Omneity® Pellets
All-In-One Vitamin & Mineral Pellet
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.
Penetration of frozen-thawed, zona-free hamster oocytes by fresh and slow-cooled stallion spermatozoa. A method for preparing stored unfrozen stallion spermatozoa for the zona-free hamster oocyte penetration test (HOPT) and a subsequent comparison of fresh and stored sperm by the HOPT were evaluated. In Experiment 1, sperm from 4 stallion ejaculates, cooled to 4 degrees C and stored for 24 h, were treated with 60, 90 and 120 microM of dilauroylphosphatidyl-choline (PC12) liposomes to initiate the acrosome reaction. The percentage of motile and acrosome-reacted (AR) sperm were recorded after 8, 15 and 30 min of incubation at 39 degrees C, by automated image analysis. Liposome concentration did n...
Variations in structural and functional changes of stallion spermatozoa in response to calcium ionophore A23187. Three experiments were conducted to assess the structural and functional changes of stallion spermatozoa in response to the calcium ionophore A23187, and to determine individual variation between stallions. In Experiment 1, changes in the acrosome of spermatozoa exposed to 7.14 microM A23187 for fixed times between 0 and 120 min were examined. There was a steady increase with time in the number of spermatozoa undergoing the acrosome reaction although the rate of increase differed between stallions. Sperm motility decreased sharply when incubation was extended beyond 30 min. In Experiment 2, th...
Chloramphenicol 3. Clinical pharmacology of systemic use in the horse. The use of chloramphenicol in the horse is now prohibited as horses are classified as food-producing animals. However, chloramphenicol has until recently been widely available for oral, intramuscular or intravenous administration. A critical appraisal of the published literature on the use of chloramphenicol in the horse clearly demonstrates that there are sound pharmacokinetic and microbiological reasons for concluding that chloramphenicol is not an appropriate antibiotic for systemic use. The short half-life of chloramphenicol in the horse, together with the broad range of minimum inhibitory...
Maturation of oocytes from normal and atretic equine ovarian follicles as affected by steroid concentrations. The ovaries of 23 mares were collected at slaughter during April-June and follicles (4-40 mm in diameter) were dissected and punctured to obtain oocytes for culture. The follicles were grouped according to histology: (a) normal, (b) showing primary and (c) secondary atresia. Antral fluid was analyzed for steroid content; oestradiol and testosterone (but not progesterone or androstenediol) were closely correlated with follicle size and histological state. Oocytes were cultured early after slaughter in Medium 199 (Difco OSI, France) or Medium B2, the highest percentage of oocytes reaching Metaph...
Direct effects of free and conjugated steroids on GnRH stimulated LH release in cultured equine anterior pituitary cells. Enzymatically dispersed anterior pituitary cells from donor mares were cultured for 48 h in alpha-modified Eagles' medium containing 10% steroid-free horse serum. The cells were then incubated for 24 h in fresh medium oestrogen followed by a 4-h incubation with or without GnRH. Media and cell extracts were analyzed for LH by radioimmunoassay. In the first series of experiments, pituitary cells from Day-3 dioestrous mares were preincubated with ethanol (control) or different concentrations of E2 (10(-11) to 10(-7) M) for 24 h prior to a 4-h incubation without (basal) or with 1.0 nM GnRH. E2 inc...
In vitro fertilization in the horse. A retrospective study. Since the first successful collection of oocytes by non-surgical puncture, there have been numerous attempts to fertilize them but few segmented embryos have resulted. The latest attempts at follicular puncture (Palmer et al., 1987) provided 159 oocytes. Oocytes found broken (18%) were probably already broken, or at least fragile, before puncture. The 41 oocytes were fertilized only with semen treated with Ionophore A23187. Following ionophore treatment of semen, 16 ova segmented (of 113 inseminated oocytes) indicating fertilization, and another 7 showed signs of fertilization but not segmenta...
In vitro steroidogenesis by granulosa cells from equine pre-ovulatory follicles. Twenty-three follicles were collected from 14 mares on specific days and grouped to represent follicles from early (Group 1; n = 6), mid (Group 2; n = 11) and late (Group 3; n = 6) oestrus, as described previously (Tucker et al., 1988). Isolated granulosa cells (GC) from each follicle were cultured in multiwell plates containing either Eagle's Minimum Essential Medium (MEM) alone, eLH (300 ng/ml), eFSH (300 ng/ml) or eLH + eFSH (300 ng/ml each), in the presence or absence of 0.5 microM testosterone. Media were collected and replaced at 24 h of culture, and 24 h later, media were again collecte...
[Local structure of cytochrome c from horse heart in solution. Conformational analysis using data of two-dimensional nuclear Overhauser effect spectroscopy]. Using the earlier suggested method the calculation of the backbone conformations of horse heart cytochrome c in oxidized (ferricytochrome c) and reduced (ferrocytochrome c) states has been performed by the two-dimensional nuclear Overhauser effect spectroscopy data. For both protein forms the secondary structure elements have been revealed and the conformations of the irregular polypeptide chain segments have been analysed. The similarity of the secondary structures of ferri- and ferrocytochrome c in solution was established from the comparison of their conformations. Small differences between...
In vitro responses of airway smooth muscle from horses with recurrent airway obstruction. The in vitro contractile and relaxant responses of tracheal smooth muscle strips (TSM) and third-generation bronchi (3B) of control horses and horses with recurrent obstructive disease (heaves) were compared. Acetylcholine (ACH) sensitivity of the diseased tissues was less than that of tissues from control horses, especially at the level of the third generation (EC50 controls 15 +/- 11 microM vs 81 +/- 17 microM for heaveys). Despite tracheal and bronchial hyporesponsiveness to ACH, these tissues from heavey horses were hyperresponsive to EFS. The inhibitory effect of isoproterenol and electri...
Trophoblastic vesicles and maternal recognition of pregnancy in mares. Research has indicated that trophoblastic vesicles (TV) formed from Day-14 equine conceptuses would prolong luteal maintenance in mares after surgical transfer to the uterus at Day 10 after ovulation. The current study assesses TV as a further model for maternal recognition of pregnancy in mares. The objectives of the study were to determine the ability of TV to prolong luteal maintenance in mares, their effect on endometrial production of prostaglandin F (PGF) in vitro, and their ability to secrete polypeptides in vitro. In contrast to our previous study (Ball et al., 1989b), transfer of TV f...
In vitro comparison of Thoroughbred and Standardbred racehorses with regard to local fatigue failure of the third metacarpal bone. In vitro local fatigue testing of the third metacarpal bone from Thoroughbred and Standardbred racehorses was performed, using fully reversed cyclic bending. Strain number of cycle data were accumulated and indicated that significant difference did not exist between the 2 breeds. It was concluded that the marked difference in the incidence of fatigue failure in the third metacarpal bone (bucked shins, dorsal metacarpal disease) seen in Thoroughbreds and Standardbreds was not related to any differences between the 2 breeds in mechanical properties of the bone.
Evaluation of measures taken by ultrasonography and caliper to estimate testicular volume and predict daily sperm output in the stallion. The semen of 26 stallions, 2-20 years of age, was collected once a day for 7 consecutive days to determine daily sperm output (DSO). After the last collection, 17 stallions were castrated. Testicular volume was estimated using two methods. Length, width, height, of each testis were measured by caliper. Length, width, height, cross-sectional area and circumference at the widest point of the testis were measured by ultrasonography. Both caliper and ultrasound measurements were first made in the live animal, and again in vitro. There were no differences in these measurements because of method or ...
Mechanical and biological effects of plate luting. Plate luting, a technique that uses polymethylmethacrylate (PMMA) interposed between the plate and the bone, as well as between the screw heads and the plate, to improve the stability of internal fixation was tested in vitro using 20 paired equine third metacarpal bones with mid-diaphyseal osteotomies plated with six-hole broad ASIF compression plates. One of each pair was luted and all specimens were tested in static overload or cyclic loading at 75% of the ultimate breaking strength using four-point bending. Although no differences were measured in static overload tests between luted and non...
Intra-follicular and peripheral steroid characteristics during vernal transition in the pony mare. This experiment investigated steroid production by ovarian tissues, in vitro, of pony mares during vernal transition from anoestrus to the breeding season. Follicular dynamics were monitored to detect the first, second, third or fourth transition follicle, greater than or equal to 30 mm diameter or the first large post luteal follicle of the breeding season. Twenty-four hours after a large follicle was detected, theca (T) and granulosa (G) tissues were harvested. Separate and co-incubations of these tissues were conducted to determine steroid production in early transition (ET), late transitio...
Use of concanavalin A for coating the membranes of stallion spermatozoa. Semen from three ejaculates from each of 4 stallions was frozen in liquid nitrogen. Morphology was evaluated by coating the spermatozoa with fluorescein-labelled Concanavalin A (FITC-ConA2) and motility was measured by computer-assisted image analysis. Coating was performed at each step of the freezing procedure (dilution, cooling, addition of glycerol and freeze-thawing) and observations were made after each step, to evaluate changes, or after subsequent steps, to determine protection provided by the coating method. All the parameters showed progressive changes during the freezing procedure. ...
Effect of prostaglandin F2 alpha on release of progesterone and leukotriene B-4 by cells from corpora lutea of mares. Corpora lutea were recovered from mares either 4 to 5 days or 12 to 13 days after ovulation. Mixed populations of luteal cells were prepared by collagenase digestion and were incubated for 24 h in the presence or absence of prostaglandin (PG) F-2 alpha (250 ng/ml). PGF-2 alpha significantly (P = 0.03) reduced progesterone secretion by cells from late diestrous corpora lutea and tended (P = 0.06) to reduce secretion by early diestrous cells. PGF-2 alpha had no significant effect on leukotriene B-4 (LTB-4) production by cells from early diestrous corpora lutea, but significantly (P = 0.03) incre...
Molecules of the early equine trophoblast. Three monoclonal antibodies raised against equine trophoblast cells were tested to determine the characteristics of the identified molecules. First, the antibodies were used to precipitate molecules from radiolabelled equine trophoblast cells of the chorionic girdle. Antibody F71.1 precipitated a molecule of 115 kDa, whereas antibodies 71.8 and 71.10 precipitated a molecule of 66 kDa. Second, 2 of the antibodies were used in an indirect immunoperoxidase assay on frozen sections of equine conceptuses of different gestational ages beginning at Day 8. Antibody F71.1 labelled trophoblast cells fro...
Lack of expression of alpha or omega interferons by the horse conceptus. Horse conceptuses were collected on Days 13, 15, 20 and 25 after ovulation. Whole conceptuses (Days 13 and 15) or extra-embryonic membranes (Days 20 and 25) were homogenized and poly-adenylated RNA (poly A RNA) was isolated by binding to oligo (dT)-cellulose. Poly A RNA (1 microgram/well) was separated by size on a denaturing 1% agarose gel and blotted onto nitrocellulose filters (northern blotting). DNA probes were prepared from plasmids containing equine alpha 1, omega 1 and omega 2 interferons and human beta actin. The presence of messenger RNA (mRNA) was detected by specific hybridization ...
Phylogeny of immune recognition: processing and presentation of structurally defined proteins in channel catfish immune responses. This work was undertaken to investigate whether or not antigen processing and presentation are important in channel catfish in vitro secondary immune responses elicited with structurally defined proteins, namely, pigeon heart cytochrome C (pCytC), hen egg lysozyme, and horse myoglobin. The use of in vitro antigen-pulsed and fixed B cells or monocytes as antigen presenting cells (APC) resulted in autologous peripheral blood leukocytes (PBL) responding with vigorous proliferation and antibody production in vitro. In addition, several long-term catfish monocyte lines have been found to function a...
Establishment of equine oviduct cell monolayers for co-culture with early equine embryos. A culture for equine oviduct epithelial cells is described. Primary cultures reached confluence in 5-8 days, forming a monolayer of polygonal cells and remaining morphologically intact for about 20 days. Subcultures were obtained by collecting cells detached spontaneously from the monolayers, and confluence was reached again after 5-7 days. Cells frozen before primary culture were confluent 10-15 days after thawing. Dishes containing confluent cells also were frozen, and some cohesive monolayers formed after thawing. Equine embryos, collected 2 days after ovulation, were cultured alone or with...
Partial purification and characterization of rhinoceros gonadotropins, growth hormone, and prolactin: comparison with the horse and sheep. The rhinoceros is an endangered species related to the horse family. Little is known of its reproductive endocrinology. The objectives of this study were to partially purify rhinoceros pituitary hormones, determine which assays could be used for their assessment, and to ascertain whether rhinoceros LH possesses the intrinsic FSH activity of equine LH. A single pituitary each from a White (1.3 g) and a Black (1.2 g) Rhinoceros was homogenized and extracted (pH 9.5), then subjected to pH and salt fractionation, and ion-exchange chromatography (DEAE and Sephadex SP-C50) to yield partially purifie...
In vitro responses of equine digital vessels to dopamine and fenoldopam. The in vitro responses of isolated vascular preparations of digital arteries and veins obtained from healthy anaesthetised horses were determined for dopamine and fenoldopam. The digital vessels were harvested, cut into 4 mm vascular segments, suspended in tissue baths and attached to force-displacement transducers. Dose-response studies between 10(-8) and 10(-4)M concentrations were performed for all drugs. The change in tension of each vascular ring was measured in grams of force. The reactivity between palmar and plantar digital vessels and baseline vascular responses were determined for do...
Changes in luteinizing hormone bioactivity associated with gonadotrophin pulses in the cycling mare. Equine plasma luteinizing hormone (LH) possesses both biological (in vitro bioassay, B) and immunological (radioimmunoassay, I) activities and the ratio of B:I varies with stage of the oestrous cycle. To estimate the contribution made by pituitary secretion and peripheral metabolism to changes in the B:I ratio, pituitary venous effluent and circulating plasma from 5 dioestrous and 2 oestrous mares were analyzed using both an in vitro bioassay and a radioimmunoassay. During dioestrus, LH was released in a pulsatile fashion with a frequency of 1.4 pulses/24 h and a pulse duration of 20-40 min (c...
Characterization of hydrophobic cores in apomyoglobin: a proton NMR spectroscopy study. A proton nuclear magnetic resonance spectroscopic study of horse apomyoglobin was undertaken in order to define the regions of myoglobin that are and that are not structurally affected by the binding of the prosthetic group. It was found that, in spite of the poor spectral resolution, a number of spin systems could be identified by using standard correlated methods. Four clusters consisting mostly of hydrophobic residues were detected by nuclear Overhauser spectroscopy, two of which involved the tryptophan side chains. Extensive similarities to nuclear Overhauser spectroscopy data collected on...
Equine infectious anemia virus derived from a molecular clone persistently infects horses. A full-length molecular clone of equine infectious anemia virus (EIAV) was isolated from a persistently infected canine fetal thymus cell line (Cf2Th). Upon transfection of equine dermis cells, the clone, designated CL22, yielded infectious EIAV particles (CL22-V) that replicated in vitro in both Cf2Th cells and an equine dermis cell strain. Horses infected with CL22-V developed an antibody response to viral proteins and possessed viral DNA in peripheral blood mononuclear cells, as determined by polymerase chain reaction assays. In addition, horses infected with CL22-V became persistently infe...
Equine granulosa-theca cell tumors express inhibin alpha- and beta A-subunit messenger ribonucleic acids and proteins. The association of equine granulosa-theca cell tumors with atrophied contralateral ovaries and abnormal estrous cycles suggests that these tumors produce hormones that affect pituitary gonadotropin production. Because inhibin, a heterodimer protein secreted by granulosa cells, decreases FSH production, we examined the presence of inhibin alpha- and beta A-subunits and their mRNAs in ovarian tumors obtained from three mares. These tumors contained neoplastic cords and nodules, multiple fluid-filled cysts, and a predominance of neoplastic granulosa cells. Reduced proteins from tumor-conditioned ...
Mechanisms of sodium and chloride transport across equine tracheal epithelium. Equine tracheal epithelium, stripped of serosal muscle, mounted in Ussing chambers, and bathed in plasmalike Ringer solution generates a serosa-positive transepithelial potential of 10-22 mV and a short-circuit current (Isc) of 70-200 microA/cm2. Mucosal amiloride (10 microM) causes a 40-60% decrease in Isc and inhibits the net transepithelial Na flux by 95%. Substitution of Cl with gluconate resulted in a 30% decrease in basal Isc. Bicarbonate substitution with 20 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid decreased the Isc by 21%. The Cl-dependent Isc was inhibited by serosal add...
Morphologic and ultrastructural evaluation of effect of ischemia and dimethyl sulfoxide on equine jejunum. Morphologic changes in equine jejunal segments subjected to 1 hour of ischemia and 1 hour of reperfusion, and protective effects of systemic administration of dimethyl sulfoxide (DMSO; 1 g/kg of body weight) were investigated in 18 ponies, using light microscopy and scanning and transmission electron microscopy. Ponies were allotted to 4 groups: group 1--control (n = 3); group 2--DMSO (n = 3); group 3--ischemia (n = 6); and group 4--ischemia and DMSO (n = 6). In each pony, 2 jejunal sections were evaluated. The first section was obtained prior to induction of ischemia, and the second was obtai...
Anatomical validation of two-dimensional echocardiography in the horse. This study was performed on 15 horses to validate the cardiac anatomy as imaged with two dimensional echocardiography (2DE) and to determine the accuracy of intracardiac measurements. 2DE images were recorded in five horses in life with a Diasonics DRF100 ultrasound system and a 3.5 MHz transducer in different tomographic planes. After slaughter, the hearts were fixed in 10 per cent formalin. To compare in vivo and in vitro results, the specimens were suspended in a water-filled tank and 2DE images were made using the same transducer positions as in life. In vitro 2DE and autopsy measurements ...
Effect of dietary alpha-linolenic acid on equine monocyte procoagulant activity and eicosanoid synthesis. To investigate the effects of an omega-3 fatty acid-enriched ration on the in vitro response of equine monocytes to endotoxin, an 8-week feeding trial was conducted in which linseed oil served as the source of the omega-3 fatty acid, alpha-linolenic acid. One group of horses was fed a control pelleted ration and the other group was fed an 8% linseed oil-enriched pelleted ration. After 8 weeks of feeding, monocytes were isolated and incubated in the presence of Escherichia coli O55:B5 endotoxin for 6 hr. After 8 weeks on the rations, the mean procoagulant activity and thromboxane B2 production ...
