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Topic:Biotechnology
Biotechnology in horses encompasses the application of biological techniques and tools to enhance equine health, performance, and reproduction. This field includes genetic engineering, cloning, and the development of vaccines and therapeutics tailored to equine physiology. Techniques such as gene editing and stem cell therapy are explored for their potential to address genetic disorders, improve tissue regeneration, and enhance disease resistance in horses. Additionally, advancements in reproductive biotechnology, such as artificial insemination and embryo transfer, contribute to genetic diversity and breeding efficiency. This page compiles peer-reviewed research studies and scholarly articles that investigate the applications, methodologies, and implications of biotechnology in equine science.
The use of electrochemically activated saline as a uterine instillation in pony mares. Twelve pony mares were randomly assigned to either a control or a treatment group and inseminated with fresh, raw semen from a single stallion of known fertility in a cross-over trial design. Pregnancy was diagnosed by transrectal ultrasound 12-14 days post-ovulation and then terminated by administration of a luteolytic dose of cloprostenol. Treatment mares received a uterine instillation of 100 ml of electrochemically activated (ECA) saline 4-12 hours post-insemination. Control mares received no treatment post-insemination. Per cycle pregnancy rate was 58.3 % in the control group and 50 % in ...
Differential gene expression of TRPM1, the potential cause of congenital stationary night blindness and coat spotting patterns (LP) in the Appaloosa horse (Equus caballus). The appaloosa coat spotting pattern in horses is caused by a single incomplete dominant gene (LP). Homozygosity for LP (LP/LP) is directly associated with congenital stationary night blindness (CSNB) in Appaloosa horses. LP maps to a 6-cM region on ECA1. We investigated the relative expression of two functional candidate genes located in this LP candidate region (TRPM1 and OCA2), as well as three other linked loci (TJP1, MTMR10, and OTUD7A) by quantitative real-time RT-PCR. No large differences were found for expression levels of TJP1, MTMR10, OTUD7A, and OCA2. However, TRPM1 (Transient Recept...
Bone marrow and lymph node evaluation. Evaluation of equine bone marrow and lymph node samples can provide the definitive diagnosis in some cases, and may provide useful information in other cases. Some newer techniques, including immunophenotyping of cells and clonality assays, provide the capability to more precisely identify cells, both as to origin and malignancy. Use of these techniques on equine bone marrow and lymph node samples, and compiling of the data, will eventually provide invaluable information about equine neoplasia that will greatly improve the ability to predict tumor behavior and response to therapy.
Somatic cell nuclear transfer in horses. The cloning of equids was achieved in 2003, several years after the birth of Dolly the sheep and also after the cloning of numerous other laboratory and farm animal species. The delay was because of the limited development in the horse of more classical-assisted reproductive techniques required for successful cloning, such as oocyte maturation and in vitro embryo production. When these technologies were developed, the application of cloning also became possible and cloned horse offspring were obtained. This review summarizes the main technical procedures that are required for cloning equids an...
Complementary DNA cloning, functional expression and characterization of a novel cytochrome P450, CYP2D50, from equine liver. Members of the CYP2D family constitute only about 2-4% of total hepatic CYP450s, however, they are responsible for the metabolism of 20-25% of commonly prescribed therapeutic compounds. CYP2D enzymes have been identified in a number of different species. However, vast differences in the metabolic activity of these enzymes have been well documented. In the horse, the presence of a member of the CYP2D family has been suggested from studies with equine liver microsomes, however its presence has not been definitively proven. In this study a cDNA encoding a novel CYP2D enzyme (CYP2D50) was cloned f...
Novel optical imaging technique to determine the 3-D orientation of collagen fibers in cartilage: variable-incidence angle polarization-sensitive optical coherence tomography. To investigate a novel optical method to determine the three dimensional (3-D) structure of articular cartilage collagen non-destructively. Methods: Polarization-sensitive optical coherence tomography was used to determine the apparent optical birefringence of articular cartilage for a number of different illumination directions. A quantitative method based on the theory of light propagation in uniaxial crystalline materials was validated on equine flexor tendon. Qualitative maps of fiber polar and azimuthal orientation at sites on the posterior and anterior segments of the equine third metaca...
Elucidation of the MD-2/TLR4 interface required for signaling by lipid IVa. LPS signals through a membrane bound-complex of the lipid binding protein MD-2 and the receptor TLR4. In this study we identify discrete regions in both MD-2 and TLR4 that are required for signaling by lipid IVa, an LPS derivative that is an agonist in horse but an antagonist in humans. We show that changes in the electrostatic surface potential of both MD-2 and TLR4 are required in order that lipid IVa can induce signaling. In MD-2, replacing horse residues 57-66 and 82-89 with the equivalent human residues confers a level of constitutive activity on horse MD-2, suggesting that conformational...
A novel application of quantile regression for identification of biomarkers exemplified by equine cartilage microarray data. Identification of biomarkers among thousands of genes arrayed for disease classification has been the subject of considerable research in recent years. These studies have focused on disease classification, comparing experimental groups of effected to normal patients. Related experiments can be done to identify tissue-restricted biomarkers, genes with a high level of expression in one tissue compared to other tissue types in the body. Results: In this study, cartilage was compared with ten other body tissues using a two color array experimental design. Thirty-seven probe sets were identified as...
Factors impacting equine sperm recovery rate and quality following cushioned centrifugation. Two experiments were conducted to investigate modifications in cushioned centrifugation of stallion semen. Specifically, the effects of tube type, centrifugation medium, cushion type, and centrifugation force on post-centrifugation sperm recovery rate and quality were evaluated. In Experiment 1, sperm recovery rate was higher (P<0.05) in conventional plastic conical-bottom tubes (103%) than in newly developed glass nipple-bottom tubes (96%) following cushioned centrifugation; however, several measures of semen quality (i.e., % total motility [MOT], % progressive motility [PMOT], curvilinear...
A commentary on domestic animals as dual-purpose models that benefit agricultural and biomedical research. Research on domestic animals (cattle, swine, sheep, goats, poultry, horses, and aquatic species) at land grant institutions is integral to improving the global competitiveness of US animal agriculture and to resolving complex animal and human diseases. However, dwindling federal and state budgets, years of stagnant funding from USDA for the Competitive State Research, Education, and Extension Service National Research Initiative (CSREES-NRI) Competitive Grants Program, significant reductions in farm animal species and in numbers at land grant institutions, and declining enrollment for graduate...
Gonadotropin-dependent regulation of the prostaglandin E2 receptor in equine preovulatory follicles during the ovulatory process in mares. The objectives of the study were to clone the primary structure of the prostaglandin E2 receptor subtype 2 (PTGER2) cDNA and to characterize its regulation in equine follicles during gonadotropin-induced ovulation. Results from DNA isolation indicated that the equine PTGER2 cDNA encodes a predicted 353-amino acid protein, which is highly similar (76-85%) to known mammalian homologues. The regulation of PTGER2 was studied by semi-quantitative RT-PCR/Southern blot using preparations of theca interna and mural granulosa cells isolated from equine follicles 0-39 hr post-treatment with human chorio...
Electrochemotherapy in veterinary oncology. Electropermeabilization is a method that uses electric field pulses to induce an electrically mediated reorganization of the plasma membrane of cells. Electrochemotherapy combines local or systemic administration of chemotherapeutic drugs such as bleomycin or cisplatin that have poor membrane permeability with electropermeabilization by direct application of electric pulses to the tumors. Preclinical studies have demonstrated excellent antitumor effectiveness of electrochemotherapy on different animal models and various tumor types, minimal toxicity, and safety of the procedure. Based on resul...
The horse pseudoautosomal region (PAR): characterization and comparison with the human, chimp and mouse PARs. The pseudoautosomal region (PAR) is a genomic segment on mammalian sex chromosomes where sequence homology mimics that seen between autosomal homologues. The region is essential for pairing and proper segregation of sex chromosomes during male meiosis. As yet, only human/chimp and mouse PARs have been characterized. The two groups of species differ dramatically in gene content and size of the PAR and therefore do not provide clues about the likely evolution and constitution of PAR among mammals. Here we characterize the equine PAR by i) isolating and arranging 71 BACs containing 129 markers (1...
The cloned equine thyrotropin receptor is hypersensitive to human chorionic gonadotropin; identification of three residues in the extracellular domain involved in ligand specificity. The receptors for TSH, LH/chorionic gonadotropin (CG), and FSH belong to the same subfamily of G protein-coupled receptors. The specificity of recognition of their cognate hormone involves a limited number of residues in the leucine-rich repeats present in the N-terminal ectodomain of the receptor. It is admitted that receptors of this subfamily coevoluted with their respective ligands. The secretion of CG is restricted to gestation of primates and Equidae. We hypothesized that, facing the challenge of a new hormone, the glycoprotein hormone receptors would have evolved differently in Equidae ...
Estrogenic activity of the equine estrogen metabolite, 4-methoxyequilenin. Oxidative metabolism of estrogens has been associated with genotoxicity. O-methylation of catechol estrogens is considered as a protective mechanism. 4-Methoxyequilenin (4-MeOEN) is the O-methylated product of 4-hydroxyequilenin (4-OHEN). 4-OHEN, the major catechol metabolite of the equine estrogens present in the most widely prescribed hormone replacement therapeutics, causes DNA damage via quinone formation. In this study, estrogen receptor (ERa) binding of 4-MeOEN was compared with estradiol (E2) and equilenin derivatives including 4-BrEN using computer modeling, estrogen response element (...
Developments in stallion semen evaluation. The conventional approach to evaluation of stallion semen dates back several decades, and includes evaluation of spermatozoal concentration, semen volume, spermatozoon morphological characteristics, and spermatozoal motility patterns initially and following in-vitro storage. While an analysis performed in this manner does have predictive value, incorporation of some more newly developed techniques may improve the predictive value of the examination. This communication addresses some newer tests that can be applied today for evaluation of semen, as well some tests that may be available in the c...
[Construction of an infectious clone of equine infectious anemia virus by N-glycosylation reverse-mutations]. To elucidate the role of N-glycosylation in fetal donkey dermal cell (FDD)-attenuated equine infectious anemia virus (EIAV), we constructed an N-glycosylation reverse-mutation molecular clone, pLGN191N236N246. This viral molecular clone was derived from the infectious clone pLGFD3-8 by site-directed mutagenesis. This clone was used to transfect fetal donkey dermal (FDD) cells. Infectious characteristics of transfectants were monitored by RT-PCR, indirect immune fluorescence and reverse transcriptase activity assay. After three passages in FDD cells, viral replications in the supernatant of cel...
[Development of sandwich ELISA for equine interferon-gamma detection]. To develop a quantitative ELISA by measuring interferon (IFN-gamma) of equine lymphocytes. Methods: Sandwich ELISA for equine IFN-gamma was developed using mAb A5 as a capture antibody and biotinylated mAb SB10 as a detection antibody. Results: The detection limit of the sandwich ELISA for equine IFN-gamma was 1 microg/L and did not show cross-reactivity with recombinant equine IL-18. Equine IFN-gamma was detected by ELISA in culture medium of the peripheral blood mononuclear cells (PBMCs) stimulated with ConA or PMA/Ionomycin. Conclusions: This method can be used to help understand the role o...
Cryobiological determinants of frozen semen quality, with special reference to stallion. Success in cryopreserving stallion semen has been very variable. Several different freezing regimes have been published. However, because extenders and procedures used in each regime have differed, direct comparison of these techniques has been very difficult, and controlled studies comparing different techniques have not been reported. A number of different factors affect sperm cryosurvival. In this article we review briefly current cryopreservation procedures for stallion semen, and then in more detail cryobiological determinants of sperm function, and mechanisms of cryoinjury and cryoprotec...
A BAC contig map over the proximal approximately 3.3 Mb region of horse chromosome 21. A total of 207 BAC clones containing 155 loci were isolated and arranged into a map of linearly ordered overlapping clones over the proximal part of horse chromosome 21 (ECA21), which corresponds to the proximal half of the short arm of human chromosome 19 (HSA19p) and part of HSA5. The clones form two contigs - each corresponding to the respective human chromosomes - that are estimated to be separated by a gap of approximately 200 kb. Of the 155 markers present in the two contigs, 141 (33 genes and 108 STS) were generated and mapped in this study. The BACs provide a 4-5x coverage of the regio...
Equine clinical cytogenetics: the past and future. Cytogenetic analyses of horses have benefited the horse industry by identifying chromosomal aberrations causing congenital abnormalities, embryonic loss and infertility. Technical advances in cytogenetics enabled the identification of chromosome specific aberrations. More recently, advances in genomic tools have been used to more precisely define chromosome abnormalities. In this report we review the history of equine clinical cytogenetics, identify historical landmarks for equine clinical cytogenetics, discuss how the current use of genomic tools has benefited this area, and how future genomi...
Potential applications of equine genomics in dissecting diseases and fertility. Following the recent development of high-resolution gene maps and generation of several basic tools and resources to use them in analyzing traits that are economically important to horse owners, genome analysis in horses is witnessing a shift towards developing an ability to analyze complex traits. The likelihood of this happening in the very near future is great, mainly because of the recent availability of the whole genome sequence in the horse. The latter has triggered the development of novel tools like SNP-chip and expression arrays that will permit rapid genome-wide analysis. While these...
Recent advances in cooled-semen technology. The majority of horse registries approve the use of artificial insemination, and horse breeding has widely taken benefit from the use of cooled-stored semen. New insights into cooled-semen technology open possibilities to reduce problems such as impaired semen quality after cooled-storage in individual stallions. The stallion itself has major impacts on quality and fertility of cooled-stored semen. Dietary supplementation of antioxidants and polyunsaturated fatty acids improves semen quality in a variety of species, but only few studies on this topic exist in the horse. Proper semen collection...
Equine ANXA2 and MMP1 expression analyses in an experimental model of normal and pathological wound repair. Wounds on horse limbs can develop exuberant granulation tissue which resembles the human keloid. Clues gained from the study of over-scarring in horses might help control fibro-proliferative disorders. Objective: The aim of the present study was to clone full-length equine ANXA2 cDNA then to study spatio-temporal expression of ANXA2 and MMP1 mRNA and protein, potential contributors to remodeling, during repair of body (normal) and limb (fibro-proliferative) wounds in an established horse wound model. Methods: Cloning of ANXA2 was achieved by screening size-selected cDNA libraries. Expression w...
Molecular cloning and characterization of equine Toll-like receptor 9. Innate immunity relies on a series of germline-encoded pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs), to detect conserved microbial components. TLR9 is typically expressed intracellularly in immune cells such as dendritic cells and recognizes unmethylated bacterial or viral cytosine-phosphate-guanine DNA (CpG-DNA). To investigate innate immune responses through TLR9 signaling pathway in horses, we cloned and characterized equine TLR9. Protein sequence analysis shows that equine TLR9 has a typically conserved cytosolic Toll/interleukin-1 receptor (TIR) domain, three l...
Stereological characteristics of the equine accessory nerve. Stereological techniques have been increasingly employed for assessment and characterization of neuromuscular diseases in humans and animals. As an adjunct to histopathology, morphometrical algorithms provide quantitative evidence of the peripheral nerve composition, thereby shedding light on its fibre characteristics and basic electrophysiological properties. In the horse, stereological investigations already have focussed on the recurrent laryngeal, deep peroneal and lateral palmar nerves (LPN). Of these, only the latter is suitable for taking biopsies in clinical settings, however, it does ...
