Laboratory methods in equine research encompass a variety of techniques and procedures used to analyze biological samples from horses to study health, disease, genetics, and physiology. These methods include hematological analyses, biochemical assays, molecular biology techniques, and microbiological cultures. Commonly utilized laboratory techniques involve blood tests for complete blood count (CBC) and serum chemistry, polymerase chain reaction (PCR) for genetic and infectious disease studies, and enzyme-linked immunosorbent assays (ELISA) for detecting specific proteins or antibodies. These methods provide valuable data that contribute to understanding equine health and disease mechanisms. This page compiles peer-reviewed research studies and scholarly articles that explore the application, development, and outcomes of laboratory methods in the context of equine research.
Montes Cortés MG, Fernández-García JL, Habela Martínez-Estéllez MÁ.Equine piroplasmoses are enzootic parasitic diseases distributed worldwide with high incidence in tropical and subtropical regions. In Spain, there is insufficient epidemiological data about equine piroplasmoses. The main aim of the present study was therefore to estimate the prevalence of Theileria equi and Babesia caballi in five regions and obtain information about the risk factors. This study was conducted in the central and south-western regions of Spain, using indirect fluorescence antibody testing (IFAT) in 3,100 sera samples from apparently healthy horses of different ages, breeds, coa...
Warnken T, Brehm R, Feige K, Huber K.The aim of the study was to analyze key proteins of the equine insulin signaling cascade and their extent of phosphorylation in biopsies from muscle tissue (MT), liver tissue (LT), and nuchal AT, subcutaneous AT, and retroperitoneal adipose tissues. This was investigated under unstimulated (B1) and intravenously insulin stimulated (B2) conditions, which were achieved by injection of insulin (0.1 IU/kg bodyweight) and glucose (150 mg/kg bodyweight). Twelve warmblood horses aged 15 ± 6.8 yr (yr), weighing 559 ± 79 kg, and with a mean body condition score of 4.7 ± 1.5 were included in the s...
Mirajkar NS, Kelley MR, Gebhart CJ.Reported herein is the draft genome sequence of equine-origin Lawsonia intracellularis strain E40504, an obligate intracellular bacterium and the etiological agent of equine proliferative enteropathy. The 1.69-Mb draft genome sequence includes 1,380 protein-coding genes and 49 RNA genes, and it lacks a genomic island reported in swine-origin L. intracellularis strain PHE/MN1-00.
Mani RJ, Thachil AJ, Ramachandran A.Accurate and timely identification of infectious etiologies is of great significance in veterinary microbiology, especially for critical diseases such as strangles, a highly contagious disease of horses caused by Streptococcus equi subsp. equi. We evaluated a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platform for use in species- and subspecies-level identification of S. equi isolates from horses and compared it with an automated biochemical system. We used 25 clinical isolates each of S. equi subsp. equi and S. equi subsp. zooepidemicus. Using ...
Aguiar FLN, Gastal GDA, Ishak GM, Gastal MO, Teixeira DIA, Feugang JM, Figueiredo JR, Gastal EL.The effect of FSH supplementation on an enriched cultured medium containing insulin (10 ng/mL) and EGF (50 ng/mL) was investigated on in vitro culture of equine ovarian biopsy tissue. Ovarian tissue fragments were collected from mares (n = 10) and distributed in the following treatments: noncultured control, cultured control, and cultured + FSH. Both treated groups were cultured for 7 or 15 days. The end points evaluated were: follicular morphology, estradiol levels in the culture medium, fluorescence intensity for TUNEL, EGFR and Ki-67 detection, and gene expression of GDF-9, BMP-15, a...
Zarski LM, High EA, Nelli RK, Bolin SR, Williams KJ, Hussey G.Equine herpesvirus 5 (EHV-5) infection is associated with pulmonary fibrosis in horses, but further studies on EHV-5 persistence in equine cells are needed to fully understand viral and host contributions to disease pathogenesis. Our aim was to develop a quantitative PCR (qPCR) assay to measure EHV-5 viral copy number in equine cell cultures, blood lymphocytes, and nasal swabs of horses. Furthermore, we used a recently developed equine primary respiratory cell culture system to study EHV-5 pathogenesis at the respiratory tract. PCR primers and a probe were designed to target gene E11 of the EH...
Sumner SM, Naskou MC, Thoresen M, Copland I, Peroni JF.Platelet preparations containing growth factors, attachment factors, and enzymes are appealing to enhance healing of injured tissues and as an alternative to xenogenic serum in cell culture media. Plateletpheresis is commonly used to collect platelets in human medicine but has not been validated in horses. Plateletpheresis to collect platelet concentrate was performed on six female, mixed breed, chemically restrained horses using commercially available apheresis equipment. Before and immediately after plateletpheresis, we performed physical examinations and collected blood for chemistry and co...
Liu S, Lian S, Yang Y, Fu C, Ma H, Xiong Z, Ling Y, Zhao C.An experiment was conducted to study the association between the single nucleotide polymorphisms (SNPs) in 5'-untranslated regions (5'-UTR) of equine chorionic gonadotropin (eCG) genes and the serum eCG levels. Methods: SNPs in 5'-UTR of eCG genes were screened across 10 horse breeds, including 7 Chinese indigenous breeds and 3 imported breeds using iPLEX chemistry, and the association between the serum eCG levels of 174 pregnant Da'an mares and their serum eCG levels (determined with ELISA) was analyzed. Results: Four SNPs were identified in the 5'-UTR of the eCGα gene, and one of them was u...
Drábková Z, Schramel JP, Kabeš R.To compare two methods of Bohr-Enghoff physiological dead space to tidal volume ratio (Vd/Vt) determination using a mixing chamber and an E-CAiOVX metabolic monitor. Methods: Prospective, clinical, method-comparison study. Methods: Twenty horses anaesthetized for elective orthopaedic procedures. Methods: Horses were anaesthetized with isoflurane in oxygen and the lungs were mechanically ventilated (Vt 15±2 mL kg). Arterial blood was sampled to provide arterial partial pressure of carbon dioxide (PaCO) for dead space calculation using a metabolic monitor. Mixed expired partial pressure of ca...
Schaafstra FJWC, van Doorn DA, Schonewille JT, van Riet MMJ, Visser P, Blok MC, Hendriks WH.Methodological aspects of digestibility measurements were studied in four Welsh pony geldings consuming haylage-based diets with increasing proportions of a pelleted concentrate according to a 4×4 Latin square design experiment. Ponies were fed four experimental, iso-energetic (net energy (NE) basis) diets (i.e. 22 MJ NE/day) with increasing proportions of a pelleted concentrate (C) in relation to haylage (H). The absolute amounts of diet dry matter fed per day were 4.48 kg of H (100H), 3.36 and 0.73 kg of H and C (75H25C), 2.24 and 1.45 kg of H and C (50H50C) and 1.12 and 2.17 kg of H and C ...
Bozorgmanesh R, Sutton-Burges JW, Tablin F.Equine whole blood collection and storage methods have been evaluated to assess red blood cell viability; however, platelet (PLT) viability has not been comprehensively assessed. Objective: The purpose of the study was to compare viability of PLTs collected in whole blood into 2 different anticoagulants. Methods: Whole blood from 6 healthy adult Thoroughbred horses was collected into citrate-phosphate-dextrose-adenine (CPDA) or acid-citrate-dextrose (ACD). Platelet count, pH, and concentrations of glucose, lactate, carbon dioxide, oxygen, bicarbonate, sodium, potassium, and chloride were measu...
ELISAs are known to have a higher diagnostic sensitivity than the agar gel immunodiffusion (AGID) when employed for serological diagnosis of equine infectious anaemia (EIA). For this purpose, an "in-house" and five commercial ELISAs available in Italy were assessed by the National Reference Centre for EIA for their analytic specificity (Sp); precocity, defined as capability of detecting first antibodies produced during a new infection; precision based on repeatability and reproducibility, estimated from the coefficient of variation (CV); accuracy, estimated from multiple K and relative Sp and ...
Suganuma K, Yamasaki S, Molefe NI, Musinguzi PS, Davaasuren B, Mossaad E, Narantsatsral S, Battur B, Battsetseg B, Inoue N.Dourine is caused by Trypanosoma equiperdum via coitus with an infected horse. Although dourine is distributed in Equidae worldwide and is listed as an internationally important animal disease by the World Organization for Animal Health (OIE), no effective treatment strategies have been established. In addition, there are no reports on drug discovery, because no drug screening system exists for this parasite. A new T. equiperdum strain was recently isolated from the genital organ of a stallion that showed typical symptoms of dourine. In the present study, we adapted T. equiperdum IVM-t1 from...
Cymerys J, Słońska A, Tucholska A, Golke A, Chmielewska A, Bańbura MW.Equine herpesvirus 1 (EHV-1), like other members of the Alphaherpesvirinae subfamily, is a neurotropic virus causing latent infections in the nervous system of the natural host. In the present study, we have investigated EHV-1 replication (wild-type Jan-E strain and Rac-H laboratory strain) during long-term infection and during the passages of the virus in cultured neurons. The studies were performed on primary murine neurons, which are an excellent in vitro model for studying neurotropism and neurovirulence of EHV-1. Using real-time cell growth analysis, we have demonstrated for the first tim...
Schnabel CL, Babasyan S, Freer H, Wagner B.Only few quantitative reports exist about the concentrations and induction of immunoglobulin A (IgA) in mucosal secretions of horses. Despite this, it is widely assumed that IgA is the predominant immunoglobulin on mucosal surfaces in the horse. Here, two new monoclonal antibodies (mAbs) against equine IgA, clones 84-1 and 161-1, were developed and characterized in detail. Both IgA mAbs specifically bound monomeric and dimeric equine IgA in different applications, such as Western blots and fluorescent bead-based assays. Cross-reactivity with other equine immunoglobulin isotypes was not observe...
Liu Y, Uboh CE, Li X, Guan F, You Y, Maylin GA, Zhu F, Soma LR.A rapid and sensitive method for simultaneous screening, quantification and confirmation of 17 barbiturates in horse plasma using liquid chromatography-tandem mass spectrometry is described. Analytes were recovered from plasma by liquid-liquid extraction using methyl tert-butyl ether, separated on a C18 column, and analyzed in negative electrospray ionization mode. Multiple-reaction monitoring was employed for screening and quantification. Confirmation for the presence of the analytes was achieved by comparing ion intensity ratio. The ranges for limits of detection, quantification and confirma...
Loux SC, Scoggin KE, Bruemmer JE, Canisso IF, Troedsson MH, Squires EL, Ball BA.MicroRNAs (miRNAs) are small, non-coding RNAs which are produced throughout the body. Individual tissues tend to have a specific expression profile and excrete many of these miRNAs into circulation. These circulating miRNAs may be diagnostically valuable biomarkers for assessing the presence of disease while minimizing invasive testing. In women, numerous circulating miRNAs have been identified which change significantly during pregnancy-related complications (e.g. chorioamnionitis, eclampsia, recurrent pregnancy loss); however, no prior work has been done in this area in the horse. To identif...
Fragkaki AG, Kioukia-Fougia N, Kiousi P, Kioussi M, Tsivou M.The artificial increase of the physical capability of horses using drugs is well known in racing and other equine sports. Both illicit and therapeutic substances are regarded as prohibited substances in competition in most countries. Some countries make distinctions for a few, specific drugs which are, however, allowed for use in other countries. The primary objective in the case of doping control is the detection of any trace of drug exposure, either parent drug or any of its metabolites, using the most powerful analytical methods which are generally based on chromatographic/mass spectrometri...
Baskerville CL, Bamford NJ, Harris PA, Bailey SR.Insulin-like growth factor-1 (IGF-1) plays several important physiological roles, and IGF-related pathways have been implicated in developmental osteochondral disease and endocrinopathic laminitis. This factor is also a downstream marker of growth hormone activity and its peptide mimetics. Unfortunately, previously used assays for measuring equine IGF-1 (radioimmunoassays and ELISAs) are no longer commercially available, and many of the kits on the market give poor results when used on horse samples. The aim of the present study was to compare three different ELISA assays (two human and one ho...
Kuwano A, Niwa H, Arai K.To establish a new system to isolate keratolytic bacteria from the hoof wall cavity () of a racehorse, we invented the horn powder agar-translucency (HoPAT) test and horn zymography (HZ). Using routine bacteriological techniques and these methods, we isolated five strains of keratolytic soil bacteria, which were then identified by means of 16S ribosomal RNA (rRNA) gene sequencing analysis. The findings from the study on the horse suggested that Brevibacterium luteolum played the main role in the local fragility of the hoof, eventually forming a in coordination with four other strains of kerat...
Wang J, Wen S, Zhao R, Qi J, Liu Z, Li W, An J, Wood C, Wang Y.The conjugation of small ubiquitin-like modifier (SUMO) to the target protein, namely, SUMOylation, is involved in the regulation of many important biological events including host-pathogen interaction. Some viruses have evolved to exploit the host SUMOylation machinery to modify their own protein. Retroviral Gag protein plays critical roles in the viral life cycle. The HIV-1 p6 and the Moloney murine leukemia virus CA have been reported to be conjugated with SUMO. In this study, we report for the first time, to our knowledge, the covalent conjugation of equine infectious anemia virus (EIAV) G...
Gim JA, Kim HS.MicroRNAs (miRNAs) are single-stranded, small RNAs (21-23 nucleotides) that function in gene silencing and translational inhibition via the RNA interference mechanism. Most miRNAs originate from host genomic regions, such as intergenic regions, introns, exons, and transposable elements (TEs). Here, we focused on the palindromic structure of medium reiteration frequencies (MERs), which are similar to precursor miRNAs. Five MER consensus sequences (MER5A1, MER53, MER81, MER91C, and MER117) were matched with paralogous transcripts predicted to be precursor miRNAs in the horse genome (equCab2) and...
Sandbaumhüter FA, Theurillat R, Bettschart-Wolfensberger R, Thormann W.The combination of ketamine and an α -receptor agonist is often used in veterinary medicine. Four different α -receptor agonists, medetomidine, detomidine, xylazine, and romifidine, which differ in their chemical structure and thus in selectivity for the α -receptor and in the sedative and analgesic potency, are typically employed during surgery of equines. Recovery following anesthesia with ketamine and an α -receptor agonist is dependent on the α -receptor agonist. This prompted us to investigate (i) the inhibition characteristics for the N-demethylation of ketamine to norketamine and (...
Chaney SB, Marsh AE, Lewis S, Carman M, Howe DK, Saville WJ, Reed SM.Equine protozoal myeloencephalitis (EPM) remains a significant central nervous system disease of horses in the American continents. Sarcocystis neurona is considered the primary causative agent and its intermediate life stages are carried by a wide host-range including raccoons (Procyon lotor) in North America. S. neurona sarcocysts mature in raccoon skeletal muscle and can produce central nervous system disease in raccoons, mirroring the clinical presentation in horses. The study aimed to develop laboratory tools whereby the life cycle and various life stages of S. neurona could be better stu...
Serafini R, Varner DD, Bissett W, Blanchard TL, Teague SR, Love CC.The effect of flash-freezing storage temperature on stallion sperm DNA has not been evaluated. Commonly, sperm are flash-frozen at various temperatures to preserve sperm DNA prior to analysis. It is unclear whether the temperature at which sperm are frozen and stored may affect the results of DNA assays. In this study, the neutral comet assay was used to evaluate the effect of flash-freezing storage temperature (freezer [-60 °C], dry ice [-78.5 °C], liquid nitrogen [-196 °C]) compared to fresh sperm DNA structure. In addition, intra- and inter-assay and intra- and inter-stallion variabil...
Heras S, Smits K, De Schauwer C, Van Soom A.Global epigenetic reprogramming is considered to be essential during embryo development to establish totipotency. In the classic model first described in the mouse, the genome-wide DNA demethylation is asymmetric between the paternal and the maternal genome. The paternal genome undergoes ten-eleven translocation (TET)-mediated active DNA demethylation, which is completed before the end of the first cell cycle. Since TET enzymes oxidize 5-methylcytosine to 5-hydroxymethylcytosine, the latter is postulated to be an intermediate stage toward DNA demethylation. The maternal genome, on the other ha...
Schnobrich MR, Gordon DL, Scoggin CF, Bradecamp EA, Canisso IF.Urine pooling, as a persistent condition, is a cause of infertility in mares due to endometrial inflammation and sperm toxicity. Identification of urometra can be challenging in mares presenting with the condition intermittently, or when urine flows into the uterus but is undetectable in the vagina. Currently, there are no reported objective methods to confirm the clinical diagnosis of urine contamination in intrauterine-fluid accumulations. Since creatinine is present in high concentrations in urine and does not diffuse across cell membranes, creatinine concentration should be increased in ma...
Steinbusch MM, Fang Y, Milner PI, Clegg PD, Young DA, Welting TJ, Peffers MJ.The development of effective treatments for the age-related disease osteoarthritis and the ability to predict disease progression has been hampered by the lack of biomarkers able to demonstrate the course of the disease. Profiling the expression patterns of small nucleolar RNAs (snoRNAs) in joint ageing and OA may provide diagnostic biomarkers and therapeutic targets. This study determined expression patterns of snoRNAs in joint ageing and OA and examined them as potential biomarkers. Using SnoRNASeq and real-time quantitative PCR (qRT-PCR) we demonstrate snoRNA expression levels in murine age...
Damm JB, Voshol H, Hård K, Kamerling JP, Vliegenthart JF.The carbohydrate chains of equine fibrinogen were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides obtained were fractionated by a combination of FPLC and HPLC and analyzed by 500-MHz 1H-NMR spectroscopy. Four monosialo and four disialo diantennary N-acetyllactosamine type of carbohydrate chains occur: (formula; see text)
Bosu WT, Van Camp SC, Miller RB, Owen RR.A five year prospective study of equine ovarian problems requiring surgical correction was undertaken at the Ontario Veterinary College. Thirty mares were studied, of which 14 had granulosa cell tumors, six were with anovulatory persistent follicular "structures", five had ovarian hemotoma, two presented ovarian hypoplasia and one each of ovarian dysgerminoma, teratoma and abscessation. The clinical signs manifested by the affected animals were varied. The affected ovaries were removed via flank or midline laparotomy or through colpotomy. Their morphology was studied and representative portion...
Musilova P, Kubickova S, Zrnova E, Horin P, Vahala J, Rubes J.Using laser microdissection we prepared a set of horse chromosome arm-specific probes. Most of the probes were generated from horse chromosomes, some of them were derived from Equus zebra hartmannae. The set of probes were hybridized onto E. grevyi chromosomes in order to establish a genome-wide chromosomal correspondence between this zebra and horse. The use of arm-specific probes provided us with more information on the mutual arrangement of the genomes than we could obtain by means of whole-chromosome paints generated by flow sorting, even if we used reciprocal painting with probe sets from...
King SS, Abughazaleh AA, Webel SK, Jones KL.Fatty acids of the n-3 type confer health benefits to humans and other species. Their importance to equine physiology could include improved exercise tolerance, decreased inflammation, and improved reproductive function. The circulating fatty acid profile and the acquisition and washout of fatty acids in response to n-3 supplementation were determined for horses in the current study. A fatty acid supplement high in eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid was fed to deliver EPA plus DHA at 0 (control), 10, 20, or 40 g/d to 16 mares (n = 4/group) for 28 d. Plasma was collected at -...
Schejter A, Ryan MD, Blizzard ER, Zhang C, Margoliash E, Feinberg BA.Contrary to most heme proteins, ferrous cytochrome c does not bind ligands such as cyanide and CO. In order to quantify this observation, the redox potential of the ferric/ferrous cytochrome c-cyanide redox couple was determined for the first time by cyclic voltammetry. Its E0' was -240 mV versus SHE, equivalent to -23.2 kJ/mol. The entropy of reaction for the reduction of the cyanide complex was also determined. From a thermodynamic cycle that included this new value for the cyt c cyanide complex E0', the binding constant of cyanide to the reduced protein was estimated to be 4.7 x 10(-3) L M(...
Sacchi L, Corona S, Gajadhar AA, Pozio E.The nurse cell-larva complex of nematodes of the genus Trichinella plays an important role in the survival of the larva in decaying muscles, frequently favouring the transmission of the parasite in extreme environmental conditions. The ultrastructure of the nurse cell-larva complex in muscles from different hosts infected with T. nativa (a walrus and a polar bear), T. spiralis (horses and humans), T. pseudospiralis (a laboratory mouse) and T. papuae (a laboratory mouse) were examined. Analysis with transmission electron microscope showed that the typical nurse cell structure was present in all...
Yao H, Osterrieder N, O'Callaghan DJ.The EICP0 gene (gene 63) of equine herpesvirus 1 (EHV-1) encodes an early regulatory protein that is a promiscuous trans-activator of all classes of viral genes. Bacterial artificial chromosome (BAC) technology and RecE/T cloning were employed to delete the EICP0 gene from EHV-1 strain KyA. Polymerase chain reaction, Southern blot analysis, and DNA sequencing confirmed the deletion of the EICP0 gene and its replacement with a kanamycin resistance gene in mutant KyA. Transfection of rabbit kidney cells with the EICP0 mutant genome produced infectious virus, indicating that the EICP0 gene is not...
Pusterla N, Mapes S, Johnson C, Slovis N, Page A, Gebhart C.The purpose of the current study was to compare the molecular detection rate of Lawsonia intracellularis between feces and rectal swabs collected from 42 foals with suspected equine proliferative enteropathy (EPE). Fecal samples and rectal swabs were processed for DNA purification by using an automated extraction system. The purified DNA was then analyzed by real-time polymerase chain reaction (PCR) for the presence of the aspartate ammonia lyase (aspA) gene of L. intracellularis. Absolute quantitation was calculated by using a standard curve for L. intracellularis and expressed as copy number...
Shao H, Robek MD, Threadgill DS, Mankowski LS, Cameron CE, Fuller FJ, Payne SL.The macrophage tropic lentivirus, equine infectious anemia virus (EIAV), encodes a dUTPase in the pol gene that is required for efficient replication in macrophages. Two naturally occurring variants of the enzyme were expressed as recombinant proteins in Escherichia coli; metal chelate affinity chromatography was used to purify histidine-tagged recombinant enzymes to greater than 80% homogeneity in a single chromatographic step. Biochemical and enzymatic analyses of these preparations suggest that this method yields dUTPase that is suitable for detailed mutational analysis. Specific activities...
Morozova-Roche LA.Calcium-binding equine lysozyme (EL) combines the structural and folding properties of c-type lysozymes and alpha-lactalbumins, connecting these two most studied subfamilies. The structural insight into its native and partially folded states is particularly illuminating in revealing the general principles of protein folding, amyloid formation and its inhibition. Among lysozymes EL forms one of the most stable molten globules and shows the most uncooperative refolding kinetics. Its partially-folded states serve as precursors for calcium-dependent self-assembly into ring-shaped and linear amyloi...
Pereira GR, Lorenzo PL, Carneiro GF, Ball BA, Gonçalves PB, Pegoraro LM, Bilodeau-Goeseels S, Kastelic JP, Casey PJ, Liu IK.The objective of this study was to test the hypothesis that equine growth hormone (eGH), in combination with insulin growth factor-I (IGF-I), influences positively in vitro nuclear and cytoplasmic maturation of equine oocytes. Cumulus-oocyte complexes were recovered from follicles that were < 25 mm in diameter, characterized by morphology and were allocated randomly as follow: (a) control (no additives); (b) 400 ng/ml eGH; (c) 200 ng/ml IGF-I; (d) eGH + IGF-I; and (e) eGH + IGF-I + 400 ng/ml anti-IGF-I antibody. Oocytes were matured for 30 h at 38.5°C in air with 5% CO2 and then stained wi...
Burns GA, Cummings JF.Sellers et al. (1979, Am. J. Phys., 237: E457-E464) proposed a pelvic flexure pacemaker mechanism to account for the bidirectional contraction waves needed to both retain ingesta within the right ventral colon for cellulose digestion and terminal fermentation and to transport the digesta distad once the process has been completed. To corroborate the presence of a pelvic flexure pacemaker, we prepared whole mount samples of the tunica muscularis from 23 horses at ten sites along the large colon, cecum and jejunum. Following smooth muscle enzymatic digestion, somata of the myenteric plexus were ...
Cook VL, Neuder LE, Blikslager AT, Jones SL.The effect of lidocaine on in vitro migration and adhesion of equine neutrophils was evaluated. Neutrophils were isolated from equine whole blood using a Percoll-gradient centrifugation protocol. Purified neutrophils were incubated with lidocaine at concentrations from 0.1 to 1000 microg/ml for 30 min at 37 degrees C, after calcein loading. Neutrophil integrin-mediated adhesion in response to stimulation with 100 nM LTB(4), 100 nM PAF, or 100 ng/ml IL-8, or integrin-mediated migration in response to stimulation with 100 nM LTB(4), 150 nM PAF, or 100 ng/ml IL-8 was assessed. Statistical signifi...
Calvete JJ, Mann K, Schäfer W, Sanz L, Reinert M, Nessau S, Raida M, Töpfer-Petersen E.We report the complete amino acid sequence of HSP-1, a major protein isolated from stallion seminal plasma or acid extracts of ejaculated spermatozoa. The protein consists of 121 amino acids organized in two types of homologous repeats arranged in the pattern AA'BB'. Each of the 13-15-residue A-type repeats contains two O-linked oligosaccharide chains. The B-type repeats span 44-47 amino acids each, are not glycosylated, and have the consensus pattern of the gelatin-binding fibronectin type-II module. This domain also occurs in the major bovine seminal plasma heparin-binding proteins PDC-109 (...
Laverty S, Okouneff S, Ionescu M, Reiner A, Pidoux I, Webber C, Rossier Y, Billinghurst RC, Poole AR.Articular osteochondrosis (OCD) occurs in both man and animals. The etiology remains to be determined. Studies of OCD lesions in animals may provide clues as to its pathogenesis. The aim of our study was to determine whether there was evidence for increased degradation namely proteoglycan (PG) release and type II collagen cleavage in articular cartilage harvested from OCD lesions. We examined ex vivo explants at post-mortem from equine OCD lesions and macroscopically normal site and age matched cartilage. These were cultured over a 10 day period in serum-free medium. Type II collagen cleavage ...
Bailly-Chouriberry L, Cormant F, Garcia P, Lönnberg M, Szwandt S, Bondesson U, Popot MA, Bonnaire Y.Recombinant human erythropoietin (rHuEPO) is a 30-34 kDa glycoprotein banned by the racing authorities. For some years this molecule has been detected in race horses in USA and in Europe, and even in racing camels. Although direct methods to differentiate horse endogenous EPO and rHuEPO have been developed either by LC-MS/MS or by isoelectric focusing (IEF) with double-blotting, the short confirmation time of such prohibited hormone in plasma remains a problem for horseracing doping control laboratories. In order to improve the rHuEPOs confirmation process in horse plasma or urine in terms of ...
Azarpeykan S, Dittmer KE.Housekeeping genes (HKGs) are used as internal controls for normalising and calculating the relative expression of target genes in RT-qPCR experiments. There is no unique universal HKG and HKGs vary among organisms and tissues, so this study aimed to determine the most stably expressed HKGs in the equine kidney. The evaluated HKGs included 18S ribosomal RNA (18S), 28S ribosomal RNA (28S), ribosomal protein L32 (RPL32), β-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), succinate dehydrogenase complex (SDHA), zeta polypeptide (YWHAZ), and hypoxanthine phosphoribosyltran...
Hara A, Taketomi T.Equine renal glycopshingolipids were composed of galactocerebroside, glucocerbroside, ceramide dihexoside, ceramide trihexoside, sulfatide, globoside I, Forssman globoside, and hematoside. Free ceramide and sphingomyelin were also found in equine kidney. Their long chain bases consisted of sphingosine, dihydrosphingosine, C18-phytosphingosine, and C20-phytosphingosine, whereas the fatty acids were separated into two groups: nonhydroxy and hydroxy fatty acids. Ceramide monohexoside was separated into five spots by TLC on borax-impregnated plates. The major component of ceramide monohexoside was...
Glöckner WM, Newman RA, Dahr W, Uhlenbruck G.Phenol extraction of horse, sheep, cow, pig and human erythrocyte membranes and human milk fat globule membranes gave glycoprotein fractions, all of which were shown by gas chromatography to contain the reduced disaccharide beta-D-galactosyl (1-3)-N-acetyl-D-galactosaminital after treatment with alkaline borohydride. Cow and pig erythrocyte membrane glycoproteins were found however to contain much lower amounts than the erythrocyte membrane glycoproteins of the other species tested. After gel filtration, a tetrasaccharide was isolated from horse and sheep glycoproteins containing the disacchar...
Benbarek H, Deby-Dupont G, Deby C, Caudron I, Mathy-Hartert M, Lamy M, Serteyn D.The activation of human polymorphonuclear leucocytes (the respiratory burst) can be studied by measuring their chemiluminescent response. This technique was adapted to equine leucocytes to investigate the effects of cell number, activator concentration, enhancers of chemiluminescence, pH, temperature and inhibitors. Leucocytes were isolated from citrated blood from healthy horses and chemiluminescence was measured with a Bio-Orbit luminometer sensitive to 900 nm light. The optimal cell density for the maximal chemiluminescent response ranged from 10(6) to 10(7) leucocytes 600 microliters-1. Ch...
Grosenbaugh DA, Gadawski JE, Muir WW.Evaluation of a portable clinical analyzer for determination of blood gas tensions, electrolyte and glucose concentrations, and Hct in a hospital setting. Methods: Prospective study. Methods: 50 dogs, 50 cats, and 28 horses, all clinically normal. Methods: Blood samples were analyzed on a portable clinical analyzer to determine concentrations of sodium, potassium, chloride, BUN, glucose, and ionized calcium and values of Hct, pH, PCO2, and PO2. Values obtained were compared with those obtained from the same blood samples, using a standard automatic analyzer (serum sodium, potassium, chloride, ...
Kinsley R, Scott SD, Daly JM.Serological assays provide an indirect route for the recognition of infectious agents via the detection of antibodies against the infectious agent of interest within serum. Serological assays for equine influenza A virus can be applied for different purposes: diagnosing infections; subtyping isolates; surveillance of circulating strains; and to evaluate the efficacy of vaccines before they reach the market. Haemagglutination inhibition (HI) and single radial haemolysis (SRH) assays are most commonly used in the equine field. This review outlines how both these assays together with virus neutra...
Schnabel CL, Fletemeyer B, Lübke S, Marti E, Wagner B, Alber G.Despite the relevance of adaptive immunity against equine pathogens antigen-specific T cell responses of horses are not well characterized and the lack of insight into T cell responses hampers the understanding of the pathogeneses of important diseases. In this study we used tetanus toxoid (TT) as a well-defined antigen to characterize antigen-reactive T cells. Six healthy adult horses received a routine booster against tetanus with an immune stimulating complex (ISCOM)-based vaccine and were followed for 28 days. TT-specific serum antibodies were quantified by ELISA and increased in all horse...
Wang N, Zhang D, Deng X, Sun Y, Wang X, Ma P, Song D.Herein we report a novel polydopamine-silver nanoparticle-polydopamine-gold (PDA-AgNPs-PDA-Au) film based surface plasmon resonance (SPR) biosensor for horse IgG detection. The PDA-AgNPs-PDA-Au film sensing platform was built on Au-film via layer-by-layer self-assembly. Ag ion was reduced in situ to AgNPs in presence of PDA. The top PDA layer can prevent AgNPs from being oxidized and connect with antibody via Schiff alkali reaction directly. The morphology and thickness of the modified gold film were characterized using scanning electron microscope and Talystep. Experimental results show that ...
Rivero JL, Talmadge RJ, Edgerton VR.In adult horses, three myosin heavy chain (MyHC) isoforms can be identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunohistochemistry using specific anti-MyHC monoclonal antibodies. This report studies the suitability of a consistent SDS-PAGE technique for quantifying MyHC profiles in homogenized cryostate sections of equine gluteus medius muscle biopsies (n = 18). The method used (previously described by R. J. Talmadge and R. R. Roy; J. Appl. Physiol. 1993, 75, 2337-2340) resolved MyHCs in three bands: I, IIB or IIX, and IIA from the fastest to the slowe...
Jülke H, Veit C, Ribitsch I, Brehm W, Ludewig E, Delling U.The purpose of this study was to evaluate the use of three different superparamagnetic iron oxide (SPIO) particles for labeling of ovine and equine bone marrow (BM)-derived multipotent stromal cells (MSCs) in vitro. MSCs were obtained from five adult sheep and horses, respectively. After three passages (p3), cells were labeled with either 1) Molday ION Rhodamine B, 2) Endorem, 3) Resovist, or 4) remained unlabeled as control. Labeling efficiency, marker retention, and long-term detectability in MRI until p7 were evaluated. Further, proliferation capacity and trilineage differentiation as indic...
Karàdi I, Kostner GM, Gries A, Nimpf J, Romics L, Malle E.Earlier studies demonstrated that lipoprotein (a), a lipoprotein of high atherogenicity, possesses proteolytic activity. In this report, we provide evidence that the lipoprotein (a)-specific antigen, apoprotein (a) is immunochemically related to plasminogen. This was demonstrated by polyclonal antisera from rabbit, sheep and horse, and with three monoclonal antibodies from mouse. Using immunospecific adsorbers against lipoprotein (a), all plasminogen could be adsorbed from lipoprotein (a)-positive and apparently lipoprotein (a)-negative plasma. As an additional similarity to plasminogen, lipop...
Lind C, Chen J, Byrjalsen I.We developed a rapid, competitive enzyme immunoassay (EIA) for measuring 25-hydroxyvitamin D3 [25(OH)D3] in serum. The EIA was based upon 25(OH)D3-3-hemisuccinate covalently coupled to secondary amino groups grafted onto the polystyrene surface of microtiter wells. Optimal coupling conditions were established, and we found that inclusion of 40 mumol/L chloramine T, an agent not previously described for use in coupling to these plates, resulted in both more reproducible coupling as well as more than a twofold increase in the coupling efficiency. Before EIA, 25(OH)D3 was extracted from the serum...
Demoruelle K, Guo B, Kao S, McDonald HM, Nikic DB, Holman SC, Wilson WW.The Haas - Drenth - Wilson (HDW) (Haas et al., 1999) theoretical model was used to correlate osmotic second virial coefficient (B) values with solubility (S) values for equine serum albumin (ESA) and ovalbumin for corresponding solution conditions. The best fit from the theoretical model was compared to experimental S versus B data. B values were experimentally measured using static light scattering. Solubilities of ESA were estimated using a sitting drop method. When the experimental data for S versus B were plotted, an excellent fit for ESA was obtained according to the HDW model. The result...
Vergari C, Ravary-Plumioën B, Evrard D, Laugier P, Mitton D, Pourcelot P, Crevier-Denoix N.Axial speed of sound (SOS) measurements have been successfully applied to noninvasively evaluate tendon load, while preliminary studies showed that this technique also has a potential clinical interest in the follow up of tendon injuries. The ultrasound propagation theory predicts that the SOS is determined by the effective stiffness, mass density and Poisson's ratio of the propagating medium. Tendon stiffness characterizes the tissue's mechanical quality, but it is often measured in quasi-static condition and for entire tendon segments, so it might not be the same as the effective stiffness w...
