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.
Dhawedkar RG, Jain NC, Mount ME, Bowling AT, Vegad JL.An enzyme-linked immunosorbent assay (ELISA) was standardised and applied for the detection of antiplatelet and antineutrophil antibodies using a heterologous system consisting of equine platelets or neutrophils and antisera raised in rabbits. The standardised technique consisted of using Immulon type 3 plate, 1 per cent gelatine as a blocking solution, poly-L-lysine buffer as a coating solution, unfixed antigen, 90 microliters test serum, horseradish peroxidase conjugated antibody and o-phenylenediamine dihydrochloride as a substrate. The number of unfixed platelets or neutrophils required fo...
Freeman DA, Butler JE, Weber JA, Geary RT, Woods GL.Oviductal and uterine embryos were collected from mares at 5 to 7 days following ovulation 1) to evaluate the effects of oviductal tissue explants on in vitro growth and development of equine embryos and 2) to study the morphologic development of equine embryos in culture. Embryos were incubated for 5 days in a medium (control group) or in medium supplemented with oviductal tissue explants (co-culture group). Embryos were evaluated and the media changed daily. Following 5 days in culture, 10 10 (100%) control embryos and 27 29 (93%) co-cultured embryos had doubled in diameter. All embryos that...
Liu DY, Lopata A, Pantke P, Baker HW.The present study demonstrates that horse and marmoset monkey sperm can bind to the human zona of salt-stored oocytes that failed to fertilize in vitro. Marmoset monkey sperm are also able to penetrate the salt-stored human zona. In contrast, human sperm do not bind to the zona of either horse or marmoset monkey oocytes. These results suggest that human sperm binding to the zona pellucida is more strictly species-specific than it is for horse and marmoset monkey sperm. In contrast, horse and marmoset monkey sperm contain receptors recognized by the human zona.
Weber JA, Freeman DA, Vanderwall DK, Woods GL.This study was conducted to identify embryonic products whose secretion was temporally associated with the oviductal transport period of the mare. Chemicals secreted by oviductal-transport-stage equine embryos were identified by incubating Day 6 or Day 7 early uterine embryos with 35S-methionine/cysteine, 3H-progesterone, or 3H-arachidonic acid for 24 h, and subsequently identifying radioactively labeled proteins (SDS-PAGE; n = 3 embryos), steroids (HPLC; n = 3 embryos), or prostaglandins (HPLC; n = 3 embryos) in the culture medium. Early uterine embryos secreted 116.1 +/- 45.5 pg of prostagla...
Obi T, Miyamoto A, Matumoto M, Ishiguro S, Nishio A.The mechanisms of histamine-induced contraction and relaxation were investigated in rings isolated from a middle part of the left descending coronary arteries of horses. Intact and endothelium-denuded preparations were compared. Rings of horse coronary arteries contracted in response to histamine in a concentration dependent manner, but some of them relaxed with lower concentrations and contracted with higher concentrations. Removal of the endothelium abolished the relaxation and potentiated the contraction. The pD2 values were 4.70 +/- 0.08 in the rings with intact endothelium and 4.95 +/- 0....
Caron JP, Eberhart SW, Nachreiner R.Articular cartilage explants from 3 horses were maintained in tissue culture to test the effects of a polysulfated glycosaminoglycan on proteoglycan biosynthesis. Cultures were exposed to concentrations of 0, 50, or 200 micrograms of the drug/ml for either 2 days or 6 days, and labeled with 35S, before measuring the content of sulfated proteoglycan in the culture media and in extracts of cartilage. In a second experiment, the explants were incubated with the isotope and subsequently exposed to the same concentrations of the polysulfated glycosaminoglycan for 4 days. Subsequently, the amount of...
Davis SW, Speer CA, Dubey JP.Asexual stages of Sarcocystis neurona were seen in cultured bovine monocytes (M617) inoculated with tissue homogenates from the spinal cord of a horse with naturally acquired protozoal myelitis. Organisms first were observed as intracytoplasmic schizonts and later as motile extracellular zoites capable of infecting surrounding M617 cells. Parasites most often occurred as clusters of merozoites dispersed throughout the host cell cytoplasm; however, schizonts also contained merozoites arranged in a radial fashion surrounding a prominent residual body. Schizonts divided by endopolygeny. The paras...
Lapan G, Awad-Masalmeh M, Hartig A, Silber R.In this study 55 strains of Taylorella equigenitalis isolated from horses of four different studs in Austria, and a comparative strain from the Federal Republic of Germany were investigated by different methods. These investigations were carried out with the help of SDS-PAGE, immunoblotting, the analyses of genomes and by proof of plasmids. Furthermore, pathogenic mechanisms such as adhesion or the formation of toxins were investigated in vitro. On the basis of the results carried out by means of SDS-PAGE and immunoblotting all tested strains of Taylorella equigenitalis were alike, whereas by ...
Rakotoarisoa L, Mironneau C, Sayet I, Mironneau J.Specific binding of the Ca++ antagonist desmethoxyverapamil, (-)-[3H]D888, to cell membranes of equine portal vein smooth muscle was inhibited in a concentration-dependent manner by guanosine 5'-O-(gamma-thio)triphosphate and ATP but was little affected by guanosine 5'-O-(beta-thio)diphosphate, noradrenaline or phorbol 12-myristate 13-acetate ester. Inhibition constants for GTP and ATP were in the range of 0.1 to 0.3 mM. From Scatchard plots and dissociation kinetic experiments, it is proposed that D888 high affinity binding sites are transferred into low affinity sites. In intact strips of ra...
Kitamoto N, Ramig RF, Matson DO, Estes MK.The production of viral antigen after infection of MA104, HepG2 (derived from human liver), and CaCo-2 (derived from human colon) cells with various cultivatable human and animal rotavirus strains was compared using immunofluorescence tests. All rotavirus strains examined expressed antigen in CaCo-2 cells and MA104 cells, but only some virus strains, namely, SA11-Cl3 (simian), RRV (simian), CU-1 (canine), and Ty1 (turkey), produced antigen in numbers of infected HepG2 cells comparable to infections in MA104 and CaCo-2 cells. Fl-14 (equine), OSU (porcine), NCDV (bovine), and Ch2 (chicken) strai...
Noiman S, Yaniv A, Tsach T, Miki T, Tronick SR, Gazit A.Nucleotide sequence analysis of a cDNA library of EIAV-infected canine cells established a complex pattern of gene expression, characterized by alternatively spliced polycistronic transcripts. The EIAV tat gene product was shown to be encoded by at least three species of mRNA which differed in their ability to trans-activate the EIAV LTR upon expression in canine cells. The most active cDNA was monocistronic, consisting of three exons. The most abundant cDNA in the library contained four exons and was identical to a polycistronic transcript previously described (Noiman et al., 1990b) which con...
Lunn DP, Holmes MA, Duffus WP.The aim of this study was to produce monoclonal antibodies (mAb) recognizing equine lymphocyte surface antigens. Fusions were conducted using BALB/c mice hyperimmunized with equine thymocytes. Hybridoma supernatants were screened by flow cytometry and positive hybridomas were cloned twice by limiting dilution. These mAb were then characterized for tissue distribution by immunohistology and flow cytometry, and by precipitation and analysis of the lymphocyte antigens which they recognized. Three mAb (CVS5, CVS4 and CVS8) are described which recognize only T lymphocytes in peripheral blood. Two-c...
Kadir FH, al-Massad FK, Fatemi SJ, Singh HK, Wilson MT, Moore GR.Reactions of reduced horse spleen ferritin with horse and Saccharomyces cerevisiae ferricytochromes c, cow ferricytochrome b5, sperm-whale metmyoglobin and Pseudomonas aeruginosa ferricytochrome c-551 were investigated by u.v.-visible spectrophotometry. In all cases the reduced ferritin reduced the ferrihaemoproteins. The rate of reduction varied from less than 0.2 M-1.s-1 for metmyoglobin to 1.1 x 10(3) M-1.s-1 for horse ferricytochrome c (0.1 M-phosphate buffer, pH 7.4, at 25 degrees C). We conclude that the mechanism of ferrihaemoprotein reduction involves long-range electron transfer throu...
May SA, Hooke RE, Lees P.Chondrocytes subjected to adverse culture conditions in vitro are stimulated to produce the eicosanoid prostaglandin E2 (PGE2) and the neutral metalloproteinase stromelysin (proteoglycanase). This indicates the potential role of the chondrocyte in cartilage degeneration in equine clinical joint disease and suggests a mechanism which may be involved in the potentiation of the effects of other inflammatory mediators. Therefore, adverse conditions within the joint, such as decreased pH in an inflammatory focus and decreased access of nutrients to deeper layers of cartilage, might contribute to th...
Redding WR, Booth LC.Equine fibroblasts and Staphylococcus aureus were exposed for 30 minutes to six dilutions of chlorhexidine gluconate, a chlorous acid-chlorine dioxide irrigation solution, a chlorous acid-chlorine dioxide disinfectant, and phosphate buffered saline controls. Cell viability was determined by trypsinizing the cells, staining them with trypan blue, and counting cells that did not take the stain. All fibroblasts were killed when exposed to 1.0% and 0.5% chlorhexidine. The survival rate of fibroblasts increased linearly with decreasing concentrations of chlorhexidine gluconate, with a peak survival...
Kot K, Tischner M.The aim of the experiment was to determine the effect of a year-to-year prolonged daylength on the patterns of equine reproductive activity and results of embryo recovery. Experiments using Konik Polski mares were conducted over four reproduction seasons. Five mares were exposed to a regimen of artificially prolonged daylength (APD) and another five mares in a control group were kept under conditions of natural daylight. Both the control and experimental groups were examined for appearance of estrus, ovulation and also for the state of their coats. A single stallion was used for breeding all o...
Fleming JT, Joshi JG.We previously showed that human brain ferritin (HBF) binds aluminum (Al) in vivo and in vitro and HBF isolated from Alzheimer's brain had more Al bound compared to aged matched controls (7). To further understand the role ferritin may play in Al neurotoxicity, we have studied in vitro the effect of Al on the function of human ferritin isolated from Alzheimer's (AD) and normal brain tissue, and compared the results with other mammalian ferritins. Al causes a concentration-dependent decrease in the initial rate of iron loading into apo-horse spleen and human brain ferritin and the rates were sim...
Grünig G, Witschi U, Winder C, Hermann M, von Fellenberg R.Supernatants of equine respiratory secretions enhanced the migration of equine neutrophils into the lower compartments of Boyden chambers. Checkerboard analysis revealed that the neutrophil migration promoting activity (NMPA) of secretion specimens was in great part caused by chemokinesis, irrespective of the neutrophil score of the specimen. The NMPA of respiratory secretions was correlated neither with the neutrophil score of the secretion specimen nor with the severity of the chronic pulmonary disease. Respiratory secretions collected while horses were kept under low dust or under dusty hou...
Horspool LJ, McKellar QA.A high performance liquid chromatography (HPLC) method was developed for the determination of seven short-chain fatty acids in equine caecal liquor. Samples were cleaned up on a Sep-pak (C18) cartridge, and the analyte was eluted from the extraction cartridge and filtered through a 0.45 micron cellulose nitrate filter. The analyte was chromatographed by ion exchange HPLC. Detection was by UV at 210 nm. Recovery from phosphate buffer (0.05 M, pH 7.0) and equine caecal liquor was 76.95% (lactic), 76.76% (valeric). The limit of (propionic), 89.35% (isobutyric), 88.73% (butyric), 80.33% (isovaleri...
Ingvast-Larsson C.A comparison between the relaxant effects of clenbuterol and theophylline on horse tracheal smooth muscle has been made in vitro. Rat tracheal smooth muscle was also investigated as a reference. The tracheal preparations were initially contracted with carbachol since the smooth muscle did not spontaneously develop tone. The response of the carbachol-contracted preparations to theophylline was the same in the two species. The response to clenbuterol varied. In only five out of eleven horses were the tracheal smooth muscles sensitive to clenbuterol (mean pD2 = 7.92 M). In the remaining six horse...
Timofeev BA, Bolotin IM, Stepanova LP, Bogdanov AA, Georgiu Kh, Malyshev SN, Petrovskiĭ , Klibanov AL, Torchilin VP.The cultures of Nuttalia eque mainly develop in the reticuloendothelial organs and so in treatment of nuttaliosis in horses and the Nuttalia carriers diamidine, an analog of imidocarb or imidozoline, was used encapsulated in liposomes. The liposomes were prepared with a modification of the phase inversion method (the lipids were dissolved in a mixture of freon-11 and chloroform). The content of the organic solvents in the preparation, as evidenced by gas liquid chromatography, was less than 0.2 per cent. The main fraction consisted of particles 1.5 to 2.5 microns in diameter. The tests on anim...
Desideri A, Stefanini S, Polizio F, Petruzzelli R, Chiancone E.Apoferritin has been selectively labeled with a maleimide nitroxide derivative at Cys-126, located in the hydrophilic 3-fold channels. Titration of this derivative with Fe(II), which gives rise to the initial Fe(III)-apoferritin complex, produces, at low metal-to-protein ratios, a decrease of the intensity of the label EPR signal due to the occurrence of a magnetic dipolar interaction. A label-metal distance ranging between 8-12 A can be estimated from titrations performed with VO(IV), which is known to bind in the 3-fold channels, and likewise produces a decrease in the label EPR signal. The ...
Fitzgerald TA, Browning GF.The sensitivity of a rotavirus serotyping enzyme-linked immunosorbent assay (ELISA) was improved by the addition of 0.5 mM CaCl2 to the washing buffer and reagent diluent. Twenty-nine of 63 (46%) previously untyped bovine and equine faecal rotavirus samples were serotyped in the modified assay. A differential response to Ca2+ ions was noted for different G-serotypes suggesting that serotyping assays performed without the inclusion of CaCl2 in the assay buffers may produce biased results.
Schaller J, Straub C, Kämpfer U, Rickli EE.The complete amino acid sequence of equine miniplasminogen (Mr 37,132, 338 residues) was determined with the aid of fragments obtained by cleavage with 2-(2-nitrophenylsulfenyl)-3-methyl-3'-bromoindolenine, cyanogen bromide or clostripain. The fragments were aligned with overlapping sequences. Sequence comparison with other species gave identities in the range of 76% (bovine) and 81% (canine), indicating the presence of the same structural and functional domains as in the other species. Sequence comparison of different miniplasminogens showed that positions 49 (Arg), 83 (Arg) and 161 (Ser) may...
MacKay RJ, King RR, Dankert JR, Reis KJ, Skelley LA.Blood monocytes and alveolar macrophages (AM) were harvested from foals (aged 46 days to 6 months) and cultured in either medium alone or medium containing 10 micrograms/ml bacterial lipopolysaccharide (LPS). After 24 h, culture supernates were collected and analyzed for cytotoxic activity on sensitized L929 cells. Both monocytes and AM that had been treated with LPS produced significantly more cytotoxic activity than the same cell type exposed to medium lacking LPS. LPS-treated macrophages secreted significantly more cytotoxic activity (120 +/- 17.8 U/ml) than did LPS-treated monocytes (47.3 ...
Padilla AW, Foote RH.Slow-cooled stallion spermatozoa, with and without seminal plasma removed by centrifugation, were diluted in Kenney's extender (KE) containing nonfat dry skim milk with glucose and antibiotics or in KE supplemented by adding a modified high-potassium Tyrode's medium (KMT). Four ejaculates from each of four stallions were collected and divided factorially across these four treatments. Percentage of motile sperm, path velocity, and linearity immediately after treatment (0 h) and after storage at 4 degrees C for 24, 48, and 72 h were evaluated objectively by use of a HTM-2030 sperm motility analy...
Stewart DR, Nevins B, Hadas E, Vandlen R.Relaxin, a polypeptide hormone normally associated with pregnancy, has been purified from many species, and the sequence determined for a growing number. Equine relaxin has been previously purified by acetone extraction, gel filtration, and ion exchange chromatographies. In an attempt to develop a more rapid and efficient method for relaxin purification, the use of affinity chromatography coupled with HPLC was explored. Monoclonal antibodies were raised against highly purified equine relaxin; large quantities of antibody were obtained by ascites production and attached to a solid phase support...
Henry MM, Moore JN.Whole blood re-calcification times were evaluated as a measure of endotoxin-associated coagulopathy in horses. First, the effects of endotoxin concentration and duration of in vitro incubation of citrated whole blood with endotoxin on the whole blood re-calcification time of blood collected from healthy horses were determined. Increasing concentrations or incubation times of endotoxin accelerated the whole blood re-calcification time. This effect was attributed mainly to increased monocyte thromboplastin activity. Second, whole blood re-calcification time, a clotting profile, plasma factor VII...
Monet-Kuntz C, Guillou F, Fontaine I, Combarnous Y.Using a suspension of seminiferous tubule cells, we had previously shown that equine FSH is superactive in the male rat, i.e. that it exhibits a higher biological potency than expected from its binding activity. In this work we investigated equine FSH superactivity in rat, pig and sheep, by comparing in each species the equine FSH with the homologous FSH, both for their binding activities (in a radioreceptor assay using a testicular membrane fraction) and for their in vitro biological potencies (in a plasminogen activator assay using a Sertoli cell-enriched population cultured on plastic). In ...
Foster SJ, Marshall DE, Houghton E, Gower DB.In on-going studies of 'classical' and ring B-unsaturated oestrogens in equine pregnancy, the products of metabolism of [2,2,4,6,6-2H(5)]-testosterone and [16,16,17-2H(3)]-5,7-androstadiene-3 beta,17 beta-diol with equine placental subcellular preparations and allantochorionic villi have been identified. Using mixtures of unlabelled and [2H]-labelled steroid substrates has allowed the unequivocal identification of metabolites by twin-ion monitoring in gas chromatography-mass spectrometry (GC-MS). Two types of incubation were used: (i) static in vitro and (ii) dynamic in vitro. The latter invol...
Olson LE, Perkowski SZ, Mason DE, Muir WW.Strips of trachealis muscle were dissected from the mid-cervical portion of the trachea from horses that were free of respiratory tract disease. The epithelium and mucosa were removed from one group of tissues and were left intact in a second group of tissues. Each tissue was suspended in a bath filled with Krebs-bicarbonate solution that was aerated with 5% CO2 in oxygen and maintained at 37 degrees C. Isometric tension was continuously recorded. The contractile response to square-wave electrical stimulations increased as frequency (3, 5, 10, 15, 20, 25, and 30 Hz), voltage (10, 15, 18, and 2...
Song X, Jiang H, Lv P, Cui K, Liu Q, Yin S, Liu H, Li Z.Oocyte in vitro maturation is necessary for the study and application of animal-assisted reproduction technology in animal reproduction and breeding. The comprehensive transcriptional profile of equine oocyte maturated in vitro has not been fully mined yet, which makes many key transcriptional events still unidentified. Here, Smart-seq2 was performed to analyse the gene expression pattern and the underlying regulatory mechanism of horse germinal vesicle (GV) and in vitro metaphase II (MII) oocytes. The results showed that 6402 genes (2640 up-regulated and 3762 down-regulated in MII samples com...
Wang Z, Yu M, Robinson NE, Broadstone RV, LeBlanc PH, Derksen FJ.The modulatory role of prostaglandin E2 (PGE2) was examined in pony tracheal smooth muscle strips. Although exogenous PGE2 inhibited the contractile response to both electrical field stimulation (EFS) and acetylcholine (ACh) in a dose-dependent manner, the concentration required to inhibit the response to EFS (10 nM) was less than that required to inhibit the response to ACh (0.1 microM). Cyclooxygenase inhibition with aspirin or meclofenamate had no effect on either the response to EFS or to ACh even though PGE2 production was inhibited. Our results demonstrate that in ponies as in other spec...
Tung JT, Venta PJ, Eberhart SW, Yuzbasiyan-Gurkan V, Alexander L, Caron JP.To determine the effects of recombinant equine interleukin -1beta (reIL-1beta) and 4 anti-inflammatory compounds on the expression and activity of cyclooxygenase (COX)-2 in cultured equine chondrocytes. Methods: Articular cartilage from 9 young adult horses. Methods: Reverse transcriptase-polymerase chain reaction methods were used to amplify a portion of equine COX-2 to prepare a cDNA probe. Northern blot analysis was used to quantify the expression of COX-2 in first-passage cultures of equine articular chondrocytes propagated in media containing dexamethasone (DEX), phenylbutazone (PBZ), pol...
Poole AW, Heath MF, Evans RJ.Pre-incubation of equine platelets in platelet-rich plasma with adenosine 5'-diphosphate (ADP) induced a reduction in aggregation responsiveness to subsequent additions of ADP. The desensitisation was shown to be homologous since the responsiveness to platelet-activating factor, thrombin, collagen, 5-hydroxytryptamine or ionomycin remained unchanged. Adenosine 5'-(beta-thio)-diphosphate (ADP beta S), a non-hydrolysable analogue of ADP, was shown to act as an agonist inducing aggregation by interaction with the ADP receptor. ADP beta S was then used in the desensitisation studies in which resid...
Johannessen MK, Skretting G, Ytrehus B, Røed KH.Endochondral bone formation is an important process in development and growth of the skeleton; still many of the mechanisms of growth cartilage remain unknown. The aim of this study was to identify genes specifically expressed in growth cartilage by constructing a subtraction cDNA library of the articular-epiphyseal cartilage complex from neonatal foal. Two hundred and eighty-four differently expressed clones, representing five novel and 37 known genes, were detected by subtraction hybridization. The tissue specificity of these genes was verified by reverse Northern analysis, and tissue distri...
Cheng FP, Fazeli AR, Voorhout WF, Tremoleda JL, Bevers MM, Colenbrander B.The aim of this study was to investigate whether mare follicular fluid (FF) induces the acrosome reaction (AR) in stallion spermatozoa and, if so, to identify the component in FF responsible for it. Furthermore, the effect of this component on sperm-zona binding and the subsequent AR was studied. Pooled FF, aspirated from the preovulatory follicles of mares in oestrous, was used and aliquots of the fluid were treated with charcoal to remove steroids (CFF). Charcoal treatment reduced the progesterone concentration in FF from 153 to < 2 ng/mL. Spermatozoa from fertile stallions collected by a...
Sampaio BFB, Nogueira BG, Souza MIL, Silva EVDCE, Zúccari CESN.The aim of this experiment was to evaluate the effects of adding ascorbic acid 2-glucoside (AA2G), a water-soluble antioxidant and stable derivative of ascorbate, to the semen extender and compare it to the addition of vitamin C (Vit. C) and the fat-soluble antioxidant α-tocopherol (α-Toh), both individually and in combination, on the seminal variables of equine sperm submitted to cooling for 72 h. We used two ejaculates from 10 stallions and evaluated them for motility, membrane integrity, chromatin fragmentation, mitochondrial activity and lipid peroxidation. In the analysis of lipid pero...
Beasley AM, Coleman GT, Kotze AC.The use of macrocyclic lactone drugs for control of equine cyathostomins is threatened by increasing levels of resistance. Detection of changes in drug sensitivity is important for effective and sustainable management of cyathostomins, however, at present such detection relies on the use of the faecal egg count reduction test, which is known to be an insensitive method. The present study therefore aimed to examine the use of a 96-well plate larval migration inhibition test for detection of resistance to macrocyclic lactone drugs in cyathostomins. We optimised conditions for migration of larvae...
Hurwitz A, Finci-Yeheskel Z, Dushnik M, Milwidsky A, Shimonovitz S, Yagel S, Adashi EY, Mayer M.This study examines the effects of interleukin-1 (IL-1) on plasminogen activator (PA) activity and prostaglandin (PG) E production in pregnant mare serum gonadotropin (PMSG)-primed granulosa cells and the potential involvement of PGE in the regulation of ovarian plasminogen activation. Methods: Granulosa cells were obtained from PMSG-primed rat (27-day-old) ovaries and cultured in serum-free conditions for 48 hours in the absence or presence of IL-1 beta (10 ng/mL) with and without transforming growth factor-beta 1 (10 ng/mL). Cellular PA activity was measured through the conversion of plasmin...
Wilsher S, Rigali F, Couto G, Camargo S, Allen WR.Historically, cryopreservation of equine embryos >300 μm gave poor pregnancy rates until researchers collapsed the blastocoele cavity and aspirated the blastocoele fluid prior to vitrification. Objective: To determine if aspiration of the blastocoele fluid prior to vitrification is essential for post warming survival. Methods: In vivo experiments. Methods: Fifty embryos were recovered on day 7-8 and washed in holding medium (HM; M-199HEPES + 20% FBS + antibiotics). Embryos were punctured using a micromanipulator mounted 30 μm biopsy needle; following this 28 had >90% of their bla...
Fitch GL, Galuppo LD, Stover SM, Willits NH.To evaluate a dorsoproximal extra-articular approach for insertion of 8.25-mm, solid-titanium, intramedullary (IM) interlocking nails into ostectomized foal third metacarpal (MC3) and third metatarsal (MT3) bones; to compare the monotonic mechanical properties of IM nail constructs with paired intact bones; and to determine the effects of age, body weight, fore- or hindlimb, and left or right limb on the mechanical testing variables. Methods: Twenty bone pairs (10 MC3, 10 MT3) collected from 10 foals of various weights and ages. Methods: One bone from each pair was randomly selected to be oste...
Yamamoto Y.NMR signals for HisB5 N(delta)H and HisEF5 N(epsilon)H protons of sperm whale and horse apomyoglobins were assigned and compared with the corresponding signals of the holoproteins in terms of pH and temperature dependence behaviors of their shifts and line widths in order to gain insight into structural difference between the apoproteins and the holoproteins. Since these protons are involved in internal hydrogen bonds at the interfaces between the B helix and the GH corner and between the EF corner and the H helix, local structures of the interfaces in these proteins have been inferred from th...
Vörös K, Holmes JR, Gibbs C.This study was designed to evaluate the accuracy of two-dimensional echocardiography (2DE) in determining the left ventricular volume (LVV) of the horse in vitro. After examining the shape of the left ventricular silicon rubber casts of four equine hearts, two modified Simpson's rule methods (Model A and Model B) as combinations of conical shapes and one biplane area-length method as a single cone (Model C) were chosen for volume calculations. One long axis and three short axis planes were used for linear and area 2DE measurements, respectively. The ventricular length (L) was calculated from t...
Suter M, Fey H.An enzyme-linked immuno sorbent assay (ELISA) for measuring horse IgE specific to ovalbumin, bencylpenicilloic acid and odinitrocarboxyphenol is described. We used a sandwich type of ELISA by which horse serum was incubated in antigen-coated tubes containing one additional polystyrene ball, followed by rabbit anti horse IgE serum. The tubes were then incubated with biotinylated goat anti rabbit globulin followed by avidin coupled to phosphatase. Endpoint titrations were compared. The ELISA is highly reproducible due to the pretreatment of the polystyrene with glutaraldehyde. The increased anti...
Parra N, Jaume M, Boscán K, Hernández A, Mijares A, González M, Alvarado Y, Restrepo J.Trypanosoma equiperdum is the causative agent of dourine, a venereal disease in horses and donkeys. This parasite has a widely distribution, is found in Africa, Asia, Southern and Eastern Europe, Russia, Mexico and Venezuela. The T. equiperdum is morphologically indistinguishable to other Trypanozoon species, however differs from other mammalian trypanosomes due to the fact that it is primarily a tissue parasite, generating cutaneous plaques, swelling of genitalia and neurological signs. The aim of this study was to evaluate the trypanocidal effectiveness of a set of derivatives of thiosemicar...
Bauquier JR, Tudor E, Bailey SR.Despite the severity and common occurrence of equine endotoxaemia, the available anti-endotoxic treatments do not effectively target key inflammatory mechanisms such as leucocyte activation and cytokine production. In this study, four compounds with potential anti-endotoxic effects, namely rolipram, azithromycin, ethyl pyruvate and metformin, were investigated in vitro using equine whole blood stimulated with bacterial lipopolysaccharide. TNF-α and IL-1β production were measured in plasma. Rolipram was the most potent inhibitor of cytokine production (IC50 0.84 and 4.68 μm for TNF-α and IL...
Zicarelli F, Tudisco R, Lotito D, Musco N, Iommelli P, Ferrara M, Calabrò S, Infascelli F, Lombardi P.Determination of digestibility represents the first step for the evaluation of the net energy content of feed for livestock animals. The aim of this study was to evaluate the in vivo digestibility and in vitro degradability of five diets characterized by different forage/concentrate ratios (F:C) in horses. The in vitro degradability was determined by the Gas Production Technique (GPT), using as an inoculum source the feces of the same subjects used for the in vivo test. Five diets consisting of poliphyte hay, straw and grains of barley and oats with a different F:C ratio [90/10 (Diet 1); 78/22...
Hornyák A, Dénes B, Szeredi L, Dencsö L, Rusvai M.Two monoclonal antibodies against the Bucyrus strain of equine arteritis virus (EAV) were produced, and according to immunoperoxidase reaction following Western blot of electrophoresed EAV structural proteins, they recognized the nucleocapsid (N) protein antigen (14-kDa protein). Besides reacting with the blotted polypeptide, the antibodies of the two clones (designated 1H1 and 4G6) selected from 576 have shown high affinity and specificity to intracellular virus antigen as well. Both antibodies reacted with the representatives of the different subtypes of equine arteritis virus providing a su...
Johnson W, Inamasu J, Yantzer B, Papangelou C, Guiot B.A soft tissue defect is often an unavoidable consequence of various surgical procedures or a result of trauma. Recently, intraoperative use of xenograft as a patch to the soft tissue defect has become popular with various products available in the market. In this study, mechanical properties of the OrthADAPTtrade mark Bioimplants (Pegasus Biologics, Irvine, CA), new xenograft products composed of collagen from equine pericardium, were evaluated individually and against an existing bioimplant product. The OrthADAPTtrade mark Bioimplants have three subtypes which differ in the degree of crosslin...
Hill AC, Polak-Vogelzang AA, Angulo AF.Acholeplasma strains were isolated from the nasopharynx of a horse (strain PN525T [T = type strain]) and the feces of a rabbit (strain B1). One clone of strain PN525T and one clone of strain B1 were examined in detail. These clones were indistinguishable from each other and were serologically distinct from the previously described Acholeplasma and Mycoplasma spp. The strains had the following properties: guanine-plus-cytosine content of 31 mol%; sterol was not required for growth, which occurred under both aerobic and anaerobic conditions; glucose was metabolized; and arginine was hydrolyzed. ...
Heath MF, Evans RJ, Poole AW, Hayes LJ, McEvoy RJ, Littler RM.The responses of equine blood platelets in citrated platelet-rich plasma to arachidonic acid, U44069 (prostaglandin endoperoxide analogue), adenosine 5'-diphosphate, platelet-activating factor or collagen were investigated by turbidimetric aggregometry. Pre-treatment of the platelets with aspirin (1 mmol/l) or paracetamol (1.3 mmol/l) abolished shape change and aggregation in response to arachidonic acid; decreased the rate of aggregation in response to collagen, with no separate effect on shape change; had no marked effect on aggregation caused by the other agonists; but in no case transforme...
Mulligan RS, Livesey JH, Evans MJ, Ellis MJ, Donald RA.It has been suggested that atrial natriuretic peptide (ANP) is the long-sought inhibitor of corticotropin (ACTH) secretion, but the evidence is conflicting. We have examined the effect of ANP and C-type natriuretic peptide (CNP) on the secretion of ACTH by perifused equine pituitary cells in an in vitro milieu intended to mimic the in vivo milieu in the horse. Corticotropin-releasing hormone (20 pM) and cortisol (0 or 100 nM) were perifused continuously and 7 pulses of arginine vasopressin (AVP; 10 nM) applied for 5 min at 30-min intervals. ANP (1 nM) or CNP (1 nM) were perifused continuously ...
Faszer K, Draper D, Green JE, Morris GJ, Grout BW.A Stirling Cycle freezer has been developed as an alternative to conventional liquid nitrogen controlled rate freezers. Horse semen samples were cooled in 0.25 ml straws and 15 ml bags in the Stirling Cycle freezer under laboratory conditions and as a portable device, powered from a car battery. For comparison, straws were frozen in a conventional liquid nitrogen controlled rate freezer. Upon thawing, motility and viability of samples frozen in the Stirling Cycle freezer were not significantly different when compared to samples frozen in the liquid nitrogen freezer. Unlike liquid nitrogen syst...
van Maanen C, Vreeswijk J, Moonen P, Brinkhof J, de Boer-Luijtze E, Terpstra C.Ten monoclonal antibodies (MAbs) were produced against equine herpes virus type 1 (EHV1). Two appeared type-specific, while the other eight were directed against epitopes common to both EHV1 and EHV4. Two MAbs directed against the glycoprotein gp2 recognized linear epitopes, as demonstrated by Western blotting. With pools of type-specific MAbs, 282 field isolates were typed in an immunoperoxidase monolayer assay (IPMA). From a total of 254 fetal or neonatal isolates, 244 (96%) were typed as EHV1, whereas 14 out of 15 (93%) respiratory tract isolates were typed as EHV4. Surprisingly, 3 out of 1...
Conforti VA, Vanderwall DK, Woods GL.The in vitro maturation (IVM) of equine oocytes is typically performed using various synthetic media; however, an optimal IVM system for equine oocytes has not been developed. The aim of the present study was to evaluate the effects of two types of follicular fluid (FF) obtained from cyclic mares and two incubation intervals for the IVM of equine cumulus-oocyte complexes (COCs). Follicular fluid was collected from medium-sized (20-29 mm diameter) and large (e30 mm; post-human chorionic gonadotrophin administration) follicles using transvaginal ultrasound-guided follicle aspiration. Compact (n ...
Ellington JE, Ball BA, Blue BJ, Wilker CE.Reliable capacitation of equine spermatozoa has been a major obstacle in the development of equine in vitro fertilization. Experiments were done to compare in vitro capacitation of equine spermatozoa by use of heparin/caffeine, calcium ionophore, uterine tube epithelial cell (UTEC)-conditioned medium, and direct culturing of spermatozoa with UTEC (coculturing). Capacitation-like changes, as determined by chlortetracycline membrane staining patterns, developed with UTEC-conditioned medium and coculturing, equivalent to that with calcium ionophore. Both of these treatments induced more (P < 0.05...
Schwarzbach SV, Melo CF, Xavier PLP, Roballo KC, Cordeiro YG, Ambrósio CE, Fukumasu H, Carregaro AB.Osteoarthritis (OA) is one of the main locomotor disorders in horses. Although nonsteroidal anti-inflammatory drugs are the first-line treatment for OA, opioids could also be used. In previous studies, opioids showed promising anti-inflammatory and analgesic effects. In this study, we aimed to investigate the effects of two opioids (morphine and methadone) against inflammation in lipopolysaccharide (LPS)-stimulated synoviocytes by analyzing microsomal prostaglandin E synthase-1 (mPGES-1) and prostaglandin-endoperoxide synthase 2 (PTGS2) expression. Synoviocytes were obtained from the joints at...
Esposito M, Astolfo A, Cipiccia S, Jones CM, Savvidis S, Ferrara JD, Endrizzi M, Dudhia J, Olivo A.Microscopic imaging of cartilage is a key tool for the study and development of treatments for osteoarthritis. When cellular and sub-cellular resolution is required, histology remains the gold standard approach, albeit limited by the lack of volumetric information as well as by processing artifacts. Cartilage imaging with the sub-cellular resolution has only been demonstrated in the synchrotron environment. Objective: To provide a proof-of-concept demonstration of the capability of a laboratory-based x-ray phase-contrast microscope to resolve sub-cellular features in a cartilage sample. Meth...
