• Title/Summary/Keyword: Species-specific Primer

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Effect of Moutan Cortex Radicis on gene expression profile of differentiated PC12 rat cells oxidative-stressed with hydrogen peroxide (모단피의 PC12 cell 산화억제 효과 및 neuronal 유전자 발현 profile 분석에 대한 연구)

  • Kim Hyun Hee;Rho Sam Woong;Na Youn Gin;Bae Hyun Su;Shin Min Kyu;Kim Chung Suk;Hong Moo Chang
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.17 no.2
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    • pp.529-541
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    • 2003
  • Yukmijihwang-tang has been widely used as an and-aging herbal medicine for hundred years in Asian countries. Numerous studies show that Yukmijihwangtang has anti-oxidative effect both in vivo and in vitro. It has been reported that Moutan Cortex Radicis extract (MCR) was the most effective herb in Yukmijihwang-tang on undifferentiated PC12 cells upon oxidative-stressed with hydrogen peroxide. The purpose of this study is to; 1) evaluate the recovery of neuronal damage by assessing the anti-oxidant effect of MCR on PC12 cells differentiated with nerve growth factor (NGF), 2) identify candidate genes responsible for anti-oxidative effect on differentiated PC12 cells by oligonucleotide chip microarray. PC12 cells, which were differentiated by treating with NGF, were treated without or with hydrogen peroxide in the presence or absence of various concentration of MCR. Cell survival was determined by using MTS assay. Measurement of intracellular reactive oxygen species (ROS) generation was determined using the H2DCFDA assay The viability of cells treated with MCR was significantly recovered from stressed PC12 cell. In addition, wide rage of concentrations of MCR shows dose-dependent inhibitory effect on ROS production in oxidative-stressed cells. Total RNAs of cells without treatment(Control group), only treated with H₂O₂ (stressed group) and treated with both H₂O₂ and of MCR (MCR group) were isolated, and cDNAs was synthesized using oligoT7(dT) primer. The fragmented cRNAs, synthesized from cDNAs, were applied to Affymetrix GeneChip Rat Neurobiology U34 Array. mRNA of Calcium/calmodulin-dependent protein kinase II delta subunit(CaMKII), neuron glucose transporter (GLUT3) and myelin/oligodendrocyte glycoprotein(MOG) were downregulated in Stressed group comparing to Control group. P2X2-5 receptor (P2X2R-5), P2X2-4 receptor (P2X2R-4), c-fos, 25 kDa synaptosomal attachment protein(SNAP-25a) and GLUT3 were downregulated, whereas A2 adenosine receptor (A2AR), cathechol-O-methyltransferase(COMT), glucose transporter 1 (GLUT1), EST223333, heme oxygenase (HO), VGF, UI-R-CO-ja-a-07-0-Ul.s1 and macrophage migration inhibitory factor (MIF) were upregulated in MCA group comparing to Control group. Expression of Putative potassium channel subunit protein (ACK4), P2X2A-5, P2X2A-4, Interferon-gamma inducing factor isoform alpha precursor (IL-18α), EST199031, P2XR, P2X2 purinoceptor isoform e (P2X2R-e), Precursor interleukin 18 (IL-18) were downregulated, whereas MOO, EST223333, GLUT-1, MIF, Neuronatin alpha, UI-R-C0-ja-a-07-0-Ul.s1, A2. adenosine receptor, COMT, neuron-specific enolase (NSE), HO, VGF, A rat novel protein which is expressed with nerve injury (E12625) were upregulated in MCR group comparing to Stressed group. The results suggest that decreased viability and AOS production of PC12 cell by H₂O₂ may be, at lease, mediated by impaired glucose transporter expression. It is implicated that the MCR treatment protect PC12 cell from oxidative stress via following mechanisms; improving glucose transport into the cell, enhancing expression of anti-oxidative genes and protecting from dopamine cytotoxicity by increment of COMT and MIF expression. The list of differentially expressed genes may implicate further insight on the action and mechanism behind the anti-oxidative effects of herbal extract Moutan Cortex Radicis.

Establishment of PCR Conditions for the Identification of Stenotrophomonas maltophilia Isolated from Boar Semen and Antimicrobial Susceptibility Patterns of the Isolates (돼지 정액에서 분리된 Stenotrophomonas maltophilia 확인을 위한 PCR 기법 개발 및 분리 균주의 항생제 감수성 양상)

  • Jung, Byeong-Yeal;Park, Bum-Soo;Kim, Ha-Young;Byun, Jae-Won;Kim, Ae-Ran;Jeon, Albert Byung-Yun;Kim, In-Cheul;Chung, Ki-Hwa
    • Journal of Life Science
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    • v.22 no.8
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    • pp.1114-1119
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    • 2012
  • Bacteria are frequently contaminated during the collection and processing procedures of boar semen. Of the contaminants, Stenotrophomonas (S.) maltophilia is a Gram-negative bacterium that is widely distributed in a variety of habitats. Although PCR assays have been developed for the detection of S. maltophilia, they cross-react with some species of Xanthomonas. In this study, we designed a primer set for the detection of S. maltophilia in order to target the chiA (GenBank accession no. NC_010943) gene. The specific PCR products were amplified from S. maltophilia only, not from other tested strains that are frequently found in semen. The detection limit of the PCR was $1.5{\times}10^3$ CFU/ml with pure-cultured S. maltophilia and $1.5{\times}10^4$ CFU/ml with S. maltophilia spiked in semen. Twenty-six (5.9%) S. maltophilia were isolated from 440 semen samples. The PCR results exhibited 98.9% agreement with a comparison of S. maltophilia isolation. Also, the sensitivity and specificity of the PCR were 100% and 98.7%, respectively. In the antimicrobial susceptibility test, S. maltophilia isolates were highly susceptible to enrofloxacin and florfenicol, while the majority of them were resistant to amoxicillin/clavulanic acid, apramycin, ceftiofur, penicillin, and spectinomycin. These results indicated that the PCR using the chiA gene was proven to be reliable and effective for the detection of S. maltophilia with high levels of sensitivity and specificity.

DETECTION OF METHICILLIN OR VANCOMYCIN-RESISTANT STAPHYLOCOCCUS AUREUS FROM DENTAL HOSPITAL (치과병원 진료실 내에서 메티실린 또는 반코마이신 저항성 Staphylococcus aureus의 검출)

  • Min, Jung-Hee;Park, Soon-Nang;Hwang, Ho-Keel;Min, Jung-Beum;Kim, Hwa-Sook;Kook, Joong-ki
    • Restorative Dentistry and Endodontics
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    • v.32 no.2
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    • pp.102-110
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    • 2007
  • The purpose of this study was to obtain the basic information for the improvement of dental environment by investigating the presence of methicillin- or vancomycin-resistant Staphylococcus aureus (MRSA or VRSA) isolated from dental health care workers (DHCWs) and environment of the Chosun University Dental Hospital (CUDH) and a private dental clinic (control group). Staphylococcus aureus (S. aureus) was isolated from anterior nares of 42 DHCWS and 38 sites, unit chairss, x-ray devices, computers, etc., at 10 departments of the CUDH and 20 DHCWs and 11 sites at the private dental clinic. S. aureus was isolated on mannitol salt agar plate and confirmed by PCR with S. aureus species-specific primer. Antimicrobial susceptibility test of clinical isolates of S. aureus against several antibiotics including methicillin (oxacillin) was performed by investigating minimum inhibitory concentration (MIC) using broth microdilution assay. In addition, PCR was performed to detect the methicillin- or vancomycin-resistant gene. The data showed that one strain of S. aureus was isolated from DHCWs of the CUDH and three strains of S. aureus was isolated from 3 samples of the private dental clinic, respectively. All of the isolates from the CUDH and the private dental clinic had resistance to penicillin G, amoxicillin and vancomycin and susceptibility to oxacillin and ciprofloxacin. The S. aureus strains were already obtained the resistance to penicillin G and amoxicillin. These results suggest that two dental clinics were under relatively safe environment.