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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.
Ultrasound B-scan image simulation, segmentation, and analysis of the equine tendon. The hypothesis is that an imaging technique based on decompression and segmentation of B-scan images with morphological operators can provide a measurement of the integrity of equine tendons. Methods: Two complementary approaches were used: (i) Simulation of B-scan images to better understand the relationship between image properties and their underlying biological structural contents and (ii) extraction and quantification from B-scan images of tendon structures identified in step (i) to diagnose the status of the superficial digital flexor tendon (SDFT) by using the proposed imaging technique...
Identification of differentially expressed genes associated with osteochondrosis in standardbred horses using RNA arbitrarily primed PCR. The aim of this study was to investigate genes for differential expression in cartilage of foals predisposed to osteochondrosis (OC). Tissue was sampled from the cranial part of the distal intermediate ridge of the tibia in the tarso-crural joint. Foals were considered predisposed to OC when parents had OC at the distal intermediate ridge of the tibia. RNA was isolated and subjected to arbitrarily primed PCR (RAP-PCR) followed by fingerprinting to screen for differentially expressed genes. By verification of results from the RAP-PCR fingerprint screening using real-time RT-PCR, we identified t...
Molecular cloning and characterization of equine thymic stromal lymphopoietin. Thymic stromal lymphopoietin (TSLP) is a novel cytokine that plays a central role in T helper 2 (Th2) cell differentiation and allergic inflammation. It is predominantly expressed by epithelial cells, and its expression is increased in patients with atopic dermatitis and asthma. Mice overexpressing TSLP in the skin develop allergic dermatitis and mice overexpressing TSLP in lungs develop asthma-like disease. However, it is not known whether TSLP plays an important role in equine allergies. Therefore, we cloned and sequenced the complete translated region of equine TSLP gene and measured its ex...
Examination of equine glandular stomach lesions for bacteria, including Helicobacter spp by fluorescence in situ hybridisation. The equine glandular stomach is commonly affected by erosion and ulceration. The aim of this study was to assess whether bacteria, including Helicobacter, could be involved in the aetiology of gastric glandular lesions seen in horses. Results: Stomach lesions, as well as normal appearing mucosa were obtained from horses slaughtered for human consumption. All samples were tested for urease activity using the Pyloritek assay, while mucosal bacterial content was evaluated using Fluorescence In Situ Hybridisation. In selected sub samples, bacteria characterisation was pursued further by cloning an...
Cloning, comparative sequence analysis and mRNA expression of calcium-transporting genes in horses. Epithelial calcium transport occurs by paracellular and transcellular mechanisms. Transcellular transport in intestinal and renal epithelia involves several transport proteins, including transient receptor potential vanilloid member 5 (TRPV5), member 6 (TRPV6), calbindin D9k (CB9), calbindin D28k (CB28), sodium calcium exchanger 1 (NCX1), plasma membrane calcium ATPase 1 (PMCA1), and the vitamin D receptor (VDR). We are interested in the horse because of its unique calcium physiology (high blood calcium, high intestinal calcium absorption, high renal excretion of calcium, low vitamin D concent...
Comparative efficacy of dermal fibroblast-mediated and direct adenoviral bone morphogenetic protein-2 gene therapy for bone regeneration in an equine rib model. Cell-mediated and direct adenoviral (Ad) vector gene therapies can induce bone regeneration, including dermal fibroblasts (DFbs). We compared two effective therapies, DFb-mediated and direct Ad vector delivery of bone morphogenetic protein-2 (BMP2), for relative efficacy in bone regeneration. Equine rib drill defects were treated by percutaneous injection of either DFb-BMP2 or an Ad-BMP2 vector. At week 6, both DFb-BMP2- and Ad-BMP2-treated rib defects had greater bone filling volume and mineral density, with DFb-BMP2 inducing greater bone volume and maturity in the cortical bone aspect of the...
Expression, purification and monoclonal antibodies preparation of recombinant equine mature interleukin-18. IL-18 is a cytokine originally discovered as an important modulator of immune responses and subsequently shown to be pleiotropic. In this report, we expressed the recombinant equine mature interleukin-18 (rEMIL-18) in E. coli and purified it by nickel affinity gel column chromatography. Purified rEMIL-18 had biological activity commensurate with recombinant human IL-18, as determined by its synergistic effect with recombinant human IL-12 (rhIL-12) on the induction of IFN-gamma gene expression in equine peripheral blood mononuclear cells (PBMC). Following intraperitoneal (i.p.) immunization of ...
Characterization of a stretch-activated potassium channel in chondrocytes. Chondrocytes possess the capacity to transduce load-induced mechanical stimuli into electrochemical signals. The aim of this study was to functionally characterize an ion channel activated in response to membrane stretch in isolated primary equine chondrocytes. We used patch-clamp electrophysiology to functionally characterize this channel and immunohistochemistry to examine its distribution in articular cartilage. In cell-attached patch experiments, the application of negative pressures to the patch pipette (in the range of 20-200 mmHg) activated ion channel currents in six of seven patches. ...
Isolation and cultivation of equine corneal keratocytes, fibroblasts and myofibroblasts. To establish an in vitro model for the investigation of equine corneal wound healing. To accomplish this goal, a protocol to isolate and culture equine corneal keratocytes, fibroblasts and myofibroblasts was developed. ANIMAL MATERIAL: Equine corneal buttons were aseptically harvested from healthy research horses undergoing humane euthanasia for reasons unrelated to this study. Slit-lamp biomicroscopy was performed prior to euthanasia by a board-certified veterinary ophthalmologist to ensure that all samples were harvested from horses free of anterior segment disease. Methods: Equine corneal s...
Adult equine bone marrow stromal cells produce a cartilage-like ECM mechanically superior to animal-matched adult chondrocytes. Our objective was to evaluate the age-dependent mechanical phenotype of bone marrow stromal cell- (BMSC-) and chondrocyte-produced cartilage-like neo-tissue and to elucidate the matrix-associated mechanisms which generate this phenotype. Cells from both immature (2-4 month-old foals) and skeletally-mature (2-5 year-old adults) mixed-breed horses were isolated from animal-matched bone marrow and cartilage tissue, encapsulated in self-assembling-peptide hydrogels, and cultured with and without TGF-beta1 supplementation. BMSCs and chondrocytes from both donor ages were encapsulated with high viab...
Gene-based vaccines: Recent developments. Gene-based vaccines are under development for a broad variety of applications, ranging from vaccines to immunotherapies for infectious diseases, cancer, autoimmune diseases and allergy. In addition, following the licensing of DNA vaccines for use in fish and horses, and DNA immunotherapy for the treatment of cancer in dogs, several veterinary uses of vaccines have been demonstrated for species ranging from fish and shrimp to cattle and horses. A variety of publications describing preclinical and clinical studies of the technologies used to increase the potency of gene-based vaccines, and resea...
Clinical update on the use of mesenchymal stem cells in equine orthopaedics. Stem cells have received much attention in recent times because of their potential to improve healing of othropaedic problems. This manuscript presents the genesis, issues and current state of stem cell treatment in equine medicine. Current literature supports the use of mesenchymal stem cells (MSCs) for treatment of orthopaedic problems.
An inactivated Vero cell-grown Japanese encephalitis vaccine formulated with Advax, a novel inulin-based adjuvant, induces protective neutralizing antibody against homologous and heterologous flaviviruses. Advax is a polysaccharide-based adjuvant that potently stimulates vaccine immunogenicity without the increased reactogenicity seen with other adjuvants. This study investigated the immunogenicity of a novel Advax-adjuvanted Vero cell culture candidate vaccine against Japanese encephalitis virus (JEV) in mice and horses. The results showed that, in mice, a two-immunization, low-dose (50 ng JEV antigen) regimen with adjuvanted vaccine produced solid neutralizing immunity comparable to that elicited with live ChimeriVax-JE immunization and superior to that elicited with tenfold higher doses of a ...
Contribution of each of four Superantigens to Streptococcus equi-induced mitogenicity, gamma interferon synthesis, and immunity. Streptococcus equi is the causative agent of strangles, the most frequently diagnosed infectious disease of horses worldwide. The disease is characterized by abscessation and swelling of the lymph nodes of the head and neck, which can literally strangle the horse to death. S. equi produces four recently acquired phage-associated bacterial superantigens (sAgs; SeeH, SeeI, SeeL, and SeeM) that share homology with the mitogenic toxins of Streptococcus pyogenes. The aim of this study was to characterize the contribution of each of these S. equi sAgs to mitogenic activity in vitro and quantify the ...
Cytochrome P450 1A-dependent activities in deer, cattle and horses. The objective of this study was to investigate and characterize the metabolic activities of CYP1A in deer, cattle and horses in comparison to those of rats using ethoxyresorufin O-deethylation (EROD) and methoxyresorufin O-demethylation (MROD) assays. We performed an inhibition study for these activities using anti-rat CYP1A1 antibody and identified that these activities were due to the CYP1A subfamily. Interspecies differences in the CYP1A-dependent activities were highly observed in this study. In particular, we found that the horse had the highest EROD and MROD activities among the examined...
Equine embryos and embryonic stem cells: defining reliable markers of pluripotency. Cartilage and tendon injuries are a significant source of animal wastage and financial loss within the horse-racing industry. Moreover, both cartilage and tendon have limited intrinsic capacity for self-repair, and the functionally inferior tissue produced within a lesion may reduce performance and increase the risk of reinjury. Stem cells offer tremendous potential for accelerating and improving tissue healing, and adult mesenchymal stem cells (MSCs) are already used to treat cartilage and tendon injuries in horses. However, MSCs are scarce in the bone marrow isolates used, have limited poten...
Detection of prohibited animal products in livestock feeds by single-strand conformation polymorphism analysis. Single-strand conformation polymorphism (SSCP) analysis of amplicons produced from a mitochondrial DNA region between the tRNA(Lys) and ATPase8 genes was applied for the detection of animal product within livestock feeds. Identification of prohibited animal (cattle, elk, sheep, deer, and goat) and nonprohibited animal (pig and horse) products from North America was possible based on the differential display of the single-stranded DNA fragments for the different animal species on SSCP gels. This method allowed specific detection and identification of mixed genomic DNA from different animal spec...
Evaluation of early cellular influences of bone morphogenetic proteins 12 and 2 on equine superficial digital flexor tenocytes and bone marrow-derived mesenchymal stem cells in vitro. To evaluate early cellular influences of bone morphogenetic protein (BMP)12 and BMP2 on equine superficial digital flexor tenocytes (SDFTNs) and equine bone marrow-derived mesenchymal stem cells (BMDMSCs). Methods: 9 adult clinically normal horses. Methods: BMDMSCs and SDFTNs were cultured in monolayer, either untreated or transduced with adenovirus encoding green fluorescent protein, adenovirus encoding BMP12, or adenovirus encoding BMP2. Cytomorphologic, cytochemical, immunocytochemical, and reverse transcriptase-quantitative PCR (RT-qPCR) analyses were performed on days 3 and 6. Genetic pro...
Automated counting of nucleated cells in equine synovial fluid without and with hyaluronidase pretreatment. Microscopy is usually used to obtain manual total and differential cell counts in equine synovial fluid. A faster, more precise method is desirable. Objective: The objectives were to compare an automated impedance method with a manual method for obtaining total and differential cell counts in equine synovial fluid and to evaluate the effect of pretreatment with hyaluronidase on automated results. Methods: Synovial fluid samples (n=48) were collected into EDTA and analyzed within 48 hours. Automated total and differential cell counts were evaluated using a Medonic CA620-VET hematology analyzer ...
Tissue engineering in wound repair: the three “R”s–repair, replace, regenerate. Horses are predisposed to traumatic wounds that can be labor intensive and expensive to manage. Skin has a considerable potential for efficient and functional repair however, while cutaneous repair is a regenerative process in the fetus, this capability declines in late gestation as inflammation and scarring alter the outcome of healing. The historical gold standard for replacement of lost skin is the autologous skin graft. However, the horse's lack of redundant donor skin limits the practicality of full-thickness grafting to smaller wounds; moreover, graft failure is relatively common in equi...
Comparison of direct and indirect methods of intra-abdominal pressure measurement in normal horses. To develop a direct method for measuring intra-abdominal pressures in the standing horse, identify a reference interval for direct intra-abdominal pressures, compare these pressures to indirect intra-abdominal pressures measured from the bladder, and determine the optimal bladder infusion volume for indirect pressure measurement. Methods: Prospective, experimental study. Methods: A university-based equine research facility. Methods: Ten healthy adult horses, 5 males and 5 females. Methods: Direct intra-abdominal pressures were measured through an intraperitoneal cannula and zeroed at the heigh...
Evaluation of shape variability of stallion sperm heads by means of image analysis and Fourier descriptors. This study quantified and evaluated the variability of sperm head shape for 10 different stallions. Sperm head shape characteristics including sperm head length to width ratio, position of the center of gravity, curvature, and degree of roundness were assessed and analysed from images using elliptic Fourier descriptors and inverse Fourier transformation. The first four principal components accounted for 88.46-92.33% of the total variance and provided a good summary of the overall data. In the case of the ejaculate with defective sperm heads the components accounted for 97.35-98.21% of variatio...
Identification of horse chestnut coat color genotype using SNaPshot. The Cantabrian Coast horse breeds of the Iberian Peninsula have mainly black or bay colored coats, but alleles responsible for a chestnut coat color run in these breeds and occasionally, chestnut horses are born. Chestnut coat color is caused by two recessive alleles, e and e(a), of the melanocortin-1 receptor gene, whereas the presence of the dominant, wild-type E allele produces black or bay coat horses. Because black or bay colored coats are considered as the purebred phenotype for most of the breeds from this region, it is important to have a fast and reliable method to detect alleles caus...
Development of a blocking ELISA using a recombinant glycoprotein for the detection of antibodies to vesicular stomatitis New Jersey virus. A recombinant glycoprotein (R-GP) of vesicular stomatitis New Jersey virus (VSV-NJ) was expressed in insect cells by a baculovirus system. Its utility as a diagnostic antigen in a blocking ELISA was investigated as an alternative to the current native GP extracted from VSV-NJ. With the cut-off value of 73% inhibition, the R-GP ELISA exhibited 99.1% specificity for naive sera from cattle and horses. It did not cross-react with VSV-Indiana (VSV-IN) positive sera and differentiated from foot-and-mouth disease and swine vesicular disease. Taken together, this is the first report that the R-GP has ...
Selection of reference genes for quantitative real-time PCR in equine in vivo and fresh and frozen-thawed in vitro blastocysts. Application of reverse transcription quantitative real-time polymerase chain reaction is very well suited to reveal differences in gene expression between in vivo and in vitro produced embryos. Ultimately, this may lead to optimized equine assisted reproductive techniques. However, for a correct interpretation of the real-time PCR results, all data must be normalized, which is most reliably achieved by calculating the geometric mean of the most stable reference genes. In this study a set of reliable reference genes was identified for equine in vivo and fresh and frozen-thawed in vitro embryos....
Identification of variables that optimize isolation and culture of multipotent mesenchymal stem cells from equine umbilical-cord blood. OBJECTIVE-To optimize the isolation and culture of mesenchymal stem cells (MSCs) from umbilical-cord blood (UCB), identify variables that predicted successful MSC isolation, and determine whether shipping, processing, and cryopreservation altered MSC viability, recovery rates, and expansion kinetics. SAMPLE POPULATION-UCB samples from 79 Thoroughbred and Quarter Horse mares. PROCEDURES-UCB samples were processed to reduce volume and remove RBCs. Nucleated cells (NCs) were cryopreserved or grown in various culture conditions to optimize MSC monolayer expansion and proliferation. Donor and UCB-s...
Detection of treponemes in canker lesions of horses by 16S rRNA clonal sequencing analysis. Equine canker is a chronic pododermatitis of the hoof in horses. Although spirochetes are detectable histopathologically in the lesions, the precise etiology remains unclear. This study reports the 16S rRNA gene sequencing of randomly selected clones based on PCR with Treponema-specific primers, using the canker lesions from two horses and healthy frog and sole from a horse. A total of 114 clones were obtained from the lesions, but no clones were detected in the healthy hoof tissues. The clones from the canker lesions examined were grouped into 19 operational taxonomic units, such as treponema...
Development of a novel equine whole transcript oligonucleotide GeneChip microarray and its use in gene expression profiling of normal articular-epiphyseal cartilage. No large scale equine microarray is available commercially to allow genomic and transcriptional profiling of the majority of genes that would define the genetic basis of equine disease. Objective: To generate a whole transcript target labelled GeneChip to interrogate the equine transcriptome and validate chip performance using RNA samples derived from organs, articular cells and normal cartilage. Methods: Equine mRNA and selected equine gene sequences derived from perfect cross-hybridisation of equine RNA on human microarray GeneChips, were used to design a custom equine gene microarray. Seque...
