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Topic:Biochemistry
The study of biochemistry in horses encompasses the chemical processes and substances that occur within equine organisms. This field investigates the molecular interactions and pathways that are fundamental to horse physiology, including metabolism, enzyme activity, and genetic expression. Key areas of interest include the examination of metabolic disorders, nutrient absorption, and the biochemical basis of muscle function and energy production. Researchers utilize biochemical analysis to understand health and disease mechanisms in horses, contributing to the development of diagnostic tools and therapeutic strategies. This page gathers peer-reviewed studies and scholarly articles that explore various biochemical processes and their implications for equine health and performance.
Distribution of glycoconjugates in the uterine tube (oviduct) of horses. To examine glycoconjugates in the isthmic and ampullar regions of the uterine tube (oviduct) of horses during estrus, diestrus, and pregnancy. Methods: Oviductal samples from 17 mares. Methods: Oviducts were collected during estrus (n = 3), diestrus (n = 3), or pregnancy (n = 3), embedded, and snap frozen in liquid nitrogen. Frozen sections (5 to 6 microns in thickness) were stained with 100 micrograms/ml of fluorescein-isothiocyanate-conjugated lectin (30 min at 38.5 C) and were evaluated by use of epifluorescence microscopy and video image analysis. Specificity of lectins was established by ...
Pharmacology of the 5-lipoxygenase inhibitors BAY Y 1015 and BAY X 1005 in the horse. Calcium ionophore A23187 induced time and concentration dependent production of immunoreactive leukotriene (LT) B4 by equine heparinized whole blood in vitro. Time dependent production of immunoreactive LTB4 by equine neutrophils and immunoreactive LTC4 by equine eosinophils in vitro was also demonstrated. The 5-lipoxygenase activating protein (FLAP) inhibitors, BAY X 1005 and BAY Y 1015, produced concentration dependent inhibition of ionophore-induced LTB4 synthesis by equine whole blood (mean +/- SEM IC50s n = 5; 6.14 +/- 0.28 microM vs. 12.30 +/- 0.75 microM for BAY Y 1015 and BAY X 1005, r...
Muscarinic signaling pathway for calcium release and calcium-activated chloride current in smooth muscle. We investigated the muscarinic activation of Ca(2+)-activated Cl- currents [ICl(Ca)] in voltage-clamped equine tracheal myocytes. The threshold of cytosolic free Ca2+ concentration ([Ca2+]i) required for activation of ICl(Ca) was 202 +/- 22 nM, and full activation of the current occurred at 771 +/- 31 nM. Hexahydro-sila-difenidol (M3 antagonist) inhibited the methacholine-induced phasic [Ca2+]i increase and ICl(Ca) in a concentration-dependent manner, whereas methoctramine (M2 antagonist) only slightly attenuated the [Ca2+]i increase and ICl(Ca) (14.8 and 21.4%, respectively), consistent with ...
Micropreparative high resolution purification of proteins by a combination of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, isoelectric focusing, and membrane blotting. We report a simple, economical, and efficient protocol for protein purification from cells. First, proteins of cell lysates were separated by standard sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electroblotted to protein-blotting membrane. The blots were stained with Coomassie blue or developed by immunoblotting to visualize specific proteins. The bands corresponding to those visible by immunoblotting were excised from the dye-stained blots and subjected to isoelectric focusing. The focused gel was stained with Coomassie blue. Finally, the stained bands were excise...
Assessment of viability and mitochondrial function of equine spermatozoa using double staining and flow cytometry. An objective double-staining method was developed to evaluate viability and mitochondrial function of stallion spermatozoa using flow cytometry. Sperm viability was assessed by propidium iodide (PI) exclusion, and mitochondrial function was measured by the intensity of rhodamine 123 (R123) fluorescence. Flow cytometry estimates of sperm viability measured by PI were equivalent (P > 0.05) to estimates made using Hoechst 33258 stain and fluorescent microscopy (% dead: 25 +/- 2.4 vs 21.5 +/- 3.5). The use of both PI and R123 was validated by addition of various proportions of freeze-shocked (m...
The acute phase serum amyloid A protein (SAA) in the horse: isolation and characterization of three isoforms. Serum amyloid A (SAA) from acute phase horse serum was isolated using hydrophobic interaction chromatography, gel filtration and ion exchange chromatography. Three SAA isoforms with different isoelectric points, i.e. SAA pI 8.0, SAA pI 9.0 and SAA pI 9.7, were identified by two-dimensional electrophoresis and further characterized with amino acid sequence analysis. These isoforms were found in similar concentrations in all animals investigated, with SAA pI 9.7 constituting about half of the total SAA content. Partial amino acid sequence analysis verified the previously published heterogeneous ...
The effect of the acute-phase response on in vitro drug metabolism and plasma protein binding in the horse. The effect of the acute-phase response (APR) on the activity of the hepatic drug-metabolizing system (DMS) and on the binding of phenylbutazone to plasma proteins was investigated in the horse. An APR was induced by intramuscular injections of Freund's complete adjuvant in five horses and, five days later, these horses together with five clinically normal horses were shot and the right ventral lobe of each liver removed. The hepatic microsomal fractions from the liver samples were isolated and significantly lower (p < 0.01) concentrations of cytochromes P450 and b5 and activities of aniline...
A simplified strong ion model for acid-base equilibria: application to horse plasma. The Henderson-Hasselbalch equation and Stewart's strong ion model are currently used to describe mammalian acid-base equilibria. Anomalies exist when the Henderson-Hasselbalch equation is applied to plasma, whereas the strong ion model does not provide a practical method for determining the total plasma concentration of nonvolatile weak acids ([Atot]) and the effective dissociation constant for plasma weak acids (Ka). A simplified strong ion model, which was developed from the assumption that plasma ions act as strong ions, volatile buffer ions (HCO-3), or nonvolatile buffer ions, indicates th...
Expression and localization of relaxin in the ovary of the mare. Immunoreactive, chromatographic and molecular techniques were used to study the expression of relaxin in mare ovaries at different stages of the oestrous cycle. Relaxin in follicular fluid ranged from 1.6 to 2.5, from 1.4 to 5.2, from 1.2 to 6.7 and from 1.0 to 3.5 ng ml-1 in small ( 2 3 4 cm) follicles, respectively, and total content of fluid relaxin per follicle increased (P < 0.05) with follicular size. When subjected to reverse phase HPLC analysis, follicular fluid yielded absorbance profiles corresponding closely to those of purified relaxin, and immunoreactive peaks in follicular f...
Determination of methocarbamol in equine serum and urine by high-performance liquid chromatography with ultraviolet detection and atmospheric pressure ionization-mass spectrometric confirmation. Urine and serum samples collected from four standard-bred mares after and oral regimen administration of methocarbamol were extracted and analyzed. The method consisted of enzyme hydrolysis followed by a one-step liquid-liquid extraction, separation on a reversed-phase (RP-18) column, and detection using an ultraviolet (UV) detector. The confirmation was carried out using a liquid chromatography-atmospheric pressure ionization-mass spectrometry (LC-API-MS) system. Maximum methocarbamol concentrations of 1498, 1734, 1547, 2322 micrograms/mL in urine and 4.9, 1.7, and 3.6 micrograms/mL in serum ...
Tachykinin receptors in the equine pelvic flexure. Tachykinins, of which substance P (SP) is the prototype, are neuropeptides which are widely distributed in the nervous systems. In the equine gut, SP is present in enteric nerves and is a powerful constrictor of enteric muscle; in other species, SP is also known to have potent vasodilatory and pro-inflammatory effects. The specific effects of SP are determined by the subtype of receptor present in the target tissue. There are 3 known subtypes of tachykinin receptors, distinguished by their relative affinities for SP and other tachykinins. The distribution of SP binding sites in the equine pelv...
In vivo determination of surface tension in the horse trachea and in vitro model studies. We measured the surface tension in the trachea of the non-anaesthetised horse from the spreading behaviour of fluid drops, using videotracheoscopy. To do this, we placed small oil drops onto the tracheal wall with a thin Teflon tubing inserted into a videocolonoscope used in humans. Either 5 ml of saline (control) or 5 ml of bovine lipid extract surfactant (BLES) at 4 mg/ml were administered. Tracheal surface tension was 31.9 +/- 0.54 mN/m (Mean +/- SEM, n = 30) in the control experiments and 24.5 +/- 0.51 mN/m (Mean +/- SEM, n = 21) in the entire trachea after the administration of BLES. Thes...
Metabolic response in skeletal muscle fibres of standardbred trotters after racing. Histochemical and biochemical analyses were performed on muscle biopsies obtained after racing from the gluteus muscle of 18 standardbred trotters. Fibre type composition and enzyme activities varied among the horses. The percentage of type IIB fibres showed a positive correlation to the lactate dehydrogenase activity and a negative correlation to the citrate synthase activity. ATP concentrations in whole muscle after racing showed a negative correlation to both lactate and IMP concentrations. Within individual fibres, ATP concentrations varied markedly, with some type II fibres having values ...
The ICP0 protein of equine herpesvirus 1 is an early protein that independently transactivates expression of all classes of viral promoters. To assess the role of the equine herpesvirus type 1 (EHV-1) ICP0 protein (EICP0) in gene regulation, a variety of molecular studies on the EICP0 gene and gene products of both the attenuated cell culture-adapted Kentucky A (KyA) strain and the Ab4p strain were conducted. These investigations revealed that (i) the ICP0 open reading frame (ORF) of the KyA virus strain is 1,257 bp in size and would encode a protein of 419 amino acids, and in comparison to the ICP0 gene (ORF63) of the Ab4p strain of 1,596 bp (E. A. Telford, M. S. Watson, K. McBride, and A. J. Davison, Virology 189:304-316, 1992), ...
Cellular heterogeneity in cathepsin D distribution in equine articular cartilage. The distribution of cathepsin D in normal equine growth cartilage has been examined immunocytochemically using an antiserum raised against human cathepsin D. The cross-reactivity and specificity of the antiserum for equine cathepsin D was confirmed, and its lysosomal localisation was demonstrated in horse skin fibroblasts by confocal scanning microscopy. Cultured horse chondrocytes were heterogenous in their expression of cathepsin D. Heterogeneity of distribution of the enzyme was also seen in chondrocytes in cartilage from different anatomical sites. A high level of cathepsin D was observed ...
Redox regulation of large conductance Ca(2+)-activated K+ channels in smooth muscle cells. The effects of sulfhydryl reduction/oxidation on the gating of large-conductance, Ca(2+)-activated K+ (maxi-K) channels were examined in excised patches from tracheal myocytes. Channel activity was modified by sulfhydryl redox agents applied to the cytosolic surface, but not the extracellular surface, of membrane patches. Sulfhydryl reducing agents dithiothreitol, beta-mercaptoethanol, and GSH augmented, whereas sulfhydryl oxidizing agents diamide, thimerosal, and 2,2'-dithiodipyridine inhibited, channel activity in a concentration-dependent manner. Channel stimulation by reduction and inhibit...
Protonophoric activity of NADH coenzyme Q reductase and ATP synthase in coupled submitochondrial particles from horse platelets. A method to prepare coupled submitochondrial particles from horse platelets is described. The method allowed us to study the protonophoric activities of both complex I and complex V following the fluorescence quenching of the monoamine 9-amino-6-chloro-2 methoxyacridine (ACMA), a probe highly sensitive to the generation of a transmembrane delta pH. We carried out a kinetic analysis of each enzyme complex studying the proton translocation and the electron transfer activities of complex I as well as the proton translocation and the ATP hydrolytic activities of complex V. A micromethod to prepare...
Characterization of prolactin- and growth hormone-binding proteins in milk and their diversity among species. The present study was undertaken to identify and characterize the diversity and species distribution of soluble prolactin binding-protein (PRL-BP) and growth hormone-binding protein (PRL-BP) in mammalian milk. We previously divided mammalian serum GH-BP into four main groups and identified a GH-BP with shared lactogenic/somatogenic properties in rabbit, horse, dog, pig and cat (Type III species). Here we describe PRL-BP in milk of Type III species and show it is relatively conserved within the group, having similar characteristics in terms of binding affinity for hGH (0.74-5.5 x 10(10) M(-1)),...
Structure of glycan moieties responsible for the extended circulatory life time of fetal bovine serum acetylcholinesterase and equine serum butyrylcholinesterase. Cholinesterases are serine hydrolases that can potentially be used as pretreatment drugs for organophosphate toxicity, as drugs to alleviate succinylcholine-induced apnea, and as detoxification agents for environmental toxins such as heroin and cocaine. The successful application of serum-derived cholinesterases as bioscavengers stems from their relatively long residence time in the circulation. To better understand the relationship between carbohydrate structure and the stability of cholinesterases in circulation, we determined the monosaccharide composition, the distribution of various oligo...
Opsonic effect of equine plasma from different donors. The ability of equine plasma from different donors to enhance phagocytic capacity was assessed in neutrophils obtained from seven foals, aged 7-8 days (Study A), and from seven adult horses (Study B). Neutrophils were allowed to phagocytize fluorescent yeast cells opsonized with plasma from one of three donors or with pooled serum, all previously frozen (-18 degrees C) and thawed. The results were analysed by flow cytometry. In study A, fresh autologous foal serum was also used for opsonization, and in study B, heat-inactivated plasma and pooled serum were used in addition to untreated samples...
Prejunctional alpha 2-adrenoceptors inhibit nitrergic neurotransmission in horse penile resistance arteries. To study the influence of alpha-adrenergic stimuli on non-adrenergic non-cholinergic (NANC) neurogenic relaxation in isolated horse penile resistance arteries. Methods: Deep intracavernous penile arteries with an internal lumen diameter of 200-500 microns., isolated from the corpus cavernosum of young horses, were mounted in microvascular myographs for isometric tension recording and electrical field stimulation (EFS) of autonomic nerve terminals. Results: In the presence of guanethidine (10(-5) M) and atropine (10(-7) M) tone of the arteries was raised by the thromboxane analogue, U46619. EFS...
Biosynthesis and possible biological roles of progestagens during equine pregnancy and in the newborn foal. Major progress on the endocrinology of the pregnant mare has been possible thanks to the catheterised equine fetal preparation developed by Marian Silver. In particular, these preparations led to the identification of the source of progestagens within the feto-placental unit and provided the impetus for further work on their biosynthesis and biological activities. The biosynthesis of the progestagens involves close interaction between the fetus, the endometrium and the placenta, and gives rise to some fundamental biochemical questions. The biological role of the progestagens is also discussed:...
The pathogenic effects of experimental cyathostome infections in ponies. Nine pony breed foals were reared indoors, then allocated to one of three groups infected with either 3.9 million (Group A) or 3.15 million (Group B) cold-conditioned third stage cyathostome larvae or kept as uninfected controls (Group C). The larvae were administered as a 'trickle' infection of 150000 larvae per dose, three times weekly. Blood biochemical and haematological analyses were performed weekly and faecal worm egg counts bi-weekly. Complete parasitological examinations were performed on all ponies at various times post-initial infection (PI): one infected animal at 9 weeks PI, four ...
Interaction of horse heart cytochrome c with lipid bilayer membranes: effects on redox potentials. Cyclic voltammetry has been used to study the effects of interactions between horse cytochrome c and solid-supported planar lipid membranes, comprised of either egg phosphatidylcholine (PC) or PC plus 20 mol.% cardiolipin (CL), on the redox potential and the electrochemical electron transfer rate between the protein and a semiconductor electrode. Experiments were performed over a wide range of cytochrome c concentrations (0-440 microM) at low (20 mM) and medium (160 mM) ionic strengths. Three types of electrochemical behavior were observed, which varied as a function of the experimental condit...
Characterization of muscarinic receptors in equine tracheal smooth muscle in vitro. This study was undertaken to assess the importance of muscarinic receptor subtypes in equine airway disease. Smooth muscle strips from the mid-cervical portion of the trachea of horses were placed in tissue baths and isometric contractile force was measured. Active force was measured in response to metacholine and the selective muscarinic receptor agonists McN-A-343 (M1-selective) and pilocarpine (M2-selective) in cumulative concentrations (10(-9)M through 10(-3)M), with and without preincubation with three or four concentrations of the selective muscarinic receptor antagonists pirenzepine (M1...
Pharmacological characterization of adrenoceptors in horse corpus cavernosum penis. 1. The presence and types of alpha and beta-adrenoceptors in the corpus cavernosum of the horse were studied in vitro by using selected ligands of adrenoceptors and isometric tension recording. 2. Noradrenaline and phenylephrine induced concentration-dependent contractions in corpus cavernosum preparations. B-HT 920 had no effect. 3. Phentolamine and prazosin produced a shift to the right of the dose-response curve of noradrenaline, while the alpha(2)-antagonist, rauwolscine had no effect on the response to noradrenaline. Phenylephrine-evoked contractions of corporal strips were significantly ...
Cloning and analysis of a Borrelia burgdorferi membrane-interactive protein exhibiting haemolytic activity. We cloned the gene encoding a membrane-interactive protein of Borrelia burgdorferi by means of its haemolytic activity in Escherichia coli. The haemolytic activity was erythrocyte-species specific, with progressively decreasing activity for erythrocytes from horse, sheep, and rabbit, respectively. Genetic analysis of the haemolytic determinant revealed two borrelia haemolysin genes, blyA and blyB, that are part of a predicted four-gene operon which is present in multiple copies on the 30 kb circular plasmid(s) of B. burgdorferi B31. blyA encodes a predicted alpha-helical 7.4 kDa protein with a...
Intramuscular bioavailability of ketoprofen lysine salt in horses. Lysine salts are often used in human pharmaceuticals to increase the solubility and absorption of acidic drugs when these are administered parenterally. In this study the intramuscular bioavailability of ketoprofen administered as the lysine salt was evaluated in horses (n = 5) treated intravenously and intramuscularly (2.2 mg/kg active substance) in a cross-over study. The absorption rate of ketoprofen administered as the lysine salt was rather low: the mean residence time increased from 31.7 min after IV injection to 128.9 min (after IM injection), and the bioavailability was high (mean 92.4...
Influence of type and breed of horse on serum osteocalcin concentration, and evaluation of the applicability of a bovine radioimmunoassay and a human immunoradiometric assay [corrected]. To evaluate applicability of a human osteocalcin (OC) immunoradiometric assay (IRMA) for use with equine serum and compare it with a bovine radioimmunoassay (RIA) previously proven valid for such samples, and to describe the effect of type and breed of horses on serum OC concentration. Methods: 100 healthy horses of either sex, classified as type I or II (draught or warmblood, respectively). Each type was represented by 2 breed groups, each comprising 25 horses. Methods: Blood samples were collected in the morning, and the serum was separated. Osteocalcin was measured, using commercially avail...
