Gene expression profiling – Clusters of possibilities

Advances in qPCR technology allow studies of increasingly large systems comprising many genes and samples. The increasing data sizes allow expression profiling both in the gene and the samples dimension while also putting higher demands on sound statistical analysis and expertise to handle and interpret its results. We distinguish between exploratory and confirmatory statistical studies. In this paper we demonstrate several techniques available for exploratory studies on a system of Xenopus laevis development from egg to tadpole. Techniques include hierarchical clustering, heatmap, principal component analysis and self-organizing maps. We stress that even though exploratory studies are excellent for generating hypotheses, results have not been proven statistically significant until an independent confirmatory study has been performed. An exploratory study may certainly be valuable in its own right, and there are often not enough resources to report both an exploratory and a confirmatory study at the same time. However, exploratory and confirmatory studies are intimately connected and we would like to raise that awareness among qPCR practitioners. We suggest that scientific reports should always have a hypothesis focus. Reports are either hypothesis generating, from an exploratory study, or hypothesis validating, from a confirmatory study, or both. In either case, we suggest the generated or validated hypotheses be specifically stated.     

Standardized descriptions of primate locomotor and postural modes

As quantitative studies on primate positional behavior accumulate the lack of a standard positional mode terminology is becoming an increasingly serious deficiency. Inconsistent use of traditional terms and inappropriate conflation of mode categories hamper interspecific and interobserver comparisons. Some workers use common terms without definition, allowing at least the possibility of misunderstanding. Other researchers coin neologisms tailored to their study species and not clearly enough defined to allow application to other species. Such neologisms may overlap, may completely encompass, or may conflate previously defined labels. The result is, at best, the proliferation of synonyms and, at worst, the creation of confusion where clarity had existed. Historical precedents have sometimes resulted in “catch-all” terms that conflate any number of kinematically different behaviors (e.g. “brachiation,” “climbing,” and “quadrumanous climbing”). We recognize three areas where distinction of positional modes has some current importance: (1) Modes that require humeral abduction should be distinguished from adducted behaviors; (2) locomotor modes that involve ascent or descent should be distinguished from horizontal locomotor modes; and (3) suspensory modes should be distinguished from supported modes. We recommend a nomenclature that is not dedicated to or derived from any one taxonomic subset of the primates. Here we define 32 primate positional modes, divided more finely into 52 postural sub-modes and 74 locomotor sub-modes.     

Assessing the impact of school-based mentoring: Common problems and solutions associated with evaluating nonprescriptive youth development programs

ABSTRACT

Like many youth development programs, most youth mentoring programs do not have prescribed practices that target specific outcomes. Because the construct of mentoring represents a broad range of potential activities, researchers face a conundrum when making generalizable causal inferences about the effects of this and similar services. On the one hand, researchers cannot make valid experimental inferences if they do not describe what they manipulate. On the other hand, experiments that include prescribed protocols do not generalize to most mentoring programs. In most cases, researchers conducting school-based mentoring program evaluations err on the side of not sufficiently specifying treatment constructs, which limits the field’s ability to make practically or theoretically useful inferences about this service. We discuss this reality in light of the fundamental logic of the experimental design and suggest several possible solutions to this conundrum. Our goal is to empower researchers to adequately specify treatments while still preserving the treatment construct validity of this and similar interventions.     

Preferential base pairing modes of T·T mismatches

It has long been recognized that T·T mismatches can adopt two different modes of exchangeable wobble base pairs in which no preferential pairing mode has been observed. In this study, we have performed a systematic nuclear magnetic resonance (NMR) investigation to study the sequence context effect on the pairing modes of T·T mismatches. Our results reveal for the first time that preferential pairing mode does exist in T·T mismatches with specific type of flanking base pairs.     

Technical note: Tree truthing: How accurate are substrate estimates in primate field studies?

Field studies of primate positional behavior typically rely on ground‐level estimates of substrate size, angle, and canopy location. These estimates potentially influence the identification of positional modes by the observer recording behaviors. In this study we aim to test ground‐level estimates against direct measurements of support angles, diameters, and canopy heights in trees at La Suerte Biological Research Station in Costa Rica. After reviewing methods that have been used by past researchers, we provide data collected within trees that are compared to estimates obtained from the ground. We climbed five trees and measured 20 supports. Four observers collected measurements of each support from different locations on the ground. Diameter estimates varied from the direct tree measures by 0–28 cm (Mean: 5.44 ± 4.55). Substrate angles varied by 1–55° (Mean: 14.76 ± 14.02). Height in the tree was best estimated using a clinometer as estimates with a two‐meter reference placed by the tree varied by 3–11 meters (Mean: 5.31 ± 2.44). We determined that the best support size estimates were those generated relative to the size of the focal animal and divided into broader categories. Support angles were best estimated in 5° increments and then checked using a Haglöf clinometer in combination with a laser pointer. We conclude that three major factors should be addressed when estimating support features: observer error (e.g., experience and distance from the target), support deformity, and how support size and angle influence the positional mode selected by a primate individual. individual. Am J Phys Anthropol, 2012. © 2012 Wiley Periodicals, Inc.     

Modal gating of human CaV2.1 (P/Q-type) calcium channels: I. The slow and the fast gating modes and their modulation by beta subunits

The single channel gating properties of human CaV2.1 (P/Q-type) calcium channels and their modulation by the auxiliary beta1b, beta2e, beta3a, and beta4a subunits were investigated with cell-attached patch-clamp recordings on HEK293 cells stably expressing human CaV2.1 channels. These calcium channels showed a complex modal gating, which is described in this and the following paper (Fellin, T., S. Luvisetto, M. Spagnolo, and D. Pietrobon. 2004. J. Gen. Physiol. 124:463-474). Here, we report the characterization of two modes of gating of human CaV2.1 channels, the slow mode and the fast mode. A channel in the two gating modes differs in mean closed times and latency to first opening (both longer in the slow mode), in voltage dependence of the open probability (larger depolarizations are necessary to open the channel in the slow mode), in kinetics of inactivation (slower in the slow mode), and voltage dependence of steady-state inactivation (occurring at less negative voltages in the slow mode). CaV2.1 channels containing any of the four beta subtypes can gate in either the slow or the fast mode, with only minor differences in the rate constants of the transitions between closed and open states within each mode. In both modes, CaV2.1 channels display different rates of inactivation and different steady-state inactivation depending on the beta subtype. The type of beta subunit also modulates the relative occurrence of the slow and the fast gating mode of CaV2.1 channels; beta3a promotes the fast mode, whereas beta4a promotes the slow mode. The prevailing mode of gating of CaV2.1 channels lacking a beta subunit is a gating mode in which the channel shows shorter mean open times, longer mean closed times, longer first latency, a much larger fraction of nulls, and activates at more positive voltages than in either the fast or slow mode.     

24-hour homeodynamic states of arterial blood pressure and pulse interval in conscious rats.

In models that describe the homeostasis of the circulation, arterial blood pressure is usually expressed as a single value, which is regarded as the set point in such systems. The aim of the study was to identify in rats from 24-h beat-to-beat recordings the value of blood pressure that describes best such a set point of the cardiovascular system. Normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR), kept on a 12:12-h lights on-off cycle, were instrumented for computerized 24-h beat-to-beat recording of mean arterial pressure (MAP) and pulse interval (PI). Three-dimensional frequency distributions were constructed by plotting for each beat its MAP vs. its PI. During the dark period, the concurrent distribution of MAP and PI showed two distinct modes while during the light period a single mode was found. Comparable patterns were found in SHRs and WKYs. These three different modes were significantly different from the mathematically calculated mean values of MAP and PI over these periods. Thus in rats the 24-h behavior of the cardiovascular system is better described by dynamic shifts between different modes (homeodynamic states) than by a single set point.     

Inactivation of single cardiac Na+ channels in three different gating modes.

In small cell-attached patches containing one and only one Na+ channel, inactivation was studied in three different gating modes, namely, the fast-inactivating F mode and the more slowly inactivating S mode and P mode with similar inactivation kinetics. In each of these modes, ensemble-averaged currents could be fitted with a Hodgkin-Huxley-type model with a single exponential for inactivation (tauh). tauh declined from 1.0 ms at -60 mV to 0.1 ms at 0 mV in the F mode, from 4.6 ms at -40 mV to 1.1 ms at 0 mV in the S mode, and from 4.5 ms at -40 mV to 0.8 ms at +20 mV in the P mode, respectively. The probability of non-empty traces (net), the mean number of openings per non-empty trace (op/tr), and the mean open probability per trace (popen) were evaluated at 4-ms test pulses. net inclined from 30% at -60 mV to 63% at 0 mV in the F mode, from 4% at -90 mV to 90% at 0 mV in the S mode, and from 2% at -60 mV to 79% at +20 mV in the P mode. op/tr declined from 1.4 at -60 mV to 1.1 at 0 mV in the F mode, from 4.0 at -60 mV to 1.2 at 0 mV in the S mode, and from 2.9 at -40 mV to 1.6 at +20 mV in the P mode. popen was bell-shaped with a maximum of 5% at -30 mV in the F mode, 48% at -50 mV in the S mode, and 16% at 0 mV in the P mode. It is concluded that 1) a switch between F and S modes may reflect a functional change of inactivation, 2) a switch between S and P modes may reflect a functional change of activation, 3) tauh is mainly determined by the latency until the first channel opening in the F mode and by the number of reopenings in the S and P modes, 4) at least in the S and P modes, inactivation is independent of pore opening, and 5) in the S mode, mainly open channels inactivate, and in the P mode, mainly closed channels inactivate.     

Varieties of exploratory experimentation in nanotoxicology

There has been relatively little effort to provide a systematic overview of different forms of exploratory experimentation (EE). The present paper examines the growing subdiscipline of nanotoxicology and suggests that it illustrates at least four ways that researchers can engage in EE: searching for regularities; developing new techniques, simulation models, and instrumentation; collecting and analyzing large swaths of data using new experimental strategies (e.g., computer-based simulation and "high-throughput" instrumentation); and structuring an entire disciplinary field around exploratory research agendas. In order to distinguish these and other activities more effectively, the paper proposes a taxonomy that includes three dimensions along which types of EE vary: (1) the aim of the experimental activity, (2) the role of theory in the activity, and (3) the methods or strategies employed for varying experimental parameters.     

Task-based phantom evaluation of cardiac catheterization imaging modes

This study aimed to quantify the dose and quality of the preprogrammed imaging modes on two cardiac angiography devices (Philips Allura FD10 Clarity and Allura FD10) using a task-specific in-house phantom, and to discuss the appropriateness of the pre-programmed settings. A Figure of Merit (FOM), defined as the squared Signal Difference to Noise Ratio (SDNR) divided by Entrance Surface Air Kerma (ESAK), was calculated for phantom inserts with different sizes and concentrations of iodine, as well as tin foils. For the Allura FD10 Clarity device, the low dose fluoroscopic mode was found to be very dose efficient, while the available ciné modes should only be used for cases with high demand for contrast and temporal resolutions. For both devices, the basic beam spectrum of the low dose fluoroscopic mode should be explored for use on other imaging modes. Ciné modes for the Allura FD10 device differ only by their spatial resolution characteristics and have almost identical dose per frame. This study also found that tin may not be a suitable replacement for iodine for research purposes due to mismatching SDNR. The number of recommendations formulated for these two devices suggests that comparative dose and image quality tests of all routinely used imaging modes should be an obligatory part of the physicists’ acceptance testing.     

Influence of Structural Symmetry on Protein Dynamics

Structural symmetry in homooligomeric proteins has intrigued many researchers over the past several decades. However, the implication of protein symmetry is still not well understood. In this study, we performed molecular dynamics (MD) simulations of two forms of trp RNA binding attenuation protein (TRAP), the wild-type 11-mer and an engineered 12-mer, having two different levels of circular symmetry. The results of the simulations showed that the inter-subunit fluctuations in the 11-mer TRAP were significantly smaller than the fluctuations in the 12-mer TRAP while the internal fluctuations were larger in the 11-mer than in the 12-mer. These differences in thermal fluctuations were interpreted by normal mode analysis and group theory. For the 12-mer TRAP, the wave nodes of the normal modes existed at the flexible interface between the subunits, while the 11-mer TRAP had its nodes within the subunits. The principal components derived from the MD simulations showed similar mode structures. These results demonstrated that the structural symmetry was an important determinant of protein dynamics in circularly symmetric homooligomeric proteins.     

A Spatiotemporal Analysis of Brazilian Science from the Perspective of Researchers’ Career Trajectories

The growth of Brazilian scientific production in recent years is remarkable, which motivates an investigation on the factors, inside and outside the country, that helped shape this wealthy research environment. This article provides a thorough analysis of the education of researchers that constitute the main Brazilian research groups, using data on about 6,000 researchers involved in the country’s National Institutes of Science and Technology (INCT) initiative. Data on the steps taken by each researcher in her education, from the bachelor’s degree to doctorate, including a possible postdoctoral experience, and employment, are extracted from an official curriculum vitae repository. The location and the time at which each career step occurred define spatiotemporal career trajectories. We then analyze such trajectories considering additional data, including the area of knowledge of the INCTs to which each researcher is associated. We found an increasing prevalence of Brazilian institutions in the education of Brazilian scientists, as the number of doctorates earned abroad is decreasing over time. Postdoctoral stages, on the other hand, often take place in Europe or in the United States. Taking an international postdoctoral position after a full education in Brazil suggests a drive towards seeking higher-level exchange and cooperation with foreign groups in a more advanced career stage. Results also show that Brazilian researchers tend to seek employment in regions that are close to the institutions at which they received their bachelor’s degrees, suggesting low mobility within the country. This study can be instrumental in defining public policies for correcting distortions, and can help other developing countries that aim to improve their national science systems.     

Multimodal action of single Na+ channels in myocardial mouse cells.

Unitary Na+ currents of myocardial mouse cells were studied at room temperature in 10 cell-attached patches, each containing one and only one channel. Small-pore patch pipettes (resistance 10-97 M omega when filled with 200% Tyrode's solution) with exceptionally thick walls were used. Observed were both rapidly inactivating (6 patches) and slowly inactivating (3 patches) Na+ currents. In one patch, a slow transition from rather fast to slow inactivation was detected over a time of 0.5 h. A short and a long component of the open-channel life time were recorded at the beginning, but only a short one at the end of the experiment. Concomitantly, the first latency was slowed. Amplitude histograms showed that the electrochemical driving force across the pore of the channel did not change during this time. In three patches, a fast and repetitive switching between different modes of Na+ channel action could be clearly identified by plotting the long-time course of the averaged current per trace. The ensemble-averaged current formed in each mode was different in kinetics and amplitude. Each mode had a characteristic mean open-channel life time and distribution of first latency, but the predominant single-channel current amplitude was unaffected by mode switches. It is concluded that two types of changes in kinetics may happen in a single Na+ channel: fast and reversible switches between different modes, and a slow loss of inactivation.     

Deduction of intracellular sub-systems from a topological description of the network

Non-linear behaviour of biochemical networks, such as intracellular gene, protein or metabolic networks, is commonly represented using graphs of the underlying topology. Nodes represent abundance of molecules and edges interactions between pairs of molecules. These graphs are linear and thus based on an implicit linearization of the kinetic reactions in one or several dynamic modes of the total system. It is common to use data from different sources -- experiments conducted under different conditions or even on different species -- meaning that the graph will be a superposition of linearizations made in many different modes. The mixing of different modes makes it hard to identify functional modules, that is sub-systems that carry out a specific biological function, since the graph will contain many interactions that do not naturally occur at the same time. The ability to establish a boundary between the sub-system and its environment is critical in the definition of a module, contrary to a motif in which only internal interactions count. Identification of functional modules should therefore be done on graphs depicting the mode in which their function is carried out, i.e. graphs that only contain edges representing interactions active in the specific mode. In general, when an interaction between two molecules is established, one should always state the mode of the system in which it is active.     

Selection in plant populations of effectively infinite size. V. Biallelic models of trioecy

A one-locus two-allele model of trioecy (presence of hermaphrodites, males and females in one population) is considered, in order to study the conditions for the persistence of this system. All possible assignments of the three sex types to the three genotypes are considered. This leads to three different modes of inheritance of trioecy, namely (a) females heterozygous, (b) males heterozygous and (c) hermaphrodites heterozygous, where in each mode each of the remaining two sex types is homozygous for one of the alleles. For mode (c) trioecy is always persistent, and the dependence of the sex ratio (for the three sex types) on the ovule and pollen fertilities and on the hermaphrodite selfing rate is specified. For the other two modes, (a) and (b), trioecy is not protected, i.e., it may not persist for any fertilities, viabilities or selfing rates. Thus, in this situation it is important to study the conditions under which the "marginal" systems of sexuality of trioecy, i.e., hermaphroditism, dioecy and gynodioecy in mode (a), and hermaphroditism, dioecy and androdioecy in mode (b), may become established. The results show that each marginal system may evolve from each other via trioecy. The evolution of dioecy is easier in mode (a) than in (b), so that female heterogamety would be expected to occur more often than male heterogamety in the present model. Under some conditions the breeding system obtained in equilibrium populations may depend on the initial genotype frequencies.—The necessity of considering modes of inheritance for sexual polymorphisms is demonstrated by comparing our results with those obtained from an evolutionary stable strategy (ESS) analysis of a purely phenotypic model.     

Limited Transfer of Newly Acquired Movement Patterns across Walking and Running in Humans

The two major modes of locomotion in humans, walking and running, may be regarded as a function of different speed (walking as slower and running as faster). Recent results using motor learning tasks in humans, as well as more direct evidence from animal models, advocate for independence in the neural control mechanisms underlying different locomotion tasks. In the current study, we investigated the possible independence of the neural mechanisms underlying human walking and running. Subjects were tested on a split-belt treadmill and adapted to walking or running on an asymmetrically driven treadmill surface. Despite the acquisition of asymmetrical movement patterns in the respective modes, the emergence of asymmetrical movement patterns in the subsequent trials was evident only within the same modes (walking after learning to walk and running after learning to run) and only partial in the opposite modes (walking after learning to run and running after learning to walk) (thus transferred only limitedly across the modes). Further, the storage of the acquired movement pattern in each mode was maintained independently of the opposite mode. Combined, these results provide indirect evidence for independence in the neural control mechanisms underlying the two locomotive modes.     

Exploring the Factor Structure of Neurocognitive Measures in Older Individuals

Here we focus on factor analysis from a best practices point of view, by investigating the factor structure of neuropsychological tests and using the results obtained to illustrate on choosing a reasonable solution. The sample (n=1051 individuals) was randomly divided into two groups: one for exploratory factor analysis (EFA) and principal component analysis (PCA), to investigate the number of factors underlying the neurocognitive variables; the second to test the “best fit” model via confirmatory factor analysis (CFA). For the exploratory step, three extraction (maximum likelihood, principal axis factoring and principal components) and two rotation (orthogonal and oblique) methods were used. The analysis methodology allowed exploring how different cognitive/psychological tests correlated/discriminated between dimensions, indicating that to capture latent structures in similar sample sizes and measures, with approximately normal data distribution, reflective models with oblimin rotation might prove the most adequate.