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Topic:Enzymes
Enzymes are biological catalysts that facilitate biochemical reactions in horses by lowering the activation energy required for these processes. They are involved in various physiological functions, including digestion, metabolism, and cellular repair. Common enzymes in equine biology include amylase, lipase, and lactate dehydrogenase, each playing a specific role in the breakdown of nutrients and energy production. The activity and concentration of these enzymes can vary in response to different physiological and pathological conditions, serving as potential indicators in veterinary diagnostics. This page compiles peer-reviewed research studies and scholarly articles that explore the function, regulation, and clinical implications of enzymes in equine health.
Cyclooxygenase-2 mRNA expression in equine nonglandular and glandular gastric mucosal biopsy specimens obtained before and after induction of gastric ulceration via intermittent feed deprivation. To measure the expression of cyclooxygenase-2 (COX-2) mRNA in gastric biopsy specimens serially obtained from horses before, during, and after an 8-day intermittent feed-deprivation trial and to investigate the mucosal location of COX-2. Methods: 9 mixed-breed horses for retrieval of gastric biopsy specimens and 16 additional horses for immunohistochemical analysis. Methods: Gastric biopsy specimens were obtained from 6 horses; 3 of these horses and 3 more participated in an intermittent feed-deprivation trial 9 weeks later. A quantitative PCR assay was used to determine the amount of COX-2 mR...
Immunotherapy for pythiosis: Effect on NTPDase activity in lymphocytes of an experimental model. NTPDase (EC 3.6.1.5) occurs in lymphocytes and plays an important role in immune function, in that hydrolyzes extracellular nucleoside tri- and/or diphosphates to form AMP. Pythium insidiosum causes the disease pythiosis, a pyogranulomatous disease of horses, dogs, cattle, cats and humans. Most antifungal drugs are ineffective against this pathogen, and immunotherapy, a treatment approach that relies on the injection of P. insidiosum antigen, has been successfully used in humans and horses to manage this disease. In this study, we investigated NTPDase activity in lymphocytes from rabbits inocu...
Cytochromes: Reactivity of the “dark side” of the heme. Ligand binding to the heme distal side is a paradigm of heme-protein biochemistry, the proximal axial ligand being in most cases a His residue. NO binds to the ferrous heme-Fe-atom giving rise to hexa-coordinated adducts (as in myoglobin and hemoglobin) with His and NO as proximal and distal axial ligands, respectively, or to penta-coordinated adducts (as in soluble guanylate cyclase) with NO as the axial distal ligand. Recently, the ferrous derivative of Alcaligenes xylosoxidans cytochrome c' (Axcyt c') and of cardiolipin-bound horse heart cytochrome c (CL-hhcyt c) have been reported to bind ...
Relationships between uterine blood flow, peripheral sex steroids, expression of endometrial estrogen receptors and nitric oxide synthases during the estrous cycle in mares. The objective of this study was to investigate the relationships between uterine perfusion and estrogen, progesterone and the uterine nitric oxide synthase (NOS) system in five trotter mares during the estrous cycle. Color Doppler sonography for measurement of uterine blood flow and collection of blood for determination of plasma estrogen and progesterone concentrations were performed on days 0 (= ovulation), 1, 5, 11 and 15 and daily during estrus (days -1 to -4) of one estrous cycle; endometrial biopsy collection for mRNA expression analysis of NOS and estrogen receptors was performed on day...
Cyclooxygenase-2 expression in animal cancers. Cyclooxygenase (COX; also known as prostaglandin endoperoxide synthase) is a key enzyme in the biochemical pathway leading to the synthesis of prostaglandins. A large amount of epidemiological and experimental evidence supports a role for COX-2, the inducible form of the enzyme, in human tumorigenesis, notably in colorectal cancer. COX-2 mediates this role through the production of PGE(2) that acts to inhibit apoptosis, promote cell proliferation, stimulate angiogenesis, and decrease immunity. Similarly, COX-2 is believed to be involved in the oncogenesis of some cancers in domestic animals. H...
Cytokine and catabolic enzyme expression in synovium, synovial fluid and articular cartilage of naturally osteoarthritic equine carpi. Understanding the expression of catabolic and anabolic genes during osteoarthritis progression should help to identify the major mediators of the disease. Objective: To compare the cytokine and anabolic marker concentrations in synovium, synovial fluid and cartilage between normal and osteoarthritic joints. Methods: Carpi from horses age 2-11 years were used. Tissues were harvested at the time of surgery or euthanasia, and RNA was isolated for RT-PCR analysis. Tumour necrosis factor alpha (TNFα), interleukin-1beta (IL-1β), aggrecanase 1 (ADAMTS-4), aggrecanase 2 (ADAMTS-5), matrix metallopro...
The effect of isotonic multiple electrolyte infusions during anesthesia on blood gas and enzymatic values in horses. General equine anesthesia is accompanied by the risk of complications and even death. The majority of general anesthetics used in veterinary practice lead to arterial hypotension, decreased capillary perfusion and ventilatory depression leading to pathological changes referred to as postanesthetic myopathy. In this study an attempt was made to analyze changes in the perioperative and postoperative acid-base equilibrium and blood enzymatic activity in horses, subject to the intensity of the applied fluid therapy. The horses were divided into two groups: I--administered Ringer's solution during ...
Morphological and biochemical changes in the blood of horses naturally infected with Gasterophilus sp. larvae. Gasterophilus sp. constitute a group of specific parasites, which larval forms are found in horses and other phylogenetically related species--donkeys, mules and zebras. Their invasion is associated with marked fluctuations of hematological and biochemical blood parameters, including the activity of many enzymes and the electrolyte concentration. The purpose of the study was to analyze changes in selected biochemical and morphological blood parameters of horses infected with botfly larvae. The experimental group was formed of horses infested by Gasterophilus sp. larvae whereas the control grou...
Electrochemical study of gelatin as a matrix for the immobilization of horse heart cytochrome c. The aim of this paper is to emphasize the strength of gelatin as a stable matrix for redox enzymes. Cyclic voltammetry has been applied for a detailed electrochemical study of horse heart cytochrome c (HHC) entrapped in a gelatin matrix immobilized on a gold electrode. The influence of the HHC concentration, the mass percentage of the gelatin and the nature of the gelatin on the electrochemical behaviour of HHC have been described in detail. In addition, attenuated total reflection infrared (ATR-IR) spectroscopy was used to prove the immobilization on a qualitative and conformational level. Th...
Definite coordination arrangement of organometallic palladium complexes accumulated on the designed interior surface of apo-ferritin. Apo-ferritin (apo-Fr) mutants are used as scaffolds to accommodate palladium (allyl) complexes. Various coordination arrangements of the Pd complexes are achieved by adjusting the positions of cysteine and histidine residues on the interior surface of the apo-Fr cage.
Expression of cytosolic phospholipase A2 in equine endometrium during the oestrous cycle and early pregnancy. Phospholipase A2 (PLA2) is a key enzyme for biosynthesis of PGF2α. Real-time RT-PCR and immunohistochemistry were used to determine transcription and cellular distribution of cytosolic PLA2 (cPLA2) in the equine endometrium during the oestrous cycle and early pregnancy. Endometrial biopsy and blood samples were collected from cycling mares on days (d) 8, 11, 15 and 18 (oestrus) (Day 0 = Day of ovulation; n = 5 for each day) and from pregnant mares (n = 4) on d15. Except for mares on d18 and some cyclic mares (n = 2) on d15 with low progesterone (P₄) concentrations (< 3.18 nm), P₄ conce...
Analysis of Ca2+/Mg2+ selectivity in alpha-lactalbumin and Ca(2+)-binding lysozyme reveals a distinct Mg(2+)-specific site in lysozyme. The triggering of Ca(2+) signaling pathways relies on Ca(2+)/Mg(2+) specificity of proteins mediating these pathways. Two homologous milk Ca(2+)-binding proteins, bovine alpha-lactalbumin (bLA) and equine lysozyme (EQL), were analyzed using the simplest "four-state" scheme of metal- and temperature-induced structural changes in a protein. The association of Ca(2+)/Mg(2+) by native proteins is entropy-driven. Both proteins exhibit strong temperature dependences of apparent affinities to Ca(2+) and Mg(2+), due to low thermal stabilities of their apo-forms and relatively high unfavorable enthalpi...
Serum thyroid hormone, insulin, glucose, triglycerides and protein concentrations in normal horses: association with topical dexamethasone usage. The aim of this study was to determine if topical application of dexamethasone affected the serum concentrations of thyroid hormones (triiodothyronine T(3) and thyroxine T(4)), glucose, triglycerides, total protein and insulin in normal horses. Ten horses were treated twice daily for 10 days with 50 g dexamethasone using an ointment formulation. Thyroid hormones and insulin were assayed using standard radioimmunoassay methods, while glucose, triglycerides and total protein were determined using a standard enzymatic method and the Biuret reaction, respectively. An increase in serum glucose and ...
Electrophoretically mediated microanalysis for the evaluation of interspecies variation in cholinesterase metabolism. This study describes an electrophoretically mediated microanalysis method, suitable for the preclinical evaluation of the hydrolysis of ester drugs by the serum of different animals and for further characterization of human-animal correlation. Dog, cat, cow, horse, sheep, rat and human serum were diluted (25%) in the appropriate buffer and replaced the enzyme solution usually used in electrophoretically mediated microanalysis methods for the study of enzyme kinetics. They were then compared in terms of the ability to hydrolyze acetylthiocholine and butyrylthiocholine (0.25 mM) by in-capillary ...
Comparative studies on bull and stallion seminal DNase activity and interaction with semen extender and spermatozoa. We performed a series of comparative studies of bull and stallion seminal plasma (SP) and its role on sperm-neutrophil binding as well as the interaction between semen extender and seminal DNase. Because of contrasting roles of SP on sperm-neutrophil binding between horses and cattle, it was suspected there were some species-specific differences on sperm interaction with SP proteins due to the variations in the natural location of semen deposition (uterus compared to vagina). Bull frozen-thawed sperm removed from egg yolk extender showed similar results to fresh sperm, but this also caused ext...
A secondary isotope effect study of equine serum butyrylcholinesterase-catalyzed hydrolysis of acetylthiocholine. beta-Secondary deuterium isotope effects have been measured for equine serum butyrylcholinesterase-catalyzed hydrolysis of acetyl-L(3)-thiocholine (L=H or (2)H). The dependencies of initial rates on isotopic substrate concentrations show close adherence to Michaelis-Menten kinetics, and yield the following isotope effects: (D3)k(cat)/K(m)=0.98+/-0.02 and (D3)k(cat)=1.10+/-0.02. The modestly inverse isotope effect on k(cat)/K(m) is consistent with partial rate limitation by a step that converts the sp(2)-hybridized ester carbonyl of the E+A reactant state into a quasi-tetrahedral transition sta...
Expression of cyclooxygenase isoforms in ulcerated tissues of the nonglandular portion of the stomach in horses. To characterize the expression of the cyclooxygenase (COX)-1 and COX-2 isoforms in naturally occurring ulcers of the nonglandular portion of the stomach in horses. SPECIMEN POPULATION: 38 specimens from ulcerated stomachs and 10 specimens from healthy stomachs. Methods: Specimens were collected at an abbatoir; for each specimen of squamous gastric mucosa, 1 portion was fixed in neutral-buffered 10% formalin for immunohistochemical analysis and another was frozen at -70 degrees C for immunoblotting analysis. Immunoreactivity to 2 antibodies, MF241 (selective for COX-1) and MF243 (selective for ...
Evaluation of the effect of phosphodiesterase on equine platelet activation and the effect of antigen challenge on platelet phosphodiesterase activity in horses with recurrent airway obstruction. To determine whether expression of equine platelet activation-dependent surface markers is influenced by phospodiesterase (PDE) isoenzyme activity and whether antigen challenge alters platelet PDE activity in horses with recurrent airway obstruction (RAO). Methods: 16 horses. Methods: 7 healthy horses were used for in vitro experiments, 6 horses with RAO were used for antigen challenge, and 6 healthy horses were used as control animals. Three of the healthy horses had also been used in the in vitro experiments. Effects of PDE inhibition and activation of adenylyl cyclase on CD41/61 and CD62P e...
Seasonal variation of plasminogen activator activity in spermatozoa and seminal plasma of boar, buck, bull and stallion. Plasminogen activators (PA) are proteolytic enzymes present in the spermatozoa and seminal plasma of various species. They play a role in the binding of the spermatozoon and its penetration through the layers surrounding the oocyte. Plasminogen activator activity (PAA) is modulated by hormones that have a seasonal variation, such as testosterone and melatonin. The present study investigates the seasonal variation of PA activity in sperm extracts and seminal plasma of four farm animal species: boar, buck, bull and stallion. Semen samples were collected every second week during a 12-month period...
Cellular localization of androgen synthesis in equine granulosa-theca cell tumors: immunohistochemical expression of 17alpha-hydroxylase/17,20-lyase cytochrome P450. Elevated blood testosterone concentrations, often accompanied by male-typical behaviors, is a common signalment of mares with granulosa-theca cell tumors (GCTCs), but no definitive information exists regarding the cellular differentiation of tumors associated with androgen secretion. This study was conducted to localize and thereby define the cellular expression of 17alpha-hydroxylase/17,20-lyase cytochrome P450 (P450c17), the enzyme most directly responsible for androgen synthesis, in 30 GTCTs and control tissues (gonads and adrenal glands) using immuno-histochemistry (IHC). Immuno-reactivity...
[The influence of pH on cholinesterase hydrolysis of alpha-naphthylacetate in the presence of some cationic detergents]. The influence of some cationic detergents on the catalytic activity of the horse blood plasma cholinesterase in reaction of hydrolysis of alpha-naphthylacetate at different pH were investigated. It was shown, that in the absence of detergents in acid pH of the reaction medium the Km value increases, but V remain constant. In the range of pH from 8.5 to 5.0 in the presence of detergents the Km and V values are not practically changed. That is why the activation of cholinesterase hydrolysis of alpha-naphthylacetate in the presence of detergents is considerably higher than that of the neutral pH....
Molecular characterization of glycogen synthase 1 and its tissue expression profile with type II hexokinase and muscle-type phosphofructokinase in horses. Muscle glycogen synthase (GYS1) is the rate-limiting enzyme in glycogen synthesis, and its activity is regulated by the phosphorylation states of certain amino acid residues encoded by the GYS1 gene. In the present study, the authors molecularly characterized the full-length equine GYS1 (eGYS1) cDNA and found that it contains a less common polyadenylation signal (AATACA). An amino acid alignment with other mammalian GYS1 showed that the phosphorylation sites in eGYS1 are completely conserved. Genomic DNA analysis revealed that the equine-specific substitutions (Glu 16 Asp and Ala 252 Thr) were...
IgG Endopeptidase SeMac does not Inhibit Opsonophagocytosis of Streptococcus equi Subspecies equi by Horse Polymorphonuclear Leukocytes. The secreted Mac protein made by group A Streptococcus (GAS) inhibits opsonophagocytosis of GAS by human polymorphonuclear leukocytes (PMNs). This protein also has the endopeptidase activity against human immunoglobulin G (IgG), and the Cys94, His262 and Asp284 are critical for the enzymatic activity. The horse pathogen Streptococcus equi subspecies equi produces a homologue of Mac (SeMac). SeMac was characterized to determine whether SeMac has IgG endopeptidase activity and inhibits opsonophagocytosis of S. equi by horse PMNs. The gene was cloned and recombinant SeMac was overexpressed in Esc...
Enantioselective CE analysis of hepatic ketamine metabolism in different species in vitro. Ketamine, an injectable anesthetic and analgesic consisting of a racemic mixture of S-and R-ketamine, is routinely used in veterinary and human medicine. Nevertheless, metabolism and pharmacokinetics of ketamine have not been characterized sufficiently in most animal species. An enantioselective CE assay for ketamine and its metabolites in microsomal preparations is described. Racemic ketamine was incubated with pooled microsomes from humans, horses and dogs over a 3 h time interval with frequent sample collection. CE data revealed that ketamine is metabolized enantioselectively to norketamine...
Simplified method to measure glucocorticoid metabolites in faeces of horses. Glucocorticoids or their metabolites can be measured in several body fluids or excreta, including plasma, saliva, urine and faeces. In recent years the measurement of glucocorticoid metabolites (GCMs) in faeces has gained increasing attention, because of its suitability for wild populations. In horses, however, the group-specific enzyme immunoassay described so far has a limited practicability due to its complex extraction procedure. Therefore, we tested the applicability of other enzyme immunoassays for glucocorticoid metabolites. The present study clearly proved that an enzyme immunoassay (E...
Effects of glucocorticoids and interleukin-1 beta on expression and activity of aggrecanases in equine chondrocytes. To determine effects of interleukin (IL)-1 beta and glucocorticoids on total glycosaminoglycan (GAG) loss and aggrecanase-mediated matrix degradation in equine cartilage. Methods: Cartilage from 24 equine cadavers free of sepsis and musculoskeletal disease. Methods: Effects of IL-1 beta, IL-1 beta with glucocorticoids (dexamethasone and triamcinolone, 10(-6) and 10(-7)M), and glucocorticoids alone on degradation of equine articular and nasal cartilage explants were assessed by measuring GAG release in media and GAG content in cartilage. Aggrecanase-mediated cleavage within the interglobular do...
Reductive nitrosylation of ferric cyanide horse heart myoglobin is limited by cyanide dissociation. Cyanide binds to ferric heme-proteins with a very high affinity, reflecting the very low dissociation rate constant (k(off)). Since no techniques are available to estimate k(off), we report herewith a method to determine k(off) based on the irreversible reductive nitrosylation reaction to trap ferric myoglobin (Mb(III)). The k(off) value for cyanide dissociation from ferric cyanide horse heart myoglobin (Mb(III)-cyanide) was determined at pH 9.2 and 20.0 degrees C. Mixing Mb(III)-cyanide and NO solutions brings about absorption spectral changes reflecting the disappearance of Mb(III)-cyanide w...
Alkaline and acid phosphatase, β-glucuronidase and electrolyte levels in fractionated stallion ejaculates. Seminal plasma (SP) is a mixture of contents from the testes, epididymides and accessory sex glands. The sperm concentration is highest in the first few jets, or fractions, of the ejaculate, and the composition of SP varies between these fractions because accessory gland secretions are released in a specific order. The aim of this study was to compare the levels of Na, Cl, K, Mg, Ca, inorganic phosphate (Pi) and the enzymes alkaline phosphatase (AP), acid phosphatase (ACP) and β-glucuronidase (BG) in the different fractions of the ejaculate and in different stallions. All semen collections we...
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...
Expression of annexin-1 in equine leucocytes and the effects of the N-terminal annexin-1 peptide, Ac2-26, on equine neutrophil superoxide production. N-terminal peptides derived from the anti-inflammatory peptide, annexin-1, inhibit neutrophil function but can also induce pro-inflammatory effects. Although equine annexin-1 has been sequenced, its cellular expression and properties have not been reported. This study has examined whether annexin-1 is present in equine leucocytes and how the N-terminal peptide, Ac2-26, affects equine neutrophil superoxide production. Annexin-1 expression in equine neutrophils and mononuclear cells and the ability of Ac2-26 to activate neutrophil p42/44 MAPK were determined by immunoblotting. Equine neutrophil ...
