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.
Adams MK, Goodrich LR, Rao S, Olea-Popelka F, Phillips N, Kisiday JD, McIlwraith CW.The 2 sites of bone marrow harvest for isolation of mesenchymal stromal cells (MSC) in the horse are the sternum and ilium. The technical procedure is based on practitioner preference, but no studies have compared MSC concentrations and growth rates between the sites in horses aged 2-5 years. Objective: The objective of this study was to compare nucleated cell counts and growth rates between the sternum and ilium and between consecutive 5 ml bone marrow aspirates. We hypothesised that there would be a higher concentration of MSCs in the sternum than the ilium, and that the first sequential a...
Ceusters JD, Serteyn DA, Minguet G, de la Rebière de Pouyade G, Romainville J, Deby-Dupont GP, Mouithys-Mickalad AA, Franck TJ.Horses are particularly sensitive and exposed to excessive inflammatory responses evolving toward an important stimulation of polymorphonuclear neutrophils (PMNs). The aim of this work was to stimulate equine neutrophils in whole blood and to evaluate their response by measuring the release of total and active myeloperoxidase (MPO) and total elastase, considered as markers of neutrophil stimulation and degranulation. Because of the critical importance of the concomitant presence of LPS and TNF-α in equine pathological situations, we combined these two natural mediators to stimulate PMN and co...
Ho EN, Kwok WH, Lau MY, Wong AS, Wan TS, Lam KK, Schiff PJ, Stewart BD.A veterinary preparation known as TB-500 and containing a synthetic version of the naturally occurring peptide LKKTETQ has emerged. The peptide segment (17)LKKTETQ(23) is the active site within the protein thymosin β(4) responsible for actin binding, cell migration and wound healing. The key ingredient of TB-500 is the peptide LKKTETQ with artificial acetylation of the N-terminus. TB-500 is claimed to promote endothelial cell differentiation, angiogenesis in dermal tissues, keratinocyte migration, collagen deposition and decrease inflammation. In order to control the misuse of TB-500 in equin...
Garner DL, Evans KM, Seidel GE.The sex of mammalian offspring can be predetermined by flow sorting relatively pure living populations of X- and Y-chromosome-bearing sperm. This method is based on precise staining of the DNA of sperm with the nucleic acid-specific fluorophore, Hoechst 33342, to differentiate between the subpopulations of X- and Y-sperm. The fluorescently stained sperm are then sex-sorted using a specialized high speed sorter, MoFlo(®) SX XDP, and collected into biologically supportive media prior to reconcentration and cryopreservation in numbers adequate for use with artificial insemination for some specie...
Durán MC, Willenbrock S, Carlson R, Feige K, Nolte I, Murua Escobar H.CD14 positive (CD14+) cells are the precursor cells of monocyte-derived dendritic cells (DCs). In horses their potent antigen-presenting capacity and ability to induce an effective immune response classify these cells suitable for several therapeutic approaches such as for equine sarcoid. However, in horses, the generation efficiency of DCs from adherent peripheral blood mononuclear cells (PBMCs) is currently still poor. Objective: Establishment of a simple short protocol to enhance DC generation in horses by using a human CD14 monoclonal antibody (mAb) and an automated magnetic activated cell...
Fossum C, Hjertner B, Olofsson KM, Lindberg R, Ahooghalandari P, Camargo MM, Bröjer J, Edner A, Nostell K.The expression of tlr4, md2 and cd14 was studied in equine blood leukocytes and in intestinal samples using real time PCR. The stability of three commonly used reference genes, glyceraldehyde-3P-dehydrogenase (GAPDH), hypoxantine ribosyltransferase (HPRT) and succinate dehydrogenase complex subunit A (SDHA), was evaluated using qbase(PLUS). The equine peripheral blood mononuclear cells (eqPBMC) examined were either stimulated in vitro with Phorbol 12-myristate 13-acetate (PMA) and ionomycin or with the CpG oligodeoxynuclotide 2216 (CpG-ODN 2216) or obtained from horses before, during and after...
Tydén E, Löfgren M, Hakhverdyan M, Tjälve H, Larsson P.In the present study, we examined the gene expression of cytochrome P450 3A (CYP3A) isoenzymes in the tracheal and bronchial mucosa and in the lung of equines using TaqMan probes. The results show that all seven CYP3A isoforms identified in the equine genome, that is, CYP3A89, CYP3A93, CYP3A94, CYP3A95, CYP3A96, CYP3A97 and CYP3A129, are expressed in the airways of the investigated horses. Though in previous studies, CYP3A129 was found to be absent in equine intestinal mucosa and liver, this CYP3A isoform is expressed in the airways of horses. The gene expression of the CYP3A isoenzymes varied...
Lange-Consiglio A, Arrighi S, Fiandanese N, Pocar P, Aralla M, Bosi G, Borromeo V, Berrini A, Meucci A, Dell'Aquila ME, Cremonesi F.There is no published information about follicular-fluid leptin concentrations or the presence of leptin and leptin receptor in the equine ovary or oocyte. Three groups of mares - adult draft mares, draft fillies and adult Standardbred mares - were included in the study. Leptin and leptin receptor were detected in all immature oocytes by immunofluorescence with higher intensity in oocytes from draft mares compared with draft fillies and Standardbred mares. After in vitro maturation a higher proportion of oocytes reached metaphase II in draft mares than in draft fillies and Standardbred mares, ...
Andersen UV, Haakansson IT, Roust T, Rhod M, Baptiste KE, Nielsen MK.Strongyle and trichostrongyle parasites are ubiquitous nematodes of grazing livestock. Several molecular diagnostic tests are based upon measuring and quantifying DNA obtained from parasite eggs. It is well known that such eggs undergo development during storage, but it remains unknown to which extent developmental stages can affect the variation of diagnostic test results. This study investigated the influence of developmental stages of strongyle eggs on the variation real-time polymerase chain reaction (PCR) results. Mixed species strongyle eggs were obtained from the faeces of a naturally i...
Salama AA, Aboulaila M, Moussa AA, Nayel MA, El-Sify A, Terkawi MA, Hassan HY, Yokoyama N, Igarashi I.Fusidic acid known to has antibacterial, antifungal, and antimalarial activities. Fusidic acid blocks translation elongation factor G gene in Plasmodium falciparum. In the present study, the inhibitory effects of fusidic acid on the in vitro growth of bovine and equine Babesia parasites were evaluated. The inhibitory effect of fusidic acid on the in vivo growth of Babesia microti was also assessed. The in vitro growth of four Babesia species that were tested was significantly inhibited (P<0.05) by micromolar concentrations of fusidic acid (IC(50) values=144.8, 17.3, 33.3, and 56.25 μM for ...
Keen JA, McGorum BC, Hillier C, Nally JE.This study investigated the effects of cortisol and insulin, hormones that affect both glycaemic status and vascular function, on the in vitro contractility of isolated healthy equine small laminar veins. Small veins (150-500 μm) draining the digital laminae from healthy horses or ponies were investigated by wire myography. Concentration response curves were constructed for noradrenaline (NA), phenylephrine (PE), endothelin-1 (ET-1) and 5-hydroxytryptamine (5-HT) in the presence of either cortisol (10(-6 ) m) or insulin (1000 μIU/mL). Cortisol significantly increased the maximum contractilit...
Muttini A, Valbonetti L, Abate M, Colosimo A, Curini V, Mauro A, Berardinelli P, Russo V, Cocciolone D, Marchisio M, Mattioli M, Tosi U....In vitro expanded and frosted ovine amniotic epithelial cells (oAECs) were evaluated for their phenotype, stemness and attitude to differentiate into tenocytes. Fifteen horses with acute tendon lesions were treated with one intralesional injection of oAECs. Tendon recovery under controlled training was monitored. In vitro expanded oAECs showed a constant proliferative ability, a conserved phenotype and stable expression profile of stemness markers. Differentiation into tenocytes was also regularly documented. US controls showed the infilling of the defect and early good alignment of the fibers...
Vallance SA, Vidal MA, Whitcomb MB, Murphy BG, Spriet M, Galuppo LD.To evaluate use of a diode laser to induce tendinopathy in the superficial digital flexor tendon (SDFT) of horses. Methods: 4 equine cadavers and 5 adult horses. Methods: Cadaveric SDFT samples were exposed to a diode laser at various energy settings to determine an appropriate energy for use in in vivo experiments; lesion size was assessed histologically. In vivo experiments involved laser energy induction of lesions in the SDFT (2 preliminary horses [0, 25, 75, and 87.5 J] and 3 study horses [0 and 125 J]) and assessment of lesions. Study duration was 21 days, and lesions were assessed clini...
Go YY, Bailey E, Timoney PJ, Shuck KM, Balasuriya UB.We investigated the correlation between in vitro susceptibility of CD3(+) T lymphocytes to equine arteritis virus (EAV) infection and establishment of persistent infection among 14 stallions following natural infections. The data showed that carrier stallions with a CD3(+) T lymphocyte susceptibility phenotype to in vitro EAV infection may be at higher risk of becoming carriers than those that lack this phenotype (P = 0.0002).
Kuhn M, Guschlbauer M, Feige K, Schluesener M, Bester K, Beyerbach M, Breves G.Equine typholocolitis is a sporadic diarrheal disease causing high mortality rates. One of the risk factors responsible for this is the oral application of the macrolide antibiotic erythromycin. The aim of the present in vitro study was to investigate whether erythromycin in combination with feed restriction provokes changes in microbial hindgut metabolism and could therefore be involved in the pathogenesis of equine typhlocolitis. As application of erythromycin and feed restriction are risk factors for equine typhlocolitis, both factors were chosen to investigate their individual and combined...
Beckelmann J, Budik S, Bartel C, Aurich C.Until now, sex determination in equine embryos has been performed by detection of Y-chromosome-specific sequences only. In the present study, expression of a Barr-body-specific marker, the X-inactivated-specific transcript (Xist) gene, whose gene product consists of RNA which coats and thereby inactivates one of the X chromosomes, was investigated in equine embryos produced in vivo. Preattachment embryos at different times after ovulation (Day 8: n = 9; Day 10: n = 12; Day 12: n = 15) were analyzed for Xist RNA expression using quantitative and qualitative reverse transcription-polymerase chai...
Ranera B, Remacha AR, Álvarez-Arguedas S, Romero A, Vázquez FJ, Zaragoza P, Martín-Burriel I, Rodellar C.Mesenchymal stem cells (MSCs) derived from bone marrow (BM-MSCs) and adipose tissue (AT-MSCs) are being applied to equine cell therapy. The physiological environment in which MSCs reside is hypoxic and does not resemble the oxygen level typically used in in vitro culture (20% O2). This work compares the growth kinetics, viability, cell cycle, phenotype and expression of pluripotency markers in both equine BM-MSCs and AT-MSCs at 5% and 20% O2. Results: At the conclusion of culture, fewer BM-MSCs were obtained in hypoxia than in normoxia as a result of significantly reduced cell division. Hypoxi...
Azab W, El-Sheikh A.Equine herpesvirus 4 (EHV-4) is an important equine pathogen that causes respiratory tract disease among horses worldwide. Glycoprotein K (gK) homologues have been identified in several alphaherpesviruses as a major player in virus entry, replication, and spread. In the present study, EHV-4 gK-deletion mutant has been generated by using bacterial artificial chromosome technology and Red mutagenesis to investigate the role of gK in EHV-4 replication. Our findings reported here show that gK is essential for virus replication in vitro and that the gK-negative strain was not able to be reconstitut...
Bidaud P, Hébert L, Barbey C, Appourchaux AC, Torelli R, Sanguinetti M, Laugier C, Petry S.Rhodococcus equi is one of the most widespread causes of disease in foals aged from 1 to 6 months. R. equi possesses antioxidant defense mechanisms to protect it from reactive oxygen metabolites such as hydrogen peroxide (H(2)O(2)) generated during the respiratory burst of phagocytic cells. These defense mechanisms include enzymes such as catalase, which detoxify hydrogen peroxide. Recently, an analysis of the R. equi 103 genome sequence revealed the presence of four potential catalase genes. We first constructed ΔkatA-, ΔkatB-, ΔkatC-and ΔkatD-deficient mutants to study the ability of R. ...
Echigoya Y, Morita S, Itou T, Sakai T.To evaluate effects of extracellular lactate on viability, shape change, lactate metabolism, and reactive oxygen species (ROS) production in equine polymorphonuclear leukocytes (PMNs). Methods: PMNs isolated from equine venous blood samples. Methods: PMNs were incubated with 0 to 300mM lactate for 30 minutes before each experiment. Viability was assessed via trypan blue exclusion. Shape change was assessed via flow cytometry and light microscopy. Relative quantification of monocarboxylic acid transporter and lactate dehydrogenase lactate dehydrogenase (LDH) isotype mRNAs was performed with a r...
Mason JB, Vandenberghe LH, Xiao R, Wilson JM, Richardson DW.To evaluate transduction efficiency of gene therapy for treatment of osteoarthritis in horses. Methods: Cartilage and synovial tissues were aseptically collected from the stifle joints of 3 Thoroughbreds; horses were 3, 7, and 12 years old and free from sepsis and long-term drug treatment and were euthanized for reasons unrelated to joint disease. Methods: Gene transfer experiments were performed with 8 recombinant adeno-associated viral vector (rAAV) serotypes in monolayer-cultured equine chondrocytes, synovial cells, and mesenchymal stromal cells and in cartilage and synovial tissues. Result...
Turner RM, Zeng W.Studies in laboratory rodents are shedding light on the pathophysiology of testicular ageing and now suggest a complicated basis for age-related declines in testicular function. A highly significant contributor to infertility may involve failure of specific and complex testicular microenvironments (niches) comprised of a variety of cellular and molecular components. Our laboratory has applied testis tissue xenografting to the study of testicular ageing in the stallion. Using this technique, we have confirmed that the disease is tissue autologous. As would be expected from a tissue autologous d...
Parera M, Martrus G, Franco S, Clotet B, Martinez MA.Canine hepacivirus (CHV) was recently identified in domestic dogs and horses. The finding that CHV is genetically the virus most closely related to hepatitis C virus (HCV) has raised the question of whether HCV might have evolved as the result of close contact between dogs and/or horses and humans. The aim of this study was to investigate whether the NS3/4A serine protease of CHV specifically cleaves human mitochondrial antiviral signaling protein (MAVS) and Toll-IL-1 receptor domain-containing adaptor inducing interferon-beta (TRIF). The proteolytic activity of CHV NS3/4A was evaluated using ...
González-Fernández L, Macías-García B, Velez IC, Varner DD, Hinrichs K.The mechanisms leading to capacitation in stallion sperm are poorly understood. The objective of our study was to define factors associated with regulation of protein tyrosine phosphorylation in stallion sperm. Stallion sperm were incubated for 4 h in modified Whitten's media with or without bicarbonate, calcium, or BSA. When sperm were incubated in air at 30×10⁶/ml at initial pH 7.25, protein tyrosine phosphorylation was detected only in medium containing 25 mM bicarbonate alone; calcium and BSA inhibited phosphorylation. Surprisingly, this inhibition did not occur when sperm were incubate...
Lagares MA, Martins HS, Carvalho IA, Oliveira CA, Souza MR, Penna CF, Cruz BC, Stahlberg R, Henry MR.Extenders with a defined composition containing only components with clearly protective effects on sperm during storage would be an advantage. The aims of the present work were to assess whether caseinate, improves cooled and frozen equine semen quality. Semen from six stallions were suspended with four different cooling extenders C1) Kenney extender; C2) 0.6 % caseinate; C3) 2.7 % caseinate ; and C4) C1 + 2.1 % caseinate, and frozen extenders: F1) INRA 82 extender; F2) 1.35 % caseinate; and F3) 2.7 % caseinate. Although there was no significant difference between the motility rate among the c...
Gambini A, Jarazo J, Olivera R, Salamone DF.The production of cloned equine embryos remains highly inefficient. Embryo aggregation has not yet been tested in the equine, and it might represent an interesting strategy to improve embryo development. This study evaluated the effect of cloned embryo aggregation on in vitro and in vivo equine embryo development. Zona-free reconstructed embryos were individually cultured in microwells (nonaggregated group) or as 2- or 3-embryo aggregates (aggregated groups). For in vitro development, they were cultured until blastocyst stage and then either fixed for Oct-4 immunocytochemical staining or maint...
Albert R, Vásárhelyi G, Bodó G, Kenyeres A, Wolf E, Papp T, Terdik T, Módis L, Felszeghy S.One of the most promising applications for the restoration of small or moderately sized focal articular lesions is mosaicplasty (MP). Although recurrent hemarthrosis is a rare complication after MP, recently, various strategies have been designed to find an effective filling material to prevent postoperative bleeding from the donor site. The porous biodegradable polymer Polyactive (PA; a polyethylene glycol terephthalate - polybutylene terephthalate copolymer) represents a promising solution in this respect. A histological evaluation of the longterm PA-filled donor sites obtained from 10 exper...
Vidale P, Magnani E, Nergadze SG, Santagostino M, Cristofari G, Smirnova A, Mondello C, Giulotto E.Many human primary somatic cells can be immortalized by inducing telomerase activity through the exogenous expression of the human telomerase catalytic subunit (hTERT). This approach has been extended to the immortalization of cell lines from several mammals. Here, we show that hTERT expression is not sufficient to immortalize primary fibroblasts from three equid species, namely donkey, Burchelli's zebra and Grevy's zebra. In vitro analysis of a reconstituted telomerase composed by hTERT and an equid RNA component of telomerase (TERC) revealed a low activity of this enzyme compared to human te...
Lindahl J, Dalin AM, Stuhtmann G, Morrell JM.One of the challenges faced by equine breeders is ensuring delivery of good quality semen doses for artificial insemination when the mare is due to ovulate. Single Layer Centrifugation (SLC) has been shown to select morphologically normal spermatozoa with intact chromatin and good progressive motility from the rest of the ejaculate, and to prolong the life of these selected spermatozoa in vitro. The objective of the present study was a proof of concept, to determine whether fertilizing ability was retained in SLC-selected spermatozoa during prolonged storage. Results: Sixteen mares were insemi...
Kol A, Walker NJ, Galuppo LD, Clark KC, Buerchler S, Bernanke A, Borjesson DL.Autologous cellular therapy products including adipose-derived stromal vascular fraction (SVF), bone marrow mononuclear cells (BMMNs), cord blood mononuclear cells (CBMNs) and platelet rich plasma are options for treatment of acute orthopaedic lesions while mesenchymal stem cells (MSCs) are culture expanded. These products may contribute to healing by secreting matrix proteins or growth factors, but they may also act on endogenous MSCs to facilitate healing. Objective: To determine the effects of cell therapy products on MSCs function in vitro. The hypothesis was that cell therapy products pro...
Cary JA, Madill S, Farnsworth K, Hayna JT, Duoos L, Fahning ML.The purpose of this study was to evaluate 2 methods of semen collection that could be used as terminal procedures in stallions with irreparable conditions, such as fractures or colic. Electroejaculation was attempted under general anesthesia. Forty-eight hours later, the ponies were castrated and 2 different epididymal sperm collection techniques were attempted by using a flushing or floating method. Additionally, the effect of supplemental seminal plasma was evaluated. Experimentally, electroejaculation was found to be a safe but ineffective method of terminal semen collection. Viable sperm c...
Volokhov DV, Gao Y, Davidson MK, Chizhikov VE.We describe two novel species of Acholeplasma sp. strain N93 and Mycoplasma sp. strain LR5794 which were isolated from the nasopharynx of a horse from the United Kingdom and from the oral cavity of a North American raccoon from Canada, respectively. These strains were phenotypically and genetically characterized and compared to other established Mycoplasma and Acholeplasma species. Both strains are facultative anaerobes, resistant to penicillin, and produce acid from glucose but do not hydrolyze arginine and urea. Both strains grew well in microaerophilic and anaerobic atmospheric conditions a...
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...
Stout TA.During the past 15 years, embryo transfer (ET) has become increasingly widespread within the sport-horse breeding industry. At present, however, the vast majority (>95%) of horse embryos are transferred fresh or after chilled storage for up to 24 h, whereas cryopreservation is rarely employed despite its obvious potential for simplifying recipient mare management and facilitating long-term storage and international transport of embryos. A number of inter-related factors have contributed to the slow development and implementation of equine embryo cryopreservation, and these include the followin...
Miszczak F, Shuck KM, Lu Z, Go YY, Zhang J, Sells S, Vabret A, Pronost S, Fortier G, Timoney PJ, Balasuriya UB.This study showed that under specifically defined conditions with respect to nucleic acid extraction method and testing reagents, a previously described real-time reverse transcription-PCR (rRT-PCR) assay (T1 assay) provides sensitivity equal to or higher than that of virus isolation for the detection of equine arteritis virus in semen.
Albrizio M, Guaricci AC, Maritato F, Sciorsci RL, Mari G, Calamita G, Lacalandra GM, Aiudi GG, Minoia R, Dell'Aquila ME, Minoia P.The development of fertilizing ability in sperm cells is associated with changes in the plasma membrane. However, to date the exact nature of sequentially activated primary receptors and channels and the signal transduction pathways derived from these remains elusive. We analyzed the expression and localization of the mu-opioid receptor in equine spermatozoa. A transcript corresponding to the third extracellular loop that selectively binds mu agonists was amplified, sequenced and compared with the known sequences in humans, rats and cattle. The amplification product showed a high degree of nuc...
Bulkeley E, Santistevan AC, Varner D, Meyers S.Low levels of intracellular reactive oxygen species (ROS) are essential for normal sperm function and are produced by sperm mitochondria as a byproduct of metabolism, but in excess, ROS can cause catastrophic cellular damage and has been correlated with infertility, poor sperm motility and abnormal morphology in humans. Stallion sperm motility is fueled predominantly by oxidative phosphorylation-produced ATP, requiring high basal rates of mitochondrial function. Consequently, whether elevated ROS production by stallion sperm is an indicator of dysfunctional or highly motile cells has been deba...
Thompson RE, Johnson AK, Prado TM, Premanandan C, Brown ME, Whitlock BK, Pukazhenthi BS.Availability of viable frozen-thawed endometrial tissues could facilitate detailed studies into physiologic and disease processes influencing the endometrium. This study was designed to investigate the cryosurvival of equine endometrial tissue. Previous studies in the human and horse have focused on cryopreservation of dissociated endometrial cells. To our knowledge, there are no studies on cryopreservation of endometrial explants. Our objectives were to 1) determine the influence of differing concentrations of the permeating cryoprotectant dimethyl sulfoxide (MeSO) on viability, structural in...
Crawford PC, Hanel RM, Levy JK.To evaluate equine IgG as a treatment for kittens with failure of passive transfer of immunity (FPT). Methods: 13 specific pathogen-free queens and their 77 kittens. Methods: Kittens were randomized at birth into 9 treatment groups. One group contained colostrum-fed (nursing) kittens; the other groups contained colostrum-deprived kittens that were administered supplemental feline or equine IgG PO or SC during the first 12 hours after birth. Blood samples were collected at serial time points from birth to 56 days of age for determination of serum IgG concentrations. The capacity of equine IgG t...
Erol E, Shaffer CL, Lubbers BV.The alarming increase in rifampin and macrolide resistance among Rhodococcus equi isolates highlights the need to identify alternative therapeutic options that can effectively control rhodococcosis in foals while limiting the further development of drug resistance. Objective: To evaluate bacterial killing, antibiotic synergism and mutant prevention concentrations (MPCs) of clarithromycin alone and in combination with doxycycline, minocycline or rifampin against clinical isolates of R equi. Methods: In vitro experiments. Methods: Bacterial time-kill, fractional inhibitory concentration (checker...
Cook VL, Holcombe SJ, Gandy JC, Corl CM, Sordillo LM.Monocytes are among the initial cells that interact with circulating LPS. Binding of LPS to monocyte surface receptors triggers an intracellular signaling cascade and results in the production of proinflammatory cytokines. Ethyl pyruvate, a stable derivative of pyruvate, has been effective in mitigating LPS induced alterations in isolated human monocytes. We hypothesized that ethyl pyruvate would suppress proinflammatory gene expression in LPS-stimulated equine monocytes without affecting cell viability. Equine monocytes were isolated from whole blood using a sediment-gradient centrifugation p...
Marshall K, Marsella R.Increased antimicrobial resistance highlights the need for alternatives to antibiotics. Bacteriophages, which are benign viruses that kill bacteria, are promising. We studied the efficacy of topical bacteriophages for treating equine staphylococcal superficial pyodermas. Eight isolates were tested against a bacteriophage bank, and a cocktail consisting of two bacteriophages was prepared. Twenty horses with clinical and cytological evidence of superficial pyoderma and confirmed infection based on swabbed culture were enrolled in the study. Each horse received both the bacteriophage cocktail a...
Griffin RA, Swegen A, Baker MA, Ogle RA, Smith N, Aitken RJ, Skerrett-Byrne DA, Fair S, Gibb Z.Stallions experience transient fluctuations in fertility throughout the breeding season. Considering pregnancy diagnoses cannot be ascertained until ~14 days postbreeding, the timely detection of decreases in stallion fertility would enhance industry economic and welfare outcomes. Therefore, this study aimed to identify the proteomic signatures reflective of short-term fertility fluctuations and to determine the biological mechanisms governing such differences. Using liquid chromatography-mass spectrometry (LC-MS/MS), we compared the proteomic profile of semen samples collected from commercia...
Grillos AS, Roach JM, de Mestre AM, Foote AK, Kinglsey NB, Mienaltowski MJ, Bellone RR.Warmblood Fragile Foal Syndrome Type 1 (WFFS) is an autosomal recessive disorder reported previously only in warmbloods and thought to be caused by a variant in the gene procollagen-lysine,2-oxoglutarate 5-dioxygenase 1 (PLOD1, c.2032G>A, p.Gly678Arg). Given the presentation of this Thoroughbred case, we hypothesised that a similar genetic mechanism caused this phenotype. Objective: To describe the pathological and genetic findings on a foal presenting to a veterinary practice in the UK with skin lesions similar to other Ehlers-Danlos Syndromes, including those documented for warmbloods with W...
Bijman J, Quinton PM.Single equine sweat glands were found to secrete for more than 1 h in vitro in response to pharmacologic secretagogues. The adrenergic agonists epinephrine and norepinephrine evoked maximal sweat rates of 2.0 nl X gland-1 X min-1. However, the concentration of norepinephrine (10(-5) M) required to evoke the maximal response was 10 times higher than that for epinephrine. Maximal sweat rates also were stimulated with the beta 2-adrenergic agonist terbutaline. This stimulation was blocked by the beta-adrenergic antagonist propranolol. Moderate sweating responses were also obtained with the alpha-...
Hudson N, Mayhew I, Pearson G.Equine dysautonomia (grass sickness) is a frequently fatal disease of horses characterised by intestinal stasis. Interstitial cells of Cajal (ICC) are the pacemakers and mediators of neurotransmission in the gastrointestinal tract. Impaired ICC-mediated control of motility has been implicated in intestinal disorders in laboratory mammals, humans and in equine grass sickness. The aim of this study was to compare the in vitro electrical properties of ileum from grass sickness cases with horses free from gastrointestinal disease. Intracellular microelectrode recordings were made from smooth muscl...
Brommer H, Laasanen MS, Brama PA, van Weeren PR, Helminen HJ, Jurvelin JS.To evaluate an arthroscopic indentation instrument (Artscan 200) for assessment of the health status of equine articular cartilage. Methods: In vitro experiment using equine isolated proximal phalanx (P1) specimens. Methods: P1 specimens from 39 horses (aged 1.5-22 years). Methods: Reproducibility was tested by determination of the coefficient of variation (CV). Dynamic modulus and cartilage degeneration index (CDI) values were measured at 2 predefined sites (site 1, joint margin; site 2, joint center) to assess the accuracy and to evaluate the relation with surface integrity. Results: CV was ...
Dias GM, Retamal CA, Tobella L, Arnholdt AC, López ML.'The highly packed chromatin of mature spermatozoa results from replacement of somatic-like histones by highly basic arginine- and cysteine-rich protamines during spermatogenesis, with additional conformational changes in chromatin structure during epididymal transit. The objective of the present study was to compare the nuclear characteristics of immature and mature epididymal stallion spermatozoa, using a variety of experimental approaches. Resistance to in vitro decondensation of chromatin, following exposure to SDS-DTT and alkaline thioglycolate, increased significantly in mature spermatoz...
Zhang Z, Wang Y, Zhu M, Wang C.The present study was conducted to evaluate the in vitro dry matter disappearance (IVDMD) and fermentation characteristics of 6 fibrous feedstuffs incubated with donkey cecal or colic microorganisms. The fibrous feeds were corn straw, wheat straw, peanut vine, peanut shell, wheat shell and wheat bran (WB), which are commonly applied in large-scale donkey farms in China. After 48 hours fermentation, the highest IVDMD occurred in WB, and the lowest occurred in peanut shell (P < .05) regardless what inoculum applied. The IVDMD was positively correlated with OM (R = 0.42, P < .01), CP (R ...
Sabban S, Ye H, Helm B.The binding of immunoglobulin E (IgE) to its high-affinity receptor (FcɛRI) is the central protein interaction in IgE-mediated allergic reactions. The cross-linking of the IgE/FcɛRI complex, through cognate allergens, on the surface of mast cells and basophil cells results in mediator release, and thus leads to the symptoms of type I hypersensitivity responses in mammals. To develop a baseline value for subsequent equine anti-allergy drug and vaccine research, the interaction of equine IgE with its high-affinity FcɛRI receptor was investigated following the cloning and expression of equine ...
Wu Z, Zheng X, Luo Y, Huo F, Dong H, Zhang G, Yu W, Tian F, He L, Chen J.The present study investigates the effects of five cryoprotectants (CPAs) and cryoprotectant combinations on the post-thaw total motility, progressive motility, viability, mitochondrial membrane potential and acrosome integrity in stallion spermatozoa. In Experiment 1, the objective was to compare the impact of different concentrations (2.5%, 3.5% and 5%) of a single CPA, including glycerol (Gly), ethylene glycol (EG), dimethyl sulphoxide (DMSO), methyl formamide (MF), and dimethylformamide (DMF) for stallion spermatozoa cryopreservation. In Experiment 2, two or more CPAs were used to assess w...
Galera PD, Ribeiro CR, Sapp HL, Coleman J, Fontes W, Brooks DE.Human amniotic membrane (AM) has been used as a biomaterial for surgical wound skin and ocular surface reconstruction for several years. Currently, equine AM has been used for corneal reconstruction in several animal species, and appears to have the same properties as human AM. Despite the observed positive healing abilities of this tissue in horses with ulcerative keratitis the proteins of equine AM have not been described. Objective: To identify proteins known to be associated with corneal healing from frozen equine AM. Methods: Placentas were acquired from healthy live foal births from a lo...
Kämpf S, Seiler E, Bujok J, Hofmann-Lehmann R, Riond B, Makhro A, Bogdanova A.Detection of hematopoietic activity in horses is a challenge due to the lack of cells carrying reticulocyte markers such as RNA remnants or CD71 in the circulation. In this study, we fractionated equine red cells according to their density and analyzed the cells forming low (L), medium (M), and high (H) density fractions for markers of aging such as membrane loss, oxidation, and alterations in the intracellular free Ca levels. Cells forming L and M fraction were highly heterogeneous in projected areas and shapes, and had higher propensity to swell in response to hypo-osmotic challenge than the...
van de Lest CH, van den Hoogen BM, van Weeren PR.The object of this study was to determine whether changes in the synovial fluid (SF) induced by in vivo loading can alter the metabolic activity of chondrocytes in vitro, and, if so, whether insulin-like growth factor-I (IGF-I) is responsible for this effect. Therefore, SF was collected from ponies after a period of box rest and after they had been exercised for a week. Normal, unloaded articular cartilage explants were cultured in 20% solutions of these SFs for 4 days and chondrocyte bioactivity was determined by glycosaminoglycan (GAG) turnover (i.e., the incorporation of 35SO4 into GAG and ...
Fossum C, Hjertner B, Olofsson KM, Lindberg R, Ahooghalandari P, Camargo MM, Bröjer J, Edner A, Nostell K.The expression of tlr4, md2 and cd14 was studied in equine blood leukocytes and in intestinal samples using real time PCR. The stability of three commonly used reference genes, glyceraldehyde-3P-dehydrogenase (GAPDH), hypoxantine ribosyltransferase (HPRT) and succinate dehydrogenase complex subunit A (SDHA), was evaluated using qbase(PLUS). The equine peripheral blood mononuclear cells (eqPBMC) examined were either stimulated in vitro with Phorbol 12-myristate 13-acetate (PMA) and ionomycin or with the CpG oligodeoxynuclotide 2216 (CpG-ODN 2216) or obtained from horses before, during and after...
Hosaka YZ, Uratsuji T, Ueda H, Uehara M, Takehana K.This aim of this study was to determine the characteristic differences in tendinocytes derived from three sites of the equine superficial digital flexor tendon (SDFT)-proximally the myotendinous junction (MTJ), mid-metacarpal (mM) and osteotendinous junction (OTJ)-in morphology, proliferation, and ability for synthesis of collagen and matrix metalloproteinases (MMPs). Little difference was observed in cell proliferation. Addition of tumor necrosis factor (TNF) alpha to the culture medium resulted in increased collagen synthesis by tendinocytes from all three sites. The amount of collagen synth...
Yasunaga S, Maeda K, Matsumura T, Kondo T, Kai K.A type-specific enzyme-linked immunosorbent assay (ELISA) using equine herpesvirus types 1 (EHV-1) and 4 (EHV-4) glycoprotein G was applied for sero-epizootiology of EHV infections in Japan. Recently, an inactivated EHV-1 vaccine has been administered to racehorses for prevention of upper respiratory disease. To examine the effect of the vaccination on the result of the ELISA, 6 horses were experimentally inoculated three times intramuscularly or intranasally with inactivated EHV-1 vaccine. Sera collected from these horses were used to the type-specific ELISA and complement-fixation (CF) test....
Gandini M.To report a technique for semiclosed 1-layer side-to-side jejunocecal anastomosis in horses. Methods: Experimental study and clinical reports. Methods: Part 1--In vitro: Intestinal specimens from 24 horses collected immediately after death at an abattoir. Part 2--CLINICAL CASES: 10 horses with clinical signs of colic requiring jejunocecostomy. Methods: Mean time to perform twelve 2-layer handsewn (HS2L) and 12 semiclosed 1-layer modified handsewn (SC) jejunocecal anastomoses was compared. Mean bursting pressures of anastomoses measured with a modified tank inflation test were compared. The SC ...
Fleming JT, Joshi JG.We previously showed that human brain ferritin (HBF) binds aluminum (Al) in vivo and in vitro and HBF isolated from Alzheimer's brain had more Al bound compared to aged matched controls (7). To further understand the role ferritin may play in Al neurotoxicity, we have studied in vitro the effect of Al on the function of human ferritin isolated from Alzheimer's (AD) and normal brain tissue, and compared the results with other mammalian ferritins. Al causes a concentration-dependent decrease in the initial rate of iron loading into apo-horse spleen and human brain ferritin and the rates were sim...
Lawson AL, Pittaway CE, Sparrow RM, Balkwill EC, Coles GC, Tilley A, Wilson AD.Anoplocephala perfoliata is the commonest equine tapeworm, the adult parasites are attached in groups close to the ileocaecal valve causing marked inflammatory pathology. This work aimed to characterize the nature of the in vivo mucosal immune response to A perfoliata, and to investigate the role of A perfoliata excretory-secretory components in modulating in vitro immune responses. Real-time PCR detected elevation of IL13 and TGFβ transcription in early-stage A perfoliata infection. In late-stage infection, IL-13, IL4 and Ifn transcripts were reduced while the regulatory cytokines, TGFβ...
