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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.
Oral administration of transgenic barley expressing a Culicoides allergen induces specific antibody response. Insect bite hypersensitivity is an immunoglobulin (Ig)E-mediated dermatitis of horses initiated by bites of midges of the genus Culicoides. Culicoides spp. are not indigenous to Iceland and the prevalence of insect bite hypersensitivity is much higher in horses born in Iceland and exported as compared to Icelandic horses born in a Culicoides rich environment. Immunotherapy is therefore needed. Objective: The aim of the study was to express an allergen from Culicoides in barley grain and investigate whether an immune response could be obtained in healthy Icelandic horses by oral treatment with ...
The Optimisation of Pseudotyped Viruses for the Characterisation of Immune Responses to Equine Influenza Virus. Pseudotyped viruses (PVs) produced by co-transfecting cells with plasmids expressing lentiviral core proteins and viral envelope proteins are potentially powerful tools for studying various aspects of equine influenza virus (EIV) biology. The aim of this study was to optimise production of equine influenza PVs. Co-transfection of the HAT protease to activate the haemagglutinin (HA) yielded a higher titre PV than TMPRSS2 with the HA from A/equine/Richmond/1/2007 (H3N8), whereas for A/equine/Newmarket/79 (H3N8), both proteases resulted in equivalent titres. TMPRSS4 was ineffective with the HA of...
Implementing an open-access CASA software for the assessment of stallion sperm motility: Relationship with other sperm quality parameters. Setting an open-access computer assisted sperm analysis (CASA) may benefit the evaluation of motility in mammalian sperm, especially when economic constraints do not allow the use of a commercial system. There have been successful attempts to develop such a device in Zebra fish sperm and the system has been used in very few studies on mammalian spermatozoa. Against this background, the present study aimed at developing an open-access CASA system for mammalian sperm using the horse as a model and based upon the Image J software previously established for Zebra fish sperm. Along with determining...
Identification of the immunogenic epitopes of the whole venom component of the Hemiscorpius lepturus scorpion using the phage display peptide library. The venom of the Hemiscorpius lepturus scorpion contains mixtures of bioactive compounds that disturb biochemical and physiological functions of the victims. Hemiscorpius lepturus envenomation is recognized as a serious health concern in tropical regions. So far, there is no preventive procedure, and the main focus is on treatment of victims with an antiserum purified from hyper-immunized horses. Although antisera can neutralize the venom, they, in some cases, lead to anaphylactic shock and even death. Selection of peptides mimicking antigenic and immunogenic epitopes of toxins from random pep...
Identification of Novel Equine (Equus caballus) Tendon Markers Using RNA Sequencing. Although several tendon-selective genes exist, they are also expressed in other musculoskeletal tissues. As cell and tissue engineering is reliant on specific molecular markers to discriminate between cell types, tendon-specific genes need to be identified. In order to accomplish this, we have used RNA sequencing (RNA-seq) to compare gene expression between tendon, bone, cartilage and ligament from horses. We identified several tendon-selective gene markers, and established eyes absent homolog 2 () and a G-protein regulated inducer of neurite outgrowth 3 () as specific tendon markers using RT-...
Isolation and characterization of equine dental pulp stem cells derived from Thoroughbred wolf teeth. Mesenchymal stem cells (MSCs) are adult multipotent stem cells that are capable of self-renewal and differentiation into multiple cell lineages. Methods for cell therapy using MSCs have been developed in equine medicine. Recently, human dental pulp stem cells (DPSCs) have drawn much attention owing to their trophic factor producing ability and minimally invasive collection methods. However, there have been no reports on equine dental pulp-derived cells (eDPCs). Therefore, the aim of this study was to isolate and characterize the eDPCs from discarded wolf teeth. Plastic-adherent spindle-shaped ...
Degradation, intra-articular retention and biocompatibility of monospheres composed of [PDLLA-PEG-PDLLA]-b-PLLA multi-block copolymers. In this study, we investigated the use of microspheres with a narrow particle size distribution ('monospheres') composed of biodegradable poly(DL-lactide)-PEG-poly(DL-lactide)-b-poly(L-lactide) multiblock copolymers that are potentially suitable for local sustained drug release in articular joints. Monospheres with sizes of 5, 15 and 30μm and a narrow particle size distribution were prepared by a micro-sieve membrane emulsification process. During in vitro degradation, less crystallinity, higher swelling and accelerated mass loss during was observed with increasing the PEG content of the poly...
The Lin28 Expression in Stallion Testes. The molecular markers for specific germ cell stages can be utilized for identifying, monitoring, and separating a particular stage of germ cells. The RNA-binding protein Lin28 is expressed in gonocytes of human fetal testes. The Lin28 expression is restricted to a very small population of spermatogonial cells in human, mice, and monkey. The main objective of this study was to investigate the expression pattern of Lin28 in stallion testes at different reproductive stages. Based on the presence or absence of full spermatogenesis and lumina in seminiferous tubules, the testicular samples were cat...
Semi-automated International Cartilage Repair Society scoring of equine articular cartilage lesions in optical coherence tomography images. Arthroscopic optical coherence tomography (OCT) is a promising tool for the detailed evaluation of articular cartilage injuries. However, OCT-based articular cartilage scoring still relies on the operator's visual estimation. Objective: To test the hypothesis that semi-automated International Cartilage Repair Society (ICRS) scoring of chondral lesions seen in OCT images could enhance intra- and interobserver agreement of scoring and its accuracy. Methods: Validation study using equine cadaver tissue. Methods: Osteochondral samples (n = 99) were prepared from 18 equine metacarpophalangeal joint...
Neutral solute transport across osteochondral interface: A finite element approach. Investigation of the solute transfer across articular cartilage and subchondral bone plate could nurture the understanding of the mechanisms of osteoarthritis (OA) progression. In the current study, we approached the transport of neutral solutes in human (slight OA) and equine (healthy) samples using both computed tomography and biphasic-solute finite element modeling. We developed a multi-zone biphasic-solute finite element model (FEM) accounting for the inhomogeneity of articular cartilage (superficial, middle and deep zones) and subchondral bone plate. Fitting the FEM model to the concentra...
Microvesicle-mediated Wnt/β-Catenin Signaling Promotes Interspecies Mammary Stem/Progenitor Cell Growth. Signaling mechanisms that regulate mammary stem/progenitor cell (MaSC) self-renewal are essential for developmental changes that occur in the mammary gland during pregnancy, lactation, and involution. We observed that equine MaSCs (eMaSCs) maintain their growth potential in culture for an indefinite period, whereas canine MaSCs (cMaSCs) lose their growth potential in long term cultures. We then used this system to investigate the role of microvesicles (MVs) in promoting self-renewal properties. We found that Wnt3a and Wnt1 were expressed at higher levels in MVs isolated from eMaSCs compared wi...
In Vitro and In Vivo Development of Horse Cloned Embryos Generated with iPSCs, Mesenchymal Stromal Cells and Fetal or Adult Fibroblasts as Nuclear Donors. The demand for equine cloning as a tool to preserve high genetic value is growing worldwide; however, nuclear transfer efficiency is still very low. To address this issue, we first evaluated the effects of time from cell fusion to activation (<1h, n = 1261; 1-2h, n = 1773; 2-3h, n = 1647) on in vitro and in vivo development of equine embryos generated by cloning. Then, we evaluated the effects of using different nuclear donor cell types in two successive experiments: I) induced pluripotent stem cells (iPSCs) vs. adult fibroblasts (AF) fused to ooplasts injected with the pluripotency-inducin...
Breakthroughs in Equine Embryo Cryopreservation. Most equine embryos are collected from the donor mare and transferred immediately as fresh embryos or shipped cooled to a recipient station for transfer within 24 hours. Very few equine embryos are frozen despite the numerous advantages of embryo cryopreservation. There are 2 major hurdles: Only the small embryos (<300 μm) provide good pregnancy rates after freezing/thawing and transfer. Also there is no good procedure for superovulating mares; thus, extra embryos for freezing are not readily available. Using either a slow cool or a vitrification method, pregnancy rates of small equine e...
Advances in Stallion Semen Cryopreservation. The use of stallion frozen semen minimizes the spread of disease, eliminates geographic barriers, and preserves the genetic material of the animal for an unlimited time. Significant progress on the frozen thawed stallion semen process and consequently fertility has been achieved over the last decade. These improvements not only increased fertility rates but also allowed cryopreservation of semen from "poor freezers." This article reviews traditional steps and new strategies for stallion semen handling and processing that are performed to overcome the deleterious effects of semen preservation a...
Defining Trends in Global Gene Expression in Arabian Horses with Cerebellar Abiotrophy. Equine cerebellar abiotrophy (CA) is a hereditary neurodegenerative disease that affects the Purkinje neurons of the cerebellum and causes ataxia in Arabian foals. Signs of CA are typically first recognized either at birth to any time up to 6Â months of age. CA is inherited as an autosomal recessive trait and is associated with a single nucleotide polymorphism (SNP) on equine chromosome 2 (13074277G>A), located in the fourth exon of TOE1 and in proximity to MUTYH on the antisense strand. We hypothesize that unraveling the functional consequences of the CA SNP using RNA-seq will elucidate the m...
Isolated effects of external bath osmolality, solute concentration, and electrical charge on solute transport across articular cartilage. The metabolic function of cartilage primarily depends on transport of solutes through diffusion mechanism. In the current study, we use contrast enhanced micro-computed tomography to determine equilibrium concentration of solutes through different cartilage zones and solute flux in the cartilage, using osteochondral plugs from equine femoral condyles. Diffusion experiments were performed with two solutes of different charge and approximately equal molecular weight, namely iodixanol (neutral) and ioxaglate (charge=-1) in order to isolate the effects of solute's charge on diffusion. Furthermore,...
Equine hepatocytes: isolation, cryopreservation, and applications to in vitro drug metabolism studies. Despite reports of the successful isolation of primary equine hepatocytes, there are no published data regarding the successful cryopreservation of these isolated cells. In this study, a detailed description of the procedures for isolation, cryopreservation, and recovery of equine hepatocytes are presented. Furthermore, the intrinsic clearance (Cl) and production of metabolites for three drugs were compared between freshly isolated and recovered cryopreserved hepatocytes. Primary equine hepatocytes were isolated using a two-step collagenase perfusion method, with an average cell yield of 2.47Â...
Characterization and angiogenic potential of xenogeneic bone grafting materials: Role of periodontal ligament cells. Adequate revascularization is a prerequisite for successful healing of periodontal bone defects. This study characterized three different xenogeneic bone grafting materials: Gen-Os of equine and porcine origins, and anorganic Bio-Oss. We also investigated their angiogenic potential. All materials were composed of poorly crystalline calcium oxide phosphate, with Bio-Oss exhibiting a carbonated phase and larger particle size and both Gen-Os showing the presence of collagen. Both Gen-Os materials significantly enhanced vascular endothelial growth factor (VEGF) secretion by PDL cells. A significan...
Isolation of endothelial colony-forming cells from blood samples collected from the jugular and cephalic veins of healthy adult horses. OBJECTIVE To evaluate optimal isolation of endothelial colony-forming cells (ECFCs) from peripheral blood of horses. SAMPLE Jugular and cephalic venous blood samples from 17 adult horses. PROCEDURES Each blood sample was divided; isolation was performed with whole blood adherence (WBA) and density gradient centrifugation (DGC). Isolated cells were characterized by uptake of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate-labeled acetylated low-density lipoprotein (DiI-Ac-LDL), vascular tubule formation, and expression of endothelial (CD34, CD105, vascular endothelial growth ...
Beta-mercaptoethanol supplementation of in vitro maturation medium does not influence nuclear and cytoplasmic maturation of equine oocytes. In vitro embryo production in the horse is still not as efficient as in other species. Oxidative stress negatively affects oocyte and embryo culture. To attenuate/minimize the oxidative stress, antioxidants such as low-molecular thiol compounds can be added to culture media. Beta-mercaptoethanol (BME) has been shown to improve maturation and embryo development in different species. The aim of this study was to investigate whether the addition to maturation medium of BME at common (0.1Â mM) and high (0.7Â mM) concentration could improve oocyte maturation also in the horse. Equine oocytes recove...
Detection of single nucleotide polymorphisms (SNP) in equine coat color genes using SNaPshotTM multiplex kit or pluronic F-108 tri-block copolymer and capillary electrophoresis. Molecular methods for the detection of mammalian coat color phenotypes have expanded greatly within the past decade. Many phenotypes are associated with a single nucleotide polymorphism mutation in the genetic sequence. Traditionally, these mutations are detected through sequencing, hybridization assays or mini-sequencing. However, these techniques can be expensive and tedious. Previously, CE-SSCP using the F-108 polymer was able to distinguish SNPs for the melanocortin-1 receptor (mc1r) coat color gene in horses (Equus caballus) that differed by one nucleotide substitution. The objective of t...
Chiral Selectivity in Inter-reactant Recognition and Electron Transfer of the Oxidation of Horse Heart Cytochrome c by Trioxalatocobaltate(III). Outer-sphere electron transfer (ET) between optically active transition-metal complexes and either other transition-metal complexes or metalloproteins is a prototype reaction for kinetic chirality. Chirality as the ratio between bimolecular rate constants of two enantiomers mostly amounts to 1.05-1.2 with either the Λ or Δ form the more reactive, but the origin of chirality in ET parameters such as work terms, electronic transmission coefficient, and nuclear reorganization free energy has not been addressed. We report a study of ET between the Λ-/Δ-[Co(Ox)3](3-) pair (Ox = oxalate) and hor...
A highly prevalent equine glycogen storage disease is explained by constitutive activation of a mutant glycogen synthase. Equine type 1 polysaccharide storage myopathy (PSSM1) is associated with a missense mutation (R309H) in the glycogen synthase (GYS1) gene, enhanced glycogen synthase (GS) activity and excessive glycogen and amylopectate inclusions in muscle. Equine muscle biochemical and recombinant enzyme kinetic assays in vitro and homology modelling in silico, were used to investigate the hypothesis that higher GS activity in affected horse muscle is caused by higher GS expression, dysregulation, or constitutive activation via a conformational change. PSSM1-affected horse muscle had significantly higher gly...
Amino acid mutations in the env gp90 protein that modify N-linked glycosylation of the Chinese EIAV vaccine strain enhance resistance to neutralizing antibodies. The Chinese EIAV vaccine is an attenuated live virus vaccine obtained by serial passage of a virulent horse isolate (EIAV) in donkeys (EIAV) and, subsequently, in donkey cells in vitro. In this study, we compare the env gene of the original horse virulent virus (EIAV) with attenuated strains serially passaged in donkey MDM (EIAV) and donkey dermal cells (EIAV). Genetic comparisons among parental and attenuated strains found that vaccine strains contained amino acid substitutions/deletions in gp90 that resulted in a loss of three potential N-linked glycosylation sites, designated g5, g9, and g1...
Reproductive stage-dependent effects of additional cryoprotectant agents for the cryopreservation of stallion germ cells. The main objective of this study was to evaluate the efficacy of an additional cryoprotectant in 10% dimethyl sulfoxide (DMSO) on cryopreserving germ cells from stallions at different reproductive stages. Testicular samples were obtained from pre-pubertal (1-1.5 yr, n=6) and post-pubertal (3-7 yr, n=5) stallions. Germ cells were isolated using a two-enzyme digestion procedure and cryopreserved in minimal essential medium alpha containing 10% fetal bovine serum and 10% DMSO with or without addition of trehalose (50, 100, or 200mM) or polyethylene glycol (PEG, 2.5, 5, or 10%). Viability, cell po...
Bone morphogenetic protein 2 stimulates chondrogenesis of equine synovial membrane-derived progenitor cells. Bone morphogenetic protein 2 (BMP-2) is critical for skeletal and cartilage development, homeostasis and repair. This study was conducted to clone and characterize equine BMP-2, develop expression constructs for equine BMP-2, and to determine whether BMP-2 can stimulate chondrogenesis of equine synovial membrane-derived progenitor cells (SMPC). Methods: Equine BMP-2 cDNA was amplified from chondrocyte RNA, and then transferred into an expression plasmid and adenoviral vector. Effective expression of equine BMP-2 was confirmed using a BMP reporter cell line. SMPC were isolated from synovium, ex...
Bone structural similarity score: a multiparametric tool to match properties of biomimetic bone substitutes with their target tissues. One of the hardest tasks in developing or selecting grafts for bone substitution surgery or tissue engineering is to match the structural and mechanical properties of tissue at the recipient site, because of the large variability of tissue properties with anatomical site, sex, age and health conditions of the patient undergoing implantation. We investigated the feasibility of defining a quantitative bone structural similarity score based on differences in the structural properties of synthetic grafts and bone tissue. Methods: Two biocompatible hydroxyapatite porous scaffolds with different nom...
Synthesis and evaluation of isometamidium-alginate nanoparticles on equine mononuclear and red blood cells. Isometamidium hydrochloride (ISMM) is an effective drug for the treatment of trypanosomosis, but it causes local and systemic toxicity. Isometamidium hydrochloride has limited therapeutic index and exhibit considerable variation in their prophylactic activities. We developed a trypanocidal nanoformulation using ISMM and polymers sodium alginate/gum acacia to enhance the efficacy of the drug at lower doses, while minimizing undesirable side effects. It was characterized by transmission electron microscopy and infrared spectroscopy for evaluation of size, morphology, functional groups, etc. In v...
