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.
Kingston JK, Bayly WM, Sellon DC, Meyers KM, Wardrop KJ.To investigate the effects of sodium citrate, low molecular weight heparin (LMWH), and prostaglandin E1 (PGE1) on aggregation, fibrinogen binding, and enumeration of equine platelets. Methods: Blood samples obtained from 4 Thoroughbreds. Methods: Blood was collected into syringes in the ratio of 9 parts blood:1 part anticoagulant. Anticoagulants used were sodium citrate, LMWH, sodium citrate and LMWH, or 300 nM PGE1/ml of anticoagulant. Platelet aggregation in response to ADP, collagen, and PGE1 was assessed, using optical aggregometry. Platelet activation was evaluated, using flow cytometry, ...
Galli C, Crotti G, Notari C, Turini P, Duchi R, Lazzari G.Embryo production by in vitro techniques has increased steadily over the years. For cattle where this technology is more advanced and is applied more, the number of in vitro produced embryos transferred to final recipients was over 30,000 in 1998. An increasing proportion of in vitro produced embryos are coming from oocytes collected from live donors by ultrasound-guided follicular aspiration (ovum pick up, OPU). This procedure allows the repeated production of embryos from live donors of particular value and is a serious alternative to superovulation. Ovum pick up is a very flexible technique...
The Journal of parasitologyApril 25, 2001
Volume 87, Issue 2 345-353 doi: 10.1645/0022-3395(2001)087[0345:COTOIO]2.0.CO;2
Dubey JP, Liddell S, Mattson D, Speert CA, Howe DK, Jenkins MC.Neospora hughesi was isolated in cell cultures inoculated with homogenate of spinal cord from a horse in Oregon. Tachyzoites of this Oregon isolate of N. hughesi were maintained continuously by cell culture passage and tachyzoites were infective to immunosuppressed mice. Gamma interferon gene knockout (KO) mice injected with tachyzoites developed fatal myocarditis and numerous tachyzoites were seen in lesions. Gerbils (Meriones unguiculatus) inoculated with tachyzoites developed antibodies (> or = 1:500) as indicated by the Neospora caninum agglutination test but did not develop clinical si...
Nie GJ, Wenzel JG.Hypoosmotic swelling (HOS) is used for assessing plasma membrane function and fertilizing capacity of human spermatozoa. However, HOS solutions and methodologies have not been evaluated specifically for assessing stallion spermatozoa. The objective of this study was to identify a HOS solution and assay conditions specifically for stallions that would maximize spermatozoal plasma membrane swelling. The HOS solutions and assay conditions, including incubation time (15 to 180 min), temperature (25 degrees vs 37 degrees C), and total number of cells examined (100, 200 or 500) were evaluated. Assay...
Arienti G, Polci A, De Cosmo A, Saccardi C, Carlini E, Palmerini CA.The semen of several mammals contains vesicles of different composition and origin. We have recently reported on the presence of lipoprotein vesicles in stallion semen. To a certain extent, these resemble human prostasomes, but differ from them in amount and composition. These horse-semen prostasome-like vesicles may be important, not only in horse reproductive physiology, but also in view of stallion semen cryopreservation. In this paper, we have studied horse-semen prostasome-like vesicles and found that they possess less saturated fatty acid than human prostasomes. Moreover, their protein p...
Davis BS, Chang GJ, Cropp B, Roehrig JT, Martin DA, Mitchell CJ, Bowen R, Bunning ML.Introduction of West Nile (WN) virus into the United States in 1999 created major human and animal health concerns. Currently, no human or veterinary vaccine is available to prevent WN viral infection, and mosquito control is the only practical strategy to combat the spread of disease. Starting with a previously designed eukaryotic expression vector, we constructed a recombinant plasmid (pCBWN) that expressed the WN virus prM and E proteins. A single intramuscular injection of pCBWN DNA induced protective immunity, preventing WN virus infection in mice and horses. Recombinant plasmid-transform...
Egerbacher M, Wolfesberger B, Gabler C.Quinolones and magnesium deficiency cause similar lesions in joint cartilage of young animals. Chondrocytes cultivated in the presence of quinolones and in Mg-free medium show severe alterations in cytoskeleton and decreased ability to adhere to the culture dish. We investigated whether Mg2+ supplementation can prevent quinolone-mediated effects on chondrocytes in vitro. Chondrocytes cultivated in Dulbecco's modified Eagle's medium/HAM's F-12 medium were treated with ciprofloxacin (80 and 160 microg/ml) and enrofloxacin (100 and 150 microg/ml). Mg2+ was added at a concentration of 0.0612 mg/ml...
Wattle OS.To determine whether there is a change in the expression of cytokeratins in the epidermal cells of the non-weight-bearing parts of the limb in horses with acute laminitis and thus determine whether the morphologic changes that develop in the periople and chestnut (torus carpeus) of horses early in acute laminitis are caused by inhibition of keratinocyte differentiation. Methods: 8 horses with acute laminitis. Methods: Tissue specimens were obtained from the chestnuts of all 8 horses and from the stratum externum of the hoof wall of 3 horses. Tissue specimens were obtained within 48 hours of th...
Fuller CJ, Barr AR, Dieppe PA.An explant system was used to investigate the hypothesis that cartilage from different equine joints might respond differently to challenge with interleukin-1alpha (IL-1alpha). Pairs of normal cartilage samples were taken from the metacarpophalangeal, proximal interphalangeal and distal interphalangeal joints of six horses. One of each pair was stimulated with 10 ng/ml human recombinant IL-1alpha for three days, and the supernatants and remaining cartilage explants were analysed for their total content of glycosaminoglycans. A significantly higher percentage of glycosaminoglycans was released ...
da Gracca Macoris D, Bertone A.The study of the influence of motion and initial intra-articular pressure (IAP) on intra-articular pressure profiles in equine cadaver metatarsophalangeal (MTP) joints was undertaken as a prelude to in vivo studies. Eleven equine cadaver MTP joints were submitted to 2 motion frequencies of 5 and 10 cycles/min of flexion and extension, simulating the condition of lower and higher (double) rates of passive motion. These frequencies were applied and pressure profiles generated with initial normal intra-articular pressure (-5 mmHg) and subsequently 30 mmHg intra-articular pressure obtained by inje...
Pepin K, Momose F, Ishida N, Nagata K.Heat shock protein 90 (Hsp90), a molecular chaperone, is ubiquitous and involved in numerous cellular processes. To contribute to the relatively small collection of vertebrate Hsp90 sequences in the gene data bank, we cloned and sequenced horse (Equus caballus) Hsp90 alpha and beta cDNAs. This enabled identification of horse-specific primers for development of a convenient PCR-based method that could monitor horse stress tolerance. We analyzed the sequence data comparatively and phylogenetically with other Hsp90 cDNA sequences, and identified vertebrate-specific and isoform-specific conserved ...
Chopineau M, Martinat N, Galet C, Guillou F, Combarnous Y.Horse LH/CG (eLH/CG) and donkey LH/CG (dkLH/CG) are strictly LH-specific in their respective homologous species. However, both bind to the FSH receptors from non-equid species, whereas the zebra hormone (zbLH/CG) does not. The FSH/LH ratio of eLH/CG and of the alphadkbetae hybrid is about tenfold higher than that of dkLH/CG and of the alphaebetadk hybrid, showing that the betae subunit contains the structural features responsible for the high FSH activity of eLH/CG. Only six amino acid positions (51, 94, 95, 102, 103 and 106) are unique to the betae subunit when compared with the betadk and be...
Scott TJ, Carnevale EM, Maclellan LJ, Scoggin CF, Squires EL.Objectives of the present study were to use oocyte transfer: 1) to compare the developmental ability of oocytes collected from ovaries of live mares with those collected from slaughterhouse ovaries; and 2) to compare the viability of oocytes matured in vivo, in vitro, or within the oviduct. Oocytes were collected by transvaginal, ultrasound-guided follicular aspiration (TVA) from live mares or from slicing slaughterhouse ovaries. Four groups of oocytes were transferred into the oviducts of recipients that were inseminated: 1) oocytes matured in vivo and collected by TVA from preovulatory folli...
Crockett EC, Graham JK, Bruemmer JE, Squires EL.The ability to ship cooled stallion semen to a facility that specializes in cryopreservation of spermatozoa would permit stallions to remain at home while their semen is cryopreserved at facilities having the equipment and expertise to freeze the semen properly. To accomplish this goal, methods must be developed to freeze cooled shipped semen. Three experiments were conducted to determine the most appropriate spermatozoal extender, package, time of centrifugation, spermatozoal concentration and length of time after collection that spermatozoa can be cooled before cryopreservation. In the first...
Mungall BA, Kyaw-Tanner M, Pollitt CC.Utilizing an in vitro laminitis explant model, we have investigated how bacterial broth cultures and purified bacterial proteases activate matrix metalloproteinases (MMPs) and alter structural integrity of cultured equine lamellar hoof explants. Four Gram-positive Streptococcus spp. and three Gram-negative bacteria all induced a dose-dependent activation of MMP-2 and MMP-9 and caused lamellar explants to separate. MMP activation was deemed to have occurred if a specific MMP inhibitor, batimastat, blocked MMP activity and prevented lamellar separation. Thermolysin and streptococcal pyrogenic ex...
Wattle O.The cytoskeleton of living keratinocytes consists mainly of cytokeratins that have polymerised into intermediate filaments. The aim of this study was to describe the expression of cytokeratins in the living epidermal cells of the weight-bearing parts of the equine hoof wall during acute spontaneous laminitis. A total of 9 hooves from 3 horses subjected to euthanasia within 48 h of the first clinical signs of laminitis were sectioned and examined. The cytokeratins in the stratum medium and stratum internum of the hoof wall were characterized by 1- and 2-dimensional gel electrophoresis, and the ...
Bailey SR, Cunningham FM.To compare adherence of stimulated and unstimulated eosinophils from allergic and normal ponies to cultured equine vascular endothelial cells (equine digital vein endothelial cells; EDVEC) and examine the effect of eosinophil-derived factor(s) on cell adherence. Methods: Eosinophil adherence to unstimulated EDVEC or EDVEC pretreated with IL-1beta or supernatants from stimulated eosinophils was measured. Supernatants were also assayed for TNFalpha and IL-1beta-like bioactivity. Results: Adherence of unstimulated and rhIL-5 (10 ng/ml)-stimulated eosinophils from allergic ponies to rhIL-1beta-tre...
Radcliffe RM, Lopez MJ, Turner TA, Watkins JP, Radcliffe CH, Markel MD.To compare the biomechanical properties of intact immature horse femurs and 3 stabilization methods in ostectomized femurs. Methods: Eighteen pairs of femurs from immature horses aged 1 to 15 months, and weighing 68 to 236 kg. Methods: Thirty-four immature horse femurs were randomly assigned to 1 of 5 test groups: 1) interlocking intramedullary nail (IIN) (n = 6); 2) IIN with a cranial dynamic compression plate (I/DCP) (n = 6); 3) 2 dynamic compression plates (2DCP) (n = 8); 4) intact femurs tested to failure in lateromedial (LM) bending (n = 6); and 5) intact femurs tested to failure in caudo...
Amavisit P, Browning GF, Lightfoot D, Church S, Anderson GA, Whithear KG, Markham PF.A rapid polymerase chain reaction (PCR) assay was developed for detecting Salmonella in faeces of horses and assessed on samples from horses admitted to a veterinary hospital. Direct detection was achieved by amplification of part of ompC after extraction of DNA from faeces using a spin column method to reduce the amount of inhibitory substances in samples. An internal positive control was included to detect false negative results. While the sensitivity of the PCR assay was less than culture when assessed on faeces inoculated with Salmonella, its sensitivity on faecal samples obtained from hor...
Dubey JP, Mattson DE, Speer CA, Hamir AN, Lindsay DS, Rosenthal BM, Kwok OC, Baker RJ, Mulrooney DM, Tornquist SJ, Gerros TC.An isolate of Sarcocystis neurona (SN7) was obtained from the spinal cord of a horse with neurologic signs. The parasite was isolated in cultures of bovine monocytes and equine spleen cells. The organism divided by endopolygeny and completed at least one asexual cycle in cell cultures in 3 days. The parasite was maintained by subpassages in bovine monocytes for 10 months when it was found to be non-pathogenic to gamma interferon knockout (KO) mice. Revival of a low passage (10th passage) of the initial isolate stored in liquid nitrogen for 18 months retained its pathogenicity for KO mice. Mero...
Speer CA, Dubey JP.The ultrastructure of Sarcocystis neurona schizonts and merozoites was studied in specimens derived from cell culture and from the brains of infected mice. Schizonts and merozoites were located in the host cell cytoplasm without a parasitophorous vacuole at any stage of development. Merozoites divided by endopolygeny. Fully formed merozoites had a pellicle, numerous polysomes and ribosomes, smooth and rough endoplasmic reticulum, 22 subpellicular microtubules, 9-16 dense granules, 25-75 micronemes, a plastid, a Golgi complex, 1-3 mitochondria, a conoid, 2 apical rings, 2 polar rings, 0-6 lipid...
Marsh AE, Mullins AL, Lakritz J.Parasite-specific incorporation of (3)H-uracil was used to assess the replication of Sarcocystis neurona, a protozoal parasite associated with equine protozoal myeloencephalitis (EPM). Anti-protozoal drugs, pyrimethamine (0.01, 0.1 and 1.0microg/ml PYR), sulfadiazine (5microg/ml; SDZ), sulfamethoxazole (5microg/ml; SMZ), diclazuril (100ng/ml; DCZ), atovaquone (0.04ng/ml; ATQ), tetracycline (5microg/ml; TET) and the herbicide glyphosate (1.5 and 4.5mM; GLY) were studied with varying S. neurona parasite densities (2x10(1)-1.2x10(6)merozoites/well). A microtiter plate format was used to test thes...
Bush JA, Freeman DE, Kline KH, Merchen NR, Fahey GC.Addition of fat to the diet of the equine is a popular method of increasing energy density of the diet while reducing feed intake. Reducing feed intake is of interest to race horse trainers because additional feed is seen as additional weight and, therefore, a hindrance to performance. Limited information is available regarding the interactions of fat with other dietary components, particularly fiber, in the equine digestive system. The effect of dietary fat on in vitro nutrient disappearance in equine cecal fluid was studied in Exp. 1 using a split-plot design within a 2 x 2 Latin square. Two...
Magnarelli LA, Ijdo JW, Van Andel AE, Wu C, Fikrig E.To develop and evaluate a polyvalent ELISA incorporating a highly specific recombinant antigen (p44) for diagnosis of granulocytic ehrlichiosis in dogs and horses. Methods: 32 dogs and 43 horses. Methods: Results of the ELISA were compared with results of indirect fluorescent antibody (IFA) staining and western immunoblotting incorporating whole-cell antigen. Results: For the canine and equine samples, percentages of samples with positive IFA staining, western immunoblotting, and ELISA results were similar. For 29 (91 %) canine samples and 30 (70%) equine samples, results of IFA staining, west...
Moses VS, Hardy J, Bertone AL, Weisbrode SE.To evaluate the effects of anti-inflammatory drugs on lipopolysaccharide (LPS)-challenged and -unchallenged equine synovial membrane in terms of production of prostaglandin E2 (PGE2) and hyaluronan, viability, and histomorphologic characteristics. Methods: Synovial membranes were collected from the carpal, tarsocrural, and femoropatellar joints of 6 adult horses. Methods: Synovial membranes from each horse were minced and pooled and explants were treated with one of the following: no drug (control), drug, LPS alone, or LPS and drug. Treatment drugs were phenylbutazone (PBZ), flunixin meglumine...
Davenport CL, Boston RC, Richardson DW.To investigate the effects of enrofloxacin and magnesium deficiency on explants of equine articular cartilage. Methods: Articular cartilage explants and cultured chondrocytes obtained from adult and neonatal horses. Methods: Full-thickness explants and cultured chondrocytes were incubated in complete or magnesium-deficient media containing enrofloxacin at concentrations of 0, 1, 5, 25, 100, and 500 microg/ml. Incorporation and release of sulfate 35S over 24 hours were used to assess glycosaminoglycan (GAG) synthesis and degradation. An assay that measured binding of dimethylmethylene blue dye ...
Venugopal CS, Moore RM, Holmes EP, Koch CE, Seahorn TL, Beadle RE.To compare responses of bronchial rings obtained from healthy horses and horses affected with summer pasture-associated obstructive pulmonary disease (SPAOPD) to selected mediators of airway hyperreactivity in vitro. Methods: Bronchial rings from 6 healthy horses and 6 horses affected with SPAOPD. Methods: Bronchial rings obtained from each group of horses were mounted in organ baths and attached to force transducers interfaced with a polygraph. After applying 2g of tension, each ring was allowed to equilibrate for 45 minutes in Tyrode's solution at 37 C. Cumulative concentration-response rela...
Adams AP, Antczak DF.A system for transplanting invasive equine trophoblast (i.e., chorionic girdle) to ectopic sites has been developed as a means to study the differentiation of this tissue and to assess maternal immune responses to the conceptus tissue in a site outside the uterus. Chorionic girdle was isolated from Day 33 to 34 conceptuses and surgically placed into the vulvar mucosa or subdermal skin of recipient mares. Biopsy specimens of the graft sites for immunohistochemical staining were taken at weekly or biweekly intervals after grafting. Serum samples were collected from each recipient and tested for ...
Merkies K, Chenier T, Plante C, Buhr MM.Viability of spermatozoa can be assessed by numerous methods, but many are slow and poorly repeatable, and subjectively assess only 100 to 200 spermatozoa per ejaculate. We collected two ejaculates from each of 4 stallions, and extended them to 50x10(6) sperm/mL in a nonfat dried milk solids glucose extender (EZ Mixin). Half the ejaculate was freeze-killed by immersing in liquid nitrogen for 10 min. Aliquots using appropriate volumes of live and freeze-killed spermatozoa provided the following ratios of live:dead spermatozoa: 100:0, 75:25, 50:50, 25:75, 0:100. We determined the viability of ea...
Stewart JM, Blakely JA, Johnson MD.Non-covalent modification of cytochrome c may have implications for electron transport and energy metabolism. We examined the interaction of various fatty acids (FAs), their coenzyme A and carnitine esters, and fatty alcohols with horse heart ferrocytochrome c. A comparison of FAs indicated a minimum chain length of 14 carbons was required for significant effect on the ferroheme chromophore and major changes in electronic spectra. Coenzyme A and carnitine esters interacted less strongly than FAs whereas long-chain alcohols did not interact with the protein. We found a single, saturable FA bind...
Bauquier JR, Tennent-Brown BS, Tudor E, Bailey SR.Doramapimod (BIRB-796-BS), is an anti-inflammatory compound, acting through p38 MAPK inhibition, but its anti-inflammatory effects have not previously been studied in the horse. Whole blood aliquots from healthy horses diluted 1:1 with cell culture medium were incubated for 21 h with 1 μg/ml of lipopolysaccharide (LPS), lipoteichoic acid (LTA) or peptidoglycan (PGN) in the presence of increasing concentrations of doramapimod (3 × 10 M to 10 M). Cell bioassays were used to measure TNF-α and IL-1β activity. Doramapimod significantly and potently inhibited TNF-α and IL-1β activity induced b...
Ebani VV, Nardoni S, Bertelloni F, Pollera C, Pistelli L, Mancianti F.The aim of the present study, the first to test clinical equine isolates, was to evaluate the antimicrobial activity of commercial essential oils (EOs) obtained from and against spp., spp., and spp. previously isolated from uteri of mares with reproductive disorders. showed the best antibacterial activity against all the selected agents, with MICs ranging from 0.07 mg/mL for subsp. to 0.29 mg/mL versus sp. and spp. isolates and versus and spp., with MICs of 0.46 mg/mL and 1.86 mg/mL, respectively. EOs showed good antibacterial activity with MICs ranging from 0.07 mg/mL ...
Yates KM, Stilwell NK, Fontenot RL, Betbeze CM.To determine the in vitro antibacterial efficacy of equine and canine autologous conditioned plasma (ACP) and amniotic membrane extract eye drops (AMEED) against aerobic bacteria common to the corneal surface. Methods: Canine (n = 4) and equine (n = 4) anticoagulated whole blood samples were sterilely collected, pooled for each species, and processed using the Arthrex ACP® Double-Syringe System. Platelet counts were performed on ACP and pooled blood. AMEED were obtained from a commercial source. An electronic medical records search (2013-2022) identified aerobic bacteria cultured from...
Barnés CM, Petoud S, Cohen SM, Raymond KN.The ability of ferritin as an Fe(II) detoxifier and Fe(III) storage protein is limited by its ability to recognize and incorporate Fe(II), which is then oxidized and mineralized at internal protein sites. The Cr(III) amine complex [Cr(N(CH(2)CH(2)NH(2))(3)(H(2)O)(OH)](2+) [abbreviated as Cr(TREN)] is a kinetically inert inhibitor of iron incorporation and mineralization in ferritin. Unlike other inhibitors, Cr(TREN) can only exchange its two aqua/hydroxy ligands. Competition studies between Cr(TREN) and Tb(III) binding have been performed in horse spleen ferritin (HoSF) to probe uptake of Fe(I...
Andrietti ALP, Durgam SS, Naumann B, Stewart M.Mesenchymal stem cells are characterized by their capacities for extensive proliferation through multiple passages and, classically, tri-lineage differentiation along osteogenic, chondrogenic and adipogenic lineages. This study was carried out to compare osteogenesis in equine bone marrow-, synovium- and adipose-derived cells, and to determine whether osteogenic capacity is reflected in the basal expression of the critical osteogenic transcription factors Runx2 and Osterix. Unassigned: Bone marrow, synovium and adipose tissue was collected from six healthy 2-year-old horses. Cells were isolate...
Javsicas LH, Giguère S, Womble AY.The objectives of this study were to determine the serum and pulmonary disposition of telithromycin in foals and to determine the minimum inhibitory concentration (MIC) of telithromycin against macrolide-susceptible and macrolide-resistant Rhodococcus equi isolates. A single dose of telithromycin (15 mg/kg of body weight) was administered to six healthy 6-10-week-old foals by the intragastric route. Activity of telithromycin was measured in serum, pulmonary epithelial lining fluid (PELF), and bronchoalveolar lavage (BAL) cells using a microbiological assay. The broth macrodilution method was u...
Venugopalan CS, Moore RM, Holmes EP, Sedrish SA.To determine the in vitro contractile responses of equine colonic arteries to angiotensin II, histamine, serotonin, norepinephrine, prostaglandin F2 alpha, vasopressin, and a thromboxane-B2-analogue. Methods: The tension generated in colonic arterial rings placed in organ baths with oxygenated Tyrode's solution at 37 degrees C after exposure to the previously mentioned chemical agents was measured using force-transducers interfaced with a polygraph. Methods: Large colon arterial rings collected from eight horses. Methods: The rings were allowed to equilibrate for 45 minutes after applying 2 g ...
Wotman KL, Chow L, Martabano B, Pezzanite LM, Dow S.Ocular surface squamous neoplasia (OSSN) is the major cause of corneal cancer in man and horses worldwide, and the prevalence of OSSN is increasing due to greater UVB exposure globally. Currently, there are no approved treatments for OSSN in either species, and most patients are managed with surgical excision or off-label treatment with locally injected interferon alpha, or topically applied cytotoxic drugs such as mitomycin C. A more broadly effective and readily applied immunotherapy could exert a significant impact on management of OSSN worldwide. We therefore evaluated the effectiveness of...
Hoffseth K, Randall C, Hansma P, Yang HT.In an attempt to study the mechanical behavior of bone under indentation, methods of analyses and experimental validations have been developed, with a selected test material. The test material chosen is from an equine cortical bone. Stress-strain relationships are first obtained from conventional mechanical property tests. A finite element simulation procedure is developed for indentation analyses. The simulation results are experimentally validated by determining (1) the maximum depth of indentation with a single cycle type of reference point indentation, and (2) the profile and depth of the ...
Boy MG, Zhang C, Antczak DF, Hamir AN, Whitlock RH.A 10-month-old Arabian foal was evaluated for a suspected immunoglobulin (Ig) M deficiency. Decreased to nondetectable concentrations of IgM, IgA, and IgG (T), and a normal concentration of IgG, were present. Results of in vitro testing of the blood lymphocyte blastogenesis showed a weak response to the B-cell mitogen, lipopolysaccharide (LPS), but normal responses to T-cell mitogens. Results of postmortem examination showed synovitis of the left tibiotarsal and both scapulohumeral joints. Atrophy and edema of the lymph nodes and lymphocyte depletion in the thymus and spleen were seen. A subac...
Ponthier J, Franck T, Detilleux J, Mottart E, Serteyn D, Deleuze S.Despite improvement of techniques, semen of 20% of stallions remains unfreezable. Recent studies focused on the impact of reactive oxygen species and oxidant enzymes on semen characteristics. Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutrophils during degranulation or after cell lysis. It is responsible for the formation of hypochlorous acid, a strong oxidant agent, which could damage spermatozoa. The aim of this study was to determine the relation between MPO concentration and characteristics of frozen semen from stallions. Thirty-five straws from different s...
Benton HP, Esquivel AV, Rice AD, Giri SN.Pirfenidone is under investigation as an anti-inflammatory and anti-fibrotic agent in several organs including lung. Since important features of arthritic conditions include inflammation and long-term damage to articular cartilage, we have investigated whether PD can suppress chondrocyte responses to bacterial lipopolysaccharide (LPS) and interleukin 1 (IL-1); modulators that induce a cascade of inflammatory responses that lead to articular joint tissue damage. PD (0 - 5microM) showed no effect on cell number or viability when incubated with high density primary equine chondrocyte cultures for...
Smits K, Goossens K, Van Soom A, Govaere J, Hoogewijs M, Peelman LJ.In vitro-produced (IVP) equine blastocysts can give rise to successful pregnancies, but their morphology and developmental rate differ from those of in vivo-derived equine blastocysts. The aim of the present study was to evaluate this difference at the genetic level. Suppression subtractive hybridisation (SSH) was used to construct a cDNA library enriched for transcripts preferentially expressed in in vivo-derived equine blastocysts compared with IVP blastocysts. Of the 62 different genes identified in this way, six genes involved in embryonic development (BEX2, FABP3, HSP90AA1, MOBKL3, MCM7 a...
Hobbs SJ, Mather J, Rolph C, Bower JA, Matuszewski B.Strains during stance on the hoof wall surface have been measured by a number of authors in vitro and in vivo. Histological structure and mechanical properties vary through the wall thickness (radially); radial strain measurements may therefore aid the understanding of mechanical function of the capsule and adjacent tissues. Objective: To develop instrumentation capable of measuring internal hoof strain, and to carry out a preliminary comparison of normal and laminitic hooves. Methods: Six forelimbs from 4 horses, including 2 with laminitis from the same horse, were tested using an Instron tes...
Barton MH, Ferguson D, Davis PJ, Moore JN.Pentoxifylline (7.5 mg/kg) was bolused intravenously to eight healthy horses and was immediately followed by infusion (1.5 mg/kg/h) for 3 h. Clinical parameters were recorded and blood samples were collected for 24 h. Plasma was separated and concentrations of pentoxifylline, its reduced metabolite I, and 6-keto-prostaglandin F1 alpha were determined. Heparinized whole blood was also incubated ex vivo with 1 ng Escherichi coli endotoxin/mL blood for 6 h before determination of plasma tumour necrosis factor activity. The peak plasma concentrations of pentoxifylline and metabolite I occurred at ...
Sullivan SN, Altmann NN, Brokken MT, Durgam SS.Primary deep digital flexor tendon (DDFT) pathologies and those accompanying degenerative changes of navicular bone fibrocartilage are major causes of lameness associated with navicular disease. Intrasynovial corticosteroids are mainstay in the treatment due to the anti-inflammatory effects, but their effect on DDFT cell biosynthesis are unknown. The objective of this study was to investigate the effects of methylprednisolone acetate (MPA) on cells isolated from the dorsal fibrocartilaginous region of forelimb DDFTs (DDFT-derived cells) of 5 horses (aged 11-17 years). Non-adherent aggregate c...
Balao da Silva CM, Spinaci M, Bucci D, Giaretta E, Peña FJ, Mari G, Galeati G.The interest on sex sorting by flow cytometry on the equine industry has been increasing over the years. In this work, three different tests were performed in order to evaluate the membrane status of sorted stallion spermatozoa: assessment of binding ability to porcine oocytes, evaluation of acrosome integrity after stimulation with A23187, and detection of tyrosine phosphorylation. These evaluations were made after incubation for 0h, 1.5h and 3h in a capacitating medium. Sorted stallion spermatozoa attached similarly to the porcine oocytes, when compared with control samples. Sorted spermatoz...
Flaminio MJ, Rush BR, Davis EG, Hennessy K, Shuman W, Wilkerson MJ.This paper describes a method for simultaneously measuring phagocytosis and oxidative burst activity in equine peripheral blood leukocytes by flow cytometry. Opsonized propidium iodide-labelled Staphylococcus aureus (PI-Sa) was used to measure the uptake of bacteria by equine phacocytes and the oxidative burst activity by oxidation of dihydrorhodamine 123. The requirements to achieve optimal activity of phagocytosis and oxidative burst are described. The advantage of the simultaneous technique is that it provides both independent and comparative values for phagocytosis and the oxidative burst,...
Boerboom D, Kerban A, Sirois J.Estradiol biosynthesis is a key biochemical trait of developing follicles. To study its regulation in equine follicles, the objectives of this study were to clone and determine the structure of equine cytochrome P450 aromatase (P450AROM), and characterize the regulation of P450AROM and P450 17alpha-hydroxylase/C17-20 lyase (P45017alpha) messenger RNAs (mRNAs) in vivo in equine preovulatory follicles isolated during hCG-induced ovulation. Two distinct P450AROM complementary DNAs (cDNAs) were isolated from an equine preovulatory follicle cDNA library. One clone was 2682 bp in length and included...
Frippiat T, Dams L, Wielick C, Delguste C, Ludwig-Begall LF, Art T, Thiry E.Viruses can be involved in respiratory disorders in horses, with limited therapeutic options. Citrate-complexed silver nanoparticles (C-AgNP) have shown bactericidal properties after in vitro nebulization. The aim of the present study was to assess the virucidal activity of C-AgNP after in vitro instillation or nebulization on equine herpesvirus-1 (EHV-1) and murine norovirus (MNV), the latter used as surrogate for small non-enveloped viruses. Both viruses were instilled or nebulized with C-AgNP of increasing concentrations, and titres were determined via TCID50 method. We demonstrated efficie...
Mateos-Trigos G, Evans RJ, Heath MF.Horses show susceptibility to platelet-related disorders. Equine platelets differ from human platelets in some of their responses, so information available about human platelets must be validated in the horse. Aggregation of platelets by ADP involves both P2Y(1) and P2Y(12) receptors on the platelet surface. We have compared the effect of the P2Y(12) antagonist, AR-C67085, on equine and human platelets in vitro using turbidimetric aggregometry to measure the rate and final extent of aggregation. Aggregation profiles, concentration-response curves and pA(2) values show that the rate of aggregat...
Müller-Schöttle F, Bogusz A, Grötzinger J, Herrler A, Krusche CA, Beier-Hellwig K, Beier HM.After its original description as a steroid-dependent protein in the rabbit uterus, uteroglobin became one of the best characterized proteins. However, detailed knowledge of its physiological role remains an enigma. In this study we investigate how its structure is phylogenetically conserved in the horse compared to other mammalian species. Northern blot analysis showed that in horses, the main expression of uteroglobin appears in lung, uterus, and prostate tissues. Western blot analysis demonstrated that the dimeric form of uteroglobin is found predominantly in biological compartments. Using ...
Jackson PS, Allen WR, Ricketts SW, Hall R.Uterine irrigation was carried out in eight Welsh pony mares using 50 ml of chlorhexidine gluconate solution diluted to give active ingredient concentrations of 0.25 per cent to 2 per cent. Treatment was repeated up to twice in mares showing no adverse effects and irritancy of treatment judged on clinical symptoms and uterine biopsy. Results indicated the inadvisability of using a higher concentration than 0.25 per cent. Three daily applications of a diluted surgical scub solution containing 2 per cent chlorhexidine gluconate to the penis of an arab stallion failed to produce symptoms or irrit...
Rebalka IA, Lindinger MI.A commercially available equine fecal blood test (FBT) claims to be able to detect the presence of blood proteins (albumin and hemoglobin) in manure. The purpose of this study was to determine the FBT test sensitivity, specificity, positive and negative predictive values (PPV, NPV), lower threshold of detection, time to obtain a visual positive result, effect of temperature and result stability in lab and field conditions. The FBT was assessed for its sensitivity and specificity for detecting pure albumin and hemoglobin at two temperatures over a range of concentrations. Time to result was mea...
Minder HP, Merritt AM, Chalupa W.Feces from 13 healthy horses and 8 horses with chronic diarrhea were subjected to an in vetro fermentation procedure that had been developed for rumen fluid. Fermentations were conducted over 6 hours in a closed system, with and without an essential amino acid (EAA) mixture being added to the basic starch-buffer medium. The addition of EAA caused no significant difference in results of fermentation of feces from healthy horses. For diarrheic animals, there was a significant (P less than 0.01) increase in gas and total volatile fatty acids production whether EAA were present or not, and alpha-a...
Santos FC, Corcini CD, Costa VG, Gheller SM, Nogueira CE, da Rosa Curcio B, Varel AS.Solid storage medium prevents cellular sedimentation, reduces metabolic demand via limiting movement, and avoids the modification of an extender composition in the sedimentary microenvironment. It has been proven to prolong spermatozoa viability in mammalians. Objective: This experiment aims to evaluate the effect of cool storage in solid phase extender on stallion sperms. Methods: Semen was collected from 10 Crioulo stallions (n=30) and submitted to treatments: control group (semen extender) and groups with gelatin addition in different concentrations (semen extender + 1%, 2% and 3%). Seminal...
Bouchey D, Evermann J, Jacob RJ.Examination of six field isolates of equine herpesvirus 3, the causative agent of equine coital exanthema, indicates that all were temperature sensitive (ts) at the body temperature, 39 degrees C, of their host (Equine asinus and callabus) when grown in cell culture. The isolates were characterized by fingerprint analysis with the restriction endonucleases XbaI, EcoRI, BamHI and Hind III to establish possible epidemiologic relatedness. Three of the six isolates may be considered related. Variation in the mobility of the BamHI-A and Hind III-K fragments indicates that a small plaque isolate may...
Klett D, Combarnous Y.In previous studies, we had shown the synergistic effect of 10 M forskolin (FSK) on the detection threshold of the cyclic AMP response to luteinizing hormones (LH) and chorionic gonadotropins (CG) from various species in the mouse Leydig tumor cell (mLTC) cell line. Independently, we started to study the effect of 10-10 M oxytocin (OXT) also on the cyclic AMP response to LH and CG preparations on these same cells and found an amplifying effect on the luminescence response caused by gonadotropins. The aim was then to explore the effects of 10-10 M OXT on the gonadotropin-induced cAMP response, ...
Cornelisse CJ, Robinson NE, Berney CA, Eberhart S, Hauptman JE, Derksen FJ.In vitro, glucocorticoids potentiate vasoconstriction of equine digital vessels to catecholamines and this has been implicated as a mechanism of glucocorticoid-induced laminitis. This observation has never been confirmed in vivo. Objective: To study the effects of glucocorticoid therapy on vasoconstrictor responsiveness in the horse in vivo. Methods: In a blinded, randomised cross-over experiment, 9 horses were treated with either dexamethasone (0.1 mg/kg bwt i.v. q. 24 h) or saline i.v. for 6 days. The changes in local average skin temperature before (baseline) and after intradermal injection...
Araujo JM, Araújo JV, Braga FR, Carvalho RO.Three isolates of predator fungi Duddingtonia flagrans (AC001), Monacrosporium thaumasium (NF34), and Arthrobotrys robusta (I-31) were assessed in in vitro test regarding the capacity of prey infective larvae (L(3)) Strongyloides westeri. Compared to control, without fungus, there was a significant decrease (P 0.01) in vitro test. Linear regression coefficients of treated and control groups were -0.21 for control, -0.32 for D. flagrans, -0.34 for M. thaumasium, and -0.22 for A. robusta. In the following, isolates AC001 and NF34 were assessed in vivo regarding the capacity of supporting the pa...
