Epidermal growth factor-induced epithelio-mesenchymal transition in human breast carcinoma cells

PMC42-LA cells display an epithelial phenotype: the cells congregate into pavement epithelial sheets in which E-cadherin and β-catenin are localized at cell-cell borders. They abundantly express cytokeratins, although 5% to 10% of the cells also express the mesenchymal marker vimentin. Stimulation of PMC42-LA cells with epidermal growth factor (EGF) leads to epithelio-mesenchymal transition-like changes including up-regulation of vimentin and down-regulation of E-cadherin. Vimentin expression is seen in virtually all cells, and this increase is abrogated by treatment of cells with an EGF receptor antagonist. The expression of the mesenchyme-associated extracellular matrix molecules fibronectin and chondroitin sulfate proteoglycan also increase in the presence of EGF. PMC42-LA cells adhere rapidly to collagen I, collagen IV, and laminin-1 substrates and markedly more slowly to fibronectin and vitronectin. EGF increases the speed of cell adhesion to most of these extracellular matrix molecules without altering the order of adhesive preference. EGF also caused a time-dependent increase in the motility of PMC42-LA cells, commensurate with the degree of vimentin staining. The increase in motility was at least partly chemokinetic, because it was evident both with and without chemoattractive stimuli. Although E-cadherin staining at cell-cell junctions disappeared in response to EGF, β-catenin persisted at the cell periphery. Further analysis revealed that N-cadherin was present at the cell-cell junctions of untreated cells and that expression was increased after EGF treatment. N- and E-cadherin are not usually coexpressed in human carcinoma cell lines but can be coexpressed in embryonic tissues, and this may signify an epithelial cell population prone to epithelio-mesenchymal-like responses.

Effect of a floating electrode on a plasma jet

Two kinds of floating electrode, floating dielectric barrier covered electrode (FDBCE) and floating pin electrode (FPE), which can enhance the performance of plasma jet are reported. The intense discharge between the floating electrode and power electrode decreased the voltage to trigger the plasma jet substantially. The transition of plasma bullet from ring shape to disk shape in the high helium concentration region happened when the floating electrode was totally inside the powered ring electrode. The enhanced electric field between propagating plasma bullet and ground electrode is the reason for this transition. The double plasma bullets happened when part of the FDBCE was outside the powered ring electrode, which is attributed to the structure and surface charge of FDBCE. As part of the FPE was outside the powered ring electrode, the return stroke resulted in a single intensified plasma channel between FPE and ground electrode.

Chaperone-usher fimbriae of Escherichia coli

Chaperone-usher (CU) fimbriae are adhesive surface organelles common to many Gram-negative bacteria. Escherichia coli genomes contain a large variety of characterised and putative CU fimbrial operons, however, the classification and annotation of individual loci remains problematic. Here we describe a classification model based on usher phylogeny and genomic locus position to categorise the CU fimbrial types of E. coli. Using the BLASTp algorithm, an iterative usher protein search was performed to identify CU fimbrial operons from 35 E. coli (and one Escherichia fergusonnii) genomes representing different pathogenic and phylogenic lineages, as well as 132 Escherichia spp. plasmids. A total of 458 CU fimbrial operons were identified, which represent 38 distinct fimbrial types based on genomic locus position and usher phylogeny. The majority of fimbrial operon types occupied a specific locus position on the E. coli chromosome; exceptions were associated with mobile genetic elements. A group of core-associated E. coli CU fimbriae were defined and include the Type 1, Yad, Yeh, Yfc, Mat, F9 and Ybg fimbriae. These genes were present as intact or disrupted operons at the same genetic locus in almost all genomes examined. Evaluation of the distribution and prevalence of CU fimbrial types among different pathogenic and phylogenic groups provides an overview of group specific fimbrial profiles and insight into the ancestry and evolution of CU fimbriae in E. coli.

Muramidases found in the foregut microbiome of the Tammar wallaby can direct cell aggregation and biofilm formation

We describe here the role of muramidases present in clones of metagenomic DNA that result in cell aggregation and biofilm formation by Escherichia coli. The metagenomic clones were obtained from uncultured Lachnospiraceae-affiliated bacteria resident in the foregut microbiome of the Tammar wallaby. One of these fosmid clones (p49C2) was chosen for more detailed studies and a variety of genetic methods were used to delimit the region responsible for the phenotype to an open reading frame of 1425 bp. Comparative sequence analysis with other fosmid clones giving rise to the same phenotype revealed the presence of muramidase homologues with the same modular composition. Phylogenetic analysis of the fosmid sequence data assigned these fosmid inserts to recently identified, but uncultured, phylogroups of Lachnospiraceae believed to be numerically dominant in the foregut microbiome of the Tammar wallaby. The muramidase is a modular protein containing putative N-acetylmuramoyl--alanine amidase and an endo-β-N-acetylglucosaminidase catalytic module, with a similar organization and functional properties to some Staphylococcal autolysins that also confer adhesive properties and biofilm formation. We also show here that the cloned muramidases result in the production of extracellular DNA, which appears to be the key for biofilm formation and autoaggregation. Collectively, these findings suggest that biofilm formation and cell aggregation in gut microbiomes might occur via the concerted action of carbohydrate-active enzymes and the production of extracellular DNA to serve as a biofilm scaffold.

Uropathogenic Escherichia coli mediated urinary tract infection

Urinary tract infection (UTI) is among the most common infectious diseases of humans and is the most common nosocomial infection in the developed world. They cause significant morbidity and mortality, with approximately 150 million cases globally per year. It is estimated that 40-50% of women and 5% of men will develop a UTI in their lifetime, and UTI accounts for more than 1 million hospitalizations and $1.6 billion in medical expenses each year in the USA. Uropathogenic E. coli (UPEC) is the primary cause of UTI. This review presents an overview of the primary virulence factors of UPEC, the major host responses to infection of the urinary tract, the emergence of specific multidrug resistant clones of UPEC, antibiotic treatment options for UPEC-mediated UTI and the current state of vaccine strategies as well as other novel anti-adhesive and prophylactic approaches to prevent UTI. New and emerging themes in UPEC research are also discussed in the context of future outlooks.

Identification of eusynstyelamide B as a potent cell cycle inhibitor following the generation and screening of an ascidian-derived extract library using a real time cell analyzer

Ascidians are marine invertebrates that have been a source of numerous cytotoxic compounds. Of the first six marine-derived drugs that made anticancer clinical trials, three originated from ascidian specimens. In order to identify new anti-neoplastic compounds, an ascidian extract library (143 samples) was generated and screened in MDA-MB-231 breast cancer cells using a real-time cell analyzer (RTCA). This resulted in 143 time-dependent cell response profiles (TCRP), which are read-outs of changes to the growth rate, morphology, and adhesive characteristics of the cell culture. Twenty-one extracts affected the TCRP of MDA-MB-231 cells and were further investigated regarding toxicity and specificity, as well as their effects on cell morphology and cell cycle. The results of these studies were used to prioritize extracts for bioassay-guided fractionation, which led to the isolation of the previously identified marine natural product, eusynstyelamide B (1). This bis-indole alkaloid was shown to display an IC50 of 5 μM in MDA-MB-231 cells. Moreover, 1 caused a strong cell cycle arrest in G2/M and induced apoptosis after 72 h treatment, making this molecule an attractive candidate for further mechanism of action studies.

Finite element analysis of CFRP strengthened steel hollow sections under tension

This paper presents a nonlinear finite element (FE) model for the analysis of very high strength (VHS) steel hollow sections wrapped by high modulus carbon fibre rein forced polymer (CFRP) sheets. The bond strength of CFRP wrapped VHS circular steel hollow section under tension is investigated using the FE model. The three dimensional FE model by Nonlinear static analysis has been carried out by Strand 7 finite element software. The model is validated by the experimental data obtained from Fawzia et al [1]. A detail parametric study has been performed to examine the effect of number of CFRP layers, different diameters of VHS steel tube and different bond lengths of CFRP sheet. The analytical model developed by Fawzia et al. [1] has been used to determine the load carrying capacity of different diameters of CFRP strengthened VHS steel tube by using the capacity from each layer of CFRP sheet. The results from FE model have found in reasonable agreement with the analytical model developed by Fawzia et al [1]. This validation was necessary because the analytical model by Fawzia et al [1] was developed by using only one diameter of VHS steel tube and fixed (five) number of CFRP layers. It can be concluded that the developed analytical model is valid for CFRP strengthened VHS steel tubes with diameter range of 38mm to 100mm and CFRP layer range of 3 to 5 layers. Based on the results it can also be concluded that the effective bond length is consistent for different diameters of steel tubes and different layers of CFRP. Three layers of CFRP is considered most effective wrapping scheme due to the cost effectiveness. Finally the distribution of longitudinal and hoop stress has been determined by the finite element model for different diameters of CFRP strengthened VHS steel tube.

Durability study of CFRP strengthened steel circular hollow section members under marine environment

Galvanic corrosion is a common phenomenon in Carbon Fibre Reinforced Polymer (CFRP) strengthened steel structures in wet environments and submerged conditions, which reduces durability by weakening the bond between the CFRP and steel substrate. CFRP materials have already been proven to have superior resistance to corrosion and chemical attacks but the adhesive and steel are generally affected by long-term exposure to moisture, especially in conjunction with salts resulting from deicing of ocean spray. This paper presents the results of a research program to improve the durability of CFRP strengthened steel circular hollow section (CHS) members by treating the steel surface with an epoxy based adhesion promoter and inserting Glass Fibre Reinforced Polymer (GFRP) as a galvanic corrosion barrier against simulated sea water. It also presents the effects of accelerated corrosion on the bond of CFRP strengthened hollow steel members. The program consisted of four CFRP strengthened steel beams and one unstrengthened steel beam. Two strengthened beams were used as control while the other two beams were exposed to a highly corrosive environment to induce accelerated corrosion. The corrosion rate was considered 10% which represents a moderate level of loss in the cross-sectional area of the steel tube throughout its intended service life. The beams were then loaded to failure under four-point bending. The research findings indicate that the accelerated corrosion adversely affected the ultimate strength of the conditioned beams and the embedded glass fibre enhanced the bond durability.

Single-cell force spectroscopy, an emerging tool to quantify cell adhesion to biomaterials

Cell adhesion receptors play a central role in sensing and integrating signals provided by the cellular environment. Thus, understanding adhesive interactions at the cell-biomaterial interface is essential to improve the design of implants that should emulate certain characteristics of the cell's natural environment. Numerous cell adhesion assays have been developed; among these, atomic force microscopy-based single-cell force spectroscopy (AFM-SCFS) provides a versatile tool to quantify cell adhesion at physiological conditions. Here we discuss how AFM-SCFS can be used to quantify the adhesion of living cells to biomaterials and give examples of using AFM-SCFS in tissue engineering and regenerative medicine. We anticipate that in the near future, AFM-SCFS will be established in the biomaterial field as an important technique to quantify cell-biomaterial interactions and thereby will contribute to the optimization of implants, scaffolds, and medical devices.

Establishing the impact of temporary tissue expanders on electron and photon beam dose distributions

Purpose: This study investigates the effects of temporary tissue expanders (TTEs) on the dose distributions in breast cancer radiotherapy treatments under a variety of conditions. Methods: Using EBT2 radiochromic film, both electron and photon beam dose distribution measurements were made for different phantoms, and beam geometries. This was done to establish a more comprehensive understanding of the implant’s perturbation effects under a wider variety of conditions. Results: The magnetic disk present in a tissue expander causes a dose reduction of approximately 20% in a photon tangent treatment and 56% in electron boost fields immediately downstream of the implant. The effects of the silicon elastomer are also much more apparent in an electron beam than a photon beam. Conclusions: Evidently, each component of the TTE attenuates the radiation beam to different degrees. This study has demonstrated that the accuracy of photon and electron treatments of post-mastectomy patients is influenced by the presence of a tissue expander for various beam orientations. The impact of TTEs on dose distributions establishes the importance of an accurately modelled high-density implant in the treatment planning system for post-mastectomy patients.

Comparative durability study of CFRP strengthened tubular steel members under cold weather

In strengthening systems, the CFRP (Carbon Fibre Reinforced Polymer) materials typically have excellent resistance against environmental conditions; however, the performance of adhesives between CFRP and steel is generally affected by various environmental conditions such as marine environment, cold and hot weather. This paper presents the comparative durability study of CFRP strengthened tubular steel structures by using two different adhesives such as MBrace saturant and Araldite K630 under four-point bending. The program consisted of testing twelve CFRP strengthened specimens having treated with epoxy based adhesion promoter, untreated surface and one unstrengthened specimen and conditioned under cold weather for 3 and 6 months to determine the environmental durability. The beams were then loaded to failure in quasi-static manner under four-point bending. The structural responses of CFRP strengthened tubular steel beams were compared in terms of failure load, stiffness and modes of failure. The research findings show that the cold weather immersion had adversely affected the durability of CFRP strengthened steel members. Design factor is also proposed to address the short-terms durability performance under cold weather.

The rapid manufacture of uniform composite multicellular-biomaterial micropellets, their assembly into macroscopic organized tissues, and potential applications in cartilage tissue engineering

We and others have published on the rapid manufacture of micropellet tissues, typically formed from 100-500 cells each. The micropellet geometry enhances cellular biological properties, and in many cases the micropellets can subsequently be utilized as building blocks to assemble complex macrotissues. Generally, micropellets are formed from cells alone, however when replicating matrix-rich tissues such as cartilage it would be ideal if matrix or biomaterials supplements could be incorporated directly into the micropellet during the manufacturing process. Herein we describe a method to efficiently incorporate donor cartilage matrix into tissue engineered cartilage micropellets. We lyophilized bovine cartilage matrix, and then shattered it into microscopic pieces having average dimensions < 10 μm diameter; we termed this microscopic donor matrix "cartilage dust (CD)". Using a microwell platform, we show that ~0.83 μg CD can be rapidly and efficiently incorporated into single multicellular aggregates formed from 180 bone marrow mesenchymal stem/stromal cells (MSC) each. The microwell platform enabled the rapid manufacture of thousands of replica composite micropellets, with each micropellet having a material/CD core and a cellular surface. This micropellet organization enabled the rapid bulking up of the micropellet core matrix content, and left an adhesive cellular outer surface. This morphological organization enabled the ready assembly of the composite micropellets into macroscopic tissues. Generically, this is a versatile method that enables the rapid and uniform integration of biomaterials into multicellular micropellets that can then be used as tissue building blocks. In this study, the addition of CD resulted in an approximate 8-fold volume increase in the micropellets, with the donor matrix functioning to contribute to an increase in total cartilage matrix content. Composite micropellets were readily assembled into macroscopic cartilage tissues; the incorporation of CD enhanced tissue size and matrix content, but did not enhance chondrogenic gene expression.

Effect of bond length on the behaviour of CFRP strengthened concrete-filled steel tubes under transverse impact

Concrete-filled steel tubular (CFST) columns have shown great potential as axial load carrying member and used widely in many mission critical infrastructures. However, attention is needed to strengthen these members where transverse impact force is expected to occur due to vehicle collisions. In this work, finite element (FE) model of carbon fibre reinforced polymer (CFRP) strengthened CFST columns are developed and the effect of CFRP bond length is investigated under transverse impact loading. Initially the numerical models have been validated by comparing impact test results from literature. The validated models are then used for detail parametric studies by varying the length of externally bonded CFRP composites. The parameters considered for this research are impact velocity, impact mass, CFRP modulus, adhesive type, and axial static loading. It has been observed that the effect of CFRP strengthening is consistent after an optimum effective bond length of CFRP wrapping. The effect of effective bond length has been studied for above parameters. The results show that, under combined axial static and transverse impact loads CFST columns can successfully prevent global buckling failure by strengthening only 34% of column length. Therefore, estimation of effective bond length is essential to utilise the CFRP composites cost effectively.

Effect of different surface profile on wear of rail steel (AS1085.1) used in Australian heavy-haul railways

The extreme diversity of conditions acting on railways necessitates a variety of experimental approaches to study the critical wear mechanisms that present themselves at the contact interface. This work investigates the effects of contact pressure and geometry in rolling-contact wear tests by using discs with different radii of curvature to simulate the varying contact conditions that may be typically found in the field. It is commonly adapted to line contact interface as it has constant contact pressure. But practical scenario of the rail wheel interface, the contact area increase and contact pressure change as tracks worn off. The tests were conducted without any significant amount of traction, but micro slip was still observed due to contact deformation. Moreover, variation of contact pressure was observed due to contact patch elongation and diameter reduction. Rolling contact fatigue, adhesive and sliding wear were observed on the curved contact interface. The development of different wear regimes and material removal phenomena were analysed using microscopic images in order to broaden the understanding of the wear mechanisms occurring in the rail-wheel contact.

Preliminary work on the detection of partial discharges in SF6 under impulse voltages

This paper presents preliminary results of an investigation into the detection of partial discharges on the rise of impulse voltages from a point-to-plane gap in SF6. A parallel RC detection impedance is placed in the earth path of a point. Computer simulations are done to determine the values of R and C that will result in the smallest impulse voltage signal and the largest discharge signal across the detection impedance. These simulations and the experimental work show that the impulse voltage signal can not be sufficiently attenuated during the rise time of the applied voltage impulse using the RC detection impedance alone. An alternative discharge detection method is proposed in which a resonant partial discharge coupler is used. Elimination of noise and the impulse voltage signal can be achieved by shorting the coupler plate to the ground plane in the middle of the disk. However, due to the bandwidth of the measuring equipment and noise from the impulse generator it was not possible to detect discharges on the rising edge of a 1.5s voltage impulse using a coupler shorted in the middle. It was found that for this particular coupler, with no shorting points, and if the rising edge of the voltage impulse is longer than 5us, (10us) PD activity can be detected on the rising edge.