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.
Misumi K, Vilim V, Clegg PD, Thompson CC, Carter SD.This study was designed to assay cartilage oligomeric matrix protein (COMP) in equine synovial fluids and to compare the concentration in synovial fluids from normal horses with joint diseased horses. The relationship between the COMP degradation and the matrix metalloproteinase activity in synovial fluids was also investigated. Methods: Using COMP antigen prepared from equine articular cartilage and murine monoclonal antibody (12C4) raised against human COMP, an inhibition ELISA was developed. COMP in equine synovial fluids from normal and diseased joints was quantified. Metalloproteinase act...
Albrecht BA, MacLeod JN, Daels PF.In the pregnant mare, luteal estrogen production increases at the onset of equine chorionic gonadotropin (eCG) secretion by endometrial cups. In previous studies, we have demonstrated that eCG stimulates luteal androgen and estrogen production in pregnant mares. To further elucidate the regulation of steroidogenesis within the equine corpus luteum (CL) of pregnancy, we examined the expression of 3beta-hydroxysteroid dehydrogenase (3beta-HSD), cytochrome P450 17alpha-hydroxylase/17,20 lyase (P450(17alpha)) and cytochrome P450 aromatase (P450(arom)) in luteal tissue samples collected during dies...
Mungall BA, Kyaw-Tanner M, Pollitt CC.Utilizing an in vitro laminitis explant model, we have investigated how bacterial broth cultures and purified bacterial proteases activate matrix metalloproteinases (MMPs) and alter structural integrity of cultured equine lamellar hoof explants. Four Gram-positive Streptococcus spp. and three Gram-negative bacteria all induced a dose-dependent activation of MMP-2 and MMP-9 and caused lamellar explants to separate. MMP activation was deemed to have occurred if a specific MMP inhibitor, batimastat, blocked MMP activity and prevented lamellar separation. Thermolysin and streptococcal pyrogenic ex...
Davenport CL, Boston RC, Richardson DW.To investigate the effects of enrofloxacin and magnesium deficiency on explants of equine articular cartilage. Methods: Articular cartilage explants and cultured chondrocytes obtained from adult and neonatal horses. Methods: Full-thickness explants and cultured chondrocytes were incubated in complete or magnesium-deficient media containing enrofloxacin at concentrations of 0, 1, 5, 25, 100, and 500 microg/ml. Incorporation and release of sulfate 35S over 24 hours were used to assess glycosaminoglycan (GAG) synthesis and degradation. An assay that measured binding of dimethylmethylene blue dye ...
Stewart JM, Blakely JA, Johnson MD.Non-covalent modification of cytochrome c may have implications for electron transport and energy metabolism. We examined the interaction of various fatty acids (FAs), their coenzyme A and carnitine esters, and fatty alcohols with horse heart ferrocytochrome c. A comparison of FAs indicated a minimum chain length of 14 carbons was required for significant effect on the ferroheme chromophore and major changes in electronic spectra. Coenzyme A and carnitine esters interacted less strongly than FAs whereas long-chain alcohols did not interact with the protein. We found a single, saturable FA bind...
Serrano AL, Quiroz-Rothe E, Rivero JL.Twenty-four 4-year-old Andalusian (Spanish breed) stallions were used to examine the plasticity of myosin heavy chain (MHC) phenotype and the metabolic profile in horse skeletal muscle with long-term endurance-exercise training and detraining. Sixteen horses underwent a training programme based on aerobic exercises for 8 months. Afterwards, they were kept in paddocks for 3 months. The remaining eight horses were used as controls. Three gluteus medius muscle biopsy samples were removed at depths of 20, 40 and 60 mm from each horse before (month 0), during (month 3) and after (month 8) training,...
Hagen SJ, Carswell CW, Sjolander EM.We have measured the effect of temperature and denaturant concentration on the rate of intrachain diffusion in an unfolded protein. After photodissociating a ligand from the heme iron of unfolded horse cytochrome c, we use transient optical absorption spectroscopy to measure the time scale of the diffusive motions that bring the heme, located at His18, into contact with its native ligand, Met80. Measuring the rate at which this 62 residue intrachain loop forms under both folding and unfolding conditions, we find a significant effect of denaturant on the chain dynamics. The diffusion of the cha...
Stradaioli G, Sylla L, Zelli R, Verini Supplizi A, Chiodi P, Arduini A, Monaci M.The reproductive characteristics and seminal carnitine and acetylcarnitine content as well as carnitine acetyltransferase activity of young Maremmano stallions (n=25) are reported. The stallions were subjected to semen collection in November and January; in each trial two ejaculates were collected 1h apart. The total motile morphologically normal spermatozoa (TMMNS) and the progressively motile spermatozoa at collection and during storage at +4 degrees C were evaluated. Seminal L-carnitine (LC), acetylcarnitine (AC), pyruvate and lactate were measured using spectrophotometric methods, whereas ...
Hamilton A, Harrington D, Sutcliffe IC.Acid phosphatases hydrolyse phosphomonoesters at acidic pH in a variety of physiological contexts. The recently defined class C family of acid phosphatases includes the 32 kDa LppC lipoprotein of Streptococcus equisimilis. To define further the distribution of acid phosphatases in the genus Streptococcus we have examined the equine pathogens Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus. Whole cell assays indicated that these organisms possess two acid phosphatases with activity optima at pH 5.0 and pH 6.0-6.5 and that only the former of these was, like LppC, resis...
Marshall DE, Mortishire-Smith RJ, Houghton E, Gower DB.Oestradiene-3,17-diol and oestratriene-3,17-diol (or the diol of Heard's ketone (3-hydroxy-5(10),6,8-oestratriene-17-one) have been extracted on a large scale from pooled urines and allantoic fluid obtained from pregnant mares. Initial purification was achieved using column chromatography, and further purification by high performance liquid chromatography or silver nitrate (argentation) thin layer chromatography. The steroids were characterised using gas chromatography-mass spectrometry. Positions of the double bonds in ring B of oestradienediol were deduced on the basis of results of ultravio...
Taylor KC, Vitello LB, Erman JE.The ionization of 4-nitroimidazole to 4-nitroimidazolate was investigated as a function of ionic strength. The apparent pKa varies from 8.99 to 9.50 between 0.001 and 1.0 M ionic strength, respectively, at 25 degrees C. The ionic strength dependence of this ionization is anomalous. The binding of 4-nitroimidazole by horse metmyoglobin was studied between pH 5.0 and 11.5 and as a function of ionic strength between 0.01 and 1.0 M. The association rate constant is pH-dependent, varying from 24 M(-1)s(-1) at pH 5 to a maximum value of 280 M(-1)s(-1) at pH 9.5 and then decreasing to 10 M(-1)s(-1) a...
Jouglin M, Robert C, Valette JP, Gavard F, Quintin-Colonna F, Denoix JM.Early detection of osteoarthritis in horses represents a challenge for equine practitioners. Several biological markers have been implicated in the pathological processes involved in articular cartilage destruction. To further document cartilage matrix proteases production, synovial fluid was collected from 14 horses (90 joints) before they were subjected to euthanasia. Growth macroscopic examination of the joints gave information on cartilage alterations. Samples were analyzed for matrix metalloproteinase (MMPs) activities by gelatin zymography and tumor necrosis factor alpha (TNF-alpha) cyto...
Rickards KJ, Page CP, Lees P, Cunningham FM.Neutrophils are recruited to the lungs of horses with chronic obstructive pulmonary disease (COPD) and exhibit increased activity after antigen challenge. Phosphodiesterase type4 (PDE4) inhibitors have been shown to attenuate human neutrophil activation. The aim of this study was to establish the PDE isoenzyme profile of equine neutrophils using isoenzyme selective inhibitors to determine if these compounds should be evaluated in horses with COPD. Total cAMP and cGMP dependent PDE activity was no different in neutrophils from normal (156.2+/-7.1 and 6.8+/-0.6 pmol/min/mg for cAMP and cGMP, res...
Svistunenko DA, Sharpe MA, Nicholls P, Blenkinsop C, Davies NA, Dunne J, Wilson MT, Cooper CE.The paramagnetic species in human metHb and horse metmyoglobin (metMb) have been studied at low temperature using EPR spectroscopy. The high-spin (HS) haem signal in aquometMb has a greater rhombic distortion than the HS metHb signal. Nevertheless, the individual line width (g=6) is smaller in metMb than in metHb, consistent with non-identical signals from the alpha and beta Hb subunits. Three low-spin (LS) haem forms are present in metHb, while metMb has only two. The major LS form in both proteins is the alkaline species (with OH(-) at the sixth co-ordination position). The minor LS forms ar...
Greenaway EC, Cunningham FM, Goode NT.Phorbol esters, which activate protein kinase C (PKC), stimulate equine eosinophil superoxide production and adherence. After showing that superoxide production could be inhibited by the nonselective PKC inhibitors, staurosporine and bisindolymaleimide I, the PKC isotypes in equine eosinophils were characterized, because evidence suggests that individual isotypes may play distinct roles in regulating eosinophil function. Western blots demonstrated that equine eosinophils expressed PKC alpha, beta, delta, epsilon, iota, and zeta. However, unlike the equine neutrophil, the majority of the PKC wa...
White SD, Rosychuk RA, Outerbridge CA, Fieseler KV, Spier S, Ihrke PJ, Chapman PL.Our objective was to determine if thiopurine methyltransferase (TPMT), the enzyme important in the metabolism of azathioprine in human beings, is detectable in red blood cell lysates (RBCL) of healthy dogs, cats, and horses. Values for TPMT activity were determined from blood collected from 20 healthy dogs, cats, and horses. The TPMT activity in each animal's RBCL was determined using a radioenzymatic end point involving TPMT-facilitated metabolism of 6-mercaptopurine to 6-methylmercaptopurine (6-MMP). One unit of TPMT activity represents the formation of 1 nmol of 6-MMP per milliliter of pack...
von Rechenberg B, Guenther H, McIlwraith CW, Leutenegger C, Frisbie DD, Akens MK, Auer JA.To define the release of nitric oxide (NO), prostaglandin E2 (PGE2), and the neutral metalloproteinases (NMPs) in horses with subchondral cystic lesions (SCL) and to study bone resorption triggered by conditioned media of fibrous tissue of SCL in vitro. Methods: Equine explant cultures of fibrous tissue of SCL, and synovial membrane and articular cartilage of normal horses and horses affected with moderate and severe osteoarthritis were performed. NO, PGE2, and NMP concentrations of media samples were measured, and osteoclast formation and activation was studied in vitro. Methods: Experiment 1...
van de Lest CH, van den Hoogen BM, van Weeren PR, Brouwers JF, van Golde LM, Barneveld A.Osteochondrosis (OC) is a disturbance in the process of endochondral ossification, a process in which cartilage is mineralised and transformed into bone. In this process different biochemical events occur, of which the cartilage component has been studied so far almost exclusively. In this study we concentrated on the biochemical characterisation of normal and osteochondrotic subchondral bone, by analysis of enzyme activities, DNA content and phospholipids (PL). In subchondral bone, lysyl oxidase and both total and bone alkaline phosphatase activity were significantly increased in all degrees ...
Ball BA, Gravance CG, Medina V, Baumber J, Liu IK.To characterize the activity of catalase in equine semen. Methods: 15 stallions of known and unknown reproductive history. Methods: Seminal plasma was collected from raw equine semen by centrifugation, and samples of seminal plasma were frozen prior to assay for catalase activity. Tissue samples (n = 3 stallions) from the bulbourethral gland, prostate gland, vesicular gland, and testis were homogenized, and cauda epididymal fluid was collected for determination of catalase activity. Catalase activity was determined as an enzyme kinetic assay by the disappearance of H2O2 as measured by ultravio...
Raulo SM, Sorsa TA, Maisi PS.To determine whether samples of tracheal epithelial lining fluid (TELF) obtained from horses have elastinolytic activity characteristic of metalloproteinases, to compare elastinolytic activity in TELF obtained from healthy horses and horses with chronic obstructive pulmonary disease (COPD), and to determine whether chemically modified tetracycline-3 (CMT-3) inhibits elastinolytic activity in TELF ANIMALS: 10 horses with COPD and 10 healthy control horses. Methods: Zymography and fluorometry were used to measure elastinolytic activity, and EDTA was used to inhibit elastinolytic activity and ver...
Forsell PK, Lindberg A, Karlsson S, Lindgren JA, Claesson HE.It has been demonstrated that equine neutrophils, but not eosinophils, require exogenous arachidonic acid for calcium ionophore A23187-induced leukotriene synthesis. Because cytosolic phospholipase A(2) (cPLA(2)) plays an essential role in leukotriene formation in leukocytes, we investigated the presence of a functional cPLA(2) in equine neutrophils. To determine whether cPLA(2) from neutrophils was catalytically active, we purified the enzyme >6,500 fold with 3% recovery from equine neutrophils. The full-length cDNA sequence encoded a 749-amino acid protein. The deduced amino acid sequence...
Yu H, Raymonda JW, McMahon TM, Campagnari AA.This article reviews the recent development of two solid-phase chemiluminescence-based techniques, fluorogenic-chemiluminescence (FCL) and electro-chemiluminescence (ECL) for detection of biological threat agents. Both techniques entail a labeled sandwich immunoassay. The objectives of this work are to develop advanced techniques for sensitive and effective detection of a target analyte, particularly in cases where the analysis includes complex samples containing multiple contaminating factors. Other important considerations in developing such detection techniques include the ease of use, the ...
Tkachenko AG, Winston GW.Preincubation of horse liver alcohol dehydrogenase (HLADH) with the oxidative agent, tert-butyl hydroperoxide (tBOOH) results in a twofold stimulation of the ethanol dehydrogenase activity of this enzyme. This stimulation was dependent on tBOOH concentration up to 100 mM; above this concentration tBOOH did not further stimulate ethanol oxidation by HLADH. Active-site-directed reagents and classical ADH binary complexes were used to probe the possible mechanism of this activating effect. The rate and extent of stimulation by tBOOH is strongly reduced by binary complexes with NAD(+) or NADH, who...
Deriusheva SE, Loginova IuA, Chiriaeva OG, Iasinetskaia NI, Efimov AM.Restriction endonuclease in situ digestion of metaphase chromosomes gives an opportunity to reveal strips with different structure within GC-rich pericentric heterochromatin of the domestic horse and the wild Przewalski horse. Blocks of heterochromatin, which are insensitive to HaeIII and brightly stained with chromomycin A3 after restriction enzyme digestion, are localized on the border with euchromatin in the majority of chromosomes of Equus caballus and E. przewalskii. In contrast to chromosome 5 of E. caballus, acrocentric chromosomes of E. prezewalskii which are homologous to this chromos...
Bosken JM, Lehner AF, Hunsucker A, Harkins JD, Woods WE, Karpiesiuk W, Carter WG, Boyles J, Fisher M, Tobin T.Isoxsuprine is routinely recovered from enzymatically-hydrolyzed, post-administration urine samples as parent isoxsuprine in equine forensic science. However, the specific identity of the material in horse urine from which isoxsuprine is recovered has never been established, although it has long been assumed to be a glucuronide conjugate (or conjugates) of isoxsuprine. Using ESI/MS/MS positive mode as an analytical tool, urine samples collected 4-8 h after isoxsuprine administration yielded a major peak at m/z 554 that was absent from control samples and resisted fragmentation to daughter ions...
Okumura M, Fujinaga T.The role of keratan sulphate (KS) as a metabolic marker of cartilage was evaluated using an in vitro model of equine articular cartilage. Articular cartilage was harvested from clinically healthy 6-month-old foals (n = 3). Chondrocytes were centrifuged and cultured as pellets. Chondrocyte pellets were stimulated by insulin-like growth factor-I alpha (IGF-I alpha) or interleukin-1 alpha (IL-1 alpha) for 2 weeks. The concentrations of sulphated glycosaminoglycans (GAG) and KS in the culture media were measured by a 1,9-dimethyl-methylene blue (DMMB) colorimetric assay and an inhibition enzyme-li...
Belin F, Goudet G, Duchamp G, Gérard N.The objective of the present study was to determine the changes in follicular fluid steroid concentrations and in granulosa cell steroidogenic enzyme expression during the follicular phase, in relation to follicular size and physiological status in the mare. Follicular fluid and follicular cells were recovered by ultrasound-guided follicular punctures either around the time of emergence of the dominant follicle, at the end of the dominant follicle growth, or at the preovulatory stage, after injection of gonadotropin to induce ovulation. Cellular relative amounts of steroidogenic acute regulato...
Okumura M, Tagami M, Fujinaga T.The role of keratan sulphate (KS) as a marker of cartilage metabolism was evaluated by using an in vitro model of equine articular cartilage. Articular cartilage was harvested from clinically healthy 6-month-old foals (n=3). Chondrocytes were centrifuged and cultured as pellets. Chondrocyte pellets were stimulated by insulin-like growth factor (IGF)-Ialpha or interleukin (IL)-1alpha for 2 weeks. The sulfated glycosaminoglycans (GAG) and antigenic KS concentrations in the culture media were measured by a 1,9-dimethyl-methylene blue (DMMB) colorimetric assay and an inhibition ELISA using a 1/14/...
Gunther MR, Sturgeon BE, Mason RP.The reaction between metmyoglobin (metMb) and hydrogen peroxide has been known since the 1950s to produce globin-centered free radicals. The direct electron spin resonance spectrum of a solution of horse metMb and hydrogen peroxide at room temperature consists of a multilined signal that decays in minutes at room temperature. Comparison of the direct ESR spectra obtained from the system under N(2)- and O(2)-saturated conditions demonstrates the presence of a peroxyl radical, identified by its g-value of 2.014. Computer simulations of the spectra recorded 3 s after the mixture of metMb and H(2)...
von Rechenberg B, McIlwraith CW, Akens MK, Frisbie DD, Leutenegger C, Auer JA.Nitric oxide (NO), prostaglandin E2 (PGE2), and the activity of neutral metalloproteinases (NMPs) were measured in conditioned media of equine synovial membrane and articular cartilage explant cultures from horses with normal joints (n = 7) and from horses affected with moderate (n = 7) or severe osteoarthritis (n = 14) as judged by macroscopic appearance. Normal articular cartilage appeared glossy and bluish-white, was of normal thickness and showed no evidence of discolouration, fibrillation or other cartilage discontinuity. Slight discolouration and fibrillation or minor clefts of the carti...
Schwarz B, Ertl R, Zimmer S, Netzmann Y, Klein D, Schwendenwein I, Hoven RV.The aim of this study was to determine the occurrence and frequency of a mutation in the gene coding for skeletal muscle glycogen synthase type 1 (GYS-1), which is the cause of equine polysaccharide storage myopathy (PSSM) type 1 in a population of 50 Haflingers. GYS-1 genotyping of 50 Haflingers was performed with a validated restriction fragment length polymorphism (RFLP) assay. The second aim was to compare resting and post-exercise muscle enzyme activities as well as parameters of glucose metabolism in blood between horses with and without the mutation. Nine of the 50 Haflingers were ident...
Siemieniuch MJ, Kozdrowski R, Szóstek-Mioduchowska AZ, Raś A, Nowak M.Endometritis is one of the most important causes of infertility in mares. Mares may suffer from endometritis outside the breeding season; however, pathological condition is often undiagnosed in anestrus. The aim of this study was to examine whether the secretion profiles of prostaglandin F2α (PGF2α), prostaglandin E2 (PGE2), and a metabolite of prostacyclin I2 (6-keto-PGF1α) differ in endometrial tissue cultures of heavy draft mares in anestrus with endometritis compared to those without endometritis. The endometrial biopsies were collected from 51 heavy draft mares. Inclusion criteria for ...
Sandholm M, Vidovic A, Puotunen-Reinert A, Sankari S, Nyholm K, Rita H.The discriminating ability of 15 parameters alone or in combinations, including results from analysis of plasma endotoxin, the Nycomed plasma D-Dimer test and phospholipase A2, were analyzed to predict morbidity and mortality in equine gastrointestinal colic. Endotoxaemia was a characteristic feature of the colic horses. The problem of adequately predicting nonsurvivors among colic horses required several parameters to be included in the logistic model: if the "classical parameters", (heart rate, respiratory rate, PCV, anion gap) were included in the model, addition of plasma D-dimer, phosphol...
Mizukami T, Abe Y, Maki K.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...
Pickles KJ, Brooks AC, Rickards KJ, Cunningham FM.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 ...
Godovac-Zimmermann J, Conti A, Liberatori J, Braunitzer G.beta-Lactoglobulin isolated from horse colostrum is heterogeneous and contains two components: beta-lactoglobulin I and beta-lactoglobulin II. These two proteins are monomeric and show differences in their electrophoretic mobilities, chain lengths and primary structures. The complete amino-acid sequence of beta-lactoglobulin II was determined by automated Edman degradation of the intact protein and of the peptides derived from these by digestion with trypsin or chymotrypsin and by chemical cleavage with cyanogen bromide. Unlike other beta-lactoglobulins which contain 162 amino acids, horse bet...
Inkerman PA, Scott K, Runnegar MT, Hamilton SE, Bennett EA, Zerner B.Chicken, sheep, and horse liver carboxylesterases have been purified by procedures involving ammonium sulfate fractionation, ion-exchange chromatography and gel filtration on Sephadex. The actual yields of the procedures described were as follows: chicken, 1 g from 2 kg of liver powder (chloroform-acetone); sheep, 200 mg from 400 g of powder (chloroform-acetone); horse, 230 mg from 800 g of powder (acetone). The purified enzymes are free of non-carboxyl-esterase protein as shown by gel electrophoresis, although they do contain electrophoretic variants. The equivalent weight of the chicken enzy...
Torbeck RL, Prieur DJ.Cartilaginous defects were created in the radiocarpal joints of 12 horses. Synovial fluid cytologic features, lysozyme activity, and beta-glucuronidase activity were monitored for 16 days. A comparison was made of plasma lysozyme and beta-glucuronidase activity and of synovial fluid lysozyme, beta-glucuronidase, and leukocyte concentrations. Plasma lysozyme was found to be independent of synovial fluid lysozyme activity. Synovial fluid lysozyme was significantly increased (P less than 0.001) in all joints with surgically induced defects (group I) compared with controls (arthrocentesis done; gr...
Stradaioli G, Sylla L, Zelli R, Verini Supplizi A, Chiodi P, Arduini A, Monaci M.The reproductive characteristics and seminal carnitine and acetylcarnitine content as well as carnitine acetyltransferase activity of young Maremmano stallions (n=25) are reported. The stallions were subjected to semen collection in November and January; in each trial two ejaculates were collected 1h apart. The total motile morphologically normal spermatozoa (TMMNS) and the progressively motile spermatozoa at collection and during storage at +4 degrees C were evaluated. Seminal L-carnitine (LC), acetylcarnitine (AC), pyruvate and lactate were measured using spectrophotometric methods, whereas ...
Herszberg B, McCue ME, Larcher T, Mata X, Vaiman A, Chaffaux S, Chérel Y, Valberg SJ, Mickelson JR, Guérin G.Glycogen storage diseases or glycogenoses are inherited diseases caused by abnormalities of enzymes that regulate the synthesis or degradation of glycogen. Deleterious mutations in many genes of the glyco(geno)lytic or the glycogenesis pathways can potentially cause a glycogenosis, and currently mutations in fourteen different genes are known to cause animal or human glycogenoses, resulting in myopathies and/or hepatic disorders. The genetic bases of two forms of glycogenosis are currently known in horses. A fatal neonatal polysystemic type IV glycogenosis, inherited recessively in affected Qu...
Pellegrini A, von Fellenberg R.The principal alpha-1-protease inhibitor of horse was fractionated by classical methods and analysed with a modified fibrinogen-agarose gel electrophoretic method of high sensitivity and resolving power. Starting with an electrophoretically homogeneous inhibitor in unfractionated serum, two isoinhibitor bands became apparent after fractionation with (NH4)2SO4 and DEAE-cellulose DE-52 ion-exchange chromatography. The isoinhibitors differed in electrophoretic migration and in the elution pattern from Sephadex G-100 gel filtration, but possessed identical antigenic determinants and enzyme specifi...
Rickards KJ, Page CP, Lees P, Cunningham FM.Neutrophils are recruited to the lungs of horses with chronic obstructive pulmonary disease (COPD) and exhibit increased activity after antigen challenge, which may contribute to inflammation and lung damage. Inhibition of phosphodiesterase isoenzymes (PDEs) has been shown to attenuate human neutrophil functions including superoxide production, leukotriene (LT)B4 biosynthesis, enzyme and chemokine release. As equine neutrophils contain predominantly the isoenzyme, PDE4, the present study was undertaken to investigate the effects of rolipram, a PDE4 inhibitor, on equine neutrophil function. For...
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...
Potempa J, Wunderlich JK, Travis J.Three structurally related but functionally different serpins from horse plasma were isolated and characterized. In spite of their identical N-terminal sequences, which show some similarity to that of human alpha 1-proteinase inhibitor, the reactive-centre loops of each of these proteins show extensive variation. Only inhibitor I, with a P1 methionine residue, resembles human alpha 1-PI with regard to (a) similarity of amino acid sequence in the vicinity of the reactive-site peptide bond, (b) broad inhibitory specificity, (c) sensitivity to oxidative inactivation and (d) high rate of reactivit...
Schmitz A, Zielinski J, Dick B, Mevissen M.Testosterone (TES) 6-β-hydroxylation is a significant metabolic step in the biotransformation of TES in human liver microsomes and reflects cytochrome P450 (CYP) 3A4/5 specific metabolic activity. Several CYP3A enzymes have been annotated in the horse genome, but functional characterization is missing. This descriptive study investigates TES metabolism in the horse liver in vitro and the qualitative contribution of three CYP3A isoforms of the horse. Metabolism of TES was investigated by using equine hepatocyte primary cultures and liver microsomes. Chemical inhibitors were used to determine t...
Morozova L, Haezebrouck P, Van Cauwelaert F.The thermal denaturation of Ca(2+)- and apo-forms of equine lysozyme was followed by using far and near UV circular dichroism and intrinsic fluorescence methods. The difference found between the temperature dependence of the ellipticity at 222 nm and 287 nm, which show two stages in the thermal transition, and those at 228 nm and 294 nm, which indicate only one stage over a wide range of temperatures reflects that different subdivisions of the protein molecule are characterized by a different stability, cooperativity and pathway of denaturation. The first transition, reflected in the increase ...
Penrod LV, Allen RE, Turner JL, Limesand SW, Arns MJ.Increased secretion of prostaglandin F(2)α (PGF(2)α) within the uterus because of uterine inflammation can cause luteolysis and result in early embryonic loss. Supplementation with polyunsaturated fatty acids (PUFAs) has been shown to influence PG production in many species, although the effects on the mare remain unknown. The present study aimed to determine fatty acid uptake in equine endometrial explants and evaluate their influence on PG secretion and expression of enzymes involved in PG synthesis in vitro. Equine endometrial explants were treated with 100 μM arachidonic acid, eicosapen...
Lindström H, Mazari AMA, Musdal Y, Mannervik B.Equine glutathione transferase A3-3 (EcaGST A3-3) belongs to the superfamily of detoxication enzymes found in all higher organisms. However, it is also the most efficient steroid double-bond isomerase known in mammals. Equus ferus caballus shares the steroidogenic pathway with Homo sapiens, which makes the horse a suitable animal model for investigations of human steroidogenesis. Inhibition of the enzyme has potential for treatment of steroid-hormone-dependent disorders. Screening of a library of FDA-approved drugs identified 16 out of 1040 compounds, which at 10 μM concentration afforded at ...
Haffner JC, Fecteau KA, Eiler H.To determine whether tetanus antitoxin, equine serum, and acetylcysteine, which are currently used in the treatment of equine corneal ulcer, inhibit the digestion of equine corneal collagen when exposed to collagenase in vitro. Methods: Corneas from 40 adult horses. Methods: Sections of equine corneas were incubated with saline, a solution of bacterial collagenase in saline, bacterial collagenase in saline plus equine tetanus antitoxin, bacterial collagenase in saline plus equine serum, or bacterial collagenase in saline plus acetylcysteine. Each one of the collagenase inhibitors was tested at...
Moslemi S, Vibet A, Papadopoulos V, Camoin L, Silberzahn P, Gaillard JL.Cytochrome P-450 aromatase was purified by five chromatographic steps from adult stallion testis. It was first separated from NADPH-cytochrome P-450 reductase (reductase) on omega-aminohexyl-Sepharose 4B then purified to homogeneity on concanavalin A-Sepharose 4B, hydroxyapatite-Sepharose 4B, DEAE-Sepharose CL-6B and on a second hydroxyapatite-Sepharose 4B. On the other hand, purifications of the equine testicular and rat liver reductases, which allowed the reconstitution of aromatase activity in vitro, were achieved for each species in one chromatographic step on an adenosine 2',5'-diphosphat...
Riley SC, Thomassen R, Bae SE, Leask R, Pedersen HG, Watson ED.Profound hormonally controlled tissue remodelling occurs in the equine ovary for follicle growth and development, and also for the alteration in follicle shape directed towards the ovulation fossa, the site where ovulation occurs. The aim of this study was to examine the spatial and temporal regulation of matrix metalloproteinases (MMP)-2 and MMP-9, important enzymes in tissue remodelling, during follicle growth, and ovulation. Using gelatin substrate zymography, we measured these MMPs in follicular fluid of large anovulatory follicles collected during spring transition, early dominant follicl...
Meyer W, Kacza J, Hornickel IN, Schoennagel B.Using immunohistochemistry and transmission electron microscopy (TEM), the esophagus epithelia of seven domesticated mammals (horse, cattle, goat, pig, dog, laboratory rat, cat) of three nutrition groups (herbivorous, omnivorous, carnivorous) were studied to get first information about energy generation, as demonstrated by succinate dehydrogenase (SDH) activities. Distinct reaction intensities could be observed in all esophageal cell layers of the different species studied reflecting moderate to strong metabolic activities. The generally strong staining in the stratum basale indicated that new...
Satué K, Fazio E, Medica P, Miguel L, Gardón JC.In the present review, the authors, based on the multiple functions performed by the liver, analyze the multiple biochemical and hematological changes as an expression of altered liver function in the horse. The liver performs important metabolic functions related to the synthesis, degradation, and excretion of various substances. Modification of these functions can be evaluated and diagnosed by determining serum concentrations of several serum analytes, including enzymes and other endogenous substances. Hepatocellular enzymes, such as sorbitol dehydrogenase-SDH and glutamate dehydrogenase-GLD...
Żak A, Siwińska N, Chełmecka E, Bażanów B, Romuk E, Adams A, Niedźwiedź A, Stygar D.The study aims to assess the impact of age, pituitary pars intermedia dysfunction (PPID) and insulin dysregulation (ID) in horses on selected oxidative stress markers. The study includes 32 horses, divided into three groups: "young" adult group (aged 8-16 years old) "geriatric" group (aged 18-24 years old) and the "PPID" group (aged 15-31 years old). The PPID group was further divided into two subgroups: PPID ID+ and PPID ID- based on presence or absence of ID. We measured serum antioxidant stress markers in all horses: total oxidant status (TOS), total antioxidant capacity (TAC), ceruloplasmi...
Higuchi T, Ichijo S, Osame S, Ohishi H.In order to clarify the cause of white muscle disease (W.M.D.) in foals, tocopherol and selenium concentrations in serum and glutathione peroxidase activities in blood were measured. Examination was made on the samples from horses affected with W.M.D., the foal kept with them in the same stable, the foals kept in the stables without affected foals, and respective mares. The heavy-breed horses in Fukuoka prefecture and Tokachi district were also examined for comparison. Serum tocopherol levels of these foals were normal because after intake of colostrum. Mares of affected foals showed lower toc...
Wilkins PA, Ducharme NG, Lowe JE, Schwark WS, Meschter C, Erb HN.To assess right colic artery blood flow and relevance of xanthine dehydrogenase/xanthine oxidase after experimentally induced strangulation obstruction and reperfusion of the colon, 5 ponies were subjected to 2.5 hours of complete ischemia of the left dorsal and ventral colons, allowed to recover from surgery, and monitored during a 48-hour reperfusion period. Five ponies were subjected to sham surgery and served as controls. All ponies had a Doppler ultrasound blood flow monitor implanted on the right colic artery near the pelvic flexure 10 to 14 days prior to the ischemic period. Colic arter...
Nilsson K, Larsson PO.Horse liver alcohol dehydrogenase was immobilized on glycerylpropyl-silica (10 micron, 1000-A pores) activated with 2,2,2-trifluoroethanesulfonyl chloride (tresyl chloride). The coupling and activity yield was almost 100%. The coenzyme-binding sites were equivalent and virtually unaffected by the immobilization process, as judged from Scatchard plots and active-site titrations. The silica-bound enzyme, packed in steel columns, was integrated with HPLC equipment and then successfully used for chromatography of adenine nucleosides, adenine nucleotides, and triazine dyes. Dissociation constants w...
Visser MB, Pollitt CC.Laminitis is known to involve deregulation of proteases and destruction of the lamellar basement membrane with the host inflammatory response also playing a role. Leukocyte infiltration has been well characterized in the black walnut model of laminitis induction, but not in carbohydrate induced models. Increased gene expression of multiple cytokines, including IL-6, has also been implicated in laminitis development. Using real time PCR, immunohistochemistry and zymography methods, we characterize leukocyte infiltration and IL-6 gene expression in oligofructose (OF) induced laminitis. As well, ...
Mohammed HO, Divers TJ, Kwak J, Omar AH, White ME, de Lahunta A.To investigate the influence of oxidative stress in terms of antioxidant capacity and lipid peroxidation on the probability of motor neuron disease (MND) in horses. Methods: 88 horses with MND (cases) and 49 controls. Methods: Blood samples were collected from all horses enrolled, and RBCs and plasma were harvested. Activity of the enzyme erythrocytic superoxide dismutase 1 (SOD1) was determined in the RBCs. Plasma concentrations of α-tocopherols and β-carotenes and activity of glutathione peroxidase were also evaluated. Degree of lipid peroxidation was measured by determining plasma concent...
Derochette S, Serteyn D, Mouithys-Mickalad A, Ceusters J, Deby-Dupont G, Neven P, Franck T.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...
