Practical Murine Hematopathology: A Comparative Review and Implications for Research

Hematologic parameters are important markers of disease in human and veterinary medicine. Biomedical research has benefited from mouse models that recapitulate such disease, thus expanding knowledge of pathogenetic mechanisms and investigative therapies that translate across species. Mice in health have many notable hematologic differences from humans and other veterinary species, including smaller erythrocytes, higher percentage of circulating reticulocytes or polychromasia, lower peripheral blood neutrophil and higher peripheral blood and bone marrow lymphocyte percentages, variable leukocyte morphologies, physiologic splenic hematopoiesis and iron storage, and more numerous and shorter-lived erythrocytes and platelets. For accurate and complete hematologic analyses of disease and response to investigative therapeutic interventions, these differences and the unique features of murine hematopathology must be understood. Here we review murine hematology and hematopathology for practical application to translational investigation.Abbreviations: GEM, genetically engineered mouse; NMB, new methylene blue; nRBC, nucleated RBC; RDW, RBC distribution width; TNCC, total nucleated cell countHematology is an important adjunct to both clinical medicine and biomedical research, with more than 1700 currently funded NIH projects¹⁰⁹ and more than 3400 research articles published over the past 5 years using mouse models.¹²⁰ There are now more than 6000 genetically engineered mouse (GEM) models of disease, with 500 new GEM created each year at the Jackson Laboratory alone, and several large projects are underway to thoroughly phenotype each new mutant mouse strain (https://www.komp.org/).^13,176 A mouse tumor database (http://tumor.informatics.jax.org/mtbwi/index.do) is available to provide information regarding mouse models of human cancer, and the Mouse Phenome Database at the Jackson Laboratory provides links to phenotypic data for many GEM models (http://phenome.jax.org/).⁸ The defined components to complete the phenotyping of GEM models have been recently reviewed.^13,157,176 In addition, 21 inbred strains of mice are commonly used for investigations into such topics as response to infectious and genetically induced disease and dietary and pharmacologic therapies. These commonly used laboratory mouse strains have, for example, inherent differences in immunology or iron trafficking, which can affect research outcomes.^16,47,137 These interstrain differences are important to recognize and understand as a component of effective study design and prior to strain selection for laboratory investigations, especially when hematologic responses to disease need to be considered.^13,16,137For any appropriately designed experiment, concurrent age-, sex-, and strain-matched control mice must be included to accurately compare the effects of a disease, genetic manipulation or therapeutic intervention;^13,155 alternatively, individual mice can be used as their own controls in some studies. Several important guidelines exist to ensure that appropriate numbers of experimental and control mice are incorporated into a study design to maximize statistical power yet minimize waste.^{13,40-42,71,72,176} During and between studies, consistent blood collection methods are essential for accurate comparative analyses. Species-appropriate hematologic instrumentation and timely analysis of fresh blood are necessary to minimize preanalytic hematologic errors.^3,37,71 Especially important for mice and their restricted available blood volume are the use of practical, accurate, species-specific, and up-to-date hematologic methods.Here we comprehensively review murine hematology and hematopathologic responses to disease in the context of biomedical research, discovery, and phenotyping studies. To maximize the opportunity for detecting phenotypes, disease, and responses to therapeutic interventions in mice, we focus on providing a practical summary of methods and analysis for accurate hematologic studies and on describing the morphologic assessment of mouse hematopathology in peripheral blood and bone marrow in ways that will be useful to those—veterinarians and researchers alike—who work with murine species.     

Asbestos in Developing Countries: Magnitude of Risk and Its Practical Implications

In developing countries, aggressive marketing of chrysotile asbestos continues as a result of restrictions on its use being imposed by the developed countries. In the Asian continent, China and India are emerging as the major users of asbestos. There is enough evidence to link chrysotile with pulmonary fibrosis and lung cancer in humans, even at low levels of exposure, hence the need to apply the Precautionary Principle for phasing out its use globally. Due to poor occupational health and safety systems in developing countries and difficulties in early detection of pulmonary malignancy related to asbestos, the statistics remain sketchy. This is hampering efforts to create pressure on policy makers and to counter the propaganda of the asbestos industry. The International Labour Office believes that more than 100,000 deaths a year occur from asbestos-related disease. In the view of studies published in Europe and Australia, the number of deaths due to such malignancies will peak around the year 2020 and could be anywhere between half a million to a million. That means more than a million deaths will occur in developing countries. At about the same time when asbestos-related deaths start to decrease in developed countries, their number will begin to rise in developing countries. This presents a major challenge to the international scientific community.     

Aerobic Granules: Microbial Landscape and Architecture,Stages, and Practical Implications

For the successful application of aerobic granules in wastewater treatment, granules containing an appropriate microbial assembly able to remove contaminants should be retained and propagated within the reactor. To manipulate and/or optimize this process, a good understanding of the formation and dynamic architecture of the granules is desirable. Models of granules often assume a spherical shape with an outer layer and an inner core, but limited information is available regarding the extent of deviations from such assumptions. We report on new imaging approaches to gain detailed insights into the structural characteristics of aerobic granules. Our approach stained all components of the granule to obtain a high quality contrast in the images; hence limitations due to thresholding in the image analysis were overcome. A three-dimensional reconstruction of the granular structure was obtained that revealed the mesoscopic impression of the cavernlike interior of the structure, showing channels and dead-end paths in detail. In “old” granules, large cavities allowed for the irrigation and growth of dense microbial colonies along the path of the channels. Hence, in some areas, paradoxically higher biomass content was observed in the inner part of the granule compared to the outer part. Microbial clusters “rooting” from the interior of the mature granule structure indicate that granules mainly grow via biomass outgrowth and not by aggregation of small particles. We identify and discuss phenomena contributing to the life cycle of aerobic granules. With our approach, volumetric tetrahedral grids are generated that may be used to validate complex models of granule formation.     

Active muscle fiber as an equivalent cardiac current generator

Equations are derived describing potentials due to an active muscle fiber in an infinite medium in terms of two surface integrals—one of the propagated action potential and the other of the membrane current density, both integrals being taken over the surface of the muscle. These equations are incorporated into an equivalent cardiac current generator in which the left ventricle (i.e. the current source) is represented by a three-dimensional wedge and the thorax (i.e. the volume conductor), by a homogeneous circular cylinder. Since this current generator expresses the body surface potentials in terms of the membrane current density and the membrane potential at any point on the surface of the electrically active muscle fiber, the calculated ECG can be correlated with theactual sources within the heart. This equivalent cardiac generator possesses many of the physical and physiological properties of cardiac muscle. The equations were evaluated numerically on a digital computer. The results indicate that equivalent cardiac current generators of this type can yield clinically significant results and that further research is necessary to investigate their properties fully.     

Time to Seed Death as a Function of the Gibbs Free Energy of Water Vapour: Practical and Theoretical Implications

Time (t) to loss of seed viability (e.g. log t or log ) is modelledin the literature as the sum of a moisture term and a quadratictemperature term, f(T,T²). The coefficients inf(T,T²) have beenshown to be ‘identical’ in orthodox seeds. I postulatethat this identity is due to a parameter common to all seedsand, consistent with that hypothesis, report a close correlation(R²=0.9998) between f(T,T²) and the Gibbs free energy of watervapour, G, over the temperature range 0–90 °C. Thehypothesis has the statistical advantage of reducing the numberof independent variables from two to one, without changing thefit. This is demonstrated from analysis of data for lettuceand barley. An explanation for this correlation of time to seeddeath with G is that water vapour is the proximate source ofenergy leading to decomposition of individual molecules, withthe ultimate result of seed death. Copyright 1999 Annals ofBotany Company Seed longevity, temperature effect, Gibbs free energy of water vapour, moisture-temperature interactions, Hordeum vulgare L., barley, Lactuca sativa L., lettuce.     

Species as Explanatory Hypotheses: Refinements and Implications

The formal definition of species as explanatory hypotheses presented by Fitzhugh (Marine Biol 26:155–165, 2005a, b) is emended. A species is an explanatory account of the occurrences of the same character(s) among gonochoristic or cross-fertilizing hermaphroditic individuals by way of character origin and subsequent fixation during tokogeny. In addition to species, biological systematics also employs hypotheses that are ontogenetic, tokogenetic, intraspecific, and phylogenetic, each of which provides explanatory hypotheses for distinctly different classes of causal questions. It is suggested that species hypotheses can not be applied to organisms with obligate asexual, parthenogenetic, and self-fertilizing modes of reproduction. Hypotheses explaining shared characters among such organisms are, instead, strictly phylogenetic. Several implications of this emended definition are examined, especially the relations between species, intraspecific, and phylogenetic hypotheses, as well as the limitations of species names to be applied to temporally different characters within populations.     

Fire acting as an increasing spatial autocorrelation force: Implications for pattern formation and ecological facilitation

Fire is an indissoluble component of ecosystems, however quantifying the effects of fire on vegetation is a challenging task as fire lies outside the typical experimental design attributes. A recent simulation study showed that under increased fire regimes positive tree–tree interactions were recorded (Bacelar et al., 2014). Data from experimental burning plots in an African savanna, the Kruger National Park, were collected across unburnt and annual burn plots. Indices of aggregation and spatial autocorrelation of the distribution of trees between different fire regimes were explored. Results show that the distribution of trees under fire were more clumped and exhibited higher spatial autocorrelation than in unburnt plots. In burnt plots spatial autocorrelation values were positive at finer scales and negative at coarser scales potentially indicating co-existence of facilitation and competition within the same ecosystem depending on the scale. The pattern derived here provides inference for (a) fire acting as an increasing aggregation & spatial autocorrelation force, (b) tree survival under fire regimes is potentially facilitated by forming patches of trees and (c) scale-dependent facilitation and competition coexisting within the same ecosystem with finer scale facilitation and coarser scale competition.     

Intracellular Localization and Conformational State of Transglutaminase 2: Implications for Cell Death

Transglutaminase 2 (TG2) is a multifunctional enzyme that has guanine nucleotide binding and GTP hydrolyzing activity in addition to its transamidating function. Studies show that TG2 is a player in mediating cell death processes. However, there is far from a consensus about the role of this enzyme in cell death processes as it appears to be dependent upon the cell type, stimuli, subcellular localization and conformational state of the enzyme. The purpose of this study was to dissect the role of TG2 in the cell death processes. To this end, we created and characterized 4 distinct point mutants of TG2, each of which differs from the wild type by its conformation or by lacking an important function. We also prepared these mutants as nuclear targeted proteins. By overexpressing mutant or wild type forms of TG2 in HEK 293 cells, we investigated the modulatory role of the protein in the cell death process in response to three stressors: thapsigargin, hyperosmotic stress and oxygen/glucose deprivation (OGD). All of the TG2 constructs, except the R580A mutant (which cannot bind guanine nucleotides and is therefore more prone to exhibit transamidating activity), either did not significantly affect the cell death processes or were protective. However in the case of the R580A mutant, cell death in response to high thapsigargin concentrations, was significantly increased. Intriguingly, nuclear localization of R580A-TG2 was sufficient to counteract the pro-death role of cytoplasmic R580A-TG2. In addition, nuclear localization of TG2 significantly facilitated its protective role against OGD. Our data support the hypothesis that the transamidation activity of TG2, which is mostly quiescent except in extreme stress conditions, is necessary for its pro-death role. In addition, nuclear localization of TG2 generally plays a key role in its protective function against cell death processes, either counteracting the detrimental effect or strengthening the protective role of the protein.     

Active chromatin and noncoding RNAs: an intimate relationship

Eukaryotic genomes are packaged into chromatin, where diverse histone modifications can demarcate chromatin domains that facilitate or block gene expression. While silent chromatin has been associated with long noncoding RNAs (lncRNAs) for some time, new studies suggest that noncoding RNAs also modulate the active chromatin state. Divergent, antisense, and enhancer-like intergenic noncoding RNAs can either activate or repress gene expression by altering histone H3 lysine 4 methylation. An emerging class of enhancer-like lncRNAs may link chromosome structure to chromatin state and establish active chromatin domains. The confluence of several new technologies promises to rapidly expand this fascinating topic of investigation.     

Pupil Size Changes as an Active Information Channel for Biofeedback Applications

Pupil size is usually regarded as a passive information channel that provides insight into cognitive and affective states but defies any further control. However, in a recent study (Ehlers et al. 2015) we demonstrate that sympathetic activity indexed by pupil dynamics allows strategic interference by means of simple cognitive techniques. Utilizing positive/negative imaginings, subjects were able to expand pupil diameter beyond baseline variations; albeit with varying degrees of success and only over brief periods. The current study provides a comprehensive replication on the basis of considerable changes to the experimental set-up. Results show that stricter methodological conditions (controlled baseline settings and specified user instructions) strengthen the reported effect, whereas overall performance increases by one standard deviation. Effects are thereby not restricted to pupillary level. Parallel recordings of skin conductance changes prove a general enhancement of induced autonomic arousal. Considering the stability of the results across studies, we conclude that pupil size information exceeds affective monitoring and may constitute an active input channel in human–computer interaction. Furthermore, since variations in pupil diameter reliably display self-induced changes in sympathetic arousal, the relevance of this parameter is strongly indicated for future approaches in clinical biofeedback.     

An Artemisinin Derivative of Praziquantel as an Orally Active Antischistosomal Agent

Background

Schistosomiasis is a major health problem in tropical and sub-tropical areas caused by species of trematode belonging to the genus Schistosoma. The treatment and control of this disease has been relying on the use of a single drug praziquantel. However, the drug resistance concern urged the development of new drugs against schistosoma. Here, we report our systematic biological evaluation of DW-3-15, a new lead compound developed based on our conjugation design rationale as an effective anti-schistosomal agent.

Methodology/Principal Findings

The antischistosomal activity of DW-3-15 was systematically evaluated in S. japonicum infected mouse model for its stage-sensitivity and dose response. The results revealed that DW-3-15 exhibited 60–85% worm reduction rate against different development stage of worm. Scanning electron microscopy (SEM) observation indicated that DW-3-15 may damage to the tegument of male schistosomes.

Conclusions/Significance

Our results demonstrated that DW-3-15 showed potent anti-schistosomal activities in vivo. The results strongly support our conjugation design strategy of artemisinin analogs and further development of DW-3-15 as a new lead compound as anti-schistosomal agent.     

Woodland Restoration in the Western Australian Wheatbelt: A Conceptual Framework Using a State and Transition Model

Woodlands dominated by Eucalyptus spp. in temperate southeastern and southwestern Australia have been extensively cleared for agriculture and are often badly degraded by livestock grazing. This has resulted in the loss of biodiversity and widespread land degradation. The continuing decline of these woodlands has become a concern for the conservation of biodiversity, and there is a growing interest among farmers, land managers, and researchers in developing techniques for restoring them. Currently few scientific guidelines exist for undertaking woodland restoration programs. We use a state and transition model to develop hypotheses on restoration strategies for salmon gum (Eucalyptus salmonophloia) woodlands. We consider that this approach provides a suitable framework for organizing knowledge and identifying areas where further information is needed, and hence provides a useful starting point for a restoration program. The model has the potential to provide a tool for land managers with which they can assess the action and effort needed to undertake woodland restoration in agricultural landscapes.     

Fabrication of Silver Chevron Arrays as an Efficient and Stable SERS Substrate: Implications in Biological Sensing

Plasmonics - Although glancing angle deposited silver substrates offer an excellent figures for surface enhanced Raman scattering (SERS) sensing, the chemical instability issues of silver...     

The Influence of Science Communication on Indigenous Climate Change Perception: Theoretical and Practical Implications

Citizens receive information on global climate change through both observation of local impacts and reception of climate science. This article presents a quantitative analysis of the interplay of these two sources of information in an indigenous population: residents of Majuro, the capital city of the Republic of the Marshall Islands. While Majuro residents’ reports of local environmental change are partly the result of firsthand observation of changing conditions, survey data robustly demonstrates that environmental change reports are also strongly influenced by awareness of climate science; scientific awareness is a better predictor of environmental change reports than exposure to the environment. This provides a rare quantitative demonstration of the openness of ‘local’ knowledge to foreign scientific information; challenges research methodologies for the study of indigenous climate change perceptions that exclude the role of scientific communication; and suggests a novel, and overlooked, rationale for the dissemination of climate science to frontline communities.     

Focal Electrical Stimulation as an Effective Sham Control for Active rTMS and Biofeedback Treatments

A valid sham control is important for determining the efficacy and effectiveness of repetitive transcranial magnetic stimulation (rTMS) as an experimental and clinical tool. Given the manner in which rTMS is applied, separately or in combination with self-regulatory approaches, and its intended impact on brain states, a valid sham control of this type may well serve as a meaningful control for biofeedback studies, where efforts to develop a credible control have often been less than ideal. This study examined the effectiveness of focal electrical stimulation of the frontalis muscle as a sham technique for blinding participants to high-frequency rTMS over the dorso-lateral prefrontal cortex (DLPFC) at durations, intensities, and schedules of stimulation similar to many clinical applications. In this within-subjects single blind design, 19 participants made guesses immediately after receiving 54 counterbalanced rTMS sessions (sham, 10 Hz, 20 Hz); 7 (13 %) of the guesses were made for sham, 31 (57 %) were made for 10 Hz, and 16 (30 %) were made for 20 Hz. Participants correctly guessed the sham condition 6 % (CI 1, 32 %) of the time, which is less than the odds of chance (i.e., of guessing at random, 33 %); correctly guessed the 10 Hz condition 66 % (CI 43, 84 %) of the time, which was greater than chance; and correctly guessed the 20 Hz condition 41 % (CI 21, 65 %) of the time, which was no different than chance. Focal electrical stimulation therefore can be an effective sham control for high-frequency rTMS of the DLPFC, as well as for active biofeedback interventions. Participants were unaware that electrical stimulation was, in fact, sham rTMS.     

Excitable Channels in Nitellopsis obtusa as Part of an Active Ion-Transport System

The present paper is a study on the rapid and the slow excitablechannels of Nitellopsis obtusa. The working hypothesis is thatduring the excitation of these cells a Ca²⁺-dependent activeion-transport system in the plasmalemma is activated (Gyenesand Bulychev, 1979; Gyenes, Bulychev, and Kurella, 1980) whichmay interact with a light-dependent active transport systemalso present in the plasmalemma. It is found that under conditionsof maximal light-induced current changes, registered in voltageclamp experiments, the amplitudes of both action current componentsare relatively small (10–15 µA cm²) and they increaseup to 100–150 µA cm^–2 during 15–30 minin the dark. Cells may also be excited chemically under conditionsof unchanged voltage across the plasmalemma. It is suggestedthat in the excitation process of Nitellopsis obtusa two typesof ion channels take part/emdash electrically excitable passiveand chemically excitable active channels-both incorporated inone proteolipid complex of a Ca²⁺-dependent ATPase of the plasmalemma.     

Changes in Antioxidant Defense Enzymes after d-amphetamine Exposure: Implications as an Animal Model of Mania

Studies have demonstrated that oxidative stress is associated with amphetamine-induced neurotoxicity, but little is known about the adaptations of antioxidant enzymes in the brain after amphetamine exposure. We studied the effects of acute and chronic amphetamine administration on superoxide dismutase (SOD) and catalase (CAT) activity, in a rodent model of mania. Male Wistar rats received either a single IP injection of d-amphetamine (1 mg/kg, 2 mg/kg, or 4 mg/kg) or vehicle (acute treatment). In the chronic treatment rats received a daily IP injection of either d-amphetamine (1 mg/kg, 2 mg/kg, or 4 mg/kg) or vehicle for 7 days. Locomotor behavior was assessed using the open field test. SOD and CAT activities were measured in the prefrontal cortex, hippocampus, and striatum. Acute and to a greater extent chronic amphetamine treatment increased locomotor behavior and affected SOD and CAT activities in the prefrontal cortex, hippocampus and striatum. Our findings suggest that amphetamine exposure is associated with an imbalance between SOD and CAT activity in the prefrontal cortex, hippocampus and striatum.     

Ground State Robustness as an Evolutionary Design Principle in Signaling Networks

The ability of an organism to survive depends on its capability to adapt to external conditions. In addition to metabolic versatility and efficient replication, reliable signal transduction is essential. As signaling systems are under permanent evolutionary pressure one may assume that their structure reflects certain functional properties. However, despite promising theoretical studies in recent years, the selective forces which shape signaling network topologies in general remain unclear. Here, we propose prevention of autoactivation as one possible evolutionary design principle. A generic framework for continuous kinetic models is used to derive topological implications of demanding a dynamically stable ground state in signaling systems. To this end graph theoretical methods are applied. The index of the underlying digraph is shown to be a key topological property which determines the so-called kinetic ground state (or off-state) robustness. The kinetic robustness depends solely on the composition of the subdigraph with the strongly connected components, which comprise all positive feedbacks in the network. The component with the highest index in the feedback family is shown to dominate the kinetic robustness of the whole network, whereas relative size and girth of these motifs are emphasized as important determinants of the component index. Moreover, depending on topological features, the maintenance of robustness differs when networks are faced with structural perturbations. This structural off-state robustness, defined as the average kinetic robustness of a network''s neighborhood, turns out to be useful since some structural features are neutral towards kinetic robustness, but show up to be supporting against structural perturbations. Among these are a low connectivity, a high divergence and a low path sum. All results are tested against real signaling networks obtained from databases. The analysis suggests that ground state robustness may serve as a rationale for some structural peculiarities found in intracellular signaling networks.