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.
Roth TL, White KL, Thompson DL, Horohov DW.It has been proposed that PGE2 is an important immunosuppressant acting at the fetal-maternal interface during pregnancy. We have previously shown that horse conceptus-conditioned medium suppresses lymphocyte proliferation. This experiment was designed to determine if horse conceptus-derived immunosuppressive activity could be attributed to PGE2 production by the trophoblast tissue. Trophoblast tissue from 21-day-old conceptuses was cut into equal sections and cultured in the presence or absence of the prostaglandin inhibitor, indomethacin. Following culture, immunosuppressive activity and the...
Weber JA, Woods GL, Freeman DA, Vanderwall DK.Prostaglandin E2 (PGE2) secreted by Day-6, Day-7, Day-8 and Day-9 equine embryos (ovulation = Day 0) during in vitro incubation was measured by radioimmunoassay. Embryonic PGE2 secretion (ng/embryo/24 hr) was detectable on Day 6 (0.27 +/- 0.39), tended to increase (P less than 0.1) on Day 7 (0.57 +/- 0.88), and increased significantly (P less than 0.05) on Day 8 (2.23 +/- 0.86) and Day 9 (4.13 +/- 0.71). Embryo diameter at the start of the incubation period was linearly correlated (P less than 0.01) to embryonic PGE2 secretion.
Niederkorn JY, Ubelaker JE, McCulley JP, Stewart GL, Meyer DR, Mellon JA, Silvany RE, He YG, Pidherney M, Martin JH.A crucial requirement for establishing corneal infection by the extracellular protozoal parasite, Acanthamoeba, is the ability of the parasite to bind to the corneal surface. In a series of in vitro studies, we examined the ability of Acanthamoeba castellanii [corrected] to adhere, invade, and damage normal, intact corneas of 11 mammalian and one avian species. A. castellanii [corrected] (80-90% trophozoites and 10-20% cysts) were incubated with corneas for 24 hours in vitro and examined by scanning electron microscopy (SEM). Results of several independent SEM experiments revealed that parasit...
Rønne M.After fluorouracil/5-bromodeoxyuridine synchronization and subsequent FPG-staining, the karyotype of 15 phenotypically normal horses displayed several breaks and gaps. Twelve bands 1q24, 4p12, 8q23, 11p12, 16q21, 17q21, 23q31, 23q32, Xp21, Xq22, Xq25 and Xq27 showed relatively frequent fragility. After thymidine/cytidine synchronization and subsequent GWL-banding the same horses display karyotypes without any fragility. Hence it is suggested that the above listed bands harbour folate and/or 5-bromodeoxyuridine sensitive fragile sites.
Lecompte F, Combarnous Y.A simple, accurate, sensitive enzyme immunoassay (EIA) has been developed that permits the measurement of equine Chorionic Gonadotropin activity in pregnant mare plasmas or serums as well as in commercial and highly-purified preparations. This assay is specific for eCG and eLH which share the same polypeptide structure but differ in their oligosaccharidic chains. The more important result is that this EIA has been found to be give data in very close agreement with the in vivo assay. Therefore this very rapid and convenient assay can be used to measure the activity of eCG/PMSG in pregnant mares...
Gentry PA, Feldman BF, O'Neill SL.The purpose of this study was to evaluate the effect of modifying commercial reagents for the laboratory evaluation of several haemostatic parameters in normal, non-pregnant mares. The routine coagulation screening assays, namely, the activated partial thromboplastin time (APTT) and the one-stage prothrombin time (PT), and the specific coagulation assays for the determination of the biological activity of Factors VII, VIII:C and IX, are discussed.
Lear TL, Trembicki KA, Ennis RB.Giemsa-11 (G-11) staining and in situ hybridization were used to identify the equine chromosome complement of horse x mouse somatic cell hybrids. The presence of horse chromosomes in somatic cell hybrids was determined by differential G-11 staining. The slides were then destained and hybridized with biotinylated total horse (Equus caballus) genomic DNA without suppression. Fluorescence detection permitted rapid confirmation of horse chromosomal DNA in the hybrid cells.
Gray PR, Derksen FJ, Robinson NE, Slocombe RF, Peters-Golden ML.We have developed an alternative method for examining equine tracheal epithelial arachidonic acid (AA) metabolism that utilizes strips of pseudostratified columnar epithelium attached to a layer of elastic tissue 80 to 130 microns thick. We compared the responses of this preparation with those of enzymatically dispersed suspensions of tracheal epithelium obtained from the same animal. Strips incubated with [3H]AA incorporated 40.8 +/- 3.6% of added radioactivity and released 2.55 +/- 0.23% of incorporated radioactivity when stimulated with 5 microM A23187. Values for the cell suspension were 5...
Hassanain MM.Investigation studies on inactivated African horsesickness vaccine using binary ethyleneimine were conducted. The inactivation process of virulent type-9 strain using the above inactivant revealed complete virus inactivation at 18, 48 and 84 h post-treatment with inactivant concentrations of 0.004, 0.003 and 0.002M, respectively, without detection of residual virus. An inactivant concentration of 0.003M is recommended and no changes in viral antigenic properties were noticed in complement fixation test. The physical parameters in oil-emulsion vaccine using the incomplete Freund's adjuvant, wer...
Ellegren H, Andersson L, Johansson M, Sandberg K.A synthetic polynucleotide (TG)n was hybridized to equine DNA digested with HinfI and hypervariable hybridization patterns were obtained. Mendelian inheritance of these DNA fingerprinting patterns was confirmed by pedigree analysis. Estimates of the probabilities of identical band patterns in unrelated individuals of different breeds (Swedish Trotters, North Swedish Trotters, Thoroughbreds and Arabians) were in the range 1 x 10(-4) - 7 x 10(-6). The variability derived with the (TG)n probe in horses was higher than what we obtained with several other commonly used probes for DNA fingerprinting...
Ericson KK, Yang TJ.During our studies of cytostatic cytokines in the mixed leukocyte culture, we found that horse serum in the medium control contained a tumor cell growth-inhibitory factor. The fraction isolated by molecular sieving and ion exchange chromatography inhibited the growth and DNA synthesis of the primary culture and passaged cell line of the canine transmissible venereal sarcoma, murine T (L5178Y) and B (P3-X63-Ag8.653) lymphoid tumor cells, murine mammary tumor cells (RIII), bovine lymphoid tumor cells (BL3), and the nontransformed cell line of baby hamster kidney cells. Nontransformed cell lines ...
Buckingham SH, Jeffcott LB, Anderson GA, McCartney RN.The in vivo precision of ultrasound velocity measurement and single photon absorptiometry for the assessment of equine bone quality is discussed. In vivo precisions for ultrasound velocity measurements were less than 0.5 per cent, whereas cortical cross-sectional area, compact bone density and modulus of elasticity were around 1 per cent, and bone mineral content and density were just over 2 per cent. Except for ultrasound velocity, substantial improvements could be achieved by taking the mean of five readings for each measurement. The long-term precision of the techniques was also high, with ...
Rashmir-Raven AM, Coyne CP, Fenwick BW, Gaughan EM, Andrews GA, DeBowes RM.The ability of polysulfated glycosaminoglycans (PSGAG) to inhibit the complement cascade was evaluated. The role of complement in inflammation and infection has been well documented. Inhibition of the complement cascade by PSGAG could explain why intra-articularly administered PSGAG diminish diarthrodial joint inflammation and potentiate septic arthritis in horses. Hemolytic complement testing was performed to evaluate the effect of PSGAG on the equine classical and alternate pathways of complement, using rabbit erythrocytes as the target cells. Concentration of PSGAG between 0.2 mg/ml and 0.6...
Fletcher JE, Tripolitis L, Beech J.The modulation of Ca2+ release by synthetic bee venom melittin was examined in equine and human terminal cisternae-containing fractions. Melittin (0.1 microM) decreased the threshold of Ca(2+)-induced Ca2+ release by 20% in equine muscle and by 36% in human muscle. If terminal cisternae fractions were first preloaded with Ca2+ to greater than about 75% of the threshold of Ca(2+)-induced Ca2+ release and then melittin added, an immediate and sustained release of Ca2+ occurred in preparations from both species. Addition of melittin after a Ca2+ preload of < 50% of the threshold of Ca(2+)-induced...
Borovanský J, Vedralová E, Hach P.Concentration of melanosomes in various tissues has been unknown because of the impracticability of their direct quantification. Using an indirect approach comprising the estimation of melanin both in freeze-dried tissue samples and in isolated melanosomes, we obtained data on the amount of melanosomes in various pigment tissues. The concentrations of melanosomes found in the tissues were relatively high, not only reflecting the dark color of pigment tissues but also explaining their capacity to perform various functions ascribed to the presence of melanin.
Willis P, Caudle AB, Fayrer-Hosken RA.Objectives of the present research were to determine the influences of types of media, sera, time and hormones on equine oocyte in vitro maturation (IVM). The following types of media and sera were evaluated: Menezo's B2 medium (B2), modified Tissue Culture Medium 199 (TCM), Defined Medium (DM), fetal calf serum (FCS), mare serum collected on the first day of estrus (MS), and mare serum collected on the day of ovulation (MSO). Resultant oocyte maturation was compared with the control: DM with bovine serum albumin (BSA). Effect of culture time (0, 15, and 32 hr) and the following hormones on oo...
Derse D, Carvalho M, Carroll R, Peterlin BM.Transcriptional regulatory mechanisms found in lentiviruses employ RNA enhancer elements called trans-activation responsive (TAR) elements. These nascent RNA stem-loops are cis-acting targets of virally encoded Tat effectors. Interactions between Tat and TAR increase the processivity of transcription complexes and lead to efficient copying of viral genomes. To study essential elements of this trans activation, peptide motifs from Tats of two distantly related lentiviruses, equine infectious anemia virus (EIAV) and human immunodeficiency virus type 1 (HIV-1), were fused to the coat protein of b...
Ekfalck A, Jones B, Obel N.After a survey of the state of laminitis research the authors conclude that none of the present concepts of the pathogenesis of laminitis unequivocally explains the basic clinical and morphological observations in this disease. There is therefore reason to consider the advances that have been made during the last decades in respect to the influence of various substances on the differentiation of cultured skin keratinocytes. The technique is available for studying hoof keratinocytes in a comparable way. Relevant literature on cultured skin keratinocytes is surveyed. Some of the results from exp...
Johnston JK, Freeman DE, Gillette D, Soma LR.Sheets of mucosa from the jejunum of healthy horses were mounted in incubation chambers and bathed with Krebs-Ringer bicarbonate solution. Changes in tissue function and histologic appearance were compared after the following conditions: (1) control conditions for 30 minutes with 95% O2/5% CO2 in the gas phase; (2) same conditions as control, except incubation with superoxide dismutase (300 U/ml) during the last 18 minutes; (3) anoxia for 15 minutes with 95% N2/5% CO2, followed by reoxygenation for 15 minutes; (4) same conditions as 3, except incubation with superoxide dismutase during reoxyge...
Su XZ, Morris DD, McGraw RA.We report the molecular cloning and nucleotide sequence of the equine gene encoding tumor necrosis factor alpha. The 2610-bp genomic sequence was derived from three overlapping polymerase chain reaction products.
Livesey JH, Carne A, Irvine CH, Ellis J, Evans MJ, Smith R, Donald RA.A 41 amino acid peptide, probably identical in structure to human corticotropin releasing factor, was isolated from 70 equine hypothalami by methanol extraction, immunoaffinity chromatography and single step of reverse phase HPLC. The amino acid sequence was determined by gas phase sequence analysis. Probable carboxyl terminal amidation was demonstrated by similar retention times for equine and human corticotropin releasing factor on reverse phase HPLC at pH 8. The likely structure of equine corticotropin releasing factor is: Ser-Glu-Glu-Pro-Pro- Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu...
Grünig G, Winder C, Hulliger C, Witschi U, Hermann M, Jungi TW, von Fellenberg R.Increases in procoagulant activities (PCA) in equine lung macrophages were induced by non-adherent blood lymphocytes which were prestimulated with phytohaemagglutinin for 48 to 72 hours or by supernatants harvested from prestimulated blood lymphocyte cultures. However, prestimulated lymphocyte suspensions themselves expressed PCA which was most probably derived from contaminating monocytes. Because non-adherent cells from lymphocyte suspensions may have attached to adherent macrophages, cells within lymphocyte suspensions might have contributed to the PCAs expressed by lymphocyte-stimulated lu...
Cochran R, Meintjes M, Reggio B, Hylan D, Carter J, Pinto C, Paccamonti D, Graff KJ, Godke RA.In vitro fertilization in horses has been less successful than anticipated owing to: (i) the inability to collect large numbers of good quality oocytes; (ii) alterations in the zona pellucida that occur during in vitro maturation of equine oocytes; and (iii) inadequate preparation of equine sperm cells. In addition, studies in humans, mice and cattle have indicated that high concentrations of glucose in culture media may inhibit embryonic development in vitro and this may also be a problem for development of equine embryos in vitro. The aims of the present study were: (i) to achieve fertilizat...
Sod GA, Mitchell CF, Hubert JD, Martin GS, Gill MS.To compare in vitro monotonic biomechanical properties of an equine spoon plate (ESP) with an axial 3-hole, 4.5 mm narrow dynamic compression plate (DCP) using 5.5 mm cortical screws in conjunction with 2 abaxial transarticular 5.5 mm cortical screws (DCP-TLS) inserted in lag fashion for equine proximal interphalangeal (PIP) joint arthrodesis. Methods: Paired in vitro biomechanical testing of 2 methods of stabilizing cadaveric adult equine forelimb PIP joints. Methods: Cadaveric adult equine forelimbs (n=18 pairs). Methods: For each forelimb pair, 1 PIP joint was stabilized with an ESP (8 hole...
Álvarez C, González N, Luño V, Gil L.The aim of this study was to evaluate the effect of trehalose and lactose extenders on ejaculated and epididymal stallion sperm vitrification. Ejaculated semen samples were collected from seven fertile stallions, and cauda epididymis samples were collected from ten stallion carcasses after slaughter. Both the ejaculated and the epididymis samples were diluted and vitrified using INRA 96® and bovine serum albumin as well as trehalose or lactose. As a control, ejaculated and epididymal samples were collected and frozen using the conventional method. Vitrification was performed by immersing sper...
Black JB, Frampton AR.Equine herpesvirus type 1 (EHV-1) is a highly transmissible pathogen that leads to a variety of clinical disease outcomes in infected horses. A major sequela that can occur after an EHV-1 infection is a neurological disease termed equine herpesvirus myeloencephalopathy (EHM). Clinical manifestations of EHM include fever, ataxia, incontinence, and partial to full paralysis, which may ultimately lead to the euthanization of the infected horse. To develop an effective treatment strategy for EHM, it is critical that the specific virus-host interactions that lead to EHM be investigated so that safe...
Udén P, Van Soest PJ.Fiber fermentation using the in situ bag technique was studied in a hay-fed cow. Entry of fine particles into bags of varying pore size, the effect of sample size, rumen contractions, bag porosity and rumen contraction (bags suspended in vitro or in situ) and obstruction of liquid flow through the bag cloth were investigated (Exp. 1). In Exp. 2 fiber degradation in vitro and in situ with 5- and 37-micron pore size bags was measured utilizing six fistulated heifers (four large: 610 kg and two small: 243 kg), two sheep and two goats (30 kg), three ponies (130 kg) and four rabbits (3.2 kg). Degra...
Potempa J.The proteolytic activity of the elastase-like proteinase from granules of horse blood leukocytes is retained on a column of Cibacron Blue-Sepharose and can be eluted with 0.5 M KSCN. During this procedure its mol. wt. is reduced from 49000 to 30000 and isoelectric point is shifted towards higher pH. The inactive protein not adsorbed on Cibacron Blue-Sepharose is strongly acidic and shows a mol. wt. of 20000. Upon mixing this protein with the modified enzyme the native proteinase is reconstituted as shown by polyacrylamide gel electrophoresis at pH 8.3 and isoelectric focusing in a sucrose grad...
Hesford F, Lazary S, Curty-Hänni K, Gerber H.A number of horse alloantisera were characterized biochemically as being directed against MHC class I or class II antigens by immunoprecipitation of the corresponding antigens from lysates of biosynthetically radioactively labelled lymphocytes and determination of their molecular weights by SDS-PAGE and fluorography. Sera recognizing A2 and A3 specificities precipitated antigens of 44,000 Daltons molecular weight (class I heavy chain), whereas sera with specificities W13, W22 and W23 precipitated antigens corresponding to class II dimers (30,000 and 32,000 Daltons). Comparison with antigens pr...
Ijpma G, Liang CQ, Kachmar L, Panariti A, Benedetti A, Lavoie JP, Lauzon AM.Isolated human airway smooth muscle (ASM) tissue contractility studies are essential for understanding the role of ASM in respiratory disease, but limited availability and cost render storage options necessary for optimal use. However, to our knowledge, no comprehensive study of cryopreservation protocols for isolated ASM has been performed to date. We tested several cryostorage protocols on equine trachealis ASM using different cryostorage media [1.8 M dimethyl sulfoxide and fetal bovine serum (FBS) or Krebs-Henseleit (KH)] and different degrees of dissection (with or without epithelium and c...
Mugnier S, Boittin S, Douet C, Monget P, Magistrini M, Goudet G.In human and rodents, sperm-zona pellucida binding is mediated by a sperm surface Galactosyltransferase that recognizes N-Acetylglucosamine residues on a glycoprotein ZPC. In large domestic mammals, the role of these molecules remains unclear: in bovine, they are involved in sperm-zona pellucida binding, whereas in porcine, they are not necessary. Our aim was to clarify the role of Galactosyltransferase and N-Acetylglucosamine residues in sperm-zona pellucida binding in ungulates. For this purpose, we analyzed the mechanism of sperm-zona pellucida interaction in a third ungulate: the horse, si...
Gimeno BF, Bariani MV, Laiz-Quiroga L, Martínez-León E, Von-Meyeren M, Rey O, Mutto AÁ, Osycka-Salut CE.Cryopreservation by negatively affecting sperm quality decreases the efficiency of assisted reproduction techniques (ARTs). Thus, we first evaluated sperm motility at different conditions for the manipulation of equine cryopreserved spermatozoa. Higher motility was observed when spermatozoa were incubated for 30 min at 30 × 106/mL compared to lower concentrations (p < 0.05) and when a short centrifugation at 200× g was performed (p < 0.05). Moreover, because sperm suitable for oocyte fertilization is released from oviduct epithelial cells (OECs), in response to the capacitation process, we e...
Frisbie DD, Sandler EA, Trotter GW, McIlwraith CW.To determine response of interleukin-1alpha (IL-1alpha)-conditioned equine articular cartilage explants to insulin-like growth factor-1 (IGF-1). Sample Population-Cartilage from the trochlea and condyles of the femur of a clinically normal 4-year-old horse. Methods: Effects of IGF-1 (0 to 500 ng/ml) after addition of IL-1alpha were evaluated by assessing matrix responses, using a sulfated glycosaminoglycan (GAG) assay, matrix 35SO4 GAG incorporation, and release of GAG. Mitogenic response was assessed by 3H-thymidine incorporation into DNA and fluorometric assay of total DNA concentration. Res...
Asmis R, Jörg A.Horse eosinophils preincubated with 3H-labelled acetate and stimulated with the Ca2+ ionophores ionomycin or A23187 form a radioactive compound, which we have shown to be 1-O-alkyl-2-[3H]acetyl-sn-glycero-3-phosphocholine (platelet-activating factor). We could detect no 1-O-acyl-2-[3H]acetyl-glycero-3-phosphocholine in the radioactive fraction. The formation of platelet-activating factor was strongly correlated to the generation of leukotriene C4, the main arachidonate metabolite in horse eosinophils, suggesting that platelet-activating factor and leukotriene C4 have a common precursor pool (1...
Montelaro RC, West M, Ivey M.Representative glycoproteins including fetuin, protein A, ovalbumin, alpha 1 acid glycoprotein, and the major glycoprotein of equine infectious anemia virus were labelled with 125I by the chloramine-T or Bolton-Hunter procedure and their binding to immobilized Con A or lentil lectin compared to untreated samples of each glycoprotein. Glycoprotein modification was no greater than one substituted residue per protein molecule. Yet the radioiodinated glycoproteins typically displayed only 0-50% of the lectin binding observed with untreated samples. These results indicate that lectin glycoprotein b...
Chung C, Wilson C, Timoney P, Adams E, Adams DS, Chung JS, Evermann JF, Shuck K, Lee SS, McGuire TC.Equine arteritis virus (EAV) causes contagious equine viral arteritis, characterized by fever, anorexia, conjunctivitis, nasal discharge, dependent edema, abortion, infrequent death in foals, and establishment of the carrier state in stallions. The World Organization for Animal Health (OIE) defines a horse as seropositive if the serum neutralization (SN) antibody titer is ≥1:4 to EAV. However, determining the SN titer is time-consuming and requires specific laboratory facilities, equipment, and technical expertise to perform. Furthermore, interpretation of the SN titer of some sera can be di...
Arosio P, Gatti G, Bolognesi M.Various precipitating agents were examined in order to crystallize horse heart and spleen ferritins. Cadmium sulfate induced the crystallization of the spleen ferritin, while 2-methyl-2,4-pentanediol and poly(ethylene glycol) only induced that of the heart ferritin. Isoelectric focusing analysis showed that the crystals grown from cadmium sulfate contained only the more acidic isoferritins, and those grown from methyl pentanediol only the less acidic isoferritins. Heart ferritin crystallizes in a cubic space group, as previously reported for spleen ferritin crystals grown from cadmium sulfate....
Gurusinghe S, Hilbert B, Trope G, Wang L, Bandara N, Strappe P.Immortalization of chondrocytes enables long term in vitro culture; however, the chondrogenic capacity of transformed cells varies, thus highlighting the need to develop a proliferative and tuneable chondrocyte cell line where hypertrophic differentiation can be controlled. In this study the SV40 large T antigen and human telomerase reverse transcriptase were employed to immortalize pooled equine chondrocytes through lentiviral vector mediated transduction either singly or on combination. Transformed chondrocytes proliferated stably over multiple passages, but resulted in significantly lower e...
Mason JB, Gurda BL, Van Wettere A, Engiles JB, Wilson JM, Richardson DW.Our long-term aim is to develop a gene therapy approach for the prevention of laminitis in the contralateral foot of horses with major musculoskeletal injuries and non-weightbearing lameness. Objective: The goal of this study was to develop a practical method to efficiently deliver therapeutic proteins deep within the equine foot. Methods: Randomised in vivo experiment. Methods: We used recombinant adeno-associated viral vectors (rAAVs) to deliver marker genes using regional limb perfusion through the palmar digital artery of the horse. Results: Vector serotypes rAAV2/1, 2/8 and 2/9 all succes...
García-Rubio I, Fittipaldi M, Trandafir F, Van Doorslaer S.In this work, we show the extreme power of multifrequency HYSCORE (hyperfine sublevel correlation spectroscopy) techniques to unravel the hyperfine interactions of the electron spin with the remote nuclei in the heme site of high-spin ferric heme proteins. Horse heart aquo-metmyoglobin was used as a model system to demonstrate the power of these techniques. Experimental evidence was collected and assigned to protons of the proximal histidine ligand, to the mesoprotons of the heme ligand, and to two different protons of the distal water ligand. The latter difference relates to the stabilization...
Rojas G, Vargas M, Robles A, Gutiérrez JM.Twenty batches of polyvalent antivenom produced at the Instituto Clodomiro Picado were analyzed for turbidity, both before and after freezing-thawing and lyophilization. Eight batches became turbid upon freezing-thawing, and this change correlated with high levels of cholesterol, triglycerides and lipoproteins, especially beta-lipoprotein. Since normal horse serum does not become turbid after freezing-thawing, despite the fact that it has high lipoprotein levels, the possibility was raised that phenol, used as a preservative during serum fractionation, might affect lipoproteins, inducing the a...
Allen GP, Cohen JC, Randall CC, O'Callaghan DJ.The replication of equine herpesvirus type 1 (EHV-1) and type 3 (EHV-3) was unimpeded in three different cell types-equine epithelial cells, equine fibroblasts, and mouse fibroblasts-which had been blocked in their capacity to synthesize host DNA by 2.5 mM hydroxyurea (HU) or 2 mM thymidine (TdR). The rate of DNA synthesis in uninfected or equine herpesvirus-infected cells in the presence of HU or TdR was measured by pulse-labeling cell samples with a labeled DNA precursor at different times after infection. DNA synthesis in uninfected cultures was completely inhibited by both compounds. Howev...
Baxter GM, Tackett RL, Moore JN.Palmar digital arteries and veins removed surgically from healthy horses under general anesthesia were cut into 4 mm vascular rings, suspended in tissue baths, and attached to force displacement transducers for continuous measurement of vascular tension. In vitro vascular responses were determined for acetylcholine, acepromazine, isoxsuprine hydrochloride (isoxsuprine), prostaglandin E2 (PGE2), and prostaglandin I2 (prostacyclin). After preconstriction with norepinephrine hydrochloride (norepinephrine), or prostaglandin F2 alpha (PGF2 alpha), the concentrations needed to produce 50% maximum re...
Balke E, Weiss R, Seipp A.A total of 110 strains of beta-hemolytic streptococci, belonging to serogroup C (Lancefield), isolated from horses (71 S. zooepidemicus, 27 S. equisimilis and 12 S. equi) as well as 5 reference strains were tested for their ability to produce hyaluronidase. The determinations were carried out in a culture test on agarose gel and in a liquid test system (turbidity test according to DiFerrante). The results of both methods used showed that the three Streptococcus species could be differentiated by the relative quantitative determination of hyaluronidase activity. S. equisimilis strains produce 5...
Balkman CE, Nixon AJ.To clone and determine molecular structure of equine stromelysin 1 (matrix metalloproteinase 3) and examine stromelysin expression in articular cartilage. SAMPLES AND PROCEDURE: Total RNA was harvested from equine arthritic cartilage specimens and was used for reverse transcription and polymerase chain reaction amplification to develop overlapping complementary DNA (cDNA) clones. Four cDNA sequences were ligated into plasmid (pGEM3Z) constructs and subcloned into bacterial expression vectors, and sequence was determined by automated dye terminator sequencing. Stromelysin mRNA expression was as...
Roels K, Smits K, Ververs C, Govaere J, D'Herde K, Van Soom A.In horse breeding, intracytoplasmic sperm injection (ICSI) has gained interest to obtain offspring from subfertile individuals. This paper presents a case report of a stallion with severe testicular degeneration. Semen analysis showed very low motility and 83.5% of detached heads. Histology of a testicular biopsy showed severely decreased spermatogenesis, while transmission electron microscopy of the sperm cells revealed no significant abnormalities. A total of 39 oocytes were fertilized by ICSI with frozen-thawed spermatozoa of this stallion: 25 oocytes with intact spermatozoa and 24 with det...
Kerfelec B, Foglizzo E, Bonicel J, Bougis PE, Chapus C.The complete sequence of the horse pancreatic lipase was elucidated by combining polypeptide chain and cDNA sequencing. Among the structural features of horse lipase, it is worth mentioning that Lys373 is not conserved. This residue, which is present in human, porcine and canine lipases, has been assumed to be involved in p-nitrophenyl acetate hydrolysis by pancreatic lipases. Kinetic investigation of the p-nitrophenyl acetate hydrolysis by the various pancreatic lipases and by the C-terminal domain (336-449) of human lipase reveals that this hydrolysis is the result of the superimposition of ...
Müller T, Chavaillaz PA, Jörg A, Grob M, Peterhans E.It was shown in several cell types that the dual lipoxygenase and cyclooxygenase inhibitor eicosatetraynoic acid but not the cyclooxygenase inhibitor acetylsalicylic acid suppressed luminol-dependent chemiluminescence. Since lipoxygenase is known to generate chemiluminescence in vitro, these observations were interpreted as evidence for a direct contribution of the lipoxygenase pathway to light emission in intact cells. We have investigated a possible contribution of the lipoxygenase to the chemiluminescence of horse eosinophils by directly comparing the formation of the byproduct chemilumines...
Rosskopf U, Noeske K, Werner E.This paper describes a double antigen ELISA (DAE) for rapid, specific and reliable assessment of the antitetanus immune status of horses and sheep. Compared with the indirect ELISA, the double antigen ELISA has the advantage of species-independent testing of sera. Thanks to its test design, it is more specific since the detected antibodies are forced to bind tetanus toxoid twice. In addition, it is very sensitive to tetanus antibodies, enabling the detection of low antibody titres, in range which is relevant for the assessment of the protective status (tetanus toxin neutralising antibodies). T...
Lorenzo PL, Liu IK, Illera JC, Picazo RA, Carneiro GF, Illera MJ, Conley AJ, Enders AC, Illera M.Epidermal growth factor (EGF) has been reported to promote different functions in mammalian ovaries, including oocyte maturation. The aim of the present study was to establish: that EGF influences oocyte maturation in ovine and equine, that a tyrosine kinase-dependent intracellular mechanism mediates EGF effect and, that EGF-R receptor is detectable in ovarian follicles by immunohistochemistry methods. Selected ovine and equine oocytes were aspirated from 2-5 mm (ovine) or 25 mm (equine) follicles and cultured in TCM 199 for 22 (ovine) or 36 hours (equine). They are then subjected to culture w...
Debord J, Verneuil B, Bollinger JC, Merle L, Dantoine T.The enzymatic hydrolysis of butyrylcholine, catalyzed by horse serum butyrylcholinesterase (EC 3.1.1.8), was studied at 37 degrees C in Tris buffer (pH 7.5) by flow microcalorimetry. A convolution procedure, using the Gamma distribution to represent the impulse response of the calorimeter, was developed to analyze the microcalorimetric curves. After correction for buffer protonation, the hydrolysis reaction was found to be slightly endothermic, with Delta H=+9.8 kJ mol(-1). Enzyme kinetics was studied with both the differential and integrated forms of the Michaelis equation with equivalent res...
Park HB, Marklund S, Jeon JT, Mickelson JR, Valberg SJ, Sandberg K, Andersson L.The PRKAG3 gene encodes a muscle-specific isoform of the regulatory gamma subunit of AMP-activated protein kinase (AMPK). A major part of the coding PRKAG3 sequence was isolated from horse muscle cDNA using reverse-transcriptase (RT)-PCR analysis. Horse-specific primers were used to amplify genomic fragments containing 12 exons. Comparative sequence analysis of horse, pig, mouse, human, Fugu, and zebrafish was performed to establish the exon/intron organization of horse PRKAG3 and to study the homology among different isoforms of AMPK gamma genes in vertebrates. The results showed conclusively...
