In vitro cytotoxicity of Selol-loaded magnetic nanocapsules against neoplastic cell lines under AC magnetic field activation

The goals of this study are to evaluate in vitro compatibility of magnetic nanomaterials and their therapeutic potential against cancer cells. Highly stable ionic magnetic fluid sample (maghemite, gamma-Fe2O3) and Selol were incorporated into polymeric nanocapsules by nanoprecipitation method. The cytotoxic effect of Selol-loaded magnetic nanocapsules was assessed on murine melanoma (B16-F10) and oral squamous cell carcinoma (OSCC) cell lines following AC magnetic field application. The influence of different nanocapsules on cell viability was investigated by colorimetric MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. In the absence of AC magnetic field Selol-loaded magnetic nanocapsules, containing 100 mu g/mL Selol plus 5 x 10(12) particle/mL, showed antitumoral activity of about 50% on B16-F10 melanoma cells while OSCC carcinoma cells demonstrated drug resistance at all concentrations of Selol and magnetic fluid (range of 100-500 mu g/mL Selol and 5 x 10(12) -2.5 x 10(13) particle/mL). On the other hand, under AC applied fields (1 MHz and 40 Oe amplitude) B16-F10 cell viability was reduced down to 40.5% (+/- 3.33) at the highest concentration of nanoencapsulated Selol. The major effect, however, was observed on OSCC cells since the cell viability drops down to about 33.3% (+/- 0.38) under application of AC magnetic field. These findings clearly indicate that the Selol-loaded magnetic nanocapsules present different toxic effects on neoplastic cell lines. Further, the cytotoxic effect was maximized under AC magnetic field application on OSCC, which emphasizes the effectiveness of the magnetohyperthermia approach. (C) 2012 American Institute of Physics. [doi: 10.1063/1.3680541]

Influence of electric fields over the nucleation ratio of the lithium-potassium sulfate and its pedagogical value

[EN]This paper presents our research about nucleation and its dependency with external conditions, as well as the internal characteristics of the solution itself. Among the research lines of our group, we has been studying the influence of electric fields over two different but related compounds: Lithium-Potassium Sulfate and Lithium-Amonium Sulfate, which both of them show a variation on the nucleation ratio when an electric field is applied during the crystal growth. Moreover, in this paper will be explained a laboratory protocol to teach universitary Science students the nucleation process itself and how it depends on external applied conditions, e.g. electric fields.

Elf magnetic field influence on ION Channels studied by Patch Clamp Technique: exposure set up and "Whole Cell" measurements on Potassium currents

The aim of this thesis was to study the effects of extremely low frequency (ELF) electromagnetic magnetic fields on potassium currents in neural cell lines ( Neuroblastoma SK-N-BE ), using the whole-cell Patch Clamp technique. Such technique is a sophisticated tool capable to investigate the electrophysiological activity at a single cell, and even at single channel level. The total potassium ion currents through the cell membrane was measured while exposing the cells to a combination of static (DC) and alternate (AC) magnetic fields according to the prediction of the so-called ‘ Ion Resonance Hypothesis ’. For this purpose we have designed and fabricated a magnetic field exposure system reaching a good compromise between magnetic field homogeneity and accessibility to the biological sample under the microscope. The magnetic field exposure system consists of three large orthogonal pairs of square coils surrounding the patch clamp set up and connected to the signal generation unit, able to generate different combinations of static and/or alternate magnetic fields. Such system was characterized in term of field distribution and uniformity through computation and direct field measurements. No statistically significant changes in the potassium ion currents through cell membrane were reveled when the cells were exposed to AC/DC magnetic field combination according to the afore mentioned ‘Ion Resonance Hypothesis’.

Modulation of NF-kappaB activation in Theileria annulata-infected cloned cell lines is associated with detection of parasite-dependent IKK signalosomes and disruption of the actin cytoskeleton

Summary Apicomplexan parasites within the genus Theileria have the ability to induce continuous proliferation and prevent apoptosis of the infected bovine leukocyte. Protection against apoptosis involves constitutive activation of the bovine transcription factor NF-kappaB in a parasite-dependent manner. Activation of NF-kappaB is thought to involve recruitment of IKK signalosomes at the surface of the macroschizont stage of the parasite, and it has been postulated that additional host proteins with adaptor or scaffolding function may be involved in signalosome formation. In this study two clonal cell lines were identified that show marked differences in the level of activated NF-kappaB. Further characterization of these lines demonstrated that elevated levels of activated NF-kappaB correlated with increased resistance to cell death and detection of parasite-associated IKK signalosomes, supporting results of our previous studies. Evidence was also provided for the existence of host- and parasite-dependent NF-kappaB activation pathways that are influenced by the architecture of the actin cytoskeleton. Despite this influence, it appears that the primary event required for formation of the parasite-dependent IKK signalosome is likely to be an interaction between a signalosome component and a parasite-encoded surface ligand.

Effects of daunorubicin, mitomycin C, azathioprine and cyclosporin A on human retinal pigmented epithelial, corneal endothelial and conjunctival cell lines

BACKGROUND: We wished to investigate the toxicity of four immunosuppressant and antimetabolic drugs, which are known to influence postoperative wound healing, on three different human ocular cell lines. METHODS: Acute toxicity to cyclosporin A, azathioprine, mitomicyn C and daunorubicin was assessed in Chang cells by monitoring their uptake of propidium iodide during a 3-h period. Chronic toxicity was assessed by monitoring the proliferation and viability of subconfluent cultures of Chang cells, human corneal endothelial cells (HCECs) and retinal pigmented epithelial (RPE) cells after continuous exposure to the drugs for 7 days. RESULTS: Acute toxicity testing revealed no obvious effects. However, the chronic toxicity tests disclosed a narrow concentration range over which cell proliferation decreased dramatically but calcein metabolism was sustained. Although the three lines reacted similarly to each agent, HCECs were the most vulnerable to daunorubicin and mitomycin. At a daunorubicin concentration of 0.05 microg/ml, a 75% decrease in calcein metabolism (P < 0.001) and a > or = 95% cell loss (P < 0.001) were observed. At a mitomycin concentration of 0.01 mug/ml, cell density decreased by 61% (P < 0.001) without a change in calcein metabolism, but at 0.1 microg/ml, the latter parameter decreased to 12% (P = 0.00014). At this concentration the proliferation of Chang and RPE cells decreased by more than 50%, whilst calcein metabolism was largely sustained. Cyclosporin inhibited cell proliferation moderately at lower concentrations (< 5 microg/ml; P=0.05) and substantially at higher ones, with a corresponding decline in calcein metabolism. Azathioprine induced a profound decrease in both parameters at concentrations above 5 microg/ml. CONCLUSION: Daunorubicin, cyclosporin and azathioprine could be used to inhibit excessive intraocular scarring after glaucoma and vitreoretinal surgery without overly reducing cell viability. The attributes of immunosuppressants lie in their combined antiproliferative and immunomodulatory effects.

Wetting and spreading of a surfactant film on solid particles: influence of sharp edges and surface irregularities

In addition to particle size and surface chemistry, the shape of particles plays an important role in their wetting and displacement by the surfactant film in the lung. The role of particle shape was the subject of our investigations using a model system consisting of a modified Langmuir-Wilhelmy surface balance. We measured the influence of sharp edges (lines) and other highly curved surfaces, including sharp corners or spikes, of different particles on the spreading of a dipalmitoylphosphatidyl (DPPC) film. The edges of cylindrical sapphire plates (circular curved edges, 1.65 mm radius) were wetted at a surface tension of 10.7 mJ/m2 (standard error (SE) = 0.45, n = 20) compared with that of 13.8 mJ/m2 (SE = 0.20, n = 20) for cubic sapphire plates (straight linear edges, edge length 3 mm) (p < 0.05). The top surfaces of the sapphire plates (cubic and cylindrical) were wetted at 8.4 mJ/m2 (SE = 0.54, n = 20) and 9.1 mJ/m2 (SE = 0.50, n = 20), respectively, but the difference was not significant (p > 0.05). The surfaces of the plates showed significantly higher resistance to spreading compared to that of the edges, as substantially lower surface tensions were required to initiate wetting (p < 0.05). Similar results were found for talc particles, were the edges of macro- and microcrystalline particles were wetted at 7.2 mJ/m2 (SE = 0.52, n = 20) and 8.2 mJ/m2 (SE = 0.30, n = 20) (p > 0.05), respectively, whereas the surfaces were wetted at 3.8 mJ/m2 (SE = 0.89, n = 20) and 5.8 mJ/m2 (SE = 0.52, n = 20) (p < 0.05), respectively. Further experiments with pollen of malvaceae and maize (spiky and fine knobbly surfaces) were wetted at 10.0 mJ/m2 (SE = 0.52, n = 10) and 22.75 mJ/m2 (SE = 0.81, n = 10), respectively (p < 0.05). These results show that resistance to spreading of a DPPC film on various surfaces is dependent on the extent these surfaces are curved. This is seen with cubic sapphire plates which have at their corners a radius of curvature of about 0.75 microm, spiky malvaceae pollen with an even smaller radius on top of their spikes, or talc with various highly curved surfaces. These highly curved surfaces resisted wetting by the DPPC film to a higher degree than more moderately curved surfaces such as those of cylindrical sapphire plates, maize pollens, or polystyrene spheres, which have a surface free energy similar to that of talc but a smooth surface. The macroscopic plane surfaces of the particles demonstrated the greatest resistance to spreading. This was explained by the extremely fine grooves in the nanometer range, as revealed by electron microscopy. In summary, to understand the effects of airborne particles retained on the surfaces of the respiratory tract, and ultimately their pathological potential, not only the particle size and surface chemistry but also the particle shape should be taken in consideration.

Establishment of murine cell lines by constitutive and conditional immortalization

Mouse cell lines were immortalized by introduction of specific immortalizing genes. Embryonic and adult animals and an embryonal stem cell line were used as a source of primary cells. The immortalizing genes were either introduced by DNA transfection or by ecotropic retrovirus transduction. Fibroblasts were obtained by expression of SV40 virus large T antigen (TAg). The properties of the resulting fibroblast cell lines were reproducible, independent of the donor mouse strains employed and the cells showed no transformed properties in vitro and did not form tumors in vivo. Endothelial cell lines were generated by Polyoma virus middle T antigen expression in primary embryonal cells. These cell lines consistently expressed relevant endothelial cell surface markers. Since the expression of the immortalizing genes was expected to strongly influence the cellular characteristics fibroblastoid cells were reversibly immortalized by using a vector that allows conditional expression of the TAg. Under inducing conditions, these cells exhibited properties that were highly similar to the properties of constitutively immortalized cells. In the absence of TAg expression, cell proliferation stops. Cell growth is resumed when TAg expression is restored. Gene expression profiling indicates that TAg influences the expression levels of more than 1000 genes that are involved in diverse cellular processes. The data show that conditionally immortalized cell lines have several advantageous properties over constitutively immortalized cells.

Effects of epidermal growth factor on the invasion activity of the oral cancer cell lines HSC3 and SAS

Epidermal growth factor (EGF) is excreted in a high concentration in human saliva and modulates the growth and differentiation of various cancer cells. To elucidate the molecular mechanisms by which EGF affects oral cancer growth and invasion, we analyzed the Matrigel invasion activity of the cultured oral cancer cell line. Cells grown under the influence of EGF were subjected to Matrigel invasion assays and cells grown in the absence of EGF were used as controls. Gelatin-zymography and Northern blot analyses quantified the invasiveness and tumorigenicity. Chloramphenicol acetyltransferase assay (CAT assay) determined the EGF stimulation of matrix metalloproteinase (MMP) expression. EGF increased the number of cells penetrating a Matrigel membrane. Gelatin-zymography and Northern blot analysis revealed that MMP9 and Ets1 expressions correlated with EGF but MMP2 was not changed. a transient transfection assay revealed that EGF increased the promoter activities of the MMP9 genes in HSC3 and SAS cells. These results suggest that EGF increases the invasion activity of oral cancer cells partly by increasing MMP9.

TWIST1 and TWIST2 promoter methylation and protein expression in tumor stroma influence the epithelial-mesenchymal transition-like tumor budding phenotype in colorectal cancer.

Tumor budding in colorectal cancer is likened to an epithelial-mesenchymal transition (EMT) characterized predominantly by loss of E-cadherin and up-regulation of E-cadherin repressors like TWIST1 and TWIST2. Here we investigate a possible epigenetic link between TWIST proteins and the tumor budding phenotype. TWIST1 and TWIST2 promoter methylation and protein expression were investigated in six cell lines and further correlated with tumor budding in patient cohort 1 (n = 185). Patient cohort 2 (n = 112) was used to assess prognostic effects. Laser capture microdissection (LCM) of tumor epithelium and stroma from low- and high-grade budding cancers was performed. In colorectal cancers, TWIST1 and TWIST2 expression was essentially restricted to stromal cells. LCM results of a high-grade budding case show positive TWIST1 and TWIST2 stroma and no methylation, while the low-grade budding case was characterized by negative stroma and strong hypermethylation. TWIST1 stromal cell staining was associated with adverse features like more advanced pT (p = 0.0044), lymph node metastasis (p = 0.0301), lymphatic vessel invasion (p = 0.0373), perineural invasion (p = 0.0109) and worse overall survival time (p = 0.0226). Stromal cells may influence tumor budding in colorectal cancers through expression of TWIST1. Hypermethylation of the tumor stroma may represent an alternative mechanism for regulation of TWIST1.

Comparative Cytotoxicity Study of Silver Nanoparticles (AgNPs) in a Variety of Rainbow Trout Cell Lines (RTL-W1, RTH-149, RTG-2) and Primary Hepatocytes.

Among all classes of nanomaterials, silver nanoparticles (AgNPs) have potentially an important ecotoxicological impact, especially in freshwater environments. Fish are particularly susceptible to the toxic effects of silver ions and, with knowledge gaps regarding the contribution of dissolution and unique particle effects to AgNP toxicity, they represent a group of vulnerable organisms. Using cell lines (RTL-W1, RTH-149, RTG-2) and primary hepatocytes of rainbow trout (Oncorhynchus mykiss) as in vitro test systems, we assessed the cytotoxicity of the representative AgNP, NM-300K, and AgNO3 as an Ag+ ion source. Lack of AgNP interference with the cytotoxicity assays (AlamarBlue, CFDA-AM, NRU assay) and their simultaneous application point to the compatibility and usefulness of such a battery of assays. The RTH-149 and RTL-W1 liver cell lines exhibited similar sensitivity as primary hepatocytes towards AgNP toxicity. Leibovitz's L-15 culture medium composition (high amino acid content) had an important influence on the behaviour and toxicity of AgNPs towards the RTL-W1 cell line. The obtained results demonstrate that, with careful consideration, such an in vitro approach can provide valuable toxicological data to be used in an integrated testing strategy for NM-300K risk assessment.

Stark Broadening of Sn III Spectral Lines of Astrophysical Interest: Predictions and Regularities

The determination of the Stark broadening parameters of Sn ions is useful for astrophysicists interested in the determination of the density of electrons in stellar atmospheres. In this paper, we report on the calculated values of the Stark broadening parameters for 171 lines of Sn iii arising from 4d105sns (n= 6–9), 4d105snp (n= 5, 6), 4d105p2, 4d105snd (n= 5–7), 4d105s4f and 4d105s5g. Stark linewidths and line shifts are presented for an electron density of 1023 m−3 and temperatures T= 11 000–75 000 K. These have been calculated using a semi-empirical approach, with a set of wavefunctions obtained from Hartree–Fock relativistic calculations, including core polarization effects. The results obtained have been compared with available experimental data. These can be used to consider the influence of Stark broadening effects in A-type stellar atmospheres

Study of the Influence of Reference System in Surface Finishing Parameters Evaluation.

In the present paper the influence of the reference system with regard to the characterization of the surface finishing is analyzed. The effect of the reference system’s choice on the most representative surface finishing parameters (e.g. roughness average Ra and root mean square values Rq) is studied. The study can also be applied to their equivalent parameters in waviness and primary profiles. Based on ISO and ASME standards, three different types of regression lines (central, mean and orthogonal) are theoretically and experimentally analyzed, identifying the validity and applicability fields of each one depending on profile’s geometry. El presente trabajo realiza un estudio de la influencia que supone la elección del sistema de referencia en la determinación los valores de los parámetros más relevantes empleados en la caracterización del acabado superficial tales como la rugosidad media aritmética Ra o la rugosidad media cuadrática Rq y sus equivalentes en los perfiles de ondulación y completo. Partiendo de la definición establecida por las normas ISO y ASME, se analizan tres tipos de líneas de regresión cuadrática (línea central, línea media y línea ortogonal), delimitando los campos de validez y de aplicación de cada una de ellas en función de la geometría del perfil. Para ello se plantean diversos tipos de perfiles y se desarrolla un estudio teórico y experimental de los mismos.

Improved design of an overhead rail current conductor for application in underground lines

Overhead rail current collector systems for railway traction offer certain features, such as low installation height and reduced maintenance, which make them predominantly suitable for use in underground train infrastructures. Due to the increased demands of modern catenary systems and higher running speeds of new vehicles, a more capable design of the conductor rail is needed. A new overhead conductor rail has been developed and its design has been patented [13]. Modern simulation and modelling techniques were used in the development approach. The new conductor rail profile has a dynamic behaviour superior to that of the system currently in use. Its innovative design permits either an increase of catenary support spacing or a higher vehicle running speed. Both options ensure savings in installation or operating costs. The simulation model used to optimise the existing conductor rail profile included both a finite element model of the catenary and a three-dimensional multi-body system model of the pantograph. The contact force that appears between pantograph and catenary was obtained in simulation. A sensitivity analysis of the key parameters that influence in catenary dynamics was carried out, finally leading to the improved design.

Comparison of vibration and rolling noise emission of resilient and solid monobloc railway wheels in underground lines

Flat or worn wheels rolling on rough or corrugated tracks can provoke airborne noise and ground-borne vibration, which can be a serious concern for nearby neighbours of urban rail transit lines. Among the various treatments used to reduce vibration and noise, resilient wheels play an important role. In conventional resilient wheels, a slightly prestressed V­shaped rubber ring is mounted between the steel wheel centre and tyre. The elastic layer enhances rolling noise and vibration suppression, as well as impact reduction on the track. In this paper the effectiveness of resilient wheels in underground lines, in comparison to monobloc ones, is assessed. The analysed resilient wheel is able to carry greater loads than standard resilient wheels used for light vehicles. It also presents a greater radial resiliency and a higher axial stiffness than conventional V­wheels. The finite element method was used in this study. A quarter car model was defined, in which the wheelset was modelled as an elastic body. Several simulations were performed in order to assess the vibrational behaviour of elastic wheels, including modal, harmonic and random vibration analysis, the latter allowing the introduction of realistic vertical track irregularities, as well as the influence of the running speed. Due to numerical problems some simplifications were needed. Parametric variations were also performed, in which the sensitivity of the whole system to variations of rubber prestress and Poisson’s ratio of the elastic material was assessed.Results are presented in the frequency domain, showing a better performance of the resilient wheels for frequencies over 200 Hz. This result reveals the ability of the analyzed design to mitigate rolling noise, but not structural vibrations, which are primarily found in the lower frequency range.

Advances in the analysis of short span railway bridges for high-speed lines

The physical model based on moving constant loads is widely used for the analysis of railway bridges. Nevertheless, the moving loads model is not well suited for the study of short bridges (L⩽20–25 m) since the results it produces (displacements and accelerations) are much greater than those obtained from more sophisticated ones. In this paper two factors are analysed which are believed to have an influence in the dynamic behaviour of short bridges. These two factors are not accounted for by the moving loads model and are the following: the distribution of the loads due to the presence of the sleepers and ballast layer, and the train–bridge interaction. In order to decide on their influence several numerical simulations have been performed. The results are presented and discussed herein.