The utility of animal models in developing immunosuppressive agents

The immune system comprises an integrated network of cellular interactions. Some responses are predictable, while others are more stochastic. While in vitro the outcome of stimulating a single type of cell may be stereotyped and reproducible, in vivo this is often not the case. This phenomenon often merits the use of animal models in predicting the impact of immunosuppressant drugs. A heavy burden of responsibility lies on the shoulders of the investigator when using animal models to study immunosuppressive agents. The principles of the three R׳s: refine (less suffering,), reduce (lower animal numbers) and replace (alternative in vitro assays) must be applied, as described elsewhere in this issue. Well designed animal model experiments have allowed us to develop all the immunosuppressive agents currently available for treating autoimmune disease and transplant recipients. In this review, we examine the common animal models used in developing immunosuppressive agents, focusing on drugs used in transplant surgery. Autoimmune diseases, such as multiple sclerosis, are covered elsewhere in this issue. We look at the utility and limitations of small and large animal models in measuring potency and toxicity of immunosuppressive therapies.

Molecular pathology of prostate cancer:the key to identifying new biomarkers of disease

Microarray technology has recently accelerated the study of the molecular events involved in prostate cancer, offering the prospect of more precise prognosis and new therapeutic strategies. This review summarises current knowledge of the molecular pathology of prostate cancer. The expression and function of numerous genes have been shown to be altered in prostate cancer. Many of these genes are involved in cell cycle regulation, steroid hormone metabolism or regulation of gene expression. The mechanisms by which androgen independence arises are discussed, including cross-activation, gene amplification and point mutations of the androgen receptor. Analysis of changes in the levels of expression of large numbers of genes during prostate cancer progression have provided a better understanding of the basis of the disease, yielding new molecular markers, such as hepsin, with potential use in diagnosis and prognosis.

Molecular Medicine and Molecular Pathology for Hematologists:II. Lets go Techno; Principles of the Molecular Technologies and their Applications in Haematology

Molecular techniques have a key role to play in laboratory and clinical haematology. Restriction enzymes allow nucleic acids to be reduced in size for subsequent analysis. In addition they allow selection of specific DNA or RNA sequences for cloning into bacterial plasmids. These plasmids are naturally occuring DNA molecules which reside in bacterial cells. They can be manipulated to act as vehicles or carriers for biologically and medically important genes, allowing the production of large amounts of cloned material for research purposes or to aid in the production of medically important recombinant molecules such as insulin. As acquired or inherited genetic changes are implicated in a wide range of haematological diseases, it is necessary to have highly specific and sensitive assays to detect these mutations. Most of these techniques rely on nucleic acid hybridisation, benefitting from the ability of DNA or RNA to bind tighly to complimentary bases in the nucleic acid structure. Production of artificial DNA molecules called probes permits nucleic acid hybridiation assays to be performed, using the techniques of southern blotting or dot blot analysis. In addition the base composition of any gene or region of DNA can be determined using DNA sequencing technology. The advent of the polymerase chain reaction (PCR) has revolutionised all aspects of medicine, but has particular relevance in haematology where easy access to biopsy material provides a wealth of material for analysis. PCR permits quick and reliable manipulation of sample material and its ability to be automated makes it an ideal tool for use in the haematology laboratory.

Molecular Medicine and Molecular Pathology for Haematologists:I. Gene Speak made Easy: An Overview of Genes and Gene Expression

Molecular Medicine and Molecular Pathology are integral parts of Haematology as we enter the new millennium. Their origins can be linked to fundamental developments in the basic sciences, particularly genetics, chemistry and biochemistry. The structure of DNA and the genetic code that it encrypts are the critical starting points to our understanding of these new disciplines. The genetic alphabet is a simple one, consisting of just 4 letters, buts its influence is crucial to human development and differentiation. The concept of a gene is not a new one but the Human Genome Project (a joint world-wide effort to characterise our entire genetic make-up) is providing an invaluable understanding of how genes function in normal cellular processes and pinpointing how disruption of these processes can lead to disease. Transcription and translation are the key events by which our genotype is converted to our phenotype (via a messenger RNA intermediate), producing the myriad proteins and enzymes which populate the cellular factory of our body. Unlike the bacterial or prokaryotic genome, the human genome contains a large amount of non coding DNA (less than 1% of our genome codes for proteins), and our genes are interrupted, with the coding regions or exons separated by non coding introns. Precise removal of the intronic material after transcription (though a process called splicing) is critical for efficient translation to occur. Incorrect splicing can lead to the generation of mutant proteins, which can have a dilaterious effect on the phenotype of the individual. Thus the 100,000-200,000 genes which are present in each cell in our body have a defined control mechanism permitting efficient and appropriate expression of proteins and enzymes and yet a single base change in just one of those genes can lead to diseases such as haemophilia or fanconis anaemia.

Diagnostic performance on briefly presented digital pathology images

Background: Identifying new and more robust assessments of proficiency/expertise (finding new "biomarkers of expertise") in histopathology is desirable for many reasons. Advances in digital pathology permit new and innovative tests such as flash viewing tests and eye tracking and slide navigation analyses that would not be possible with a traditional microscope. The main purpose of this study was to examine the usefulness of time-restricted testing of expertise in histopathology using digital images.
Methods: 19 novices (undergraduate medical students), 18 intermediates (trainees), and 19 experts (consultants) were invited to give their opinion on 20 general histopathology cases after 1 s and 10 s viewing times. Differences in performance between groups were measured and the internal reliability of the test was calculated.
Results: There were highly significant differences in performance between the groups using the Fisher's least significant difference method for multiple comparisons. Differences between groups were consistently greater in the 10-s than the 1-s test. The Kuder-Richardson 20 internal reliability coefficients were very high for both tests: 0.905 for the 1-s test and 0.926 for the 10-s test. Consultants had levels of diagnostic accuracy of 72% at 1 s and 83% at 10 s.
Conclusions: Time-restricted tests using digital images have the potential to be extremely reliable tests of diagnostic proficiency in histopathology. A 10-s viewing test may be more reliable than a 1-s test. Over-reliance on "at a glance" diagnoses in histopathology is a potential source of medical error due to over-confidence bias and premature closure.

Farm animal welfare:Assessing risks attributable to the prenatal environment

An ever-expanding scientific literature highlights the impact of the prenatal environment on many areas of biology. Across all major farmed species, experimental studies have clearly shown that prenatal experiences can have a substantial impact on outcomes relevant to later health, welfare and productivity. In particular, stress or sub-optimal nutrition experienced by the mother during pregnancy has been shown to have wide-ranging and important effects on how her offspring cope with their social, physical and infectious environment. Variation in the conditions for development provided by the reproductive tract or egg, for instance by altered nutritional supply or hormonal exposure, may therefore explain a large degree of variation in many welfare- and productivity-relevant traits. The scientific literature suggests a number of management practices for pre-birth/hatch individuals that could compromise their later welfare. Such studies may have relevance for the welfare of animals under human care, depending on the extent to which real life conditions involve exposure to these practices. Overall, the findings highlight the importance of extending the focus on animal welfare to include the prenatal period, an aspect which until recently has been largely neglected. © 2012 Universities Federation for Animal Welfare The Old School.

Integrating personality research and animal contest theory:Aggressiveness in the green swordtail xiphophorus helleri

Aggression occurs when individuals compete over limiting resources. While theoretical studies have long placed a strong emphasis on context-specificity of aggression, there is increasing recognition that consistent behavioural differences exist among individuals, and that aggressiveness may be an important component of individual personality. Though empirical studies tend to focus on one aspect or the other, we suggest there is merit in modelling both within- and among-individual variation in agonistic behaviour simultaneously. Here, we demonstrate how this can be achieved using multivariate linear mixed effect models. Using data from repeated mirror trials and dyadic interactions of male green swordtails, Xiphophorus helleri, we show repeatable components of (co)variation in a suite of agonistic behaviour that is broadly consistent with a major axis of variation in aggressiveness. We also show that observed focal behaviour is dependent on opponent effects, which can themselves be repeatable but were more generally found to be context specific. In particular, our models show that within-individual variation in agonistic behaviour is explained, at least in part, by the relative size of a live opponent as predicted by contest theory. Finally, we suggest several additional applications of the multivariate models demonstrated here. These include testing the recently queried functional equivalence of alternative experimental approaches, (e.g., mirror trials, dyadic interaction tests) for assaying individual aggressiveness. © 2011 Wilson et al.

Comprehensive molecular pathology analysis of small bowel adenocarcinoma reveals novel targets with potential for clinical utility

Small bowel accounts for only 0.5% of cancer cases in the US but incidence rates have been rising at 2.4% per year over the past decade. One-third of these are adenocarcinomas but little is known about their molecular pathology and no molecular markers are available for clinical use. Using a retrospective 28 patient matched normal-tumor cohort, next-generation sequencing, gene expression arrays and CpG methylation arrays were used for molecular profiling. Next-generation sequencing identified novel mutations in IDH1, CDH1, KIT, FGFR2, FLT3, NPM1, PTEN, MET, AKT1, RET, NOTCH1 and ERBB4. Array data revealed 17% of CpGs and 5% of RNA transcripts assayed to be differentially methylated and expressed respectively (p < 0.01). Merging gene expression and DNA methylation data revealed CHN2 as consistently hypermethylated and downregulated in this disease (Spearman -0.71, p < 0.001). Mutations in TP53 which were found in more than half of the cohort (15/28) and Kazald1 hypomethylation were both were indicative of poor survival (p = 0.03, HR = 3.2 and p = 0.01, HR = 4.9 respectively). By integrating high-throughput mutational, gene expression and DNA methylation data, this study reveals for the first time the distinct molecular profile of small bowel adenocarcinoma and highlights potential clinically exploitable markers.

An interview with Manuel Salto-Tellez on diagnostic pathology: the future is morphomolecular

Professor Manuel Salto-Tellez of Queen’s University, Belfast, Northern Ireland is an expert histopathologist and molecular diagnostician. Professor Salto-Tellez is a lead investigator at the Northern Ireland Molecular Pathology Laboratory and also serves as a member of the Editorial Advisory Board for Expert Review of Molecular Diagnostics. In this interview, he proposes directions for the future of molecular pathology and molecular diagnostics, integrating all aspects of pathology toward a common goal.

Guidance for laboratories performing molecular pathology for cancer patients

Molecular testing is becoming an important part of the diagnosis of any patient with cancer. The challenge to laboratories is to meet this need, using reliable methods and processes to ensure that patients receive a timely and accurate report on which their treatment will be based. The aim of this paper is to provide minimum requirements for the management of molecular pathology laboratories. This general guidance should be augmented by the specific guidance available for different tumour types and tests. Preanalytical considerations are important, and careful consideration of the way in which specimens are obtained and reach the laboratory is necessary. Sample receipt and handling follow standard operating procedures, but some alterations may be necessary if molecular testing is to be performed, for instance to control tissue fixation. DNA and RNA extraction can be standardised and should be checked for quality and quantity of output on a regular basis. The choice of analytical method(s) depends on clinical requirements, desired turnaround time, and expertise available. Internal quality control, regular internal audit of the whole testing process, laboratory accreditation, and continual participation in external quality assessment schemes are prerequisites for delivery of a reliable service. A molecular pathology report should accurately convey the information the clinician needs to treat the patient with sufficient information to allow for correct interpretation of the result. Molecular pathology is developing rapidly, and further detailed evidence-based recommendations are required for many of the topics covered here.

Alzheimer's disease-like pathology has transient effects on the brain and blood metabolome

The pathogenesis of Alzheimer's disease (AD) is complex involving multiple contributing factors. The extent to which AD pathology impacts upon the metabolome is still not understood, nor is it known how disturbances change as the disease progresses. For the first time we have profiled longitudinally (6, 8, 10, 12 and 18 months) both the brain and plasma metabolome of APP/PS1 double transgenic and wild type (WT) mice. A total of 187 metabolites were quantified using a targeted metabolomics methodology. Multivariate statistical analysis produced models that distinguished APP/PS1 from WT mice at 8, 10 and 12 months.Metabolic pathway analysis found perturbed polyamine metabolism in both brain and blood plasma. There were other disturbances in essential amino acids,branched chain amino acids and also in the neurotransmitter serotonin.Pronounced imbalances in phospholipid and acylcarnitine homeostasis was evident in two age groups. AD-like pathology therefore impacts greatly on both the brain and blood metabolomes, although there appears to be a clear temporal sequence whereby changes to brain metabolites precede those in blood.

Prying into the intimate secrets of animal lives; software beyond hardware for comprehensive annotation in 'Daily Diary' tags

BACKGROUND: Smart tags attached to freely-roaming animals recording multiple parameters at infra-second rates are becoming commonplace, and are transforming our understanding of the way wild animals behave. Interpretation of such data is complex and currently limits the ability of biologists to realise the value of their recorded information.

DESCRIPTION: This work presents Framework4, an all-encompassing software suite which operates on smart sensor data to determine the 4 key elements considered pivotal for movement analysis from such tags (Endangered Species Res 4: 123-37, 2008). These are; animal trajectory, behaviour, energy expenditure and quantification of the environment in which the animal moves. The program transforms smart sensor data into dead-reckoned movements, template-matched behaviours, dynamic body acceleration-derived energetics and position-linked environmental data before outputting it all into a single file. Biologists are thus left with a single data set where animal actions and environmental conditions can be linked across time and space.

CONCLUSIONS: Framework4 is a user-friendly software that assists biologists in elucidating 4 key aspects of wild animal ecology using data derived from tags with multiple sensors recording at high rates. Its use should enhance the ability of biologists to derive meaningful data rapidly from complex data.