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Hexene-functionalized graphitic nanoplatelets (He- f-GN) were easily prepared using a mechanochemical reaction between solid graphite and liquid 1-hexene. The He- f-GN exhibited outstanding properties (e.g., high specific surface area, high crystallinity and so on) and could be well distributed in various solvents including formic acid. The He- f-GN/Nylon 6_X nanocomposites were simply prepared using the solution method, and showed excellent mechanical properties and thermal stability compared with the neat Nylon 6. Specifically, the tensile strength and Young’s modulus of the He- f-GN/Nylon 6_1 nanocomposites increased by approximately 32.5% and 33.7%, respectively, compared to the neat Nylon 6 due to the special properties of the He- f-GN and its excellent compatibility with Nylon 6 chains. The He- f-GN also acts as nucleation sites, increasing the crystallinity of Nylon 6, and generated hydrogen bonds with the amide groups of the Nylon 6. The new filler, He- f-GN, provides an effective way to increase the performance of polymer, demonstrating good application prospects.

Although polymers are very important and vastly used materials, their physical properties are limited. Therefore, they are reinforced with fillers to relieve diverse restrictions and expand their application areas. The exceptional properties of graphene make it an interesting material with huge potential for application in various industries and devices. The interfacial interaction between graphene and the polymer matrix improved the uniform graphene dispersion in the polymer matrix, enhancing the general nanocomposite performance. Therefore, graphene functionalization is essential to enhance the interfacial interaction, maintain excellent properties, and obstruct graphene agglomeration. Many studies have reported that graphene/polymer nanocomposites have exceptional properties that enable diverse applications. The use of graphene/polymer nanocomposites is expected to increase sustainably and to transform from a basic to an advanced material to offer optimum solutions to industry and consumers.

In robust optimization, an optimum solution of a system is obtained when some uncertainties exist in the system. The uncertainty can be defined by probabilistic characteristics or deterministic intervals (uncertainty ranges or tolerances) that are the main concern in this study. An insensitive objective function is obtained with regard to the uncertainties or the worst case is considered for the objective function within the intervals in robust optimization. A supreme value within the uncertainty interval is minimized. The worst case approach has been extensively utilized in the linear programming (LP) community. However, the method solved only small scale problems of structural optimization where nonlinear programming (NLP) is employed. In this research, a novel worst case approach is proposed to solve large scale problems of structural optimization. An uncertainty interval is defined by a tolerance range of a design variable or problem parameter. A supreme value is obtained by optimization of the objective function subject to the intervals, and this process yields an inner loop. The supremum is minimized in the outer loop. Linearization of the inner loop is proposed to save the computational time for optimization. This technique can be easily extended for constraints with uncertainty intervals because the worst case of a constraint should be satisfied. The optimum sensitivity is utilized for the sensitivity of a supremum in the outer loop. Three examples including a mathematical example and two structural applications are presented to validate the proposed idea.

The pharmacological effects of Rosa hybrida are well known in the cosmetics industry. However, the role of Rosa hybrida in cardiovascular biology had not previously been investigated, to the best of our knowledge. The aim of the present study was to elucidate the effect of water extract of Rosa hybrida (WERH) on platelet‑derived growth factor (PDGF)-stimulated vascular smooth muscle cells (VSMCs). VSMC proliferation, which was stimulated by PDGF, was inhibited in a non-toxic manner by WERH treatment, which also diminished the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and AKT. Treatment with WERH also induced G1-phase cell cycle arrest, which was due to the decreased expression of cyclins and cyclin-dependent kinases (CDKs), and induced p21WAF1 expression in PDGF-stimulated VSMCs. Moreover, WERH treatment suppressed the migration and invasion of VSMCs stimulated with PDGF. Treatment with WERH abolished the expression of matrix metalloproteinase-9 (MMP-9) and decreased the binding activity of nuclear factor-κB (NF-κB), activator protein-1 (AP-1), and specificity protein 1 (Sp1) motifs in PDGF-stimulated VSMCs. WERH treatment inhibited the proliferation of PDGF‑stimulated VSMCs through p21WAF1‑mediated G1-phase cell cycle arrest, by decreasing the kinase activity of cyclin/CDK complexes. Furthermore, WERH suppressed the PDGF-induced phosphorylation of ERK1/2 and AKT in VSMCs. Finally, treatment with WERH impeded the migration and invasion of VSMCs stimulated by PDGF by downregulating MMP-9 expression and a reduction in NF-κB, AP-1 and Sp1 activity. These results provide new insights into the effects of WERH on PDGF-stimulated VSMCs, and we suggest that WERH has the potential to act as a novel agent for the prevention and/or treatment of vascular diseases.

To accommodate growing demands on either heavy steel structures or unique buildings with irregular configurations, built-up wide-flange steel (BWS) beams are being popularly used in modern steel construction. In current fabrication practices of BWS members, high-performance steels produced in steelmaking factories under the thermo-mechanical control process (TMCP) are typically utilized to achieve proper welding performances. However, since its basic unit price is quite higher than typical hot-rolled steel materials, this study introduced a hybrid BWS section for cost saving with no performance degradation, where high-performance TMCP steel was used in flanges, and conventional hot-rolled steel was adopted in web plate. To verify the tensile performances of a hybrid BWS section with non-uniform properties, split T tension and Charpy impact tests were conducted, and flexural tests were also carried out on hybrid and homogeneous BWS beam members. On this basis, it was confirmed that the structural performance of the hybrid BWS member is comparable with that of the conventional one with a uniform section property.

The diagrid structural system has been in the spotlight because of its superiority in terms of its resistance to lateral forces when applied to skyscrapers. In the diagrid structural system, most columns can be eliminated because the vertical loads (gravity loads) and horizontal loads (lateral loads) are delivered simultaneously due to the triangular shape of the diagrid module. Despite this increasing interest, few studies have examined the connection shape and details, which has made it difficult to employ the system to the buildings. In this study, the structural safety of the node connections in a circular steel tube diagrid structural system, which has been considered in the Cyclone Tower in Korea (Seven stories below and fifty-one above the ground), was evaluated using finite element analysis and 4 full-scale specimens. The parameters assessed were the extended length (20, 40, and 60 mm) and thickness (40 and 50 mm)

Received 19 April 2014; received in revised form 14 May 2014; accepted 3 June 2014ABSTRACTEngineering applications of global optimization techniques are recently abundant in the litera-ture and it may be caused by both new methodologies arising and faster computers coming out.Many of the optimization techniques are based on natural or biological phenomena. This studyput focus on enhancing the performace of Cuckoo Search (CS) among them since it has theleast number of parameters to tune. The proposed enhancement can be achieved by applyingsurrogate-based optimization at every cycle of CS, which fortifies the exploitation capability ofthe original method. The enhanced algorithm has been applied several engineering design prob-lems with constraints. The proposed method shows comparable or superior performance to theoriginal method.Key Words: Approximation, Cuckoo Search (CS), Design of experiments (DOE), Global optimi-zation, Metamodel, Surrogate-based optimization (SBO)

Inflammatory cytokines may be a critical component of epithelial cancer progression. We examined the role of interleukin (IL)-5 in the migration of bladder cancer cells. The expression of IL-5 and its receptor IL-5Rα was enhanced in patients with muscle invasive bladder cancers (MIBC), and then it was detected in bladder cancer cell lines 5637 and T-24. IL-5 increased migration and MMP-9 expression via activation of transcription factors NF-κB and AP-1, and induced activation of ERK1/2 and Jak-Stat signaling in both cells. Treatment with ERK1/2 inhibitor U0126 significantly inhibited induction of migration, MMP-9 expression, and activation of NF-κB and AP-1 in IL-5-treated cells. However, none of the Jak inhibitors affected the IL-5-induced migration of bladder cancer cells. Moreover, gene knockdown for IL-5Rα, using siRNA transfection, suppressed migration, ERK1/2 activation, MMP-9 expression, as well as the binding activation of NF-κB and AP-1 in IL-5-treated bladder cancer cells. Similar results were observed in βc siRNA (si-βc) transfected cells. Unexpectedly, IL-5 treatment resulted in significant induction of p21WAF1 in both cell lines. The p21WAF1-specific small interfering RNA inhibited IL-5-induced cell migration, ERK activity, MMP-9 expression, and activation of NF-κB and AP-1 in bladder cancer cells. The effects of IL-5-induced cell responses were confirmed by transfection of IL-5 gene, which demonstrated that p21WAF1 participates in the induction of cell migration, leading to an increase in ERK1/2-mediated MMP-9 expression through activation of NF-κB and AP-1 in IL-5-treated bladder cancer cells. These unexpected results provide a theoretical basis for the therapeutic targeting of IL-5 in bladder cancer.

Various types of reinforcement splicing methods have been developed and implemented in reinforced concrete construction projects for achieving the continuity of reinforcements. Due to the complicated reinforcement arrangements and the difficulties in securing bar spacing, the traditional lap splicing method, which has been widely used in reinforced concrete constructions, often shows low constructability and difficulties in quality control. Also, lap spliced regions are likely to be over-reinforced, which may not be desirable in seismic design. On the other hand, mechanical splicing methods can offer simple and clear arrangements of reinforcement. In order to utilize the couplers for the ribbed-deformed bars, however, additional screw processing at the ends of reinforcing bars is typically required, which often lead to performance degradations of reinforced concrete members due to the lack of workmanship in screw processing or in adjusting the length of reinforcing bars. On the contrary, the use of screw-ribbed reinforcements can easily solve these issues on the mechanical splicing methods, because it does not require the screw process on the bar. In this study, the mechanical coupler suitable for the screw-ribbed reinforcements has been developed, in which any gap between the reinforcements and sleeve device can be removed by grouting high-flow inorganic mortar. This study presents the uniaxial tension tests on the screw-ribbed reinforcement with the mechanical sleeve devices and the cyclic loading tests on RC columns with the developed coupler. The test results show that the mechanical sleeve connection developed in this study has an excellent splicing performance, and that it is applicable to reinforced concrete columns with a proper confinement by hoop reinforcement.

The concept of robust optimization is based on Taguchi's method. Especially, robustness indices of objective function pursue an insensitive and conservative design when there are variations on design variables and parameters. To accomplish the purpose, various robustness indices on the objective function have been developed. However, it can be caused limitations to develop the robustness index, because there is difference between the Taguchi's method and robust optimization. In this paper, an investigation is performed to identify the characteristics and the drawbacks of the previous studies. To achieve the purpose, evaluations are conducted by using the examples which have both a deterministic optimum and a robust optimum. Moreover, a new viewpoint as well as a robustness index using a supremum value of the objective function is proposed based on the investigation.

The thorns of Gleditsia sinensis are a traditional Oriental medicine used for the treatment of swelling, suppuration, carbuncle and skin diseases. In the present study, we identified a novel molecular mechanism by which an ethanol extract of Gleditsia sinensis thorns (EEGS) inhibits the growth of the SNU-5 human gastric cancer cell line. EEGS treatment inhibited cell growth and was associated with G1 phase cell cycle arrest at a concentration of 400 µg/ml (IC50) in SNU-5 cells. Treatment with EEGS also stimulated p21WAF1 expression, which significantly decreased the expression of cyclins and cyclin-dependent kinases (CDKs). Further study suggested that p38 MAP kinase pathways may be involved in the inhibition of cell proliferation through p21WAF1‑dependent G1 phase cell cycle arrest in EEGS-treated cells. In addition, NF-κB and AP-1 transcription factor binding sites were identified as the cis-elements for tumor necrosis factor-α (TNF-α)-induced matrix metalloproteinase-9 (MMP-9) expression in SNU-5 cells, as determined by gel-shift assay. Treatment of cells with EEGS suppressed MMP-9 expression induced by TNF-α via a decrease in the binding activity of both NF-κB and AP-1 motifs. These data demonstrate that EEGS-mediated inhibition of cell growth appears to involve the activation of p38 MAP kinase, subsequently leading to the induction of p21WAF1 and the downregulation of cyclin D1/CDK4 and cyclin E/CDK2 complexes. Moreover, EEGS strongly inhibited TNF-α-induced MMP-9 expression by impeding the DNA binding activity of NF-κB and AP-1. Overall, these results provide a potential mechanism for EEGS in the treatment of gastric cancer.

Received 8 June 2012; received in revised form 5 September 2012; accepted 6 September 2012ABSTRACTMost engineering design problems require analyses or simulations to evaluate objective func-tions. However, a single simulation can take many hours or even days to finish for many realworld problems. As a result, design optimization becomes impossible since they require hun-dreds or thousands of simulation evaluations. The surrogate-based optimization (SBO) strategybecame a remedy for such computationally expensive analyses and simulations. A surrogate-based optimization strategy has been developed in this study in order to improve global optimi-zation performance. The strategy is a heuristic algorithm and it exploits not only multiple surro-gates, but also multiple optimizers. Multiple optimizations of multiple surrogate models yieldmultiple candidate design points of optima. During the sequential sampling process, the algo-rithm ranks candidate design points, selects the points as many as specified, and builds theimproved surrogate model. Various mathematical functions with different numbers of designvariables are chosen to compare the proposed method with the other most recent algorithm,MSEGO. The proposed method shows superior performance to the other method.Key words: Approximation, Design of Experiments (DOE), Global optimization, Meta-model,Sequential sampling, Simulation, Surrogate, Surrogate-based optimization (SBO)

Vehicle collisions frequently happen at a low speed. Insurance companies and the Research Council for Automobile Repairs both require reduction of repair costs and improvement in occupant safety in a low-speed crash. In order to reduce repair costs, an energy absorbing device such as the crash box is usually installed. The crash box is a thin-walled structure attached between the vehicle bumper structure and the side rail. The determination of the crash box geometry is quite important to absorb the impact energy, since the installation space of the crash box is not very large. In this research, a design procedure to determine the cross-sectional dimensions is proposed to enhance the energy absorption capability of the crash box. The proposed process has two steps. In the first step, the cross-sectional dimensions for the conceptual design are determined in two ways. One is a parameter study using discrete design with an orthogonal array. The cross-sectional dimensions of the crash box are selected among the available cross-sections, such as a circle or a polygon. The cross-sectional dimensions are determined by the analysis of the mean from the discrete design with an orthogonal array. The other is topology optimization, which is performed to determine the cross-section of the crash box to maximize the absorbed strain energy based on the Research Council for Automobile Repairs test conditions. The equivalent static loads method for non linear static response structural optimization is employed to solve the formulated topology optimization problems. The cross sections of the crash box are determined from the results of the conceptual design. In the second step, the detailed design processes are performed by using discrete design with an orthogonal array for the models that are selected in the first step. The detailed shapes of the new crash boxes are determined from the detailed design. The optimization problem for the crash box is formulated considering the geometric constraints of fitting into the given space for the crash box. Three new types of crash box are suggested, with detailed shapes from the proposed design procedure.

Although interleukin-28A (IL-28A) is believed to have an antiviral effect, its role in tumor migration requires further examination. The present study was intended to verify the effect of IL-28A on the migration of UMUC-3 bladder cancer cells. IL-28A and its receptor IL-28AR1 mRNA were detected in UMUC-3 cells. Although exogenous IL-28A showed no effect on cell proliferation, a wound-healing migration assay showed that the migration of UMUC-3 cells was induced by IL-28A. Furthermore, treatment of the cells with IL-28A significantly promoted MMP-9 expression via binding activities of NF-κB and AP-1. IL-28A also induced the activation of p38 MAPK and Jak2-Stat2 signaling. Using the p38 MAPK inhibitor SB203580 and the dominant-negative plasmid DN-p38, we found evidence that the inhibition of p38 MAPK signaling suppressed the effects of IL-28A including wound-healing migration and MMP-9 expression by activation of NF-κB and AP-1 binding in UMUC-3 cells. However, Jak-2 inhibition by AG490 did not affect IL-28A-induced migration of UMUC-3 cells. Collectively, we suggest for the first time that the p38 MAPK pathway mediates IL-28A-induced cell migration through MMP-9 expression by activating NF-κB and AP-1 binding motifs.

The reliability based design optimization (RBDO) approach requires high computing cost to consider uncertainties. In order to reduce the design cost, the single loop single vector (SLSV) approach has been developed for RBDO. This method can reduce the cost in calculating deign sensitivity by elimination of the nested optimization process. However, this process causes the increment of the instability or inaccuracy of the method according to the problem characteristics. Therefore, the method may not give accurate solution or the robustness of the solution is not guaranteed. Especially, when the function is concave, the process frequently diverges. In this research, the concept of the conjugate gradient method for unconstrained optimization is utilized to develop a new single loop single vector method. The conjugate gradient is calculated with gradient directions at the most probable points (MPP) of previous cycles. Mathematical examples are solved for the verification of the proposed method. The numeri cal performances of the obtained results are compared to those of other RBDO methods. The SLSV approach using conjugate gradient is not greatly influenced by the problem characteristics and improves its convergence capability.

The thorns of Gleditsia sinensis are used as a medicinal herb in China and Korea. However, the mechanisms responsible for the antitumor effects of the water extract of Gleditsia sinensis thorns (WEGS) remain unknown. HCT116 cells treated with the WEGS at a dose of 800 μg/mL (IC50) showed a significant decrease in cell growth and an increase in cell cycle arrest during the G2/M-phase. G2/M-phase arrest was correlated with increased p53 levels and down-regulation of the check-point proteins, cyclinB1, Cdc2 and Cdc25c. In addition, treatment with WEGS induced phosphorylation of extracellular signal-regulated kinase (ERK), p38 MAP kinase and JNK (c-Jun N-terminal kinases). Moreover, inhibition of ERK by treatment of cells with the ERK-specific inhibitor PD98059 blocked WEGS-mediated p53 expression. Similarly, blockage of ERK function in the WEGS-treated cells reversed cell-growth inhibition and decreased cell cycle proteins. Finally, in vivo WEGS treatment significantly inhibited the growth of HCT116 tumor cell xenografts in nude mice with no negative side effects, including loss of body weight. These results describe the molecular mechanisms whereby the WEGS might inhibit proliferation of colon cancer both in vitro and in vivo, suggesting that WEGS has potential as an anticancer agent for the treatment of malignancies. Copyright © 2010 John Wiley & Sons, Ltd.

Antibacterial and biofilm removal activity of a new podoviridae Staphylococcus aureus bacteriophage (SAP-2), which belongs to the phi29-like phage genus of the Podoviridae family, and a cell-wall-degrading enzyme (SAL-2), which is derived from bacteriophage SAP-2, have been characterized. The cell-wall-degrading enzyme SAL-2 was expressed in Escherichia coli in a soluble form using a low-temperature culture. The cell-wall-degrading enzyme SAL-2 had specific lytic activity against S. aureus, including methicillin-resistant strains, and showed a minimum inhibitory concentration of about 1 microg/ml. In addition, this enzyme showed a broader spectrum of activity within the Staphylococcus genus compared with bacteriophage SAP-2 in its ability to remove the S. aureus biofilms. Thus, the cell-wall-degrading enzyme SAL-2 can be used to prevent and treat biofilm-associated S. aureus infections either on its own or in combination with other cell-wall-degrading enzymes with anti-S. aureus activity.

요 약 본 논문은 중공을 가진 판재 두 매를 결합하기 위하여 중공 주위를 따라 기존 레이저 용접법 대신 기계적 프레스 결합법을 적용하는 것에 관한 연구이다. 이를 통해 레이저 용접을 적용했을 때 불가피하게 발생하는 열 변형을 효과적으로 없앨 수 있다. 유한요소해석을 통하여 중공형 판재를 기계적으로 결합시킬 수 있는 금형 설계 방법을 제안하였다. 기계적 결합력을 최대화시키는 데 관련 있는 다섯 가지 설계인자를 선택하여 다꾸치 실험법을 적용한 결과 성형 깊이와 펀치모서리 반경이 가장 크게 영향을 미치는 인자로 나타났다 .Abstract This study is to apply the mechanical press joining method to join two kinds of sheet metals with circular holes by mechanical pressing instead of laser beam. Usage of the mechanical pressing avoids the thermal deformation of sheet metals which occurs inevitably in laser joining. A die design has been proposed to make the mechanical press joining applicable with finite element analysis. Five design factors related to the joining force have been selected and applied to the Taguchi method for optimization. Among five factors, 'Forming Depth' and 'Punch Corner Radius' have been revealed to be the most influential ones.

This study investigated the anticancer activity of Magnolia officinalis on urinary bladder cancer in vitro and in vivo, and elucidated the mechanism of its activity. An aqueous extract of M. officinalis inhibited cell viability and DNA synthesis in cultured human urinary bladder cancer 5637 cells. Inhibition of proliferation was the result of apoptotic induction, because FACS analyses of 5637 cells treated with M. officinalis showed a sub-G1 phase accumulation. M. officinalis extract also increased cytoplasmic DNA-histone complex dose-dependently. These inhibitory effects were associated with the upregulation of proapoptotic molecules Bax, cytochrome c and caspase 3. Treatment of 5637 cells with M. officinalis extract suppressed the expression of matrix metalloproteinase 2 (MMP-2) and MMP-9, as revealed by zymographic and immunoblot analyses. When M. officinalis extract was given to mice simultaneously with the carcinogen N-butyl-N-(4-hydroxybutyl) nitrosamine, which induces urinary bladder tumors, the size of the induced tumors was smaller. Finally, histological data indicated that the histological grade of carcinoma and the depth of invasion were dramatically decreased by treatment with M. officinalis extract in mice with N-butyl-N-(4-hydroxybutyl) nitrosamine-induced urinary bladder tumors. In conclusion, the findings showed that M. officinalis extract exhibited potential chemopreventive activity against urinary bladder tumor in vitro and in vivo.

Magnolol has been reported to play a role in antitumor activity. However, the relevant pathway integrating cell cycle regulation and signaling pathways involved in growth inhibition in cancer cells remains to be identified. In the present study, magnolol treatment of these cells resulted in significant dose-dependent growth inhibition together with apoptosis, G1- and G2/M-phase cell cycle arrest at a 60 microM (IC50) dose in 5637 bladder cancer cells. In addition, magnolol treatment strongly induced p27KIP1 expression, and down-regulated expression of cyclin-dependent kinases (CDKs) and cyclins. Moreover, treatment with magnolol-induced phosphorylation of ERK, p38 MAP kinase, and JNK. Among the pathway inhibitors examined, only PD98059, an ERK-specific inhibitor, blocked magnolol-dependent p27KIP1 expression. Blockade of ERK function consistently reversed magnolol-mediated inhibition of cell proliferation and decreased G2/M cell cycle proteins, but not G1 cell cycle proteins. Furthermore, magnolol treatment increased both Ras and Raf activation. Transfection of cells with dominant negative Ras (RasN17) and Raf (RafS621A) mutant genes suppressed magnolol-induced ERK activity and p27KIP1 expression. Finally, the magnolol-induced reduction in cell proliferation and G2/M cell cycle proteins was also abolished in the presence of RasN17 and RafS621A mutant genes. These data demonstrate that the Ras/Raf/ERK pathway participates in p27KIP1 induction, leading to a decrease in the levels of cyclin B1/Cdc2 complexes and magnolol-dependent inhibition of cell growth. Overall, these novel findings concerning the molecular mechanisms of magnolol in 5637 bladder cancer cells provide a theoretical basis for therapeutic treatment of malignancies.

The present study identified a novel mechanism for the effects of sanguinarine in vascular smooth muscle cells (VSMC). Sanguinarine treatment of VSMC resulted in significant growth inhibition as a result of G1-phase cell-cycle arrest mediated by induction of p27KIP1 expression, and resulted in a down-regulation of the expression of cyclins and CDKs in VSMC. Moreover, sanguinarine-induced inhibition of cell growth appeared to be linked to activation of Ras/ERK through p27KIP1-mediated G1-phase cell-cycle arrest. Overall, the unexpected effects of sanguinarine treatment in VSMC provide a theoretical basis for clinical use of therapeutic agents in the treatment of atherosclerosis.

A parallel computing technique was applied to large scale structure analysis or aerodynamic design and it is a essential element in reducing the huge computation time for large scale design problem. We can use a many computers for reducing the analysis time of multidisciplinary design optimization. But previous MDO frameworks can not support a parallel design process technique so still existing which calls an analysis program continuously. In this paper, We developed a MDO framework(MLR) which supports a parallel design process to solve sequential analysis call. Finally, three sample cases are presented to show the efficiency of design time using the suggested MDO framework.

Malfunctions or critical fatigue problems often occur in mistuned periodic structural systems since their vibration responses may become much larger than those of perfectly tuned periodic systems. These are called vibration localization phenomena and it is of great importance to accurately predict the localization phenomena for safe and reliable designs of the periodic structural systems. In this study, a simple discrete system which represents periodic structural systems is employed to analyze the vibration localization phenomena. The statistical effects of mistuning, stiffness coupling, and damping on the vibration localization phenomena are investigated through Monte Carlo simulation. It is found that the probability of vibration localization was significantly influenced by the statistical properties except the standard deviation of coupling stiffness.

The role of inflammatory cytokine interleukin-20 (IL-20) has not yet been studied in cancer biology. Here, we demonstrated up-regulation of both IL-20 and IL-20R1 in muscle-invasive bladder cancer patients. The expressions of IL-20 and IL-20R1 were observed in bladder cancer 5637 and T-24 cells. We found that IL-20 significantly increased the expression of matrix metalloproteinase (MMP)-9 via binding activity of NF-κB and AP-1 in bladder cancer cells and stimulated the activation of ERK1/2, JNK, p38 MAPK, and JAK-STAT signaling. Among the pathways examined, only ERK1/2 inhibitor U0126 significantly inhibited IL-20-induced migration and invasion. Moreover, siRNA knockdown of IL-20R1 suppressed migration, invasion, ERK1/2 activation, and NF-κB-mediated MMP-9 expression induced by IL-20. Unexpectedly, the cell cycle inhibitor p21(WAF1) was induced by IL-20 treatment without altering cell cycle progression. Blockade of p21(WAF1) function by siRNA reversed migration, invasion, activation of ERK signaling, MMP-9 expression, and activation of NF-κB in IL-20-treated cells. In addition, IL-20 induced the activation of IκB kinase, the degradation and phosphorylation of IκBα, and NF-κB p65 nuclear translocation, which was regulated by ERK1/2. IL-20 stimulated the recruitment of p65 to the MMP-9 promoter region. Finally, the IL-20-induced migration and invasion of cells was confirmed by IL-20 gene transfection and by addition of anti-IL-20 antibody. This is the first report that p21(WAF1) is involved in ERK1/2-mediated MMP-9 expression via increased binding activity of NF-κB, which resulted in the induction of migration in IL-20/IL-20R1 dyad-induced bladder cancer cells. These unexpected results might provide a critical new target for the treatment of bladder cancer.

We used gene expression profiling to identify inflammatory cytokines that correlate with bladder cancer development. Gene expression profiles of the tissue samples were investigated using cDNA microarrays that contained 103 non-muscle invasive bladder cancers (NMIBC), 62 muscle invasive bladder cancers (MIBC), 58 samples of histologically normal-looking surrounding tissues, and 10 normal, healthy subjects who served as the control cohort for comparison. We grouped the data-sets according to biological characterizations and focused on immune response genes with at least 2-fold differential expression in MIBC vs. controls. The experimental data-set identified 36 immune-related genes that were significantly altered in MIBC samples. In addition, 10 genes were up-regulated and 26 genes were down-regulated in MIBC samples compared with the normal tissues. Among the 10 up-regulated molecules examined, the capacity for both wound-healing migration and invasion was enhanced in response to IL-5, IL-20, and IL-28A in bladder cancer cell lines (253J and EJ cells), compared with untreated cells. The expression levels of IL-5, IL-20, and IL-28A were increased in patients with MIBC. All 3 cytokines and their receptors were produced in bladder cancer cell lines, as determined by real-time PCR, immunoblot analysis and confocal immunofluorescence. Up-regulation of MMP-2 and MMP-9 was found after IL-5, IL-20, and IL-28A stimulation in both cell types. Moreover, an EMSA assay showed that treatment with IL-5, IL-20, and IL-28A induced activation of the transcription factors NF-κB and AP-1 that regulate the MMP-9 promoter. Finally, activation of MAPK and Jak-Stat signaling was observed after the addition of IL-5, IL-20, and IL-28A to bladder cancer cells. This study suggests the presence of specific inflammatory cytokine (IL-5, IL-20, and IL-28A)-mediated association in bladder cancer development. All 3 cytokines may be important new molecular targets for the modulation of migration and invasion in bladder cancer.

The thorns of Gleditsia sinensis, which are extensively used as a medicinal herb in Asian countries, have been reported to exert various pharmacological effects. However, the anti-atherogenic effect of Gleditsia sinensis thorns has never been investigated. In the present study, we investigated the role and effect of the ethanol extract of Gleditsia sinensis thorns (EEGS) on cultured vascular smooth muscle cells (VSMC). Treatment of VSMC with EEGS led to a significant decrease in cell growth by arresting cells in the G2/M-phase of the cell cycle, which was associated with up-regulated p21WAF1 levels and suppression of G2/M cell cycle regulators, cyclinB1, Cdc2 and Cdc25c. In addition, EEGS treatment led to the induction of extracellular signal-regulated kinase1/2 (ERK1/2), p38 MAPK, and JNK (c-Jun N-terminal kinases) activation. EEGS-induced p21WAF1 expression was blocked by treatment with the p38 MAPK-specific inhibitor SB203580. SB203580 also markedly recovered the inhibition of cell growth and decrease in cell cycle proteins in EEGS-treated VSMC. Moreover, EEGS inhibited matrix metalloproteinase-9 (MMP-9) expression induced by tumor necrosis factor-α (TNF-α) in VSMC. Finally, an electrophoresis mobility shift assay demonstrated that EEGS suppressed expression of transcription factor, nuclear factor kappaB (NF-κB) and activator protein-1 (AP-1), which are essential cis-elements for the MMP-9 promoter in TNF-α-treated VSMC. These results demonstrate that EEGS exerts a potent inhibitory effect on cell proliferation and MMP-9 expression in VSMC. These unexpected novel findings represent theoretical data for the preventive and therapeutic use of EEGS for the treatment of atherosclerosis disease.

Interleukin (IL)-28A, also called IFN-λ2, is a member of the classIIcytokine family and has demonstrated anti-proliferative and anti-viral signals. The present study demonstrated migration inducement of IL-28A-treated bladder cancer cells - a novel function. RNA microarray analysis showed an enhanced expression of IL-28A and its receptor IL-28AR1 in muscle invasive urothelial carcinoma in a human bladder. Strong expression of IL-28A and IL-28AR1 was detected in bladder cancer tissues and cell lines (5637, T-24, and HT1376 cells), as determined by real-time PCR and immunoblot analysis. IL-28A treatment induced migration of bladder cancer cells, independent of the cell growth. IL-28AR1-specific small interfering RNA (si-IL-28AR1) inhibited the induction of migration in IL-28A-treated cells. IL-28A treatment stimulated the expression of matrix metalloproteinases-9 (MMP-9) via activation of transcription factor NF-κB. Gene knockdown for MMP-9 and the p65 subunit of NF-κB, using siRNA transfection, suppressed wound healing migration in IL-28A-treated bladder cancer cells. Also, treatment with IL-28A induced activation of mitogen-activated protein kinase (MAPK) in bladder cancer cells. MAPK function blockage by a MAPK-specific inhibitor showed that MAPK phosphorylation is required for IL-28A-induced MMP-9 expression through activation of NF-κB. The transient transfection of bladder cancer cells with ERK1/2 knock down (si-ERK1/2) and dominant negative p38 (DNp38) suppressed IL-28A-induced migration. IL-28A-induced induction of MMP-9 expression, MAPK activation, and DNA binding activity of NF-κB was abolished in the presence of IL-28A neutralizing antibody or by transfection of si-IL-28AR1. These results show that IL-28A/IL-28AR1 dyad-induced wound healing migration requires NF-κB-mediated MMP-9 expression by MAPK activation.

Esculetin, a potent non-competitive inhibitor of lipoxygenase, has been shown to inhibit vascular smooth muscle cell (VSMC) proliferation. However, the effect of esculetin on the matrix metalloproteinase-9 (MMP-9) regulation responsible for cell migration and invasion has not been previously investigated. The results of the present study showed the esculetin (12.5-25 µg/ml) induced the inhibition of migration and invasion in tumor necrosis factor-α (TNF-α)-treated VSMC, as demonstrated by a matrigel invasion assay and wound healing analysis. However, esculetin did not affect cell viability in TNF-α-treated VSMC under 0-25 µg/ml concentration conditions. In addition, both zymographic and immunoblot experiments showed that esculetin suppressed the TNF-α-induced expression of MMP-9 in VSMC in a dose-dependent manner. Furthermore, the treatment of cells with esculetin decreased the activity of the TNF-α-induced MMP-9 promoter, which was driven by a luciferase reporter. Finally, esculetin reduced the binding activities of nuclear factor-κB (NF-κB) and activator protein-1 (AP-1), which are cis-elements present in the promoter of the MMP-9 gene, in TNF-α-treated VSMC. Taken together, these results demonstrated that esculetin decreased the migration and invasion of cells by suppressing MMP-9 expression, which subsequently reduced the binding activities of NF-κB and AP-1 in TNF-α-treated VSMC. These novel findings provide basic information for effective therapeutic treatment with esculetin for atherosclerotic disease.

Decursin, a pyranocoumarin isolated from the Korean Angelica gigas root, has demonstrated anti-cancer properties. In the present study, we found that decursin inhibited cell viability in cultured human urinary bladder cancer 235J cells and colon cancer HCT116 cells. The inhibited proliferation was due to apoptotic induction, because both cells treated with decursin dose-dependently showed a sub-G1 phase accumulation and an increased cytoplasmic DNA-histone complex. Cell death caused by decursin was also associated with the down-regulation of anti-apoptotic factor Bcl-2 and the up-regulation of pro-apoptotic molecules cytochrome c, caspase 3 and Bax. Treatment of both types of cancer cells with decursin resulted in G1-phase cell cycle arrest, as revealed by FACS analyses. In addition, decursin increased protein levels of p21WAF1 with a decrease in cyclins and cyclin dependent kinases (CDKs). Furthermore, decursin induced the activation of extracellular signal-regulated kinases (ERK) in both cancer cell lines, with the notable exceptions of c-Jun N-terminal kinase (JNK) and p38 mitogen activated protein (MAP) kinase. Finally, pretreatment with ERK-specific inhibitor PD98059 reversed decursin-induced p21WAF1 expression and decursin-inhibited cell growth. Thus, these findings suggest that decursin has potential therapeutic efficacy for the treatment of bladder and colon cancer.

The thorns of Gleditsia sinensis have traditionally been used in the treatment of several diseases, which includes their use as anti-tumor agents, but there has been no scientific evidence of this anti-tumor effect. However, the present study has identified a novel mechanism for the anti-tumor effect of Gleditsia sinensis thorns in the treatment of colon cancer. Treatment with the ethanol extract of Gleditsia sinensis thorns (EEGS) resulted in significant growth inhibition together with G2/M-phase cell cycle arrest at a dose of 600 microg/ml (IC50) in HCT116 cells. In addition, treatment with EEGS induced p27 expression and down-regulated expression of cyclins and cyclin-dependent kinases. Moreover, EEGS treatment induced phosphorylation of extracellular signal-regulated kinases (ERK), p38 MAP kinase and JNK (c-Jun N-terminal kinases). Among the pathways examined, only PD98059 (ERK-specific inhibitor) abolished EEGS-dependent p27 expression. Similarly, suppression of ERK function reversed EEGS-mediated cell proliferation inhibition and decreased cell cycle proteins. In addition, tumor necrosis factor-alpha (TNF-alpha)-induced matrix metalloproteinase-9 (MMP-9) expression was inhibited by EEGS treatment via decreased transcriptional activity of both activator protein-1 (AP-1) and nuclear factor-kappaB. Finally, EEGS treatment significantly reduced tumor sizes in HCT116 cell-xenografted tumor tissues, which was associated with the changed levels of ERK phosphorylation, p27 and MMP-9 expression. Overall, these results have identified a novel molecular mechanism for EEGS in the treatment of colon cancer and might provide a theoretical basis for the potential therapeutic use of EEGS in the treatment of malignancies.

Cordycepin (3'-deoxyadenosine) has many anti-cancer properties. However, neither its molecular mechanism nor its molecular targets are well understood. In the present study, we investigated novel molecular mechanisms for the anti-tumor effects of cordycepin in human colon cancer HCT116 cells. After treatment of cells with cordycepin, dose-dependent cell growth inhibition was observed at an IC(50) value of 200muM. Cordycepin treatment resulted in G2/M-phase cell-cycle arrest, which was associated with increased p21WAF1 levels and reduced amounts of cyclin B1, Cdc2, and Cdc25c in a p53-independent pathway. Moreover, cordycepin treatment induced activation of JNK (c-Jun N-terminal kinases). Pretreatment with SP600125, a JNK-specific inhibitor, abrogated cordycepin-mediated p21WAF1 expression, cell growth inhibition, and reduced cell-cycle proteins. Furthermore, JNK1 inhibition by small interfering RNA (siRNA) produced similar results: suppression of cordycepin-induced p21WAF1 expression, decreased cell growth, and reduced cell-cycle proteins. Together, these results suggest a critical role for JNK1 activation in cordycepin-induced inhibition of cell growth and G2/M-phase arrest in human colon cancer cells.

Cordycepin (3'-deoxyadenosine), a bioactive compound of Cordyceps militaris, has many pharmacological activities. The present study reveals novel molecular mechanisms for the anti-tumor effects of cordycepin in two different bladder cancer cell lines, 5637 and T-24 cells. Cordycepin treatment, at a dose of 200 microM (IC(50)) during cell-cycle progression resulted in significant and dose-dependent growth inhibition, which was largely due to G2/M-phase arrest, and resulted in an up-regulation of p21WAF1 expression, independent of the p53 pathway. Moreover, treatment with cordycepin-induced phosphorylation of JNK (c-Jun N-terminal kinases). Blockade of JNK function using SP6001259 (JNK-specific inhibitor) and small interfering RNA (si-JNK1) rescued cordycepin-dependent p21WAF1 expression, inhibited cell growth, and decreased cell cycle proteins. These results suggest that cordycepin could be an effective treatment for bladder cancer.

Cellular and molecular events in vascular smooth muscle cells (VSMC) from Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) were investigated. SHR-derived VSMC showed increased proliferative capacity and MAP kinase levels in comparison with WKY-derived VSMC. Flow cytometry analysis revealed that progression from G1 to S phase was faster in SHR-derived VSMC in response to tumor necrosis factor-alpha (TNF-alpha) as compared with cells from WKY. The G1 cell cycle-associated proteins such as cyclin D1, cyclin E, CDK2 and CDK4, and kinase activities associated with CDK2 and CDK4, were increased in SHR-derived VSMC. In addition, CDK inhibitor p21 was elevated in SHR-derived cells. Matrix metalloproteinase-9 (MMP-9) expression and migration were also increased in response to TNF-alpha in SHR-derived cells. This increase was characterized by the up-regulation of MMP-9, which was transcriptionally regulated at the AP-1 and NF-kappaB sites in the MMP-9 promoter. These results suggest that the hypertensive-associated increase in VSMC proliferative capacity, G1 to S-phase cell-cycle progress and MMP-9 expression may play a role in vascular remodeling in hypertension.