Application of the cDNA-AFLP method for studying gene expression in Fibrobacter succinogenes S85 exposed to 134.2 kHz electromagnetic field

AbstractMany biological effects related to the exposure of cells and tissues to electromagnetic fields have been reported in the literature, including those influencing DNAs and RNAs structure and ..

Exercise-induced up-regulation of MMP-1 and IL-8 genes in endurance horses

<p>Abstract</p> <p>Background</p> <p>The stress response is a critical factor in the training of equine athletes; it is important for performance and for protection of the animal against physio-pathological disorders.</p> <p>In this study, the molecular mechanisms involved in the response to acute and strenuous exercise were investigated using peripheral blood mononuclear cells (PBMCs).</p> <p>Results</p> <p>Quantitative real-time PCR (qRT-PCR) was used to detect modifications in transcription levels of the genes for matrix metalloproteinase-1 (<it>MMP-1</it>) and interleukin 8 (<it>IL-8</it>), which were derived from previous genome-wide expression analysis. Significant up-regulation of these two genes was found in 10 horses that had completed a race of 90–120 km in a time-course experimental design.</p> <p>Conclusion</p> <p>These results suggest that <it>MMP-1 </it>and <it>IL-8 </it>are both involved in the exercise-induced stress response, and this represents a starting point from which to understand the adaptive responses to this phenomenon.</p

Exercise induced stress in horses: Selection of the most stable reference genes for quantitative RT-PCR normalization

<p>Abstract</p> <p>Background</p> <p>Adequate stress response is a critical factor during athlete horses' training and is central to our capacity to obtain better performances while safeguarding animal welfare.</p> <p>In order to investigate the molecular mechanisms underlying this process, several studies have been conducted that take advantage of microarray and quantitative real-time PCR (qRT-PCR) technologies to analyse the expression of candidate genes involved in the cellular stress response.</p> <p>Appropriate application of qRT-PCR, however, requires the use of reference genes whose level of expression is not affected by the test, by general physiological conditions or by inter-individual variability.</p> <p>Results</p> <p>The expression of nine potential reference genes was evaluated in lymphocytes of ten endurance horses during strenuous exercise. These genes were tested by qRT-PCR and ranked according to the stability of their expression using three different methods (implemented in <it>geNorm</it>, <it>NormFinder </it>and <it>BestKeeper</it>). Succinate dehydrogenase complex subunit A (<it>SDHA</it>) and hypoxanthine phosphoribosyltransferase (<it>HPRT</it>) always ranked as the two most stably expressed genes. On the other hand, glyceraldehyde-3-phosphate dehydrogenase (<it>GAPDH</it>), transferrin receptor (<it>TFRC</it>) and ribosomal protein L32 (<it>RPL32</it>) were constantly classified as the less reliable controls.</p> <p>Conclusion</p> <p>This study underlines the importance of a careful selection of reference genes for qRT-PCR studies of exercise induced stress in horses. Our results, based on different algorithms and analytical procedures, clearly indicate <it>SDHA </it>and <it>HPRT </it>as the most stable reference genes of our pool.</p

Immune Response in Young Thoroughbred Racehorses under Training

Training has a great impact on the physiology of an athlete and, like all stressful stimuli, can trigger an innate immune response and inflammation, which is part of a wider coping strategy of the host to restore homeostasis. The Thoroughbred racehorse is a valid animal model to investigate these changes thanks to its homogeneous training and highly selected genetic background. The aim of this study was to investigate modifications of the innate immune response and inflammation in young untrained Thoroughbred racehorses during the first training season through haematological and molecular investigations. Twenty-nine Thoroughbred racehorses were followed during their incremental 3-month sprint exercise schedule. Blood collection was performed at time 0 (T0; before starting the intense training period), 30 days after T0 (T30), and 90 days after T0 (T90). Haematological parameters (red and white blood cells, haemoglobin, and platelets) were evaluated and haematocrit (HCT), mean corpuscular haemoglobin concentration (MCHC), and red cells width distribution + standard deviation (RDW-SD) were calculated. Moreover, via RT-qPCR, we investigated the expression of, Interleukin 1β (IL-1β), Interleukin 4 (IL-4) Interleukin 6 (IL-6), Interleukin 2 (IL-2), Interleukin 3 (IL-3), Interleukin 5 (IL-5) Interleukin 8 (IL-8), Trasformig Growth Factor β and α (TGF-β), Tumor necrosis factor α (TNF-α), and Interferon γ (IFN-γ)genes. Main corpuscular volume (MCV) showed a significant (p = 0.008) increase at T90. Main corpuscular haemoglobin (MCH) and haemoglobin concentration (MCHC) values were significantly augmented at both T30 (p &lt; 0.001) and T90 (p &lt; 0.001). Basophils were significant increased at T30 (p = 0.02) and eosinophils were significantly increased at T90 (p = 0.03). Significant differences in gene expression were found for all the genes under study, with the exception of IFN-γ and TNF-α. In particular, IL-2 (T30, p = 0.011; T90, p = 0.015), IL-4 (T30, p = 0.009; T90, p &lt; 0.001), and IL-8 (T30, p &lt; 0.001; T90, p &lt; 0.001) genes were significantly upregulated at both T30 and T90 with respect to T0, TGF-β was intensely downregulated at T30 (p &lt; 0.001), IL-5 gene expression was significantly decreased at T90 (p = 0.001), while IL-1β (p = 0.005) and IL-3 (p = 0.001) expression was strongly augmented at the same time. This study highlighted long-term adjustments of O2 transport capability that can be reasonably traced back to exercise adaptation. Moreover, the observed changes of granulocyte numbers and functions and inflammatory cytokine gene expression confirm a major role of the innate immune system in the response to the complex of stressful stimuli experienced during the training period

Transcriptome Analysis of Canine Cutaneous Melanoma and Melanocytoma Reveals a Modulation of Genes Regulating Extracellular Matrix Metabolism and Cell Cycle

Interactions between tumor cells and tumor microenvironment are considered critical in carcinogenesis, tumor invasion and metastasis. To examine transcriptome changes and to explore the relationship with tumor microenvironment in canine cutaneous melanocytoma and melanoma, we extracted RNA from formalin-fixed, paraffin-embedded (FFPE) specimens and analyzed them by means of RNA-seq for transcriptional analysis. Melanocytoma and melanoma samples were compared to detect differential gene expressions and significant enriched pathways were explored to reveal functional relations between differentially expressed genes. The study demonstrated a differential expression of 60 genes in melanomas compared to melanocytomas. The differentially expressed genes cluster in the extracellular matrix-receptor interaction, protein digestion and absorption, focal adhesion and PI3K-Akt (phosphoinositide 3-kinase/protein kinase B) signaling pathways. Genes encoding for several collagen proteins were more commonly differentially expressed. Results of the RNA-seq were validated by qRT-PCR and protein expression of some target molecules was investigated by means of immunohistochemistry. We hypothesize that the developing melanoma actively promotes collagen metabolism and extracellular matrix remodeling as well as enhancing cell proliferation and survival contributing to disease progression and metastasis. In this study, we also detected unidentified genes in human melanoma expression studies and uncover new candidate drug targets for further testing in canine melanoma

Polysynovitis in a horse due to Borrelia burgdorferi sensu lato infection – Case study

Lyme borreliosis (LB) is a multi-systemic tick-borne disease affecting both humans and animals, including horses, and is caused by a group of interrelated spirochetes classified within the Borrelia burgdorferi sensu lato (s.l.) complex. Despite the high reported seroprevalence in the European equine population for B. burgdorferi s.l., to-date no documented clinical cases have been described. A 6-year-old Paint gelding was referred with a history of three weeks of fever, intermittent lameness and digital flexor tendon sheath effusion of the right hind limb. Based on a strict diagnostic protocol, which included serological tests for infectious diseases and molecular investigations, a final diagnosis was made of polysynovitis due to B. burgdorferi s.l. infection. An unreported aspect observed in this case was the absence of the pathogen DNA in two of the affected joints. To the authors' knowledge, the case described represents the first documented clinical case of equine LB in Italy. Moreover, the absence of pathogen DNA in two of the affected joints observed in this case revealed a possible similarity with the same condition described in humans, where an immunomediated pathogenesis for arthropathy due to B. burgdorferi s.l. infection is suspected. Since humans and horses share the same habitat, this report supports the role of the horse as potential sentinel for human biological risk

Gene co-expression networks in liver and muscle transcriptome reveal sex-specific gene expression in lambs fed with a mix of essential oils

Background: Essential oil (EO) dietary supplementation is a new strategy to improve animal health. EO compounds have antiparasitic, antimicrobial, antiviral, antimycotic, antioxidant and anti-inflammatory proprieties. Nutrigenomics investigations represent innovative approaches in understanding the relation between diet effect and gene expression related to the animal performance. Few nutrigenomics studies have used a high-throughput RNA-Sequencing (RNA-Seq) approach, despite great potential of RNA-Seq data in gene expression quantification and in co-expression network analyses. Our aim is to use the potential of RNA-Sequencing data in order to evaluate the effect of an EO supplementary diet on gene expression in both lamb liver and muscle. Results: Using a treatment and sex interaction model, 13 and 4 differentially expressed genes were identified in liver and muscle respectively. Sex-specific differentially expressed (DE) genes were identified in both sexes. Using network based analysis, different clusters of co-expressed genes that were highly correlated to the diet were detected in males vs. females, in agreement with DE analysis. A total of five regulatory genes in liver tissue associated to EO diet were identified: DNAJB9, MANF, UFM1, CTNNLA1 and NFX1. Our study reveals a sex-dependent effect of EO diet in both tissues, and an influence on the expression of genes mainly involved in immune, inflammatory and stress pathway. Conclusion: Our analysis suggests a sex-dependent effect of the EO dietary supplementation on the expression profile of both liver and muscle tissues. We hypothesize that the presence of EOs could have beneficial effects on wellness of male lamb and further analyses are needed to understand the biological mechanisms behind the different effect of EO metabolites based on sex. Using lamb as a model for nutrigenomics studies, it could be interesting to investigate the effects of EO diets in other species and in humans

Exploring the Italian equine gene pool via high-throughput genotyping

Introduction: The Italian peninsula is in the center of the Mediterranean area, and historically it has been a hub for numerous human populations, cultures, and also animal species that enriched the hosted biodiversity. Horses are no exception to this phenomenon, with the peculiarity that the gene pool has been impacted by warfare and subsequent “colonization”. In this study, using a comprehensive dataset for almost the entire Italian equine population, in addition to the most influential cosmopolitan breeds, we describe the current status of the modern Italian gene pool.Materials and Methods: The Italian dataset comprised 1,308 individuals and 22 breeds genotyped at a 70 k density that was merged with publicly available data to facilitate comparison with the global equine diversity. After quality control and supervised subsampling to ensure consistency among breeds, the merged dataset with the global equine diversity contained data for 1,333 individuals from 54 populations. Multidimensional scaling, admixture, gene flow, and effective population size were analyzed.Results and Discussion: The results show that some of the native Italian breeds preserve distinct gene pools, potentially because of adaptation to the different geographical contexts of the peninsula. Nevertheless, the comparison with international breeds highlights the presence of strong gene flow from renowned breeds into several Italian breeds, probably due to historical introgression. Coldblood breeds with stronger genetic identity were indeed well differentiated from warmblood breeds, which are highly admixed. Other breeds showed further peculiarities due to their breeding history. Finally, we observed some breeds that exist more on cultural, traditional, and geographical point of view than due to actual genetic distinctiveness

Why make the effort? Exploring Recovery College Engagement

Purpose - Whilst there is growing evidence to suggest that the Recovery College (RC) environment supports students towards their mental health recovery (Meddings et al., 2015b), students’ initial motivations for engagement, alongside factors that may hinder or support attendance, have yet to be exclusively explored. Design - All new RC students were invited to take part in a semi-structured interview three months following their enrolment. Four participants completed an interview which were later analysed using Thematic Analysis. Findings - Four themes emerged within analysis: Making the effort; Being “too unwell”; Friendly Environment and Glad I came. These are discussed alongside the literature, and it is proposed that whilst there is a substantial struggle involved in engagement with a RC, likely related to mental health and social factors, the RC environment, peer support and support of the tutors helps students to overcome the impact of this. Research limitations / implications - Due to the small sample size and exploratory stance of this study, additional research into the complexities around engagement with RCs is strongly recommended. Only students who had attended at least one RC course chose to participate in this study, therefore an under-researched population of non-attendees may provide a valuable contribution to further understanding. Originality / value - This is one of the first studies to qualitatively explore factors which may support, or hinder, initial and ongoing engagement with a RC. It is proposed that a greater understanding of these important issues could be used to increase RC accessibility and inclusion