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.
Dyrda K, Orzołek A, Ner-Kluza J, Wysocki P.Phosphorylation and dephosphorylation of proteins are considered to be the most important processes in sperm maturation during epididymal transit. The main aim of this study was to isolate and identify phosphoproteins from the epididymal milieu obtained from reproductively mature stallions during and out of the breeding season. With the use of 1D-PAGE and nanoLC-MS/MS, we identified phosphoproteins that fulfil various functions: regulatory, transport, motility, ubiquitination, chaperone, antioxidant, apoptotic and enzymatic. Moreover, we characterized tyrosine, serine and threonine phosphoryla...
Gu XL, Dai FY, Xiao X, Li GZ, Zhang LM, Qu WJ.The effect of Ageratina adenophora on pathological characteristics of the liver and lungs as well as serum biochemical parameters in horses were investigated. Ten horses without ingestion history of Ageratina adenophora were classified into the control group, and 10 poisoned but survived horses with 3 months ingestion history were set as the case group. Results showed that serum AST, ALT, ALP, magnesium and phosphorus were elevated significantly, while creatinine was decreased remarkably. Hematoxylin and eosin staining of liver tissues showed diffuse swelling or destruction of hepatocytes, nar...
Catalán J, Yánez-Ortiz I, Tvarijonaviciute A, González-Arostegui LG, Rubio CP, Yeste M, Miró J, Barranco I.This study investigated whether the activities of the antioxidant components of donkey seminal plasma (SP)-both enzymatic (superoxide dismutase (SOD), catalase-like (CAT), glutathione peroxidase-like (GPX), and paraoxonase type 1 (PON1)) and non-enzymatic (measured in terms of total thiol, copper-reducing antioxidant capacity (CUPRAC), ferric-reducing ability of plasma (FRAP), and Trolox equivalent antioxidant capacity (TEAC))-and oxidative stress index (OSI) are related to sperm cryotolerance. For this purpose, 15 ejaculates from jackasses (one per individual) were collected and split into tw...
Wang L, Unger L, Sharif H, Eriksson S, Gerber V, Rönnberg H.Thymidine kinase 1 (TK1) plays a key role in the synthesis of deoxythymidine triphosphate (dTTP) and is thus important for DNA replication and cell proliferation. The expression of TK1 is highest during S-phase, and it is rapidly degraded after mitosis. In cancer cells, TK1 is upregulated, resulting in leakage of excess TK1 into the blood. Consequently, serum TK1 has been used as a diagnostic and prognostic cancer biomarker, mainly in human medicine. The aims of this work were to characterize equine TK1 and to evaluate its suitability as a serum biomarker for equine lymphoma. Results: Equine T...
Fojtík L, Fiala J, Pompach P, Chmelík J, Matoušek V, Beier P, Kukačka Z, Novák P.Covalent labeling of proteins in combination with mass spectrometry has been established as a complementary technique to classical structural methods, such as X-ray, NMR, or cryogenic electron microscopy (Cryo-EM), used for protein structure determination. Although the current covalent labeling techniques enable the protein solvent accessible areas with sufficient spatial resolution to be monitored, there is still high demand for alternative, less complicated, and inexpensive approaches. Here, we introduce a new covalent labeling method based on fast fluoroalkylation of proteins (FFAP). FFAP u...
Autry JM, Svensson B, Carlson SF, Chen Z, Cornea RL, Thomas DD, Valberg SJ.We have analyzed the enzymatic activity of the sarcoplasmic reticulum (SR) Ca2+-transporting ATPase (SERCA) from the horse gluteal muscle. Horses are bred for peak athletic performance yet exhibit a high incidence of exertional rhabdomyolysis, with elevated levels of cytosolic Ca2+ proposed as a correlative linkage. We recently reported an improved protocol for isolating SR vesicles from horse muscle; these horse SR vesicles contain an abundant level of SERCA and only trace-levels of sarcolipin (SLN), the inhibitory peptide subunit of SERCA in mammalian fast-twitch skeletal muscle. Here, we re...
Rouge M, Elkhatib R, Delalande C, Cognié J, Reigner F, Barriere P, Deleuze S, Cousty M, Legendre F, Galera P, Hanoux V, Bouraima-Lelong H.Although vitamin D acts in various biological processes, it plays a critical role in the maintenance of bone health, and regulates calcium homeostasis. In humans and rodents, the main tissues involved in vitamin D metabolism are the liver and the kidneys, however it has been shown that the testis has strongly participated in its bioactivation. Indeed, in these different species, enzymes metabolizing vitamin D (CYP27A1, CYP27B1 and CYP2R1) have been demonstrated in this tissue. Moreover, men with hypogonadism have shown a decrease in circulating levels of vitamin D. In equine species, the castr...
Henry ML, Velez-Irizarry D, Pagan JD, Sordillo L, Gandy J, Valberg SJ.Horses have one of the highest skeletal muscle oxidative capacities amongst mammals, which, combined with a high glycolytic capacity, could perturb redox status during maximal exercise. We determined the effect of 30 d of oral coenzyme Q10 and N-acetyl-cysteine supplementation (NACQ) on muscle glutathione (GSH), cysteine, ROS, and coenzyme Q10 concentrations, and the muscle proteome, in seven maximally exercising Thoroughbred horses using a placebo and randomized cross-over design. Gluteal muscle biopsies were obtained the day before and 1 h after maximal exercise. Concentrations of GSH, cyste...
Sander J, Terhardt M, Janzen N.In horses, congenital defects of energy production from long-chain fatty acids have not been described so far. In contrast, inhibition of fatty acid degradation caused by the toxins hypoglycin A and methylenecyclopropylglycine from various maple species are observed frequently. These non-proteinogenic aminoacids are passed on placentally to fetuses or with collostrum or milk to newborn foals. Nevertheless, newborn foals become very rarely symptomatic. Vertical transmission apparently is not sufficient to induce clinical disease without a particular genetic constellation being present. One of t...
Ganeshpurkar A, Singh R, Kumar D, Gutti G, Sardana D, Shivhare S, Singh RB, Kumar A, Singh SK.Machine learning (ML), an emerging field in drug design, has the potential to predict toxicity, shape-based analysis of inhibitors, scoring function (SF) etc. In the present study, a homology model, docking protocol, and a dedicated SF have been developed to identify the inhibitors of horse butyrylcholinesterase (BChE) enzyme. Horse BChE enzyme has homology with human BChE and is a substitute for the screening of inhibitors. The developed homology model was validated and the active site residues were identified from Cavityplus to generate grid box for docking. The validation of docking invol...
Stanley S, Deng D, Van den Berg K, Foo HC.The abuse of performance-enhancing catecholamine-based stimulants, such as levodopa, is controlled in horse racing through the application of a regulatory threshold for the common major metabolite. However, catechol-O-methyltransferase (COMT) enzyme inhibitors can be used to restrict the catalysis of the stimulant, and so the concurrent administration of both substances would be a viable strategy to enhance racing performance while removing the risk of exceeding the threshold. A 200 mg dose of nitecapone, a COMT inhibitor, was administered to a Thoroughbred horse, and we have analysed the bl...
Suchan T, Kusliy MA, Khan N, Chauvey L, Tonasso-Calvière L, Schiavinato S, Southon J, Keller M, Kitagawa K, Krause J, Bessudnov AN, Bessudnov AA....DNA hybridization-capture techniques allow researchers to focus their sequencing efforts on preselected genomic regions. This feature is especially useful when analysing ancient DNA (aDNA) extracts, which are often dominated by exogenous environmental sources. Here, we assessed, for the first time, the performance of hyRAD as an inexpensive and design-free alternative to commercial capture protocols to obtain authentic aDNA data from osseous remains. HyRAD relies on double enzymatic restriction of fresh DNA extracts to produce RNA probes that cover only a fraction of the genome and can serve a...
Wehrman RF, Genschel U, Charli A, Kanthasamy AG, Allbaugh RA, Ben-Shlomo G.To establish a physiologically relevant ex vivo model of equine corneal epithelial wound healing. Methods: Fourteen equine corneas were randomly assigned to one of two groups: wounded (n = 8) or unwounded (n = 6) controls. In the wounded group, the axial corneal epithelium was removed by applying a 6 mm filter paper disk soaked in 1N-NaOH for 60 s. Corneas were subsequently cultured using an air-liquid interface model. Evaluation of corneal healing was performed daily, and culture medium was collected. Corneas were randomly assigned to undergo processing via histopathology and RNAscope i...
Kósa CA, Nagy K, Szenci O, Baska-Vincze B, Andrásofszky E, Szép R, Keresztesi Á, Mircean M, Taulescu M, Kutasi O.A severe form of recurrent exertional rhabdomyolysis occurs enzootically in a well-defined region of Transylvania, Harghita county. At the highest lying two settlements (more than 800 m above sea level), the prevalence of equine rhabdomyolysis is between 17 and 23%, while in the neighbouring villages in the valley it is less than 2%. The objective of our study was to clarify the role of selenium and vitamin E in the high prevalence of rhabdomyolysis in that region. Soil and hay samples were collected from each area to evaluate mineral content. Ten horses from the non-affected and 20 horses fro...
Fasanello DC, Su J, Deng S, Yin R, Colville MJ, Berenson JM, Kelly CM, Freer H, Rollins A, Wagner B, Rivas F, Hall AR, Rahbar E, DeAngelis PL....TNF-α-stimulated gene 6 (TSG-6) protein, a TNF-α-responsive hyaladherin, possesses enzymatic activity that can catalyze covalent crosslinks of the polysaccharide hyaluronic acid (HA) to another protein to form heavy chain-hyaluronic acid (HC-HA) complexes in pathological conditions such as osteoarthritis (OA). Here, we examined HA synthase and inflammatory gene expression; synovial fluid HA, TNF-α, and viscosity; and TSG-6-mediated HC-HA complex formation in an equine OA model. The objectives of this study were to (1) evaluate the TNF-α-TSG-6-HC-HA signaling pathway across multiple joint t...
Delvescovo B, Tomlinson J, DeNotta S, Hodge E, Bookbinder L, Mohammed HO, Divers TJ.Serum biochemistry results and presence of fibrosis on liver biopsies are frequently used as prognostic indicators in horses with liver dysfunction. The objective of this retrospective multicenter study was to determine if the magnitude of abnormal liver specific biochemical tests such as bile acids (BA), direct bilirubin and gamma-glutamyltransferase (GGT), or the presence of fibrosis reported on liver biopsies was associated with prognosis in horses with liver dysfunction. Eighty-two horses older than one year, examined at four referral hospitals in the eastern United States, with BA values ...
Ellerbrock RE, Podico G, Scoggin KE, Ball BA, Carossino M, Canisso IF.The expression pattern and distribution of sex steroid receptors and steroidogenic enzymes during development of the equine accessory sex glands has not previously been described. We hypothesized that equine steroidogenic enzyme and sex steroid receptor expression is dependent on reproductive status. Accessory sex glands were harvested from mature stallions, pre-pubertal colts, geldings, and fetuses. Expression of mRNA for estrogen receptor 1 (ESR1), estrogen receptor 2 (ESR2), androgen receptor (AR), 3β-Hydroxysteroid dehydrogenase/Δ5-4 isomerase (3βHSD), P450,17α hydroxylase, 17-20 lyase...
Hales EN, Habib H, Favro G, Katzman S, Sakai RR, Marquardt S, Bordbari MH, Ming-Whitfield B, Peterson J, Dahlgren AR, Rivas V, Ramirez CA, Peng S....Equine neuroaxonal dystrophy/equine degenerative myeloencephalopathy (eNAD/EDM) is an inherited neurodegenerative disorder associated with a vitamin E deficiency within the first year of life. Vitamin E consists of 8 isoforms metabolized by the CYP4F2 enzyme. No antemortem diagnostic test currently exists for eNAD/EDM. Objective: Based on the association of α-tocopherol deficiency with the development of eNAD/EDM, we hypothesized that the rate of α-tocopherol, but not γ-tocopherol or tocotrienol metabolism, would be increased in eNAD/EDM-affected horses. Methods: Vitamin E metabolism: Proof...
Juśkiewicz J, Fotschki B, Jaworska J, Siemieniuch M.Gastrointestinal microbiota play a key role in the nutrients digestion and hence maintaining animal health and welfare. The diet offered to the animals in captivity may differ considerably from that on natural pastures. In a stabled maintenance system, horses have a limited choice of habitat and feed. Time spend for feeding is relevant for equine welfare because the reduction of the time devoted for foraging may be responsible for inducing gastric inflammation and ulceration. Therefore, in the present study, it was hypothesized that fecal bacterial fermentative processes differ between free-ro...
Mouithys-Mickalad A, Storms N, Franck T, Ceusters J, de la Rebière de Pouyade G, Deby-Dupont G, Serteyn D.Experimental laminitis, characterized by a failure of the dermal-epidermal interface of the foot, can be induced in horses by the oral administration of a black walnut extract (BWE). In the early phase of this severe and painful disease, an activation of neutrophil occurs, with the release of myeloperoxidase (MPO), a pro-oxidant enzyme of neutrophils, in plasma, skin, and laminar tissue. Juglone, a naphthoquinone derivative endowed with redox properties, is found in walnuts and has been incriminated in this neutrophil activation. We report for the first time the inhibitory activity of juglone ...
Ortega Granda O, Valle C, Shannon A, Decroly E, Canard B, Coutard B, Rabah N.Venezuelan equine encephalitis virus (VEEV) is a reemerging arthropod-borne virus causing encephalitis in humans and domesticated animals. VEEV possesses a positive single-stranded RNA genome capped at its 5' end. The capping process is performed by the nonstructural protein nsP1, which bears methyl and guanylyltransferase activities. The capping reaction starts with the methylation of GTP. The generated mGTP is complexed to the enzyme to form an mGMP-nsP1 covalent intermediate. The mGMP is then transferred onto the 5'-diphosphate end of the viral RNA. Here, we explore the specificities of the...
Ellero N, Lanci A, Ferlizza E, Andreani G, Mariella J, Isani G, Castagnetti C.The matrix metalloproteinases (MMPs) are a family of enzymes involved in extracellular matrix remodeling. MMPs are secreted in a latent form and activated by local and infiltrating cells. MMP-2 and -9 are the most studied in reproduction and have been detected in bovine, ovine, equine and human placenta. There is only one study on MMPs in the equine amniotic fluid (AF) reporting a decrease in the activity of MMP-2 in case of premature delivery. The aim of this study was focused on MMP-2 and -9 activity in AF collected at parturition from mares with normal or high-risk pregnancy. High-risk preg...
Haralambus R, Florczyk A, Sigl E, Gültekin S, Vogl C, Brandt S, Schnierer M, Gamerith C, Jenner F.Synovial sepsis is a commonly occurring, potentially career-ending or even life-threatening orthopaedic emergency. Diagnosis of synovial sepsis is currently primarily based on synovial fluid analysis, which often leaves diagnostic ambiguity due to overlap of clinicopathological parameters between septic and aseptic inflammatory synovitis. Objective: To evaluate the reliability of lysozyme (LYS), myeloperoxidase (MPO) and elastase (ELT) as biomarkers for synovial sepsis in horses using a photometric assay to measure increased enzyme activity. Methods: Prospective, single-blinded, analytical, cl...
Di Stefano DA, Vinci R, Capparè P, Gherlone EF.Enzyme-deantigenic equine bone (EDEB) is a substitute of autogenous bone. Mixing it with plasma rich in growth factors (PRGF) seems a viable option to achieve enhanced bone formation in alveolar bone augmentation surgeries. This retrospective study aims to first report the histomorphometric and clinical outcomes achieved when using the EDEB/PRGF mixture for performing sinus augmentation procedures followed by delayed implant placement. Records of 11 patients who underwent 14 sinus augmentation surgeries using EDEB/PRGF followed by delayed implant placement were retrospectively collected and an...
Fouché N, Oesch S, Gerber V, Richter H, Howard J, Peters LM.Sorbitol dehydrogenase (SDH) activity is one of the most sensitive and specific markers for hepatocellular injury in horses, but its reported lability makes it impractical for use in many clinical settings. To date, stability of SDH in equine samples has only been evaluated in a limited number of studies in serum samples of horses with activities within reference intervals. The objective of the study was to determine pre-analytical stability of equine SDH activity in heparinized plasma stored at different temperatures for up to 72 h. Twenty client-owned horses admitted to a veterinary teaching...
Sandbaumhüter FA, Gittel C, Larenza-Menzies MP, Theurillat R, Thormann W, Braun C.The enantioselectivity of the pharmacokinetics of methadone was investigated in anesthetized Shetland ponies after a single intravenous (0.5 mg/kg methadone hydrochloride; n = 6) or constant rate infusion (0.25 mg/kg bolus followed by 0.25 mg/kg/h methadone hydrochloride; n = 3) administration of racemic methadone. Plasma concentrations of l-methadone and d-methadone and their major metabolites, l- and d-2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), respectively, were analyzed by CE with highly sulfated γ-cyclodextrin as chiral selector and electrokinetic analyte injection from...
Amaral A, Fernandes C, Szóstek-Mioduchowska A, Rebordão MR, Skarzynski DJ, Ferreira-Dias G.Endometrosis is a reproductive pathology that is responsible for mare infertility. Our recent studies have focused on the involvement of neutrophil extracellular traps enzymes, such as elastase (ELA), in the development of equine endometrosis. Noscapine (NOSC) is an alkaloid derived from poppy opium with anticough, antistroke, anticancer, and antifibrotic properties. The present work investigates the putative inhibitory in vitro effect of NOSC on collagen type I alpha 2 chain () mRNA and COL1 protein relative abundance induced by ELA in endometrial explants of mares in the follicular or mid-lu...
Siwińska N, Żak A, Pasławska U.Diagnosis of acute kidney injury (AKI) in horses is difficult at the subclinical stage, due to nonspecific clinical signs. The aim of this study was to evaluate the concentrations of selected serum and urinary biomarkers in healthy horses, horses at risk of AKI, and those with clinical AKI. Methods: Thirty healthy horses, 30 horses at risk of AKI and 11 horses with clinical AKI and azotaemia were included in the study. Serum and urinary neutrophil gelatinase-associated lipocalin (NGAL) and cystatin C were measured using commercially available enzyme immunoassay tests. Results: The median and (...
Budik S, Walter I, Leitner MC, Ertl R, Aurich C.In the horse, mobility of the conceptus is required for maternal recognition of pregnancy depending on secretion of prostaglandins by the conceptus. The aim of this study was to determine the expression and localization of key enzymes of the different pathways leading to synthesis of prostaglandin E2 and F2α in the equine conceptus during the mobility phase. Enzyme expression was analyzed via quantitative RT-PCR in total RNA samples of equine conceptuses collected on days 10 (n = 5), 12 (n = 12), 14 (n = 5) and 16 (n = 7) from healthy mares. Relative abundance of cyclooxygenase ()-2 mRNA was ...
Capistrano da Silva E, Gibson DJ, Jeong S, Zimmerman KL, Smith-Fleming KM, Martins BDC.Matrix metalloproteinases (MMPs)-2 and -9 are present in corneal ulcers, and an imbalance between MMPs and tissue inhibitors of metalloproteinases (TIMPs) leads to further corneal degradation. Amniotic membrane homogenate (AMH) has proteolytic properties beneficial for corneal healing, but it is unknown whether AMH possesses TIMPs or effectively inhibits MMP-2 and MMP-9 activity. Objective: To determine if bovine and equine AMH reduce in vitro MMP-2 and MMP-9 activities associated with the presence of TIMPs. Methods: Undiluted and diluted twofold series (0-fold to 16-fold dilutions) of equine ...
Hildebrand DP, Burk DL, Maurus R, Ferrer JC, Brayer GD, Mauk AG.The spectroscopic and structural properties of the His93Tyr variant of horse heart myoglobin have been studied to assess the effects of replacing the proximal His residue of this protein with a tyrosyl residue as occurs in catalases from various sources. The variant in the ferric form exhibits electronic spectra that are independent of pH between pH 7 and 10, and it exhibits changes in absorption maxima and intensity that are consistent with a five-coordinate heme iron center at the active site. The EPR spectrum of the variant is that of a high-spin, rhombic system similar to that reported for...
Legacki EL, Ball BA, Corbin CJ, Loux SC, Scoggin KE, Stanley SD, Conley AJ.Equine fetuses have substantial circulating pregnenolone concentrations and thus have been postulated to provide significant substrate for placental 5α-reduced pregnane production, but the fetal site of pregnenolone synthesis remains unclear. The current studies investigated steroid concentrations in blood, adrenal glands, gonads and placenta from fetuses (4, 6, 9 and 10 months of gestational age (GA)), as well as tissue steroidogenic enzyme transcript levels. Pregnenolone and dehydroepiandrosterone (DHEA) were the most abundant steroids in fetal blood, pregnenolone was consistently higher bu...
Fojtík L, Fiala J, Pompach P, Chmelík J, Matoušek V, Beier P, Kukačka Z, Novák P.Covalent labeling of proteins in combination with mass spectrometry has been established as a complementary technique to classical structural methods, such as X-ray, NMR, or cryogenic electron microscopy (Cryo-EM), used for protein structure determination. Although the current covalent labeling techniques enable the protein solvent accessible areas with sufficient spatial resolution to be monitored, there is still high demand for alternative, less complicated, and inexpensive approaches. Here, we introduce a new covalent labeling method based on fast fluoroalkylation of proteins (FFAP). FFAP u...
Szóstek AZ, Siemieniuch MJ, Lukasik K, Galvão AM, Ferreira-Dias GM, Skarzynski DJ.Accurate regulation of the reproductive cycle and successful implantation depend on proper functioning of the endometrium. The aim of this study was to determine whether mRNA transcription of specific enzymes responsible for prostaglandin (PG) synthesis (prostaglandin-endoperoxide synthase, PTGS-2; prostaglandin F(2α) synthase, PGFS; and prostaglandin E(2) synthases, PGES) and PG concentrations in endometrial extracts would change in moderate (Kenney's Category II) and severe phases of fibrosis (Kenney's Category III; endometrosis), compared with healthy endometrium (Kenney's Category I), dur...
Papas M, Arroyo L, Bassols A, Catalán J, Bonilla-Correal S, Gacem S, Yeste M, Miró J.This study compared the activities of four antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; glutathione peroxidase, GPX; and glutathione reductase, GSR) in the seminal plasma of stallions and jackasses. Eighteen stallion ejaculates and 24 jack ejaculates were collected through an artificial vagina. Seminal plasma was obtained by several centrifugations at 3000×g and 4 °C for 10 min, and activities of SOD, CAT, GPX and GSR were subsequently determined. We also evaluated whether the collecting season had any influence on the activities of these four enzymes in both stallions ...
Toledano M, Asady S, Toledano-Osorio M, García-Godoy F, Serrera-Figallo MA, Benítez-García JA, Osorio R.Native collagen-based membranes are used to guide bone regeneration; but due to their rapid biodegradation, this treatment is often unpredictable. The purpose of this study was to investigate the biodegradability of natural collagen membranes. Three non-cross-linked resorbable collagen barrier membranes were tested: Derma Fina (porcine dermis), Evolution Standard (equine pericardium) and Duo-Teck (equine lyophilized collagen felt). 10 × 10 mm pieces of membranes were submitted to three different degradation procedures: (1) hydrolytic degradation in phosphate buffer solution, (2) enzyme resist...
Gadsby PM, Peterson J, Foote N, Greenwood C, Thomson AJ.Magnetic-circular-dichroism (m.c.d.) spectra over the wavelength range 300-2000 nm at room temperature and at 4.2K of horse heart cytochrome c are reported at a series of pH values between 7.8 and 11.0, encompassing the alkaline transition. The effect of glassing agents on the e.p.r. spectrum at various pH values is also reported. Comparison of these results with spectra obtained for the n-butylamine adduct of soybean leghaemoglobin support the hypothesis that lysine is the sixth ligand in the alkaline form of horse heart cytochrome c. The m.c.d. and e.p.r. spectra of horse heart cytochrome c ...
Kang CH, Ferguson-Miller S, Margoliash E.1. The steady state kinetics for the oxidation of ferrocytochrome c by yeast cytochrome c peroxidase are biphasic under most conditions. The same biphasic kinetics were observed for yeast iso-1, yeast iso-2, horse, tuna, and cicada cytochromes c. On changing ionic strength, buffer anions, and pH, the apparent Km values for the initial phase (Km1) varied relatively little while the corresponding apparent maximal velocities varied over a much larger range. 2. The highest apparent Vmax1 for horse cytochrome c is attained at relatively low pH (congruent to 6.0) and low ionic strength (congruent to...
Abdullaev ZKh, Bodrova ME, Chernyak BV, Dolgikh DA, Kluck RM, Pereverzev MO, Arseniev AS, Efremov RG, Kirpichnikov MP, Mokhova EN, Newmeyer DD....A cytochrome c mutant lacking apoptogenic function but competent in electron transfer and antioxidant activities has been constructed. To this end, mutant species of horse and yeast cytochromes c with substitutions in the N-terminal alpha-helix or position 72 were obtained. It was found that yeast cytochrome c was much less effective than the horse protein in activating respiration of rat liver mitoplasts deficient in endogenous cytochrome c as well as in inhibition of H(2)O(2) production by the initial segment of the respiratory chain of intact rat heart mitochondria. The major role in the di...
Franck T, Grulke S, Deby-Dupont G, Deby C, Duvivier H, Peters F, Serteyn D.Equine inflammatory disease is accompanied by a neutrophil activation resulting in the release of granulocytic enzyme myeloperoxidase (MPO). To measure MPO in horse plasma as marker of neutrophil activation, the authors purified equine neutrophil MPO and developed a specific enzyme immunoassay using 2 specific polyclonal antibodies obtained from rabbit (primary antibody) and guinea pig (secondary antibody). The sandwich complex "primary antibody-MPO-secondary antibody" was detected using a goat anti-guinea pig immunoglobulin antibody conjugated to alkaline phosphatase. The enzyme-linked immuno...
Gusson F, Carletti M, Albo AG, Dacasto M, Nebbia C.To complete a studyaimed at investigating the pattern of the basal activities of liver xenobioticmetabolizing enzymes in major and minor species intended for meat production, microsomal carboxylesterases and some conjugating enzyme activities were determined and compared in liver preparations from horses, cattle, pigs, rabbits and broiler chicks, using the rat as a reference species. Horses and broiler chicks exhibited a lower microsomal carboxylesterase activity towards indophenyl or p-nitrophenyl acetate than that measured in cattle or pig subfractions. Among food-producing species, the rate...
Baumber J, Sabeur K, Vo A, Ball BA.The objective of this study was to examine the influence of reactive oxygen species (ROS) on equine sperm capacitation. Motile equine spermatozoa were separated on a discontinuous Percoll gradient, resuspended at 10 x 10(6)ml in Tyrode's medium supplemented with BSA (0.5%) and polyvinyl alcohol (0.5%) and incubated at 39 degrees C for 2h with or without the xanthine (X; 0.1mM)-xanthine oxidase (XO; 0.01 U/ml) system or NADPH (0.25 mM). The importance of hydrogen peroxide or superoxide for capacitation was determined by the addition of catalase (CAT; 150 U/ml) or superoxide dismutase (SOD; 150 ...
Valberg SJ, Ward TL, Rush B, Kinde H, Hiraragi H, Nahey D, Fyfe J, Mickelson JR.Seven related Quarter Horse foals that died by 7 weeks of age were examined for glycogen branching enzyme (GBE) deficiency. Clinical signs varied from stillbirth, transient flexural limb deformities, seizures, and respiratory or cardiac failure to persistent recumbency. Leukopenia (5 of 5 foals) as well as high serum creatine kinase (CK; 5 of 5), aspartate transaminase (AST; 4 of 4), and gamma glutamyl transferase (GGT; 5 of 5) activities were present in most foals, and intermittent hypoglycemia was present in 2 foals. Gross postmortem lesions were minor, except for pulmonary edema in 2 foals....
Siess W, Lapetina EG.Phospholipase C has been studied in homogenates, total particulate and soluble fractions of horse and human platelets. This enzyme, assayed with exogenous L-3-phosphatidyl[14C]inositol, is predominantly localized in the soluble fraction and its distribution parallels that of lactate dehydrogenase. A small percentage of activity present in the particulate fraction seems to be due to contamination with soluble enzyme. Enzyme from horse and human platelets appears identical, having a Km of 0.10-0.15 mM, acid pH optimum (pH 5.5) and showing Ca2+-dependency and weak inhibition by deoxycholate. Anal...
Kindig CA, McDonough P, Erickson HH, Poole DC.Within the moderate exercise intensity domain, the speed of oxygen uptake (V(O(2))) kinetics at the transition to a higher metabolic rate is thought to be limited by an inertia of the oxidative machinery. Nitric oxide (NO)-induced inhibition of O(2) consumption within the electron transport chain may contribute to this inertia. This investigation tested the hypothesis that a reduction or removal of any such NO effect via infusion of N(omega)-nitro-L-arginine methyl ester (L-NAME; a NOS inhibitor) would speed V(O(2)) kinetics at the onset of moderate exercise. Five Thoroughbred geldings underwe...
Boyer RF, Clark HM, LaRoche AP.The reductive release of ferritin iron by several naturally occurring o-diphenols was studied. The initial rate of iron release was quantified by spectrophotometric measurement of the Fe(ferrozine)3(2+) complex, which absorbs maximally at 562 nm. The initial rate of iron release was dependent upon o-diphenol concentration, but not on the concentration of the chromophoric chelating agent, ferrozine, Stoichiometric measurements resulted in a ratio of 2Fe(II) released per molecule of o-diphenol. The series of o-diphenols studied included, caffeic acid, chlorogenic acid, dihydrocaffeic acid, 3,4-d...
Gustchina A, Kervinen J, Powell DJ, Zdanov A, Kay J, Wlodawer A.Equine infectious anemia virus (EIAV), the causative agent of infectious anemia in horses, is a member of the lentiviral family. The virus-encoded proteinase (PR) processes viral polyproteins into functional molecules during replication and it also cleaves viral nucleocapsid protein during infection. The X-ray structure of a complex of the 154G mutant of EIAV PR with the inhibitor HBY-793 was solved at 1.8 A resolution and refined to a crystallographic R-factor of 0.136. The molecule is a dimer in which the monomers are related by a crystallographic twofold axis. Although both the enzyme and t...
Goubran Botros H, Poncet P, Rabillon J, Fontaine T, Laval JM, David B.A new allergen from horse dander, Equ c 5 has been purified. Its biochemical and biophysical properties have been characterized and compared with those of Equ c 1, Equ c 2 and Equ c 4. Their molecular masses, determined by mass spectrometry, were 22 kDa for Equ c 1, 16 kDa for Equ c 2, 18.7 kDa for Equ c 4 and 16.7 kDa for Equ c 5. Their pI values were between 3.8 and 5.25. Equ c 2 and Equ c 5 are not glycosylated, while Equ c 4 contains a tri-antennary tri-sialylated N-linked glycan. Linkages of terminal N-acetylneuraminic acid to galactose were: alpha-(2-->6) in Equ c 4, and both alpha-(2...
Couto MA, Harwig SS, Lehrer RI.Equine neutrophil antimicrobial peptide 2 (eNAP-2), a recently described antimicrobial peptide isolated from equine neutrophils, was found to selectively inactivate microbial serine proteases (subtilisin A and proteinase K) without inhibiting mammalian serine proteases (human neutrophil elastase, human cathepsin G, and bovine pancreatic trypsin). Although the primary structure of eNAP-2 resembled that of several known antiproteases that belong to the 4-disulfide core peptide family, this pattern of selectivity is unique. eNAP-2 formed a noncovalent complex with native subtilisin A or proteinas...
Lee CM, Kisiday JD, McIlwraith CW, Grodzinsky AJ, Frisbie DD.It is well documented that osteoarthritis (OA) can develop following traumatic joint injury and is the leading cause of lameness and subsequent wastage of equine athletes. Although much research of injury induced OA has focused on cartilage, OA is a disease that affects the whole joint organ. Methods: In this study, we investigated the impact of synovial cells on the progression of an OA phenotype in injured articular cartilage. Injured and control cartilage were cultured in the presence of synoviocytes extracted from normal equine synovium. Synoviocytes and cartilage were evaluated for catabo...
Gunn HM.Histochemical profiles of individual muscle fibres were established using myosin adenosine triphosphatase (myosin ATPase), succinate dehydrogenase (SDHase), and glycogen phosphorylase (GPase) reactions in three muscles (semitendinosus, diaphragm, and pectoralis transversus) of the horse and dog. The major histochemical difference between fibres lies in their myosin ATPase activity; fibres can be subdivided into those with a high and those with a low activity. In horse muscle, all fibres have a high activity of GPase. In the diaphragm and pectoralis transversus, all fibres have a high SDHase ac...
Griko YV, Freire E, Privalov G, van Dael H, Privalov PL.The energetics of the temperature-induced unfolding of equine lysozyme was studied calorimetrically and compared with that of two structurally homologous proteins: hen egg white lysozyme and alpha-lactalbumin. The structure of each of these proteins is characterized by the presence of a deep cleft that divides the molecule into two regions called the alpha and beta domains. In equine lysozyme and alpha-lactalbumin the latter domain specifically binds Ca2+. It is shown that, in contrast to hen egg white lysozyme in which the alpha and beta domains unfold as a single cooperative unit, in equine ...
Thorpe CT, Stark RJ, Goodship AE, Birch HL.Damage to the flexor tendons, particularly the superficial digital flexor tendon (SDFT), is one of the most common musculoskeletal injuries sustained by horses competing in all disciplines. Our previous work has shown that SDFTs from different individuals show a wide variation in mechanical strengths; this is important clinically as it may relate to predisposition to injury. The high mechanical strength of tendon relies on the correct orientation of collagen molecules within fibrils and stabilisation by the formation of chemical cross-links between collagen molecules. It is not known whether t...
Ascenzi P, Ciaccio C, Sinibaldi F, Santucci R, Coletta M.Upon interaction with bovine heart cardiolipin (CL), horse heart cytochrome c (cytc) changes its tertiary structure disrupting the heme-Fe-Met80 distal bond, reduces drastically the midpoint potential out of the range required for its physiological role, binds CO and NO with high affinity, and displays peroxidase activity. Here, the effect of CL on peroxynitrite isomerization by ferric cytc (cytc-Fe(III)) is reported. In the absence of CL, hexa-coordinated cytc does not catalyze peroxynitrite isomerization. In contrast, CL facilitates cytc-Fe(III)-mediated isomerization of peroxynitrite in a d...
Ollivier FJ, Brooks DE, Kallberg ME, Komaromy AM, Lassaline ME, Andrew SE, Gelatt KN, Stevens GR, Blalock TD, van Setten GB, Schultz GS.To examine in vitro effects of various antiproteolytic compounds on activity of matrix metalloproteinase (MMP)-2 and -9 in the tear film of horses with active corneal ulcers. Methods: Samples of tear film obtained from the eyes of 34 horses with active ulcerative keratitis. Methods: Horses were sedated, and tear samples were collected from the lower fornix of 34 ulcerated eyes by use of capillary tubes. The protease inhibitors 0.2% EDTA, 0.1% doxycycline, 10% N-acetylcysteine (NAC), 0.1% solution of a modified dipeptide that contains hydroxamic acid (ie, ilomostat), 0.1% alpha1-proteinase inhi...
Brama PA, van den Boom R, DeGroott J, Kiers GH, van Weeren PR.Matrix metalloproteinases (MMPs) are considered candidate biomarkers for both physiological and pathological tissue remodelling because of their key role in articular cartilage homeostasis. As disruption of the collagenous architecture is thought to be pivotal in chronic degenerative diseases such as osteoarthritis (OA), the collagenases form an interesting subset of the MMPs. The significance of any biomarker in synovial fluid (SF) can be assessed properly only when fluctuations in patterns induced by physiological processes such as development and growth, and by external influences and inter...
Esparza I, Brock JH.The effect of trypsin digestion on iron-saturated and iron-free (apo) human, rabbit, bovine, pig and horse tranferrins has been studied. Iron-binding fragments were produced only from iron-saturated pig and bovine transferrins although some cleavage of the polypeptide chain occurred in all cases. The apo-transferrins were generally degraded to a greater extent than the corresponding iron-saturated proteins. The ability of the different transferrins to donate iron to rabbit reticulocytes varied in the order rabbit approximately pig greater than human approximately horse greater than bovine. Try...
Johansson J, von Bahr-Lindström H, Jeck R, Woenckhaus C, Jörnvall H.The primary structure of the mitochondrial form of horse liver aldehyde dehydrogenase has been determined, utilizing peptide analyses and homology with other enzyme forms. The subunit exhibits N-terminal heterogeneity in size similar to that for the corresponding human mitochondrial protein, the longest form having 500 residues. Catalase was identified as a contaminant of the preparations. All four pairs within a set of aldehyde dehydrogenases can now be compared, including the same two species variants (horse and human) for both the cytosolic and mitochondrial enzyme, revealing characteristic...
Gatti JL, Druart X, Guérin Y, Dacheux F, Dacheux JL.SDS-PAGE analysis of luminal fluid from the ram testis and epididymis revealed a protein of about 105 kDa in the fluid in the caput epididymal region. The molecular mass of this fluid protein shifted from 105 kDa to 94 kDa in the distal caput epididymidis and remained at 94 kDa in the lower regions of the epididymis. The possible sperm origin of this protein was suggested by the decrease in intensity of a 105-kDa compound on the sperm plasma membrane extract and by its total disappearance from the fluid of animals with impaired sperm production caused by scrotal heating. The 94-kDa protein was...
