• Title/Summary/Keyword: MTT method.

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SURFACE CHARACTERISTICS AND BIOLOGICAL RESPONSES OF HYDROXYAPATITE COATING ON TITANIUM BY HYDROTHERMAL METHOD: AN IN VITRO STUDY

  • Kim, Dong-Seok;Kim, Chang-Whe;Jang, Kyung-Soo;Lim, Young-Jun
    • The Journal of Korean Academy of Prosthodontics
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    • v.43 no.3
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    • pp.363-378
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    • 2005
  • Statement of problem. Hydroxyapatite(HA) coated titanium surfaces have not yet showed the reliable osseointegration in various conditions. Purpose. This study was aimed to investigate microstructures, chemical composition, and surface roughness of the surface coated by the hydrothermal method and to evaluate the effect of hydrothermal coating on the cell attachment, as well as cell proliferation. Material and Methods. Commercially pure(c.p.) titanium discs were used as substrates. The HA coating on c.p. titanium discs by hydrothermal method was performed in 0.12M HCl solution mixed with HA(group I) and 0.1M NaOH solution mixed with HA(group II). GroupⅠ was heated at 180 $^{\circ}C$ for 24, 48, and 72 hours. GroupⅡ was heated at 180 $^{\circ}C$ for 12, 24, and 36 hours. And the treated surfaces were evaluated by Scanning electron microscopy(SEM), Energy dispersive X-ray spectroscopy(EDS), X-ray photoelectron spectroscopy(XPS), X-ray diffraction method(XRD), Confocal laser scanning microscopy(CLSM). And SEM of fibroblast and 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT) assay were used for cellular responses of the treated surfaces. Results. The color of surface changed in both groups after the hydrothermal process. SEM images showed that coating pattern was homogeneous in group II, while inhomogeneous in group I. H72 had rosette-like precipitates. The crystalline structure grew gradually in group II, according to extending treatment period. The long needle-like crystals were prominent in N36. Calcium(Ca) and phosphorus(P) were not detected in H24 and H48 in EDS. In all specimens of group II and H72, Ca was found. Ca and P were identified in all treated groups through the analysis of XPS, but they were amorphous. Surface roughness did not increase in both groups after hydrothermal treatment. The values of surface roughness were not significantly different between groups I and II. According to the SEM images of fibroblasts, cell attachments were oriented and spread well in both treated groups, while they were not in the control group. However, no substantial amount of difference was found between groups I and II. Conclusions. In this study during the hydrothermal process procedure, coating characteristics, including the HA precipitates, crystal growth, and crystalline phases, were more satisfactory in NaOH treated group than in HCl treated group. Still, the biological responses of the modified surface by this method were not fully understood for the two tested groups did not differ significantly. Therefore, more continuous research on the relationship between the surface features and cellular responses seems to be in need.

Study on the Anticancer & Inhitory Effects of Somamsan (소암산의 항암효과 및 혈관신생억제에 미치는 영향)

  • Kim Yoong Su;Lee Seong Won;Choo Young Kug;Jung Kyu Yong;Ahn Seong Hun;Jeong Woo Yeal;Woo Won Hong
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.17 no.1
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    • pp.77-84
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    • 2003
  • Cancer, which is expressed in various forms, is one of the leading causes of human death, Soamsan (SAS) is composed of ten medicinal herb, the prescription was made according to the principles of Oriental traditional medicine based on the concept of synergic effects and interaction of among the components. SAS has been used for the cancer therapy, but the mechanism of it's effect is not well known. In the present study, the cytotoxic effect of the SAS water extract on cancer cell lines was investigated by the method of MTT in A549 cell lines and the anti-angiogenic effect was shown in the assay of chorioallantoic membrane (CAM) and in the cornea of rat administerd orally with SAS water extraction. The viability of A549 cell lines was not affected by the whole extract of SAS but the n-Hexan fraction of SAS water extract showed strong cytotoxicity which was not seemed to be done by the apoptotic mechanism. SAS water extract showed inhibition effects of angiogenesis induced in the cornea of rat and CAM assay. As the above results, it is suggested that SAS can be a candidate for new prescription for cancer therapy.

ACRYLAMIDE-INDUCED NEOPLASTIC TRANSFORMATION OF HUMAN EPITHELIAL CELLS IN CULTURE (Acrylamide가 인체상피세포의 발암화에 미치는 영향)

  • Kim, Sang-Kyu;Kim, Jin-Wook;Kim, Chin-Soo
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.34 no.6
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    • pp.602-610
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    • 2008
  • Purpose Acrylamide is present in significant quantities in a wide range of commonly consumed human foods. Carcinogenic risk of acrylamide through the consumption of food is a great public concern and in controversy, but it is not properly addressed due to the lack of evidence in humans. While a plenty of data is available on the carcinogenicity in animal models, the studies in humans are limited. Thus, the present study attempted to examine the carcinogenic potentials of acrylamide on the human epithelial cell, which is the target cell origin of the most cancers. Material and method & Result 1. Acrylamide was not cytotoxic up to $100{\mu}M$ as measured by MTT and LDH assays, indicating a relatively low toxicity of this substance in human epithelial cells. 2. The parameters of neoplastic cellular transformation such as cell saturation density, soft-agar colony formation and cell aggregation were analyzed to examine the carcinogenic potential of acrylamide. 3. The neoplastic transformation was further increased with the co-treatment of TPA 4. Antioxidants blocked the generation of Reactive Oxygen Species(ROS) and the GSH depleting agent dramatically increased the ROS production. 5. mRNA levels of fibronectin following acrylamide exposure was increased in a dose-dependent manner, indicating a possible biomarker of acrylamide-induced cellular transformation. Conclusion The present study will provide a valuable basis to compare the interspecies differences in response to carcinogenic potentials of acrylamide. The data on the interspecies differences are essential element in human risk assessment. Thus, our results obtained from the human epithelial cells will contribute to improving the risk assessment of human neoplasm including oral cancer.

Fabrication of Luminescent Hydroxyapatite Microspheres for Drug Loading and Release

  • Park, So Yeon;Kwon, Seung Lee;Baek, Mi Yeon;Yoo, Jin Sun;Kim, Min-Cheol;Jung, Hyun Suk
    • Proceedings of the Korean Vacuum Society Conference
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    • 2016.02a
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    • pp.360.1-360.1
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    • 2016
  • Hydroxyapatite (Ca10(PO4)6(OH)2, HAP) particles have attracted a great deal of attention in biomedical fields due to their good biocompatibility, bioactivity and fairly broad applications as drug delivery, dental implant, bone cement, and etc. Thus, many researchers have made an effort to add new functionalities such as luminescence, drug delivery, and bone regeneration properties up to HAP powders by controlling their nanostructure as well as composition. In this research, the mesoporous strontium substituted HAP (Sr-HAP) microspheres were synthesized using a hydrothermal method. In this synthesis, aspartic acid monomers were utilized to form microsphere by controlling surface energy of HAP particles and Sr ions were substituted into Ca ion sites, which induced luminescence property in HAP powders. Moreover, the change in the amount of Sr substitution was found to influence the particle size, morphology, and concurrently surface area, which led to changing drug loading as well as drug release property. The amount of Sr influences the morphology, luminescent properties, particle size, surface area cell viability and drug loading property, which are investigated by SEM, TEM, XRD, FTIR, BET, XPS and in vitro test such as MTT assay and drug release test. In particular, the multifunctional Sr-HAP with molar ratios of 0.25 (Sr/(Ca+Sr)) possessed the strongest luminescent property as well as the superior drug loading and sustained release properties that were correspondent with large surface area and pore size. Our study indicates that the fabricated multifunctional Sr-HAP microspheres are quite useful for bone regeneration and drug delivery.

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Targeting HSP90 Gene Expression with 17-DMAG Nanoparticles in Breast Cancer Cells

  • Mellatyar, Hassan;Talaei, Sona;Nejati-Koshki, Kazem;Akbarzadeh, Abolfazl
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.5
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    • pp.2453-2457
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    • 2016
  • Background: Dysregulation of HSP90 gene expression is known to take place in breast cancer. Here we used D,L-lactic-co-glycolic acid-polyethylene glycol-17-dimethylaminoethylamino-17-demethoxy geldanamycin (PLGA-PEG-17DMAG) complexes and free 17-DMAG to inhibit the expression of HSP90 gene in the T47D breast cancer cell line. The purpose was to determine whether nanoencapsulating 17DMAG improves the anti-cancer effects as compared to free 17DMAG. Materials and Methods: The T47D breast cancer cell line was grown in RPMI 1640 supplemented with 10% FBS. Encapsulation of 17DMAG was conducted through a double emulsion method and properties of copolymers were characterized by Fourier transform infrared spectroscopy and H nuclear magnetic resonance spectroscopy. Assessment of drug cytotoxicity was by MTT assay. After treatment of T47D cells with a given amount of drug, RNA was extracted and cDNA was synthesized. In order to assess HSP90 gene expression, real-time PCR was performed. Results: Taking into account drug load, IC50 was significant decreased in nanocapsulated 17DMAG in comparison with free 17DMAG. This finding was associated with decrease of HSP90 gene expression. Conclusions: PLGA-PEG-17DMAG complexes can be more effective than free 17DMAG in down-regulating of HSP90 expression, at the saesm time exerting more potent cytotoxic effects. Therefore, PLGA-PEG could be a superior carrier for this type of hydrophobic agent.

THE EFFECTS OF HONOKIOL AND MAGNOLOL ON THE ANTIMICROBIAL, BACTERIAL COLLAGENASE ACTIVITY, CYTOTOXICITY AND CYTOKINE PRODUCTION (Magnolol과 Honokiol이 항균, 교원질 분해효소, 세포독성 및 Cytokine생산에 미치는 영향)

  • Jang, Beom-Seok;Son, Seong-Heai;Chung, Chong-Pyoung;Bae, Ki-Hwan
    • Journal of Periodontal and Implant Science
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    • v.23 no.1
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    • pp.145-158
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    • 1993
  • The oral microbiota such as P. gingivalis, P. intermedia and A. actinomycetemcomitans play a primary role in the initiation and progression of the periodontal disease. The purpose of this study was to evaluate the antimicrobial effects and inhibitory effects of honokiol and magnolol on the bacterial collagenase activity, cytotoxicity and cytokine production of periodontopathic microorganisms. The antimicrobial activities of honokiol and magnolol was evaluted with minimum inhibition concentration. Honokiol was more active than magnolol, but less than chlorhexidine on antimicrobial activity. The inhibitory effects of magnolol and honokiol on the collagenolytic activity and cytotoxicity were evaluated using a Collagenokit CLN-100 and rapid colorimetric assay (MTT method) for cellular growth and survival of gingival fibroblast and periodontalligament cell and $[^3H]-thymidine$ incorporation for the gingival epithelial cell. The inhibitory effects on the collagenolytic activity was the highest in chlorhexidine, and the lowest in magnolol. Magnolol had the lowest cytotoxic effect and chlorhexidine had the highest. The inhibitory effects on cytokine production was evaluated using $interleukin-1{\beta}$ ELISA kit (Cistron Biotech.), IL-6, $TNF-{\alpha}$ ELISA kit (Genzyme) and inhibitory effects were higher than bacterial LPS and there is no difference among the honokiol, magnolol and chlorhexidine. From these results, the antimicrobial and antienzymatic activities of honokiol and magnolol were seemed to inhibit bacterial growth and enzyme activities with lesser cytotoxic activities. Therefore, it was suggested that honokiol and magnolol are very effective antimicrobial agents on periodontal pathogens.

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A Study of the Effect of Platelet-Rich Plasma on the Cellular Proliferation and Differentiation of Osteoblast Cell Line (혈소판농축혈장이 조골세포주의 세포증식 및 분화에 미치는 영향에 대한 연구)

  • Jung, Tae-Wook;Jang, Kyung-Soo;Kim, Chang-Whe;Kim, Yung-Soo
    • Journal of Dental Rehabilitation and Applied Science
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    • v.20 no.1
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    • pp.31-41
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    • 2004
  • The osseointegration in implant therapy is achieved following general wound healing mechanism. Platelet play a major role in wound healing process. In addition to blood clot formation, they secrete many growth factors which regulate the attachment, proliferation and differentiation of nearly all cell types. The use of these growth factors is now known to be very effective methods to improve the cellular activity. Platelet-rich plasma which is made with the newly developed technique concentrating platelets 3-folds or more is also proven to be very effective method to stimulate and accelerate the healing of bone and soft tissue. Previous study proved that platelet-rich plasma enhanced the cellular attachment by inducing fibronectin, vitronectin from osteoblast. So, this study was aimed to investigate the effect of platelet-rich plasma on the cellular proliferation and differentiation in vitro. The effect on the proliferation was evaluated by MTT assay. To evaluate autocrine and paracrine effect, conditioned medium was made and compared. By measuring alkaline phosphatase activity, the effect on the cellular differentiation was evaluated. The results were as following: The cellular proliferation of osteoblast cell line increased depending on the concentration of platelet-rich plasma and conditioned medium. The alkaline phosphatase activity increased depending on the concentration of platelet-rich plasma and conditioned medium. These findings imply that platelet-rich plasma enhance the cellular proliferation and differentiation and maximize the cellular activity by using the autocrine and paracrine effect.

Antioxidative and Antimutagenic Effects of Ganoderma lucidum Krast Extracts (영지버섯(Ganoderma lucidum Krast) 추출물의 항산화 및 항돌연변이 효과)

  • Oh Se-In;Lee Mee-Sook
    • The Korean Journal of Food And Nutrition
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    • v.18 no.1
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    • pp.54-62
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    • 2005
  • The antioxidative effect, antimutagenic capacity and inhibitory effect of cancer cell proliferation in ethanol extracts of Ganoderma lucidum were studied for suggestion of prevention and dietetic treatment of chronic diseases and development of antioxidative, antimutagenic and anticancer functional food by employing biological and biochemical assay. The IC/sub 50/ of MDA with BSA conjugation reaction, lipid peroxidation and scavenging effect on DPPH radical in ethanol extracts of Ganoderma lucidum showed 1026.21 mg/mL, 0.152 mg/mL and 0.412 mg/mL, respectively. So, the most effective antioxidative capacity in ethanol extracts of Ganoderma lucidum was the lipid peroxidation, among the method used this study. The indirect and direct antimutagenic effects of ethanol extracts of Ganoderma lucidum were examined by Ames test using Salmonella typimurium TA98 and TA100. The inhibition rates on indirect mutagenicity mediated by 2-Anthramine showed 100% in the Salmonella typimurium TA98 and 98.74% in the Salmonella typimurium TA100 and the direct mutagenicity mediated by sodium azide in Salmonella typimurium TA100 was 82.96%. But, the inhibitory effect on indirect mutagenicity mediated by 2-Nitrofluorene in Salmonella typimurium TA98 was low(7.81%). The inhibitory Effect of Ganoderma lucidum ethanol extracts on cell proliferation in Hela and MCF-7 by MTT test were 74.36% in HeLa and 73.90% in MCF-7 at the 0.50 mg/assay concentration and IC/sub 50/ were 0.163 mg/mL and 0.196 mg/mL respectively. From this result, it is suggested that Ganoderma lucidum is believed to have a possible antioxidative, antimutagenic and anticancer capacities.

The Effects of Bee Venom on PLA2, COX-2, iNOS, AA and PG in RAW 264.7 Cells (봉약침액(蜂藥鍼液)이 PLA2, COX-2, iNOS, AA 및 미치는 영향(影響)에 관(關)한 연구(硏究))

  • Ha, Seang-Jong;Lee, Seong-No;Jo, Hyun-Chul;Kim, Kee-Hyun
    • Journal of Pharmacopuncture
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    • v.5 no.2
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    • pp.40-51
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    • 2002
  • Objectives : The purpose of this study was to investigate the effect of Bee Venom on the lipopolysaccharide-induced expression phospholipase $A_2$, cyclooxygenase-2 and inducible nitrogen oxide synthase, and the generation of arachidonic acid, prostaglandin D2 and E2 in RAW 264.7 cells, a murine macrophage cell line. Methods : The expression of phospholipase $A_2$, cyclooxygenase and inducible nitrogen oxide synthase was determined by western blotting with corresponding antibodies, and the generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ was assayed by ELISA method in RAW 264.7 cells. The non-toxic concentrations (0.1 to $5\;{\mu}g/ml$) of bee venom determined by MTT assay, were used in this study. Results : 1. Bee venom inhibited lipopolysaccharide-induced expression of phospholipase $A_2$ in a dose dependent manner after 48 hours treatment. 2. Bee venom inhibited lipopolysaccharide-induced expression of cyclooxygenase-2 in a dose dependent manner after 24 and 48 hours treatment. 3. Bee venom inhibited lipopolysaccharide-induced expression of inducible nitrogen oxidesynthase in a dose dependent manner after 48 hours treatment. 4. The generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ was not much affected by the treatment of bee venom on the lipopolysaccharide-induced generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ in RAW 264.7 cells.

In Vitro and In Vivo Evaluation of Composite Scaffold of BCP, Bioglass and Gelatin for Bone Tissue Engineering

  • Kim, Woo Seok;Nath, Subrata Deb;Bae, Jun Sang;Padalhin, Andrew;Kim, Boram;Song, Myeong Jin;Min, Young Ki
    • Korean Journal of Materials Research
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    • v.24 no.6
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    • pp.310-318
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    • 2014
  • In this experiment, a highly porous scaffold of biphasic calcium phosphate (BCP) was prepared using the spongereplica method. The BCP scaffold was coated with 58S bioactive glass (BG) and sintered for a second time. The resulting scaffold was coated with gelatin (Gel) and cross-linked with [3-(3-dimethyl aminopropyl) carbodiimide] and N-Hydroxysuccinamide (EDC-NHS). The initial average pore size of the scaffold ranged from 300 to $700{\mu}m$, with more than 85 % porosity. The coating of BG and Gel had a significant effect on the scaffold-pore size, decreasing scaffold porosity while increasing mechanical strength. The material and surface properties were evaluated by means of several experiments involving scanning electron microscopy (SEM), energy-dispersive X-ray (EDX) and X-ray diffraction (XRD). Cytotoxicity was evaluated using MTT assay and confocal imaging of MC3T3-E1 pre-osteoblast cells cultured in vitro. Three types of scaffold (BCP, BCP-BG and BCP-BG-Gel) were implanted in a rat skull for in vivo evaluation. After 8 weeks of implantation, bone regeneration occurred in all three types of sample. Interestingly, regeneration was found to be greater (geometrically and physiologically) for neat BCP scaffolds than for two other kinds of composite scaffolds. However, the other two types of scaffolds were still better than the control (i.e., defect without treatment).