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Topic:Biochemistry
The study of biochemistry in horses encompasses the chemical processes and substances that occur within equine organisms. This field investigates the molecular interactions and pathways that are fundamental to horse physiology, including metabolism, enzyme activity, and genetic expression. Key areas of interest include the examination of metabolic disorders, nutrient absorption, and the biochemical basis of muscle function and energy production. Researchers utilize biochemical analysis to understand health and disease mechanisms in horses, contributing to the development of diagnostic tools and therapeutic strategies. This page gathers peer-reviewed studies and scholarly articles that explore various biochemical processes and their implications for equine health and performance.
Effect of urea and alkylureas on the stability and structural fluctuation of the M80-containing Ω-loop of horse cytochrome c. The effect of denaturants on the structural fluctuation of M80-containing Ω-loop of ferrocytochrome c was determined by measuring the rate coefficient of CO-association with ferrocytochrome c under varying concentrations of urea and alkylureas (methylurea (MU), N,N'-dimethylurea (DMU), ethylurea (EU), tetramethylurea (TMU)) at pH7.0, 25°C. As denaturant concentration is increased within the subdenaturing limit, the CO-association reaction is decelerated indicating that subdenaturing concentrations of denaturant reduce the structural fluctuation of the Ω-loop. Structural fluctuation of the ...
Determination of pergolide in horse plasma by UPLC-MS/MS for pharmacokinetic applications. Pergolide, an ergot-derived dopamine D2 receptor agonist, is used extensively as an orally administered treatment for pituitary pars intermedia dysfunction (PPID) in horses. One of the barriers associated with pergolide determinations in plasma for pharmacokinetic applications has been the technically demanding requirement for sensitivity. The objective of our work was to develop a simple assay for the determination of pergolide in plasma and demonstrate its potential application in the study of pergolide pharmacokinetics (PK) in horses. A UPLC-MS/MS assay was developed with a simple sample pr...
Apparent digestibility of broken rice in horses using in vivo and in vitro methods. The aim of this study was to assess the apparent digestibility of broken rice using total collection of feces and the pepsin-cellulase in vitro technique to provide updated and more accurate digestion coefficients for this by-product when fed to horses. The in vivo digestibility trial was consecutively performed, using five adult geldings, weighing 555.6 kg on average. First, hay was given as the only feedstuff, while second, the experimental diet consisted of the same hay plus broken rice at a forage-to-concentrate ratio of 70/30 (on dry matter (DM) basis). Feces were collected over 6 days pr...
The Genome of the Predominant Equine Lactobacillus Species, Lactobacillus equi, Is Reflective of Its Lifestyle Adaptations to an Herbivorous Host. We report the draft genome sequence of Lactobacillus equi strain DPC6820, isolated from equine feces. L. equi is a predominant Lactobacillus species in the horse hindgut microbiota. An examination of the genome identified genes and enzymes highlighting L. equi adaptations to the herbivorous gastrointestinal tract of the horse, including fructan hydrolases. This genome sequence may help us further understand the microbial ecology of the equine hindgut and the influence lactobacilli have on it.
Long-range electron transfer with myoglobin immobilized at Au/mixed-SAM junctions: mechanistic impact of the strong protein confinement. Horse muscle myoglobin (Mb) was tightly immobilized at Au-deposited ~15-Å-thick mixed-type (1:1) alkanethiol SAMs, HS-(CH₂)₁₁-COOH/HS-(CH₂)₁₁-OH, and placed in contact with buffered H₂O or D₂O solutions. Fast-scan cyclic voltammetry (CV) and a Marcus-equation-based analysis were applied to determine unimolecular standard rate constants and reorganization free energies for electron transfer (ET), under variable-temperature (15-55 °C) and -pressure (0.01-150 MPa) conditions. The CV signal was surprisingly stable and reproducible even after multiple temperature and pressure cycl...
Membrane culture and reduced oxygen tension enhances cartilage matrix formation from equine cord blood mesenchymal stromal cells in vitro. Ongoing research is aimed at increasing cartilage tissue yield and quality from multipotent mesenchymal stromal cells (MSC) for the purpose of treating cartilage damage in horses. Low oxygen culture has been shown to enhance chondrogenesis, and novel membrane culture has been proposed to increase tissue yield and homogeneity. The objective of this study was to evaluate and compare the effect of reduced oxygen and membrane culture during in vitro chondrogenesis of equine cord blood (CB) MSC. Methods: CB-MSC (n = 5 foals) were expanded at 21% oxygen prior to 3-week differentiation in membrane or...
Immunophenotypic characterization and tenogenic differentiation of mesenchymal stromal cells isolated from equine umbilical cord blood. Mesenchymal stem cells (MSCs) isolated from umbilical cord blood (UCB) in equines have not been well characterized with respect to the expression of pluripotency and mesenchymal markers and for tenogenic differentiation potential in vitro. The plastic adherent fibroblast-like cells isolated from 13 out of 20 UCB samples could proliferate till passage 20. The cells expressed pluripotency markers (OCT4, NANOG, and SOX2) and MSC surface markers (CD90, CD73, and CD105) by RT-PCR, but did not express CD34, CD45, and CD14. On immunocytochemistry, the isolated cells showed expression of CD90 and CD73...
During cooled storage the extender influences processed autophagy marker light chain 3 (LC3B) of stallion spermatozoa. To investigate the role of the processed autophagy marker light chain 3 (LC3B) protein in sperm survival in stallion semen processing during cooled storage, split ejaculates were diluted in two different extenders, KMT and INRA 96, and LC3B processing and sperm quality evaluated during incubation at 5°C for five days. After 3 days of incubation there was a drop in total motility in both extenders, although the percentage of progressive motile sperm was greater (P<0.05) in samples extended in INRA96. On Day 5 of cooled storage all sperm parameters decreased significantly independent of the ...
Elevations in serum muscle enzyme activities in racehorses due to unaccustomed exercise and training. Hereditary muscular disease is well described in racehorses, yet little is known about traumatic muscle disease associated with unaccustomed exercise or training. The objective of the study was to compare sedentary horses, racehorses undergoing training for the first time (unaccustomed exercise), and experienced racehorses during a training season (accustomed exercise) to investigate the effect of exercise and training on serum muscle enzyme activities and other variables. Horses were sampled prior to exercise for serum activities of aspartate amino transferase (AST), creatine kinase and other...
Effects of quinapril on angiotensin converting enzyme and plasma renin activity as well as pharmacokinetic parameters of quinapril and its active metabolite, quinaprilat, after intravenous and oral administration to mature horses. Angiotensin converting enzyme (ACE) inhibitors improve survival and quality of life in human patients and small animals with cardiovascular and renal disease. There is limited information regarding their effects in horses. Objective: The purpose of this study was to determine the pharmacokinetics of quinapril and its effects on ACE and renin in horses. Methods: Experimental study using healthy mature horses. Methods: Six healthy horses were administered quinapril at 120 mg i.v., 120 mg per os and 240 mg per os in a 3-way crossover design. Blood was collected for measurement of quinapril ...
Detection and pharmacokinetics of three formulations of firocoxib following multiple administrations to horses. The use of firocoxib in horses and its ability to affect performance and potential to allow a horse to compete when it otherwise should not, necessitates establishing appropriate withdrawal time guidelines prior to performance. Objective: To describe plasma concentrations and characterise the pharmacokinetics of 3 firocoxib formulations following multiple administrations of the label dose, with respect to recommended plasma thresholds for performance horses. Methods: Balanced 3-way crossover prospective study. Methods: Nine healthy mature horses were administered firocoxib injectable solution ...
Proteomic analysis of tendon extracellular matrix reveals disease stage-specific fragmentation and differential cleavage of COMP (cartilage oligomeric matrix protein). During inflammatory processes the extracellular matrix (ECM) is extensively remodeled, and many of the constituent components are released as proteolytically cleaved fragments. These degradative processes are better documented for inflammatory joint diseases than tendinopathy even though the pathogenesis has many similarities. The aims of this study were to investigate the proteomic composition of injured tendons during early and late disease stages to identify disease-specific cleavage patterns of the ECM protein cartilage oligomeric matrix protein (COMP). In addition to characterizing fragme...
Thiol-disulfide proteins of stallion epididymal spermatozoa. Thiol groups of cysteine residues represent redox centers involved in multiple biological functions. It has been postulated that changes in the redox status of mammalian epididymal spermatozoa contribute to the sperm maturation process. The present work shows the thiol-disulfide protein profile of stallion epididymal spermatozoa achieved by two-dimension electrophoresis and MALDI-TOF/TOF mass spectrometry of proteins labeled with a thiol-reactive fluorescent tag, monobromobimane. Our results have shown the formation of disulfide bonds in several sperm protein fractions during the epididymal ma...
Characterization of arginase expression by equine neutrophils. Neutrophils are the predominant cells recruited in the airways of horses suffering from heaves. These cells have been shown to express arginase in some species. The metabolism of l-arginine is thought to be involved in chronic inflammation, and airway obstruction and remodeling. The aim of this study was to assess the expression, regulation, activity, and functional role of arginase isoforms in equine neutrophils. Arginase I, arginase II, ornithine decarboxylase (ODC) and ornithine aminotransferase (OAT) expression were assessed in resting and stimulated (IL-4, LPS/fMLP, PMA; 5 and 18 h) blood...
Measurement of Carbonic Anhydrase I and II Isozymes in Feces as a Marker of Occult Blood in Horses with Intestinal Tract Bleeding. Although endoscopy is the definitive diagnostic method for the detection of colonic ulcers, the equipment required for performing the test is costly and difficult to use. Therefore, a simple cost-effective and reliable screening test for intestinal tract bleeding is needed. To this end, we measured carbonic anhydrase isozymes (CA-I and CA-II) originating from erythrocytes by ELISA in order to determine if they could be used as markers of occult blood in feces. For fecal extract preparation, 2 g of feces were mixed with 4 ml of 0.01 M Tris-HCl (pH 8.0) containing 0.01% thimerosal. The concentra...
Characterization of bovine ruminal and equine cecal microbial populations enriched for enhanced nitro-toxin metabolizing activity. The nitrotoxins 3-nitro-1-propionic acid (NPA) and 3-nitro-1-propanol (NPOH) are produced by a wide variety of leguminous plants, including over 150 different species and varieties of Astragalus potentially grazed by livestock. These toxins are known to be detoxified by at least one ruminal bacterium but detoxification by bacteria from other gut habitats is not known. In the present study, mixed populations of bovine ruminal and equine cecal microbes were enriched for NPA-metabolizing bacteria via consecutive 24-72 h culture in a basal minimal rumen fluid-based medium supplemented with 4.2 mM ...
Hematological and biochemical reference values for the endangered kiso horse. To establish blood and biochemical references for the endangered Kiso horse, blood samples were collected from 111 adult Kiso horses, 74.5% of the existing breed. The samples were analyzed for 23 hematological and biochemical parameters to determine their means and standard deviations (SD). We compared the mean ± 2SD with the reference values cited in one of the most commonly used veterinary textbooks in Japan. The hematology of Kiso horses is characterized by lower erythrocyte count and hematocrit and hemoglobin levels. In addition, their serum biochemistry showed lower levels of aspartate t...
Case-Study Investigation of Equine Maternity via PCR-RFLP: A Biochemistry Laboratory Experiment. A simple and robust biochemistry laboratory experiment is described that uses restriction fragment length polymorphism (RFLP) of polymerase chain reaction (PCR) products to verify the identity of a potentially valuable horse. During the first laboratory period, students purify DNA from equine samples and amplify two loci of mitochondrial DNA. During the second laboratory period, students digest PCR products with restriction enzymes and analyze the fragment sizes through agarose gel electrophoresis. An optional step of validating DNA extracts through realtime PCR can expand the experiment to th...
Capacitation in the presence of methyl-β-cyclodextrin results in enhanced zona pellucida-binding ability of stallion spermatozoa. While IVF has been widely successful in many domesticated species, the development of a robust IVF system for the horse remains an elusive and highly valued goal. A major impediment to the development of equine IVF is the fact that optimised conditions for the capacitation of equine spermatozoa are yet to be developed. Conversely, it is known that stallion spermatozoa are particularly susceptible to damage arising as a consequence of capacitation-like changes induced prematurely in response to semen handling and transport conditions. To address these limitations, this study sought to develop a...
IgE and IgG epitope mapping by microarray peptide-immunoassay reveals the importance and diversity of the immune response to the IgG3 equine immunoglobulin. The presence of whole horse IgG in therapeutic snake antivenom preparations of high purity is a contamination that can cause IgE-mediated allergic reactions in patients. In this study, the immunodominant IgE and IgG-binding epitopes in horse heavy chain IgG3 were mapped using arrays of overlapping peptides synthesized directly onto activated cellulose membranes. Pooled human sera from patients with and without horse antivenom allergies were used to probe the membrane. We have demonstrated that, for both cases, individuals produce antibodies to epitopes of sequential amino acids of horse heavy ...
Effect of period, water temperature and agitation on loss of water-soluble carbohydrates and protein from grass hay: implications for equine feeding management. The effects of different water-soaking treatments on removal of water-soluble carbohydrates (WSC), WSC constituents and protein from four UK hays were determined. Hays were soaked in water for up to 16 hours at mean temperatures of 8°C, 16°C, in hot tap water (initially 49°C) or agitated and rinsed in clean water at 16°C. Initial hay WSC contents ranged from 154 to 216 g/kg dry matter. Losses of WSC from hays after 16 hours soaking at 8°C, 16°C, 16°C plus agitation and 49°C averaged 28, 46, 49 and 44 per cent, respectively. Corresponding percentage losses of fructan were 16, 37, 39 and...
Detection of endogenous cortisol in equine tears and blood at rest and after simulated stress. To determine whether cortisol is present in equine tears at rest and during simulated stress and compare tear cortisol to serum free and total cortisol. Methods: Fourteen healthy adult horses were included. Paired tear total cortisol and serum total and free cortisol concentrations were measured with ELISA, chemiluminescent immunoassay, and ultrafiltration methodology, respectively, in 10 horses at rest once daily for five consecutive days. In an additional four horses, paired tear and serum samples were collected for cortisol measurement before and after adrenocorticotropic hormone (ACTH) sti...
Effect of Cholesterol and Equex-STM Addition to an Egg Yolk Extender on Pure Spanish Stallion Cryopreserved Sperm. Cholesterol and Equex-STM are frequently added to different commercial and experimental extenders improving postthawing sperm quality. Doses of 125-150 mM of cholesterol from pig liver and 0.5-0.7% of Equex-STM were evaluated in a standard eggyolk extender (Martin et al., 1979). Six ejaculates per stallion from six pure Spanish stallions (6-8 years old) were collected in Martin's extender (B) and different mixtures of 125 mM-0.5% (I), 125 mM-0.7% (II), 150 mM-0.5% (III), and 150 mM-0.7% (IV) were added to original Martin's extender. Samples were frozen in 0.5 mL straws (100 × 10(6...
Distribution of MLH1 foci in horse male synaptonemal complex. Advances in molecular cytogenetics have provided the opportunity to study events during prophase I of meiosis. Immunofluorescent localization of different meiotic protein components were used to characterize the early stages of the first meiotic division in horse spermatocytes. The frequency and distribution of recombination events during prophase I were investigated using the mutL homolog 1 (MLH1) protein that is known to be associated with these events. The frequency and distribution of MLH1 foci were investigated in pachytene nuclei of 6 fertile stallions, and the average relative synaptone...
Comparison of paired serum and lithium heparin plasma samples for the measurement of serum amyloid A in horses using an automated turbidimetric immunoassay. This study aimed to evaluate whether equine serum amyloid A (SAA) concentrations could be reliably measured in plasma with a turbidimetric immunoassay previously validated for equine SAA concentrations in serum. Paired serum and lithium-heparin samples obtained from 40 horses were evaluated. No difference was found in SAA concentrations between serum and plasma using a paired t test (P=0.48). The correlation between paired samples was 0.97 (Spearman's rank P<0.0001; 95% confidence interval 0.95-0.99). Passing-Bablok regression analyses revealed no differences between paired samples. Bland-A...
Computed tomography-guided tissue engineering of upper airway cartilage. Normal laryngeal function has a large impact on quality of life, and dysfunction can be life threatening. In general, airway obstructions arise from a reduction in neuromuscular function or a decrease in mechanical stiffness of the structures of the upper airway. These reductions decrease the ability of the airway to resist inspiratory or expiratory pressures, causing laryngeal collapse. We propose to restore airway patency through methods that replace damaged tissue and improve the stiffness of airway structures. A number of recent studies have utilized image-guided approaches to create cell-...
Detection and quantification of dermorphin and selected analogs in equine urine. Dermorphin, a hepta-peptide with potent analgesic properties, is classified as a doping agent in equine racing. Since its discovery, a number of biologically active structural analogs have been synthesized and made commercially available so there is a need for reliable methods of detection. Results: A sensitive detection method was developed for dermorphin and six analogs in equine urine. Peptide enrichment was achieved using weak cation exchange with subsequent separation and detection by nano-UHPLC-MS/MS. Method validation parameters included: specificity, linearity (5-10000 pg/ml), recovery...
ASVCP guidelines: allowable total error guidelines for biochemistry. As all laboratory equipment ages and contains components that may degrade with time, initial and periodically scheduled performance assessment is required to verify accurate and precise results over the life of the instrument. As veterinary patients may present to general practitioners and then to referral hospitals (both of which may each perform in-clinic laboratory analyses using different instruments), and given that general practitioners may send samples to reference laboratories, there is a need for comparability of results across instruments and methods. Allowable total error (TEa ) is ...
Comparative Labeling of Equine and Ovine Multipotent Stromal Cells With Superparamagnetic Iron Oxide Particles for Magnetic Resonance Imaging In Vitro. The purpose of this study was to evaluate the use of three different superparamagnetic iron oxide (SPIO) particles for labeling of ovine and equine bone marrow (BM)-derived multipotent stromal cells (MSCs) in vitro. MSCs were obtained from five adult sheep and horses, respectively. After three passages (p3), cells were labeled with either 1) Molday ION Rhodamine B, 2) Endorem, 3) Resovist, or 4) remained unlabeled as control. Labeling efficiency, marker retention, and long-term detectability in MRI until p7 were evaluated. Further, proliferation capacity and trilineage differentiation as indic...
Molecular characterization and evolutionary analysis of horse BAFF-R, a tumor necrosis factor receptor related to B-cell survival. B-cell survival depends on signals induced by B-cell activating factor (BAFF) that binds to the BAFF receptor (BAFF-R). Herein, a BAFF-R homolog was identified in a horse (Equus caballus). The horse BAFF-R gene, located on chromosome 28, spans 1444 base pairs and encodes a 183-amino acid protein. The protein is structurally conserved, in which the DxL motif plays an important role in binding to BAFF. Furthermore, the horse BAFF-R extracellular domain was expressed and purified, which specifically bound to His6-sBAFF and had the capability of blocking the function of His6-sBAFF in vitro. Finall...
