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.
Choi YH, Chung YG, Walker SC, Westhusin ME, Hinrichs K.This study was conducted to evaluate the effects of insulin-like growth factor I (IGF-I) and other media factors during oocyte maturation, and the presence of different compositions of amino acids in embryo culture medium, on the development of equine embryos. Oocytes recovered from slaughterhouse-derived ovaries were matured in vitro for 24 h and those with a polar body were subjected to intracytoplasmic sperm injection (ICSI) or nuclear transfer with adult fibroblasts (NT). For ICSI embryos, there were no significant differences in rates of morphological cleavage, cleavage with normal nuclei...
Nebbia C, Dacasto M, Rossetto Giaccherino A, Giuliano Albo A, Carletti M.The apoprotein expression and the catalytic activities of cytochrome P450s involved in the biotransformation of xenobiotics were investigated in horse liver microsomes and compared with those of food producing (cattle, pigs, broiler chicks, and rabbits) and laboratory species (rats). Western blot analysis revealed the presence of proteins immunorelated to rat CYP 1A, CYP 2B, CYP 2E, and CYP 3A subfamilies in hepatic microsomes from horses and from any other examined species. With the exception of the N-demethylation of N-nitrosodimethylamine in broiler chicks, all the recorded interspecies dif...
Pedersen HG, Watson ED, Telfer EE.Follicular atresia has been examined previously by various biochemical and histological methods. The aim of this study was to compare, for the first time, detection of granulosa cell apoptosis by biochemical DNA analysis and microscopic examination of fresh granulosa cell morphology with the established method of detecting atresia by histology in equine follicles. DNA extracted from granulosa cells was examined by staining with ethidium bromide and end-labelling with [(32)P]dideoxy-ATP, which labels the free 3'-end of DNA fragments. In 25 of 26 follicles (96%) there was agreement between end-l...
Richardson DW, Dodge GR.To assess the effects of glucocorticoids on the expression of multiple matrix-related genes in normal and cytokine-treated cultured equine articular chondrocytes in a phenotypically correct suspension culture. Methods: Articular cartilage harvested from the joints of 15 foals, 7 yearling horses, and 16 adult horses. Methods: Glucocorticoids (dexamethasone, prednisolone, triamcinolone) at 10(-10) to 10(-4) M. Methods: Equine articular chondrocytes maintained in suspension cultures were treated with glucocorticoids with and without human recombinant interleukin 1-beta (IL1-beta) and tumor necros...
Kreusch S, Schwedler S, Tautkus B, Cumme GA, Horn A.An UV spectrophotometric method for protein determination using microplates is described. Using the SPECTRAmax PLUS reader, the UVStar 96- and 384-well microplates and a 96 or 384 parallel channel liquid handling technique, large-scale determinations can be performed with intraassay precision better than 3% CV (coefficient of variation) in the range from 1 to 8000 microg of protein/ml, measuring at 205, 215, and 280 nm and using different volume-dependent light-path lengths. Since the absorbance coefficient at 205 nm is found to be 30 ml/(mgxcm) for eight different proteins with a CV of 5.6% o...
O Morris D, Lindborg S.Sixteen healthy horses with no history of skin or respiratory disease were used for an intradermal testing (IDT) threshold study, in order to determine the concentrations of 13 commercial allergenic insect extracts most appropriate for IDT. Five dilutions of each extract were used, which included the manufacturer's recommended concentrations for equine IDT, plus one dilution higher and three lower than these standard concentrations. Allergens tested included caddisfly (Trichoptera spp.), mayfly (Ephemeroptera spp.), horsefly (Tabanus spp.), deerfly (Chrysops spp.), fire ant (Solenopsis invicta...
Chou CC, Vickroy TW.To determine the presence of adenosine receptor subtypes A1 and A2a in equine forebrain tissues and to characterize the interactions of caffeine and its metabolites with adenosine receptors in the CNS of horses. Methods: Brain tissue specimens obtained during necropsy from 5 adult male research horses. Methods: Membrane-enriched homogenates from cerebral cortex and striatum were evaluated by radioligand binding assays with the A1-selective ligand [3H]DPCPX and the A2a-selective ligand [3H]ZM241385. Functional responses to adenosine receptor agonists and antagonists were determined by a nucleot...
Quintero-Moreno A, Miró J, Teresa Rigau A, Rodríguez-Gil JE.The aim of this study was to test the presence of separate sperm subpopulations, with specific motility characteristics, in stallion ejaculates by using a computer-assisted semen motility analysis (CASA) system. Motility data were analyzed with a hierarchical clustering of variables based on a correlation or covariance matrix to select like parameters of sperm motility descriptors that better explain the kinetics of spermatozoa. The statistical analyses clustered the whole motile sperm population in both fresh and 24 h stored ejaculates into four separate groups. There were significant differe...
Seralini GE, Tomilin A, Auvray P, Nativelle-Serpentini C, Sourdaine P, Moslemi S.We characterized testicular equine aromatase and its expression. A 2707 bp cDNA was isolated, it encoded a polypeptide of 503 residues with a deduced molecular mass of 57.8 kDa. The sequence features were those of a cytochrome P450 aromatase, with a 78% polypeptide identity with the human counterpart. The gene has a minimal length of 74 kb comprising at least 9 exons and expresses a 2.8 kb mRNA in the testis. Transient cDNA transfections in E293 cells and in vitro translations in a reticulocyte lysate system allowed aromatase protein and activity detections. The activity increased with androst...
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...
Kavak A, Johannisson A, Lundeheim N, Rodriguez-Martinez H, Aidnik M, Einarsson S.Methods to evaluate the quality of frozen-thawed stallion semen are still needed, particularly those considering the sperm function. The present study evaluated sperm motility, membrane and acrosome integrity and the capacitation status of frozen-thawed spermatozoa from seven Tori and six Estonian breed stallions by way of computer assisted sperm analysis (CASA), a triple fluorophore stain combination and Merocyanine 540, respectively, the latter ones using flow cytometry. Two ejaculates from each stallion were cryopreserved using the Hannover method in 0.5 ml plastic straws. Two straws per ej...
Qureshi SH, Moza B, Yadav S, Ahmad F.The denaturation of bovine and horse cytochromes-c by weak salt denaturants (LiCl and CaCl(2)) was measured at 25 degrees C by observing changes in molar absorbance at 400 nm (Delta epsilon(400)) and circular dichroism (CD) at 222 and 409 nm. Measurements of Delta epsilon(400) and mean residue ellipticity at 409 nm ([theta](409)) gave a biphasic transition for both modes of denaturation of cytochromes-c. It has been observed that the first denaturation phase, N (native) conformation X (intermediate) conformation and the second denaturation phase, X conformation D (denatured) conformation are...
Research in microbiologyFebruary 11, 2003
Volume 154, Issue 1 29-35 doi: 10.1016/s0923-2508(02)00003-7
Silva TA, Rodrigues PH, Ribeiro RN, Noronha FS, Farias Lde M, Carvalho MA.The influence of growth medium, hemin and menadione, blood source and atmosphere of incubation on the expression of hemolytic activity of 25 strains of Prevotella intermedia and Prevotella nigrescens was evaluated. The best hemolytic activity was observed for samples of both species growing in brain heart infusion agar and incubated in Brewer-like anaerobic jars for 48 h. Hemolysis was less intense and occurred later in the presence of hemin and menadione in solid media. beta-Hemolysis was detected for medium supplemented with horse or human blood and alpha-hemolysis was observed when sheep bl...
Hu WG, Alvi AZ, Fulton RE, Suresh MR, Nagata LP.A recombinant gene encoding a single-chain variable fragment (scFv) antibody against Venezuelan equine encephalitis virus (VEE) was cloned into a prokaryotic T7 RNA polymerase-regulated expression vector. A streptavidin-binding peptide gene fused to a 6His tag was attached downstream to the scFv gene. The recombinant fusion protein was expressed in bacteria as inclusion bodies that were subsequently solubilized with 8 M urea and renatured by an arginine system. Purification of the fusion protein was achieved by immobilized metal affinity chromatography. Enzyme-linked immunosorbent assay (ELISA...
Tayyab S, Khan NJ, Khan MA, Kumar Y.Bilirubin (BR) binding properties of serum albumins from different mammalian species viz. human (HSA), equine (ESA), dog (DSA) and guinea pig (GPSA) were studied by absorption, fluorescence and CD spectroscopy. Whereas, a complex of BR with ESA produced maximum change, GPSA-BR complex showed weaker interaction as reflected from absorption and fluorescence spectroscopic data. Conformational analysis of these albumins by near- and far-UV CD spectra suggested similar structural characteristics (both secondary and tertiary structures) for ESA and HSA, whereas, DSA and GPSA had lower amounts of sec...
Neild DM, Gadella BM, Chaves MG, Miragaya MH, Colenbrander B, Agüero A.Many theories have been postulated concerning the possible effects of cryopreservation on spermatozoa, including suggestions the freeze-thawing process produces membranes that have greater fluidity and are more fusogenic, thus inducing changes similar to those of capacitation. The main objectives of this study were to determine at what stage of the freeze-thaw process membrane changes occur and whether evaluation with chlortetracycline (CTC) stain could predict the freezability of stallion sperm. Sperm viability and state of capacitation were simultaneously evaluated using CTC and Hoechst 3325...
Moussa M, Duchamp G, Mahla R, Bruyas JF, Daels PF.Equine embryos have been successfully transferred after 24h cooled storage in Ham's F-10. The aim of this study was to compare the viability of equine embryos in vitro and in vivo after 6 and 24h cooled storage using three media and to examine the relationship between embryo size and viability after 24h cooled storage. In Experiment 1, the viability of embryos was evaluated using DAPI-staining after 0, 6 or 24h in Ham's F-10, 24h in Emcare embryo holding solution (EHS) or 24h in ViGro holding plus (VHP) (n=10/group). The mean number of dead cells was similar for embryos stored in Ham's F-10, E...
Rappocciolo G, Birch J, Ellis SA.There is good evidence that cytotoxic T lymphocytes play an important role in the clearance of equine herpesvirus-1 (EHV1) in horses. We have demonstrated that, in common with other alphaherpesviruses, EHV1 infection can lead to dramatic down-regulation of MHC class I expression at the cell surface, a common strategy for pathogen evasion of the host immune response. This down-regulation is specific for MHC class I and does not reflect a general shut-off of host-cell protein synthesis. The use of monoclonal antibodies that recognize different MHC class I epitopes has demonstrated that the effec...
Dagleish MP, Brazil TJ, Scudamore CL.Lipopolysaccharide (LPS) and N-formyl-methionyl-leucyl-phenylalanine (fMLP)-like peptides are Gram-negative bacterial cell wall components which, when released into the peripheral circulation in endotoxaemia, have the potential to activate leucocytes. In vitro, equine neutrophils require priming with LPS in order to generate reactive oxygen intermediates (ROI) in response to fMLP. Objective: The aim of this study was to examine whether the release of other neutrophil products is similarly dependent on prior priming with LPS. In particular, neutrophil elastase (NE), a potent proteolytic enzyme,...
Maury W, Wright PJ, Bradley S.A novel strain of equine infectious anemia virus (EIAV) called vMA-1c that rapidly and specifically killed infected equine fibroblasts (ED cells) but not other infectible cell lines was established. This strain was generated from an avirulent, noncytopathic strain of EIAV, MA-1. Studies with this new cytolytic strain of virus have permitted us to define viral parameters associated with EIAV-induced cell killing and begin to explore the mechanism. vMA-1c infection resulted in induction of rapid cell death, enhanced fusogenic activity, and increased rates of spread in equine fibroblasts compared...
Li GP, Seidel GE, Squires EL.Five experiments were designed to study the fertilizability and development of bovine oocytes fertilized by intracytoplasmic sperm injection (ICSI) with stallion spermatozoa. Experiment 1 determined the time required for pronuclear formation after ICSI. Equine sperm head decondensation began 3 h after ICSI; 42% were decondensed 6 h after ICSI. Male pronuclei (MPN) began to form 12 h after ICSI. Female pronuclei (FPN), however, formed as early as 6 h after ICSI. In Experiment 2, ionomycin, ionomycin plus 6-dimethylaminopurine (DMAP), and thimerosal were used to activate ICSI ova. None of the IC...
Vatistas NJ, Nieto JE, Van Hoogmoed L, Gardner I, Snyder JR.To evaluate the efficacy of an isolated perfusion circuit and the effect of ischemia-reperfusion on mucosal permeability of the jejunum. Methods: In vitro study of intestinal mucosal permeability. Methods: Twelve healthy adult horses. Methods: A control segment of jejunum was placed in an isolated perfusion circuit for 240 minutes and mucosal permeability was measured. After detecting no deleterious effects of the isolated system on the control intestine, low flow ischemia was created in experimental segments for 20, 40, 60 and 90 minutes followed by 60 minutes of reperfusion and mucosal perme...
Brinsko SP, Blanchard TL, Rigby SL, Love CC, Varner DD.The aim of this study was to determine if dead spermatozoa reduced motility or membrane integrity of live spermatozoa in fresh and cooled-stored equine semen. Three ejaculates from each of three stallions were centrifuged and virtually all seminal plasma was removed. Spermatozoa were resuspended to 25 x 10(6) spermatozoa/ml with EZ-Mixin CST extender and 10% autologous seminal plasma, then divided into aliquots to which 0 (control), 10, 25, 50, or 75% (v/v) dead spermatozoa were added. Dead spermatozoa preparations contained 25 x 10(6) spermatozoa/ml and 10% seminal plasma from pooled ejaculat...
Love LB, Choi YH, Love CC, Varner DD, Hinrichs K.Two experiments were conducted to determine the effects of storage on equine ovaries or isolated oocytes. Ovaries were collected at an abattoir and were maintained at room temperature during collection and transport (3-9h total). After arrival at the laboratory, ovaries were divided into three groups: immediate oocyte collection (control), storage at room temperature overnight (15-18 h) before oocyte collection, or storage at 4 degrees C overnight before oocyte collection. Collected oocytes were cultured in maturation medium for 24h. There was a significant increase in the proportion of oocyte...
Dell'Aquila ME, Albrizio M, Maritato F, Minoia P, Hinrichs K.Follicle atresia and granulosa cell apoptosis may be related to oocyte meiotic and developmental competence. We analyzed the relationships among granulosa cell apoptosis, initial cumulus morphology, oocyte nuclear maturation in vitro, and pronucleus formation after intracytoplasmic sperm injection (ICSI) in the horse. For each follicle, the size was measured and granulosa cells were used for DNA laddering analysis. Oocytes were evaluated for cumulus morphology, cultured for in vitro maturation, and submitted to ICSI. Apoptosis was categorized as absent, intermediate, or advanced according to t...
Törneke K, Ingvast-Larsson C, Boström A, Appelgren LE.The distribution of muscarinic receptors in equine airways was investigated using autoradiography. Frozen sections of tissue from six different levels in the bronchial tree, from the trachea to the distal bronchioles, were incubated in vitro with 1.5 nmol/L of the muscarinic receptor antagonist 1-[N-methyl-3H]scopolamine methyl chloride (3H-NMS). In addition, the subtype pattern of muscarinic receptors was investigated in equine tracheal smooth muscle using radioligand binding with methoctramine, tripinamidc, 4-DAMP-methiodide and pirenzipine as competitors against the binding of 1.3 nmol/L 3H...
Benarafa C, Collins ME, Hamblin AS, Cunningham FM.The chemokine eotaxin is involved in the recruitment of eosinophils and T helper 2 lymphocytes in human allergic diseases, and drugs that block its activity, including eotaxin receptor (CCR3) antagonists, are being developed. The authors have recently cloned the horse ortholog of eotaxin and shown that it can induce equine eosinophil migration and activation in vitro. Moreover, eotaxin mRNA expression was upregulated in cultured horse dermal fibroblasts exposed to equine interleukin-4, suggesting a possible source of this eosinophil chemoattractant in equine skin. The results of this study sho...
Filho JS, Corcini CD, Santos FCC, Anciuti AN, Gatti NLS, Anastacio E, Mielke R, Nogueira CEW, Curcio BR, Varela AS. BACKGROUND: Supplementation of sperm diluents to reduce the damage caused by the freeze-thaw cycle is broadly used in equine semen cryopreservation. Objective: The present study aimed at determining the most appropriate quercetin supplementation in equine freezing extender. Methods: Quercetin at four different concentrations (0.25, 0.5, 0.75 or 1 mM) was added in the sperm freezing diluent before the freeze-thaw cycle. The spermatozoa population was analyzed by flow cytometry and a statistical analysis was conducted to detect significant differences between control and treated samples. R...
Grigolo B, Desando G, Cavallo C, Zini N, Ghisu S, Facchini A.Association of biomaterials with autologous cells can provide a new generation of implantable devices for cartilage repair. An ideal scaffold should possess a preformed three-dimensional shape, fix the cells to the damaged area and prevent their migration into the articular cavity. Furthermore, the constructs should have sufficient mechanical strength to facilitate handling in a clinical setting and stimulate the uniform spreading of cells and a phenotype re-differentiation process. The aim of this study was to verify the ability of an equine collagen membrane to support the growth of human ch...
McD￮ LA, Stover SM, Taylor KT.To determine the monotonic mechanical properties of osteotomized adult equine tibiae stabilized with two dynamic compression plates (DCP) and to compare the mechanical properties with those of intact tibiae and in vivo loads. Methods: The compressive, bending, and torsional mechanical properties of plated and intact tibiae were assessed in vitro. Methods: Twelve pairs of adult equine tibiae. Methods: Tibiae were loaded in axial compression, craniocaudal 3-point bending, or torsion in external rotation in a single cycle to failure. Mechanical properties were determined from load-displacement da...
Leid RW, Coley SC, Blanchard DP, Perryman LE.The equine alternative complement pathway has been partially characterized and compared to the equine classical activation pathway. A dose-dependent lysis of RbRBC was observed with peak lytic values noted within 10 minutes at 37 degrees C when rabbit red blood cells (RbRBC) were used as an alternative pathway activator. Sheep red blood cells (SRBC) sensitized with rabbit hemolysin or partially purified equine IgM antibodies were equally sensitive to lysis. Dilution of the commercial hemolysin by 1/5 reduced lysis from 90% to 38% in the presence of constant cell numbers. Hemolysis of SRBC peak...
Giudicelli J, Boudouard M, Delqué P, Vannier C, Sudaka P.Neutral alpha-D-glucosidase (alpha-D-glucoside glucohydrolase, EC 3.2.1.20) from horse kidney brush-border membranes was solubilized using Emulphogene BC 720 and purified by an affinity chromatography technique. The enzyme preparation (390-fold purified), which was free of other known microvillus hydrolases, exhibited one precipitate line in crossed immunoelectrophoresis and migrated as a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Several criteria (charge-shift crossed immunoelectrophoresis and hydrophobic chromatography) revealed the purified detergent form of t...
Chin WW, Maizel JV, Habener JF.The sizes of the human and subhuman alpha-subunits of the glycoprotein hormones differ by four amino acids (hCG alpha, 92 amino acids; murine, equine, bovine, and ovine alpha, 96 amino acids). The shortening of the human alpha-subunit has been attributed to posttranslational proteolysis. We have recently determined the nucleotide sequences of the mRNAs encoding the precursors of the alpha-subunit of mouse TSH and rat gonadotropins using recombinant DNA techniques. In this report, we have compared these nucleotide sequences and their deduced amino acid sequences with those of the pre- alpha-sub...
Degueurce C, Chateau H, Jerbi H, Crevier-Denoix N, Pourcelot P, Audigié F, Pasqui-Boutard V, Geiger D, Denoix JM.The proximal interphalangeal joint (PIPJ) has always been considered as a low-motion joint with an almost constant angle during loading of the limb. Until very recently, its motion was not taken into account in kinematic studies in vivo. Recent in vivo studies yielded surprisingly high ranges of motion in this joint. The aim of this study was to measure, in terms of the 3 possible rotations (flexion/extension, collateromotion and axial rotation), the movements of the PIPJ in vitro during limb loading in a neutral position (500-6000 N) and after the addition of heel and toe wedges (6 degrees an...
Biegel U, Mevissen M, Schuller S, Ruess K, Christen O, Ayrle H, Koch C, Walkenhorst M.Cancer is a common disease in humans and in companion animals and treatment is challenging. The aim of this systematic review was to identify and assess the potential use of Viscum album L. extracts (VAE) for treatment of neoplastic diseases in companion animals. Peer-reviewed animal, in vivo and in vitro studies were included, considering the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement and A Measurement Tool to Assess Systematic Reviews (AMSTAR). Overall, 6,148 references were identified. Following a predefined protocol, 114 full-text references were ...
Romagnano A, Richer CL.Cell synchronization was used to obtain an adequate percentage of very long chromosomes in equine mitotic spreads. Reported here is our variation, adapted to horse chromosomes, of a method using excess thymidine followed by bromodeoxyuridine incorporation. This technique routinely yields excellent quality cells, predominantly in prometaphase and prophase. Among other differences with the standard technique, this method does not use Colcemid, which, in addition to inhibiting spindle fiber formation, also increases chromosome contraction resulting in thicker and thus fewer bands. Consequently, h...
Patterson SD, Bell K.An affinity method was developed to investigate the interaction between protease and protease inhibitor by incorporating a protease incubation step into a two-dimensional electrophoretic separation of the plasma protease inhibitory proteins. This involved the application of the isoelectric focusing gel to filter paper saturated in the protease of choice before being placed on the second-dimensional polyacrylamide electrophoresis gel. General protein staining or immunoblotting was used to detect the protein or ligand in the complex. An in situ oxidation method was developed using the reagent ch...
Brogna R, Fan J, Sieme H, Wolkers WF, Oldenhof H.Even though dried sperm is not viable, it can be used for fertilization as long as its chromatin remains intact. In this study, we investigated drying- and temperature-induced conformational changes of nucleic acids and stallion sperm chromatin. Sperm was diluted in preservation formulations with and without sugar/albumin and subjected to convective drying at elevated temperatures on glass substrates. Accumulation of reactive oxygen species was studied during storage at different temperatures, and the sperm chromatin structure assay was used to assess DNA damage. Fourier transform infrared spe...
Álvarez C, Gil L, González N, Olaciregui M, Luño V.The rise of assisted reproduction techniques in equine medicine has fostered investigations that seek to optimize methods to increase fertility rates. Since cryopreservation continues to give low values of viability in stallions, the handling and preservation of the sperm is of vital importance. This reduction of fertility makes it essential for farmers to find new options that ensure reliability in the use of these techniques. The main objective of this study was to assess the effect of INRA 96® (manufactured commercial extender for cooling of Equine semen) as an extender for cryopreservatio...
Kisailus EC, Kabat EA.The specificity of Bandeiraea simplicifolia lectin I (BS I) has been studied by competitive-binding assays (CBA) using tritium-labeled human B and hog A substances. Blood-group B substances isolated from horse gastric mucosae and from human ovarian-cyst fluids were much better inhibitors of binding of tritiated blood-group B substance to insoluble BS I-Sepharose 2B than were human blood-group A substances from saliva and ovarian-cyst fluid. A and B active blood-group substances showed the same range of potency in inhibiting binding of tritium-labeled hog A substance to BS I-Sepharose 2B. CBA w...
Ziegler A, Olzhausen J, Hamza E, Stojiljkovic A, Stoffel MH, Garbani M, Rhyner C, Marti E.Allergen-specific immunotherapy (AIT) constitutes the only curative approach for allergy treatment. There is need for improvement of AIT in veterinary medicine, such as in horses suffering from insect bite hypersensitivity, an IgE-mediated dermatitis to Culicoides. Dendritic cell (DC)-targeting represents an efficient method to increase antigen immunogenicity. It is studied primarily for its use in improvement of cancer therapy and vaccines, but may also be useful for improving AIT efficacy. Immunomodulators, like the Toll-like receptor 4 (TLR-4) agonist monophosphoryl lipid-A (MPLA) has been ...
Miyamoto A, Kanda J, Nishio A.Transmural nerve stimulation (TNS) induced relaxations in porcine and bovine basilar arteries which were abolished by tetrodotoxin (TTX) and by L-nitro-arginine (LNAG). However, TNS induced contractions in equine basilar artery which were abolished by TTX and by guanethidine, but not by LNAG. These results suggest that the TNS-induced contractions of equine basilar arteries may be mediated by norepinephrine release.
Sissener TR, Squires EL, Clay CM.Prostaglandin F2alpha secretion by the uterine endometrium between Days 13 and 14 postovulation causes luteal regression in mares. A mechanism involving interruption or suppression of this secretion causes pregnancy to be maintained. The present study was designed to determine the age of the conceptus when maximal suppression of PGF2alpha secretion occurs. Mares were examined daily during estrus with ultrasonography (day 0 = day of ovulation). Conceptus tissues were recovered nonsurgically on Days 9 (n = 7), 12 (n = 5), 13 (n = 5), and 16 (n = 7) and uterine biopsies on Day 14. Both uterine an...
Evans MJ, Kitson NE, Alexander SL, Irvine CH, Turner JE, Perkins NR, Livesey JH.We wish to use a gonadotrophin-releasing hormone (GnRH) antagonist in the mare as a tool for investigating the control of the oestrous cycle. The aim of this study was to test the effectiveness of the antagonist cetrorelix by testing both in vitro, using perifused equine anterior pituitary cells, and in vivo in seasonally acyclic mares. Pituitary cells were prepared and after 3-4 days incubation, loaded onto columns and given four pulses of GnRH (at 0, 30, 60 and 90 min; dose-response study). After the second GnRH pulse, infusion of cetrorelix began (0, 100, 1000 and 2000 pmol/l) and continued...
Bustamante-Filho IC, Rosa AP, Van der Linden LS, Pederzolli CD, Neves AP, Dutra-Filho CS, Jobim MI, Mattos RC.The use of stallion semen collected from cauda epididymis for AI has increased due to the new protocols available for cryopreservation. Preserving the genetic material from valuable males that suffer sudden death or other events that prematurely end the stallion's reproductive life is an important strategy for Stud breeding management. While protecting spermatozoa from oxidative stress and infectious agents, the epididymis promotes the enhancement of sperm cell morphology and changes in membrane protein profile, increasing its fertility potential. The epididymal fluid must be a balanced redox ...
Ferreira HN, Ferreira-Silva JC, Rocha JM, Farras MC, Calixto M, Moura MT, Alvarenga MA, Oliveira AL.Semen cryopreservation causes DNA damage, thus requiring continuous monitoring. Objective: To compare two assays for sperm DNA fragmentation (SDF) from stallions with contrasting semen freezability. Methods: Thirteen stallions were classified as good semen freezers (GSF) or bad semen freezers (BSF). Ejaculates were cryopreserved with three diluents. Semen was subject to SDF evaluation using the sperm chromatin structure assay (SCSA) and Halomax after thawing (0 h) and after a 4 h thermoresistance test. Results: On semen of BSF, analysis by SCSA was similar between evaluations, but Halomax show...
do Nascimento AD, Marques JCC, Cezar ARR, Batista AM, Kastelic JP, Câmara DR.Although equine blastocysts ≤ 300 µm in diameter can be successfully vitrified, larger equine blastocysts are not good candidates for cryopreservation. As Na, K-ATPase is involved in maintaining blastocyst expansion, perhaps inhibition of this enzyme would be a viable method of reducing blastocyst diameter prior to cryopreservation. Objectives were to evaluate effects of ouabain-induced inhibition of Na, K-ATPase in equine blastocysts. Sixteen mares were ultrasonographically monitored, given deslorelin acetate to induce ovulation, and inseminated. Embryos (D7 and D9) were harvested and Na, ...
Ainsworth DM, Reyner CL.To examine the effects of in vitro exposure to solutions of autologous horse blood (AHB) and autologous horse serum (AHS) on expressions of selected cytokine genes in equine primary bronchial epithelial cell (BEC) cultures and to contrast these responses to those induced in BEC cultures by endotoxin and hay dust. Methods: BEC cultures established from bronchi of 6 healthy horses. Methods: 5-day-old BEC cultures were treated with PBS solution, AHB (2 concentrations), AHS, hay dust solution, and lipopolysaccharide solution for 24 hours. Gene expressions of interleukin (IL)-8, IL-1β, chemokine (...
Weiss S, Janett F, Burger D, Hässig M, Thun R.The aim of the present study was to investigate the influence of various centrifugation methods on sperm loss and quality of frozen-thawed semen. From at a total of 8 Warmblood stallions of the National Stud Farm in Avenches, 3 ejaculates each were collected and seminal plasma was removed using 3 different centrifugation regimes. In method I (reference method) centrifugation occurred by a speed of 600 x g during 10 minutes. In method II 1000 x g was used during 2 minutes while in method III centrifugation was performed by 2000 x g during 2 minutes. After centrifugation 90%, of the supernatant ...
Oldenhof H, Heutelbeck A, Blässe AK, Bollwein H, Martinsson G, Wolkers WF, Sieme H.The aim of this study was to evaluate inter-individual variability in osmotic properties of stallion spermatozoa and its correlation with cryosurvival. In addition, temperature dependency of hypo-osmotic tolerance and membrane fluidity were studied. Stallion sperm membranes exhibited good resistance towards hypotonic stress in the 15-30 °C temperature range, whereas membrane stability was found to be decreased at 4 and 37 °C. Bull spermatozoa showed greater hypo-osmotic tolerance compared with stallion spermatozoa, especially at temperatures above 30 °C, which coincided with decreased membr...
Salinas C, Barriga K, Albornoz A, Alarcon P, Quiroga J, Uberti B, Sarmiento J, Henriquez C, Ehrenfeld P, Burgos RA, Moran G.Neutrophils display an array of biological functions including the formation of neutrophil extracellular traps (NETs), web-like structures specialized in trapping, neutralizing, killing and preventing microbial dissemination within the host. However, NETs contribute to a number of inflammatory pathologies, including severe equine asthma. Tamoxifen (TX) is a selective estrogen receptor modulator which belongs to the triphenylethyllenes group of molecules, and which is used as a treatment in all stages of estrogen-positive human breast cancer. Our previous results suggest that tamoxifen can modu...
Pozzoli C, Bertini S, Poli E, Placenza G, Menozzi A.β-adrenoceptor agonists are considered the most effective drugs to counteract bronchoconstriction in horses with asthma, but only clenbuterol is commonly employed in clinical practice. We evaluated the effects of different selective β agonists: clenbuterol, ritodrine, salbutamol, and fenoterol on the contractions of isolated bronchial muscle of horses induced by electrical field stimulation (EFS), carbachol, histamine, and KCl. All β agonists reduced the amplitude of contraction induced by the different stimuli but with variable efficacy and potency. Fenoterol and salbutamol were more effec...
Morrell JM, Timoney P, Klein C, Shuck K, Campos J, Troedsson M.Several countries have adopted strategies for preventing and/or controlling equine viral arteritis based on vaccination and restricting the breeding activities of carrier stallions. However, in some cases, carrier stallions are only identified after they have transmitted virus to a mare. Therefore, a mechanism for separating virus from spermatozoa in the semen of carrier stallions would facilitate control measures for preventing disease transmission. In this study, the use of several modifications of single-layer centrifugation (SLC, SLC with an inner tube and double SLC) through Androcoll-E, ...
Seethanathan P, Bottoms GD, Schafer K.Direct effects of endotoxin (lipopolysaccharide [LPS]) on equine WBC are known to stimulate the release of a variety of mediators including thromboxane, prostacyclin, and leukotrienes. In this study, 0.1 microgram of LPS/ml stimulated an early increase in tumor necrosis factor, succeeded by an increase in interleukin-1, but concentrations of LPS up to 5.0 micrograms/ml caused no significant increase in superoxide anion release. The concentration of LPS (0.1 microgram/ml) used in this experiment was in the range of concentrations measured in plasma of some horses with gastrointestinal problems....
de Grauw JC.Diseases affecting synovial joints are a major cause of chronic disability both in humans and in companion animal species, most notably dogs and horses. As progressive deterioration of the articular cartilage is the hallmark of degenerative joint disease or osteoarthritis, research efforts traditionally tended to focus primarily on cartilage pathology. However, in recent years it has become clear that synovial joints should be considered intricate organs in their own right, with each of the constituent tissues (cartilage, bone, and synovial membrane) interacting with each other both in health ...
Davis JL, Marshall JF, Papich MG, Blikslager AT, Campbell NB.The purpose of this study was to determine the pharmacokinetics of deracoxib following oral administration to horses. In addition, in vitro equine whole blood cyclooxygenase (COX) selectivity assays were performed. Six healthy adult horses were administered deracoxib (2 mg/kg) orally. Plasma samples were collected prior to drug administration (time 0), and 10, 20, 40 min and 1, 1.5, 2, 4, 6, 8, 12, 24, and 48 h after administration for analysis with high pressure liquid chromatography using ultraviolet detection. Following PO administration, deracoxib had a long elimination half-life (t(...
Piuko K, Bravo IG, Müller M.In the present study we describe the isolation and characterization of putative equine granzyme B for which we propose the designation 'eqGrzmB'. Sequence analysis revealed characteristic features of a GrzmB protease such as the presence of a signal (leader-) peptide and an activation di-peptide. The isolated eqGrzmB is functionally active when expressed in human or in insect cells. Furthermore, exchange of any of three putative active site amino acids, which are highly conserved along granzyme B enzymes, led to a complete loss of enzymatic activity in the newly identified eqGrzmB. Phylogeneti...
