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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.
Equine laminitis: membrane type matrix metalloproteinase-1 (MMP-14) is involved in acute phase onset. Enzymatic separation at the hoof lamellar dermal-epidermal interface may play a role in the development of laminitis and characterising and locating matrix metalloproteinases (MMPs) and their inhibitors (tissue inhibitors of MMPs or TIMPs) in lamellar tissues may further understanding of pathogenesis. Objective: To clone and sequence the cDNA encoding lamellar MMP-14 and TIMP-2, and quantify their transcription in normal and laminitic tissue; and to develop antibody to locate MMP-14 in lamellar tissues. Methods: Tissue samples were obtained from an oligofructose induced model of laminitis. Tot...
Molecular cloning and characterization of the alphaX subunit from CD11c/CD18 horse integrin. This work reports the cloning and sequence determination of the horse alpha subunit of the integrin CD11c/CD18, a marker of dendritic cells. A cDNA clone of 4582 base pairs was obtained. It encodes a protein segment of 1086 amino acid residues of the extracellular domain with 10 potential sites of glycosylation, a transmembrane domain of 32 residues and a C-terminal cytoplasmic tail of 24 residues. A phylogenetic analysis of this integrin shows close similarity (83%) with that of Canis familiaris.
In vitro cell compatibility and antibacterial activity of microencapsulated doxycycline designed for improved localized therapy of septic arthritis. For the treatment of septic arthritis in large animals, the local application of antibiotics as a slow release system may be an appropriate means to reach high local bioactivity and low systemic side effects and drug residues. In this study, doxycycline microspheres were developed and tested in vitro for their drug-release properties, suitability for intra-articular application and antimicrobial activity. Methods: The development of a slow release system was achieved by microencapsulation of the drug into poly(lactide-co-glycolide) microspheres by a novel ultrasonic atomization method. Drug el...
Detection of secondary biomarker of met-eGH as a strategy to screen for somatotropin misuse in horseracing. Since the Australian commercialisation of the recombinant equine growth hormone (reGH) in 1998 (EquiGen-5), Bresagen), this reGH, which differs only from eGH by an additional methionine at the N-terminal end (met-eGH), is worldwide suspected to be administered to racehorses as a doping agent. Indeed, the use of this biological drug is considered as a threat to horseracing since it acts both on growth, development or reproductive functions, and on the improvement of performances. In this work, we describe two reliable techniques based on surface plasmon resonance biosensor immunoassay (SPR-BIA)...
Liposome-mediated uptake of exogenous DNA by equine spermatozoa and applications in sperm-mediated gene transfer. Sperm-mediated gene transfer has been reported as a method for production of transgenic animals in a variety of species, and this technique represents a possible method for production of transgenic equids. Objective: To evaluate the uptake of exogenous DNA (enhanced green fluorescent protein; pEGFP) by equine spermatozoa and to assess the ability of transfected spermatozoa to introduce this transgene into early equine embryos. Methods: To evaluate incorporation of pEGFP into equine spermatozoa, washed spermatozoa were incubated with 32P-pEGFP, with or without lipofection. Spermatozoa were also...
Construction of a recombinant plasmid as reaction control in routine PCR for detection of contagious equine metritis (CEM-PCR). Contagious equine metritis (CEM) is a highly contagious bacterial venereal disease of horses caused by Taylorella equigenitalis. CEM-PCR is a semi-nested PCR method for detecting this bacterium. Although this technique is regarded as a sensitive diagnostic method for CEM, there are risks of it generating false positive and false negative results. In this study, we constructed a recombinant plasmid (CEM-POS) as reaction control to assure adequate PCR reaction and prevent false positive results caused by contamination of the reaction control in routine CEM-PCR examinations. CEM-POS was construct...
Cloning, expression and purification of envelope proteins E1 and E2 of western equine encephalitis virus and potential use of them as antigens in immunoassays. The genes encoding envelope proteins E1 and E2 of western equine encephalitis virus (WEEV) were respectively cloned into a prokaryotic T7 RNA polymerase-regulated expression vector. The recombinant C-terminal 6xHis-tagged WEEV E1 and E2 were expressed in bacteria as inclusion bodies that were subsequently solubilized with 8M urea, purified by immobilized metal ion affinity chromatography and finally refolded using an arginine system. The purified 6xHis-tagged proteins showed 50kDa bands as revealed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, consistent with the expected size...
Analysis of factor VIII mediated suppression of lentiviral vector titres. Effective gene therapy for haemophilia A necessitates a vector system that is not subject to a pre-existing immune response, has adequate coding capacity, gives long-term expression and preferably can target non-dividing cells. Vector systems based on lentiviruses such as equine infectious anaemia virus (EIAV) fulfil these criteria for the delivery of factor VIII (FVIII). We have found that B domain-deleted (BDD) FVIII protein inhibits functional viral particle production when co-expressed with the EIAV vector system. Although particle numbers (as measured by reverse transcriptase activity) ar...
Tissue engineering: chondrocyte culture on type 1 collagen support. Cytohistological and immunohistochemical study. The scope of our study is to evaluate the possibility of cultivating and expanding human chondrocytes and seeding them on pure equine type I collagen support. Our results show that human articular cartilaginous cells can multiply and grow on type I collagen substrate with production of extracellular matrix. This type of chondrocyte culture on a support can be used for repairing cartilaginous lesions since they show a correct morphology (evaluated by cytological and histological methods) and a suitable differentiation and phenotype as shown by Alcian PAS staining to indicate the presence of muc...
Glanders: off to the races with Burkholderia mallei. Burkholderia mallei, the etiologic agent of the disease known as glanders, is primarily a disease affecting horses and is transmitted to humans by direct contact with infected animals. The use of B. mallei as a biological weapon has been reported and currently, there is no vaccine available for either humans or animals. Despite the history and highly infective nature of B. mallei, as well as its potential use as a bio-weapon, B. mallei research to understand the pathogenesis and the host responses to infection remains limited. Therefore, this minireview will focus on current efforts to elucida...
Cloning and structural analysis of equine platelet endothelial cell adhesion molecule (PECAM, CD31) and vascular cell adhesion molecule-1 (VCAM-1, CD106). Platelet endothelial cell adhesion molecule (PECAM, CD31) and vascular cell adhesion molecule-1 (VCAM-1, CD106) are essential for leukocyte emigration and diapedesis. PECAM is an essential histologic marker of endothelial cells; VCAM-1 is a prototype marker for endothelial cell activation. In this study, equine PECAM and VCAM mRNA were cloned and sequenced. Both genes are highly conserved amongst several species. This study also revealed conserved structural and regulatory motifs, emphasizing the importance of these genes' physiological roles in immunological responses.
Development of a fluorescent-microsphere immunoassay for detection of antibodies specific to equine arteritis virus and comparison with the virus neutralization test. The development and validation of a microsphere immunoassay (MIA) to detect equine antibodies to the major structural proteins of equine arteritis virus (EAV) are described. The assay development process was based on the cloning and expression of genes for full-length individual major structural proteins (GP5 amino acids 1 to 255 [GP5(1-255)], M(1-162), and N(1-110)), as well as partial sequences of these structural proteins (GP5(1-116), GP5(75-112), GP5(55-98), M(88-162), and N(1-69)) that constituted putative antigenic regions. Purified recombinant viral proteins expressed in Escherichia col...
Expression of cell-surface antigens and embryonic stem cell pluripotency genes in equine blastocysts. Embryonic stem-like (ES-like) cells have now been derived from the inner cell mass (ICM) of horse embryos at the blastocyst stage. Because they have been shown to express cell-surface antigens found in both human and mouse ES cells, the present study investigated gene expression patterns in day-7 horse blastocysts from which the horse ES-like cells had been derived originally. The genes studied included Oct-4, stage-specific embryonic antigen-1 (SSEA-1), SSEA-3, SSEA-4, tumor rejection antigen-1-60 (TRA-1-60), TRA-1-81, and alkaline phosphatase activity, and whereas all three of the SSEA antig...
Equine umbilical cord blood contains a population of stem cells that express Oct4 and differentiate into mesodermal and endodermal cell types. Mesenchymal stem cells (MSCs) offer promise as therapeutic aids in the repair of tendon, ligament, and bone damage suffered by sport horses. The objective of the study was to identify and characterize stem-like cells from newborn foal umbilical cord blood (UCB). UCB was collected and MSC isolated using human reagents. The cells exhibit a fibroblast-like morphology and express the stem cell markers Oct4, SSEA-1, Tra1-60 and Tra1-81. Culture of the cells in tissue-specific differentiation media leads to the formation of cell types characteristic of mesodermal and endodermal origins. Chondrogenic...
Establishment of a novel equine cell line for isolation and propagation of equine herpesviruses. In the present study, an equine-derived cell line was established by transfecting primary fetal horse kidney (FHK) cells with expression plasmid encoding simian virus 40 (SV40) large T antigen and then cloning them by limiting dilution. The cloned cell line, named FHK-Tcl3, grew well and could be propagated over 30 times by splitting them 1:3. Equine herpesvirus (EHV)-1 and EHV-4 replicated well in FHK-Tcl3. EHV-2 and EHV-4 were isolated from samples collected from horses in the field using FHK-Tcl3, and EHV-3 also propagated in FHK-Tcl3. These results indicated that this novel cell line, FHK-...
Isolation and characterization of bone marrow-derived equine mesenchymal stem cells. To isolate and characterize bone marrow-derived equine mesenchymal stem cells (MSCs) for possible future therapeutic applications in horses. Methods: Equine MSCs were isolated from bone marrow aspirates obtained from the sternum of 30 donor horses. Methods: Cells were cultured in medium (alpha-minimum essential medium) with a fetal calf serum content of 20%. Equine MSC features were analyzed to determine selfrenewing and differentiation capacity. For potential therapeutic applications, the migratory potential of equine MSCs was determined. An adenoviral vector was used to determine the transdu...
A hidden Markov model-based stride segmentation technique applied to equine inertial sensor trunk movement data. Inertial sensors are now sufficiently small and lightweight to be used for the collection of large datasets of both humans and animals. However, processing of these large datasets requires a certain degree of automation to achieve realistic workloads. Hidden Markov models (HMMs) are widely used stochastic pattern recognition tools and enable classification of non-stationary data. Here we apply HMMs to identify and segment into strides, data collected from a trunk-mounted six degrees of freedom inertial sensor in galloping Thoroughbred racehorses. A data set comprising mixed gait sequences from...
Estimation of genetic parameters and prediction of breeding values for multivariate threshold and continuous data in a simulated horse population using Gibbs sampling and residual maximum likelihood. Simulated horse data were used to compare multivariate estimation of genetic parameters and prediction of breeding values (BV) for categorical, continuous and molecular genetic data using linear animal models via residual maximum likelihood (REML) and best linear unbiased prediction (BLUP) and mixed linear-threshold animal models via Gibbs sampling (GS). Simulation included additive genetic values, residuals and fixed effects for one continuous trait, liabilities of four binary traits, and quantitative trait locus (QTL) effects and genetic markers with different recombination rates and polymor...
A comparison between freezing methods for the cryopreservation of stallion spermatozoa. The effects of sperm freezing concentration (40 x 10(6)mL(-1) vs. 400 x 10(6)mL(-1)), straw size (0.25 mL vs. 0.5 mL) and freezing method (liquid nitrogen vapour in a Styrofoam box vs. programmable freezing machine) were evaluated in a 2 x 2 x 2 factorial experimental design using 3 split ejaculates from each of 4 stallions. Immediately after thawing, the total motility and forward progressive motility of spermatozoa frozen at a concentration of 40 x 10(6)mL(-1) was higher than for spermatozoa frozen at 400 x 10(6)mL(-1). No significant differences were observed in the semen parameters assesse...
Molecular diagnosis of certain nematode infections can save life and beauty, and preserve breeds of socially relevant and sporting animals. The recognition that the health and welfare of some humans are improved through contact and relationships with animals is now established. Two commonly recognized assistance animals are dogs and horses. Both provide therapeutic benefits to humans with some physical and mental illnesses and both assist people with disabilities. Moreover, the public and scientific attention to the health and conservation of many animal breeds is also increasing worldwide. In the past few years, two potentially life-threatening nematode infections that can induce tumours or tumour-like masses in canids and equids...
The judgement of Solomon and journal status. In the preceding 2 issues, March and May, editorials were presented
regarding the generality of peer review and the relationship of
authors to readership. In this issue, the outcome of peer review and
content of the Journal is considered as a logical end-point of
endeavours of authors and editors. Science is a hard taskmaster for
both of these participants; it is about what is; and not about what
would be nice, desirable or, even, applicable. The content of any
scientific Journal is presented in order to advance knowledge and is
thereby constrained by correspondingly strict disciplines...
Characterization of the equine glycogen debranching enzyme gene (AGL): Genomic and cDNA structure, localization, polymorphism and expression. Glycogen debranching enzyme (AGL) is a multifunctional enzyme acting in the glycogen degradation pathway. In humans, the AGL activity deficiency causes a type III glycogen storage disease (Cori-Forbes disease). One particularity of AGL gene expression lies in the multiple alternative splicing in its 5' region. The AGL gene was localized on ECA5q14-q15. The sequence of the equine cDNA was determined to be 7.5 kb in length with an open reading frame of 4602 bp. The gene is 69 kb long and contains 35 exons. The equine AGL gene has an ubiquitous expression and presents five tissue-dependent cDNA v...
Purification of equine IgG using membrane based enhanced hybrid bioseparation technique: a potential method for manufacturing hyperimmune antibody. Hyperimmune equine IgG is widely used as antivenom and anti-rabies agents. This article discusses a membrane based enhanced hybrid bioseparation technique for efficient and scalable purification of equine immunoglobulin G (IgG) from horse serum. This technique is an improved version of a standard hybrid bioseparation technique developed within our group earlier for fractionation of human plasma proteins (Ghosh. 2004. J Membr Sci 237: 109-117). In the presence of a high antichaotropic salt concentration, equine IgG is selectively and reversibly captured within a stirred cell membrane module fro...
Cell phenotypic variation in normal and damaged tendons. Injuries to tendons are common in both human athletes as well as in animals, such as the horse, which are used for competitive purposes. Furthermore, such injuries are also increasing in prevalence in the ageing, sedentary population. Tendon diseases often respond poorly to treatment and require lengthy periods of rehabilitation. The tendon has a unique extracellular matrix, which has developed to withstand the mechanical demands of such tensile-load bearing structures. Following injury, any repair process is inadequate and results in tissue that is distinct from original tendon tissue. There ...
Multilineage differentiation potential of equine blood-derived fibroblast-like cells. Tissue engineering (TE) has emerged as a promising new therapy for the treatment of damaged tissues and organs. Adult stem cells are considered as an attractive candidate cell type for cell-based TE. Mesenchymal stem cells (MSC) have been isolated from a variety of tissues and tested for differentiation into different cell lineages. While clinical trials still await the use of human MSC, horse tendon injuries are already being treated with autologous bone marrow-derived MSC. Given that the bone marrow is not an optimal source for MSC due to the painful and risk-containing sampling procedure, i...
Characterization and differentiation of equine umbilical cord-derived matrix cells. Stem cells are being evaluated in numerous human clinical trials and are commercially used in veterinary medicine to treat horses and dogs. Stem cell differentiation, homing to disease sites, growth and cytokine factor modulation, and low antigenicity contribute to their therapeutic success. Bone marrow and adipose tissue are the two most common sources of adult-derived stem cells in animals. We report on the existence of an alternative source of primitive, multipotent stem cells from the equine umbilical cord cellular matrix (Wharton's jelly). Equine umbilical cord matrix (EUCM) cells can be ...
Stem cells in veterinary medicine–attempts at regenerating equine tendon after injury. Stem cells have evoked considerable excitement in the animal-owning public because of the promise that stem cell technology could deliver tissue regeneration for injuries for which natural repair mechanisms do not deliver functional recovery and for which current therapeutic strategies have minimal effectiveness. This review focuses on the current use of stem cells within veterinary medicine, whose practitioners have used mesenchymal stem cells (MSCs), recovered from either bone marrow or adipose tissue, in clinical cases primarily to treat strain-induced tendon injury in the horse. The backgr...
Ruthenium anticancer drugs and proteins: a study of the interactions of the ruthenium(III) complex imidazolium trans-[tetrachloro(dimethyl sulfoxide)(imidazole)ruthenate(III)] with hen egg white lysozyme and horse heart cytochrome c. The interactions with protein targets of the ruthenium(III) complex imidazolium trans-[tetrachloro(dimethyl sulfoxide)(imidazole)ruthenate(III)], NAMI-A, an effective anticancer and antimetastatic agent now in clinical trials, deserve great attention as they are believed to be at the basis of the mechanism of action of this innovative molecule. Here, we report on the reactions of NAMI-A with two well-known model proteins, namely, hen egg white lysozyme and horse heart cytochrome c; these reactions were investigated by a variety of physicochemical methods, including optical spectroscopy, (1)H N...
Production of cloned horse foals using roscovitine-treated donor cells and activation with sperm extract and/or ionomycin. We evaluated the effect of different activation treatments on the production of blastocysts and foals by nuclear transfer. Donor cells were prepared using roscovitine treatment, which has previously been associated with increased production of viable offspring. All activation treatments were followed by culture in 6-dimethylaminopurine (6-DMAP) for 4 h. In experiment 1, blastocyst production after activation by injection of sperm extract followed by treatment with ionomycin was significantly higher than that for activation with a serial treatment of ionomycin, 6-DMAP, and ionomycin (12.5 vs 2....
