• Title/Summary/Keyword: Molecular approach

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Spinal Fusion Based on Ex Vivo Gene Therapy Using Recombinant Human BMP Adenoviruses (사람 골 형성 단백질 Ex vivo 유전자 치료법을 이용한 척추 유합)

  • Kim, Gi-Beom;Kim, Jae-Ryong;Ahn, Myun-Hwan;Seo, Jae-Sung
    • Journal of Yeungnam Medical Science
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    • v.24 no.2
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    • pp.262-274
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    • 2007
  • Purpose : Bone morphogenetic proteins (BMPs) play an important role in the formation of cartilage and bone, as well as regulating the growth of chondroblasts and osteoblasts. In this study, we investigated whether recombinant human BMP adenoviruses are available for ex vivo gene therapy, using human fibroblasts and human bone marrow stromal cells in an animal spinal fusion model. Materials and Methods : Human fibroblasts and human bone marrow stromal cells were transduced with recombinant BMP-2 adenovirus (AdBMP-2) or recombinant BMP-7 adenovirus (AdBMP-7), referred to as AdBMP-7/BMSC, AdBMP-2/BMSC, AdBMP-7/HuFb, and AdBMP-2/HuFb. We showed that each cell secreted active BMPs by alkaline phosphatase staining. Since AdBMP-2 or AdBMP-7 tranducing cells were injected into the paravertebral muscle of athymic nude mice, at 4 weeks and 7 weeks, we confirmed that new bone formation occurred by induction of spinal fusion on radiographs and histochemical staining. Results : In the region where the AdBMP-7/BMSC was injected, new bone formation was observed in all cases and spinal fusion was induced in two of these. AdBMP-2/BMSC induced bone formation and spinal fusion occurred among one of five. However, in the region where AdBMP/HuFb was injected, neither bone formation nor spinal fusion was observed. Conclusion : The osteoinductivity of AdBMP-7 was superior to that of AdBMP-2. In addition, the human bone marrow stromal cells were more efficient than the human fibroblasts for bone formation and spinal fusion. Therefore, the results of this study suggest that AdBMP-7/BMSC would be the most useful approach to ex vivo gene therapy for an animal spinal fusion model.

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Transduced PEP-1-AMPK inhibits the LPS-induced expression of COX-2 and iNOS in Raw264.7 cells

  • Shin, Min-Jea;Lee, Yeom-Pyo;Kim, Dae-Won;An, Jae-Jin;Jang, Sang-Ho;Cho, Sung-Min;Sheen, Seung-Hoon;Lee, Hae-Ran;Kweon, Hae-Yong;Kang, Seok-Woo;Lee, Kwang-Gill;Park, Jin-Seu;Eum, Won-Sik;Cho, Yong-Jun;Choi, Soo-Young
    • BMB Reports
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    • v.43 no.1
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    • pp.40-45
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    • 2010
  • AMP-activated protein kinase (AMPK) is a heterotrimeric enzyme that plays a central role in cellular metabolic stress. Modulation of nitric oxide (NO) and cyclooxygenase-2 (COX-2) is considered a promising approach for the treatment of inflammation and neuronal diseases. In this study, the AMPK gene was fused in-frame with PEP-1 peptide in a bacterial expression vector to produce a PEP-1-AMPK fusion protein. Expressed and purified PEP-1-AMPK fusion proteins were transduced efficiently into macrophage Raw 264.7 cells in a time- and dose-dependent manner. Furthermore, transduced PEP-1-AMPK fusion protein markedly inhibited LPS-induced iNOS and COX-2 expression. These results suggest that the PEP-1-AMPK fusion protein can be used for the protein therapy of COX-2 and NO-related disorders such as inflammation and neuronal diseases.

Integrative Study on PPARGC1A: Hypothalamic Expression of Ppargc1a in ob/ob Mice and Association between PPARGC1A and Obesity in Korean Population

  • Hong, Mee-Suk;Kim, Hye-Kyung;Shin, Dong-Hoon;Song, Dae-Kyu;Ban, Ju Yeon;Kim, Bum Shik;Chung, Joo-Ho
    • Molecular & Cellular Toxicology
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    • v.4 no.4
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    • pp.318-322
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    • 2008
  • Obesity is an increasing worldwide health problem that is strongly related to the imbalance of food intake and energy metabolism. It was well-known that several substances in the hypothalamus regulate food intake and energy metabolism. We planned an integrative study to elucidate the mechanism of the development of obesity. Firstly, to find candidate genes with the marvelous effect, the different expression in the hypothalamus between ob/ob and 48-h fasting mice was investigated by using DNA microarray technology. As a result, we found 3 genes [peroxisome proliferator activated receptor, gamma, coactivator 1 alpha (Ppargc1a), calmodulin 1 (Calm1), and complexin 2 (Cplx2)] showing the different hypothalamic expression between ob/ob and 48-h fasting mice. Secondly, a genetic approach on PPARGC1A gene was performed, because PPARGC1A acts as a transcriptional coactivator and a metabolic regulator. Two hundred forty three obese female patients with body mass index (BMI)${\geq}$25 and 285 control female subjects with BMI 18 to<23 were recruited according to the Classification of Korean Society for the Study of Obesity. Among the coding single nucleotide polymorphisms (cSNPs) of PPARGC1A, 2 missense SNPs (rs8192678, Gly482Ser; rs3736265, Thr612Met) and 1 synonymous SNP (rs3755863, Thr528Thr) were selected, and analyzed by PCR-RFLP and pyrosequencing. For the analysis of genetic data, chi-square ($X^2$) test and EH program were used. The rs8192678 was significantly associated with obese women (P<0.0006; odds ratio, 1.5327; 95% confidence interval, 1.2006-1.9568). Haplotypes also showed significant association with obese women ($X^2$=33.28, P<0.0008). These results suggest that PPARGC1A might be related to the development of obesity.

Comparative analysis of two methods of laser induced boron isotopes separation

  • K.A., Lyakhov;Lee, H.J.
    • Proceedings of the Korean Vacuum Society Conference
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    • 2011.02a
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    • pp.407-408
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    • 2011
  • Natural boron consists of two stable isotopes 10B and 11B with natural abundance of 18.8 atom percent of 10B and 81.2 atom percent of 11B. The thermal neutron absorption cross-section for 10B and 11B are 3837 barn and 0.005 barn respectively. 10B enriched specific compounds are used for control rods and as a reactor coolant additives. In this work 2 methods for boron enrichment were analysed: 1) Gas irradiation in static conditions. Dissociation occurs due to multiphoton absorption by specific isotopes in appropriately tuned laser field. IR shifted laser pulses are usually used in combination with increasing the laser intensity also improves selectivity up to some degree. In order to prevent recombination of dissociated molecules BCl3 is mixed with H2S 2) SILARC method. Advantages of this method: a) Gas cooling is helpful to split and shrink boron isotopes absorption bands. In order to achieve better selectivity BCl3 gas has to be substantially rarefied (~0.01%-5%) in mixture with carrier gas. b) Laser intensity is lower than in the first method. Some preliminary calculations of dissociation and recombination with carrier gas molecules energetics for both methods will be demonstrated Boron separation in SILARC method can be represented as multistage process: 1) Mixture of BCl3 with carrier gas is putted in reservoir 2) Gas overcooling due to expansion through Laval nozzle 3) IR multiphoton absorption by gas irradiated by specifically tuned laser field with subsequent gradual gas condensation in outlet chamber It is planned to develop software which includes these stages. This software will rely on the following available software based on quantum molecular dynamics in external quantized field: 1) WavePacket: Each particle is treated semiclassicaly based on Wigner transform method 2) Turbomole: It is based on local density methods like density of functional methods (DFT) and its improvement- coupled clusters approach (CC) to take into account quantum correlation. These models will be used to extract information concerning kinetic coefficients, and their dependence on applied external field. Information on radiative corrections to equation of state induced by laser field which take into account possible phase transition (or crossover?) can be also revealed. This mixed phase equation of state with quantum corrections will be further used in hydrodynamical simulations. Moreover results of these hydrodynamical simulations can be compared with results of CFD calculations. The first reasonable question to ask before starting the CFD simulations is whether turbulent effects are significant or not, and how to model turbulence? The questions of laser beam parameters and outlet chamber geometry which are most optimal to make all gas volume irradiated is also discussed. Relationship between enrichment factor and stagnation pressure and temperature based on experimental data is also reported.

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Bacterial Diversity and its Phylogenetic Analysis in Lake Sapgyo (삽교호의 세균 다양성과 계통분류학적 분석)

  • Kim, Myeong;Jeon, Eun-Hyeong;An, Tae-Yeong
    • Korean Journal of Microbiology
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    • v.39 no.4
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    • pp.272-276
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    • 2003
  • Sapgyo Lake is an artificial freshwater reservoir which is located to the midwest of Korea and is the main water reservoir for industry and agriculture of the region. In this study we investigated environmental factors and the change of bacterial community with the influence of surrounding inflow water and the seasonal variation using the molecular ecological approach. Water samples were collected at front of the dike in May and August, 2001. Bacterial genomic DNAs were extracted directly and purified for the amplification of bacterial 16S rDNA. Clone libraries of the 16S rDNA were constructed using pGEM-T easy vector and RFLP analysis was performed to make a group as OTUs with 4 base recognizing enzymes (MspI and HaeIII). The estimated values of richness in August sample was higher than in May. Thirty-three of 153 clones in May and thirty-eight of 131 clones in August were sequenced from forward region of bacterial 16S rDNA for about 600~800 bp. Proteobacteria, Cytophaga, gram positive bacteria and Verrucomicrobia were observed both months. Especially, Planctomyces, cyanobacteria and chloroplast appeared in August when algal bloom occurred. On the whole investigation, Sapgyo lake showed a typical community structure of estuarine and was influenced by heterochthonous organic matters from the surrounding stream.

Surface-modified Nanoparticle Additives for Wear Resistant Water-based Coatings for Galvanized Steel Plates

  • Becker-Willinger, Carsten;Heppe, Gisela;Opsoelder, Michael;Veith, H.C. Michael;Cho, Jae-Dong;Lee, Jae-Ryung
    • Corrosion Science and Technology
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    • v.9 no.4
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    • pp.147-152
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    • 2010
  • Conventional paints for conversion coating applications in steel production derived mainly from water-based polymer dispersions containing several additives actually show good general performance, but suffer from poor scratch and abrasion resistance during use. The reason for this is because the relatively soft organic binder matrix dominates the mechanical surface properties. In order to maintain the high quality and decorative function of coated steel sheets, the mechanical performance of the surface needs to be improved significantly. In fact the wear resistance should be enhanced without affecting the optical appearance of the coatings by using appropriate nanoparticulate additives. In this direction, nanocomposite coating compositions (Nanomer$^{(R)}$) have been derived from water-based polymer dispersions with an increasing amount of surface-modified nanoparticles in aqueous dispersion in order to monitor the effect of degree of filling with rigid nanoparticles. The surface of nanoparticles has been modified for optimum compatibility with the polymer matrix in order to achieve homogeneous nanoparticle dispersion over the matrix. This approach has been extended in such a way that a more expanded hybrid network has been condensed on the nanoparticle surface by a hydrolytic condensation reaction in addition to the quasi-monolayer type small molecular surface modification. It was expected that this additional modification will lead to more intensive cross-linking in coating systems resulting in further improved scratch-resistance compared to simple addition of nanoparticles with quasi-monolayer surface modification. The resulting compositions have been coated on zinc-galvanized steel and cured. The wear resistance and the corrosion protection of the modified coating systems have been tested in dependence on the compositional change, the type of surface modification as well as the mixing conditions with different shear forces. It has been found out that for loading levels up to 50 wt.-% nanoparticles, the mechanical wear resistance remains almost unaffected compared to the unmodified resin. In addition, the corrosion resistance remained unaffected even after $180^{\circ}$ bending test showing that the flexibility of coating was not decreased by nanoparticle addition. Electron microscopy showed that the inorganic nanoparticles do not penetrate into the organic resin droplets during the mixing process but rather formed agglomerates outside the polymer droplet phase resulting in quite moderate cross linking while curing, because of viscosity. The proposed mechanisms of composite formation and cross linking could explain the poor effect regarding improvement of mechanical wear resistance and help to set up new synthesis strategies for improved nanocomposite morphologies, which should provide increased wear resistance.

Design Sensitivity Analysis of Coupled MD-Continuum Systems Using Bridging Scale Approach (브리징 스케일 기법을 이용한 분자동역학-연속체 연성 시스템의 설계민감도 해석)

  • Cha, Song-Hyun;Ha, Seung-Hyun;Cho, Seonho
    • Journal of the Computational Structural Engineering Institute of Korea
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    • v.27 no.3
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    • pp.137-145
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    • 2014
  • We present a design sensitivity analysis(DSA) method for multiscale problems based on bridging scale decomposition. In this paper, we utilize a bridging scale method for the coupled system analysis. Since the analysis of full MD systems requires huge amount of computational costs, a coupled system of MD-level and continuum-level simulation is usually preferred. The information exchange between the MD and continuum levels is taken place at the MD-continuum boundary. In the bridging scale method, a generalized Langevin equation(GLE) is introduced for the reduced MD system and the GLE force using a time history kernel is applied at the boundary atoms in the MD system. Therefore, we can separately analyze the MD and continuum level simulations, which can accelerate the computing process. Once the simulation of coupled problems is successful, the need for the DSA is naturally arising for the optimization of macro-scale design, where the macro scale performance of the system is maximized considering the micro scale effects. The finite difference sensitivity is impractical for the gradient based optimization of large scale problems due to the restriction of computing costs but the analytical sensitivity for the coupled system is always accurate. In this study, we derive the analytical design sensitivity to verify the accuracy and applicability to the design optimization of the coupled system.

Transfection of Mesenchymal Stem Cells with the FGF-2 Gene Improves Their Survival Under Hypoxic Conditions

  • Song, Heesang;Kwon, Kihwan;Lim, Soyeon;Kang, Seok-Min;Ko, Young-Guk;Xu, ZhengZhe;Chung, Ji Hyung;Kim, Byung-Soo;Lee, Hakbae;Joung, Boyoung;Park, Sungha;Choi, Donghoon;Jang, Yangsoo;Chung, Nam-Sik;Yoo, Kyung-Jong;Hwang, Ki-Chul
    • Molecules and Cells
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    • v.19 no.3
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    • pp.402-407
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    • 2005
  • Bone marrow mesenchymal stem cells (MSCs) have shown potential for cardiac repair following myocardial injury, but this approach is limited by their poor viability after transplantation. To reduce cell loss after transplantation, we introduced the fibroblast growth factor-2 (FGF-2) gene ex vivo before transplantation. The isolated MSCs produced colonies with a fibroblast-like morphology in 2 weeks; over 95% expressed CD71, and 28% expressed the cardiomyocyte-specific transcription factor, Nkx2.5, as well as ${\alpha}$-skeletal actin, Nkx2.5, and GATA4. In hypoxic culture, the FGF-2-transfected MSCs (FGF-2-MSCs) secreted increased levels of FGF-2 and displayed a threefold increase in viability, as well as increased expression of the anti-apoptotic gene, Bcl2, and reduced DNA laddering. They had functional adrenergic receptors, like cardiomyocytes, and exposure to norepinephrine led to phosphorylation of ERK1/2. Viable cells persisted 4 weeks after implantation of $5.0{\times}10^5$ FGF-2-MSCs into infarcted myocardia. Expression of cardiac troponin T (CTn T) and a voltage-gated $Ca^{2+}$ channel (CaV2.1) increased, and new blood vessels formed. These data suggest that genetic modification of MSCs before transplantation could be useful for treating myocardial infarction and end-stage cardiac failure.

A Study on the Speciadon of a Fresh Water Fish Zacco temminckL VII. Vadation of Mitochondrial DNA between 2 Types of Zacco temmincki (갈겨니(Zacco temminki)의 진화에 관한 연구 VII.갈겨니 2 Type의 Mitochondrial DNA변이)

  • Lee, Hei-Yung;Yang, Suh-Yung;Paik, Sang-Gi;Park, Chang-Shin;Yu, Sung-Lim;Lee, Sung-Keun
    • The Korean Journal of Zoology
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    • v.31 no.3
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    • pp.236-242
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    • 1988
  • Mitochondrial DNAs of two Mdh allelotypes of the dark chub, Z. temmincki inhabiting in Korean fresh water, were analysed. Samples of each type were collected from four populations, and the fragment patterns for mtdNA of each type were explained from 7 of the eleven restriction enzymes with hexanucleotide recognition site. Genome size was approximately 16.7 kilobases. The highly typical mtdNA fragments of each type were discovered in digestion profiles produced by Eco RI and Pst I enzyrnes. The comparisons of restriction fragment patterns and relative digestion maps permitted the estimation of fragment homology (F) and nucleotide sequence divergence(p). Between the two identical types, sequence divergence(p) was 0.128(MS), and 0.045(MM), ; between the two different types, 0.195 (range 0.177-0.226). These result may provide a distinct difference more than the value derived from allozyrne analysis, and a powerful new molecular approach for assessing genetic-evolutionary relationship among fishes.

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Enhanced Transdermal Delivery of Drug Compounds Using Scalable and Deformable Ethosomes (에토좀 입자크기와 멤브레인 특성 조절을 통한 약물의 경피흡수능 향상)

  • An, Eun-Jung;Shim, Jong-Won;Choi, Jang-Won;Kim, Jin-Woong;Park, Won-Seok;Kim, Han-Kon;Park, Ki-Dong;Han, Sung-Sik
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.36 no.2
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    • pp.105-113
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    • 2010
  • This study introduces a flexible approach to enhance skin permeation by using ethosomes with deformable lipid membranes as well as controllable sizes. To demonstrate this, a set of ethosomes encapsulating an anti-hair loss ingredient, Triaminodil$^{TM}$, as a model drug, were fabricated with varying their size, which was achieved by solely applying the different level of mechanical energy, while maintaining their chemical composition. After characterization of the ethosomes with dynamic light scattering, transmission electron microscopy, and deformability measurements, it was found that their membrane deformability depended on the particle size. Moreover, studies on in vitro skin permeation and murine anagen induction allowed us to figure out that the membrane deformability of ethosomes essentially affects delivery efficiency of Triaminodil$^{TM}$ through the skin. It was noticeable in our study that there existed an optimum particle size that can not only maximize the delivery of the drug through the skin, but also increase its actual dermatological activity. These findings offer a useful basis for understanding how ethosomes should be designed to improve delivery efficiency of encapsulated drugs therein in the aspects of changing their length scales and membrane properties.