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.
Moreno CR, Santschi EM, Younkin JT, Larson RL, Litsky AS.To determine the failure method of simulated equine medial femoral condyle (MFC) subchondral bone defects under compression and the influence of screw placement on failure resistance. Methods: In vitro study. Methods: Composite disks (CD) simulating the moduli of yearling bone in the MFC. Methods: Four CD conditions were tested, all with a 12.7 mm void (n = 6 per condition): intact (no void), void only, void with a 4.5 mm screw placed in neutral fashion, and void with a 4.5 mm screw placed in lag fashion. Composite disks of each condition were tested under monotonic compression to 6000...
Monteiro RA, Cunha RM, Guerra MMP, de Almeida VM, Peña-Alfaro CE, Silva SV.The aim of this study was to test equine semen cryopreservation techniques for the conservation of donkey germplasm. Ejaculates of three male donkeys were used (n = 18; six ejaculates per donkey; six repetitions), collected by the artificial vagina method. To remove the seminal plasma, the ejaculates were split and submitted to filtration or centrifugation methods. To assess the freezing method, each fraction was submitted to the automated system or the conventional system, and groups were formed: automated centrifuge, automated filtrate, conventional centrifuge and conventional filtrate. Af...
Lançoni R, Celeghini ECC, Gonella-Diaza AM, Júnior VG, de Carvalho CPT, Zoca GB, Garcia-Oliveros LN, Batissaco L, Oliveira LZ, de Arruda RP.This study aimed to assess the semen ubiquitin levels of stallions with good (GF) and poor semen freezability (PF) and to evaluate the relationship between sperm ubiquitination and sperm morphological defects. Five ejaculates from eight adult stallions (n = 40) were collected and cryopreserved. Then, the ubiquitin level in equine sperm cells was assessed by immunohistochemistry with epifluorescence microscopy, and sperm morphology was assessed by differential interference contrast microscopy. Sperm cells were classified according to the intensity (classification 1: from I to IV; I = very l...
Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Kusano K, Nagata SI.Processed pseudogenes, also known as retrocopy genes, are copies of messenger RNAs that have been reverse transcribed into DNA and inserted into the genome. In this study, we identified 62 processed pseudogene candidates as intron-less genes from whole-genome sequencing (WGS) data of Thoroughbred horses using delly structural variation software. The 62 processed pseudogene candidates were confirmed by PCR amplification of intron-less products. A total of 11 processed pseudogenes were confirmed in the genome of all 23 analysed horses, whereas three processed pseudogenes with structures of ATP11...
Fernández-Hernández P, García-Marín LJ, Bragado MJ, Domingo A, González-Fernández L, Macías-García B.In the horse, a repeatable protocol for in vitro fertilization has not been developed, possibly due to incomplete sperm capacitation. We have previously identified the metabolites present in equine oviductal fluid (OF). We aimed to test the effects of different metabolites found in equine oviductal fluid on quality parameters of frozen-thawed spermatozoa. Different concentrations of myoinositol (5-25 mM), lactate (6-60 mM), glycine (0.1-5 mM), β-alanine (1-6 mM), and histamine (0.05-0.4 mM) were added independently to modified Whitten's medium (pH = 7.25). Thawed equine spermatozoa (three s...
Briski O, Salamone DF.During the past 2 decades, intracytoplasmic sperm injection (ICSI) has become a routine technique for clinical applications in humans. The widespread use among domestic species, however, has been limited to horses. In horses, ICSI is used to reproduce elite individuals and, as well as in humans, to mitigate or even circumvent reproductive barriers. Failures in superovulation and conventional in vitro fertilization (IVF) have been the main reason for the use of this technology in horses. In pigs, ICSI has been successfully used to produce transgenic animals. A series of factors have resulted in...
Rizk MA, El-Sayed SAE, Eltaysh R, Igarashi I.Equine piroplasmosis is a tick-transmitted disease that is considered one of the most serious infectious diseases affecting equines. Searching for novel antipiroplasm drugs remains indispensable due to the emergence and spreading of resistant piroplasm parasites against the limited currently used drugs, diminazene aceturate and imidocarb dipropionate. Therefore, novel drugs with specified targets need to be identified and exploited. The inhibitory effects of Medicines for Malaria Venture (MMV) Malaria Box compounds with potent in vitro anti-equine piroplasmosis activity were evaluated against ...
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...
Hagen A, Holland H, Brandt VP, Doll CU, Häußler TC, Melzer M, Moellerberndt J, Lehmann H, Burk J.Platelet lysate (PL) is an attractive platelet-based therapeutic tool and has shown promise as xeno-free replacement for fetal bovine serum (FBS) in human and equine mesenchymal stromal cell (MSC) culture. Here, we established a scalable buffy-coat-based protocol for canine PL (cPL) production (n = 12). The cPL was tested in canine adipose MSC (n = 5) culture compared to FBS. For further comparison, equine adipose MSC (n = 5) were cultured with analogous equine PL (ePL) or FBS. During canine blood processing, platelet and transforming growth factor-β1 concentrations increased ( < 0.05 and < ...
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 ...
Lanier CJ, Taintor JS, Christopherson PW, Spangler EA.Acidemia in sick or injured horses is often due to lactic acid accumulation. Alterations in platelet function and hemostasis are among numerous deleterious effects caused by decreased physiologic pH. Objective: We aimed to evaluate the effect of hyperlactatemia and resultant acidemia on platelet aggregation in equine whole blood using impedance aggregometry. Methods: Platelet aggregation was measured using the Multiplate analyzer in whole blood from 34 healthy horses at baseline and after in vitro addition of lactic acid to adjust the pH. Platelet aggregation of each sample was quantified by t...
Avellar HK, Lutter JD, Ganta CK, Beard W, Smith JR, Jonnalagadda N, Peloquin S, Kang Q, Ayub K.Septic arthritis is considered a medical emergency. Disease following bacterial colonization can lead to significant morbidity and mortality and requires costly treatment. Antimicrobial properties of regenerative therapies, including mesenchymal stromal cells and platelet products, have been researched extensively in human medicine. Although fewer studies have been conducted in veterinary species, they have shown promising results. The purpose of this study was to evaluate bacterial suppression by equine platelet lysate (EPL) and adipose-derived mesenchymal stromal cells (ASCs) in vitro. We hy...
Leemans B, Bromfield EG, Stout TAE, Vos M, Van Der Ham H, Van Beek R, Van Soom A, Gadella BM, Henning H.We describe the development of two methods for obtaining confluent monolayers of polarized, differentiated equine oviduct epithelial cells (EOEC) in Transwell inserts and microfluidic chips. EOECs from the ampulla were isolated post-mortem and seeded either (1) directly onto a microporous membrane as differentiated EOECs (direct seeding protocol) or (2) first cultured to a confluent de-differentiated monolayer in conventional wells, then trypsinized and seeded onto a microporous membrane (re-differentiation protocol). Maintenance or induction of EOEC differentiation in these systems was achiev...
Johnson JP, Vinardell T, David F.Ultrasound-guided injections can be used for a wide variety of conditions in the horse, including both diagnostic and therapeutic applications. Benefits of ultrasound guidance include more accurate deposition of injectate compared with blind approaches. Improved identification of vital structures, including nerves and blood vessels, allows their avoidance and thus reduces procedure-associated complications. Validation of such ultrasound-guided techniques has shown that they can be easily learnt by inexperienced veterinarians, assuming a proper knowledge of the sonographic anatomy. In many case...
Schellenberg LM, Regenthal R, Abraham G.This research paper investigates the influence of the Rho kinase (ROCK) inhibitor Y-27632 on β2-adrenoceptor density and the generation of a cellular messenger called cAMP in horse bronchial cells. The […]
Timms M, Steel R.The proteotypic human EPO peptides YLLEAK (T4), SLTTLLR (T11), TITADTFR (T14), and VYSNFLR (T17) are often used to confirm the presence of recombinant human EPO (rhEPO) in equine samples. Each of these peptides contains one or more isomeric leucine or isoleucine amino acids, raising the possibility that a simple leucine/isoleucine substitution could lead to a false identification when compared with a rhEPO reference standard. To examine this possibility variants of these four peptides were analysed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). These studies indicate that confir...
Macêdo IN, Arruda LCP, de Santana BB, de Moura TCM, Guerra MMP, Bezerra DG, Carneiro GF, Silva SV.The objective of this study was to evaluate the effects of the addition of different concentrations of ozone to quarter horse semen submitted to cryopreservation. Six ejaculates from four stallions were collected and were divided in four experimental groups: a control group (BotuCRIO® extender) and three other groups with BotuCRIO® ozonized at concentrations of 6, 8 and 12 μg of O/mL. The semen samples were diluted (200 x 10 spermatozoa/mL), filled in straws and frozen. After thawing (37 ºC, 30s), the samples were evaluated at 0, 30 and 60 minutes of incubation regarding sperm kinetics by ...
Lisboa FP, Silvestre WP, Castro JO, Martins GV, Segabinazzi LGTM, Pauletti GF, Dell'Aqua JA.This study aimed to evaluate the in vitro antimicrobial activity of essential oils (EO) from Ocimum basilicum (basil), Rosmarinus officinalis (rosemary), and Cymbopogon citratus (lemongrass) on endometritis-causing microorganisms in mares. Serial concentrations of the EO from 30.00 mg/mL to 0.47 mg/mL were tested. The major compounds of O. basilicum EO were linalyl acetate (33.32 wt.%) and citronellal (25.06 wt.%); of R. officinalis EO were borneol (26.48 wt.%), trans-β-ocimene (16.76 wt.%), camphene (12.45 wt.%), and α-phellandrene (11.08 wt.%); and of C. citratus EO were geranial (45.96 ...
Cutler C, Viljanto M, Taylor P, Hincks P, Biddle S, Van Eenoo P.Selective androgen receptor modulators, SARMs, are a large class of compounds developed to provide therapeutic anabolic effects with minimal androgenic side effects. A wide range of these compounds are available to purchase online and thus provide the potential for abuse in sports. Knowledge of the metabolism of these compounds is essential to aid their detection in doping control samples. In vitro models allow a quick, cost-effective response where administration studies are yet to be carried out. In this study, the equine phase I metabolism of the non-steroidal SARMs GSK2881078, LGD-2226, LG...
Lee OJ, Koch TG.Inflammation-associated disorders are significant causes of morbidity in horses. Equine single-donor mesenchymal stromal cells (sdMSCs) hold promise as cell-therapy candidates due to their secretory nonprogenitor functions. This has been demonstrated by mononuclear cell suppression assays (MSAs) showing that sdMSCs are blood mononuclear cell (BMC) suppressive in vitro. sdMSCs derived from umbilical cord blood are of clinical interest due to their ease of procurement, multipotency, and immunomodulatory ability. Due to the inherent donor-to-donor heterogeneity of MSCs, the development of robust ...
Spinaci M, Nerozzi C, Mislei B, Blanco-Prieto O, Mari G, Galeati G, Bucci D.The growing and widespread use of glyphosate-based herbicides (GBHs) has raised an intense public debate about the impact of environmental contamination on animal and human health, including male fertility. The aim of this study was to deepen the impact of glyphosate (Gly) and GBHs on mammalian sperm investigating the effect of in vitro exposure of stallion spermatozoa to Gly and to its commercial formulation Roundup® (R). Spermatozoa were incubated at 37 °C with different Gly or R concentrations (from 0.5 to 720 μg/mL Gly or R at the same Gly-equivalent concentrations). After 1 h of in...
Yánez-Ortiz I, Catalán J, Delgado-Bermúdez A, Carluccio A, Miró J, Yeste M.In donkeys, the use of frozen-thawed sperm for artificial insemination (AI) leads to low fertility rates. Furthermore, donkey sperm produce a large amount of reactive oxygen species (ROS), and post-AI inflammation induces the formation of neutrophil extracellular traps (NETosis), which further generates many more ROS. These high ROS levels may induce lipid peroxidation in the sperm plasma membrane, thus affecting its integrity. Enzymatic and non-enzymatic antioxidants, mainly found in the seminal plasma (SP), are responsible for maintaining the redox balance. However, this fluid is removed pri...
Fojtík L, Fiala J, Pompach P, Chmelík J, Matoušek V, Beier P, Kukačka Z, Novák P.Covalent labeling of proteins in combination with mass spectrometry has been established as a complementary technique to classical structural methods, such as X-ray, NMR, or cryogenic electron microscopy (Cryo-EM), used for protein structure determination. Although the current covalent labeling techniques enable the protein solvent accessible areas with sufficient spatial resolution to be monitored, there is still high demand for alternative, less complicated, and inexpensive approaches. Here, we introduce a new covalent labeling method based on fast fluoroalkylation of proteins (FFAP). FFAP u...
Journal of biomechanicsNovember 27, 2021
Volume 130 110888 doi: 10.1016/j.jbiomech.2021.110888
Shaffer SK, Shelly K, Garcia TC, Samol MA, Hill AE, Fyhrie DP, Stover SM.Proximal sesamoid bone (PSB) fractures in racehorses are likely fatigue fractures that occur due to repetitive loads and stress remodeling. The loading circumstances that may induce damage in the PSBs are not well understood. The goal of this study was to determine in three-dimensions, PSB motions relative to the opposing metacarpal condyle during simulated mid-stance loads. Seven equine cadaveric forelimbs were axially loaded in a material testing system to simulate standing and mid-stance walk, trot, and gallop load conditions (1.8-10.5 kN). Joint angles were determined by tracking the posit...
Moura TCM, Arruda LCP, Araújo Silva RAJ, Silva RPF, Oliveira AS, Tobal LFM, Batista AM, Carneiro GF, Guerra MMP.The aim of this study was to investigate the effects of sucrose on post-thawed equine semen quality. Semen samples (n = 24) were collected from six stallions. They were diluted (200 × 10 sperm/mL) in a freezing medium based on skimmed milk, egg yolk, dimethylformamide, and supplemented with sucrose at concentrations of 0 (Control), 25, 50, and 100 mM and in a commercial extender (BotuCrio). Subsequently, they were filled in straws (0.5 mL) and subjected to freezing and storage (-196°C). Immediately after thawing (37°C, 30 seconds), semen samples were evaluated for kinetics (CASA), plasm...
Doll CU, Niebert S, Burk J.Tendon lesions are common sporting injuries in humans and horses alike. The healing process of acute tendon lesions frequently results in fibrosis and chronic disease. In horses, local mesenchymal stromal cell (MSC) injection is an accepted therapeutic strategy with positive influence on acute lesions. Concerning the use of MSCs in chronic tendon disease, data are scarce but suggest less therapeutic benefit. However, it has been shown that MSCs can have a positive effect on fibrotic tissue. Therefore, we aimed to elucidate the interplay of MSCs and healthy or chronically diseased tendon matrix...
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 ...
Philip M, Karakka Kal AK, Subhahar MB, Karatt TK, Mathew B, Perwad Z.Performance-enhancing substances and methods have become a serious problem in competitive sports. The hypoxia-inducible factor (HIF) stabilizers can enhance the organism's capacity for molecular oxygen transport and are likely to be abused as performance-enhancing agents in sports. This paper describes the metabolic conversion of the popular hypoxia inducible factor-prolyl hydroxylase (HIF-PH) inhibitors, namely, daprodustat, desidustat, and vadadustat using equine liver microsomes, determined on a QExactive high-resolution mass spectrometer. During this study, a total of 10 metabolites for da...
Bork S, Yokoyama N, Matsuo T, Claveria FG, Fujisaki K, Igarashi I.We evaluated the growth inhibitory effect of triclosan, which has recently been reported to inhibit the growth of Plasmodium species and Toxoplasma gondii, on bovine and equine Babesia parasites in in vitro cultures The growth of Babesia bovis and B. bigemina was significantly inhibited in the presence of 100 microg/ml of triclosan, while B. caballi and B. equi were susceptible to as low as 50 microg/ml. Babesia bigemina and B. caballi were completely cleared as early as on the first and second day of the treatment, respectively. These parasites did not exhibit any growth in the subsequent fiv...
Milner PI, Wilkins RJ, Gibson JS.To examine the effect of O(2) and the role, and source, of reactive oxygen species (ROS) on pH regulation in articular chondrocytes. Methods: Cartilage from equine metacarpo/tarsophalangeal joints was digested (collagenase) to isolate chondrocytes and loaded with 2',7'-bis-2-(carboxyethyl)-5(6)-carboxylfluorescein, a pH-sensitive fluorophore. O(2) tension was maintained using Eschweiler tonometers and a Wosthoff gas mixer. Cells were exposed to agents which alter ROS levels, mitochondrial inhibitors and/or inhibitors of protein phosphorylation. ROS levels were determined by dichlorofluorescein...
Morris GJ, Faszer K, Green JE, Draper D, Grout BW, Fonseca F.The cellular damage that spermatozoa encounter at rapid rates of cooling has often been attributed to the formation of intracellular ice. However, no direct evidence of intracellular ice has been presented. An alternative mechanism has been proposed by Morris (2006) that cell damage is a result of an osmotic imbalance encountered during thawing. This paper examines whether intracellular ice forms during rapid cooling or if an alternative mechanism is present. Horse spermatozoa were cooled at a range of cooling rates from 0.3 to 3,000 degrees C/min in the presence of a cryoprotectant. The ultra...
Miyabara S, Yuda Y, Kasashima Y, Kuwano A, Arai K.Tenomodulin has been recognized as a biomarker for tendon differentiation, and its gene expression is regulated by several transcription factors including Scleraxis and Mohawk. In this study, we found a novel regulatory mechanism of tenomodulin expression. Equine bone marrow-derived mesenchymal stem cells (BMSCs) in monolayer culture showed a low mRNA level of tenomodulin in comparison with the level in the tendon. When cultured in collagen gel containing a glycogen synthase kinase-3 (GSK-3) inhibitor (BIO), expression of tenomodulin in BMSCs increased up to the level in the tendon. Participat...
O'Rourke JP, Newbound GC, Kohn DB, Olsen JC, Bunnell BA.This report compares gene transfer efficiencies as well as durations and levels of gene expression for human immunodeficiency virus (HIV) and equine infectious anemia virus (EIAV) lentiviral vectors in a variety of human cell types in vitro. EIAV and HIV vectors transduced equivalent numbers of proliferating and G1/S- and G2/M-arrested cells, and both had very low efficiencies of transduction into G0-arrested cells. Analysis of the levels of both the enhanced green fluorescent protein (EGFP) and mRNA demonstrated that the HIV-transduced cells expressed greater levels of EGFP protein and RNA th...
Gao W, Chan Y, You M, Lacap-Bugler DC, Leung WK, Watt RM.This study explored the range of bacterial taxa present within healthy subgingival (below the gum-line) niches in the horse oral cavity using 16S rRNA gene amplicon pyrosequencing. Pooled subgingival plaque samples were collected from approximately 200 sulcus sites from two horses (EQ1, EQ2) for analysis. A total of 14,260 quality-filtered pyrosequencing reads were obtained, which were assigned to 3875 operational taxonomic units (OTUs; 99% identity cut-off); 1907 OTUs for EQ1 and 2156 OTUs for EQ2. Diverse taxa from 12 phyla were identified, including Actinobacteria (3.17%), Bacteroidetes (25...
Bressan FF, Bassanezze V, de Figueiredo Pessôa LV, Sacramento CB, Malta TM, Kashima S, Fantinato Neto P, Strefezzi RF, Pieri NCG, Krieger JE....Induced pluripotent stem cells (iPSCs) have enormous potential in developmental biology studies and in cellular therapies. Although extensively studied and characterized in human and murine models, iPSCs from animals other than mice lack reproducible results. Herein, we describe the generation of robust iPSCs from equine and bovine cells through lentiviral transduction of murine or human transcription factors Oct4, Sox2, Klf4, and c-Myc and from human and murine cells using similar protocols, even when different supplementations were used. The iPSCs were analyzed regarding morphology, gene and...
LaPolt PS, Tilly JL, Aihara T, Nishimori K, Hsueh AJ.The actions of gonadotropins on ovarian differentiation are associated with dynamic changes in gonadotropin receptor content, presumably due to modulation of receptor gene expression. The present studies used a reverse transcription-polymerase chain reaction to obtain a rat FSH receptor cDNA fragment, followed by synthesis of a labeled cRNA probe to examine the regulation of FSH receptor mRNA levels during follicular maturation, ovulation, and luteinization. Northern blot analysis of ovarian RNA with the FSH receptor probe revealed two predominant hybridization signals of 7.0 and 2.5 kilobases...
Borjesson DL, Peroni JF.This article focuses on the emerging field of equine regenerative medicine with an emphasis on the use of mesenchymal stem cells (MSCs) for orthopedic diseases. We detail laboratory procedures and protocols for tissue handling and MSC isolation, characterization, expansion, and cryopreservation from bone marrow, fat, and placental tissues. We provide an overview of current clinical uses for equine MSCs and how MSCs function to heal tissues. Current laboratory practices in equine regenerative medicine mirror those in the human field. However, the translational use of autologous and allogeneic M...
Gilger BC, Salmon JH, Wilkie DA, Cruysberg LP, Kim J, Hayat M, Kim H, Kim S, Yuan P, Lee SS, Harrington SM, Murray PR, Edelhauser HF, Csaky KG....To determine the feasibility, safety, and effectiveness of an episcleral or deep scleral lamellar sustained release cyclosporine (CsA) device in a naturally occurring animal model of uveitis. Methods: A two-compartment perfusion chamber was used to assess in vitro human and equine scleral permeability of fluorescein, dexamethasone-fluorescein, or CsA. A biodegradable, matrix-reservoir CsA implant was designed, and release rates of CsA were determined in vitro. Tissue CsA levels were measured in eyes with the implant. Horses with equine recurrent uveitis (ERU) received episcleral or deep sclera...
Clérico G, Taminelli G, Veronesi JC, Polola J, Pagura N, Pinto C, Sansinena M.Oocyte vitrification is considered experimental in the horse with only three live foals reported. The oxidative conditions induced by vitrification could in part explain the poor results and melatonin, a powerful antioxidant, could stimulate ROS metabolization and restore mitochondrial function in these oocytes. Our objective was to determine the oxidative status of vitrified equine oocytes and to analyze the effect of melatonin on mitochondrial-specific ROS (mROS), oocyte maturation, ICSI embryo development and viability. Immature, abattoir-derived oocytes were held for 15 h and vitrified in...
Blanchard-Channell M, Moore PF, Stott JL.Two monoclonal antibodies (mAb), UC F6G-3 and UC F13C-5, were characterized as being specific for the apparent equine homologues of CD3 and CD5, respectively. Both antibodies exhibited characteristics of pan-T-lymphocyte markers based upon immunohistology and two-colour flow cytometry. UC F6G-3 precipitated a complex of proteins (up to seven) with molecular weights ranging from 18,000 to 42,000, similar to the human and murine CD3 complex. Upon further dissociation of the precipitated complex, two proteins were identified with molecular weights of 22,000 and 27,000. Immobilized UC F6G-3 was ef...
Riesenberg A, Feßler AT, Erol E, Prenger-Berninghoff E, Stamm I, Böse R, Heusinger A, Klarmann D, Werckenthin C, Schwarz S.The aim of this study was to determine the MICs of 32 antimicrobial agents for 200 isolates of Rhodococcus equi of animal origin by applying a recently described broth microdilution protocol, and to investigate isolates with distinctly elevated rifampicin MICs for the genetic basis of rifampicin resistance. Methods: The study included 200 R. equi isolates, including 160 isolates from horses and 40 isolates from other animal sources, from the USA and Europe. MIC testing of 32 antimicrobial agents or combinations thereof followed a recently published protocol. A novel PCR protocol for the joint ...
Kishi M, Arimura Y, Ikuta K, Shoya Y, Lai PK, Kakinuma M.A cDNA fragment of the Borna disease virus (BDV) open reading frame II (ORF-II), which encodes a 24-kDa phosphoprotein (p24 [P protein]), was amplified from total RNA of peripheral blood mononuclear cells (PBMC) from three psychiatric inpatients. The amplified cDNA fragments were cloned, sequenced, and analyzed. A total of 15 clones, 5 from each patient, were studied. Intrapatient divergencies of the BDV ORF-II nucleotide sequence were 4.2 to 7.3%, 4.8 to 7.3%, and 2.8 to 7.1% for the three patients, leading to differences of 7.7 to 14.5%, 10.3 to 17.1%, and 6.0 to 16.2%, respectively, in the ...
Riemersa DJ, Schamhardt HC.A clamp designed for holding tendons in force/elongation studies is described. No slippage occurred when tensile forces up to 13,800 N were applied to horses digital flexor tendons fixed in this clamp.
Smits K, De Coninck DI, Van Nieuwerburgh F, Govaere J, Van Poucke M, Peelman L, Deforce D, Van Soom A.Although the equine oviduct clearly affects early embryo development and the selective transport of equine embryos through the oviduct indicates a reciprocal interaction, the influence of the embryo on gene expression in the oviduct remains to be determined in the horse. The aim of this study was to examine this by means of RNA sequencing. Four days after ovulation, epithelial cells ipsilateral and contralateral to the ovulation side from five cyclic and five pregnant mares were collected from the oviduct. RNA was extracted, samples were sequenced, and data analysis was performed to determine ...
González-Fernández ML, Pérez-Castrillo S, Sánchez-Lázaro JA, Prieto-Fernández JG, López-González ME, Lobato-Pérez S, Colaço BJ, Olivera ER....OBJECTIVE To assess the ability to regenerate an equine meniscus by use of a collagen repair patch (scaffold) seeded with mesenchymal stem cells (MSCs) derived from bone marrow (BM) or adipose tissue (AT). SAMPLE 6 female Hispano-Breton horses between 4 and 7 years of age; MSCs from BM and AT were obtained for the in vitro experiment, and the horses were subsequently used for the in vivo experiment. PROCEDURES Similarities and differences between MSCs derived from BM or AT were investigated in vitro by use of cell culture. In vivo assessment involved use of a meniscus defect and implantation o...
Camargo Garbin L, Lopez C, Carmona JU.In the 1990s, the role of platelets in inflammation and tissue healing was finally recognized. Since then, the clinical use of platelet-derived products (hemocomponents), such as, platelet-rich plasma (PRP), markedly increased. The promise of a more economical option of a disease-modifying treatment led to the intensive and continuous research of PRP products and to its widespread clinical use. A number of protocols and commercial kits have been developed with the intention of creating a more practical and reliable option for clinical use in equine patients. Still, the direct comparison betwee...
Jörg A, Pasquier JM, Klebanoff SJ.Eosinophil peroxidase (donor: hydrogen-peroxidase oxidoreductase, EC 1.11.1.7) was isolated in a highly purified form (415/280 nm ratio, 1.05) from horse peripheral blood eosinophil. Eosinophil peroxidase was extracted from intact eosinophils (98-100% purity) or isolated eosinophil granules with 0.05 M acetate buffer (pH 4.7)/0.18 M NaCl and purified by chromatography on Sephadex G-200 and carboxymethylcellulose. Final elution was with 0.05 M acetate buffer (pH 4.7)/ 1 M NaCl. Horse eosinophil peroxidase is a strongly basic protein with bacterial properties when combined with H2O2 and iodide, ...
David F, Farley J, Huang H, Lavoie JP, Laverty S.To evaluate mRNA expression of several proinflammatory and anti-inflammatory cytokines and chemokines in equine unstimulated and interleukin-1beta (IL-1beta)-stimulated chondrocytes. Methods: In vitro experiment using equine chondrocyte cultures. Methods: Whole articular cartilage from metacarpophalangeal joints (n=5 horses; 10 fetlocks). Methods: Chondrocyte monolayer cultures were established from digested adult equine articular cartilage and stimulated with 5 ng/mL of recombinant human IL-1beta. RNA was extracted from the cells 24 hours after stimulation. IL-1beta, IL-4, IL-6, IL-8, tumor n...
Edmunds DH, Whittaker DK, Green RM.The progress of artificial caries-like lesions created in human, bovine, equine, and ovine enamel has been studied. Lesions were produced by exposure to an acid gel system or by 5-day exposure to a sequential batch culture technique using Streptococcus mutans NCTC 10832. Longitudinal ground sections were prepared. The lesions were of similar appearance in all species when examined in polarized light. The depth in human enamel was approximately half that in the animal species. Microradiography confirmed subsurface demineralization in all four species. Similar depth ratios were seen in the scann...
Renshaw RW, Glaser AL, Van Campen H, Weiland F, Dubovi EJ.A virus that could not be identified as a previously known equine virus was isolated from the mononuclear cells of a horse. Electron microscopy revealed enveloped virions with nucleocapsid structures characteristic of viruses in the Paramyxoviridae family. The virus failed to hemabsorb chicken or guinea pig red blood cells and lacked neuraminidase activity. Two viral genes were isolated from a cDNA expression library. Multiple sequence alignments of one gene indicated an average identity of 45% as compared to Morbillivirus N protein sequences. A weaker relationship was found with Tupaia paramy...
Tecles F, Cerón JJ.Whole blood cholinesterase was measured using acetyl-, butyryl- and propionylthiocholine as substrates in 10 healthy adult dogs, cats, horses, pigs, goats, sheep and cows, in order to determine and characterise the cholinesterase activity in whole blood of the main domestic animals. An in vitro exposure test with two anticholinesterase compounds, the organophosphate insecticide coumaphos and the carbamate insecticide imidocarb, was also performed. In whole blood of ruminants and pigs, acetylthiocholine yielded the highest cholinesterase activity and other substrates were poorly hydrolysed; in ...
Dobrinski I, Smith TT, Suarez SS, Ball BA.Interaction of equine spermatozoa with oviductal epithelial cells (OEC) prolongs sperm viability and maintains low intracellular calcium concentration ([Ca2+]i) in spermatozoa. Experiments were designed to investigate 1) whether release of spermatozoa from OEC in vitro is associated with elevated [Ca2+]i and 2) whether soluble products from OEC or direct membrane contact between spermatozoa and OEC mediates the effects of OEC on sperm [Ca2+]i. In the first experiment, changes in [Ca2+]i in spermatozoa loaded with indo-1 acetoxymethylester were determined in motile spermatozoa released from OEC...
Nixon AJ, Lust G, Vernier-Singer M.Equine articular chondrocytes were isolated from explant cartilage cultures by digestion in a 0.075% collagenase solution for 15 to 19 hours. Cartilage from late-term fetal and neonatal foals resulted in mean chondrocyte yield of 51.99 x 10(6) cells/g of cartilage (wet weight), compared with a yield of 17.83 x 10(6) cells/g for foals 3 to 12 months old. Propagation of chondrocytes in monolayer and 3-dimensional culture was accomplished, using Ham's F-12 as the basal medium, with supplements of fetal bovine serum (10%), ascorbic acid, alpha-ketoglutarate, and L-glutamine. The medium was buffere...
Glynn ER, Londono AS, Zinn SA, Hoagland TA, Govoni KE.The use of equine bone marrow mesenchymal stem cells (BMSC) is a novel method to improve fracture healing in horses. However, additional research is needed to identify optimal culture conditions and to determine the mechanisms involved in regulating BMSC differentiation into osteoblasts. The objectives of the experiments were to determine: 1) if autologous or commercial serum is better for proliferation and differentiation of equine BMSC into osteoblasts, and 2) the expression of key transcription factors during the differentiation of equine BMSC into osteoblasts. Equine BMSC were isolated fro...
Liebscher G, Vanchangiri K, Mueller T, Feige K, Cavalleri JM, Paschke R.Betulinic acid, a pentacyclic triterpene, and its derivatives are promising compounds for cancer treatment in humans. Melanoma is not only a problem for humans but also for grey horses as they have a high potential of developing melanoma lesions coupled to the mutation causing their phenotype. Current chemotherapeutic treatment carries the risk of adverse health effects for the horse owner or the treating veterinarian by exposure to antineoplastic compounds. Most treatments have low prospects for systemic tumor regression. Thus, a new therapy is needed. In this in vitro study, Betulinic acid a...
Gonzales DS, Jones JM, Pinyopummintr T, Carnevale EM, Ginther OJ, Shapiro SS, Bavister BD.The behaviour of bovine, equine and human blastocysts was studied in vitro by time-lapse videomicrography and computer imaging. This study revealed that cytoplasmic extensions of the trophectoderm ['trophectoderm projections' (TEP)] were expressed by embryos of all three species, prior to or during zona escape. Bovine and human blastocysts escaped their zonae with a combination of blastocoele expansion, collapse and re-expansion coupled with the penetration of the zona pellucida by TEP. In equine embryos, after several cycles of blastocoele expansion and collapse, trophectoderm ruptured the zo...
Raabe MR, Issel CJ, Montelaro RC.Equine infectious anemia virus (EIAV) has been shown to infect cells of monocyte/macrophage lineage. These primary cells are intrinsically difficult to obtain, to purify and to culture in vitro for extended periods of time. As a result, most in vitro studies concerning this lentivirus make use of primary equine fibroblasts or transformed canine or feline cell lines. We describe methods that yield reproducibly pure cultures of equine blood monocytes from peripheral blood mononuclear cells. The in vitro differentiation of these cells into mature equine macrophage was verified using various cytoc...
van de Water SG, van Gennip RG, Potgieter CA, Wright IM, van Rijn PA.African horse sickness virus (AHSV) is a virus species in the genus Orbivirus of the family Reoviridae. There are nine serotypes of AHSV showing different levels of cross neutralization. AHSV is transmitted by species of Culicoides biting midges and causes African horse sickness (AHS) in equids, with a mortality rate of up to 95% in naive horses. AHS has become a serious threat for countries outside Africa, since endemic Culicoides species in moderate climates appear to be competent vectors for the related bluetongue virus (BTV). To control AHS, live-attenuated vaccines (LAVs) are used in Afri...
