• 제목/요약/키워드: Cell-based assay

검색결과 663건 처리시간 0.027초

Isolation and Characterization of PERV-C env from Domestic Pig in Korea

  • Park, Sung-Han;Bae, Eun-Hye;Park, Sang-Min;Park, Jin-Woo;Lim, Mi-Suk;Jung, Yong-Tae
    • Journal of Microbiology and Biotechnology
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    • 제18권10호
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    • pp.1735-1740
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    • 2008
  • Clone PERV-C (A3) env was isolated from the genomic DNA of domestic pig (Sus scrofa domesticus) in Korea to investigate the molecular properties of PERV-C. The nucleic acid homologies between the PERV-MSL (type C) reference and the PERV-C(A3) clone was 99% for env, but a single base pair deletion was found in the transmembrane (TM) region of the env open reading frame. To examine the functional characteristics of truncated PERV-C env, we constructed a replication-incompetent retroviral vector by replacing the env gene of the pCL-Eco retrovirus vector with PERV-C env. A retroviral vector bearing PERV-C/A chimeric envelopes was also created to complement the TM defect. Our results indicated that truncated PERV-C env was not infectious in human cells as expected. Interestingly, however, the vector with the PERV-C/A envelope was able to infect 293 cells. This observation suggests that recombination within PERV-C TM could render PERV-C infectious in humans. To further characterize PERV-C/A envelopes, we constructed an infectious molecular clone by using a PCR-based technique. This infectious molecular clone will be useful to examine more specific regions that are critical for human cell tropism.

Comparative Biodegradation of HDPE and LDPE Using an Indigenously Developed Microbial Consortium

  • Satlewal, Alok;Soni, Ravindra;Zaidi, Mgh;Shouche, Yogesh;Goel, Reeta
    • Journal of Microbiology and Biotechnology
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    • 제18권3호
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    • pp.477-482
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    • 2008
  • A variety of bacterial strains were isolated from waste disposal sites of Uttaranchal, India, and some from artificially developed soil beds containing maleic anhydride, glucose, and small pieces of polyethylene. Primary screening of isolates was done based on their ability to utilize high- and low-density polyethylenes (HDPE/LDPE) as a primary carbon source. Thereafter, a consortium was developed using potential strains. Furthermore, a biodegradation assay was carried out in 500-ml flasks containing minimal broth (250ml) and HDPE/LDPE at 5mg/ml concentration. After incubation for two weeks, degraded samples were recovered through filtration and subsequent evaporation. Fourier transform infrared spectroscopy (FTIR) and simultaneous thermogravimetric-differential thermogravimetry-differential thermal analysis (TG-DTG-DTA) were used to analyze these samples. Results showed that consortium-treated HDPE (considered to be more inert relative to LDPE) was degraded to a greater extent (22.41% weight loss) in comparison with LDPE (21.70% weight loss), whereas, in the case of untreated samples, weight loss was more for LDPE than HDPE (4.5% and 2.5%, respectively) at $400^{\circ}C$. Therefore, this study suggests that polyethylene could be degraded by utilizing microbial consortia in an eco-friendly manner.

Enhancement of Antibacterial Activity of Short Tryptophan-rich Antimicrobial Peptide Pac-525 by Replacing Trp with His(chx)

  • Ahn, Mija;Rajasekaran, Ganesan;Gunasekaran, P.;Ryu, Eun Kyoung;Lee, Ga-Hyang;Hyun, Jae-Kyung;Cheong, Chaejoon;Kim, Nam-Hyung;Shin, Song Yub;Bang, Jeong-Kyu
    • Bulletin of the Korean Chemical Society
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    • 제35권9호
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    • pp.2818-2824
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    • 2014
  • Trp residue is considered as one of the important constituents in antimicrobial peptides (AMPs) as it presence secured good activities in many cases. However, it is preferable to be changed because of their sensitivity towards light. We have synthesized the short Trp-rich AMP Pac-525 and its analogues to investigate the possibility of His(chx) as possible replacement analogue for Trp in AMPs. Based on the assay result of the antibacterial activity including anti-MRSA activity, His(chx) is considered as good candidate for the Trp replacement. Through these study, we found that His(chx) had several merits to design therapeutic antimicrobial agents compare to Trp in terms of i) increasing antibacterial activity without hemolytic activity, ii) successful in designing the short peptide (only four residues), iii) having anti-MRSA activity, iv) overcoming the light sensitivity. Furthermore, transmission electron microscopy (TEM) and dye leakage experiments suggested that P11 and P16 containing His(chx) kill bacteria via forming pore/ion channels on bacterial cell membranes.

동의보감에 수재된 오계(烏鷄)에 대한 생리활성 연구 (The Study on Biological Activities of Yeonsan Ogye listed on Dong-ui-bo-gam)

  • 김진우;심부용;최학주;이해진;김동희
    • 대한본초학회지
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    • 제30권5호
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    • pp.23-28
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    • 2015
  • Objectives : The aim of this study is to investigate cell viability, anti-inflammatory, antioxidant, immunoenhancing activity using various extracts of Yeonsan Ogye.Methods : In order to evaluate cytotoxicity, MTT assay was performed. We investigated production levels of pro-inflammatory cytokines such as tumor necrosis factor (TNF)-αand interleukin (IL)-6, and nitric oxide(NO) in LPS-induced RAW 264.7 cells. NO production in RAW 264.7 cells was measured by using Griess reagent. Cytokines including IL-6 and TNF-αwere measured by Luminex and ROS was measured by Flow cytometry.Results : No cytotoxicity of various extracts of Yeonsan Ogye was observed in RAW 264.7 cells. Productions of ROS in RAW 264.7 cells were increased from extraction of bones and decreased from extraction of skin. Also, productions of NO in RAW 264.7 cells were increased to bone extract and decreased at skin extract. In addition, productions of pro-inflammatory cytokines (IL-6 and TNF-α) in LPS-induced RAW 264.7 cells were decreased at skin, meat extracts, respectively. Finally, the levels of immune-related cytokines (IL-6 and TNF-α) were increased compared to those of the normal group.Conclusions : It is concluded that Yeonsan Ogye extracts seem to have significant biological activities likes anti-inflammatory, antioxidant, immuno-enhancing etc. These results may be developed as a raw material for new health food and new therapeutics to ease the symptoms related with inflammatory and oxidative stress. In terms of oriental traditional medicine, we expect that it contribute to building of EBM (Evidence-Based Medicine) from the this result.

수세미오이(Sponge-gourd: Luffa cylindrica L.) 추출물의 항산화, 항돌연변이 및 암세포 증식 억제 효과 (Antioxidative and Antimutagenic Effects and Hyperplasia Inhibitory Activity of Cancer Cells from Luffa cylindrica Extracts)

  • 오세인;이미숙
    • 한국식품영양학회지
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    • 제25권4호
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    • pp.888-896
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    • 2012
  • Considering the dearth of information regarding the medicinal properties of Luffa cylindrica, we assessed the antioxidative, antimutagenic and hyperplasia inhibitory activity of cancer cells from Luffa cylindrica extracts by employing biological and biochemical assays. Ethanol extracts of Luffa cylindrica inhibited MDA-BSA (malondialdehyde-bovine serum albumin) conjugation reaction ($66.38{\pm}2.65$), DPPH (1,1-diphenyl-2-picryl-hydrazyl) radical production ($60.13{\pm}0.42$) and lipid peroxidation ($56.04{\pm}3.24$). In this study, Luffa cylindrica is believed to exert possible antioxidative effects. The direct and indirect antimutagenic effects of the ethanol extracts of Luffa cylindrica were examined by the Ames test using Salmonella typimurium TA98 and TA100. The inhibitory effects on indirect and direct mutagenicity shows an weak tendency, particularly in direct mutagenicity mediated by 2-nitrofluorene in Salmonella typimurium TA98 ($5.82{\pm}5.74$) and in indirect mutagenicity mediated by 2-anthramine in Salmonella typimurium TA100 ($5.76{\pm}2.15$). The ethanol extracts of Luffa cylindrica on cancer cell hyperplasia inhibitory activity via MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay exerted cytotoxic effects on Hela cells ($55.83{\pm}3.83$) and MCF-7 cells ($33.03{\pm}2.09$), which were used in this study. Based on these results, it believed that the ethanol extracts of Luffa cylindrica have antioxidative capacities as well as hyperplasia inhibitory activity of cancer cells. Furthemore, Luffa cylindrica is a candidate for the prevention and dietetic treatment of chronic diseases and for the development of functional food.

Acinetobacter sp.1의 일산화탄소 산화효소의 특성 (Carbon Monoxide Dehydrogenase in Cell Extracts of an Acinetobacter Isolate)

  • 조진원;김영민
    • 미생물학회지
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    • 제24권2호
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    • pp.133-140
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    • 1986
  • 일산화탄소를 이용하여 자가영양적으로 성장한 Acinetobacter sp. 1 의 세포추출액은 혐 기성 실험조건하에서 thionin, methylene blue, 2,6-dichlorophenol-indophenol둥올 일산화탄소의 산회를 위한 전자수용체로 사용할 수 있었으나 NAD, NADP, FAD, 또는 FMN등은 천자수용체로 이용하지 못하였다. 이 세균에 존재하는 일산화탄소 산화효소는 유도효소로 밝혀졌고, pH 7.5와 $60^{\circ}C$에서 최대의 활성을 나타내었다. 이 효소의 활성화에너지는 6.1kcal/mol (25.5 kJ/mol)이며 일산화탄소에 대한 Km값은 $154{\mu}M$로 밝혀졌다. 그리고 잘 알려진 몇가지 금속 chelat tIng agent와 2가의 양이온들은 이 효소의 활성에 거의 영향을 미치지 않았는데 $Cu^{2+}$ 이온만은 이 효소의 활성을 완전히 억제시켰다. 또한 이 효소는 포도당과 숙신산에 의해 활성이 저해되었으며, hydrogenase의 활성도 나타내었다. 그리고 Acinetobacter sp. 1의 일산화탄소 산화효소는 Pseudomonas carboxydohydrogena의 일산화탄소 산화효소와 연역학적인 연관성이 없는 것으로 나타났다.

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장기적인 인삼섭취가 인체의 혈전 관련 요인에 미치는 영향 (The Effect of the Long-Term Ginseng Intakes on Serum Lipids Profile and Hemostatic Factors in Human)

  • 이정희
    • Journal of Nutrition and Health
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    • 제28권9호
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    • pp.862-871
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    • 1995
  • This study was designed to investigate the effectiveness of ginseng intakes in modifying serum lipid profiles and plasma clotting factors. The participants in this study were 47 normal healthy volunteers(men 24, women 23) with an age range of 35-49 years and a mean age of 41 years residing in Taejon. Based on the diet intakes, subjects were classed into one of three groups : control, vegetarian, and ginseng consumed over 3-4 years. There was no significant difference in their physical characteristics. Dietary calorie intakes were not significantly different in subjects. The ratio of energy intake in the control and ginseng consumed group was 63-64% : 20-21% : 15-16%(Cho : Fat : Pro), but 70-73% : 13-14% :14-15%(Cho : Fat : Pro) in the vegetarians. The intakes of animal food in the vegetarian was significantly lower than the control and ginseng consumed group in men. The ratio of P/S(1.27) was the highest in the vegetarians. Venous blood samples were taken for serum lipid profiling, plasma clotting assay and platelet function. The concentration of serum triglyceride in the men ginseng group is significantly lower than those of the men control group. Serum lipid profiles values of the men ginseng group, such as total cholesterol and phospholipid were lower those of the men control group, but higher those of the men vegetarian group. the serum lipid profile in the women were not significant, but total cholesterol, triglyceride and LDL cholesterol levels in the ginseng groups were low. The concentration of HDL cholesterol was not significantly different. Platelet cell count and platelet aggregation were low in the ginseng groups. APTT(Activated Partial thromboplastin time) was significantly elongated in ginseng groups in the normal range. In seems that the major beneficial effects of ginseng intakes in especially men were on the blood concentrations of triglyceride, total cholesterol and elongation of plasma clotting time.

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Anti-oxidative, Nitric Oxide Inhibitory Activities and Irritation Test of the Fermented Opuntia humifusa Cladodes

  • Chang, Zhi-Qiang;Hwang, Mi-Hyun;Oh, Byung-Chul;Lee, Sam-Pin;Rhee, Man-Hee;Kim, Kil-Soo;Jeong, Kyu-Shik;Kim, Jong-Choon;Park, Seung-Chun
    • Toxicological Research
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    • 제22권3호
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    • pp.229-235
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    • 2006
  • Opuntia humifusa is a member of the Cactaceae family. In the present study, the antioxidant, nitric oxide(NO) inhibitory activities and potential irritation response of the fermented Opuntia humifusa cladodes(FOH) were investigated for cosmetic use. Antioxidant activities were tested using 1,1-diphenyl-2-picrylhydrazyl(DPPH) and xanthine oxidase assay, we found that FOH could scavenge DPPH free radicals and inhibit xanthine oxidase activity in a dose dependent manner, with $IC_{50}$ of 2599.46${\mu}g/ml$ and 721.38${\mu}g/ml$, respectively. To investigate the possible anti-inflammatory effects of FOH, RAW 264.7 macrophages were pretreated with FOH($0{\sim}400{\mu}g/ml$) for 30 min and then treated with LPS for 24 h. We found that cell number did not vary significantly with the treatment of FOH, and FOH did not show any inhibitory effects on LPS-induced NO production. After application of FOH to rabbits for skin and eye irritation test, the experimental sites did not show any response compared to the control. FOH were considered to be a non-irritant to the skin and eye. Based on the above information, we suggest that FOH can be considered to be a non-irritant base cosmetic material for safely use.

Generation and Expression in Plants of a Single-Chain Variable Fragment Antibody Against the Immunodominant Membrane Protein of Candidatus Phytoplasma Aurantifolia

  • Shahryari, F.;Safarnejad, M.R.;Shams-Bakhsh, M.;Schillberg, S.;Nolke, G.
    • Journal of Microbiology and Biotechnology
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    • 제23권8호
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    • pp.1047-1054
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    • 2013
  • Witches' broom of lime is a disease caused by Candidatus Phytoplasma aurantifolia, which represents the most significant global threat to the production of lime trees (Citrus aurantifolia). Conventional disease management strategies have shown little success, and new approaches based on genetic engineering need to be considered. The expression of recombinant antibodies and fragments thereof in plant cells is a powerful approach that can be used to suppress plant pathogens. We have developed a single-chain variable fragment antibody (scFvIMP6) against the immunodominant membrane protein (IMP) of witches' broom phytoplasma and expressed it in different plant cell compartments. We isolated scFvIMP6 from a naïve scFv phage display library and expressed it in bacteria to demonstrate its binding activity against both recombinant IMP and intact phytoplasma cells. The expression of scFvIMP6 in plants was evaluated by transferring the scFvIMP6 cDNA to plant expression vectors featuring constitutive or phloem specific promoters in cassettes with or without secretion signals, therefore causing the protein to accumulate either in the cytosol or apoplast. All constructs were transiently expressed in Nicotiana benthamiana by agroinfiltration, and antibodies of the anticipated size were detected by immunoblotting. Plant-derived scFvIMP6 was purified by affinity chromatography, and specific binding to recombinant IMP was demonstrated by enzyme-linked immunosorbent assay. Our results indicate that scFvIMP6 binds with high activity and can be used for the detection of Ca. Phytoplasma aurantifolia and is also a suitable candidate for stable expression in lime trees to suppress witches' broom of lime.

Neuroprotective effect of caffeic acid phenethyl ester in 3-nitropropionic acid-induced striatal neurotoxicity

  • Bak, Jia;Kim, Hee Jung;Kim, Seong Yun;Choi, Yun-Sik
    • The Korean Journal of Physiology and Pharmacology
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    • 제20권3호
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    • pp.279-286
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    • 2016
  • Caffeic acid phenethyl ester (CAPE), derived from honeybee hives, is a bioactive compound with strong antioxidant activity. This study was designed to test the neuroprotective effect of CAPE in 3-nitropropionic acid (3NP)-induced striatal neurotoxicity, a chemical model of Huntington's disease (HD). Initially, to test CAPE's antioxidant activity, a 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid (ABTS) antioxidant assay was employed, and CAPE showed a strong direct radical-scavenging effect. In addition, CAPE provided protection from 3NP-induced neuronal cell death in cultured striatal neurons. Based on these observations, the in vivo therapeutic potential of CAPE in 3NP-induced HD was tested. For this purpose, male C57BL/6 mice were repeatedly given 3NP to induce HD-like pathogenesis, and 30 mg/kg of CAPE or vehicle (5% dimethyl sulfoxide and 95% peanut oil) was administered daily. CAPE did not cause changes in body weight, but it reduced mortality by 29%. In addition, compared to the vehicle-treated group, robustly reduced striatal damage was observed in the CAPE-treated animals, and the 3NP-induced behavioral deficits on the rotarod test were significantly rescued after the CAPE treatment. Furthermore, immunohistochemical data showed that immunoreactivity to glial fibrillary acidic protein (GFAP) and CD45, markers for astrocyte and microglia activation, respectively, were strikingly reduced. Combined, these data unequivocally indicate that CAPE has a strong antioxidant effect and can be used as a potential therapeutic agent against HD.