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.
Aldehyde Dehydrogenase Plays a Pivotal Role in the Maintenance of Stallion Sperm Motility. Although stallion spermatozoa produce significant quantities of reactive oxygen species, a lag between 4-hydroxynonenal (4HNE) adduction and the loss of motility in stallion spermatozoa suggests the presence of a robust aldehyde detoxification mechanism. Because there is a paucity of studies characterizing the role of aldehyde dehydrogenase (ALDH) in sperm functionality, the aim of this study was to ascertain the relationship between 4HNE production and motility and ALDH expression by stallion spermatozoa. PCR analysis revealed the presence of the ALDH1A3, ALDH1B1, and ALDH2 isoforms in these ...
HSP-1/2, a major horse seminal plasma protein, acts as a chaperone against oxidative stress. The major protein of equine seminal plasma, HSP-1/2 exhibits chaperone-like activity and protects a variety of target proteins against thermal and chemical stress conditions. Here, we show that HSP-1/2 is able to protect enzymes such as alcohol dehydrogenase and glucose-6-phosphate dehydrogenase against H2O2 induced stress, clearly demonstrating that HSP-1/2 acts as a chaperone against oxidative stress. Further, the present studies show that HSP-1/2 also inhibits lipid (linoleic acid) peroxidation by hydroxyl radicals in vitro. These results are of great significance considering that so far l...
Early cathepsin K degradation of type II collagen in vitro and in vivo in articular cartilage. To characterize the initial events in the cleavage of type II collagen mediated by cathepsin K and demonstrate the presence of the resulting products in human and equine articular osteoarthritic cartilage. Equine type II collagen was digested with cathepsin K and the cleavage products characterized by mass spectrometry. Anti-neoepitope antibodies were raised against the most N-terminal cleavage products and used to investigate the progress of collagen cleavage, in vitro, and the presence of cathepsin K-derived products in equine and human osteoarthritic cartilage. Six cathepsin K cleavage sit...
Characterization of equine cytochrome P450: role of CYP3A in the metabolism of diazepam. Research on drug metabolism and pharmacokinetics in large animal species including the horse is scarce because of the challenges in conducting in vivo studies. The metabolic reactions catalyzed by cytochrome P450s (CYPs) are central to drug pharmacokinetics. This study elucidated the characteristics of equine CYPs using diazepam (DZP) as a model compound as this drug is widely used as an anesthetic and sedative in horses, and is principally metabolized by CYPs. Diazepam metabolic activities were measured in vitro using horse and rat liver microsomes to clarify the species differences in enzy...
Trypanosoma evansi contains two auxiliary enzymes of glycolytic metabolism: Phosphoenolpyruvate carboxykinase and pyruvate phosphate dikinase. Trypanosoma evansi is a monomorphic protist that can infect horses and other animal species of economic importance for man. Like the bloodstream form of the closely related species Trypanosoma brucei, T. evansi depends exclusively on glycolysis for its free-energy generation. In T. evansi as in other kinetoplastid organisms, the enzymes of the major part of the glycolytic pathway are present within organelles called glycosomes, which are authentic but specialized peroxisomes. Since T. evansi does not undergo stage-dependent differentiations, it occurs only as bloodstream forms, it has been ...
Serum muscle-derived enzymes response during show jumping competition in horse. The effect of two jumping competitions, performed in two consecutive weekends, on serum creatine phosphokinase (CPK), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH), urea, creatinine (CREA) concentrations were evaluated in 12 healthy jumper horses. Methods: Blood sampling was performed before the 1(st) day of competition (T0), at the end of each show (J1, J2), on the day after the competition (T1); the same sampling plan was followed during the second weekend (J3, J4 and T2). Results: One-way repeated measures analysis of variance showed an increase in CPK at J1 and J2 respe...
Cathepsin K inhibition renders equine bone marrow nucleated cells hypo-responsive to LPS and unmethylated CpG stimulation in vitro. Cathepsin K (CatK) is an important enzyme regulating bone degradation and has been shown to contribute to the immune response. We have studied two inflammatory models in equine bone marrow nucleated cells (BMNCs); the LPS and the unmethylated CpG stimulation with the following objectives to: 1.determine whether CatK inhibition will alter the cytokine secretion by stimulated BMNCs; specifically IL-1β, IL-6, and TNF-α, and 2.determine the changes in BMNCs surface markers' expression and MHC II molecule under CatK inhibition. Cathepsin K inhibition promoted BMNCs viability and reduced cell apop...
Evidence for a PGF2α auto-amplification system in the endometrium in mares. In mares, prostaglandin F2α (PGF2α) secreted from the endometrium is a major luteolysin. Some domestic animals have an auto-amplification system in which PGF2α can stimulate its own production. Here, we investigated whether this is also the case in mares. In an in vivo study, mares at the mid-luteal phase (days 6-8 of estrous cycle) were injected i.m. with cloprostenol (250 µg) and blood samples were collected at fixed intervals until 72 h after treatment. Progesterone (P4) concentrations started decreasing 45 min after the injection and continued to decrease up to 24 h (P < 0.05). In t...
Pharmacokinetics and Pulmonary Distribution of Clarithromycin and Rifampicin after Concomitant and Consecutive Administration in Foals. Drug interactions often result from multiple pharmacokinetic changes, such as after rifampicin (RIF) and clarithromycin (CLA) in the treatment of abscessing lung diseases. Comedication of RIF may interact with CLA disposition by either induction of presystemic elimination processes and/or inhibition of uptake mechanisms because it regulates gene transcription and modulates function of various CYP enzymes, multidrug efflux and uptake transporters for which CLA is a substrate. To distinguish the transcriptional changes from the modulating interaction components upon CLA absorption and pulmonary ...
Production of serum amyloid A in equine articular chondrocytes and fibroblast-like synoviocytes treated with proinflammatory cytokines and its effects on the two cell types in culture. To investigate the role of the major equine acute phase protein serum amyloid A (SAA) in inflammation of equine intraarticular tissues. Methods: Articular chondrocytes and fibroblast-like synoviocytes (FLSs) from 8 horses (4 horses/cell type). Methods: Chondrocytes and FLSs were stimulated in vitro for various periods up to 48 hours with cytokines (recombinant interleukin [IL]-1β, IL-6, tumor necrosis factor-α, or a combination of all 3 [IIT]) or with recombinant SAA. Gene expression of SAA, IL-6, matrix metalloproteinases (MMP)-1 and -3, and cartilage-derived retinoic acid-sensitive protein...
Nodal Promotes Functional Luteolysis via Down-Regulation of Progesterone and Prostaglandins E2 and Promotion of PGF2α Synthetic Pathways in Mare Corpus Luteum. In the present work, we investigated the role of Nodal, an embryonic morphogen from the TGFβ superfamily in corpus luteum (CL) secretory activity using cells isolated from equine CL as a model. Expression pattern of Nodal and its receptors activin receptor A type IIB (ACVR2B), activin receptor-like kinase (Alk)-7, and Alk4, as well as the Nodal physiological role, demonstrate the involvement of this pathway in functional luteolysis. Nodal and its receptors were immune localized in small and large luteal cells and endothelial cells, except ACVR2B, which was not detected in the endothelium. Nod...
Effect of firocoxib on cyclooxygenase 2, microsomal prostaglandin E2 synthase 1, and cytosolic phospholipase A2 gene expression in equine mononuclear cells. To validate primer sets for use in reverse transcription quantitative PCR assays to measure gene expression of cytosolic phospholipase A2 (cPLA2) and microsomal prostaglandin E2 synthase 1 (mPGES1) in equine mononuclear cells and determine the effects of firocoxib, a selective cyclooxygenase 2 (COX-2) inhibitor, on COX-2, cPLA2, and mPGES1 gene expression following incubation of mononuclear cells with lipopolysaccharide (LPS). Methods: 8 healthy adult horses. Methods: Peripheral blood mononuclear cells were isolated by density gradient centrifugation and incubated at 37°C with medium alone, f...
Evaluation of species differences and the effects of storage duration and temperature on the anticollagenase efficacy of canine, feline, and equine serum on in vitro corneal degradation. OBJECTIVE To evaluate species differences and effects of storage duration and temperature on the anticollagenase efficacy of canine, feline, and equine serum on in vitro corneal degradation. SAMPLES Corneas and serum from dogs, cats, and horses. PROCEDURES Clinically normal corneas from dogs, cats, and horses were harvested within 2 hours after euthanasia. Serum samples from dogs, cats, and horses were collected and pooled by species. Corneal specimens were incubated with collagenase derived from Clostridium histolyticum, 5mM calcium chloride in saline (0.9% NaCl) solution, and feline, canine,...
Clinical Research Abstracts of the British Equine Veterinary Association Congress 2015. Colic remains a life-threatening condition in the horse. Ischaemia and reperfusion following correction of small intestinal strangulation may produce oxidative stress. The ability to withstand oxidative stress depends on antioxidant levels and may be linked to horse survival. Objective: To measure peripheral antioxidant levels in horses undergoing exploratory laparotomy with small intestinal strangulation. Methods: Case-control study. Methods: Blood and plasma were collected from horses undergoing exploratory laparotomy for small intestinal strangulation and stored at -80°C. Controls involved...
Increased susceptibility of skin from HERDA (Hereditary Equine Regional Dermal Asthenia)-affected horses to bacterial collagenase degradation: a potential contributing factor to the clinical signs of HERDA. Hereditary equine regional dermal asthenia (HERDA) is a genetic disorder of collagen resulting in fragile, hyper-extensible skin and ulcerative lesions. The predominance of skin lesions have been shown to occur on the dorsum of HERDA-affected horses. While this has been postulated to be due to increased exposure to sunlight of these areas, the precise pathological mechanism which causes this to occur is unclear. Objective: We hypothesized that an increase in collagenase activity, that has been associated with the exposure of dermal fibroblasts to sunlight, will significantly degrade the materi...
Characterization of alkaline phosphatase activity in seminal plasma and in fresh and frozen-thawed stallion spermatozoa. Alkaline phosphatase (AP) has been studied in several situations to elucidate its role in reproductive biology of the male from different mammalian species; at present, its role in horse sperm physiology is not clear. The aim of the present work was to measure AP activity in seminal plasma and sperm extracts from freshly ejaculated as well as in frozen-thawed stallion spermatozoa and to verify whether relationship exists between AP activity and sperm quality parameters. Our data on 40 freshly ejaculated samples from 10 different stallions demonstrate that the main source of AP activity is semi...
Differences in extracellular matrix remodeling in the placenta of mares that retain fetal membranes and mares that deliver fetal membranes physiologically. In mammals, placenta separation at term may involve degradation of the extracellular matrix by matrix metalloproteinases (MMPs). The activity of MMPs is modulated by TIMPs. We hypothesized that the placentas of mares that deliver fetal membranes physiologically and those that retain fetal membranes (FMR) differ in terms of histology; mRNA expression of MMP-2 and MMP-9; protein expression of MMP-2, MMP-9, and TIMP-2; and the potential activity of both MMPs. Methods: Placenta biopsies were taken from mares (n = 9; 4 FMR, 5 controls) immediately after foal expulsion. Retention was defined as fail...
Evidence for a Shared Mechanism in the Formation of Urea-Induced Kinetic and Equilibrium Intermediates of Horse Apomyoglobin from Ultrarapid Mixing Experiments. In this study, the equivalence of the kinetic mechanisms of the formation of urea-induced kinetic folding intermediates and non-native equilibrium states was investigated in apomyoglobin. Despite having similar structural properties, equilibrium and kinetic intermediates accumulate under different conditions and via different mechanisms, and it remains unknown whether their formation involves shared or distinct kinetic mechanisms. To investigate the potential mechanisms of formation, the refolding and unfolding kinetics of horse apomyoglobin were measured by continuous- and stopped-flow fluore...
EquiNox2: A new method to measure NADPH oxidase activity and to study effect of inhibitors and their interactions with the enzyme. Excessive neutrophil stimulation and reactive oxygen species (ROS) production are involved in numerous human or horse pathologies. The modulation of the neutrophil NADPH oxidase (NOX) has a great therapeutic potential since this enzyme produces superoxide anion whose most of the other ROS derive. The measurement of NOX activity by cell-free systems is often used to test potential inhibitors of the enzyme. A major drawback of this technique is the possible interferences between inhibitors and the probe, ferricytochrome c, used to measure the activity. We designed the "EquiNox2", a new pharmacol...
Characterization of neopeptides in equine articular cartilage degradation. Osteoarthritis is characterized by a loss of extracellular matrix that leads to cartilage degradation and joint space narrowing. Specific proteases, including the aggrecanases ADAMTS-4 and matrix metalloproteinase 3, are important in initiating and promoting cartilage degradation in osteoarthritis. This study investigated protease-specific and disease-specific cleavage patterns of particular extracellular matrix proteins by comparing new peptide fragments, neopeptides, in specific exogenous protease-driven digestion of a crude cartilage proteoglycan extract and an in-vitro model of early osteo...
The activity and inhibition of poly(ADP-ribose) polymerase-1 in equine peripheral blood mononuclear cells in vitro. To evaluate the poly (ADP-ribose) polymerase-1 (PARP1) enzyme and its inhibition in horses and explore its potential as a novel therapeutic target for equine intestinal ischemia-reperfusion injury by (1) identifying poly (ADP-ribose) (PAR) as an indication of PARP1 activation in equine cells using available immunoblot analytical techniques, (2) inducing PARP1 activation in an in vitro oxidative DNA damage model, (3) and demonstrating the inhibition of PARP1 in equine cells using commercially available PARP1 inhibitors. Methods: Experimental study. Methods: Blood samples were collected from sys...
Fermentation characteristics and angiotensin I-converting enzyme-inhibitory activity of Lactobacillus helveticus isolate H9 in cow milk, soy milk, and mare milk. Lactobacillus helveticus isolate H9 demonstrated high angiotensin I-converting enzyme (ACE)-inhibitory activity in previous research. Here, we evaluated the fermentation characteristics (pH, titratable acidity, free amino nitrogen, and viable bacterial counts), ACE-inhibitory activity, and contents of Val-Pro-Pro (VPP) and Ile-Pro-Pro (IPP) peptides of stored yogurt (4°C for 28 d) fermented by L. helveticus isolate H9 (initially inoculated at 4 concentrations), from cow, mare, and soy milks. During storage, the pH and titratable acidity remained stable in yogurts produced from all milk types ...
Characterization and comparison of the bacterial microbiota in different gastrointestinal tract compartments in horses. The advance of new sequencing technologies has allowed more comprehensive characterization of complex microbial communities, including the ones inhabiting the intestinal tract. The presence of extreme environmental filters, such as low pH, digestive enzymes and anaerobic conditions along the tract, acts on the selection of unique bacteria in each compartment. The intestinal microbiota has an enormous impact on the maintenance of health. However, data about the bacteria present in the different intestinal compartments of horses are sparse. In this study, high throughput sequencing was used to c...
Species-specific effects on non-enzymatic metmyoglobin reduction in vitro. Our objectives were to determine the non-enzymatic metmyoglobin reduction properties of bovine, porcine, and equine myoglobins and to characterize the effects of pre-incubation of 4-hydroxy-2-nonenal (HNE) with myoglobins on non-enzymatic metmyoglobin reduction in vitro. Purified bovine, porcine, and equine metmyoglobins (0.05 mM) were reduced at pH 5.6 and 7.4 in the presence or absence of HNE. Rates of metmyoglobin reduction were monitored by spectrophotometry, and myoglobin adducts were characterized by high-resolution mass-spectrometry. Results showed that the species origins of individual...
Effects of isoflurane and sevoflurane on the neutrophil myeloperoxidase system of horses. Volatile anaesthestics have shown to modulate the oxidative response of polymorphonuclear neutrophils (PMNs). We investigated the effects of isoflurane and sevoflurane on the degranulation of total and active myeloperoxidase (MPO) from horse PMNs and their direct interaction with MPO activity. Whole blood from horse was incubated in 1 and 2 minimal alveolar concentrations (MAC) of isoflurane or sevoflurane for 1h and PMNs were stimulated with cytochalasin B (CB) plus N-formyl-méthionyl-leucyl-phenylalanine (fMLP). After stimulation, the plasma was collected to measure total and active MPO by ...
Characterization of an L-amino acid oxidase in equine spermatozoa. This study demonstrates for the first time the presence of an L-amino acid oxidase (LAAO) enzyme in equine spermatozoa that is able to generate significant amounts of reactive oxygen species (ROS) and create a state of oxidative stress. RT-PCR analysis revealed that the mRNA for this enzyme was present in the equine testis and spermatozoa, while immunocytochemical studies demonstrated that the mature LAAO protein was located in the sperm head, particularly in the acrosomal and postacrosomal domains. Experimental studies demonstrated that the aromatic amino acids (L-phenylalanine > L-tryptop...
Crosslinkable hydrogels derived from cartilage, meniscus, and tendon tissue. Decellularized tissues have proven to be versatile matrices for the engineering of tissues and organs. These matrices usually consist of collagens, matrix-specific proteins, and a set of largely undefined growth factors and signaling molecules. Although several decellularized tissues have found their way to clinical applications, their use in the engineering of cartilage tissue has only been explored to a limited extent. We set out to generate hydrogels from several tissue-derived matrices, as hydrogels are the current preferred cell carriers for cartilage repair. Equine cartilage, meniscus, a...
Effectiveness of training programmes used in two stables of thoroughbred race horses. The purpose of this study was to compare the training methods used in two stables and their effects on selected blood parameters and race results. A total number of 36 thoroughbred race horses was examined in two groups, trained by two trainers. Twenty-four horses (group A) were trained at Sluzewiec and the remaining twelve horses (group B) were kept and trained in a private stable. The experiment lasted for five months. The activities of CPK (creatine phosphokinase) and AST (aspartate aminotransferase) and the concentration of LA (lactic acid) were determined. The speed was controlled and rec...
Evaluation of contact activation of citrated equine whole blood during storage and effects of contact activation on results of recalcification-initiated thromboelastometry. To evaluate the degree of activation of the contact pathway in citrated equine whole blood over holding times ≤ 30 minutes and assess effects of contact activation on recalcification-initiated thromboelastometry. Methods: 11 healthy adult mixed-breed horses. Methods: Blood was collected by atraumatic jugular venipuncture into prewarmed evacuated siliconized glass tubes containing citrate anticoagulant and held at 37°C for ≤ 30 minutes. Thromboelastometry was performed with an in vitro viscoelasticity (thromboelastometry) monitoring system. Factor XII and factor XI procoagulant activities ...
Serum thymidine kinase activity in clinically healthy and diseased horses: a potential marker for lymphoma. Serum thymidine kinase (sTK) activity is a tumour marker used as a prognostic indicator for lymphoma in humans, dogs and cats. The aim of this study was to evaluate the clinical utility of sTK as a biomarker for lymphoma in horses. Serum samples were collected from clinically normal horses (n = 37), horses with lymphoma (n = 23), horses with non-haematopoietic neoplasia (n = 9) and horses with inflammatory disease (n = 14). sTK was measured using a radioenzyme assay. A reference cut-off value of <2.7 U/L (mean + 2 standard deviations, SDs) was established using data fro...