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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.
Red blood cell deformability and aggregation behaviour in different animal species. Comparative animal studies showed the wide variation of whole blood and plasma viscosity, and erythrocyte aggregation among mammalian species. Whole blood viscosity and red blood cell aggregation is influenced by red cell fluidity. To evaluate differences in erythrocyte deformability in mammals, three species were investigated, whose erythrocytes have a different aggregation property: horse, as a species with high, dog with medium, and sheep with almost unmeasurable aggregation tendency. Erythrocyte deformability was tested ektacytometrically (Elongation Index [EI], LORCA, Mechatronics, Hoorn,...
Aquaporin water channels AQP1 and AQP3, are expressed in equine articular chondrocytes. Chondrocytes exist in an unusual and highly variable ionic and osmotic environment in the extracellular matrix of articular cartilage. Alterations to the ionic and osmotic environment of chondrocytes influence the volume and ionic content of the cells, which, in turn, modifies the rate at which extracellular matrix macromolecules are synthesized and degraded. Thus, regulation of the water and solute content of chondrocytes will profoundly affect their anabolic and catabolic functions. The water content of cells is effectively influenced by the abundance of aquaporin (AQP) water channels. Recen...
Evaluation of alternative cryoprotectants for preserving stallion spermatozoa. Although use of cryopreserved stallion spermatozoa is currently accepted by many breed registries, utilization of this technique remains limited due to poor fertility for some stallions. One reason for these results is osmotic stress that spermatozoa experiences when the cryoprotectant (glycerol) is added to the cells prior to freezing and removal from the cells after thawing. In an effort to minimize osmotic damage, alternative cryoprotectants, having lower molecular weights and greater membrane permeability than glycerol, were evaluated to determine their effectiveness for cryopreserving sta...
Application of the comet assay for investigation of oxidative DNA damage in equine peripheral blood mononuclear cells. Oxidative stress occurs when antioxidant defense mechanisms are overwhelmed by free radicals and may lead to DNA damage, which has been implicated in processes such as aging and diseases such as cancer. The two main techniques presently used to quantify DNA damage are measurement of 8-hydroxydeoxyguanosine and the Comet assay (also known as single-cell gel electrophoresis). The aim of this study was to apply the comet assay to equine peripheral blood mononuclear cells (PBMCs) and identify two conditions in which we hypothesized that oxidative DNA damage would be increased in PBMCs: aging and e...
Effects of feeding meals with various soluble-carbohydrate content on muscle glycogen synthesis after exercise in horses. To determine effects of feeding diets with various soluble-carbohydrate (CHO) content on rates of muscle glycogen synthesis after exercise in horses. Methods: 7 fit horses. Methods: In a 3-way crossover study, horses received each of 3 isocaloric diets (a high soluble CHO [HC] diet, a low soluble CHO [LC] diet, or a mixed soluble CHO [MC] diet). For each diet, horses were subjected to glycogen-depleting exercise, followed by feeding of the HC, LC, or MC diet at 8-hour intervals for 72 hours. Results: Feeding the HC diet resulted in a significantly higher glycemic response for 72 hours and sign...
Electrospray ionization mass spectrometric characterization and quantitation of xanthine derivatives using isotopically labelled analogues: an application for equine doping control analysis. Isotope-dilution mass spectrometry has been employed successfully in numerous fields of analytical chemistry enabling the establishment of fast and reliable procedures. In equine sports, xanthine derivatives such as caffeine and theobromine are prohibited, and doping control laboratories analyze horse urine specimens regarding these illicit performance-enhancing drugs. Theobromine has to exceed a threshold level of 2 microg/mL, hence a robust and reliable quantitation is required. Stably deuterated theobromine and caffeine were synthesized by the reaction of xanthine or theobromine with iodome...
Investigation of the expression and localization of glucose transporter 4 and fatty acid translocase/CD36 in equine skeletal muscle. To investigate the expression and localization of glucose transporter 4 (GLUT4) and fatty acid translocase (FAT/CD36) in equine skeletal muscle. Methods: Muscle biopsy specimens obtained from 5 healthy Dutch Warmblood horses. Methods: Percutaneous biopsy specimens were obtained from the vastus lateralis, pectoralis descendens, and triceps brachii muscles. Cryosections were stained with combinations of GLUT4 and myosin heavy chain (MHC) specific antibodies or FAT/CD36 and MHC antibodies to assess the fiber specific expression of GLUT4 and FAT/CD36 in equine skeletal muscle via indirect immunofl...
Expression of the full-length and alternatively spliced equine luteinizing hormone/chorionic gonadotropin receptor mRNAs in the primary corpus luteum and fetal gonads during pregnancy. The full-length equine luteinizing hormone/chorionic gonadotropin (LH/CG) receptor (eLH/CG-RA) cDNA and two alternatively spliced isoforms (eLH/CG-RB,C) were isolated from luteal tissue and characterized using a combination of reverse transcription-polymerase chain reaction (RT-PCR) and 5'-rapid amplification of cDNA ends. The 680-amino acid full sequence of eLH/CG-RA displayed 87-92% homology with other mammalian LH/CG-Rs. The eLH/CG-RB and eLH/CG-RC cDNA isoforms were truncated from the 3'-end of exon X: eLH/CG-RB spliced out of frame into the last exon whereas eLH/CG-RC contained an in-fram...
Effect of matrix depleting agents on the expression of chondrocyte metabolism by equine chondrocytes. This study was carried out to investigate the effect of two enzymes (collagenase and chondroitinase) and two cytokines/metabolites (interleukin-1beta and retinoic acid) of known catabolic activity on the expression of cartilage metabolism/phenotype in equine articular cartilage. Articular cartilage explants from 11 horses (5-13 years old) were treated for 48 h and assayed for total sulphated glycosaminoglycan (GAG), the incorporation of 35S-sulphate, collagen degradation and mRNA expression of the proteoglycans collagen II, collagen IIA, collagen III, collagen IX, collagen X, collagen XI and g...
Proteomic tools to characterize the protein fraction of Equidae milk. The principal components of the protein fraction in pony mare's milk have been successfully identified and partially characterized using proteomic tools. Skimmed pony mare's milk was fractionated by either reversed phase-high-performance liquid chromatography (RP-HPLC) on a C4 column or a bi-dimensional separation technique coupling RP-HPLC in the first dimension and sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE) in the second dimension (two-dimensional RP-HPLC/SDS-PAGE). The fractions thus obtained were analyzed by Edman N-terminal microsequencing and mass determination, wit...
Detection of endogenous boldenone in the entire male horses. Boldenone (1,2-dehydrotestosterone) is a common veterinary anabolic agent. Its structure is very similar to testosterone. Testosterone is endogenous in the horse, whereas there has been no report concerning the detection of endogenous boldenone. This paper reports the direct observation of sulphate conjugate of boldenone in equine urine from entires. The detection procedures involved solid-phase extraction, immunoaffinity column (IAC) purification, and then LC-MS-MS analysis on a Q-ToF instrument. The identification of boldenone sulphate has provided direct evidence for the endogenous nature o...
Sequence of horse (Equus caballus) apoA-II. Another example of a dimer forming apolipoprotein. Apolipoprotein A-II, the second major apolipoprotein of human HDL, also has been observed in a variety of mammals; however, it is either present in trace amounts or absent in other mammals. In humans and chimpanzee, and probably in other great apes, apoA-II with a cysteine at residue 6 is able to form a homodimer. In other primates as well as other mammals, apoA-II, lacking a cysteine residue, is monomeric. However, horse HDL has been reported to contain dimeric apoA-II that following reduction forms monomers. In this report, we extend these observations by reporting on the first complete sequ...
Daily rhythm of circulating fat soluble vitamin concentration (A, D, E and K) in the horse. BACKGROUND: Many physiological processes of mammalian species exhibit daily rhythmicity. An intrinsic relationship exists between fat soluble vitamins (A, D, E and K) and several body functions. Few investigations on the rhythmic pattern of vitamins in domestic animals have been carried out. The present study evaluated the circadian rhythmicity of fat soluble vitamins in the horse. METHODS: Blood samples from 5 Thoroughbred mares were collected at four-hour intervals over a 48-hour period (starting at 8:00 hours on day 1 and finishing at 4:00 on day 2) via an intravenous cannula inserted into ...
Validated capillary electrophoretic method for the analysis of ivermectin in plasma after intragastric administration in pigs and horses. A capillary electrophoretic (CE) method has been developed for the determination of ivermectin (CAS 70288-86-7), a new generation drug with antiparasitic activity, in pig and horse plasma. The method was statistically validated for its linearity, accuracy, precision and selectivity. The linear range was from 1 to 30 ng mL(-1) with correlation coefficients greater than 0.999. The limit of detection was 0.3 ng mL(-1), while the quantitative limit was 1 ng mL(-1), using a 0.5 mL sample size. The validated procedure was used to determination of pharmacokinetic parameters of ivermectin after ingest...
Effect of L-carnitine administration on the seminal characteristics of oligoasthenospermic stallions. The effect of orally administered l-carnitine on the quality of semen obtained from stallions with different semen qualities was investigated. Four stallions with proven fertility (high motility group, HM) and with normal seminal characteristics (>50% progressive motility and > 80 x 10(6) spermatozoa/ml), and four questionable breeders (low motility group, LM) with <50% of sperm progressive motility and < 80 x 10(6) spermatozoa/ml, received p.o. 20 g of l-carnitine for 60 days. Blood and semen samples were collected before treatment (T0) and after 30 (T1) and 60 days (T2). Semen ev...
Peripartal endocrinology in the mare and foetus. The endocrine profiles in the periparturient mares are dominated by increasing concentrations of progestagens and decreasing oestrogens. These hormones are produced by precursors from the foetus, metabolized by the placenta and act primarily on the maternal uterus. The circulating concentrations of hormones in maternal plasma, generally, represent a small proportion of those metabolized by the foetus and utero-placental tissues. There is clear evidence that the foetal hypothalamo-pituitary-adrenal (HPA) axis initiates the process of foetal maturation and the hormonal cascade which culminates i...
Fat digestibility in Equus caballus follows increasing first-order kinetics. The digestibility of ether extract varies greatly from forages to grains and further to added fats consisting mainly of triglycerides. This variation has been attributed to two main factors, the presence of nonhydrolyzable substances in the ether extract, especially in leafy foods, and the dilution of endogenous fecal fat. A compilation of results from 188 equine digestion balance observations on five basal feeds and 18 test feeds with added fats demonstrated a true digestibility of fat approaching 100% and an endogenous fecal fat of 0.22 g x d(-1) x kg BW(-1). The results revealed that nonhyd...
The apparent digestibility of phytate phosphorus and the influence of supplemental phytase in horses. Availability of phytate-bound P as influenced by supplemental phytase was studied in eight horses consuming four diets in a 4 x 4 Latin square design experiment. The treatments were a control (containing a low P level, 18.4 g/d) and three high-P diets. These diets contained P as monocalcium phosphate (MCP; 43.7 g/d), myoinositol hexakisphosphate in the form of wheat and rice bran (MIHP; 41.8 g/d), or MIHP with microbial phytase (MIHPP; 42.5 g/d). The proportions of phytate-bound P were 3, 1, 55, and 56% for the control, MCP, MIHP, and MIHPP, respectively. The MIHPP diet was supplemented with 3...
Expression of equine interleukin-18 by baculovirus expression system and its biologic activity. The equine interleukin-18 (IL-18) cDNA that contains the coding sequence was cloned and a recombinant baculovirus, named AcEIL-18, was constructed. The recombinant protein of the equine IL-18 was expressed by AcEIL-18 and its expression was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Insect cells infected with AcEIL-18 secreted a precursor IL-18 with 24 kilo dalton (kDa) into the culture supernatant. Western blot analysis showed that mature equine IL-18 about 18 kDa was also confirmed without co-expression of caspase-1. Culture supern...
Cumulus expansion, nuclear maturation and connexin 43, cyclooxygenase-2 and FSH receptor mRNA expression in equine cumulus-oocyte complexes cultured in vitro in the presence of FSH and precursors for hyaluronic acid synthesis. The aim of this study was to investigate cumulus expansion, nuclear maturation and expression of connexin 43, cyclooxygenase-2 and FSH receptor transcripts in equine cumuli oophori during in vivo and in vitro maturation in the presence of equine FSH (eFSH) and precursors for hyaluronic acid synthesis. Equine cumulus-oocyte complexes (COC) were cultured in a control defined medium supplemented with eFSH (0 to 5 micrograms/ml), Fetal Calf Serum (FCS), precursors for hyaluronic acid synthesis or glutamine according to the experiments. After in vitro maturation, the cumulus expansion rate was incr...
Tracing axons of peripheral nerves in rats: a potential technique to study the equine recurrent laryngeal nerve. To study the fascicular anatomy of peripheral nerves, three different groups of retrograde axonal tracers were evaluated: fluorophores, horseradish peroxidase conjugated to subunit B of cholera toxin (CT-HRP), and adeno-associated virus (AAV). The hindlimb nerves in rats served as a model to identify the most efficient tracer in regard to labeling axons within peripheral nerves. The rat's tibial and common peroneal nerves were injected with the different tracers and the sciatic nerve was subsequently examined for evidence of labeled axons. The CT-HRP clearly provided the best results in this r...
Enzyme activities in some types of peripheral leukocytes of thoroughbred race horses before and after the races. Glucose, free fatty acid (FFA), triglyceride (TG) and immunoreactive insulin (IRI) concentrations in plasma and activities of enzymes related to energy metabolism in some types of peripheral leukocytes were measured in thoroughbred race horses before and after racing. Glucose, FFA, TG and IRI concentrations and enzyme activities did not change significantly in plasma. However, the activities of cytosolic hexokinase, malate dehydrogenase (MDH), mitochondrial MDH and aspartate aminotransferase decreased significantly in leukocytes of the horses after the races. The cytosolic ratio of MDH/LDH act...
Detection of enrofloxacin and its metabolite ciprofloxacin in equine hair. Hair analysis to detect drug administration has not been studied extensively in horses. This study aimed to (a) develop an analytical method for enrofloxacin and its metabolite ciprofloxacin in mane and tail hair, (b) relate measured values to doses, routes of administration, hair colour, and (c) demonstrate long-term detectability. Samples were extracted in trifluoroacetic acid at 70 degrees C. Extracts were cleaned-up by solid-phase extraction and analysed by high-performance liquid chromatography with UV-diode array detection. Analyte recoveries were > 87%. Horses were sampled after therape...
Identification of amino-acids in the alpha-subunit first and third loops that are crucial for the heterospecific follicle-stimulating hormone activity of equid luteinizing hormone/choriogonadotropin. To identify amino-acids in the alpha-subunit important for expression of heterospecific FSH activity of horse (e) LH/choriogonadotropin (CG) (eLH) and donkey (dk) LH/CG (dkLH) (FSH/LH ratio ten times higher for eLH than for dkLH); this FSH activity absolutely requires an equid (donkey or horse) alpha-subunit combined with an equid beta-LH subunit. Methods: Chimeric alpha-subunits possessing the first 63 amino-acids of the porcine (p) and the last 33 amino-acids of the donkey alpha-subunit (alphap-dk) and the inverse (alphadk-p) were constructed. Porcine-specific amino-acids were introduced by ...
Detection of surfactant protein A (SP-A) and surfactant protein D (SP-D) in equine synovial fluid with immunoblotting. Once considered unique to the lung, surfactant proteins have been clearly identified in the intestine and peritoneum and are suggested to exist in several other organs. In the lung, surfactant proteins assist in the formation of a monolayer of surface-active phospholipid at the liquid-air interface of the alveolar lining, reducing the surface tension at this surface. In contrast, surface-active phospholipid adsorbed to articular surfaces has been identified as the load-bearing boundary lubricant of the joint. This raises the question of whether surfactant proteins in synovial fluid (SF) are re...
[Concentrations of L-carnitine and parameters of lipid metabolism during estrus in broodmares]. In the present study the concentrations of L-carnitine (total carnitine, free carnitine, and acyl carnitine) and several parameters of the lipid metabolism were measured during the estrus in 10 broodmares. The carnitine concentrations varied in a wide range between the mares. The differences of the mean carnitine concentrations during the estrus did not reach the level of significance (P < 0.05). There was no relationship between the concentrations of total and free carnitine to the follicle size; however, the concentration of acyl carnitine was significantly correlated (r = -0.42). Because...
Phage display of the Equine arteritis virus nsp1 ZF domain and examination of its metal interactions. A putative zinc finger (ZF) domain in the Equine arteritis virus (EAV) nsp1 protein was described recently to be required for viral transcription. The nsp1 ZF (50 aa) was expressed on the surface of M13KE gIII phage, fused to the N terminus of the phage pIII protein. To evaluate the functionality of the ZF domain, a binding assay was developed, based on the use of immobilized Ni(2+) ions (Ni-NTA). Phages displaying ZF bound significantly better to Ni-NTA than did phages displaying negative-control peptides, which also contained metal-coordinating residues. Also, binding of ZF-displaying phages...
Germ cell apoptosis in the testes of normal stallions. Apoptosis in testicular germ cells has been demonstrated in many mammalian species. However, little is known about the stallion (Equus caballus) and rates of apoptosis during spermatogenesis. Morphological and biochemical features of apoptosis reported in other species were used to confirm that the TdT-mediated dUTP Nick end labeling (TUNEL) assay is an acceptable method for identification and quantification of apoptotic germ cells in histological tissue sections from stallion testis. Seminiferous tubules from eight stallions with normal testis size and semen quality were evaluated according t...
Estrogen metabolism in the equine conceptus and endometrium during early pregnancy in relation to estrogen concentrations in yolk-sac fluid. Because estradiol (E(2)) production by the early equine conceptus is considered crucial to the establishment of pregnancy, the amounts of E(2), estrone (E(1)), and their sulfates (E(2)S, E(1)S) were measured by RIA in yolk-sac fluid of 63 conceptuses collected by transcervical lavage over the period of 11-26 days after ovulation. Amounts increased significantly with age of conceptus, especially for E(1)S. Then, the metabolism of E(2), which may be highly relevant for its action, was examined in the conceptus and endometrium over the period when the conceptus ceases to migrate within the uterus...
Metabolism of MDCK cells during cell growth and influenza virus production in large-scale microcarrier culture. The production of equine influenza in Madin-Darby canine kidney (MDCK) cells in large-scale microcarrier culture is described with detailed on- and off-line analytical data during cell growth and virus replication. Metabolite concentration profiles for glucose, glutamine, lactate and ammonium are shown. Lactate and ammonium concentrations were always below inhibiting levels. Concentration profiles for essential and non-essential amino acids of the cell culture medium are discussed. During cell growth proline was released into the medium with a significant rate while two amino acids, serine and...
