Topic:In Vitro Research
In vitro research involving horses refers to the study of equine cells, tissues, or biological molecules outside their normal biological context, typically in controlled laboratory environments. This research approach allows scientists to investigate cellular processes, molecular interactions, and the effects of various treatments without the ethical and logistical complexities of in vivo studies. In vitro studies contribute to understanding equine physiology, pathology, and pharmacology by providing insights into cellular responses to pathogens, drugs, and other stimuli. This page compiles peer-reviewed research studies and scholarly articles that explore various in vitro methodologies and their applications in equine science, including cell culture techniques, molecular assays, and drug efficacy testing.
Application of high-performance liquid chromatography–inductively coupled plasma mass spectrometry to the investigation of cadmium speciation in pig kidney following cooking and in vitro gastro-intestinal digestion. The speciation of cadmium in retail pig kidney has been examined by size-exclusion chromatography (SEC) coupled directly to inductively coupled plasma mass spectrometry (ICP-MS). Approximately 35% of the cadmium from uncooked kidney was soluble after aqueous extraction at pH 8 and SEC - ICP-MS revealed three discrete peaks whose retention times corresponded to estimated relative molecular masses of 1.2 x 10(6), 7.0 x 10(4) and 6 x 10(3)-9 x 10(3). In the cooked kidney, 35% of the Cd was soluble and was all associated with a peak of a relative molecular mass (Mr) of 6 x 10(3)-9 x 10(3). After s...
Functional and ligand binding studies suggest heterogeneity of platelet prostacyclin receptors. 1. This study describes attempts to compare prostacyclin (IP-) receptors in human, pig, horse, rabbit and rat platelets and in circular muscle of human, rabbit and dog mesenteric and pig gastroepiploic arteries. Three stable prostacyclin analogues, iloprost, cicaprost and 6a-carba-prostacyclin (6a-carba-PGI2) and a prostaglandin endoperoxide analogue EP 157 (previously shown to mimic prostacyclin on human platelets) were used. 2. Our main conclusion is that prostacyclin receptors on human, pig and horse platelets are similar in nature, but distinct from those on rabbit and rat platelets. Funct...
Cell mediated immune responses in ponies following infection with equine influenza virus (H3N8): the influence of induction culture conditions on the properties of cytotoxic effector cells. Cytotoxic cell precursors and/or cytotoxic memory cells were demonstrated in the peripheral blood of ponies after aerosol infection with influenza A/equine/Newmarket/79 (H3N8). In order to reveal their cytotoxic potential, peripheral blood mononuclear cells required a secondary antigenic stimulation. In vitro induced cytotoxic cells showed activity against influenza infected target cells in a 3-4 h 51Cr-release assay. The reactivity of cytotoxic cells was markedly influenced by the conditions of the secondary induction culture. If high concentrations of exogenous crude equine IL-2 were used, v...
The effect of immunity to core lipopolysaccharides (LPS) on the production of thromboxane and prostacyclin by equine peritoneal macrophages. An experiment was designed to determine whether a change in the ability of macrophages to respond to lipopolysaccharides (LPS) of gram-negative bacteria was involved in the development of cross-reactive immunity to endotoxemia. The endotoxin-induced production of thromboxane A2(TxA2) and prostacyclin (PGI2) by peritoneal macrophages from horses which were hyperimmunized against the common core region of LPS were compared to those in unimmunized horses. Bacterins used for induction of core LPS immunity were prepared from the J-5 mutant of Escherichia coli 0111:B4, and the R 595 mutant of Salmon...
The lipoxygenase pathway and chemiluminescence in horse eosinophilic leukocytes. It was shown in several cell types that the dual lipoxygenase and cyclooxygenase inhibitor eicosatetraynoic acid but not the cyclooxygenase inhibitor acetylsalicylic acid suppressed luminol-dependent chemiluminescence. Since lipoxygenase is known to generate chemiluminescence in vitro, these observations were interpreted as evidence for a direct contribution of the lipoxygenase pathway to light emission in intact cells. We have investigated a possible contribution of the lipoxygenase to the chemiluminescence of horse eosinophils by directly comparing the formation of the byproduct chemilumines...
Comparative toxicity of the horse eosinophil peroxidase-H2O2-halide system and granule basic proteins. Stimulated eosinophils release cytotoxic granule constituents, including eosinophil peroxidase (EPO) and a group of granule basic proteins (GBP). EPO reacts with H2O2 formed by the respiratory burst and a halide to form cytotoxic oxidants. The relative potency of the EPO-H2O2-halide system and the GBP is considered here. Horse eosinophils were induced to degranulate, the degranulation products were separated by chromatography on Sephadex G-50 and comparable volumes of the column fractions were tested for toxicity to Escherichia coli and the schistosomula of Schistosoma mansoni in the presence ...
Mössbauer spectroscopic study of the initial stages of iron-core formation in horse spleen apoferritin: evidence for both isolated Fe(III) atoms and oxo-bridged Fe(III) dimers as early intermediates. Ferritin stores iron within a hollow protein shell as a polynuclear Fe(III) hydrous oxide core. Although iron uptake into ferritin has been studied previously, the early stages in the creation of the core need to be clarified. These are dealt with in this paper by using Mössbauer spectroscopy, a technique that enables several types of Fe(II) and Fe(III) to be distinguished. Systematic Mössbauer studies were performed on samples prepared by adding 57Fe(II) atoms to apoferritin as a function of pH (5.6-7.0), n [the number of Fe/molecule (4-480)], and tf (the time the samples were held at room ...
DNA polymorphism analysis of hereditary multiple exostoses in horses. Genomic DNA polymorphisms obtained by restriction fragment-length polymorphism from healthy horses and horses with hereditary multiple exostoses were analyzed. These DNA were digested by 12 restriction enzymes and were hybridized against 6 isotopically labeled oncogene probes. Hybridization was not detected with the viral oncogene, v-ras, which indicated this oncogene was absent in the equine genome. Oncogenes (c-raf-1, c-fes, c-myb, c-myc, and c-sis) were present and had similar hybridization patterns and signal intensities in DNA from healthy horses and horses with hereditary multiple exosto...
Diagnostic aids for the detection of urine in the equine ejaculate. An experiment was conducted to evaluate three commercially available test kits, the Azostix, Multistix and Uric-acid test, for the detection of urine in the equine ejaculate. Azostix, which tests for urea nitrogen, consistently detected urine in the equine ejaculate. Urine contamination was evident when a color change occurred in the reagent pad, going from yellow to green after 10 sec of exposure. The sensitivity of Azostix to urea nitrogen in contaminated samples was 39 mg/dl. The Multistix test kit also successfully detected urine in semen. In the Multistix nitrite pad the color changed fro...
Equine peritoneal macrophage production of thromboxane and prostacyclin in response to platelet activating factor and its receptor antagonist SRI 63-441. The formation of eicosanoids may be a primary route through which platelet activating factor (PAF) exerts its effects during endotoxemia. Since endotoxemia is a common cause of death in horses, a study was conducted to determine whether PAF could stimulate equine macrophage release of thromboxane A2 (TxA2) and prostacyclin (PGI2) and whether a PAF-receptor antagonist would alter macrophage eicosanoid synthesis. Equine peritoneal macrophages were cultured from clinically normal horses and exposed to various concentrations of PAF, the PAF-receptor antagonist SRI 63-441, endotoxin, or a combinati...
In-vitro plasma protein binding of propafenone and protein profile in eight mammalian species. The protein binding of propafenone in vitro was assessed in plasma of mouse, rat, rabbit, dog, sheep, man, cow, and horse at two concentration levels. In all species and at both concentrations propafenone was found highly bound (86-99%) to plasma proteins. No significant relationship was found between free propafenone and the plasma protein fractions. A concentration-dependency was seen in plasma of mouse, sheep, man, and horse, in which the free fraction of propafenone became larger on raising the concentration. Qualitative and quantitative differences were observed in the protein plasma prof...
Serum angiotensin converting enzyme activity and response to angiotensin I in horses. The activity of serum angiotensin converting enzyme (ACE) for healthy horses was 64 +/- 13 mUnits/ml. In vitro, equine serum ACE was sensitive to the following inhibitors (IC50): enalapril (570 nM or 215 ng/ml), captopril (190 nM or 41.3 ng/ml), and enalaprilat (6 nM or 2.1 ng/ml). The intravenous (i.v.) administration of angiotensin I to six healthy horses produced a dose proportional pressor response. The maximal increase in mean arterial pressure over baseline values was 65.6 mmHg at angiotensin I doses of 500 ng/kg bodyweight (bwt). The attenuation of this response to angiotensin I was fur...
Observations on the colic motor complex in a pony with a small intestinal obstruction. Characteristic motility patterns were seen throughout the gastrointestinal tract in a pony prepared chronically with electromechanical recording devices after developing a simple obstruction of the small intestine. Gross distension of the stomach with fluid produced loss of gastric contractile activity and a chaotic electrogram. These changes were reversed instantaneously when the stomach was decompressed. In the jejunum, proximal to the obstruction, the unique 'colic motor complex' was observed with contractions of longer duration arranged in characteristic pulses of activity. The left dorsal...
The sequence-specific assignment of the 1H-NMR spectrum of an enzyme, horse-muscle acylphosphatase. A complete range of two-dimensional NMR experiments was used for the assignment of the 1H-NMR spectrum of horse muscle acylphosphatase. Firstly the spin systems of some easily identifiable amino acid side chains were assigned. These side chains involved all the aromatic residues and all the leucine, valine, isoleucine, threonine, alanine, proline as well as some of the glycine residues. Analysis of nuclear Overhauser enhancement spectra in our previous work had identified the sequential and long-range patterns characteristics for secondary structure elements. This result had also provided the ...
Change in host cell tropism associated with in vitro replication of equine infectious anemia virus. Similar to other human and animal lentiviruses, equine infectious anemia virus (EIAV) is detectable in vivo in cells of the monocyte-macrophage lineage. Owing to their short-lived nature, horse peripheral blood macrophage cultures (HMC) are rarely used for in vitro propagation of EIAV, and equine dermal (ED) or kidney cell cultures, which can be repeatedly passed in vitro, are used in most studies. However, wild-type isolates of EIAV will not grow in these cell types without extensive adaptation, a process which may attenuate viral virulence. To better define the effect of host cell tropism on...
Castor-oil induced diarrhoea in ponies: a model for acute colitis. A reproducible, reversible model of colitis induced in ponies by administering castor oil (2.5 ml/kg bodyweight [bwt] per os) was characterised by abdominal pain, fever, watery diarrhoea, dehydration, hypovolaemia, toxaemia, leucopenia, decreased serum Cl, Na and K levels and metabolic acidosis. The signs were most severe between 24 and 48 h post induction, stabilisation was frequently observed after 72 h, although diarrhoea could persist beyond 96 h. Morphological and in vitro transport studies (right ventral colon) were conducted on tissues from animals destroyed at 24, 48 and 72 h. In the c...
Androgen synthesis and aromatization by equine corpus luteum microsomes. Whereas mare corpus luteum does not produce androgens or estrogens in vivo, the incubation of mare corpus luteum microsomes with progesterone and NADPH resulted in 17 alpha-hydroxyprogesterone and estrogen production with a small yield of androstenedione. In the presence of an aromatase inhibitor (4-hydroxyandrostenedione), 17 alpha-hydroxyprogesterone and androstenedione were accumulated. Aromatization of testosterone and androstenedione occurred via stereospecific loss of the 1 beta, 2 beta hydrogen atoms and was inhibited by MgCl2, KCl, and EDTA. The Km of estrogen synthetase from equine co...
Transendoscopic neodymium:yttrium aluminum garnet laser irradiation in horses. A neodymium:yttrium aluminum garnet (Nd:YAG) laser was used to study effects of applying laser irradiation transendoscopically to the corniculate process of the arytenoid cartilage in horses. Dosimetry was established initially in vitro in 10 corniculate cartilages that were irradiated and examined histologically to determine penetration depths at selected power settings. Eleven horses were given xylazine IV and butorphoral tartrate IV, and their left ventricle and corniculate process were irradiated. Six horses had left laryngeal hemiplegia and were euthanatized and necropsied 14 weeks after ...
(+)-[3H]-PN 200-110 binding to cell membranes and intact strips of portal vein smooth muscle: characterization and modulation by membrane potential and divalent cations. 1. Specific binding of the calcium-antagonist dihydropyridine derivative, (+)-[3H]-PN 200-110 (isradipine), to cell membranes of equine portal vein smooth muscle was compared with binding to intact strips isolated from rat portal veins. 2. Specific binding to vascular smooth muscle membranes was of high affinity, saturable and reversible. The dissociation constant obtained from association and dissociation kinetics of (+)-[3H]-PN 200-110 was similar to that obtained from equilibrium binding and competition experiments. 3. Specific binding of (+)-[3H]-PN 200-110 was completely displaced by unla...
Tubuloreticular inclusions in equine connective tissue neoplasms. Abnormal irregularly branched and anastomosing tubules within cisternae of endoplasmic reticulum were observed by transmission electron microscopy in tumour cells comprising connective tissue neoplasms (sarcoids) from three horses and a mule. These tubuloreticular inclusions were also observed in cultured tumour cells from one of these horses examined, but were not detected in fibroblasts (fibrocytes), epidermis, or vascular endothelial cells in skin biopsy specimens from five clinically healthy horses, nor in one additional equine connective tissue neoplasm.
A spectrophotometric procedure for the determination of objective measurements of equine spermatozoan motility. A spectrophotometric procedure was developed and evaluated for the objective measurement of equine spermatozoan motility. A 100 mul sample of a sperm suspension, prepared by the removal of seminal plasma, was layered under a column of optically clear medium in a specially designed spectrophotometric cuvette maintained at 37 degrees C. Changes in light transmittance above the interface of the sperm suspension and medium were recorded on chart paper. As sperm cells swam into the medium, a decrease in light transmittance was recorded as a deflection on the chart paper. Chart recordings were analy...
[The possibility of using equine serum albumin in place of bovine serum albumin and ovalbumin in radioimmunological and immunoenzyme analyses and in virological practice]. Horse serum albumin has been shown to meet the requirements to protein preparations for microanalysis and thus to be suitable for use in kits of reagents for the radioimmunological determination of insulin and myoglobin, for the determination of tick-borne encephalitis virus antigen by the method of the enzyme immunoassay and for the stabilization of proteins in the hemagglutination test and the hemagglutination inhibition test.
Reactivity of equine palmar digital arteries and veins to vasodilating agents. Palmar digital arteries and veins removed surgically from healthy horses under general anesthesia were cut into 4 mm vascular rings, suspended in tissue baths, and attached to force displacement transducers for continuous measurement of vascular tension. In vitro vascular responses were determined for acetylcholine, acepromazine, isoxsuprine hydrochloride (isoxsuprine), prostaglandin E2 (PGE2), and prostaglandin I2 (prostacyclin). After preconstriction with norepinephrine hydrochloride (norepinephrine), or prostaglandin F2 alpha (PGF2 alpha), the concentrations needed to produce 50% maximum re...
The preparation and biochemical characterization of intact capsids of equine infectious anemia virus. Capsids of equine infectious anemia virus have been isolated as cone-shaped particles 60 x 120 nm in size. Detergent treatment of whole virus followed by two cycles of rate-zonal centrifugation in Ficoll produces these capsids in a yield of approximately 10%. The major protein components are the gag-encoded p11 nucleocapsid protein and p26 capsid protein, which are present in equimolar amounts. Substantial cleavage of p11 to p6 and p4 can be observed under conditions where the viral protease packaged in the capsid is enzymatically active.
Nonequivalence of the two subunits of horse erythrocyte glutathione transferase in their reaction with sulfhydryl reagents. Glutathione transferase (EC 2.5.1.18) from horse erythrocytes has been purified and some molecular and kinetic properties have been investigated. It appears to be a dimeric protein composed of subunits of about 23 kDa, indistinguishable either in sodium dodecyl sulfate or in urea electrophoresis. Amino acid composition, substrate specificities, sensitivity to inhibitors, CD spectra, and immunological studies provide evidence that the horse enzyme is related to the pi class transferases. This enzyme has only two reactive thiol groups/dimer whose integrity appears to be essential for the activit...
In vitro reactivity of digital arteries and veins to vasoconstrictive mediators in healthy horses and in horses with early laminitis. The in vitro reactivity of vasoconstrictive mediators that are implicated in acute laminitis was determined in palmar and plantar digital arteries and veins obtained from healthy horses and in palmar digital vessels of horses with early laminitis (Obel grade I). To obtain baseline reactivity data, 3 experiments were conducted, using healthy horses: (1) the reactivity of palmar and plantar digital arteries and veins to angiotensin II, norepinephrine, and 5-hydroxytryptamine (serotonin) were compared; (2) the direct effects of bacterial endotoxin on vascular reactivity were assessed; and (3) the...
Synthesis and aromatization of 19-norandrogens in the stallion testis. The results of the measurement of 19-nortestosterone in the testiscular artery and vein of the stallion, the very low levels of this steroid in the peripheral blood of geldings and the similar patterns of increase in the peripheral levels of 19-nortestosterone and testosterone after hCG stimulation, show that 19-nortestosterone, like testosterone, is essentially synthesized in the testis. This testicular origin was confirmed by the ability of testicular tissue to synthesize 19-norandrogens from [4-14C]androgens in vitro. 19-Nortestosterone was 50% conjugated in the peripheral blood and almost ...
Surface antigens on equine sarcoid cells and normal dermal fibroblasts as assessed by xenogeneic antisera. To characterise the expression of surface antigens on equine sarcoid cells compared to normal equine fibroblasts, immune sera were produced in rabbits against transformed cells of a virus-containing sarcoid cell line (Mc-1) and normal dermal fibroblasts, respectively. The specificities of the sera were analysed by antibody-dependent cellular cytotoxicity against 51Cr-labelled target cells using human lymphocytes as effector cells. Anti-Mc-1 antiserum induced strong cytotoxicity against transformed cells of two sarcoid cell lines (Mc-1 and Bay Mc-1), whereas the cytotoxicity against transformed...
[Ruminal 15N-straw degradation in sacco and the in vivo digestibility of 15N from straw in the sheep and pony]. The nylon-bag method was applied for determination of the rumen degradation of dry matter and nitrogen of 15N-labelled wheat straw. For the experiment 4 wethers fitted with a rumen cannula were used. The bags containing 15N straw were introduced into the rumen and withdrawn 3, 6 or 12 h after incubation. In a second experiment the apparent 15N-digestibility of the same straw was determined in wether and pony. The dry matter disappearance varied between 6 and 23%. For 15N-labelled straw the disappearance of 15N was higher than that of total N. 12 h after incubation 71% of 15N and only 25% of to...