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.
Villa L, Gazzonis AL, Diezma-Diaz C, Perlotti C, Zanzani SA, Ferrucci F, Álvarez-García G, Manfredi MT.Besnoitiosis is an emerging parasitic disease of equids. Italy is one of the few European countries where the circulation of Besnoitia spp. antibodies was demonstrated. In this study, a case of clinical besnoitiosis in two donkeys in northern Italy is reported. The two animals were clinically examined. Serum and blood samples were analyzed for the detection of Besnoitia spp. antibodies and for hematology, biochemistry, and enzyme activity, respectively. ITS-1 PCR and sequencing were carried out on DNA extracted from skin biopsies. Clinical examination revealed numerous scleral pearls in eyes o...
Borba LA, Nogueira CEW, Bruhn FRP, da Silva GC, Feijó LS, Canisso IF, Curcio BDR.Neonatal sepsis is a leading cause of neonatal death during the first-week postfoaling. Despite recent advancements in the diagnosis and treatment of sepsis in the newborn foal, the non-specific clinical signs and subtle nature of this disease may result in delayed diagnosis until severe progression of the disease; thus, early detection of sepsis remains critical for a favourable outcome. This study aimed to identify early blood markers as predictive of sepsis on foals. Thirty-five foals were allocated into three groups: healthy control foals (n=7) and foals born from mares with placentitis: s...
Gretler SR, Finno CJ, Kass PH, Knych HK.In humans, the drug metabolizing enzyme CYP2D6 is highly polymorphic resulting in substantial differences in the metabolism of drugs including anti-arrhythmics, neuroleptics, and opioids. The objective of this study was to phenotype a population of 100 horses from five different breeds and assess differences in the metabolic activity of the equine CYP2D6 homolog using codeine as a probe drug. Administration of a probe drug is a common method used for patient phenotyping in human medicine, whereby the ratio of parent drug to metabolite (metabolic ratio, MR) can be used to compare relative enzym...
Rivera Del Alamo MM, Katila T, Palviainen M, Reilas T.Intrauterine devices block luteolysis in cyclic mares, but the underlying mechanism is unknown. To clarify the mechanisms, the protein profile of the endometrial secretome was analyzed using two-dimensional difference gel electrophoresis (2D-DIGE). Twenty-seven mares were classified according to whether they were inseminated (AI) or had an intrauterine device (IUD), a water-filled plastic sphere, inserted into the uterus on Day 3 after ovulation. Uterine lavage fluids were collected on Day 15 from pregnant inseminated mares (AI-P; n = 8), non-pregnant inseminated mares (AI-N; n = 4), and m...
Kareskoski M, Vakkamäki J, Laukkanen K, Palviainen M, Johannisson A, Katila T.Matrix metalloproteinase (MMP)-2 and MMP-9 are gelatinases that take part in several reproductive processes. The aim of this study was to measure levels of MMP-2 and MMP-9 in fractionated stallion ejaculates, and to evaluate the association between these components and semen quality, and sperm longevity during cooled storage. Semen quality were assessed separately for sperm-rich fractions (HIGH), sperm-poor fractions (LOW), and whole ejaculate samples (WE) from 33 stallions. After cooled storage with SP either present or removed, sperm motility and DFI were determined. The relative activity of...
Amaral A, Fernandes C, Rebordão MR, Szóstek-Mioduchowska A, Lukasik K, Pinto-Bravo P, Telo da Gama L, Jan Skarzynski D, Ferreira-Dias G.Neutrophils can originate neutrophil extracellular traps (NETs). Myeloperoxidase (MPO) is a peroxidase found in NETs associated to equine endometrosis and can be inhibited by 4-aminobenzoic acid hydrazide (ABAH). Metallopeptidases (MMPs) participate in extracellular matrix stability and fibrosis development. The objectives of this in vitro work were to investigate, in explants of mare's endometrium, (i) the ABAH capacity to inhibit MPO-induced collagen type I (COL1) expression; and (ii) the action of MPO and ABAH on the expression and gelatinolytic activity of MMP-2/-9. Explants retrieved from...
Du G, Feng W, Cai H, Ma Z, Liu X, Yuan C, Shi J, Zhang B.Composting has been a rational method to dispose of horse or wildlife animal manures, especially in the developed cities with horse clubs and wildlife parks. However, few studies have focused on the mechanism and improvement methods for composting the horse or wildlife animal manures. In this study, we investigated the effect of exogenous compound enzymes on thermophilic composting, which could potentially support the management of horse and wildlife animal manures. With the presence of exogenous enzymes, the duration of high temperature (> 60 °C) was significantly prolonged (p <...
Current drug metabolismDecember 19, 2020
Volume 22, Issue 3 215-223 doi: 10.2174/1389200221666201217141025
Hu SX, Ernst K, Benner CP, Feenstra KL.Pharmacokinetic and pharmacodynamic assessment of ester-containing drugs can be impacted by hydrolysis of the drugs in plasma samples post blood collection. The impact is different in the plasma of different species. Objective: This study evaluated the stability of a prodrug, ketoprofen methylester (KME), in commercially purchased and freshly collected plasma of mouse, rat, dog, cat, pig, sheep, cattle and horse. Methods: KME hydrolysis was determined following its incubation in commercially purchased and freshly collected plasma of those species. Different esterase inhibitors were evaluated f...
Rossi G, Richardson A, Jamaludin H, Secombe C.Paraoxonase-1 (PON-1) activity is a new inflammatory and oxidative marker. Technical effects and biological factors could affect the accuracy of PON-1 activity measurement. We investigated the effects of storage at different temperatures, repeated freeze-thaw cycles, interferences from hemolytic, lipemic, and icteric samples, and seasonal effects on PON-1 activity in horses. We evaluated 2 substrates with an automated spectrophotometer. Ten equine serum samples were stored under different conditions. Although storage at room (21°C) or refrigeration (4°C) temperature induced a statistically s...
Scavone D, Sgorbini M, Borges AS, Oliveira-Filho JP, Vitale V, Paltrinieri S.Paraoxonase-1 (PON-1) is an antioxidant enzyme, whose activity decreases during the acute phase response in many species. Little is known about PON-1 and its role as a negative acute phase protein during septic inflammation in horses, but promising findings about its utility in diagnosing SIRS and predicting the outcome in diseased horses, were recently highlighted. The objective of the study was to investigate the behaviour of PON-1 in horses after experimentally induced endotoxemia. To this aim, PON-1 activity was measured on 66 plasma samples collected from six clinically healthy mares, pre...
Škerlová J, Ismail A, Lindström H, Sjödin B, Mannervik B, Stenmark P.Organotin compounds are highly toxic environmental pollutants with neurotoxic and endocrine-disrupting effects. They are potent inhibitors of glutathione transferases (GSTs), thus impeding their detoxication and antioxidant functions. Several GSTs, including equine GST A3-3 (EcaGST A3-3), exhibit steroid double-bond isomerase activity and are involved in the biosynthesis of testosterone and progesterone. We have performed enzyme kinetics analyses of the inhibition of EcaGST A3-3 by organotin compounds. We have also solved crystal structures of EcaGST A3-3 in complexes with glutathione, and wit...
Contreras-Aguilar MD, Tvarijonaviciute A, Monkeviciene I, Martín-Cuervo M, González-Arostegui LG, Franco-Martínez L, Cerón JJ, Tecles F....Measurement of adenosine deaminase (ADA) can provide information about cell-mediated immunity. This report's objective was to study the enzymatic activity of total ADA (tADA) and its isoenzymes ADA1 and ADA2 in canine, equine, porcine, and bovine serum and saliva and their changes in different inflammatory situations in each species. Besides, an automated method for ADA2 measurement was developed and validated. Results: tADA was present in serum and saliva of healthy animals of the four species. Erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) concentration of 0.47 mM was needed for ADA1 inhibit...
Journal of proteomicsOctober 8, 2020
Volume 231 104007 doi: 10.1016/j.jprot.2020.104007
Donkey's milk (DM) has been extensively investigated as a valuable substitute of breast milk, often suitable to manage cow's milk protein allergy in infants. DM exhibits potent inhibitory properties against numerous microbial species. Although oligosaccharides and lipids might contribute to the antimicrobial potential, the current inventory of proteins is not able to justify the low count of microorganisms generally observed in DM. The shotgun proteomic analysis of fractionated DM disclosed a set of 94 gene products, 41% of which have documented antimicrobial activity or are involved in transf...
de Roodt AR, Lanari LC, Lago NR, Bustillo S, Litwin S, Morón-Goñi F, Gould EG, van Grootheest JH, Dokmetjian JC, Dolab JA, Irazú L, Damin CF.Samples of Apis mellifera mellifera venom from different hives in two regions of the Buenos Aires province and its pool were analyzed for their lethal potency, myotoxic, defibrinogenating, hemolytic and inflammatory-edematizing activity and for the histological alterations they produce in the heart, lungs, kidneys, skeletal muscle and liver of mice. In vitro studies focused on the venom's hemolytic activity in different systems and species (horse, man, sheep and rabbit), the cytotoxicity in cellular lines, and on the proteolytic and coagulant activity in plasma and fibrinogen. Hemolytic activi...
Autry JM, Karim CB, Cocco M, Carlson SF, Thomas DD, Valberg SJ.We have analyzed protein expression and enzyme activity of the sarcoplasmic reticulum Ca2+-transporting ATPase (SERCA) in horse gluteal muscle. Horses exhibit a high incidence of recurrent exertional rhabdomyolysis, with myosolic Ca2+ proposed, but yet to be established, as the underlying cause. To better assess Ca2+ regulatory mechanisms, we developed an improved protocol for isolating sarcoplasmic reticulum (SR) vesicles from horse skeletal muscle, based on mechanical homogenization and optimized parameters for differential centrifugation. Immunoblotting identified the peak subcellular fract...
Shihadih DS, Harris TR, Kodani SD, Hwang SH, Lee KSS, Mavangira V, Hamamoto B, Guedes A, Hammock BD, Morisseau C.The veterinary pharmacopeia available to treat pain and inflammation is limited in number, target of action and efficacy. Inhibitors of soluble epoxide hydrolase (sEH) are a new class of anti-inflammatory, pro-resolving and analgesic drugs being tested in humans that have demonstrated efficacy in laboratory animals. They block the hydrolysis, and thus, increase endogenous concentrations of analgesic and anti-inflammatory signaling molecules called epoxy-fatty acids. Here, we screened a library of 2,300 inhibitors of the sEH human against partially purified feline, canine and equine hepatic sEH...
Contreras-Aguilar MD, Lamy E, Escribano D, Cerón JJ, Tecles F, Quiles AJ, Hevia ML.This study aims to evaluate the circadian and circannual variations in a panel of analytes in horse saliva that have been previously described as biomarkers related to stress and disease, in order to interpret them correctly when they are measured in this species. This panel of analytes integrated cortisol, salivary alpha-amylase (sAA), lipase (Lip), total esterase (TEA), butyrylcholinesterase (BChE), adenosine deaminase (ADA), γ-glutamyl transferase (gGT), creatine kinase (CK), urea, total bilirubin, total protein (TP), and phosphorus. These analytes were measured in saliva obtained from a p...
Löfgren M, Larsson P, Lindberg R, Hörnaeus K, Tydén E.Mild, moderate and severe equine asthma is a problem for equine welfare. The aetiology of the disease is not known in detail but is likely multi-factorial. One important factor may be inhaled dust which carries harmful substances which may be bioactivated and thus can lead to local inflammation in the airways. The aim of this study was to investigate gene expression and protein localisation of cytochrome P450 (CYP) enzymes, superoxide dismutase and glutathione-S-transferases (GST) involved in bioactivation and detoxification of harmful substances in lungs of horses with or without histological...
Knych HK, Finno CJ, Baden R, Arthur RM, McKemie DS.The in vivo metabolism and pharmacokinetics of flunixin meglumine and phenylbutazone have been extensively characterized; however, there are no published reports describing the in vitro metabolism, specifically the enzymes responsible for the biotransformation of these compounds in horses. Due to their widespread use and, therefore, increased potential for drug-drug interactions and widespread differences in drug disposition, this study aims to build on the limited current knowledge regarding P450-mediated metabolism in horses. Drugs were incubated with equine liver microsomes and a panel of r...
Hamamoto-Hardman BD, Baden RW, McKemie DS, Knych HK.Uridine diphospho-glucuronosyltransferases (UGTs) are membrane-bound enzymes that catalyze the conjugation of glucuronic acid onto a diverse set of xenobiotics. Horses efficiently and extensively glucuronidate a number of xenobiotics, including opioids, making UGTs an important group of drug-metabolizing enzymes for the clearance of drugs. Recombinant enzymes have allowed researchers to characterize the metabolism of a variety of drugs. The primary objective was to clone, express and characterize equine UGTs using drugs characterized as UGT substrates in other species. A secondary objective wa...
Bażanów BA, Chełmecka E, Romuk E, Stygar DM.Hucul and Arabian horses differ in the physiological constitution and exposition to environmental conditions. Oxidative stress plays a pathogenic role in many diseases and enables further injuries. The objective of this study was to compare the levels of enzymatic and nonenzymatic oxidative stress markers in Hucul horses living in seminatural conditions and in commercially handled Arabian horses. We tested the serum samples for total superoxide dismutase (total SOD), Cu-Zn-superoxide dismutase (CuZnSOD), and Mn-dependent superoxide dismutase (MnSOD) activity; for lipofuscin (LPS), ceruloplasmi...
França de Souza D, Alonso MA, Brito MM, Meirelles MG, Francischini MCP, Nichi M, Fernandes CB.In newborns, exposure to the extrauterine environment with high oxygen tension and sudden pulmonary adaptation leads to an increase in reactive oxygen species (ROS). ROS have several physiological roles, which are essential for neonatal development, however, when unbalanced, these highly unstable molecules can cause cellular destabilisation, compromising vital processes. Objective: To characterise the oxidative status in healthy equine neonates, evaluating an indicator of lipid peroxidation and both enzymatic and nonenzymatic antioxidant systems, during the first week of life. Methods: Experim...
Alamaary MS, Haron AW, Hiew MWH, Ali M.Present study aimed to investigate the effect of adding antioxidants, cysteine and ascorbic acid on the levels of glutamic oxaloacetic transaminase (GOT), glutamic-pyruvate (GPT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and γ-glutamyl transpeptidase (GGT) enzymes of post-thawed stallion sperm. Ten ejaculates were collected each from four healthy stallions and cryopreserved using HF-20 freezing extender containing either 0 mg/ml cysteine or ascorbic acid, 0.5 mg/ml cysteine and 0.5 mg/ml ascorbic acid. All samples in freezing extender containing cysteine or ascorbic acid or ...
Pasolini MP, Pezzella R, Santoro P, Cocchia N, Greco M, Del Prete C, Della Valle G, Auletta L.Equine exertional rhabdomyolysis (ER) is a well-recognized clinical syndrome affecting racehorses. Prevalence analysis of ER showed that female sex was a significant risk factor. The aim of this research was to evaluate the differences and correlations in the serum activity of muscle enzymes and the stage of the estrous cycle in ER-susceptible and control (C) mares. Serum muscle enzyme activity before and after exercise and sex hormones were analyzed in the two groups of mares. Ten cyclic ER and 10 cyclic C mares were examined weekly for 4 weeks. During diestrus, ER horses had significantly hi...
Aitken RJ, Gregoratos D, Kutzera L, Towney E, Lin M, Wilkins A, Gibb Z.MTT is widely used in biology as a probe for cell viability by virtue of its ability to generate deposits of insoluble formazan at sites of intense oxidoreductase activity. This response is generally held to reflect mitochondrial redox activity; however, extra-mitochondrial MTT reduction has also been recorded in certain cell types. Given this background, we set out to determine the major sites of formazan deposition in mammalian spermatozoa. In the mouse, most MTT reduction took place within the extensive mitochondrial gyres, with a single minor site of formazan deposition on the sperm head. ...
Gautier C, Scarlet D, Ertl R, Walter I, Wulf M, Nagel C, Aurich J, Aurich C.The aim of the present study was to characterise key enzymes involved in polyunsaturated fatty acid (PUFA) synthesis in the testis and epididymis collected from 2-year-old healthy warmblood stallions (n=10). The mRNA expression of fatty acid synthase, the Δ9-, Δ6-, Δ5- and Δ4-desaturases and elongases 6, 5 and 2 (encoded by the fatty acid synthase (FASN), the stearoyl-CoA desaturase (SCD), the fatty acid desaturase 2 (FADS2), the fatty acid desaturase 1 (FADS1), the delta 4-desaturase, sphingolipid 1 (DEGS1), ELOVL fatty acid elongase 6(ELOVL6), ELOVL fatty acid elongase 5 (ELOVL5), ELOVL ...
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...
Gadaj A, Ventura E, Healy J, Botrè F, Sterk SS, Buckley T, Mooney MH.Selective androgen receptor modulators (SARMs) represent non-steroidal agents commonly abused in human and animal (i.e. equine, canine) sports, with potential for further misuse as growth promoting agents in livestock-based farming. As a direct response to the real and possible implications of illicit application in both sport as well as food production systems, this study incorporated enzymatic hydrolysis (β-glucuronidase/arylsulfatase) into a previously established protocol while maintaining the minimal volume (200 µL) of urine sample required to detect SARMs encompassing various pharmacop...
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...
Amaral A, Fernandes C, Rebordão MR, Szóstek-Mioduchowska A, Lukasik K, Gawronska-Kozak B, Telo da Gama L, Skarzynski DJ, Ferreira-Dias G.Neutrophil extracellular traps (NETs) fight endometritis, and elastase (ELA), a protease found in NETs, might induce collagen type I (COL1) accumulation in equine endometrium. Metallopeptidases (MMPs) are involved in extracellular matrix balance. The aim was to evaluate the effects of ELA and sivelestat (selective elastase inhibitor) on MMP-2 and MMP-9 expression and gelatinolytic activity, as well as the potential inhibitory effect of sivelestat on ELA-induced COL1 in equine endometrium. Endometrial explants from follicular (FP) and mid-luteal (MLP) phases were treated for 24 or 48 h with ELA...
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...
Boerboom D, Brown KA, Vaillancourt D, Poitras P, Goff AK, Watanabe K, Doré M, Sirois J.Luteolysis in domestic species is mediated by the release of luteolytic pulses of prostaglandin (PG) F(2alpha) by the uterus at the end of diestrus, which must be suppressed by the conceptus to permit maternal recognition of pregnancy. In many species, including the horse, both the conceptus and the endometrium also synthesize PGE(2), which may antagonize PGF(2alpha) by playing a luteotropic and/or antiluteolytic role. While the release of PGE(2) and PGF(2alpha) by the equine endometrium in late diestrus and early pregnancy has been previously studied, the underlying prostaglandin synthase gen...
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...
Garvican ER, Vaughan-Thomas A, Redmond C, Gabriel N, Clegg PD.The plasma serine protease activated protein C (APC) is synthesized by human chondrocytes at sites of pathological cartilage fibrillation. APC levels are increased in osteoarthritis (OA) synovial fluid, and in vitro APC has been shown to synergize with interleukin-1beta (IL-1) to promote degradation from ovine cartilage. A model of equine cartilage degradation was established and used to explore corticosteroid activities. Intraarticular corticosteroids are a commonly prescribed treatment for joint disease, however their role in disease modification remains unclear. APC synergized with IL-1 or ...
Vinardell T, Dejica V, Poole AR, Mort JS, Richard H, Laverty S.The mechanisms leading to degeneration of articular cartilage in osteoarthritis (OA) are complex and not yet fully understood. Cathepsin K (CK) is a cysteine protease which can also cleave the triple helix of type II collagen. This exposes a neoepitope that can now be identified by specific antibodies. The aim of this study was to obtain evidence suggesting a role for CK in naturally occurring equine OA in both lesional and peri-lesional regions. Methods: Articular cartilages (n=12 horses; 5 healthy, 7 OA) were harvested from animals postmortem. A gross macroscopic examination, histologic (Saf...
Brophy PM, MacKintosh N, Morphew RM.Anthelmintics are the cornerstone of parasitic helminth control. Surprisingly, understanding of the biochemical pathways used by parasitic helminths to detoxify anthelmintics is fragmented, despite the increasing global threat of anthelmintic resistance within the ruminant and equine industries. Reductionist biochemistry has likely over-estimated the enzymatic role of glutathione transferases in anthelmintic metabolism and neglected the potential role of the cytochrome P-450 superfamily (CYPs). Proteomic technologies offers the opportunity to support genomics, reverse genetics and pharmacokine...
Raulo SM, Sorsa T, Tervahartiala T, Pirilä E, Maisi P.Gelatinolytic activity was analysed to study whether elevated activity previously found at the tracheal level of the respiratory tract of horses with chronic obstructive pulmonary disease (COPD) could also be found at the lower part of the respiratory tract. Furthermore, presence and significance of the gelatinolytic matrix metalloproteinases (MMPs) MMP-2 and MMP-9 in respiratory secretions of healthy and COPD horses were determined. Elevated gelatinolytic matrix metalloproteinases were detected in bronchoalveolar and tracheobronchial secretions from COPD horses. The main pathologically elevat...
