• Title/Summary/Keyword: Co-incubation

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Comparative study of 2-nitroimidazole-fluorophore-conjugated derivatives with pimonidazole for imaging tumor hypoxia

  • Seelam, Sudhakara Reddy;Hong, Mi Kyung;Lee, Yun-Sang;Jeong, Jae Min
    • Journal of Radiopharmaceuticals and Molecular Probes
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    • v.5 no.2
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    • pp.101-112
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    • 2019
  • Herein, 2-nitroimidazole-fluorophore conjugates were synthesized by linking 2-nitroimidazole and FITC or RITC via thiourea bonds. The prepared derivatives were stable for 2 h in Dulbecco's modified Eagle's medium (DMEM) at 37 ℃. The novel conjugates were studied for their in vitro uptake under hypoxic conditions using U87MG and CT-26 cell lines, showing significantly higher uptakes in hypoxic than normoxic cells. Immunohistochemical analysis confirmed hypoxia in U87MG and CT-26 xenografted tumor tissues. Moreover, the prepared conjugates were evaluated by in vivo experiments after intravenous injection in U87MG and CT-26 xenografted mice. Hypoxia was confirmed by immunohistochemistry of the prepared derivatives with co-injected pimonidazole. Confocal microscopy of the prepared derivatives showed strong fluorescence in hypoxic tumor tissues correlated with the pimonidazole distribution. This suggested that the 2-nitroimidazole-fluorophore conjugates are promising optical imaging probes for tumor hypoxia and are promising substitutes for pimonidazole immunohistochemistry, which requires a multi-step procedure of incubation involving antibody, second antibody, dye, hydrogen peroxide, and multiple washing steps.

Evaluations of Poly{vinyl alcohol)/Alginate Hydrogels Cross-linked by r-ray Irradiation Technique

  • Nam, Sang-Yong;Nho, Young-Chang;Chae, Gue-Tae;Jang, Hong-Seok;Suh, Tae-Suk;Ahn, Woong-Shick;Ryu, Kyu-Eun;Chun, Heung-Jae
    • Macromolecular Research
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    • v.12 no.2
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    • pp.219-224
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    • 2004
  • In this work, we prepared hydrogels for wound dressing from a mixture of poly(vinyl alcohol) (PVA) and alginate using the $\^$60/Co ${\gamma}$-ray irradiation technique. We examined the physical properties of these hydrogels, including gelation, water absorptivity, and gel strength, to evaluate the applicability of these hydrogels for wound dressings. The biocompatibility of these hydrogels was also evaluated in vitro, in cultures of mouse fibroblasts, and in vivo, by subcutaneous implantation studies in rats. The gel content and strength increased upon increasing the radiation dose and upon decreasing the concentration of alginate. The degree of swelling was inversely proportional to the gel content and strength. The degree of cytotoxicity of the ${\gamma}$-ray-treated hydrogels was ca. 60% compared to the (-) control (serum) after 1 day of incubation. When the incubations were prolonged up to 2 days, the toxicity of all the samples decreased remarkably and reached that of the control. Subcutaneous implantation studies in rats indicated that foreign body reactions occurring around the implanted hydrogels were moderate and became minimal upon increasing the implantation time.

Isolation and Characterization of Aniline-Degrading Bacteria

  • Kahng, Hyung-Yeel;Kim, Seung-Il;Woo, Mi-Jeong;Park, Yong-Keun;Lee, Yung-Nok
    • Korean Journal of Microbiology
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    • v.30 no.3
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    • pp.199-206
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    • 1992
  • Six isolated strains degrading aniline were selected, identified and designated as pseudomonas putida K6, Pseudomonas acidovorans K82, Achromobacter gr. D. V. K24, Achromobacter xylosocidans K4, Moraxella sp. K21 and Moraxella sp. K22. All of them degraded 1000 ppm aniline completely within 30 to 36 hours. Most of these strains are resistant to antibiotics more than one, but Moraxella sp. has not any antibiotic marker tested. Most strains except for P. acidovorans K82 were shown to have resistance to the heavy metal ions such as Ni, Cu, Li, Ba, Co, etc. but not to Hg to which only P. putida K6 was resistant. M. sp. K21 was capable of degrading aniline to a maximum concentration of 2500 ppm without any repression. The incubation of the cell in limited pH ranges (4-8) had no great effect on aniline degradation. The addition of bactopeptone to the minimal media promoted the speed of aniline degradation, but the addition of glucose rather repressed the rate of aniline degradation. Through enzyme assay, A. gr. D. V. K 24 was shown to degrade aniline through artho-pathway and formed .betha.-ketoadipate as intermediate metabolite.

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Biological Methanol Production by a Type II Methanotroph Methylocystis bryophila

  • Patel, Sanjay K.S.;Mardina, Primata;Kim, Sang-Yong;Lee, Jung-Kul;Kim, In-Won
    • Journal of Microbiology and Biotechnology
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    • v.26 no.4
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    • pp.717-724
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    • 2016
  • Methane (CH4) is the most abundant component in natural gas. To reduce its harmful environmental effect as a greenhouse gas, CH4 can be utilized as a low-cost feed for the synthesis of methanol by methanotrophs. In this study, several methanotrophs were examined for their ability to produce methanol from CH4; including Methylocella silvestris, Methylocystis bryophila, Methyloferula stellata, and Methylomonas methanica. Among these methanotrophs, M. bryophila exhibited the highest methanol production. The optimum process parameters aided in significant enhancement of methanol production up to 4.63 mM. Maximum methanol production was observed at pH 6.8, 30℃, 175 rpm, 100 mM phosphate buffer, 50 mM MgCl2 as a methanol dehydrogenase inhibitor, 50% CH4 concentration, 24 h of incubation, and 9 mg of dry cell mass ml-1 inoculum load, respectively. Optimization of the process parameters, screening of methanol dehydrogenase inhibitors, and supplementation with formate resulted in significant improvements in methanol production using M. bryophila. This report suggests, for the first time, the potential of using M. bryophila for industrial methanol production from CH4.

Inhibitory Effect of Medicinal Plant Extract on Cell Toxicity and Interleukin-8 Production Induced by Helicobacter pylori (Helicobacter pylori에 의한 세포독성 및 Interleukin-8 생성에 미치는 생약혼합물의 영향)

  • Kwon, Dong-Yeul;Gan, Cai;Shon, Yun-Hee;Nam, Kyung-Soo
    • Korean Journal of Pharmacognosy
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    • v.33 no.2 s.129
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    • pp.124-129
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    • 2002
  • The effects of Helicobacter pylori and medicinal plants extract (Leweifang) on the viability and interleukin(IL)-8 production of gastric epithelial cell were investigated. Cells viability was significantly decreased when they incubated with H. pylori or H. pylori toxin. Co-incubation with Leweifang increased H. pylori or H. pylori toxin-inhibited cell growth in a concentration-dependent manner. The production of IL-8 was greatly increased in H. pylori-infected KATO III gastric epithelial cells in a concentration- and time-dependent manner. The increased production of IL-8 was significantly inhibited by Leweifang $(1,000{\sim}5,000{\mu}g/ml)$. These results indicate that Leweifang has protective effect on H. pylori-inhibited cell growth and H. pylori-induced gastric mucosal cell inflammation by suppressing the production of inflammatory cytokine (IL-8) from gastric epithelial cells.

End-to-end Structural Restriction of α-Synuclein and Its Influence on Amyloid Fibril Formation

  • Hong, Chul-Suk;Park, Jae Hyung;Choe, Young-Jun;Paik, Seung R.
    • Bulletin of the Korean Chemical Society
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    • v.35 no.12
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    • pp.3542-3546
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    • 2014
  • Relationship between molecular freedom of amyloidogenic protein and its self-assembly into amyloid fibrils has been evaluated with ${\alpha}$-synuclein, an intrinsically unfolded protein related to Parkinson's disease, by restricting its structural plasticity through an end-to-end disulfide bond formation between two newly introduced cysteine residues on the N- and C-termini. Although the resulting circular form of ${\alpha}$-synuclein exhibited an impaired fibrillation propensity, the restriction did not completely block the protein's interactive core since co-incubation with wild-type ${\alpha}$-synuclein dramatically facilitated the fibrillation by producing distinctive forms of amyloid fibrils. The suppressed fibrillation propensity was instantly restored as the structural restriction was unleashed with ${\beta}$-mercaptoethanol. Conformational flexibility of the accreting amyloidogenic protein to pre-existing seeds has been demonstrated to be critical for fibrillar extension process by exerting structural adjustment to a complementary structure for the assembly.

Involvement of Endoplasmic Reticulum Stress in Palmitate-induced Apoptosis in HepG2 Cells

  • Cho, Hyang-Ki;Lee, Jin-Young;Jang, Yu-Mi;Kwon, Young-Hye
    • Toxicological Research
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    • v.24 no.2
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    • pp.129-135
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    • 2008
  • The results of recent studies indicate that high levels of free fatty acids(FFAs) and adipokines may be the main causes of non-alcoholic liver disease; however, the molecular mechanism that links FFAs to lipotoxicity remains unclear. In the present study, we treated HepG2 cells with FFA(either palmitate or oleate) to investigate the mechanisms involved in lipotoxicity in the liver cells. We also treated cells with palmitate in the presence of a chemical chaperone, 4-phenylbutyric acid(PBA), to confirm the involvement of ER stress in lipotoxicity. Palmitate significantly induced cytotoxicity in dose- and time-dependent manners. Apoptosis was also significantly induced by palmitate as measured by caspase-3 activity and DAPI staining. Palmitate led to increased expressions of the spliced form of X-box-protein(Xbp)-1 mRNA and C/EBP homologous transcription factor(CHOP) protein, suggesting activation of the unfolded-protein response. PBA co-incubation significantly attenuated apoptosis induced by palmitate. The above data demonstrate that high levels of palmitate induce apoptosis via the mediation of ER stress in the liver cells and that chemical chaperones act to modulate ER stress and accompanying apoptosis.

Effects of teat dipping disinfectant containing biodegradable iodophor microspheres on preventing dry period intramammary infection in dairy cows (생분해성 iodophor microsphere 함유 유두침지 소독제의 건유기 젖소에서의 유방내 신규감염 예방 효과)

  • Hwang, Cheol-Yong;Kim, Jong-Min;Youn, Hwa-Young;Han, Bo;Han, Hong-Ryul
    • Korean Journal of Veterinary Research
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    • v.44 no.1
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    • pp.113-120
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    • 2004
  • 1% iodophor loaded microspheres of PLGA (Poly[DL-Lactide-co-Glycolide]) were prepared by solvent evaporation method and were applied to the cows on dry period for evaluating it's preventive effects on intramammary infections. The morphology of the microspheres were evaluated using scanning electron microscopy and their releasing patterns were investigated. On investigating idophor releasing patterns of the microsphere, burst releasing pattern was detected until 2 days after in vitro incubation and sustained releasing was observed until 4 weeks. In field trial of teat dipping solution containing idophor loaded microspheres in dry cows showed significant preventive effects of intramammary infection caused by S. aureus, S. agalactiae, coagulase negative Staphylococci and coliform bacteria (p<0.05).

Cytoprotective effect of polysaccharide isolated from different mushrooms against 7-ketocholesterol induced damage in mouse liver cell line (BNL CL. 2)

  • Kim, Joo-Shin;Chung, Hau Yin;Na, Keun
    • Nutrition Research and Practice
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    • v.1 no.3
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    • pp.180-183
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    • 2007
  • Cytoprotective ability of polysaccharides isolated from different edible mushrooms was investigated on the 7-ketocholesterol-induced damaged cell line. Polysaccharide extracts from six different edible mushrooms-Flammulina velutipes, Peurotus ostreatus, Lentinus edodes, Agrocybe aegerita, Agaricus blazei, and Cordyceps militaris- were prepared by hot water extraction and alcohol precipitation. Cytoprotective ability was evaluated by measuring the viable cells of the normal embryonic liver cell line (BNL CL. 2) in the presence of 7-ketocholesterol. At $80\;{\mu}g/mL$ of 7-ketocholesterol, cytotoxicity was very high with a loss of 98% of viable cells after 20 h of incubation. With the addition of $200\;{\mu}g/mL$ of each polysaccharide isolate to the cell line containing $80\;{\mu}g/mL$ of 7-ketocholesterol, polysaccharide isolates from both Flammulina velutipes and Peurotus ostreatus could significantly inhibit the 7-ketochoelsterol-induced cytotoxicity in the cells. But other polysaccharide isolates were not effective in inhibiting cell damage caused by the oxLDL-induced cytotoxicity.

Cloning, Expression, and Purification of Exoinulinase from Bacillus sp. snu-7

  • Kim, Kyoung-Yun;Koo, Bong-Seong;Jo, Do-Hyun;Kim, Su-Il
    • Journal of Microbiology and Biotechnology
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    • v.14 no.2
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    • pp.344-349
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    • 2004
  • A gene encoding inulin-degrading enzyme of Bacillus sp. snu-7 with ORF of 1536 nucleotides was cloned. And it was overexpressed as His-tagged protein in E. coli BL21(DE3) pLysS using pRSET B vector containing mature enzyme sequence. Maximum enzyme production was achieved by IPTG (0.1 mM) induction at $OD_{600}$ 1.2 and $30^{\circ}C$ followed by 6 h incubation. The expressed protein purified through immobilized metal affinity chromatography showed molecular mass of 60 kDa on SDS-PAGE. Results of thin-layer chromatography using inulin as a substrate showed the enzyme to be an exotype inulinase capable of producing only monomeric fructose as a product. $K_m$ and $k_{cat}$, for the hydrolyses of inulin and sucrose were $2.28\pm0.08$ mM and 358.05$\pm$20.38 $min^{-l}$, and 22.02$\pm$0.41 mM and 4619.11$\pm$215.12 $$min^{-1}, respectively. Optimal activity of the exoinulinase occurred at pH 7.0 and $50^{\circ}C$.