other $IL-1{\beta}$ treatment groups, whereas treatment with 10 and 100 ng/mL $IL-1{\beta}$ decreased. Treatment with 2 IU/mL hCG increased PA activity compared with the other treatment groups, although there were no significant differences between the hCG and control groups. In conclusion, the activity and mRNA expression of PAs were differently regulated by the hormone/cytokine and its concentration in porcine endometrial cells. Therefore, understanding PA regulatory mechanisms may help to improve the reproductive potential of domestic animals.

Serologic Survey of Toxoplasmosis in Seoul and Jeju-do, and a Brief Review of Its Seroprevalence in Korea

  • Lim, Hyemi;Lee, Sang-Eun;Jung, Bong-Kwang;Kim, Min-Ki;Lee, Mi Youn;Nam, Ho-Woo;Shin, Jong-Gyun;Yun, Cheong-Ha;Cho, Han-Ik;Shin, Eun-Hee;Chai, Jong-Yil
    • Parasites, Hosts and Diseases
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    • v.50 no.4
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    • pp.287-293
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    • 2012
  • Knowledge of the prevalence of human Toxoplasma gondii infection is required in the Republic of Korea. In this study, we surveyed the seroprevalence of T. gondii infection and analyzed the risk factors associated with seropositivity among residents in 2 administrative districts; Seoul and the island of Jeju-do, which have contrasting epidemiologic characteristics. Sera and blood collected from 2,150 residents (1,114 in Seoul and 1,036 in Jeju-do) were checked for IgG antibody titers using ELISA and for the T. gondii B1 gene using PCR. In addition, participants completed a questionnaire that solicited information on gender, age, occupation, eating habits, history of contact with animals, and travel abroad. The T. gondii B1 gene was not detected in all residents examined. However, ELISA showed 8.0% (89 of 1,114 sera) positive for IgG antibodies against T. gondii in Seoul and 11.3% (117 of 1,036 sera) in Jeju-do. In both districts, the positive rates were higher in males than in females, and those 40-79 years of age showed higher rates than other ages. In Seoul, residents older than 70 years of age showed the highest positive rate, 14.9%, whereas in Jeju-do the highest prevalence, 15.6%, was in those in their sixties. The higher seropositive rate in Jeju-do than in Seoul may be related to eating habits and occupations. The present results and a review of related literature are indicative of an increased seroprevalence of T. gondii in Korea in recent years.