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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.
Dimethylformamide improves the in vitro characteristics of thawed stallion spermatozoa reducing sublethal damage. A total of 42 ejaculates were used in the experiment; six ejaculates per stallion, obtained from seven Pure Spanish stallions (PRE), were split and frozen in freezing media with different concentrations and combinations of cryoprotectant (CPA): (i) Cáceres (skim milk based extender) containing 2.5% glycerol (2.5GL), (ii) Cáceres containing 1.5% glycerol and 1.5% dimethylformamide (1.5%GL-1.5%DMFA), (iii) Cáceres extender supplemented with 1.5% glycerol and 2.5% dimethylformamide (1.5%GL-2.5%DMFA) and (iv) Cáceres extender supplemented with 4% dimethylformamide (4%DMFA). After at least 4 we...
Acute phase protein concentrations after limited distance and long distance endurance rides in horses. Acute phase proteins (APP) have been described as useful for assessing health in human and animal patients, as they closely reflect the acute phase reaction (APR). In humans and dogs a reaction analogous to APR has also been described after prolonged or strenuous exercise. The aim of this study was to determine, if similar reactions occur in endurance horses after limited and long distance rides. Seventeen horses that successfully completed various distance competitions were tested. Routine haematological and biochemical tests were performed and the concentrations of serum amyloid A (SAA), C-r...
Identification of major milk fat globule membrane proteins from pony mare milk highlights the molecular diversity of lactadherin across species. Although several studies have been devoted to the colloidal and soluble protein fractions of mare milk (caseins and whey proteins), to date little is known about the milk fat globule membrane (MFGM) protein fraction from mare milk. The objective of this study was thus to describe MFGM proteins from Equidae milk and to compare those proteins to already described MFGM proteins from cow and goat milk. Major MFGM proteins (namely, xanthine oxidase, butyrophilin, lactadherin, and adipophilin) already described in cow or goat milk were identified in mare milk using mass spectrometry. However, specie...
Assessment of reactive oxygen species production in cultured equine skeletal myoblasts in response to conditions of anoxia followed by reoxygenation with or without exposure to peroxidases. To culture equine myoblasts from muscle microbiopsy specimens, examine myoblast production of reactive oxygen species (ROS) in conditions of anoxia followed by reoxygenation, and assess the effects of horseradish peroxidase (HRP) and myeloperoxidase (MPO) on ROS production. Methods: 5 healthy horses (5 to 15 years old). Methods: Equine skeletal myoblast cultures were derived from 1 or 2 microbiopsy specimens obtained from a triceps brachii muscle of each horse. Cultured myoblasts were exposed to conditions of anoxia followed by reoxygenation or to conditions of normoxia (control cells). Cell p...
Change of Ferritin-binding Activity in the Serum of Foal after Birth. In mammal circulation, various ferritin-binding proteins (FBPs) are thought to be involved in the clearance of circulating ferritin after complex formation with it. However, horse FBPs are known to cause inhibitory effects on ferritin immunoassay due to the concealment of the ferritin molecule to anti-ferritin antibodies used in the ferritin immunoassay. These inhibitory effects are eliminated by heat treatment of horse serum at 75°C for 15 min. The inhibitory effects on ferritin immunoassay in the sera of ten foal sera (5 females and 5 males) from 1 to 18 months were detected during all peri...
Comparison of the principal proteins in bovine, caprine, buffalo, equine and camel milk. Proteomic analysis of bovine, caprine, buffalo, equine and camel milk highlighted significant interspecies differences. Camel milk was found to be devoid of β-lactoglobulin, whereas β-lactoglobulin was the major whey protein in bovine, buffalo, caprine, and equine milk. Five different isoforms of κ-casein were found in camel milk, analogous to the micro-heterogeneity observed for bovine κ-casein. Several spots observed in 2D-electrophoretograms of milk of all species could tentatively be identified as polypeptides arising from the enzymatic hydrolysis of caseins. The understanding gained f...
Optimizing human in vivo dosing and delivery of β-alanine supplements for muscle carnosine synthesis. Interest into the effects of carnosine on cellular metabolism is rapidly expanding. The first study to demonstrate in humans that chronic β-alanine (BA) supplementation (~3-6 g BA/day for ~4 weeks) can result in significantly augmented muscle carnosine concentrations (>50%) was only recently published. BA supplementation is potentially poised for application beyond the niche exercise and performance-enhancement field and into other more clinical populations. When examining all BA supplementation studies that directly measure muscle carnosine (n=8), there is a significant linear correlation...
The role of diffusion in ferritin-induced relaxation enhancement of protons. The influence of proton diffusion on nuclear magnetic resonance (NMR) relaxation was investigated in the presence of horse spleen ferritin at 7 T. Binary mixtures of water and glycerol were used to control diffusion within the range of 0.6-2.0 × 10(-9)m(2)/s, which was confirmed by pulsed gradient techniques. The effect of chemical exchange by hydrolysis between water and glycerol on relaxation was characterized with Carr-Purcell-Meiboom-Gill (CPMG) dispersion experiments. The relaxation rate enhancement of the protons due to ferritin was found to be inversely proportional to the diffusion co...
Effects of cell storage and passage on basal and oxytocin-regulated prostaglandin secretion by equine endometrial epithelial and stromal cells. Cell cultures are useful for determining the responses of specific cell types to various factors under controlled conditions and for obtaining a better understanding of in vivo physiologic processes. The aims of the present study were (i) to establish methodologies for isolation, culture and cryopreservation of equine endometrial epithelial and stromal cells; and (ii) to determine the effect of passage and cryopreservation on endometrial cell physiology, based on their basal and oxytocin (OT)-stimulated prostaglandin (PG) release. Epithelial and stromal cells were obtained by enzymatic digesti...
Monoclonal antibodies to equine CD23 identify the low-affinity receptor for IgE on subpopulations of IgM+ and IgG1+ B-cells in horses. CD23, also called FcεRII, is the low-affinity receptor for IgE and has first been described as a major receptor regulating IgE responses. In addition, CD23 also binds to CD21, integrins and MHC class II molecules and thus has a much wider functional role in immune regulation ranging from involvement in antigen-presentation to multiple cytokine-like functions of soluble CD23. The role of CD23 during immune responses of the horse is less well understood. Here, we expressed equine CD23 in mammalian cells using a novel IL-4 expression system. Expression resulted in high yield of recombinant IL-4/...
Cytokine production and proliferation upon in vitro oligodeoxyribonucleotide stimulation of equine peripheral blood mononuclear cells. Synthetic oligodeoxyribonucleotides (ODN) may prove useful immune modulators in equine medicine. It is however important to assess the effects of each specific ODN in the species it is intended to be used in. The present study therefore aimed to evaluate some ODN for induction of cytokine production; i.e. type I interferons (IFN), IFN-γ, tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β), and proliferation of equine peripheral blood mononuclear cells (PBMC). A panel of four ODN containing unmethylated cytosine-guanosine sequences (CpG) was used: ODN 1 and ODN 8 repre...
Mass spectral measurements of the apoHDL in horse (Equus caballus) cerebrospinal fluid. As a continuation of our proteogenomic studies of equine apolipoproteins, we have obtained molecular masses for several of the apolipoproteins associated with the HDL in horse cerebrospinal fluid (CSF). Using electrospray-ionization mass spectrometry (ESI-MS), we report on values for apolipoproteins, A-I and A-II, as well as acylated apoA-I. In comparison with our previously published data on equine plasma apolipoproteins, there appears to be a higher percentage of acylated apoA-I in the CSF than in plasma. As was the case in plasma, apoA-II circulates as a homodimer. These studies also reveal...
Toxicity of glycerol for the stallion spermatozoa: effects on membrane integrity and cytoskeleton, lipid peroxidation and mitochondrial membrane potential. Glycerol is, to date, the most widely used cryoprotectant to freeze stallion spermatozoa at concentrations between 2% and 5%. Cryoprotectant toxicity has been claimed to be the single most limiting factor for the success of cryopreservation. In order to evaluate the toxic effects of the concentrations of glycerol used in practice, stallion spermatozoa were incubated in Biggers Whitten and Whittingham (BWW) media supplemented with 0%, 0.5%, 1.5%, 2.5%, 3.5%, and 5% glycerol. In two additional experiments, a hyposmotic (75 mOsm/kg) and a hyperosmotic (900 mOsm/kg) control media were included. Sp...
Effect of growth factors on the migration of equine oral and limb fibroblasts using an in vitro scratch assay. The objective of this study was to determine the effect of platelet derived growth factor BB (PDGF), epidermal growth factor (EGF), transforming growth factor β1 (TGFβ1), insulin like growth factor-1 (IGF-1) and fibroblast growth factor-2 (FGF-2) on the proliferation and migration of equine oral mucosa and leg skin fibroblast cell lines, using an in vitro scratch assay. Fibroblasts from the two sites were firstly grown to confluence and then an area of cells removed (cell void area). Cell migration alone (with the addition of the mitosis inhibitor mitomycin-C to the culture media) and prolif...
Evaluation of direct in vivo gene transfer in an equine metacarpal IV ostectomy model using an adenoviral vector encoding the bone morphogenetic protein-2 and protein-7 gene. To evaluate gene transfer in an equine metacarpal IV (MCIV) ostectomy model using adenoviral vectors encoding the human bone morphogenetic protein-2 and protein-7 gene (Ad-BMP-2/-7). Methods: Healthy adult horses (n = 15). Methods: A plate stabilized, critical size 1.5 cm ostectomy was created in left and right MCIV. The ostectomy site was injected with either Ad-green fluorescent protein (Ad-GFP) or Ad-hBMP-2/-7 at completion of surgery; the same treatment was assigned to both the left and right forelimb of each horse (n = 5 horses/group). Bone healing was evaluated radiographically every 2 w...
Long term intensive exercise training leads to a higher plasma malate/lactate dehydrogenase (M/L) ratio and increased level of lipid mobilization in horses. Continuous high intensity training may induce alterations to enzyme activities related to glucose and lipid metabolism in horses. In our study, five Thoroughbred race horses (3 male and 2 female, avg age=5 yrs old) were compared against five riding horses (1 male, 1 female, 3 gelding, avg age=13 yrs old) in terms of energy metabolism, by examining plasma malate (MDH) and lactate (LDH) dehydrogenase activities and M/L ratio. MDH is involved in NADH and ATP generation, whereas LDH can convert NADH back into NAD(+) for ATP generation. An increase in plasma M/L ratio can reflect heightened energy ...
Repeated post-exercise administration with a mixture of leucine and glucose alters the plasma amino acid profile in Standardbred trotters. The branched chain amino acid leucine is a potent stimulator of insulin secretion. Used in combination with glucose it can increase the insulin response and the post exercise re-synthesis of glycogen in man. Decreased plasma amino acid concentrations have been reported after intravenous or per oral administration of leucine in man as well as after a single per oral dose in horses. In man, a negative correlation between the insulin response and the concentrations of isoleucine, valine and methionine have been shown but results from horses are lacking. This study aims to determine the effect of ...
Differential gene expression of CYP3A isoforms in equine liver and intestines. Recently, seven CYP3A isoforms - CYP3A89, CYP3A93, CYP3A94, CYP3A95, CYP3A96, CYP3A97 and CYP129 - have been isolated from the horse genome. In this study, we have examined the hepatic and intestinal gene expression of these CYP3A isoforms using TaqMan probes. We have also studied the enzyme activity using luciferin-isopropyl acetal (LIPA) as a substrate. The results show a differential gene expression of the CYP3A isoforms in the liver and intestines in horses. In the liver, CYP3A89, CYP3A94, CYP3A96 and CYP3A97 were highly expressed, while in the intestine there were only two dominating isof...
Effect of a solution of hyaluronic acid-chondroitin sulfate-N-acetyl glucosamine on the repair response of cartilage to single-impact load damage. To investigate effects of 1% hyaluronic acid-chondroitin sulfate-N-acetyl glucosamine (HCNAG) on the damage repair response in equine articular cartilage. Methods: Articular cartilage from 9 clinically normal adult horses. Methods: Full-thickness cartilage disks were harvested from the third metacarpal bone. Cartilage was single-impact loaded (SIL) with 0.175 J at 0.7 m/s and cultured in DMEM plus 1 % (vol/vol) HCNAG or fibroblastic growth factor (FGF)-2 (50 ng/mL). Histologic and immunohistochemical techniques were used to identify tissue architecture and apoptotic cells and to immunolocalize...
Autophagy and apoptosis have a role in the survival or death of stallion spermatozoa during conservation in refrigeration. Apoptosis has been recognized as a cause of sperm death during cryopreservation and a cause of infertility in humans, however there is no data on its role in sperm death during conservation in refrigeration; autophagy has not been described to date in mature sperm. We investigated the role of apoptosis and autophagy during cooled storage of stallion spermatozoa. Samples from seven stallions were split; half of the ejaculate was processed by single layer centrifugation, while the other half was extended unprocessed, and stored at 5°C for five days. During the time of storage, sperm motility (C...
Identification and characterization of equine herpesvirus type 1 pUL56 and its role in virus-induced downregulation of major histocompatibility complex class I. Major histocompatibility complex class I (MHC-I) molecules play an important role in host immunity to infection by presenting antigenic peptides to cytotoxic T lymphocytes (CTLs), which recognize and destroy virus-infected cells. Members of the Herpesviridae have developed multiple mechanisms to avoid CTL recognition by virtue of downregulation of MHC-I on the cell surface. We report here on an immunomodulatory protein involved in this process, pUL56, which is encoded by ORF1 of equine herpesvirus type 1 (EHV-1), an alphaherpesvirus. We show that EHV-1 pUL56 is a phosphorylated early protein w...
Isolation, characterization and differentiation of mesenchymal stem cells from amniotic fluid, umbilical cord blood and Wharton’s jelly in the horse. Mesenchymal stem cells (MSCs) have been derived from multiple sources of the horse including umbilical cord blood (UCB) and amnion. This work aimed to identify and characterize stem cells from equine amniotic fluid (AF), CB and Wharton's Jelly (WJ). Samples were obtained from 13 mares at labour. AF and CB cells were isolated by centrifugation, while WJ was prepared by incubating with an enzymatic solution for 2 h. All cell lines were cultured in DMEM/TCM199 plus fetal bovine serum. Fibroblast-like cells were observed in 7/10 (70%) AF, 6/8 (75%) CB and 8/12 (66.7%) WJ samples. Statistically ...
In vivo osteoinductivity of gelatin β-tri-calcium phosphate sponge and bone morphogenetic protein-2 on an equine third metacarpal bone defect. This study evaluated the therapeutic effects of a gelatin-β-TCP sponge (sponge) incorporating BMP-2 (BMP-2/sponge) on bone regeneration in equines. Six bone defects were created in third metacarpals of five thoroughbred horses, and a total of six treatments were applied in a randomized manner. The treatments were BMP-2/sponge, BMP-2/gelatin hydrogel sheet (sheet), free BMP-2, bFGF/sheet, plain sponge, and plain sheet. The defects were monitored for 16 weeks by radiography and then examined by histological analysis. Radiographic evaluation scores of bone regeneration revealed significantly gre...
Effect of Hoechst 33342 on stallion spermatozoa incubated in KMT or Tyrodes modified INRA96. The only known means of effectively separating populations of X and Y bearing sperms is the Beltsville sexing technology. The technology implies that each individual sperm is interrogated for DNA content, measuring the intensity of the fluorescence after staining the spermatozoa with Hoechst 33342. Because there are no data regarding the effect of the staining on stallion sperm, ejaculates were incubated up to 90 min in presence of 0, 4.5, 9, 22.5, 31.5, 45, 54, 67.5, 76.5 and 90 μM of Hoechst 33342, in two media, KMT or INRA-Tyrodes. After 40 and 90 min of incubation, motility (CASA) and mem...
Identification of ORF sequences and exercise-induced expression change in thoroughbred horse OXCT1 gene. In the mitochondrial matrix, the OXCT1 gene catalyzes the reversible transfer of coenzyme A from succinyl-CoA to acetoacetate in a reaction related to energy production from ketone bodies. Here, horse OXCT1 gene containing coenzyme A transferase domain was identified in the transcriptome analysis of cDNAs derived from skeletal muscles. Horse OXCT1 gene consisted of 1761 [corrected] nucleotide sequences with an open reading frame of 1560 nucleotides encoding a protein of 520 putative amino acid residues.The number of non-synonymous substitutions was lower than the number of synonymous substitut...
The influence of road transport on the activities of glutathione reductase, glutathione peroxidase, and glutathione-S-transferase in equine erythrocytes. Transport of horses may have significant impact on serum biochemical and hematologic analytes and resistance to infection. Objective: The aim of our study was to assess the influence of transport stress on selected enzymatic antioxidants in equine blood. Methods: The study was conducted on a group of 60 horses of different breeds and ranging in age from 4 to 10 years. Venous blood was collected immediately before loading horses onto trailers for 8 hours of transport (I), immediately after unloading them from the trailer (II), and after subsequent stall rest for 24 hours (III). Hemolysates of b...
Effect of corticotherapy on proteomics of endometrial fluid from mares susceptible to persistent postbreeding endometritis. The objective was to determine the effects of corticotherapy, in the presence and absence of uterine inflammation, on proteomics of endometrial fluid from mares susceptible to endometritis. In 11 mares, estrus was induced seven times with 5 mg PGF(2α) given at 14-day intervals. The first estrus was a control (no treatment). During the third estrus, mares received glucocorticoid (GC) treatment (20 mg isoflupredone acetate) every 12 h, for three consecutive days. The fifth estrus was the Infected treatment (intrauterine infusion of 1 × 10(9) colony-forming unit/mL Streptococcus equi subspecies...
The biology of equine mesenchymal stem cells: phenotypic characterization, cell surface markers and multilineage differentiation. Mesenchymal stem cells (MSCs) are multipotent stem cells that can give rise to a range of connective tissue cells including osteoblasts, chondrocytes and adipocytes. MSCs have been isolated from humans and a variety of animal species including rodents, dogs, horses and rabbits. There is currently no consensus on how these cells are identified and characterized. This is partly due to the lack of standardized specific cell surface markers for MSCs. The aim of this review is to examine the literature on equine MSCs and establish whether there is a well-defined phenotype for these cells. Equine MS...
Daily endogenous cortisol production and hydrocortisone pharmacokinetics in adult horses and neonatal foals. To compare daily endogenous cortisol production rate and the pharmacokinetics of an i.v. bolus of hydrocortisone between neonatal foals and adult horses. Methods: 10 healthy full-term 2- to 4-day-old foals and 7 healthy adult horses. Methods: Blood samples were collected from each horse every 15 to 20 minutes for 24 hours for determination of 24-hour mean cortisol concentration. Afterward, dexamethasone (0.08 mg/kg) was administered i.v. to suppress endogenous cortisol production. Twelve hours afterward, hydrocortisone sodium succinate (1.0 mg/kg) was administered as a rapid i.v. bolus and ser...
