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.
Caffrey CR, Ryan MF.An excretory-secretory (ES) preparation derived from adult Strongylus vulgaris in vitro was assessed for proteolytic activity using azocasein and synthetic, fluorogenic, peptide substrates. Fractionation was by molecular sieve fast protein liquid chromatography (molecular sieve FPLC) and resolution by gelatin-substrate sodium dodecyl sulphate-polyacrylamide gel electrophoresis (gelatin-substrate SDS-PAGE). The cysteine proteinase activator, dithiothreitol (DTT), enhanced azocaseinolysis and hydrolysis of carbobenzoxy-phenylalanyl-arginine-7-amido-4-methylcoumarin (Z-Phe-Arg-NMec) by the ES pre...
Jackman BR, Moore JN, Barton MH, Morris DD.The purpose of this study was to investigate the in vitro effects of flunixin meglumine, a cyclo-oxygenase inhibitor, and ketoprofen, a reported cyclo-oxygenase and lipoxygenase inhibitor, on the synthesis of cyclo-oxygenase end-products thromboxane B2 and prostaglandin E2, lipoxygenase derived 12-hydroxyeicosatetraenoic acid, tumor necrosis factor and tissue factor. Six adult horses were each randomly administered flunixin meglumine (1.1 mg/kg) or ketoprofen (2.2 mg/kg) intravenously every 12 hours with the drug treatments separated by two weeks. Blood samples were obtained prior to initiatin...
Lloyd E, Mauk AG.Expression of recombinant horse heart myoglobin in Escherichia coli has been found to result in the production of both native and variable amounts (approximately 16-17% total) of two sulphmyoglobin isomers. The recombinant sulphmyoglobin produced consists primarily of the A and B isomers as identified by 1H NMR spectroscopy with no evidence for production of the C isomer. Conversion of recombinant sulphmyoglobin to the native protein can be achieved by reconstitution with protohaem IX. The possible relationship of this observation to recombinant expression of other heme proteins is discussed.
Todhunter RJ, Wootton JA, Lust G, Minor RR.Collagen type I was purified from equine skin and flexor tendon, and type II collagen was purified from equine articular cartilage. The proteoglycans in these tissues were extracted, using guanidine HCl; the collagens were solubilized, using pepsin digestion, then were selectively precipitated with NaCl. Gel electrophoresis indicated that the precipitates contained only type I or type II collagen. Amino acid analysis indicated that collagen constituted > 97% of the total protein in the precipitates. Hydroxylation of proline was 42.0 +/- 0.6% (mean +/- SEM) in alpha 1(I) and alpha 2(I), and was...
St-Laurent G, Morin G, Archambault D.A reverse transcription (RT)-PCR assay was developed for the detection of equine arteritis virus (EAV) in cell culture supernatant and in horse semen. Four different sets of oligonucleotide primers complementary to sequences located in the 3' end of the polymerase gene (open reading frame [ORF] 1b) and to sequences representing the entire ORFs 3, 4, and 7, which encode for nonstructural (ORFs 3 and 4) or viral nucleocapsid (ORF 7) proteins, were compared for their abilities to amplify the targeted EAV sequences by the RT-PCR procedure. The sensitivities of the RT-PCR for amplification of EAV s...
Windberger U, Ribitsch V, Resch KL, Losert U.There is little data on blood viscosity in different animals. Therefore a comparison was performed between five species by an in-vitro method using oscillatory flow. At shear rates from 1 to 150/sec the viscous and the elastic component of the complex blood viscosity was highest in pig, followed by horse, dog, ox, and sheep. Plasma viscosity and plasma density were similar in dog and sheep and were higher in ox. The differences in whole blood viscosity were obviously related to the variable interspecies hematocrit, and may also be influenced by different aggregation tendencies. Aggregation in ...
Bochröder B, Schubert R, Bödeker D.Discs of stripped mucosa from the proximal ventral colon were prepared immediately after slaughter of 8 Shetland cross-breed ponies. The mucosae were fixed in incubation chambers and used in incubation experiments to study the transmucosal fluxes of the amino acids lysine, histidine and arginine (150 min) and of ammonia (90 min). When the amino acid concentrations in the mucosal solution were in the physiological range (2.8-3.0 mmol/l) no transport to the serosal side of the tissue was found. When the concentrations were raised 10-fold, less than 2% of the mucosal amino acid pool was recovered...
Turell MJ, Beaman JR, Neely GW.The vector competence of Aedes taeniorhynchus (Wiedemann) and four strains of Aedes albopictus (Skuse) was assessed for eastern equine encephalitis (EEE) virus isolated from Ae. albopictus collected in Polk County, Florida. Both species became infected with and transmitted EEE virus by bite after feeding on 1-d-old chicks that had been inoculated with EEE virus (viremia = 10(10.1) plaque-forming units [PFU] per ml of blood). However, when fed on an older chick with a lower viremia (viremia = 10(6.1) PFU per ml of blood), Ae. albopictus was significantly more susceptible to infection (90%, n = ...
Hendrickson DA, Nixon AJ, Erb HN, Lust G.Equine neonatal chondrocytes were cultured in three-dimensional fibrin matrices under conditions of immediate implantation or implantation following monolayer culture for 6 days, and 3 cell concentrations (1 x 10(5), 1 x 10(6), and 5 x 10(6) chondrocytes/cm3). Equine fibrinogen was collected by cryoprecipitation and polymerized by use of activated bovine thrombin. The fibrin implants were harvested and analyzed histologically and biochemically at 3, 7, and 14 days after the chondrocytes were implanted in fibrin. The differentiation ratio (ratio of rounded, chondrocyte-like cells to stellate, f...
Katayama Y, Oikawa M, Kaneko M, Yoshihara T, Yoshikawa H, Yoshikawa T.Three monoclonal antibodies capable of individually recognizing chondrocytes, osteoblasts and osteocytes were prepared. EB-1 reacted with a 55-kDa antigen on the chondrocyte membrane, EB-2 with a 110-kDa antigen on the membrane of osteoblasts and/or partial osteocytes, and EB-3 with a 130-kDa antigen on the membrane of osteocytes. These monoclonal antibodies may be useful probes for studying the differentiation and maturation of osteogenic cells.
Pilling A, Davison AJ, Telford EA, Meredith DM.Glycoprotein 45 is a major envelope glycoprotein of equine herpesvirus type 1. The gene encoding this protein is located between map units 0.615 and 0.636 on the virus genome and evidence has suggested that it is encoded by gene 52, one of four genes within this region. Using PCR we have amplified gene 52 and subsequently cloned it into a mammalian expression vector under the control of the human cytomegalovirus immediate early gene promoter. The gene was expressed in COS-7 cells and its product was detected by immunofluorescence and Western blotting. The results indicate that glycoprotein 45 ...
Zientara S, Sailleau C, Moulay S, Cruciere C.A single tube reverse transcription-polymerase chain reaction (RT-PCR) method for detection of African horse sickness virus (AHSV) in splenic tissues from infected horses is described. Double stranded RNA was extracted from infected organs of horses and used to produce complementary DNA (cDNA) with the two primers selected for the PCR. The 1179 bp amplified product (the segment 7 which encodes for VP 7), detected by electrophoresis on agarose gel and ethidium bromide staining, was hydrolysed with eight restriction endonucleases for characterization of the AHSV. The sensitivity of this method i...
Kettler MK, Weil MR, Mascotti K, Perryman LE.A group of diseases termed combined immunodeficiency (CID) results in a severe form of immunodeficiency. While CID in humans has two genetics bases, in Arabian it is inherited in an autosomal recessive manner. Kettler et al. (1989) determined that uric acid was significantly (p 0.05) between carrier and non-carrier horse's serum levels of hypoxanthine or xanthine. These data, combined with our previous ones suggest that an enzymatic lesion in the purine salvage pathway may occur at the urate oxidase step. ZUSAMMENFASSUNG: Serum-Hypoxanthin- und -Xanthin-Spiegel in Pferden mit Heterozygotie fÃ...
Spiers S, May SA, Bennett D, Edwards GB.Isolated equine blood and articular cells were investigated for proteolytic enzyme production by means of gel filtration and analysis on 14C-acetylated collagen and casein substrates. Significant amounts of collagenase and caseinase activity were produced by cultured synoviocytes stimulated with equine interleukin 1, although large amounts of collagenase also originated from neutrophils.
Schubert R, Flachowsky G, Bochröder B.Chopped wheat straw was homogeneously mixed with urine of horses (5.75 gN per 1, 16.88 atom-% 15N-excess) and airtightly stored in plastic containers for 6 months. Three rumen fistulated sheep and goats each were fed with untreated or urine treated straw. Concentrate was added to straw. Untreated and urine treated straw were given in nylon bags and incubated in the rumen of sheep and goats for 1, 3, 6, 12, 24, 48 and 72 hours. A three compartment exponential function was used to fit the measurements of 15N-excess and 15N-amount of bag content. The curves and the calculated partial Y-values of ...
Weiss DJ, Evanson OA, Geor RJ.The effects of various concentrations of furosemide and pentoxifylline on equine RBC in vitro were evaluated to facilitate better understanding of the potential effects of these drugs on blood flow properties. Furosemide induced increased mean cell volume (MCV), increased RBC potassium concentration, increased whole blood viscosity, and decreased the RBC filtrability. These data indicate that furosemide may block the RBC membrane transport pathways resulting in potassium and water retention. The increase in size and the resultant decrease in the surface-area-to-volume ratio may have caused the...
Ronen N.Urinary tract pressure profiles were evaluated in 6 clinically normal geldings over 3 consecutive days. This was performed by introducing a 1.3 m-long cuffed catheter into the urinary tract, under xylazine sedation (0.8 mg/kg, iv). The method was reproducible. The mean (+/- sd) intra-vesicular pressure (IVP) and maximal urethral closure pressures (MUCP) were 10.3 (+/- 1.7) and 129.8 (+/- 19.6) cmH2O, respectively, and the ratio between MUCP and IVP was 13.2 (+/- 2.5). A gelding with urinary incontinence showed a significantly lower MUCP (73.4 cmH2O), and an MUCP to IVP ratio of 8.0. It was con...
Fischer B, Rose-Hellekant TA, Sheffield LG, Bertics PJ, Bavister BD.Preimplantation embryos of the pig (Days 11 to 15), cow (Days 14 to 16), sheep (Day 14) and pony (Day 16) bind epidermal growth factor (EGF) specifically. Binding was not detected in embryos of the rabbit at Day 5 or 6 or the hamster at Day 3. Transforming growth factor-alpha displaced [(125)I] EGF in pig, cow and pony embryos almost as much as unlabeled EGF. The binding affinities of EGF ranged from 12 to 233 pM in pig and cow embryos. The range of species and binding features indicate that the EGF family may play a significant role in mammalian preimplantation development.
Hochi S, Fujimoto T, Braun J, Oguri N.The objective of this study was to investigate the in vitro and in vivo developmental abilities of equine embryos cryopreserved by vitrification. Twenty-eight embryos were recovered from Native pony and Thoroughbred mares at Days 5 to 7 by nonsurgical uterine flushing (detection of ovulation=Day 0). The vitrification solution contained 40% ethylene glycol, 18% Ficoll, and 0.3 M sucrose in PBS. The embryos were placed for 1 to 2 min in vitrification solution (Group 1) or following exposure to 20% ethylene glycol in PBS for 10 to 20 min (Groups 2 and 3). Single embryos were loaded in 0.25-ml str...
Willis P, Caudle AB, Fayrer-Hosken RA.Transmission electron microscopy (TEM) was used to evaluate the fine structure of equine oocytes cultured in vitro. Oocytes obtained by follicular aspiration were cultured for either zero or 15 hr. After treatment oocytes were processed either by light microscopy (nuclear evaluation) or TEM (cytoplasmic evaluation). Those oocytes cultured for 15 hr were incubated in modified TCM 199 with 15% (v/v) mare serum (day of ovulation) at 39 +/- 0.2 degree C. Evaluation using TEM revealed that cortical granules were present in all oocytes. However, zero-time oocytes contained few cortical granules, and...
McDonnell SM, Oristaglio Turner RM.Imipramine-induced ex copula ejaculates (11) and fractionated in copula ejaculates were collected from each of 5 pony stallions for freezing in 5-ml straws (6), using a modified Kenney glucose skim-milk extender (2). Initial post-thaw total and progressive motilities and daily post-thaw total and progressive motilities, as well as the number of days to reach 0 progressively motile spermatozoa, were also similar for the 2 methods of collection. The percentage of morphologically normal spermatozoa both before freezing and after thawing were also similar for in copula and ex copula ejaculates. Co...
Campbell-Thompson M.Equine oxyntic mucosal cells were obtained by sequential exposure to pronase and collagenase. Acid production by parietal cells was assessed by uptake of [14C]aminopyrine (AP), a weak base that accumulates in intracellular acidic spaces. Incubation for various times revealed a maximal AP uptake at 10 minutes for histamine and carbachol. Similar secretagogue responses were observed for parietal cells from the mucosal cell preparation or after enrichment by elutriation. Histamine and isobutyl-methylxanthine (IBMX) stimulated AP uptake with a dose-dependent response and maximal effective concentr...
Braun J, Sakai M, Hochi S, Oguri N.The suitability of ejaculated and epididymal stallion spermatozoa for cooled storage (5 degrees C) and cryopreservation was examined in 5 ejaculates from each of 6 stallions and in spermatozoa recovered from the cauda epididymidis after castration of these stallions. The percentage of progressively motile spermatozoa, examined by subjective estimation (cooled samples) or by computerized analysis (frozen-thawed samples), was used as parameter. In ejaculated semen samples containing 5 and 25% seminal plasma in a skim milk glucose extender, the lower amount of seminal plasma supported spermatozoa...
Morris EA, Treadwell BV.The effect of interleukin 1 (IL-1) on equine articular cartilage was investigated, using a cartilage explant culture system. Measurement of [35S]O4 incorporation revealed synthesis of matrix proteoglycan by cartilage to be decreased 45, 59.7, and 37.5% after 1, 3, and 5 days, respectively, in culture in the presence of 5 U of IL-1/ml. There was no change in proteoglycan degradation as determined by measurement of [35S]O4 release into the culture medium. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of cartilage-conditioned medium indicated that exposure of cartilage to IL-1 caused ...
Miyamoto A, Nishio A.The vasomotor effects of histamine on isolated bovine and equine basilar arteries were examined. Histamine induced contractions in both these preparations. The maximal response to and pEC50 value for histamine of the equine artery were larger than those of bovine tissue. Similar results were obtained with endothelium-denuded basilar arteries. Diphenhydramine (H1-receptor antagonist) inhibited histamine-induced contractions of the basilar arteries from both species in a concentration-dependent manner and its pA2 values (with 95% confidence limits) were 7.61 (7.39-7.83) and 8.15 (8.01-8.29) for ...
Roser JF, McCue PM, Hoye E.An overnight double antibody RIA, employing a rabbit antiserum raised to bovine 31 kDa inhibin (rAs-#1989, NICHD) and purified bovine 31 kDa inhibin (bINH-I-90/1, NICHD) as trace and standard, was validated to measure immunoreactive inhibin (iINH) concentrations in equine peripheral plasma, follicular fluid (FF), ovarian vein (OV) plasma, testicular tissue extracts (TTE) and testicular vein (TV) plasma. The dynamic relationship of iINH and follicle stimulating hormone (FSH) was investigated during the estrous cycle of the mare and the annual reproductive cycle of the stallion. In the RIA, para...
Lebedev SG, Lebedeva LF.A medium containing egg yolk, mare's milk and/or modified PBS was used to culture Day-8 to 8.5 equine blastocysts. Twenty-one variants of the medium containing different concentrations of the 3 components were prepared. Embryos were recovered nonsurgically and placed into the media at 37 degrees C for 24 h. A total of 45 embryos was cultured; of these 7 died in culture and 13 showed inadequate development at the onset, while 25 continued to grow in the media. It was established that embryos grew best in media containing 20 to 60% yolk, 20 to 60% mare's milk and/or 20 to 60% PBS. It was found e...
Dippert KD, Jasko DJ, Seidel GE, Squires EL.Embryo recovery per ovulation has been shown to be lower in superovulated mares than in untreated controls. The objectives of this study were to 1) determine whether follicles stimulated with superovulatory treatment ovulate or luteinize without ovulation, 2) determine fertilization rates of oocytes in oviducts of superovulated and control mares, and 3) evaluate viability of early stage embryos from superovulated and control mares when cultured in equine oviductal cell-conditioned medium. Cyclic mares were randomly assigned to 1 of 2 groups (n=14 per group) on the day of ovulation (Day 0): Gro...
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...
Wagner B, Siebenkotten G, Leibold W, Radbruch A.Due to characteristic clinical signs, immunoglobulins of isotype E (IgE) are believed to be involved in several allergic diseases of the horse. To date, closer investigations have been hampered by the fact that neither purified equine IgE nor anti-equine IgE monoclonal antibodies were available for IgE isotype determination. As an approach to solve this problem, we constructed a stable cell line (EqE6) that expresses recombinant equi-murine IgE specific for 4-(hydroxy-3-nitro-phenyl) acetyl (NP). Biochemical analysis of the purified protein revealed a highly glycosilated IgE monomer of approxi...
Nakken G, Kirk J, Fjordbakk CT.Autologous conditioned serum (ACS), i.e serum enriched with anti-inflammatory cytokines and growth factors, is a popular orthobiologic therapy used in equine practice. Costly specialized tubes containing glass beads are commonly used for ACS production. The objective of this in vitro study was to compare cytokine and growth factor levels in equine serum after incubation in three different tubes: commercial plastic ACS tubes (COMM); sterile 50Â ml plastic centrifugation tubes (CEN); and 10Â ml plastic vacutainer tubes (VAC). Blood from 15 healthy horses was incubated in the different tubes at 3...
Klinc P, Kosec M, Majdic G.The success rate of artificial insemination following the freezing of stallion semen is limited; therefore, improving the stallion semen quality after the freezing and thawing process is a necessary objective. Objective: To investigate the influence of glass bead column separation on the freezability of stallion semen. Objective: Glass beads in a column separator remove damaged and dead spermatozoa in the ejaculate during centrifugation. Methods: In total, 50 ejaculates from 6 Lipizzaner stallions were studied. Each ejaculate was divided into 2 parts, one half processed following standard proc...
Schneider DA, Parks AH, Eades SC, Tackett RL.To compare in vitro smooth muscle relaxation of palmar digital vessels from healthy horses with those from horses in the prodromal stage of experimentally (carbohydrate) induced laminitis. Methods: 16 adult horses. Methods: Segments of palmar digital vessels were obtained from 5 healthy horses and 6 horses given carbohydrate. Vascular rings from the palmar digital artery and vein were suspended in individual organ baths containing buffer solution and indomethacin; isometric tension was recorded, and contraction and relaxation were compared. Smooth muscle contraction in response to cumulative a...
Watson ED, Aubrey ES, Zanecosky HG, Sertich PL.Glandular epithelial and stromal cells were isolated from the endometrium of mares by collagenase digestion and were incubated on plastic for 7-9 days until the cells formed confluent monolayers. The cells differed in morphology: epithelial cells appeared polyhedral and stromal cells were spindle like. The monolayers were incubated in the presence and absence of oxytocin. Medium was removed from wells after 2, 8 and 24 h of incubation. Concentrations of prostaglandin F (PGF) in the medium increased significantly during this time. Glandular epithelial cells produced significantly more PGF than ...
Hendrickson DA, Nixon AJ, Erb HN, Lust G.Equine neonatal chondrocytes were cultured in three-dimensional fibrin matrices under conditions of immediate implantation or implantation following monolayer culture for 6 days, and 3 cell concentrations (1 x 10(5), 1 x 10(6), and 5 x 10(6) chondrocytes/cm3). Equine fibrinogen was collected by cryoprecipitation and polymerized by use of activated bovine thrombin. The fibrin implants were harvested and analyzed histologically and biochemically at 3, 7, and 14 days after the chondrocytes were implanted in fibrin. The differentiation ratio (ratio of rounded, chondrocyte-like cells to stellate, f...
Clulow JR, Buss H, Evans G, Sieme H, Rath D, Morris LH, Maxwell WM.Sex-sorted, frozen-thawed stallion spermatozoa remain out of reach of commercial horse breeders because of the low efficiency of the sex-sorting process and unacceptable fertility rates after insemination. Two experiments were designed to test the effects of alternative staining and freezing media to improve the viability of sex-sorted frozen-thawed stallion spermatozoa. Experiment 1 compared two freezing media, INRA 82(®) and a modified lactose-ethylenediaminetetraacetic acid (EDTA), for the cryopreservation of sex-sorted stallion spermatozoa. No significant differences between the two freez...
Scotty NC, Evans TJ, Giuliano E, Johnson PJ, Rottinghaus GE, Fothergill AW, Cutler TJ.Lufenuron is a benzoylphenyl urea-derived insecticide that has been recently introduced as a novel treatment for fungal infections in horses. The purposes of this study were to determine (1) the in vitro efficacy of lufenuron against Aspergillus and Fusarium spp. and (2) the ability of lufenuron to reach efficacious blood concentrations after PO administration in horses. Fungal colonies isolated from diseased equine corneas were tested against lufenuron solutions up to 700 microg/mL. Twenty-one adult horses received 1 of 3 PO lufenuron treatment regimens: 5 mg/kg body weight (BW) q24h for 3 da...
Nunamaker DM, Richardson DW, Butterweck DM.Plate luting, a technique that uses polymethylmethacrylate (PMMA) interposed between the plate and the bone, as well as between the screw heads and the plate, to improve the stability of internal fixation was tested in vitro using 20 paired equine third metacarpal bones with mid-diaphyseal osteotomies plated with six-hole broad ASIF compression plates. One of each pair was luted and all specimens were tested in static overload or cyclic loading at 75% of the ultimate breaking strength using four-point bending. Although no differences were measured in static overload tests between luted and non...
Palma F, Ligi F, Soverchia C.This work is a comparative study of Na(+)-K+ and Ca(2+)-Mg2+ ATPase associated with the erythrocyte plasma membranes in different mammals. The method used to test the activity of these enzymes is based on quantitative measurements of ADP released during the reaction with HPLC: the chromatographic type is an Ion-Pair Reversed Phase. We have found that the levels of Ca2+ stimulated ATPase are higher than those of Na(+)-K+ ATPase in red blood cells of all the different mammalian species, with the only exception being lamb erythrocytes where the values of both the ATPase activities are almost equa...
Torres LEC, Florez CO, Oliveira JG, Vieira GD, Ribeiro IS, Keller KM, Leme FOP, Fantini P, Maranhão RPA.In equine ophthalmology, ulcerative keratitis is among the most common conditions and, in general, arises as a consequence of some trauma suffered. Secondarily, subsequent contamination by pathogenic or resident bacteria of the horse's ocular microbiota may have undesirable consequences. Under physiological conditions, the normal microbiota coexists with the immune status of the host, serving as a barrier, ensuring the health of the ocular surface, and inhibiting the proliferation of pathogens. However, in the imbalance of immune barriers, the normal microbiota can become pathogenic and lead t...
Saini S, Singha H, Siwach P, Tripathi BN.Interleukin (IL)-4 and IL-10 activate plethora of immune cells and induce the humoral immune response. However, recombinant version of horse IL-4 and IL-10 has not been investigated to understand their immunomodulating activities. This study aimed to produce recombinant horse mature IL-4 and IL-10 in . Immune-modulating activities of recombinant horse IL-4 and IL-10 were investigated in peripheral blood mononuclear cells (PBMCs). Methods: Equine PBMCs were stimulated with recombinant IL-4 and IL-10. A proliferation of PBMCs was measured by XTT assay and cytokines induction was measured by enzy...
Riemersma DJ, Lammertink JL.A calibration method is presented by which the signals of mercury-in-silastic strain gauges (MISS), implanted in the tendons of in vitro loaded equine hindlegs, were converted to tendon loads. The relationships between MISS-signals and tendon loads were obtained from tensile-force tests applied to the tendons. Special attention was paid to the correction of the MISS-signals for amplitude-shifts resulting from internal repositioning of the MISS after tendon isolation and temperature differences. Shift corrections equivalent to tendon strains up to 2.8% were necessary in the in vitro experiment....
O'Brien MA, Holmes MA, Duffus WP.Anti-tetanus toxoid (TT) antibody (Ig) levels in the supernatant of cultured, pre-immunised equine peripheral blood mononuclear cells (PBMC) were measured by an indirect enzyme-linked immunoabsorbent assay (ELISA). Optimal anti-TT Ig production occurred at concentrations of stimulating, purified TT of between 0.001 and 0.1 micrograms ml-1, which varied depending on the cell concentration. Optimal anti-TT Ig production was most consistently produced when the cell concentration was 5 x 10(6) ml-1. At this cell concentration maximal anti-TT Ig was induced using 0.1 micrograms ml-1 TT. At a cell c...
Lindmark DG, Jarroll EL, Timoney PJ, Shin SJ.The energy metabolism of the English E-CMO strain of contagious equine metritis bacterium was studied in whole cells and cell extracts. This bacterium appears to have an active Krebs cycle and probably obtains energy by oxidative phosphorylation since glycolysis and the hexose monophosphate pathways appear to be absent. These conclusions are based on the findings that [U-14C]glucose incorporation by this bacterium is below the level of detection, and that respiration is stimulated by Krebs cycle intermediates (i.e., malate, citrate, and succinate), but not by glucose, fructose, maltose, or suc...
Schirmer EC, Farooqui J, Polak PE, Szuchet S.Cell-substratum adhesion plays a crucial part in the cascade of events that control growth or turn on and consummate a differentiation program. We are investigating the molecular basis of oligodendrocyte (OLG) cytodifferentiation, employing pure cultures of OLGs isolated from postmyelination brains. We have shown that such OLGs will regenerate in vitro and reenact the ontogenic development of myelin, but to do so they need a signal. Adherence to a polylysine surface in the presence of 20% horse serum generates such a signal. Among the events that are turned on upon OLG adhesion is the phosphor...
Millard JT, Chuang E, Lucas JS, Nagy EE, Davis GT.A simple and robust biochemistry laboratory experiment is described that uses restriction fragment length polymorphism (RFLP) of polymerase chain reaction (PCR) products to verify the identity of a potentially valuable horse. During the first laboratory period, students purify DNA from equine samples and amplify two loci of mitochondrial DNA. During the second laboratory period, students digest PCR products with restriction enzymes and analyze the fragment sizes through agarose gel electrophoresis. An optional step of validating DNA extracts through realtime PCR can expand the experiment to th...
May SA, Hooke RE, Lees P.There is increasing evidence that the proteoglycan-degrading neutral metalloproteinase, stromelysin, is a key enzyme in the pathogenesis of osteoarthrosis. Equine synovial lining cells were stimulated in vitro to produce stromelysin, and phenylbutazone, flunixin, betamethasone, sodium hyaluronate and polysulphated glycosaminoglycan (PSGAG) were tested for their ability to inhibit the action of this enzyme on 14C-labelled casein substrate. Only PSGAG possessed inhibitory activity at concentrations likely to be achieved therapeutically in the equine fetlock joint.
Wang ZW, Yu MF, Robinson NE.Muscarinic autoreceptors on horse airway cholinergic nerves were studied by examining the effects of muscarinic receptor antagonists on electrical field stimulation (EFS)-induced acetylcholine (ACh) release in trachealis preparations. All the antagonists including atropine (non-selective), pirenzepine (M1-selective), AF-DX 116 (M2-selective), and hexahydrosiladifenidol (M3-selective) augmented ACh release concentration-dependently. The augmentation was not due to displacement of ACh molecules from tissue receptors into the bath liquid because incubation with atropine after EFS had no influence...
Zibura AE, Cullen MA, Rutledge H, Lassalle L, Salmon JH, Gilger BC, Westermeyer HD.Determine optimal iontophoresis times for riboflavin delivery to the corneal stroma across different species and compare these to corneal injection. Methods: Ex vivo horse, dog, rabbit, and pig globes were treated with riboflavin administered with either iontophoresis for 2.5-20Â minutes with or without corneal epithelium; or with purpose-designed precise corneal injection (PCI) application with intact epithelium. Immediately following riboflavin administration, samples were harvested, frozen, and sectioned. Riboflavin penetration was imaged using fluorescence microscopy. Results: Horse sample...
Gillis C, Sharkey N, Stover SM, Pool RR, Meagher DM, Willits N.Ultrasonographic cross sectional area (CSA) measurements of equine superficial digital flexor (SDF) tendon were obtained to determine the feasibility of ultrasonography for CSA measurement of tendon in vivo and in vitro. Ultrasonographic measurements were compared with a more traditional CSA measurement method, ink-blot analysis. In addition, values for ultrasonographic SDF tendon mean echogenicity were obtained in vivo and in vitro. The left forelimb SDF tendons of 23 horses were evaluated ultrasonographically. Cross sectional images were acquired at 4-cm intervals distal to the base of the a...
Bass LD, Denniston DJ, Maclellan LJ, McCue PM, Seidel GE, Squires EL.Equine embryos (n=43) were recovered nonsurgically 7-8 days after ovulation and randomly assigned to be cryopreserved in one of two cryoprotectants: 48% (15M) methanol (n=22) or 10% (136 M) glycerol (n=21). Embryos (300-1000 microm) were measured at five intervals after exposure to glycerol (0, 2, 5, 10 and 15 min) or methanol (0, 15, 35, 75 and 10 min) to determine changes (%) in diameter over time (+/-S.D.). Embryos were loaded into 0.25-ml plastic straws, sealed, placed in a programmable cell freezer and cooled from room temperature (22 degrees C) to -6 degrees C. Straws were then seeded, h...
Knych HK, McKemie DS, Kass PH, Stanley SD, Blea J.Ketoprofen is an anti-inflammatory drug that is commonly administered to racehorses for the alleviation of musculoskeletal pain and inflammation. This study represents a comprehensive examination of the metabolism (in vivo and in vitro), pharmacokinetics and ex vivo pharmacodynamics, of ketoprofen in horses. The in vitro metabolism as well as specific enzymes responsible for metabolism was determined by incubating liver microsomes and recombinant CYP450 and UGT enzymes with ketoprofen. For the in vivo portion, 15 horses were administered a single intravenous dose of 2.2-mg/kg ketoprofen. Blood...
Barton MH, Hurley D, Norton N, Heusner G, Costa L, Jones S, Byars D, Watanabe K.Lactoferrin is a colostral glycoprotein with antimicrobial properties. Objective: (1) Serum lactoferrin and immunoglobulin G (IgG) concentrations are correlated and increase in healthy foals after ingestion of colostrum; (2) compared to healthy foals, ill foals will have lower lactoferrin concentrations that correlate with their IgG concentration, neutrophil count, the diagnosis of sepsis, and survival; and (3) plasma concentrations of lactoferrin will be less than serum concentrations. Methods: Healthy foals (n = 16), mature horses (n = 10), and ill foals 1-4 days old (n = 111) that were exam...
Frontiers in allergyNovember 26, 2021
Volume 2 762026 doi: 10.3389/falgy.2021.762026
Ben Hamouda S, Miglino MA, de Sá Schiavo Matias G, Beauchamp G, Lavoie JP.Asthma is associated with increased deposition and altered phenotype of airway smooth muscle (ASM) cells. However, little is known about the processes responsible for these changes. It has been suggested that alterations of the extracellular matrix (ECM) contribute to the remodeling of ASM cells in asthma. Three-dimensional matrices allow the study of complex cellular responses to different stimuli in a close-to-natural environment. Thus, we investigated the ultrastructural and genic variations of ASM cells cultured on acellular asthmatic and control bronchial matrices. We studied horses, as ...
Patan-Zugaj B, Gauff FC, Egerbacher M, Licka TF.OBJECTIVE To investigate the effect of lipopolysaccharide (LPS) on type VII collagen- cleaving matrix metalloproteinases (MMPs) in the lamellar tissue of extracorporeally perfused equine limbs. SAMPLE 10 right forelimbs and 3 left forelimbs collected from 10 adult horses after slaughter at a licensed abattoir. PROCEDURES Extracorporeal perfusion of the isolated equine limbs was performed for 10 hours under physiologic conditions (control-perfused limbs; n = 5) and with the addition of 80 ng of LPS/L of perfusate (LPS-perfused limbs; 5). Lamellar tissue specimens were then collected from the do...
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...
Murray A, Pearson GT, Cottrell DF.Small intestine was taken from the caudal flexure of the duodenum and the terminal ileum proximal to the ileocaecal fold of 25 horses, 9 with acute grass sickness (AGS), 12 with subacute grass sickness (SAGS) and 12 with chronic grass sickness (CGS). The motility in the samples was measured isometrically either within 1 h of death or after storage for 24 h at 4 degree C. In control tissue, noradrenaline produced contractions of muscle strips which did not involve a muscarinic cholinergic mechanism and which were unaffected by the alpha 1 antagonist prazosin but were blocked by the alpha 2 anta...
