Method for generation of homogeneous multicellular tumor spheroids applicable to a wide variety of cell types

Multicellular tumor spheroids (MCTS) are used as organotypic models of normal and solid tumor tissue. Traditional techniques for generating MCTS, such as growth on nonadherent surfaces, in suspension, or on scaffolds, have a number of drawbacks, including the need for manual selection to achieve a homogeneous population and the use of nonphysiological matrix compounds. In this study we describe a mild method for the generation of MCTS, in which individual spheroids form in hanging drops suspended from a microtiter plate. The method has been successfully applied to a broad range of cell lines and shows nearly 100% efficiency (i.e., one spheroid per drop). Using the hepatoma cell line, HepG2, the hanging drop method generated well-rounded MCTS with a narrow size distribution (coefficient of variation [CV] 10% to 15%, compared with 40% to 60% for growth on nonadherent surfaces). Structural analysis of HepG2 and a mammary gland adenocarcinoma cell line, MCF-7, composed spheroids, revealed highly organized, three-dimensional, tissue-like structures with an extensive extracellular matrix. The hanging drop method represents an attractive alternative for MCTS production, because it is mild, can be applied to a wide variety of cell lines, and can produce spheroids of a homogeneous size without the need for sieving or manual selection. The method has applications for basic studies of physiology and metabolism, tumor biology, toxicology, cellular organization, and the development of bioartificial tissue. (C) 2003 Wiley Periodicals, Inc.

Stoichiometric and kinetic characterisation of Nitrobacter in mixed culture by decoupling the growth and energy generation processes

The growth, maintenance and lysis processes of Nitrobacter were characterised. A Nitrobacter culture was enriched in a sequencing batch reactor (SBR). Fluorescent in situ hybridisation showed that Nitrobacter constituted 73% of the bacterial population. Batch tests were carried out to measure the oxygen uptake rate and/or nitrite consumption rate when both nitrite and CO2 were in excess, and in the absence of either of these two substrates. The results obtained, along with the SBR performance data, allowed the determination of the maintenance coefficient and in situ cell lysis rate of Nitrobacter. Nitrobacter spends a significant amount of energy for maintenance, which varies considerably with the specific growth rate. At maximum growth, Nitrobacter consume nitrite at a rate of 0.042 mgN/mgCOD(biomass)center dot h for maintenance purposes, which increases more than threefold to 0.143 mgN/mgCOD(biomass)center dot h in the absence of growth. In the SBR, where Nitrobacter grew at 40% of its maximum growth rate, a maintenance coefficient of 0.113 mgN/mgCOD center dot h was found, resulting in 42% of the total amount of nitrite being consumed for maintenance. The above three maintenance coefficient values obtained at different growth rates appear to support the maintenance model proposed in Pirt (1982). The in situ lysis rate of Nitrobacter was determined to be 0.07/day under aerobic conditions at 22 C and pH 7.3. Further, the maximum specific growth rate of Nitrobacter was estimated to be 0.02/h (0.48/day). The affinity constant of Nitrobacter with respect to nitrite was determined to be 1.50 mgNO(2)(-)-N/L, independent of the presence or absence of CO2. (c) 2006 Wiley Periodicals, Inc.

Stoichiometric and kinetic characterisation of Nitrosomonas sp in mixed culture by decoupling the growth and energy generation processes

A novel method that relies on the decoupling of the energy production and biosynthesis processes was used to characterise the maintenance, cell lysis and growth processes of Nitrosomonas sp. A Nitrosolnonas culture was enriched in a sequencing batch reactor (SBR) with ammonium as the sole energy source. Fluorescent in situ hybridization (FISH) showed that Nitrosomonas bound to the NEU probe constituted 82% of the bacterial population, while no other known ammonium or nitrite oxidizing bacteria were detected. Batch tests were carried out under conditions that both ammonium and CO, were in excess, and in the absence of one of these two substrates. The oxygen uptake rate and nitrite production rate were measured during these batch tests. The results obtained from these batch tests, along with the SBR performance data, allowed the determination of the maintenance coefficient and the in situ cell lysis rate, as well as the maximum specific growth rate of the Nitrosomonas culture. It is shown that, during normal growth, the Nitrosomonas culture spends approximately 65% of the energy generated for maintenance. The maintenance coefficient was determined to be 0.14 - 0.16 mgN mgCOD(biomass)(-1) h(-1), and was shown to be independent of the specific growth rate. The in situ lysis rate and the maximum specific growth rate of the Nitrosomonas culture were determined to be 0.26 and 1.0 day(-1) (0.043 h(-1)), respectively, under aerobic conditions at 30 degrees C and pH7. (c) 2006 Elsevier B.V. All rights reserved.

A Nonstationary Model of Newborn EEG

The detection of seizure in the newborn is a critical aspect of neurological research. Current automatic detection techniques are difficult to assess due to the problems associated with acquiring and labelling newborn electroencephalogram (EEG) data. A realistic model for newborn EEG would allow confident development, assessment and comparison of these detection techniques. This paper presents a model for newborn EEG that accounts for its self-similar and non-stationary nature. The model consists of background and seizure sub-models. The newborn EEG background model is based on the short-time power spectrum with a time-varying power law. The relationship between the fractal dimension and the power law of a power spectrum is utilized for accurate estimation of the short-time power law exponent. The newborn EEG seizure model is based on a well-known time-frequency signal model. This model addresses all significant time-frequency characteristics of newborn EEG seizure which include; multiple components or harmonics, piecewise linear instantaneous frequency laws and harmonic amplitude modulation. Estimates of the parameters of both models are shown to be random and are modelled using the data from a total of 500 background epochs and 204 seizure epochs. The newborn EEG background and seizure models are validated against real newborn EEG data using the correlation coefficient. The results show that the output of the proposed models has a higher correlation with real newborn EEG than currently accepted models (a 10% and 38% improvement for background and seizure models, respectively).

Effects of temperature-dependent viscosity variation on entropy generation, heat and fluid flow through a porous-saturated duct of rectangular cross-section

Effect of temperature-dependent viscosity on fully developed forced convection in a duct of rectangular cross-section occupied by a fluid-saturated porous medium is investigated analytically. The Darcy flow model is applied and the viscosity-temperature relation is assumed to be an inverse-linear one. The case of uniform heat flux on the walls, i.e. the H boundary condition in the terminology of Kays and Crawford, is treated. For the case of a fluid whose viscosity decreases with temperature, it is found that the effect of the variation is to increase the Nusselt number for heated walls. Having found the velocity and the temperature distribution, the second law of thermodynamics is invoked to find the local and average entropy generation rate. Expressions for the entropy generation rate, the Bejan number, the heat transfer irreversibility, and the fluid flow irreversibility are presented in terms of the Brinkman number, the Péclet number, the viscosity variation number, the dimensionless wall heat flux, and the aspect ratio (width to height ratio). These expressions let a parametric study of the problem based on which it is observed that the entropy generated due to flow in a duct of square cross-section is more than those of rectangular counterparts while increasing the aspect ratio decreases the entropy generation rate similar to what previously reported for the clear flow case.

Heat transfer and entropy generation optimization of forced convection in a porous-saturated duct of rectangular cross-section

We investigate analytically the first and the second law characteristics of fully developed forced convection inside a porous-saturated duct of rectangular cross-section. The Darcy-Brinkman flow model is employed. Three different types of thermal boundary conditions are examined. Expressions for the Nusselt number, the Bejan number, and the dimensionless entropy generation rate are presented in terms of the system parameters. The conclusions of this analytical study will make it possible to compare, evaluate, and optimize alternative rectangular duct design options in terms of heat transfer, pressure drop, and entropy generation. (c) 2006 Elsevier Ltd. All rights reserved.

Entropy generation for forced convection in a porous saturated circular tube with uniform wall temperature

A numerical study is reported to investigate both the First and the Second Law of Thermodynamics for thermally developing forced convection in a circular tube filled by a saturated porous medium, with uniform wall temperature, and with the effects of viscous dissipation included. A theoretical analysis is also presented to study the problem for the asymptotic region applying the perturbation solution of the Brinkman momentum equation reported by Hooman and Kani [1]. Expressions are reported for the temperature profile, the Nusselt number, the Bejan number, and the dimensionless entropy generation rate in the asymptotic region. Numerical results are found to be in good agreement with theoretical counterparts.

The Coefficient of Variance as an index of L2 lexical processing skill

The Coefficient of Variance (mean standard deviation/mean Response time) is a measure of response time variability that corrects for differences in mean Response time (RT) (Segalowitz & Segalowitz, 1993). A positive correlation between decreasing mean RTs and CVs (rCV-RT) has been proposed as an indicator of L2 automaticity and more generally as an index of processing efficiency. The current study evaluates this claim by examining lexical decision performance by individuals from three levels of English proficiency (Intermediate ESL, Advanced ESL and L1 controls) on stimuli from four levels of item familiarity, as defined by frequency of occurrence. A three-phase model of skill development defined by changing rCV-RT.values was tested. Results showed that RTs and CVs systematically decreased as a function of increasing proficiency and frequency levels, with the rCV-RT serving as a stable indicator of individual differences in lexical decision performance. The rCV-RT and automaticity/restructuring account is discussed in light of the findings. The CV is also evaluated as a more general quantitative index of processing efficiency in the L2.

Analysis of heat transfer and entropy generation for a thermally developing Brinkman–Brinkman forced convection problem in a rectangular duct with isoflux walls

Heat transfer and entropy generation analysis of the thermally developing forced convection in a porous-saturated duct of rectangular cross-section, with walls maintained at a constant and uniform heat flux, is investigated based on the Brinkman flow model. The classical Galerkin method is used to obtain the fully developed velocity distribution. To solve the thermal energy equation, with the effects of viscous dissipation being included, the Extended Weighted Residuals Method (EWRM) is applied. The local (three dimensional) temperature field is solved by utilizing the Green’s function solution based on the EWRM where symbolic algebra is being used for convenience in presentation. Following the computation of the temperature field, expressions are presented for the local Nusselt number and the bulk temperature as a function of the dimensionless longitudinal coordinate, the aspect ratio, the Darcy number, the viscosity ratio, and the Brinkman number. With the velocity and temperature field being determined, the Second Law (of Thermodynamics) aspect of the problem is also investigated. Approximate closed form solutions are also presented for two limiting cases of MDa values. It is observed that decreasing the aspect ratio and MDa values increases the entropy generation rate.

High-resolution bulk density images, using calibrated X-ray radiography of impregnated soil slices

Bulk density of undisturbed soil samples can be measured using computed tomography (CT) techniques with a spatial resolution of about 1 mm. However, this technique may not be readily accessible. On the other hand, x-ray radiographs have only been considered as qualitative images to describe morphological features. A calibration procedure was set up to generate two-dimensional, high-resolution bulk density images from x-ray radiographs made with a conventional x-ray diffraction apparatus. Test bricks were made to assess the accuracy of the method. Slices of impregnated soil samples were made using hardsetting seedbeds that had been gamma scanned at 5-mm depth increments in a previous study. The calibration procedure involved three stages: (i) calibration of the image grey levels in terms of glass thickness using a staircase made from glass cover slips, (ii) measurement of ratio between the soil and resin mass attenuation coefficients and the glass mass attenuation coefficient, using compacted bricks of known thickness and bulk density, and (iii) image correction accounting for the heterogeneity of the irradiation field. The procedure was simple, rapid, and the equipment was easily accessible. The accuracy of the bulk density determination was good (mean relative error 0.015), The bulk density images showed a good spatial resolution, so that many structural details could be observed. The depth functions were consistent with both the global shrinkage and the gamma probe data previously obtained. The suggested method would be easily applied to the new fuzzy set approach of soil structure, which requires generation of bulk density images. Also, it would be an invaluable tool for studies requiring high-resolution bulk density measurement, such as studies on soil surface crusts.

Prefrontal cortex involvement in selective letter generation: A functional magnetic resonance imaging study

Cerebral responses to alternating periods of a control task and a selective letter generation paradigm were investigated with functional Magnetic Resonance Imaging (fMRI). Subjects selectively generated letters from four designated sets of six letters from the English language alphabet, with the instruction that they were not to produce letters in alphabetical order either forward or backward, repeat or alternate letters. Performance during this condition was compared with that of a control condition in which subjects recited the same letters in alphabetical order. Analyses revealed significant and extensive foci of activation in a number of cerebral regions including mid-dorsolateral frontal cortex, inferior frontal gyrus, precuneus, supramarginal gyrus, and cerebellum during the selective letter generation condition. These findings are discussed with respect to recent positron emission tomography (PET) and fMRI studies of verbal working memory and encoding/retrieval in episodic memory.

Size of the first spring generation of Helicoverpa punctigera (Wallengren) (Lepidoptera : Noctuidae) and winter rain in central Australia

Serious infestations of Helicoverpa punctigera are experienced yearly in the eastern cropping regions of Australia. Regression analysis was used to determine whether the size of the first generation in spring (G(1)), which is comprised mostly of immigrants from inland Australia, was related to monthly rainfall in inland winter breeding areas. Data from two long series of light-trap catches at Narrabri in New South Wales (NSW) and Turretfield in South Australia (SA) were used in the analyses. The size of G1 at Narrabri in each year was significantly regressed on the amount of rainfall in western Queensland and NSW in May and June. The size of G1 at Turretfield each year was significantly regressed on the amount of rain in May, June and July in western Queensland and NSW and also in the desert of central Western Australia. Low r(2) values of the regressions suggest that rainfall data for more sites, as well as biological and other physical factors, such as temperature, evaporation, and prevailing wind systems, may need to be included to improve forecasts of the potential magnitude of the infestations in coastal cropping regions.