• Title/Summary/Keyword: MTT method.

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The Effect of the Moutan Radicis Cortex on Expression of CD81 and GFAP in Injured Astrocyte (목단피(牧丹皮)가 손상된 성상신경세포의 CD81 및 GFAP의 발현에 미치는 영향)

  • Moon, Sung-Jin;Seon, Ki-Moon;Lim, Jin-Young;Song, Bong-Keun
    • The Journal of Internal Korean Medicine
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    • v.30 no.1
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    • pp.24-35
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    • 2009
  • Object : In conditions of brain infarction, irreversible axon damage occurs in the central nerve system (CNS), because gliosis makes physical and mechanical barriers. If gliosis formation could be suppressed, irreversible axon damage would be reduced. This could mean that an injured CNS could be regenerated. CD81 and GFAP have close relationships to gliosis. The increase in glial cells at CNS injury gives rise to the expression of CD81 and GFAP. CD81 was postulated to play a central role in the process of CNS scar formation. Method : In this study, the author investigated the effect of the water extract of the Moutan Radicis Cortex on regulation of CD81 and GFAP expression in injured CNS cells. MTT assay was used to examine cell viability, while RT-PCR and ELISA methods were carried out to measure the expression of CD81 and GFAP in the astrocyte. Results : We observed that water extract of the Moutan Radicis Cortex increased cell viability under hypoxia induced by $CoCl_2$ and suppressed the expression of CD81 and GFAP up-regulated by hypoxia. Conclusion : These results suggest that the Moutan Redicis Cortex could promote neural regeneration as a consequence of protecting CNS cells from hypoxia and suppressing the reactive gliosis following CNS injury.

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Purification of Total Ginsesides with Macroporous Resins and Their Biological Activities

  • Li, Huayue;Jin, Haizhu;Lee, Dong-Geun;Lee, Jae-Hwa;Lee, Sang-Hyeon;Ha, Bae-Jin;Ha, Jong-Myung
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.20 no.5
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    • pp.1321-1326
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    • 2006
  • Total ginsenosides were purified and their antioxidant, antibacterial and anticancer activities were measured. The crude extracts of ginseng, which were extracted with 75% ethanol by ultrasonification method, were firstly purified on AB-8 macroporous adsorption column to remove water soluble impurities, and decolored on Amberlite IRA 900 Cl anion-exchange column. Then, they were purified on Amberlite XAD16 adsorption column to delete the non-polar impurities. Total ginsenosides contents of the purified extracts were 79.4%, 71.7% and 72.5% in cultured wild ginseng, red ginseng and white ginseng, which were significantly increased than those of crude extracts. All of the three extracts showed concentration-dependant scavenging activities against DPPH radicals, among which white ginseng showed the most powerful activity. Cultured wild ginseng roots showed strongest effect against both B. subtilis PM 125(Gram-positive) and E. coli D31 (Gram-negative) bacteria, while red ginseng and white ginseng only showed the activity against B. subtilis. According to the result of the MTT assay, ail of the three extracts inhibited the growth of U-937 human hohistiocytic lympma cell, which were significantly different (p < 0.05) when compared to the control.

In Vitro Antioxidant and Repaired Effects of the Extracts from Pueraria thunbergiana Roots (갈근 추출물의 항산화와 수복효과에 관한 연구)

  • Shin, Sun-Woo;Shin, Joung-Mi;Yook, Chan-Nam;Na, Young-Soon;Baek, Seung-Hwa
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.20 no.4
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    • pp.909-913
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    • 2006
  • The antioxidative activity was observed in the ethyl acetate extract $(IC_{50},\;119{\pm}0.16\;{\mu}g/mL)$ from Pueraria thunbergiana. The DPPH radical scavenging effect of this extract was comparable with that of synthetic antioxdant, BHA $(IC_{50},\;88.39{\pm}1.1\;{\mu}g/mL){\times}\;10^{-3}\;{\mu}M$. Pueraria thunbergiana extracts against microorganisms were evaluated in terms of the minimum inhibitory concentrations (MIC). In general, C. albicans was stronger antimicrobial activity than the other microorganisms. in vitro the antitoxic activity of extracts of Pueraria thunbergiana on NIH 373 fibroblasts was evaluated by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide) method. Generally, detoxification effect by the extracts of Pueraria thunbergiana increased in proportion to the concentrations. When $10\;{\mu}g/mL$ of the hexane extract of Pueraria thunbergiana was treated it showed the highest antitoxic effects (p<0.01). These results suggest that the crude extract of P. thunbergiana has a potential therapeutic activity.

Astaxanthin induces migration in human skin keratinocytes via Rac1 activation and RhoA inhibition

  • Ritto, Dakanda;Tanasawet, Supita;Singkhorn, Sawana;Klaypradit, Wanwimol;Hutamekalin, Pilaiwanwadee;Tipmanee, Varomyalin;Sukketsiri, Wanida
    • Nutrition Research and Practice
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    • v.11 no.4
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    • pp.275-280
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    • 2017
  • BACKGROUND/OBJECTIVES: Re-epithelialization has an important role in skin wound healing. Astaxanthin (ASX), a carotenoid found in crustaceans including shrimp, crab, and salmon, has been widely used for skin protection. Therefore, we investigated the effects of ASX on proliferation and migration of human skin keratinocyte cells and explored the mechanism associated with that migration. MATERIAL/METHOD: HaCaT keratinocyte cells were exposed to $0.25-1{\mu}g/mL$ of ASX. Proliferation of keratinocytes was analyzed by using MTT assays and flow cytometry. Keratinocyte migration was determined by using a scratch wound-healing assay. A mechanism for regulation of migration was explored via immunocytochemistry and western blot analysis. RESULTS: Our results suggest that ASX produces no significant toxicity in human keratinocyte cells. Cell-cycle analysis on ASX-treated keratinocytes demonstrated a significant increase in keratinocyte cell proliferation at the S phase. In addition, ASX increased keratinocyte motility across the wound space in a time-dependent manner. The mechanism by which ASX increased keratinocyte migration was associated with induction of filopodia and formation of lamellipodia, as well as with increased Cdc42 and Rac1 activation and decreased RhoA activation. CONCLUSIONS: ASX stimulates the migration of keratinocytes through Cdc42, Rac1 activation and RhoA inhibition. ASX has a positive role in the re-epithelialization of wounds. Our results may encourage further in vivo and clinical study into the development of ASX as a potential agent for wound repair.

The Immunomodulatory Activity of Mori folium, the Leaf of Morus alba L., in RAW 264.7 Macrophages in Vitro

  • Kwon, Da Hye;Cheon, Ji Min;Choi, Eun-Ok;Jeong, Jin Woo;Lee, Ki Won;Kim, Ki Young;Kim, Sung Goo;Kim, Suhkmann;Hong, Su Hyun;Park, Cheol;Hwang, Hye-Jin;Choi, Yung Hyun
    • Journal of Cancer Prevention
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    • v.21 no.3
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    • pp.144-151
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    • 2016
  • Background: Immunoregulatory elements have emerged as useful immunotherapeutic agents against cancer. In traditional medicine, Mori folium, the leaf of Morus alba L. (Moraceae), has been used for various medicinal purposes; however, the immunomodulatory effects have not been fully identified. We evaluated the immunoenhancing potential of water extract of Mori folium (WEMF) in murine RAW264.7 macrophages. Methods: RAW264.7 cells were treated with WEMF for 24 hours and cell viability was detected by an MTT method. Nitric oxide (NO) levels in the culture supernatants were assayed using Griess reagent. The productions of prostaglandin $E_2$ ($PGE_2$) and immune-related cytokines was measured using ELISA detection kits. The mRNA and protein expression levels of Inducible NO synthase, COX-2, and cytokines were assayed by reverse transcription-PCR and Western blotting, respectively. The effect of WEMF on phagocytic activity was measured using a Phagocytosis Assay Kit. Results: WEMF significantly stimulated the production of NO and $PGE_2$ as immune response parameters at noncytotoxic concentrations, which was associated with the increased expression of inducible NO synthase and COX-2. The release and expression of cytokines, such as $TNF-{\alpha}$, interleukin $(IL)-1{\beta}$, IL-6, and IL-10, were also significantly increased in response to treatment with WEMF. Moreover, WEMF promoted the macrophagic differentiation of RAW264.7 cells and the resulting phagocytosis activity. Conclusions: WEMF has the potential to modulate the immune function by regulating immunological parameters. Further studies are needed to identify the active compounds and to support the use of WEMF as an immune stimulant.

Apoptotic Effect of ARTEMISIAE CAPILLARIS HERBA and COPTIDIS RHIZOMA Extracts on MIA PaCa-2 Cells (인진과 황련 추출물의 췌장암 세포주 MIA PaCa-2에 대한 세포사멸 효과)

  • Joo, Hyun-A;Bae, Hyeon-Jin;Hwang, Chung-Yeon
    • The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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    • v.27 no.4
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    • pp.158-176
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    • 2014
  • Purpose : The purpose of this study is to investigate the effect of ARTEMISIAE CAPILLARIS HERBA and COPTIDIS RHIZOMA Extracts on cell death in pancreatic cancer cells. Method : Human-derived pancreatic cancer cell line, MIA PaCa-2 cells were treated by Prescription A with various concentrations and the cytotoxicity was determined by MTT assay. To investigate the effects of Prescription A on pancreatic cancer cells, the staining of Annexin V/PI, cell cycle arrest, nuclear chromatin condensation and the production of reactive oxygen species (ROS) were examined. The effect of Prescription A's effective components, ARTEMISIAE CAPILLARIS HERBA and COPTIDIS RHIZOMA Extracts on cell death were also observed. Results : The viability of MIA PaCa-2 cells treated with Prescription A were decreased in a dose dependent manner. Prescription A induced cell death in MIA PaCa-2 cells as shown by result of Annexin V/PI double staining, chromatin condensation and cell cycle arrest. In addition, production of ROS was increased by Prescription A treatment, suggesting that ROS induced by Prescription A mediated cell death. Furthermore, Prescription A's effective components, ARTEMISIAE CAPILLARIS HERBA and COPTIDIS RHIZOMA Extracts were also induced apoptosis of MIA PaCa-2 cells through ROS production. Conclusion : These results suggest that Prescription A's effective components, ARTEMISIAE CAPILLARIS HERBA and COPTIDIS RHIZOMA Extracts induced death of MIA PaCa-2 through ROS production.

Antioxidant and Neuronal cell protective effects of Stachys sieboldii Miq. according to drying methods (건조방법에 따른 초석잠(Stachys sieboldii Miq.)의 항산화 활성 및 신경세포 보호효과)

  • Oh, Hee-Kyung
    • Journal of the Korean Applied Science and Technology
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    • v.35 no.4
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    • pp.1320-1326
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    • 2018
  • This study investigated the antioxidant and neuronal cell protective effects of activity of water and 70% ethanol extracts from Stachys sieboldii Miq. according to different drying method(hot air drying and freeze-drying). The total flavonoid and total polyphenol content in water extracts was significantly higher after freeze-drying compared to hot air drying(p<0.05). DPPH and ABTS radical scavenging were increased in a dose-dependent manner. The DPPH and ABTS radical scavenging activity of water extract were significantly higher after free-drying compared to hot air drying(p<0.05). In a cell viability using MTT, the water extract according to hot air drying and freeze-drying of Stachys sieboldii Miq. showed protective effect against H2O2-induced nurotoxicity. The results suggest that the water extracts of Stachys sieboldii Miq. after freeze-drying has antioxidant activities and may be useful for neurodegenerative disorders.

Anti-inflammatory and Anti-gastritis Effect of Sogunjung-tang Mixed Soft Extract (소건중탕 혼합단미연조엑스의 항염증 및 항위염 효과)

  • Kim, Myoung-Jin;Choi, Hye-Min;Nam, Won-Hee;Kim, Se-Jin;Son, Su-Mi;Kim, Jung-Ok
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.35 no.6
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    • pp.235-241
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    • 2021
  • Sogunjung-tang (SGJT) is traditional herbal prescriptions used to treat abdominal pain. In this study, we evaluated the effect on inflammation and gastritis through SGJT formulation development. SGJT composition herbal medicine was boiled in water at 95~100℃ for 3 hours and then concentrated. Sogunjung-tang mix soft extract (STM) was prepared using pharmaceutical excipients such as purified water, sodium benzoate, β-cyclodextrin and CMC-Na. Anti-inflammatory experiment was conducted using STM and lipopolysaccharide (LPS) in RAW 264.7 cells. Cell survival was measured by MTT method. Nitric oxide (NO) was measured using griess reagents, and pro inflammatory cytokines were measured by enzyme-linked immunosorbent assay and RT-PCR. Also, we verified STM validity in acute gastritis using the ICR mouse. STM was administered oral for three days. And 150 mM HCl in 60% ethanol was oral administered 0.5 mL one hour after the last drug administration. Mice were sacrificed 1 hour after 150 mM HCl in 60% ethanol administration. The gastric mucosa was visually observed. STM were not toxic in RAW 264.7 cells. Treatment of STM inhibited the production of NO and inflammatory cytokine at the protein and mRNA levels. Also, in the acute gastritis model with the mouse, the treatment of STM improved gastric mucosal bleeding and edema. In summary, it was confirmed that the treatment of STM exerts anti-inflammatory and anti-gastritis effects. Therefore, we suggest that STM may provide a preclinical evidence for the prevention and treatment of inflammatory and gastritis diseases.

Physicochemical, Antibacterial Properties, and Compatibility of ZnO-NP/Chitosan/β-Glycerophosphate Composite Hydrogels

  • Huang, Pingping;Su, Wen;Han, Rui;Lin, Hao;Yang, Jing;Xu, Libin;Ma, Lei
    • Journal of Microbiology and Biotechnology
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    • v.32 no.4
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    • pp.522-530
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    • 2022
  • In this study we aimed to develop novel ZnO-NP/chitosan/β-glycerophosphate (ZnO-NP/CS/β-GP) antibacterial hydrogels for biomedical applications. According to the mass fraction ratio of ZnO-NPs to chitosan, mixtures of 1, 3, and 5% ZnO-NPs/CS/β-GP were prepared. Using the test-tube inversion method, scanning electron microscopy and Fourier-transform infrared spectroscopy, the influence of ZnO-NPs on gelation time, chemical composition, and cross-sectional microstructures were evaluated. Adding ZnO-NPs significantly improved the hydrogel's antibacterial activity as determined by bacteriostatic zone and colony counting. The hydrogel's bacteriostatic mechanism was investigated using live/dead fluorescent staining and scanning electron microscopy. In addition, crystal violet staining and MTT assay demonstrated that ZnO-NPs/CS/β-GP exhibited good antibacterial activity in inhibiting the formation of biofilms and eradicating existing biofilms. CCK-8 and live/dead cell staining methods revealed that the cell viability of gingival fibroblasts (L929) cocultured with hydrogel in each group was above 90% after 24, 48, and 72 h. These results suggest that ZnO-NPs improve the temperature sensitivity and bacteriostatic performance of chitosan/β-glycerophosphate (CS/β-GP), which could be injected into the periodontal pocket in solution form and quickly transformed into hydrogel adhesion on the gingiva, allowing for a straightforward and convenient procedure. In conclusion, ZnO-NP/CS/β-GP thermosensitive hydrogels could be expected to be utilized as adjuvant drugs for clinical prevention and treatment of peri-implant inflammation.

Inhibition of Pathogenic Bacteria and Fungi by Natural Phenoxazinone from Octopus Ommochrome Pigments

  • Lewis-Lujan, Lidianys Maria;Rosas-Burgos, Ema Carina;Ezquerra-Brauer, Josafat Marina;Burboa-Zazueta, Maria Guadalupe;Assanga, Simon Bernard Iloki;del Castillo-Castro, Teresa;Penton, Giselle;Plascencia-Jatomea, Maribel
    • Journal of Microbiology and Biotechnology
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    • v.32 no.8
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    • pp.989-1002
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    • 2022
  • Cephalopods, in particular octopus (Octopus vulgaris), have the ability to alter their appearance or body pattern by showing a wide range of camouflage by virtue of their chromatophores, which contain nanostructured granules of ommochrome pigments. Recently, the antioxidant and antimicrobial activities of ommochromes have become of great interest; therefore, in this study, the pH-dependent redox effect of the extraction solvent on the antioxidant potential and the structural characterization of the pigments were evaluated. Cell viability was determined by the microdilution method in broth by turbidity, MTT, resazurin, as well as fluorescence microscopy kit assays. A Live/Dead Double Staining Kit and an ROS Kit were used to elucidate the possible inhibitory mechanisms of ommochromes against bacterial and fungal strains. The results obtained revealed that the redox state alters the color changes of the ommochromes and is dependent on the pH in the extraction solvent. Natural phenoxazinone (ommochromes) is moderately toxic to the pathogens Staphylococcus aureus, Bacillus subtilis, Salmonella Typhimurium and Candida albicans, while the species Pseudomonas aeruginosa and Pseudomonas fluorescens, and the filamentous fungi Aspergillus parasiticus, Alternaria spp. and Fusarium verticillioides, were tolerant to these pigments. UV/visible spectral scanning and Fourier- transform infrared spectroscopy (FTIR) suggest the presence of reduced ommatin in methanol/ HCl extract with high intrinsic fluorescence.