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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.
Variation of an epitope of keratan sulphate and total glycosaminoglycans in normal equine joints. An epitope of keratan sulphate (KS) and total glycosaminoglycans (GAG) were measured in synovial fluid samples from joints of 53 horses immediately following humane destruction. Internal examination of the joints post mortem ensured that there was no gross evidence of osteoarthritis or other joint disease. Joints sampled were distal interphalangeal (DIP), proximal interphalangeal (PIP), metacarpophalangeal (MCP), metatarsophalangeal (MTP), tarsometatarsal (TMT), tarsocrural (TC), femoropatellar (FP) and antebrachiocarpal (ABC) joints. The age of each horse was assessed by examination of the te...
Studies into aromatase activity associated with fetal allantochorionic and maternal endometrial tissues of equine placenta. Identification of metabolites by gas chromatography mass spectrometry. Maternal endometrial and fetal allantochorionic tissues were separated manually from the placentae of seven healthy thoroughbred and three pony mares, ranging in gestational age from 100 to 318 days. The homogeneity of subcellular fractions prepared from these tissues was assessed initially using the marker enzymes, succinate dehydrogenase, NADPH cytochrome C reductase and lactate dehydrogenase for the mitochondrial, microsomal and cytosolic fractions, respectively. Light microscopy and histochemical analysis demonstrated that the separated fetal allantochorionic membrane, which is made up of ...
Role of carbohydrates in the attachment of equine spermatozoa to uterine tubal (oviductal) epithelial cells in vitro. To test the hypotheses that the attachment of equine spermatozoa to uterine tubal (oviductal) epithelial cells (OEC) in vitro is mediated by glycoproteins, and that proteins with carbohydrate-binding properties are present in the periacrosomal plasma membrane of equine spermatozoa. Methods: 4 reproductively sound stallions, and 1 mare in estrus. Methods: In experiment 1a, fluorescent-labeled spermatozoa were cocultured with monolayers of OEC in the presence of 50 mM glucose, fructose, galactose, mannose, N-acetyl glucosamine, N-acetyl galactosamine, or N-acetyl neuraminic acid, or 10 mg of fet...
Modulation of matrix metalloprotease 13 (collagenase 3) gene expression in equine chondrocytes by interleukin 1 and corticosteroids. To determine whether matrix metalloprotease 13 (MMP-13; collagenase 3) is produced by equine chondrocytes and to investigate modulation of its expression by recombinant human interleukin 1 beta (rhIL-1 beta) and corticosteroids. Methods: Equine chondrocytes in monolayer culture were stimulated with rhIL-1 beta. Total RNA was extracted, purified, and reverse transcribed into DNA. Using appropriate primers, a putative MMP-13 fragment was amplified by polymerase chain reaction, and cloned into a bacterial vector. The resultant fragment was purified and sequenced, then was used to prepare a digoxi...
Immunolocalization of aromatase P-450 in ovarian tissue from pregnant and nonpregnant mares and in ovarian tumours. Aromatase P-450 (P-450arom) is a crucial regulatory enzyme that is necessary for conversion of androgens to oestrogens. Corpora lutea and follicles were obtained from the ovaries of cyclic mares and from mares at day 20 and days 40-70 of pregnancy. The presence of P-450arom within specific cell types was investigated by immunostaining to determine potential sites of oestrogen synthesis. Immunoreactivity for P-450arom was confined to the granulosa layer of non-atretic follicles > 5 mm in diameter and to corpora lutea at all stages of the oestrous cycle and during pregnancy. These findings co...
Purification, crystallization and preliminary crystallographic analysis of mare lactoferrin. Lactoferrin is an iron-binding glycoprotein with a molecular weight of 80 kDa. The protein has two iron binding sites. It has two structural lobes, each housing one Fe(3+) and the synergistic CO(3)(2-) ion. The protein was isolated from the colostrum/milk of mares maintained at National Research Centre on Equines, Hisar, India. The purified samples of the protein were crystallized using a microdialysis method. The protein was dialysed against low ionic strength buffer solution. Several crystal forms were obtained, out of which three were characterized which have cell dimensions as follows. For...
Use of peanut agglutinin to assess the acrosomal status and the zona pellucida-induced acrosome reaction in stallion spermatozoa. Peanut agglutinin (PNA) was used to assess the sperm acrosomal status and the acrosome reaction during gamete interaction in the equine species. PNA exclusively binds to the outer acrosomal membrane of stallion spermatozoa, as was established by transmission electron microscopy. Fluorescein isothiocyanate-PNA (FITC-PNA) labeling was used to monitor sperm acrosomal changes during a prolonged incubation period of 24 hours and during a 2-hours incubation in the presence of 5 microM calcium ionophore A23187. In addition, after a 4-hours preincubation in SP-TALP medium, sperm samples were incubated...
Expression of Vitreoscilla hemoglobin is superior to horse heart myoglobin or yeast flavohemoglobin expression for enhancing Escherichia coli growth in a microaerobic bioreactor. Expression of a gene encoding hemoglobin (VHb) from the aerobic bacterium Vitreoscilla sp. in several organisms, including Escherichia coli, has been shown to improve microaerobic cell growth and enhance oxygen-dependent product formation. The suitability of VHb to enhance microaerobic metabolism has been suggested to depend on its unusual oxygen binding characteristics. To examine whether hemoproteins of other origins can also elicit the positive effects VHb exerts in microaerobic E. coli cells, we subcloned the genes encoding Vitreoscilla VHb, horse heart myoglobin (HMb), and yeast flavohemo...
Study of the haemolytic process and receptors of thermostable direct haemolysin from Vibrio parahaemolyticus. The haemolytic action of 125I-labelled thermostable direct haemolysin from Vibrio parahaemolyticus was studied on human and equine erythrocytes. In the first step, the haemolysin bound to the membranes of both erythrocyte species. This binding seemed temperature-independent. Then, for human erythrocytes, haemolysin produced cell disruption, and haemoglobin was released. Following this step, haemolysin was also released in a temperature-dependent manner. In contrast, equine erythrocytes were not disrupted, and no release of haemolysin occurred. The receptors of labelled haemolysin were analysed...
The complete mitochondrial DNA (mtDNA) of the donkey and mtDNA comparisons among four closely related mammalian species-pairs. The nucleotide sequence of the complete mitochondrial genome of the donkey, Equus asinus, was determined. The length of the molecule is 16,670 bp. The length, however, is not absolute due to pronounced heteroplasmy caused by variable numbers of two types of repetitive motifs in the control region. The sequence of the repeats is (a) 5'-CACACCCA and (b) 5'-TGCGCGCA, respectively. The order of (a) and (b) can be expressed as {n[2(a)+(b)]+m(a)}. In 32 different clones analyzed the number of n and m ranged from 0 to 9 and 1 to 7. The two rRNA genes, the 13 peptide-coding genes, and the 22 tRNA gene...
Identification of an alternatively spliced transcript of equine interleukin-1 beta. Using lipopolysaccharide (LPS)-stimulated equine peripheral blood mononuclear cell (PBMC) cDNA as a template, we performed polymerase chain reaction (PCR) amplification with equine interleukin-1 beta (IL-1 beta) specific primers. Electrophoresis of the PCR product on agarose gel revealed an additional smaller fragment that hybridized with an equine IL-1 beta cDNA probe. Sequencing of this fragment demonstrated that it was shorter than normal equine IL-1 beta cDNA by 162 nucleotides, which corresponded to exon 5 of the human and murine IL-1 beta genes. The deletion of 162 nucleotides did not re...
Interaction of GroEL with conformational states of horse cytochrome c. GroEL interacts with proteins in denatured states and promotes their efficient folding. To understand the conformational features required for the substrate, we studied the interactions of GroEL with various derivatives of horse cytochrome c including porphyrin-cytochrome c, apo-cytochrome c, and the three fragments containing the heme group, i.e. fragments 1-65, 1-38 and 11-21. Size-exclusion chromatography was performed, taking advantage of the heme absorption of the fluorescence label. Under low-salt conditions, significant binding to GroEL was observed for porphyrin-cytochrome c, apo-cytoc...
Antagonistic effects of atipamezole on medetomidine-induced sedation in horses. The antagonistic effects of atipamezole (20, 40, 60, 80, and 100 micrograms/kg i.v.) on medetomidine (10 micrograms/kg i.v.)-induced sedation were evaluated in horses. Although 20 and 40 micrograms/kg of atipamezole were not sufficient to reverse the sedation, 60 micrograms/kg did effectively reverse the sedation. Atipamezole at 80 micrograms/kg was more potent, and significantly shortened the duration of sedation without any apparent side effects, but a higher dose of 100 micrograms/kg was not more effective than 80 micrograms/kg. The possible use of atipamezole as a reversal agent may enhanc...
Myosin heavy chain isoforms in adult equine skeletal muscle: an immunohistochemical and electrophoretic study. The aim of this study was to characterize the myosin heavy chain (MyHC) isoforms present in equine skeletal muscle. Methods: Muscle biopsies were removed from the superficial region of the gluteus medius muscle of five mature horses and analyzed by immunohistochemistry (using a battery of monoclonal antibodies specific for rat MyHC isoforms) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results: Immunohistochemistry allowed subdivision of three different muscle fiber populations containing a single MyHC, one slow and two fast, and two hybrid populations, one containing slow an...
Activation of apical P2U purine receptors permits inhibition of adrenaline-evoked cyclic AMP accumulation in cultured equine sweat gland epithelial cells. Experiments were undertaken using cultured equine sweat gland epithelial cells that express purine receptors belonging to the P2U subclass which allow the selective agonist uridine triphosphate (UTP) to increase the concentration of intracellular free Ca2+ ([Ca2+]i). Experiments using pertussis toxin (Ptx), which inactivates certain guanine-nucleotide-binding proteins (G-proteins), showed that this response consisted of Ptx-sensitive and Ptx-resistant components, and immunochemical analyses of the G-protein alpha subunits present in the cells showed that both Ptx-sensitive (alpha i1-3) and Ptx...
Variability of cardiomyocyte DNA content, ploidy level and nuclear number in mammalian hearts. DNA content, ploidy level, cell size and nuclear number were investigated in 54 mammalian hearts from nine species. DNA content was determined biochemically and ploidy level of cells was studied by the means of Feulgen cytophotometry. Nuclear number was calculated by a new method, while cell size was determined by using ocular micrometry. In most mammals diploid cell nuclei predominate. Higher ploidy levels were found in the human and the pig hearts. The total amount of DNA correlated with the myocardial weight. Eight million heart muscle cell nuclei were found in mice (myocardial weight 160 m...
Excitatory prejunctional beta 2-adrenoceptor distribution within equine airway cholinergic nerves. We examined the effect of activation of beta 2-adrenoceptor (AR) by isoproterenol (ISO) on acetylcholine (ACh) release evoked by electrical field stimulation (EFS: 20 V, 0.5 Hz, 0.5 msec) from cholinergic nerves in five regions of equine airways. We also tested if the effect of ISO was dependent on epithelium or prostanoids by examining the effect of ISO on ACh release in the presence and absence of epithelium or cyclooxygenase inhibition. Trachealis strips or bronchial rings were preincubated for 60 min with 10(-7) M atropine, 10(-6) M neostigmine, and 10(-5) M guanethidine. The ACh amount wa...
Nucleologenesis and ribonucleic acid synthesis in preimplantation equine embryos. The nucleolus is believed to be the active site of rRNA synthesis in all eukaryotic cells. In preimplantation embryos, the embryonic genome is apparently more or less silent up to a species-specific developmental stage at which a major burst of transcription occurs. Here we report on nucleologenesis and some ultrastructural aspects of the onset of RNA synthesis in equine embryos during in vivo development. The zygotes and embryos up to blastocyst stages were surgically recovered from normally cycling mares. Mares were induced to ovulate by treatment with 3000 IU hCG and inseminated 20 and 34 h...
[Enantioselectivity in the excretion of glucuronides of carprofen in man, dogs and horses]. After administration of the racemic drug, the stereoselective quantification of the enantiomers of free and conjugated carprofen was performed in human plasma and in plasma, urine and bile of dogs and horses. In humans, the plasma profile of free carprofen and its glucuronides is not stereoselective and the glucuronides excreted in urine are close to a racemate. In dogs and horses on the contrary, the R(-) enantiomer of the free drug is predominant in plasma, while urine and/or bile concentrations of the glucuronides are high in comparison to plasma with a strong selectivity for the S(+) enant...
Characterization of polypeptides synthesized and secreted by oviductal epithelial cell explants obtained from young, fertile and aged, subfertile mares. To compare the electrophoretic patterns of proteins synthesized and secreted by oviductal epithelial cell (OEC) explants obtained from young, fertile and aged, subfertile mares. Methods: Young, fertile (n = 5; 2 to 7 years old) and aged, subfertile (n = 5; 17 to 24 years old) mares. Methods: 2-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and computerized densitometry. Results: Variation in the synthesis and secretion of polypeptides from young, fertile mare OEC (YOEC) and aged, subfertile mare OEC (AOEC) was evidenced by differences in the intensity of radiolabeled pol...
Equine cytochrome P450 aromatase exhibits an estrogen 2-hydroxylase activity in vitro. Aromatase (estrogen synthetase) is a steroidogenic enzyme complex which catalyzes the conversion of androgens to estrogens (termed aromatization). This enzyme was purified from adult equine testis to homogeneity by five chromatographic steps. The ability of purified and reconstituted equine aromatase to exhibit an estrogen 2-hydroxylase activity was tested and compared to testosterone aromatization. Enzymatic activities were assessed by tritiated water release from labelled estradiol and testosterone. Kinetic analysis of estradiol 2-hydroxylation showed an apparent K(m) of 23 microM and a V(ma...
Pharmacokinetics of ketoprofen in the donkey (Equus asinus). The pharmacokinetic parameters of ketoprofen were determined in four donkeys after a single intravenous injection of a dose of 2.2 mg/kg body weight. The total body clearance (ClB) was 414.0 +/- 98.70 ml/h/kg (mean +/- SD), the volume of distribution at steady state (Vss) 263.10 +/- 55.43 ml/kg and the elimination half-life 1.30 +/- 0.75 h. These values were compared to those obtained in horses.
Daily fluctuations of haematology and blood biochemistry in horses fed varying levels of protein. Changes in the plasma biochemistry of 12 stallions were studied over seven 24 h periods. Twelve healthy male horses of 4 different breeds (Thoroughbred [TB], Anglo-Arabo-Sardo [AAS], Avelignese [Av] and Maremmano [M]) were divided into 3 experimental groups and fed with an isoenergetic maintenance ration with different protein levels. The same amount of food was given daily at 0730 and 1930 h. After 20 days of pretrial, a series of 6 blood samples were collected daily from the jugular vein every 4 h for 7 days. The samples were analysed for packed cell volume (PCV), haemoglobin, plasma glucose...
Maintenance of equine articular cartilage explants in serum-free and serum-supplemented media, compared with that in a commercial supplemented medium. To evaluate the effects of a commercially defined, serum-free medium additive on equine articular cartilage explants, compared with effects of serum-free and serum-supplemented media. Methods: Articular cartilage from a 3-year-old, mixed breed horse euthanatized for reasons other than musculoskeletal disease or sepsis. Methods: Media were changed every 48 hours, and the glycosaminoglycan (GAG) content was determined in media collected at each time point. Glycosaminoglycan synthesis by explant chondrocytes, and residual GAG content of articular cartilage (as a measure of explant GAG loss) were ...
Molecular diffusion into horse spleen ferritin: a nitroxide radical spin probe study. Electron paramagnetic resonance spectroscopy and gel permeation chromatography were employed to study the molecular diffusion of a number of small nitroxide spin probes (approximately 7-9 A diameter) into the central cavity of the iron-storage protein ferritin. Charge and polarity of these radicals play a critical role in the diffusion process. The negatively charged radical 4-carboxy-2,2,6,6-tetramethylpiperidine-N-oxyl (4-carboxy-TEMPO) does not penetrate the cavity whereas the positively charged 4-amino-TEMPO and 3-(aminomethyl)-proxyl radical and polar 4-hydroxy-TEMPO radical do. Unlike th...
Calcium buffering is required to maintain bone stiffness in saline solution. This work determined whether mineral dissolution due to prolonged testing or storage of bone specimens in normal saline would alter their elastic modulus. In one experiment, small pieces of equine third metacarpal bone were soaked in normal saline supplemented with varying amounts of CaCl2. Changing Ca ion concentrations in the bath were monitored and the equilibrium concentration was determined. In a second experiment, the elastic moduli of twenty 4 x 10 x 100 mm equine third metacarpal beams were determined non-destructively in four-point bending. Half the beams were then soaked for 10 days ...
Measurement of bone specific alkaline phosphatase in the horse: a comparison of two techniques. For many years total alkaline phosphatase (AP) activity in serum has been used to monitor bone metabolism in different species. However, total AP lacks bone specificity because the total activity in serum is made up of several isoenzymes, of which the liver and bone isoforms predominate. The aim of the present study was to evaluate an immunoradiometric assay for measuring bone specific alkaline phosphatase (BAP) in horses. BAP, a specific marker of bone formation, was measured in sera from thoroughbred horses by using a previously characterised wheat germ lectin (WGL) precipitation assay and a...
