Aspects of in vitro and in vivo research approaches directed towards identifying probiotics and prebiotics for human use

The microbiota of the human gastrointestinal tract plays a key role in nutrition and health. Through the process of fermentation, gut bacteria metabolize various substrates (principally dietary components) to end products such as short-chain fatty acids and gases. This anaerobic metabolism is thought to contribute positively toward host daily energy requirements. However, under certain circumstances, the fermentative process may produce undesirable metabolites. This may cause the onset of gut disorders that can be manifest through both acute and chronic conditions. Moreover, the gut flora may become contaminated by transient pathogens that serve further to upset the normal community structure. There has been a recent increase in the use of dietary components that help to maintain, or even improve, the gut microflora "balance." Probiotics are live microbial feed supplements added to appropriate food vehicles (usually fermented milks), whereas prebiotics are dietary carbohydrates that have a selective metabolism in the colon and serve to increase numbers of bacteria seen as desirable. Because of their purported health-promoting properties, lactic acid-producing bacteria, including bifidobacteria, are the usual target organisms. The market value and biological potential of both approaches are enormous. This article will summarize how efficacious types can be identified.

Dietary components may prevent mutation-related diseases in humans

Since it is not always possible to reduce human exposure to mutagens, attempts have been directed to identify potential antimutagens and anticarcinogens for use in protecting the population against environmental disease. The purpose of this paper is to provide the reader with information about the antimutagenic and anticarcinogenic potentials of some dietary constituents and foods widely consumed in Brazil, and to reinforce diet as a key factor in determining genomic stability and preventing human diseases. In this report, we have summarized data that show interactive effects between some dietary components and specific chemical mutagens or carcinogens using in vitro and in vivo short- or medium-term assays. The summary indicates that certain dietary compounds may be useful agents for disease prevention. (C) 2003 Elsevier B.V. All rights reserved.

Mixing-demixing transition and collapse of a vortex state in a quasi-two-dimensional boson-fermion mixture

We investigate the mixing-demixing transition and the collapse in a quasi-two-dimensional degenerate boson-fermion mixture (DBFM) with a bosonic vortex. We solve numerically a quantum-hydrodynamic model based on a new density functional which accurately takes into account the dimensional crossover. It is demonstrated that with the increase of interspecies repulsion, a mixed state of DBFM could turn into a demixed state. The system collapses for interspecies attraction above a critical value which depends on the vortex quantum number. For interspecies attraction just below this critical limit there is almost complete mixing of boson and fermion components. Such mixed and demixed states of a DBFM could be experimentally realized by varying an external magnetic field near a boson-fermion Feshbach resonance, which will result in a continuous variation of interspecies interaction.

Female fertility in a Guzerat dairy subpopulation: Heterogeneity of variance components for calving intervals

The objectives of the present study were to determine if variance components of calving intervals varied with age at calving and if considering calving intervals as a longitudinal trait would be a useful approach for fertility analysis of Zebu dairy herds. With these purposes, calving records from females born from 1940 to 2006 in a Guzerat dairy subpopulation in Brazil were analyzed. The fixed effects of contemporary groups, formed by year and farm at birth or at calving, and the regressions of age at calving, equivalent inbreeding coefficient and day of the year on the studied traits were considered in the statistical models. In one approach, calving intervals (Cl) were analyzed as a single trait, by fitting a statistical model on which both animal and permanent environment effects were adjusted for the effect of age at calving by random regression. In a second approach, a four-trait analysis was conducted, including age at first calving (AFC) and three different female categories for the calving intervals: first calving females; young females (less than 80 months old, but not first calving); or mature females (80 months old or more). Finally, a two-trait analysis was performed, also including AFC and Cl, but calving intervals were regarded as a single trait in a repeatability model. Additionally, the ranking of sires was compared among approaches. Calving intervals decreased with age until females were about 80 months old, remaining nearly constant after that age. A quasi-linear increase of 11.5 days on the calving intervals was observed for each 10% increase in the female's equivalent inbreeding coefficient. The heritability of AFC was 0.37. For Cl. the genetic-phenotypic variance ratios ranged from 0.064 to 0.141, depending on the approach and on ages at calving. Differences among genetic variance components for calving intervals were observed along the animal's lifetime. Those differences confirmed the longitudinal aspect of that trait, indicating the importance of such consideration when accessing fertility of Zebu dairy females, especially in situations where the available information relies on their calving intervals. Spearman rank correlations among approaches ranged from 0.90 to 0.95, and changes observed in the ranking of sires suggested that the genetic progress of the population could be affected by the approach chosen for the analysis of calving intervals. (C) 2012 Elsevier ay. All rights reserved.

Identification of novel components of NAD-utilizing metabolic pathways and prediction of their biochemical functions

Nicotinamide adenine dinucleotide (NAD) is a ubiquitous cofactor participating in numerous redox reactions. It is also a substrate for regulatory modifications of proteins and nucleic acids via the addition of ADP-ribose moieties or removal of acyl groups by transfer to ADP-ribose. In this study, we use in-depth sequence, structure and genomic context analysis to uncover new enzymes and substrate-binding proteins in NAD-utilizing metabolic and macromolecular modification systems. We predict that Escherichia coli YbiA and related families of domains from diverse bacteria, eukaryotes, large DNA viruses and single strand RNA viruses are previously unrecognized components of NAD-utilizing pathways that probably operate on ADP-ribose derivatives. Using contextual analysis we show that some of these proteins potentially act in RNA repair, where NAD is used to remove 2'-3' cyclic phosphodiester linkages. Likewise, we predict that another family of YbiA-related enzymes is likely to comprise a novel NAD-dependent ADP-ribosylation system for proteins, in conjunction with a previously unrecognized ADP-ribosyltransferase. A similar ADP-ribosyltransferase is also coupled with MACRO or ADP-ribosylglycohydrolase domain proteins in other related systems, suggesting that all these novel systems are likely to comprise pairs of ADP-ribosylation and ribosylglycohydrolase enzymes analogous to the DraG-DraT system, and a novel group of bacterial polymorphic toxins. We present evidence that some of these coupled ADP-ribosyltransferases/ribosylglycohydrolases are likely to regulate certain restriction modification enzymes in bacteria. The ADP-ribosyltransferases found in these, the bacterial polymorphic toxin and host-directed toxin systems of bacteria such as Waddlia also throw light on the evolution of this fold and the origin of eukaryotic polyADP-ribosyltransferases and NEURL4-like ARTs, which might be involved in centrosomal assembly. We also infer a novel biosynthetic pathway that might be involved in the synthesis of a nicotinate-derived compound in conjunction with an asparagine synthetase and AMPylating peptide ligase. We use the data derived from this analysis to understand the origin and early evolutionary trajectories of key NAD-utilizing enzymes and present targets for future biochemical investigations.

Salivary antibodies directed against outer membrane proteins of Moraxella catarrhalis in healthy adults

Moraxella catarrhalis is a major mucosal pathogen of the human respiratory tract, but the mucosal immune response directed against surface components of this organism has not been characterized in detail. The aim of this study was to investigate the salivary immunoglobulin A (IgA) response toward outer membrane proteins (OMP) of M. catarrhalis in healthy adults, the group of individuals least likely to be colonized and thus most likely to display mucosal immunity. Unstimulated saliva samples collected from 14 healthy adult volunteers were subjected to IgA immunoblot analysis with OMP preparations of M. catarrhalis strain O35E. Immunoblot analysis revealed a consistent pattern of IgA reactivity, with the appearance of five major bands located at >250, 200, 120, 80, and 60 kDa. Eleven (79%) of 14 saliva samples elicited reactivity to all five bands. Immunoblot analysis with a set of isogenic knockout mutants lacking the expression of individual OMP was used to determine the identities of OMP giving rise to IgA bands. Human saliva was shown consistently to exhibit IgA-binding activity for oligomeric UspA2 (>250 kDa), hemagglutinin (200 kDa), monomeric UspA1 (120 kDa), transferrin-binding protein B (TbpB), monomeric UspA2, CopB, and presumably OMP CD. TbpB, oligomeric UspA2, and CopB formed a cluster of bands at about 80 kDa. These data indicate that the human salivary IgA response is directed consistently against a small number of major OMP, some of which are presently considered vaccine candidates. The functional properties of these mucosal antibodies remain to be elucidated.

Follicle-forming cat thyroid cell lines synthesizing extracellular matrix and basal membrane components: a new tool for the study of thyroidal morphogenesis

Interactions between follicular epithelial cells and extracellular matrix (ECM) are supposed to play an important role in the development and maintenance of thyroid tissue architecture. In the present study we have therefore investigated the synthesis of ECM components by a feline thyroid cell line which is able to form follicle-like structures in vitro, and also in v-ras-transfected and control-transfected sublines. Transfections were performed by lipofection with pZSR (viral Harvey ras gene; neo) and pSV2-neo (control, neo only) plasmids. We have adapted a semisolid culture system composed exclusively of polymerized alginate and therefore devoid of ECM components. Feline cells embedded in alginate gels as single cells and cultured for up to 90 days formed cell clusters within 10 days. Follicle-like structures were formed in the original cell lines and also in the v-ras- and control-transfected cells. Differences in proliferation rates were observed, the v-ras-transfected cells growing up to two to three times faster than the non-transfected cells. Immunostaining was done using rabbit first antibodies directed against mouse collagen IV, human fibronectin, laminin (tumor Engelbreth-Holm-Swarm laminin), perlecan and other ECM components. For comparison, immunostaining was also performed on cryosections of nodular goiters of six hyperthyroid cats. The cell lines and their transfected clones stained strongly positive for collagen IV and fibronectin, and positively but less strongly for laminin and perlecan. The cat goiter tissue stained positively for collagen IV, laminin, perlecan, and fibronectin, and positive staining for S-laminin (containing the beta2-chain) was seen in blood vessel walls in this tissue. In conclusion, cat cell lines grow three-dimensionally in alginate beads over several weeks, they form follicle-like structures and express the same ECM components as the native cat goiter tissue. Transfection with v-ras does increase proliferation rate, but does not fundamentally alter formation of follicle-like structures and ECM expression. Alginate gel culture is a promising new tool for the study of follicular morphogenesis, polarity, the expression pattern of ECM components and of the interaction between thyrocytes and ECM. It avoids interference caused by gels composed of ECM components.

Concomitant lipopolysaccharide-induced transfer of blood-derived components including immunoglobulins into milk

During a mammary immune response, the integrity of the blood-milk barrier is negatively affected and becomes leaky. The aim of the present study was to demonstrate the blood origin, and to investigate changes in the concentration, of various constituents including immunoglobulins in blood and milk during the early phase of lipopolysaccharide (LPS)-induced mastitis. Five lactating dairy cows received continuous β-hydroxybutyrate (BHBA) clamp infusions to maintain elevated BHBA blood concentrations (1.5 to 2.0 mmol/L) from 48 h before and 8h after LPS administration. One udder quarter was infused with 200 μg of Escherichia coli LPS. A second quarter served as control. Milk and blood samples were taken hourly for 8h postchallenge (PC). The somatic cell count in LPS-challenged quarters was increased from 4h PC to the end of the experiment compared with control quarters. In LPS-challenged quarters, l-lactate, BHBA, lactate dehydrogenase (LDH), IgG(1), and IgG(2) were increased at 3h PC and remained elevated until the end of experiment (8h PC) compared with control quarters. In addition, the optical density values in milk in a nonquantitative ELISA for antibodies directed against bluetongue virus (used as a measure of nonspecific antibody transfer; all animals were vaccinated) increased and, thus, indicates an increase in these antibodies in response to LPS treatment. l-Lactate concentration also increased in blood 2h PC and in the milk of control quarters during the experiment from 3h PC. A second experiment was conducted in vitro to investigate a possible contribution from destructed milk cells to l-lactate concentration and activity of LDH in milk. Aliquots of milk samples (n=8) were frozen (-20°C) or disrupted with ultrasound, respectively. Freeze thawing and ultrasound treatment increased LDH in milk samples, but had no effect on l-lactate concentrations. Results suggest that intramammary infusion of LPS induces a systemic response, as evidenced by an elevation of blood l-lactate concentration. The concomitant changes of all investigated components suggest that they were blood derived. However, the increase in blood components in the milk is not necessarily supportive of the mammary immune system, and likely a side effect of reduced blood-milk barrier integrity.

Strong Connectivity in Directed Hypergraphs and its Application to the Atomic Decomposition of Ontologies

Los hipergrafos dirigidos se han empleado en problemas relacionados con lógica proposicional, bases de datos relacionales, linguística computacional y aprendizaje automático. Los hipergrafos dirigidos han sido también utilizados como alternativa a los grafos (bipartitos) dirigidos para facilitar el estudio de las interacciones entre componentes de sistemas complejos que no pueden ser fácilmente modelados usando exclusivamente relaciones binarias. En este contexto, este tipo de representación es conocida como hiper-redes. Un hipergrafo dirigido es una generalización de un grafo dirigido especialmente adecuado para la representación de relaciones de muchos a muchos. Mientras que una arista en un grafo dirigido define una relación entre dos de sus nodos, una hiperarista en un hipergrafo dirigido define una relación entre dos conjuntos de sus nodos. La conexión fuerte es una relación de equivalencia que divide el conjunto de nodos de un hipergrafo dirigido en particiones y cada partición define una clase de equivalencia conocida como componente fuertemente conexo. El estudio de los componentes fuertemente conexos de un hipergrafo dirigido puede ayudar a conseguir una mejor comprensión de la estructura de este tipo de hipergrafos cuando su tamaño es considerable. En el caso de grafo dirigidos, existen algoritmos muy eficientes para el cálculo de los componentes fuertemente conexos en grafos de gran tamaño. Gracias a estos algoritmos, se ha podido averiguar que la estructura de la WWW tiene forma de “pajarita”, donde más del 70% del los nodos están distribuidos en tres grandes conjuntos y uno de ellos es un componente fuertemente conexo. Este tipo de estructura ha sido también observada en redes complejas en otras áreas como la biología. Estudios de naturaleza similar no han podido ser realizados en hipergrafos dirigidos porque no existe algoritmos capaces de calcular los componentes fuertemente conexos de este tipo de hipergrafos. En esta tesis doctoral, hemos investigado como calcular los componentes fuertemente conexos de un hipergrafo dirigido. En concreto, hemos desarrollado dos algoritmos para este problema y hemos determinado que son correctos y cuál es su complejidad computacional. Ambos algoritmos han sido evaluados empíricamente para comparar sus tiempos de ejecución. Para la evaluación, hemos producido una selección de hipergrafos dirigidos generados de forma aleatoria inspirados en modelos muy conocidos de grafos aleatorios como Erdos-Renyi, Newman-Watts-Strogatz and Barabasi-Albert. Varias optimizaciones para ambos algoritmos han sido implementadas y analizadas en la tesis. En concreto, colapsar los componentes fuertemente conexos del grafo dirigido que se puede construir eliminando ciertas hiperaristas complejas del hipergrafo dirigido original, mejora notablemente los tiempos de ejecucion de los algoritmos para varios de los hipergrafos utilizados en la evaluación. Aparte de los ejemplos de aplicación mencionados anteriormente, los hipergrafos dirigidos han sido también empleados en el área de representación de conocimiento. En concreto, este tipo de hipergrafos se han usado para el cálculo de módulos de ontologías. Una ontología puede ser definida como un conjunto de axiomas que especifican formalmente un conjunto de símbolos y sus relaciones, mientras que un modulo puede ser entendido como un subconjunto de axiomas de la ontología que recoge todo el conocimiento que almacena la ontología sobre un conjunto especifico de símbolos y sus relaciones. En la tesis nos hemos centrado solamente en módulos que han sido calculados usando la técnica de localidad sintáctica. Debido a que las ontologías pueden ser muy grandes, el cálculo de módulos puede facilitar las tareas de re-utilización y mantenimiento de dichas ontologías. Sin embargo, analizar todos los posibles módulos de una ontología es, en general, muy costoso porque el numero de módulos crece de forma exponencial con respecto al número de símbolos y de axiomas de la ontología. Afortunadamente, los axiomas de una ontología pueden ser divididos en particiones conocidas como átomos. Cada átomo representa un conjunto máximo de axiomas que siempre aparecen juntos en un modulo. La decomposición atómica de una ontología es definida como un grafo dirigido de tal forma que cada nodo del grafo corresponde con un átomo y cada arista define una dependencia entre una pareja de átomos. En esta tesis introducimos el concepto de“axiom dependency hypergraph” que generaliza el concepto de descomposición atómica de una ontología. Un modulo en una ontología correspondería con un componente conexo en este tipo de hipergrafos y un átomo de una ontología con un componente fuertemente conexo. Hemos adaptado la implementación de nuestros algoritmos para que funcionen también con axiom dependency hypergraphs y poder de esa forma calcular los átomos de una ontología. Para demostrar la viabilidad de esta idea, hemos incorporado nuestros algoritmos en una aplicación que hemos desarrollado para la extracción de módulos y la descomposición atómica de ontologías. A la aplicación la hemos llamado HyS y hemos estudiado sus tiempos de ejecución usando una selección de ontologías muy conocidas del área biomédica, la mayoría disponibles en el portal de Internet NCBO. Los resultados de la evaluación muestran que los tiempos de ejecución de HyS son mucho mejores que las aplicaciones más rápidas conocidas. ABSTRACT Directed hypergraphs are an intuitive modelling formalism that have been used in problems related to propositional logic, relational databases, computational linguistic and machine learning. Directed hypergraphs are also presented as an alternative to directed (bipartite) graphs to facilitate the study of the interactions between components of complex systems that cannot naturally be modelled as binary relations. In this context, they are known as hyper-networks. A directed hypergraph is a generalization of a directed graph suitable for representing many-to-many relationships. While an edge in a directed graph defines a relation between two nodes of the graph, a hyperedge in a directed hypergraph defines a relation between two sets of nodes. Strong-connectivity is an equivalence relation that induces a partition of the set of nodes of a directed hypergraph into strongly-connected components. These components can be collapsed into single nodes. As result, the size of the original hypergraph can significantly be reduced if the strongly-connected components have many nodes. This approach might contribute to better understand how the nodes of a hypergraph are connected, in particular when the hypergraphs are large. In the case of directed graphs, there are efficient algorithms that can be used to compute the strongly-connected components of large graphs. For instance, it has been shown that the macroscopic structure of the World Wide Web can be represented as a “bow-tie” diagram where more than 70% of the nodes are distributed into three large sets and one of these sets is a large strongly-connected component. This particular structure has been also observed in complex networks in other fields such as, e.g., biology. Similar studies cannot be conducted in a directed hypergraph because there does not exist any algorithm for computing the strongly-connected components of the hypergraph. In this thesis, we investigate ways to compute the strongly-connected components of directed hypergraphs. We present two new algorithms and we show their correctness and computational complexity. One of these algorithms is inspired by Tarjan’s algorithm for directed graphs. The second algorithm follows a simple approach to compute the stronglyconnected components. This approach is based on the fact that two nodes of a graph that are strongly-connected can also reach the same nodes. In other words, the connected component of each node is the same. Both algorithms are empirically evaluated to compare their performances. To this end, we have produced a selection of random directed hypergraphs inspired by existent and well-known random graphs models like Erd˝os-Renyi and Newman-Watts-Strogatz. Besides the application examples that we mentioned earlier, directed hypergraphs have also been employed in the field of knowledge representation. In particular, they have been used to compute the modules of an ontology. An ontology is defined as a collection of axioms that provides a formal specification of a set of terms and their relationships; and a module is a subset of an ontology that completely captures the meaning of certain terms as defined in the ontology. In particular, we focus on the modules computed using the notion of syntactic locality. As ontologies can be very large, the computation of modules facilitates the reuse and maintenance of these ontologies. Analysing all modules of an ontology, however, is in general not feasible as the number of modules grows exponentially in the number of terms and axioms of the ontology. Nevertheless, the modules can succinctly be represented using the Atomic Decomposition of an ontology. Using this representation, an ontology can be partitioned into atoms, which are maximal sets of axioms that co-occur in every module. The Atomic Decomposition is then defined as a directed graph such that each node correspond to an atom and each edge represents a dependency relation between two atoms. In this thesis, we introduce the notion of an axiom dependency hypergraph which is a generalization of the atomic decomposition of an ontology. A module in the ontology corresponds to a connected component in the hypergraph, and the atoms of the ontology to the strongly-connected components. We apply our algorithms for directed hypergraphs to axiom dependency hypergraphs and in this manner, we compute the atoms of an ontology. To demonstrate the viability of this approach, we have implemented the algorithms in the application HyS which computes the modules of ontologies and calculate their atomic decomposition. In the thesis, we provide an experimental evaluation of HyS with a selection of large and prominent biomedical ontologies, most of which are available in the NCBO Bioportal. HyS outperforms state-of-the-art implementations in the tasks of extracting modules and computing the atomic decomposition of these ontologies.

A computationally directed screen identifying interacting coiled coils from Saccharomyces cerevisiae

Computational methods can frequently identify protein-interaction motifs in otherwise uncharacterized open reading frames. However, the identification of candidate ligands for these motifs (e.g., so that partnering can be determined experimentally in a directed manner) is often beyond the scope of current computational capabilities. One exception is provided by the coiled-coil interaction motif, which consists of two or more α helices that wrap around each other: the ligands for coiled-coil sequences are generally other coiled-coil sequences, thereby greatly simplifying the motif/ligand recognition problem. Here, we describe a two-step approach to identifying protein–protein interactions mediated by two-stranded coiled coils that occur in Saccharomyces cerevisiae. Coiled coils from the yeast genome are first predicted computationally, by using the multicoil program, and associations between coiled coils are then determined experimentally by using the yeast two-hybrid assay. We report 213 unique interactions between 162 putative coiled-coil sequences. We evaluate the resulting interactions, focusing on associations identified between components of the spindle pole body (the yeast centrosome).

Mapping protein-protein interactions by affinity-directed mass spectrometry.

A precise and rapid method for identifying sites of interaction between proteins was demonstrated; the basis of the method is direct mass spectrometric readout from the complex to determine the specific components of the proteins that interact--a method termed affinity-directed mass spectrometry. The strategy was used to define the region of interaction of a protein growth factor with a monoclonal antibody. A combination of proteolytic digestion and affinity-directed mass spectrometry was used to rapidly determine the approximate location of a continuous binding epitope within the growth factor. The precise boundaries of the binding epitope were determined by affinity-directed mass spectrometric analysis of sets of synthetic peptide ladders that span the approximate binding region. In addition to the mapping of such linear epitopes, affinity-directed mass spectrometry can be applied to the mapping of other types of molecule-molecule contacts, including ligand-receptor and protein-oligonucleotide interactions.

A quasi-elastic aquifer deformational behavior: Madrid aquifer case study

The purpose of this paper is to analyze the quasi-elastic deformational behavior that has been induced by groundwater withdrawal of the Tertiary detrital aquifer of Madrid (Spain). The spatial and temporal evolution of ground surface displacement was estimated by processing two datasets of radar satellite images (SAR) using Persistent Scatterer Interferometry (PSI). The first SAR dataset was acquired between April 1992 and November 2000 by ERS-1 and ERS-2 satellites, and the second one by the ENVISAT satellite between August 2002 and September 2010. The spatial distribution of PSI measurements reveals that the magnitude of the displacement increases gradually towards the center of the well field area, where approximately 80 mm of maximum cumulated displacement is registered. The correlation analysis made between displacement and piezometric time series provides a correlation coefficient greater than 85% for all the wells. The elastic and inelastic components of measured displacements were separated, observing that the elastic component is, on average, more than 4 times the inelastic component for the studied period. Moreover, the hysteresis loops on the stress–strain plots indicate that the response is in the elastic range. These results demonstrate the quasi-elastic behavior of the aquifer. During the aquifer recovery phase ground surface uplift almost recovers from the subsidence experienced during the preceding extraction phase. Taking into account this unique aquifer system, a one dimensional elastic model was calibrated in the period 1997–2000. Subsequently, the model was used to predict the ground surface movements during the period 1992–2010. Modeled displacements were validated with PSI displacement measurements, exhibiting an error of 13% on average, related with the inelastic component of deformation occurring as a long-term trend in low permeability fine-grained units. This result further demonstrates the quasi-elastic deformational behavior of this unique aquifer system.

Canonical simplification of finite objects, well quasi-ordered by tree embedding /

"UILU-ENG 79 1727."

Inter- and intralaboratory variation of in vitro diffusion cell measurements: An international multicenter study using quasi-standardized methods and materials

In vitro measurements of skin absorption are an increasingly important aspect of regulatory studies, product support claims, and formulation screening. However, such measurements are significantly affected by skin variability. The purpose of this study was to determine inter- and intralaboratory variation in diffusion cell measurements caused by factors other than skin. This was attained through the use of an artificial (silicone rubber) rate-limiting membrane and the provision of materials including a standard penetrant, methyl paraben (MP), and a minimally prescriptive protocol to each of the 18 participating laboratories. Standardized calculations of MP flux were determined from the data submitted by each laboratory by applying a predefined mathematical model. This was deemed necessary to eliminate any interlaboratory variation caused by different methods of flux calculations. Average fluxes of MP calculated and reported by each laboratory (60 +/- 27 mug cm(-2) h(-1), n = 25, range 27-101) were in agreement with the standardized calculations of MP flux (60 +/- 21 mug cm(-2) h(-1), range 19-120). The coefficient of variation between laboratories was approximately 35% and was manifest as a fourfold difference between the lowest and highest average flux values and a sixfold difference between the lowest and highest individual flux values. Intra-laboratory variation was lower, averaging 10% for five individuals using the same equipment within a single laboratory. Further studies should be performed to clarify the exact components responsible for nonskin-related variability in diffusion cell measurements. It is clear that further developments of in vitro methodologies for measuring skin absorption are required. (C) 2005 Wiley-Liss, Inc.

The effect of stimulus location on the major components of the visual evoked response

The topographical distribution of the pattern reversal Visual Evoked Response (VER) was recorded from a localised montage of 20 electrodes over the visual cortex. The response was recorded after stimulation with a black and white checkerboard stimulus. The effect of field location on the major components was investigated in 11 subjects (age range (23-55). The major components of the half field response were; a negative around 75ms (N75) followed by a positivity around 80ms (P80), then a positivity around 100ms (P100) followed by another positivity at around 120ms (P120) and a negativity at approximately 145ms (N145). No effect of field size could be demonstrated on either the amplitude or latency of the late negativity, N145. No significant effect of field size or location was shown on the latency of the P100 response. A delay previously shown in the upper half field response was therefore not substantiated. In contrast the amplitude of the major positivity, P100 was significantly affected by the field size and location. The amplitude of both P100 and N145 were significantly reduced following upper field stimulation when compared with the lower field response. No significant amplitude difference between the upper and lower field responses was demonstrated using electroretinography, the amplitude may therefore be reduced as a result of the ventral position of the upper field representation on the visual cortex. The lateral half field VEP was compared with the distribution of the visual evoked magnetic response (VEMR). The distribution of the VEMR supported the proposal that the paradoxical lateralisation of the VEP half field response is the result of the source being directed ipsilaterally. The morphology of the VEP following octant and double octant stimulation suggests that the response is generated in the striate cortex, with a reversal in response distribution following stimulation of the upper vertical and horizontal meridia.