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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.
Molecular cloning of equine 17beta-hydroxysteroid dehydrogenase type 1 and its downregulation during follicular luteinization in vivo. The type 1 form of 17beta-hydroxysteroid dehydrogenase (17betaHSD1) was the first isoform to be identified and is capable of converting estrone to 17beta-estradiol. This study was aimed at characterizing the molecular structure of the equine 17betaHSD1 gene and cDNA, as well as its molecular regulation during human chorionic gonadotropin (hCG)-induced follicular luteinization/ovulation in vivo. The equine 17betaHSD1 gene was cloned from an equine genomic library and shown to have a conserved genomic structure composed of six exons. Its cDNA sequence was also identified and coded for a 308 amin...
Detecting early kidney damage in horses with colic by measuring matrix metalloproteinase -9 and -2, other enzymes, urinary glucose and total proteins. The aim of the study was to investigate urine matrix metalloproteinase (MMP-2 and -9) activity, alkaline phosphatase/creatinine (U-AP/Cr) and gamma-glutamyl-transpeptidase/creatinine (U-GGT/Cr) ratios, glucose concentration, and urine protein/creatinine (U-Prot/Cr) ratio and to compare data with plasma MMP-2 and -9 activity, cystatin-C and creatinine concentrations in colic horses and healthy controls. Horses with surgical colic (n = 5) were compared to healthy stallions (n = 7) that came for castration. Blood and urine samples were collected. MMP gelatinolytic activity was measured by zymogra...
Influence of general anaesthesia on the pharmacokinetics of intravenous fentanyl and its primary metabolite in horses. In order to evaluate its potential as an adjunct to inhalant anaesthesia in horses, the pharmacokinetics of fentanyl must first be determined. Objective: To describe the pharmacokinetics of fentanyl and its metabolite, N-[1-(2-phenethyl-4-piperidinyl)maloanilinic acid (PMA), after i.v. administration of a single dose to horses that were awake in Treatment 1 and anaesthetised with isoflurane in Treatment 2. Methods: A balanced crossover design was used (n = 4/group). During Treatment 1, horses received a single dose of fentanyl (4 microg/kg bwt, i.v.) and during Treatment 2, they were anaesthet...
Laminar xanthine oxidase, superoxide dismutase and catalase activities in the prodromal stage of black-walnut induced equine laminitis. REASONS FOR STUDY: Xanthine oxidase (XO)-dependent production of superoxide anion and hydrogen peroxide, a characteristic of ischaemia-reperfusion injury, may contribute to the development of equine laminitis. Objective: To determine the levels of XO and antioxidant enzymes (catalase, superoxide dismutase [SOD]) in the digital laminae of normal horses (CON) and horses in the developmental stage of laminitis using the black walnut extract (BWE) model. Methods: Healthy horses (n = 12) were administered BWE (BWE group, n = 6), or water (CON group, n = 6) through a nasogastric tube. At the onset o...
Determination of lidocaine and its two N-desethylated metabolites in dog and horse plasma by high-performance liquid chromatography combined with electrospray ionization tandem mass spectrometry. A sensitive method for the quantification of lidocaine and its metabolites, monoethylglycinexylidide (MEGX) and glycinexylidide (GX), in animal plasma using high-performance liquid chromatography combined with electrospray ionization mass spectrometry is described. The sample preparation includes a liquid-liquid extraction with methyl tert-butylmethyl ether after addition of 2M sodium hydroxide. Ethylmethylglycinexylidide (EMGX) is used as an internal standard. For chromatographic separation, an ODS Hypersil column was used. Isocratic elution was achieved with 0.01 M ammonium acetate and aceto...
Stereoselective pharmacokinetics of ketamine and norketamine after racemic ketamine or S-ketamine administration during isoflurane anaesthesia in Shetland ponies. The arterial pharmacokinetics of ketamine and norketamine enantiomers after racemic ketamine or S-ketamine i.v. administration were evaluated in seven gelding ponies in a crossover study (2-month interval). Methods: Anaesthesia was induced with isoflurane in oxygen via a face-mask and then maintained at each pony's individual MAC. Racemic ketamine (2.2 mg kg(-1)) or S-ketamine (1.1 mg kg(-1)) was injected in the right jugular vein. Blood samples were collected from the right carotid artery before and at 1, 2, 4, 8, 16, 32, 64, and 128 min after ketamine administration. Ketamine and norketamine...
Urinary excretion of 5(10)-estrene-3beta,17alpha-diol and estrone by the female horse: complementary indicators of early pregnancy screened with regard to a putative anabolic doping practice. Rules of horse racing stipulate that pregnant mares may compete under definite conditions of date, because early pregnant status may be misused for the sake of enhancing physical performance by putative anabolic steroid action. Screening for pregnancy is generally performed by plasma equine gonadotrophin (eCG) immunoassay, which covers the period between Days 40 and 120. In common screening for urinary anabolic steroids performed by gas chromatography-mass spectrometry, inclusion of two complementary criteria, i.e. the evaluation of total conjugates of 5(10)-estrene-3beta,17alpha-diol (EED) an...
Comparison of plasma biochemical parameters in Thoroughbred and Purebred Arabian horses during the same-intensity exercise. The objective of the present study was to compare changes of blood parameters induced by the same work and performed in the same conditions in two racehorse breeds, Thoroughbred and Purebred Arabian. The effect of moderate-intensity exercise was studied in 20 stallions--ten Thoroughbreds, aged 2-3 years and the same number of Purebred Arabians, 3-4 years old. All the horses were administrated the same effort test consisting in 1200 m gallop at a speed typical of the daily training sessions. Three jugular venous blood samples were collected for each horse: at rest, just after the end of the gal...
Quantitative determination of the macrolide antibiotic tulathromycin in plasma and broncho-alveolar cells of foals using tandem mass spectrometry. The long-acting antibiotic tulathromycin is on the marked for treatment of pulmonary infection of cattle, swine and horses. To measure disposition and distribution of tulathromycin in foals, a high throughput method was developed for horse plasma (calibration range: 0.006-0.8 microg/mL) and broncho-alveolar cells (calibration range: 0.1-4.0 microg/10(9)cells) using tandem mass spectrometry. Tulathromycin was extracted from plasma and broncho-alveolar fluid using cation exchange cartridges with acetonitrile/ammonia (95:5, v/v). The chromatography was performed isocratically with a mobile phase ...
Neutrophil myeloperoxidase measurements in plasma, laminar tissue, and skin of horses given black walnut extract. To compare measurements of myeloperoxidase (MPO) in plasma, laminar tissues, and skin obtained from control horses and horses given black walnut heartwood extract (BWHE). Methods: 22 healthy 5- to 15-year-old horses. Methods: Horses were randomly assigned to 4 groups as follows: a control group given water (n = 5) and 3 experimental groups given BWHE (17) via nasogastric intubation. Experimental groups consisted of 5, 6, and 6 horses that received BWHE and were euthanatized at 1.5, 3, and 12 hours after intubation, respectively. Control horses were euthanatized at 12 hours after intubation. Pl...
In vitro cultivation of Plasmodium falciparum: studies with modified medium supplemented with ALBUMAX II and various animal sera. RPNI, a combination of three commercially available growth media (RPMI-1640, NCTC-135 and IMDM) has been found to support long term continuous cultivation of 3D7 strain of Plasmodium falciparum in the presence of 10% bovine calf serum. During the present study, the suitability of this medium was evaluated for the development of P. falciparum in the presence of horse, goat and rabbit sera as well as various concentrations of ALBUMAX II. RPNI medium supplemented with 10% bovine calf serum (RPNI-BCS) was used as control. The cultures were maintained in candle jars protocol and parasitaemia was mo...
In vitro effects of oxygen on physico-chemical properties of horse erythrocyte membrane. Whether direct exposure to different concentrations (0%, 13%, 100%) of oxygen may affect horse erythrocyte membrane fluidity (EMF) and fatty acid (FA) composition was studied during 1 (T60) and 2h (T120) exposure. EMF was investigated at the head group level and hydrophobic core thanks to phosphorus nucleus 31 ((31)P) nuclear magnetic resonance ((31)P NMR) and electronic paramagnetic resonance (EPR) using two spin probes: 5-nitroxydestearic acid and 16-doxylstearic acid. Lipid structure of the membranes was studied by gas liquid chromatography. 4-Hydroxy-2E-nonenal was also analyzed as a marke...
Vehicle effects on the in vitro penetration of testosterone through equine skin. The effects of three vehicles, phosphate-buffered saline (PBS), ethanol (50% in PBS w/w) and propylene glycol (50% in PBS w/w) on in vitro transdermal penetration of testosterone was investigated in the horse. Skin was harvested from the thorax of five Thoroughbred horses after euthanasia and stored at -20 degrees C until required. The skin was then defrosted and placed into Franz-type diffusion cells, which were maintained at approximately 32 degrees C by a water bath. Saturated solutions of testosterone, containing trace amounts of radiolabelled [14C]testosterone, in each vehicle were applie...
Inhibitor-free DNA for real-time PCR analysis of synovial fluid from horses, cattle and pigs. The potential of five different commercial DNA isolation methods to remove real-time PCR inhibitors from the synovial fluid of horses, cattle and pigs was investigated. All kits with the exception of one included a silica column-based purification of the DNA. With the fifth kit, DNA purification is achieved by removing contaminating macromolecules by a desalting process. We used a recently developed method based on comparison of the real-time PCR signal of an artificial target incorporated into each PCR reaction in the presence of the isolated DNA from the sample, and in control samples contai...
Variability of plasma melatonin level in pony mares (Equus caballus), comparison with the hybrid: mules and with jennies (Equus asinus). In long-day breeders like horses, the length of nocturnal melatonin secretion is the main messenger of photoperiod. Previous studies have shown that the nocturnal jugular melatonin concentration is lower in horses, than in mules but is unknown in donkeys. The aim of this study was to estimate the inter-animal variability of plasma melatonin concentration in domestic mares and to compare this concentration with those observed in domestic jennies and in their hybrid mules. In the autumn, blood samples were collected at 22 h, 23 h, 0 h and 1 h during 2 nights at 3 weeks intervals, in 110 pony mar...
Segment-dependent expression of muscarinic acetylcholine receptors and G-protein coupling in the equine respiratory tract. Muscarinic receptors are considered to be of comparable clinical importance in chronic obstructive pulmonary disease (COPD) in equines and in humans. At present, data are scarce on the expression and distribution of probable subtypes of these receptors and their signalling pathways in airway segments, including lung parenchyma and bronchial and tracheal epithelium with the underlying smooth muscle in horses. Specific [N-methyl-3H]scopolamine chloride ([3H]NMS) binding to all three tissues was saturable and of high affinity, with KD values ranging between 1.6+/-0.7 and 1.9+/-0.3 nmol/L. [3H]NMS...
Novel findings regarding Glut-4 expression in adipose tissue and muscle in horses–a preliminary report. One of the hallmarks of insulin resistance is a reduction in glucose transporter-4 (Glut-4) expression in adipose tissue but not in skeletal muscle. However, while Glut-4 has been demonstrated in skeletal and cardiac muscles in horses it has not been demonstrated in adipose tissue. The initial objectives of the present study were: (1) to test the hypothesis that Glut-4 expression would vary between selected key skeletal muscles; (2) to test the hypothesis that it would also vary between representative adipose tissue depots, and (3) to see whether expression would be greater in adipose tissue c...
Reliability of 1,9-dimethylmethylene blue tests in comparison to agarose gel electrophoresis for quantification of urinary glycosaminoglycans. The relevance of glycosaminoglycan determination in biological fluids is gradually gaining importance in the literature. Nevertheless, the results obtained by different methods vary widely. We evaluated 1,9-dimethylmethylene blue (DMB) dye-binding assays for quantification of urinary glycosaminoglycans, in comparison to densitometry after agarose gel electrophoresis. Methods: Urinary glycosaminoglycans from different mammalian species were quantified by 3 different DMB dye-binding assays. The results were compared to those obtained by densitometry after agarose gel electrophoresis of glycosami...
Defining cytochemical markers for different cell types in the equine retina. The major cell types in the mammalian retina are photoreceptors, amacrine, horizontal, bipolar, ganglion and Mueller glial cells. Most of the specific cell types are conserved, but cytochemical markers vary between species. The aim of our study was to characterize cytochemically distinctive markers for different cell types in the equine retina. We were able to define specific markers for equine Mueller glial cells and photoreceptor cells. Furthermore, we describe markers for large ganglion cells, horizontal and amacrine cells and a subpopulation of bipolar cells. Additionally, discrimination b...
Effects of sodium hyaluronate and methylprednisolone acetate on proteoglycan metabolism in equine articular chondrocytes treated with interleukin-1. To determine the effects of sodium hyaluronate (HA) in combination with methylprednisolone acetate (MPA) on interleukin-1 (IL-1)-induced inflammation in equine articular cartilage pellets. Methods: Chondrocytes collected from 7 horses euthanatized for problems unrelated to the musculoskeletal system. Methods: Chondrocyte pellets were treated with medium (negative control); medium containing IL-1 (positive control); or medium containing IL-1 with MPA only (0.05 or 0.5 mg/mL), HA only (0.2 or 2 mg/mL), or MPA (0.05 or 0.5 mg/mL) and HA (0.2 or 2 mg/mL) in combination. Proteoglycan (PG) synthesis...
Studies related to the origin of C18 neutral steroids isolated from extracts of urine from the male horse: the identification of urinary 19-oic acids and their decarboxylation to produce estr-4-en-17beta-ol-3-one (19-nortestosterone) and estr-4-ene-3,17-dione (19-norandrost-4-ene-3,17-dione) during sample processing. For almost two decades we have known that enzymatic hydrolysis of "normal" urine samples from the entire male horse using Escherichia coli (E. coli) followed by solvolysis (ethyl acetate:methanol:sulphuric acid) results in the detection of significant amounts of estr-4-ene-3,17-dione (19-norandrost-4-ene-3,17-dione) along with estr-4-en-17beta-ol-3-one (19-nortestosterone, nandrolone) in extracts of the hydrolysed urine and that both steroids are isolated from the solvolysis fraction. This solvolysis process is targeted at the steroid sulphates. Also we have shown that 19-norandrost-4-ene-3,17...
Proline scanning mutagenesis reveals non-native fold in the molten globule state of equine beta-lactoglobulin. The secondary structure in the molten globule state (an equilibrium analogue of a burst-phase folding intermediate) of equine beta-lactoglobulin was investigated by changes in the circular dichroic spectrum induced by a series of site-directed proline substitutions. The results challenge the structural picture obtained from previous hydrogen/deuterium exchange experiments. A stable non-native alpha-helix was found to exist in the region corresponding to the eighth strand (H strand) in the native structure, where the backbone amide protons are the most strongly protected from exchange. Therefor...
Factors influencing pre-race serum concentration of total carbon dioxide in Thoroughbred horses racing in California. Many racing jurisdictions monitor pre-race serum concentration of total carbon dioxide (TCO2) among racing horses. To our knowledge, factors influencing concentration of TCO2 among horses participating in racing have not been systematically evaluated and reported. Objective: To determine if characteristics of horses and racing conditions routinely recorded were significantly associated with pre-race concentration of TCO2, while accounting for and estimating effects of trainer and horse. Methods: Pre-race serum TCO2 concentrations from 5028 starts made by 2,349 horses trained by 287 trainers at...
Cartilage oligomeric matrix protein and hyaluronan levels in synovial fluid from horses with osteoarthritis of the tarsometatarsal joint compared to a control population. Quantification of cartilage oligomeric matrix protein (COMP) levels within synovial fluid from the tarsometatarsal joint has not previously been reported and an effective synovial fluid marker would allow monitoring of disease progression and treatment. Objective: To quantify levels of COMP and hyaluronan (HA) in synovial fluid from the tarsometatarsal joint, identify differences in levels from horses with osteoarthritis (OA) of the tarsometatarsal joint compared to a control population and to correlate levels with radiographic changes in horses with OA. Methods: Synovial fluid was collected f...
In vitro effects of fungi isolated from equine hooves on primary human keratinocytes. The effects of two dermatophytes (Microsporum gypseum and Trichophyton mentagrophytes) and four moulds (Scopulariopsis brevicaulis, Alternaria alternata, Geotrichum candidum and Penicillium spp.) on living keratinocyte cultures were examined in vitro using primary human keratinocytes. Rates of apoptosis of infected cells were determined using a colorimetric TUNEL system which detects the characteristic nuclear DNA fragmentation of apoptotic cells. The cytotoxicity of the individual fungi was tested by quantitatively measuring cytosolic enzyme lactate dehydrogenase, released upon cell lysis, in...
Effects of exercise and oral antioxidant supplementation enriched in (n-3) fatty acids on blood oxidant markers and erythrocyte membrane fluidity in horses. The aim of this study was to investigate in a placebo-controlled field study the effect of a (n-3)-vitamin supplementation on erythrocyte membrane fluidity (EMF), oxidant/antioxidant markers and plasmatic omega3/omega6 fatty acid ratio (FAR) in 12 eventing horses. Venous blood was sampled at rest before (PRE) and after (POST) a three week treatment period with either the supplement (group S, n=6) or a placebo (group P, n=6) as well as after 15min (POST E15') and 24h (POST E24h) after a standardised exercise test. The following markers were analysed: EMF, plasma antioxidant capacity of water an...
Heterophile antibody interference in a solid phase sandwich immunoassay for detection of equine growth hormone in plasma. Heterophile antibodies (HAs) present in serum recognize animal immunoglobulins and are one of the most unpredictable causes of false results in immunoassays. However, no study has yet reported their interference on the diagnostic reliability of immunochemical analyses on horse plasma. Recently, we developed a sandwich ELISA for detection of equine growth hormone (eGH) in plasma. In a pilot study to measure basal eGH levels (blood samples were drawn from 13 horses every 10 min for 1h), we noted one horse with abnormally high eGH (>100 ng/mL). We demonstrate here that this plasma eGH level wa...
Proteolytic maturation of replicase polyprotein pp1a by the nsp4 main proteinase is essential for equine arteritis virus replication and includes internal cleavage of nsp7. The positive-stranded RNA genome of the arterivirus Equine arteritis virus (order Nidovirales) encodes the partially overlapping replicase polyproteins pp1a (1727 aa) and pp1ab (3175 aa). Previously, three viral proteinases were reported to cleave these large polyproteins into 12 non-structural proteins (nsps). The chymotrypsin-like viral main proteinase residing in nsp4 is responsible for eight of these cleavages. Processing of the C-terminal half of pp1a (the nsp3-8 region) was postulated to occur following either of two alternative proteolytic pathways (the 'major' and 'minor' pathways). He...
Direct-injection screening for acidic drugs in plasma and neutral drugs in equine urine by differential-gradient LC-LC coupled MS/MS. Direct-injection LC-LC hybrid tandem MS methods have been developed for undertaking broad-based screening for acidic drugs in protein-precipitated plasma and neutral doping agents in equine urine. In both analyses, analytes present in the matrix were trapped using a HLB extraction column before being refocused and separated on a Chromolith RP-18e monolithic analytical column using a controlled differential gradient generated by proportional dilution of the first column's eluent with water. Each method has been optimised by the adoption of a mobile phase and gradient that was tailored to enhanc...
