Cyclosporin A and steroid therapy in sixty-six cadaver kidney recipients

From nine to 18 months ago, 66 patients were given 67 randomly matched cadaveric kidneys with cyclosporin A and steroid therapy. Nine of the recipients were undergoing retransplantation. The over-all kidney survival rate to date has been 77.6 per cent, and 78.8 per cent of the recipients are dialysis-free. The patient mortality in this learning phase was 13.3 per cent. Nephrotoxicity, hepatotoxicity and other side-effects of cyclosporin A could usually be dealt with by dosage adjustments, making feasible the chronic use of this agent. One B-cell immunoblastic sarcoma was encountered which was monoclonal. It was not responsible for death. Another patient had a perforation of the intestine from a lymphoproliferative reaction in which the B cells were polyclonal. After jejunal resection a year ago, there were no further complications. This lesion was not classified as a lymphoma. Both lymphoproliferative lesions were associated with a rise in antibody to viral capsid antigen of Epstein-Barr virus. Results of this study have verified the effectiveness and relative safety of cyclosporin A with steroids for immunosuppression in human recipients of cadaveric kidneys

Transcriptional adaptations following exercise in Thoroughbred horse skeletal muscle highlights molecular mechanisms that lead to muscle hypertrophy

<p>Abstract</p> <p>Background</p> <p>Selection for exercise-adapted phenotypes in the Thoroughbred racehorse has provided a valuable model system to understand molecular responses to exercise in skeletal muscle. Exercise stimulates immediate early molecular responses as well as delayed responses during recovery, resulting in a return to homeostasis and enabling long term adaptation. Global mRNA expression during the immediate-response period has not previously been reported in skeletal muscle following exercise in any species. Also, global gene expression changes in equine skeletal muscle following exercise have not been reported. Therefore, to identify novel genes and key regulatory pathways responsible for exercise adaptation we have used equine-specific cDNA microarrays to examine global mRNA expression in skeletal muscle from a cohort of Thoroughbred horses (<it>n = </it>8) at three time points (before exercise, immediately post-exercise, and four hours post-exercise) following a single bout of treadmill exercise.</p> <p>Results</p> <p>Skeletal muscle biopsies were taken from the <it>gluteus medius </it>before (T₀), immediately after (T₁) and four hours after (T₂) exercise. Statistically significant differences in mRNA abundance between time points (T₀<it>vs </it>T₁and T₀<it>vs </it>T₂) were determined using the empirical Bayes moderated <it>t</it>-test in the Bioconductor package Linear Models for Microarray Data (LIMMA) and the expression of a select panel of genes was validated using real time quantitative reverse transcription PCR (qRT-PCR). While only two genes had increased expression at T₁(<it>P </it>< 0.05), by T₂932 genes had increased (<it>P </it>< 0.05) and 562 genes had decreased expression (<it>P </it>< 0.05). Functional analysis of genes differentially expressed during the recovery phase (T₂) revealed an over-representation of genes localized to the actin cytoskeleton and with functions in the MAPK signalling, focal adhesion, insulin signalling, mTOR signaling, p53 signaling and Type II diabetes mellitus pathways. At T₁, using a less stringent statistical approach, we observed an over-representation of genes involved in the stress response, metabolism and intracellular signaling. These findings suggest that protein synthesis, mechanosensation and muscle remodeling contribute to skeletal muscle adaptation towards improved integrity and hypertrophy.</p> <p>Conclusions</p> <p>This is the first study to characterize global mRNA expression profiles in equine skeletal muscle using an equine-specific microarray platform. Here we reveal novel genes and mechanisms that are temporally expressed following exercise providing new knowledge about the early and late molecular responses to exercise in the equine skeletal muscle transcriptome.</p

RNA Sequencing (RNA-Seq) Reveals Extremely Low Levels of Reticulocyte-Derived Globin Gene Transcripts in Peripheral Blood From Horses (Equus caballus) and Cattle (Bos taurus)

RNA-seq has emerged as an important technology for measuring gene expression in peripheral blood samples collected from humans and other vertebrate species. In particular, transcriptomics analyses of whole blood can be used to study immunobiology and develop novel biomarkers of infectious disease. However, an obstacle to these methods in many mammalian species is the presence of reticulocyte-derived globin mRNAs in large quantities, which can complicate RNA-seq library sequencing and impede detection of other mRNA transcripts. A range of supplementary procedures for targeted depletion of globin transcripts have, therefore, been developed to alleviate this problem. Here, we use comparative analyses of RNA-seq data sets generated from human, porcine, equine, and bovine peripheral blood to systematically assess the impact of globin mRNA on routine transcriptome profiling of whole blood in cattle and horses. The results of these analyses demonstrate that total RNA isolated from equine and bovine peripheral blood contains very low levels of globin mRNA transcripts, thereby negating the need for globin depletion and greatly simplifying blood-based transcriptomic studies in these two domestic species

Tracing Cattle Breeds with Principal Components Analysis Ancestry Informative SNPs

The recent release of the Bovine HapMap dataset represents the most detailed survey of bovine genetic diversity to date, providing an important resource for the design and development of livestock production. We studied this dataset, comprising more than 30,000 Single Nucleotide Polymorphisms (SNPs) for 19 breeds (13 taurine, three zebu, and three hybrid breeds), seeking to identify small panels of genetic markers that can be used to trace the breed of unknown cattle samples. Taking advantage of the power of Principal Components Analysis and algorithms that we have recently described for the selection of Ancestry Informative Markers from genomewide datasets, we present a decision-tree which can be used to accurately infer the origin of individual cattle. In doing so, we present a thorough examination of population genetic structure in modern bovine breeds. Performing extensive cross-validation experiments, we demonstrate that 250-500 carefully selected SNPs suffice in order to achieve close to 100% prediction accuracy of individual ancestry, when this particular set of 19 breeds is considered. Our methods, coupled with the dense genotypic data that is becoming increasingly available, have the potential to become a valuable tool and have considerable impact in worldwide livestock production. They can be used to inform the design of studies of the genetic basis of economically important traits in cattle, as well as breeding programs and efforts to conserve biodiversity. Furthermore, the SNPs that we have identified can provide a reliable solution for the traceability of breed-specific branded products

A rapid and sensitive intracellular flow cytometric assay to identify Theileria parva infection within target cells

Theileria parva is an intracellular protozoan parasite transmitted by ticks that causes a fatal lymphoproliferative disease of cattle known as East Coast Fever. Vaccination against the disease currently relies on inoculation of the infective sporozoite stage of the parasite and simultaneous treatment with long-acting formulations of oxytetracycline. Sporozoites are maintained as frozen stabilates of triturated infected ticks and the method requires accurate titration of stabilates to determine appropriate dose rates. Titration has traditionally been undertaken in cattle and requires large numbers of animals because of individual variation in susceptibility to infection. An alternative tissue culture-based method is laborious and time consuming. We have developed a flow cytometric method for quantifying the infectivity of sporozoite stabilates in vitro based on the detection of intracellular parasite antigen. The method allows clear identification of parasitized cells with a high degree of sensitivity and specificity. Analysis of infected cells between 48 and 72 h post-infection clearly defines the potential transforming capability of different stabilates

Genome sequencing of the extinct Eurasian wild aurochs, Bos primigenius, illuminates the phylogeography and evolution of cattle

Background Domestication of the now-extinct wild aurochs, Bos primigenius, gave rise to the two major domestic extant cattle taxa, B. taurus and B. indicus. While previous genetic studies have shed some light on the evolutionary relationships between European aurochs and modern cattle, important questions remain unanswered, including the phylogenetic status of aurochs, whether gene flow from aurochs into early domestic populations occurred, and which genomic regions were subject to selection processes during and after domestication. Here, we address these questions using whole-genome sequencing data generated from an approximately 6,750-year-old British aurochs bone and genome sequence data from 81 additional cattle plus genome-wide single nucleotide polymorphism data from a diverse panel of 1,225 modern animals. Results Phylogenomic analyses place the aurochs as a distinct outgroup to the domestic B. taurus lineage, supporting the predominant Near Eastern origin of European cattle. Conversely, traditional British and Irish breeds share more genetic variants with this aurochs specimen than other European populations, supporting localized gene flow from aurochs into the ancestors of modern British and Irish cattle, perhaps through purposeful restocking by early herders in Britain. Finally, the functions of genes showing evidence for positive selection in B. taurus are enriched for neurobiology, growth, metabolism and immunobiology, suggesting that these biological processes have been important in the domestication of cattle. Conclusions This work provides important new information regarding the origins and functional evolution of modern cattle, revealing that the interface between early European domestic populations and wild aurochs was significantly more complex than previously thought

Location of the CD8 T Cell Epitope within the Antigenic Precursor Determines Immunogenicity and Protection against the Toxoplasma gondii Parasite

CD8 T cells protect the host from disease caused by intracellular pathogens, such as the Toxoplasma gondii (T. gondii) protozoan parasite. Despite the complexity of the T. gondii proteome, CD8 T cell responses are restricted to only a small number of peptide epitopes derived from a limited set of antigenic precursors. This phenomenon is known as immunodominance and is key to effective vaccine design. However, the mechanisms that determine the immunogenicity and immunodominance hierarchy of parasite antigens are not well understood.Here, using genetically modified parasites, we show that parasite burden is controlled by the immunodominant GRA6-specific CD8 T cell response but not by responses to the subdominant GRA4- and ROP7-derived epitopes. Remarkably, optimal processing and immunodominance were determined by the location of the peptide epitope at the C-terminus of the GRA6 antigenic precursor. In contrast, immunodominance could not be explained by the peptide affinity for the MHC I molecule or the frequency of T cell precursors in the naive animals. Our results reveal the molecular requirements for optimal presentation of an intracellular parasite antigen and for eliciting protective CD8 T cells. © 2013 Feliu et al

Immunization with one Theileria parva strain results in similar level of CTL strain-specificity and protection compared to immunization with the three-component Muguga cocktail in MHC-matched animals

Abstract Background The tick-borne protozoan parasite Theileria parva causes a usually fatal cattle disease known as East Coast fever in sub-Saharan Africa, with devastating consequences for poor small-holder farmers. Immunity to T. parva, believed to be mediated by a cytotoxic T lymphocyte (CTL) response, is induced following natural infection and after vaccination with a live vaccine, known as the Infection and Treatment Method (ITM). The most commonly used version of ITM is a combination of parasites derived from three isolates (Muguga, Kiambu 5 and Serengeti-transformed), known as the “Muguga cocktail”. The use of a vaccine comprising several strains is believed to be required to induce a broad immune response effective against field challenge. In this study we investigated whether immunization with the Muguga cocktail induces a broader CTL response than immunization with a single strain (Muguga). Results Four MHC haplotype-matched pairs of cattle were immunized with either the trivalent Muguga cocktail or the single Muguga strain. CTL specificity was assessed on a panel of five different strains, and clonal responses to these strains were also assessed in one of the MHC-matched pairs. We did not find evidence for a broader CTL response in animals immunized with the Muguga cocktail compared to those immunized with the Muguga strain alone, in either the bulk or clonal CTL analyses. This was supported by an in vivo trial in which all vaccinated animals survived challenge with a lethal dose of the Muguga cocktail vaccine stabilate. Conclusion We did not observe any substantial differences in the immunity generated from animals immunized with either Muguga alone or the Muguga cocktail in the animals tested here, corroborating earlier results showing limited antigenic diversity in the Muguga cocktail. These results may warrant further field studies using single T. parva strains as future vaccine candidates

Large-Scale Mitochondrial DNA Analysis of the Domestic Goat Reveals Six Haplogroups with High Diversity

Background. From the beginning of domestication, the transportation of domestic animals resulted in genetic and demographic processes that explain their present distribution and genetic structure. Thus studying the present genetic diversity helps to better understand the history of domestic species. Methodology/Principal Findings. The genetic diversity of domestic goats has been characterized with 2430 individuals from all over the old world, including 946 new individuals from regions poorly studied until now (mainly the Fertile Crescent). These individuals represented 1540 haplotypes for the HVI segment of the mitochondrial DNA (mtDNA) control region. This large-scale study allowed the establishment of a clear nomenclature of the goat maternal haplogroups. Only five of the six previously defined groups of haplotypes were divergent enough to be considered as different haplogroups. Moreover a new mitochondrial group has been localized around the Fertile Crescent. All groups showed very high haplotype diversity. Most of this diversity was distributed among groups and within geographic regions. The weak geographic structure may result from the worldwide distribution of the dominant A haplogroup (more than 90% of the individuals). The large-scale distribution of other haplogroups (except one), may be related to human migration. The recent fragmentation of local goat populations into discrete breeds is not detectable with mitochondrial markers. The estimation of demographic parameters from mismatch analyses showed that all groups had a recent demographic expansion corresponding roughly to the period when domestication took place. But even with a large data set it remains difficult to give relative dates of expansion for different haplogroups because of large confidence intervals. Conclusions/Significance. We propose standard criteria for the definition of the different haplogroups based on the result of mismatch analysis and on the use of sequences of reference. Such a method could be also applied for clarifying the nomenclature of mitochondrial haplogroups in other domestic species

Genome-wide transcriptional profiling of peripheral blood leukocytes from cattle infected with Mycobacterium bovis reveals suppression of host immune genes

Background Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB), a pathological infection with significant economic impact. Recent studies have highlighted the role of functional genomics to better understand the molecular mechanisms governing the host immune response to M. bovis infection. Furthermore, these studies may enable the identification of novel transcriptional markers of BTB that can augment current diagnostic tests and surveillance programmes. In the present study, we have analysed the transcriptome of peripheral blood leukocytes (PBL) from eight M. bovis-infected and eight control non-infected age-matched and sex-matched Holstein-Friesian cattle using the Affymetrix® GeneChip® Bovine Genome Array with 24,072 gene probe sets representing more than 23,000 gene transcripts. Results Control and infected animals had similar mean white blood cell counts. However, the mean number of lymphocytes was significantly increased in the infected group relative to the control group (P = 0.001), while the mean number of monocytes was significantly decreased in the BTB group (P = 0.002). Hierarchical clustering analysis using gene expression data from all 5,388 detectable mRNA transcripts unambiguously partitioned the animals according to their disease status. In total, 2,960 gene transcripts were differentially expressed (DE) between the infected and control animal groups (adjusted P-value threshold ≤ 0.05); with the number of gene transcripts showing decreased relative expression (1,563) exceeding those displaying increased relative expression (1,397). Systems analysis using the Ingenuity® Systems Pathway Analysis (IPA) Knowledge Base revealed an over-representation of DE genes involved in the immune response functional category. More specifically, 64.5% of genes in the affects immune response subcategory displayed decreased relative expression levels in the infected animals compared to the control group. Conclusions This study demonstrates that genome-wide transcriptional profiling of PBL can distinguish active M. bovis-infected animals from control non-infected animals. Furthermore, the results obtained support previous investigations demonstrating that mycobacterial infection is associated with host transcriptional suppression. These data support the use of transcriptomic technologies to enable the identification of robust, reliable transcriptional markers of active M. bovis infection.This work was supported by Investigator Grants from Science Foundation Ireland (Nos: SFI/01/F.1/B028 and SFI/08/IN.1/B2038), a Research Stimulus Grant from the Department of Agriculture, Fisheries and Food (No: RSF 06 405) and a European Union Framework 7 Project Grant (No: KBBE-211602-MACROSYS). KEK is supported by the Irish Research Council for Science, Engineering and Technology (IRCSET) funded Bioinformatics and Systems Biology PhD Programme http://bioinfo-casl.ucd.ie/PhD