Proteins are essential macromolecules that play diverse roles in the physiology and health of horses. They are composed of amino acids and are involved in various biological processes, including tissue growth, repair, and the synthesis of enzymes and hormones. Dietary proteins are a key component of equine nutrition, influencing muscle development, immune function, and overall performance. Horses require a balanced intake of essential amino acids, which must be obtained through their diet, as they cannot be synthesized endogenously. This page compiles peer-reviewed research studies and scholarly articles that explore the types, functions, and dietary requirements of proteins in horses, as well as their impact on equine health and performance.
Bourebaba L, Serwotka-Suszczak A, Bourebaba N, Zyzak M, Marycz K.Hyperactivation of protein tyrosine phosphatase (PTP1B) has been associated with several metabolic malfunctions ranging from insulin resistance, metaflammation, lipotoxicity, and hyperglycaemia. Liver metabolism failure has been proposed as a core element in underlying endocrine disorders through persistent inflammation and highly fibrotic phenotype. Unassigned: In this study, the outcomes of PTP1B inhibition using trodusquemine (MSI-1436) on key equine metabolic syndrome (EMS)-related alterations including inflammation, fibrosis, and glucose uptake have been analyzed in liver explants collect...
Alshut F, Venner M, Martinsson G, Vervuert I.Electrolyte supplementation may be a risk factor for gastric mucosal lesions, but relevant evidence is limited in horses. Objective: Investigate the effects of PO sodium chloride (NaCl) supplementation on the gastric mucosa of exercising horses. We hypothesized that NaCl supplementation would neither cause nor exacerbate existing gastric mucosal damage. Methods: Fifteen 3-year-old healthy Warmblood stallions from a stud farm. Methods: Placebo-controlled study with a crossover design. Horses were fed either a NaCl pellet at a dosage adequate to replace the electrolyte losses in 10 L sweat or ...
Riihimäki M, Fegraeus K, Nordlund J, Waern I, Wernersson S, Akula S, Hellman L, Raine A.Equine asthma (EA) is a heterogenous, complex disease, with a significant negative impact on horse welfare and performance. EA and human asthma share fundamental similarities, making EA a useful model for studying the disease. One relevant sample type for investigating chronic lung inflammation is bronchoalveolar lavage fluid (BALF), which provides a snapshot of the immune cells present in the alveolar space. To investigate the immune cell landscape of the respiratory tract in horses with mild-to-moderate equine asthma (mEA) and healthy controls, single-cell RNA sequencing was conducted on equ...
Jung HS, Yun JU, Jung MJ, Song HS, Kim YB, Kim Y, Kim JG, Jeon CO, Roh SW, Whon TW, Lee SH.A novel, anaerobic, Gram-stain-positive coccoid strain, CBA3646, was isolated from the faeces of a thoroughbred racehorse. Phylogenetic analysis based on 16S rRNA gene sequencing yielded results indicative of CBA3646 representing a member of the genus , with the species most closely related to it being DSM 20463, with a similarity of 94.79 %. DNA-DNA relatedness and average nucleotide identity values between CBA3646 and DSM 20463 were 21.4 and 67.6 %, respectively. CBA3646 has a circular chromosomal genome of 1 709 189 bp (45.5 mol% DNA G+C content), containing 1652 genes in tota...
Jammes M, Cassé F, Velot E, Bianchi A, Audigié F, Contentin R, Galéra P.Osteoarthritis (OA) is a widespread osteoarticular pathology characterized by progressive hyaline cartilage degradation, exposing horses to impaired well-being, premature career termination, alongside substantial financial losses for horse owners. Among the new therapeutic strategies for OA, using mesenchymal stromal cell (MSC)-derived exosomes (MSC-exos) appears to be a promising option for conveying MSC therapeutic potential, yet avoiding the limitations inherent to cell therapy. Here, we first purified and characterized exosomes from MSCs by membrane affinity capture (MAC) and size-exclusio...
Borovikov S, Ryskeldina A, Tursunov K, Syzdykova A, Akibekov O. abortion in mares is caused by subspecies serovar infection and is characterized by premature (abortion) or non-viable fetus birth. Although all horses are susceptible to infection, the condition is more often clinically manifested in pregnant mares, with most abortions recorded in young females. In addition, nonspecific clinical disease signs and poorly sensitive and effective bacteriological diagnostic methods hinder rapid and reliable infection diagnoses. Immunochemical methods such as enzyme-linked immunosorbent assay (ELISA) and immunochromatography assays can facilitate effective and...
Bourebaba N, Sikora M, Qasem B, Bourebaba L, Marycz K.Equine metabolic syndrome (EMS), which encompasses insulin resistance, low-grade inflammation and predisposition to laminitis is a critical endocrine disorder among the most prevalent conditions affecting horses from different breeds. According to the most recent research, low human sex hormone-binding globulin (SHBG) serum levels correlate with an increased risk of obesity, insulin resistance and diabetes, and may contribute to overall metabolic dysregulations. This study aimed to test whether exogenous SHBG could protect EMS affected adipose-derived stromal stem cells (EqASC) from apoptosis,...
Tau RL, Ferreccio C, Bachir N, Torales F, Romera SA, Maidana SS.High-throughput sequencing of genomes has expanded our knowledge of the Alphaherpesvirinae, a widely extended subfamily of DNA viruses that recombine to increase their genetic diversity. It has been acknowledged that equid herpesvirus 1 (EHV-1) and equid herpesvirus 4 (EHV-4), two alphaherpesviruses with an economic impact on the horse industry, can recombine. This work aimed to analyze interspecific recombination between all equid alphaherpesvirus species, using genomes of EHV-1, EHV-3, EHV-4, EHV-6, EHV-8, and EHV-9 available in GenBank. 14 events of recombination by RDP4 and Simplot between...
da Silva CMB, Cano FEM, Gaitskell-Phillips G, Vega FJP.Flow cytometry is a powerful tool for the analysis of cell samples formed of multipopulations, such as spermatozoa. In recent years, multiparametric cytometers have evolved, allowing the study of different cellular characteristics, such as protein expression, DNA analysis, or mitochondrial activity. Whether using traditional fluorescent dyes or fluorophore-conjugated antibodies, each cell or cellular component is individually stained, the sample is analyzed at high velocities, and then is displayed and interpreted in a dot-plot. We hereby describe the procedure to perform a multiparametric flo...
Jammes M, Contentin R, Audigié F, Cassé F, Galéra P. Osteoarthritis (OA) is an invalidating articular disease characterized by cartilage degradation and inflammatory events. In horses, OA is associated with up to 60% of lameness and leads to reduced animal welfare along with extensive economic losses; currently, there are no curative therapies to treat OA. The mesenchymal stromal cell (MSC) secretome exhibits anti-inflammatory properties, making it an attractive candidate for improving the management of OA. In this study, we determined the best storage conditions for conditioned media (CMs) and tested whether priming MSCs with cytokines can enh...
Xing J, Xie L, Qi X, Liu G, Akhtar MF, Li X, Bou G, Bai D, Zhao Y, Dugarjaviin M, Zhang X.The proliferation and differentiation of skeletal muscle satellite cells (SCs) are necessary for the development of mature skeletal muscle. Leucine (Leu) is both an essential amino acid (EAA) and a branched-chain amino acid (BCAA), which has attracted worldwide attention due to its ability to repair and become new fibers. We separated the equine SCs into the control group (CON) and the Leu-supplemented group (LEU), which the cells were cultured in Leu-deprived and Leu-supplemented media respectively. We combined the transcriptome (RNA-Seq) and quantitative proteome (TMT) profiling analyses on ...
Vidal Moreno de Vega C, Lemmens D, de Meeûs d'Argenteuil C, Boshuizen B, de Maré L, Leybaert L, Goethals K, de Oliveira JE, Hosotani G, Deforce D....Important changes in glucose transporter (GLUT) expression should be expected if the glucose influx plays a pivotal role in fuelling or connecting metabolic pathways that are upregulated in response to exercise. The aim was to assess GLUT4, 8, and 12 dynamics in response to training and acute exercise. Sixteen untrained Standardbred mares (3-4 year) performed an incremental SET at the start and end of 8 weeks harness training. M. pectoralis (PM) and M. vastus lateralis (VL) muscle biopsies were taken before and after each SET, allowing for comparing rest and acute samples in untrained (UT) ...
Bockisch F, Taubert J, Coenen M, Vervuert I.The German Society of Nutrition Physiology has proposed a new protein evaluation system for horse feeds to estimate pre-cecally digestible crude protein (pcdCP) and amino acids (pcdAA) from chemical properties. A total of 71 feeds for horses were chemically tested and evaluated according to the new protein evaluation system. A feeding trial with eight horses tested whether differences in estimated pcdAA and neutral detergent soluble CP (NDSCP) in the diet were reflected by post-prandial (ppr) kinetics of plasma lysine (Lys) by feeding a complementary feed (control = CTRL) with 1.02 g Lys/100 k...
Iddagoda J, Gunasekara P, Handunnetti S, Jeewandara C, Karunatilake C, Malavige GN, de Silva R, Dasanayake D.It is clinically important to identify allergens in Artocarpus heterophyllus (jackfruit), Moringa oleifera (moringa), Trianthema portulacastrum (horse purslane) and Syzygium samarangense (rose apple). This study included 7 patients who developed anaphylaxis to jackfruit (1), moringa (2), horse purslane (3) and rose apple (1). We sought to determine allergens in the edible ripening stages of jackfruit (tender, mature, and ripened jackfruit) and seeds, edible parts of moringa (seeds, seedpod, flesh inside seedpod, and leaves), horse purslane leaves and ripened rose apple fruit. The persistence o...
Kolyganova TI, Arzumanyan VG, Matvienko MA, Rodionova AA, Korshunova DS, Shatunova PO, Yastrebova NE.Antimicrobial activity of milk whey in different mammals against Candida albicans yeast cells was studied by a spectrophotometric method. The activity increased in the order goat→horse→camel→cow→human→mouse. The level of whey activity in mice was higher by 3 and 10 times than in humans and goats, respectively. Similar changes were noted for activity of the whey fraction <100 kDa containing a complex of antimicrobial polypeptides, and there was a direct correlation between these two parameters (r=0.881; p<0.05). The total activity of whey had a high degree of correlation with th...
Bergmann R, Schroedl W, Müller U, Baums CG.Streptococcus equi subsp. zooepidemicus (SEZ) is a major equine pathogen that causes pneumonia, abortion, and polyarthritis. It can also cause invasive infections in humans. SEZ expresses the M-like protein SzM, which recruits host proteins such as fibrinogen to the bacterial surface. Equine SEZ strain C2, which binds only comparably low amounts of human fibrinogen in comparison to human SEZ strain C33, was previously shown to proliferate in equine and human blood. As the expression of SzM_C2 was necessary for survival in blood, this study investigated the working hypothesis that SzM_C2 inhibi...
Mogielnicka-Brzozowska M, Woźniak MJ, Cichowska AW, Fraser L, Kraziński B, Strzezek R, Zielińska D.Seminal plasma (SP) proteins are responsible for sperm functional quality. Developing a reliable method to determine the degree of oxidative damage of these proteins is important for establishing semen fertilizing ability. The main aim of the study was to verify the applicability of protein carbonyl derivatives measurement in the SP of canine and stallion, using a method with 2,4-dinitrophenylhydrazine (DNPH). The research material consisted of ejaculates obtained from eight English Springer Spaniels, and from seven half-blood stallions during the breeding and non-breeding season. The content ...
Cittadini A, Domínguez R, Sarriés MV, Pateiro M, Lorenzo JM.This study aimed to evaluate the effect of the total or partial replacement of NaCl by a commercial low-salt mixture Pansalt® (T1) or Kombu (L. ochroleuca) seaweed powder (T2), respectively, on the quality properties of dry-cured foal "cecina". Salt reformulation unaffected fat percentages, decreased (P < 0.001) moisture and ash contents, while protein amounts increased (P < 0.001) in the T2 batch. Considering the physicochemical variables, any significant differences were observed among batches, except for a*, water activity, and springiness values. T1 and T2 samples presented a si...
Ferreira LVO, Kamura BDC, Oliveira JPM, Chimenes ND, Carvalho M, Santos LAD, Dias-Melicio LA, Amorim RL, Amorim RM.Schwann cells (SCs) are essential for the regenerative processes of peripheral nerve injuries. However, their use in cell therapy is limited. In this context, several studies have demonstrated the ability of mesenchymal stem cells (MSCs) to transdifferentiate into Schwann-like cells (SLCs) using chemical protocols or co-culture with SCs. Here, we describe for the first time the in vitro transdifferentiation potential of MSCs derived from equine adipose tissue (AT) and equine bone marrow (BM) into SLCs using a practical method. In this study, the facial nerve of a horse was collected, cut into ...
Jaeser M, Wunderlich C, Henle T.To study the protein-bound glycans of equine κ-casein, equine sodium caseinate was first obtained from raw mare's milk by acid precipitation and then fractionated by cation-exchange chromatography. The oligosaccharides of the obtained equine κ-casein were analyzed by RP-HPLC-UV-HRMS after β-elimination with simultaneous derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP). In addition to the acidic tetrasaccharide derivative Neu5Ac-Gal-[Neu5Ac]-GalNAc-2PMP known from bovine κ-casein, the acidic pentasaccharide derivative Neu5Ac-Gal-[Gal-GlcNAc]-GalNAc-2PMP was identified as the most ab...
Hayes AM, Kastl B, Perry E, Moore AR, Springer NL.Myeloma-related disorders, including multiple myeloma, extramedullary plasmacytoma, and solid osseous plasmacytoma, are rare in horses. Clinical complaints for myeloma-related disorders are nonspecific, and when present, M-protein location is more variable on protein electrophoresis in horses relative to dogs and cats. Here, we describe a case of a 15-year-old Thoroughbred mare who presented with recurrent blepharitis. Marked hyperglobulinemia was an incidental finding on routine hematologic and biochemical testing. Bone marrow aspiration consisted of >30% plasma cells, and serum protein el...
Miller LMJ, Woodward EM, Campos JR, Squires EL, Troedsson MHT.Sperm protein at 22 kDa has been associated with fertility. Objective: The objectives of this study were to determine (1) the localization pattern of SP22 on ejaculated and caudal epididymal equine spermatozoa and in epididymal fluid, and to (2) characterize SP22 protein and mRNA expression in testicular and epididymal tissues in response to heat-induced testicular degeneration. Methods: Semen was collected before and after hemi-castration, as well as prior to and following insulation of the remaining testes, and tissue specimens were collected for analysis. Results: Histopathology confirmed ...
Arfuso F, Piccione G, Guttadauro A, Monteverde V, Giudice E, Giannetto C.Inflammaging or chronic, low-grade inflammation is a phenomenon characterizing age-related pathologies and natural processes in aging tissues. This study aimed to investigate the effect of age on the serum levels of C-reactive protein and protein electrophoretic pattern in horses. Thirty Italian Saddle horses from the same horse training center, were enrolled and divided in three equal groups according to their age: Group A (2-4 years), Group B (7-10 years), Group C (15-20 years). Blood samples were collected from each horse to evaluate hematological parameters and the serum concentration of C...
Terhaar HM, Henriksen ML, Mehaffy C, Hess A, McMullen RJ.The objective of this study was to use shotgun label-free tandem mass spectrometry (LF-MS/MS) to evaluate aqueous humor (AH) from horses with uveitis (UH) compared to ophthalmologically healthy horses (HH). Methods: Twelve horses diagnosed with uveitis based on ophthalmic examination and six ophthalmologically healthy horses (postmortem) purchased for teaching purposes. Methods: All horses received a complete ophthalmic examination and physical exam. Aqueous paracentesis was performed on all horses and AH total protein concentrations were measured with nanodrop (TPn) and refractometry (TPr). A...
Bronstein A, Marx A.Comparison of myoglobin structures reveals that protein isolated from horse heart consistently adopts an alternate turn conformation in comparison to its homologues. Analysis of hundreds of high-resolution structures discounts crystallization conditions or the surrounding amino acid protein environment as explaining this difference, that is also not captured by the AlphaFold prediction. Rather, a water molecule is identified as stabilizing the conformation in the horse heart structure, which immediately reverts to the whale conformation in molecular dynamics simulations excluding that structur...
Istanbullugil FR, Risvanli A, Salikov R, Bayraktar M, Kadiraliyeva N, Zhunushova A, Yilmaz O, Yuksel BF, Turanli M, Uz M, Asinbekovic MI.Immune compatibility between mare and foal is one of the important topics of reproductive immunology. At this point, although there are many studies on antibodies, there are not many publications on the relationship between the cytokine levels of mare, foal and milk and the effects of milk composition on this relationship. Here we investigate the relationship between Th1/Th2 cytokine balance and milk composition in Kyrgyz mares and foals. Samples were taken soon after the foal was born and on days 5, 10 and 20 after birth. Th1 (IFN-γ, IL-2, TNF-α) and Th2 (IL-4, IL-5, IL-10) cytokine levels ...
Alim S, Laitaoja M, Pawar SS, Talluri TR, Jänis J, Swamy MJ.Major proteins of the seminal plasma in a variety of mammals such as bovine PDC-109, equine HSP-1/2, and donkey DSP-1 contain fibronectin type-II (FnII) domains and are referred to as FnII family proteins. To further our understanding on these proteins, we carried out detailed studies on DSP-3, another FnII protein of donkey seminal plasma. High-resolution mass-spectrometric studies revealed that DSP-3 contains 106 amino acid residues and is heterogeneously glycosylated with multiple acetylations on the glycans. Interestingly, high homology was observed between DSP-1 and HSP-1 (118 identical r...
Bourebaba L, Serwotka-Suszczak A, Pielok A, Sikora M, Mularczyk M, Marycz K.Equine metabolic syndrome (EMS) is a multifactorial pathology gathering insulin resistance, low-grade inflammation and past or chronic laminitis. Among the several molecular mechanisms underlying EMS pathogenesis, increased negative insulin signalling regulation mediated by protein tyrosine phosphatase 1 B (PTP1B) has emerged as a critical axis in the development of liver insulin resistance and general metabolic distress associated to increased ER stress, inflammation and disrupted autophagy. Thus, the use of PTP1B selective inhibitors such as MSI-1436 might be considered as a golden therapeut...
Fedorka CE, Ali HE, Troedsson MHT.Galectins are proteins that bind to glycans in targeted cells and function in cell-to-cell signaling throughout the body. Galectins have been found to be involved in various reproductive processes, including placental dysfunction, but this has not been investigated in the horse. Therefore, the objective of this study was to assess alterations in galectin expression of the abnormal placenta in pregnant mares. Next-generation RNA sequencing was performed on the postpartum chorioallantois of two placental pathologies following clinical cases of ascending placentitis (n = 7) and focal mucoid place...
Semanchik PL, Wesolowski LT, Ryan PJ, White-Springer SH, Fluckey JD.Skeletal muscle is a highly dynamic organ that is essential for locomotion as well as endocrine regulation in all populations of horses. However, despite the importance of adequate muscle development and maintenance, the mechanisms underlying protein anabolism in horses on different diets, exercise programs, and at different life stages remain obscure. Mechanistic target of rapamycin (mTOR) is a key component of the protein synthesis pathway and is regulated by biological factors such as insulin and amino acid availability. Providing a diet ample in vital amino acids, such as leucine and gluta...
Fouchécourt S, Métayer S, Locatelli A, Dacheux F, Dacheux JL.Proteins present in and secreted into the lumen of various regions of the stallion epididymis were characterized qualitatively and quantitatively by two-dimensional electrophoresis. Using this proteomic approach, 201 proteins were found in the lumen and 117 were found that were secreted by the epithelium in various parts of the organ. Eighteen proteins made up 92.6% of the total epididymal secretory activity, lactoferrin (41.2%) and clusterin (24.8%) being the most abundant. Procathepsin D, HE1/CTP (cholesterol transfer protein), GPX (glutathione peroxidase), beta-N-acetyl-hexosaminidase, and ...
Gregoire C, Rosinski-Chupin I, Rabillon J, Alzari PM, David B, Dandeu JP.The gene encoding the major horse allergen, designated Equus caballus allergen 1 (Equ c1), was cloned from total cDNA of sublingual salivary glands by reverse transcription-polymerase chain reaction using synthetic degenerate oligonucleotides deduced from N-terminal and internal peptide sequences of the glycosylated hair dandruff protein. A recombinant form of the protein, with a polyhistidine tail, was expressed in Escherichia coli and purified by immobilized metal affinity chromatography. The recombinant protein is able to induce a passive cutaneous anaphylaxis reaction in rat, and it behave...
Peffers MJ, McDermott B, Clegg PD, Riggs CM.The aim of the study was to characterise the protein complement of synovial fluid (SF) in health and osteoarthritis (OA) using liquid chromatography mass spectrometry (LC-MS/MS) following peptide-based depletion of high abundance proteins. Methods: SF was used from nine normal and nine OA Thoroughbred horses. Samples were analysed with LC-MS/MS using a NanoAcquity™ LC coupled to an LTQ Orbitrap Velos. In order to enrich the lower-abundance protein fractions protein equalisation was first undertaken using ProteoMiner™. Progenesis-QI™ LC-MS software was used for label-free quantification. ...
Egberink HF, Ederveen J, Montelaro RC, Pedersen NC, Horzinek MC, Koolen MJ.Feline immunodeficiency virus (FIV) grown in cat lymphocyte and thymocyte cultures was labelled with L-[35S]methionine or [3H]glucosamine and virus-coded proteins were identified using immunoprecipitation. Polypeptides with apparent Mr values of 15K, 24K, 43K, 50K, 120K and 160K were detected. An additional polypeptide of 10K was detected by Western blot analysis. The two highest Mr species sometimes appeared as one band, of which only the 120K polypeptide was glycosylated. In the presence of tunicamycin gp120 was no longer detectable and a non-glycosylated precursor of 75K was found instead. ...
Chance MR, Sagi I, Wirt MD, Frisbie SM, Scheuring E, Chen E, Bess JW, Henderson LE, Arthur LO, South TL.Zinc finger arrays have been established as a critical structural feature of proteins involved in DNA recognition. Retroviral nucleocapsid proteins, which are involved in the binding of viral RNA, contain conserved cysteine-rich arrays that have been suggested to coordinate zinc. We provide metalloprotein structural data from an intact virus preparation that validate this hypothesis. Extended x-ray absorption fine structure (EXAFS) spectroscopy of well-characterized and active preparations of equine infectious anemia virus, compared with a peptide with known coordination and in combination wit...
Tang YD, Na L, Zhu CH, Shen N, Yang F, Fu XQ, Wang YH, Fu LH, Wang JY, Lin YZ, Wang XF, Wang X, Zhou JH, Li CY.Viperin is an endoplasmic reticulum (ER)-associated multifunctional protein that regulates virus replication and possesses broad antiviral activity. In many cases, viperin interferes with the trafficking and budding of viral structural proteins by distorting the membrane transportation system. The lentivirus equine infectious anemia virus (EIAV) has been studied extensively. In this study, we examined the restrictive effect of equine viperin (eViperin) on EIAV replication and investigated the possible molecular basis of this restriction to obtain insights into the effect of this cellular facto...
González-Fernández L, Macías-García B, Loux SC, Varner DD, Hinrichs K.Protein tyrosine phosphorylation (PY) is a hallmark of sperm capacitation. In stallion sperm, calcium inhibits PY at pH 7.8). These results show for the first time a role for calcium/calmodulin-dependent kinases in PRKA-dependent sperm PY; a non-PRKA-dependent pathway regulating sperm PY; and the apparent involvement of the FAK family of protein tyrosine kinases downstream in both pathways.
MacLeod JN, Burton-Wurster N, Gu DN, Lust G.Fibronectin is an extracellular matrix glycoprotein encoded by a single gene. Alternative RNA splicing has been reported at three sites, ED (extra type III domain)-A, ED-B, and the variable or V region. Articular cartilage fibronectin monomers are rarely (ED-A)+, but approximately 25% are (ED-B)+. RNA gel electrophoresis and Northern blot analysis identified two (ED-B)+ and two (ED-B)- fibronectin transcripts in cartilage, each pair differing by approximately 750 bases. This difference results from a previously unreported RNA splicing pattern that eliminates not only the V region but also nucl...
O'Callaghan DJ, Hyde JM, Gentry GA, Randall CC.Infection of exponential-phase suspension cultures of mouse fibroblast cells (L-M) with equine abortion virus (EAV) resulted in inhibition of cell growth and marked alterations in host metabolic processes. The synthesis of deoxyribonucleic acid (DNA) and ribonucleic acid was inhibited within 4 hr after infection and was suppressed by more than 90% by the time of maximal virus replication (14 to 18 hr). The overall rate of protein synthesis, however, was similar in uninfected and virus-producing cells as determined by measurements of net protein and isotope incorporation. The time course of vir...
Payne SL, Rushlow K, Dhruva BR, Issel CJ, Montelaro RC.Previous characterizations of equine infectious anemia virus (EIAV) glycoprotein variation by DNA sequence analysis and epitope mapping using monoclonal antibodies (MAbs) have revealed the presence of conserved and variable regions within the EIAV env gene. To extend these studies, fragments of the EIAV envelope proteins gp90 and gp45 were expressed in Escherichia coli and used in Western blot analysis with a diverse panel of equine immune sera to identify antigenic segments. All sera from EIAV-infected animals reacted with the carboxyl terminal portion of gp90 and the amino terminal portion o...
Malisauskas M, Zamotin V, Jass J, Noppe W, Dobson CM, Morozova-Roche LA.The calcium-binding equine lysozyme has been found to undergo conversion into amyloid fibrils during incubation in solution at acidic pH. At pH 4.5 and 57 degrees C, where equine lysozyme forms a partially unfolded molten globule state, the protein forms protofilaments with a width of ca. 2 nm. In the absence of Ca(2+) the protofilaments are present as annular structures with a diameter of 40-50 nm. In the presence of 10 mM CaCl(2) the protofilaments of equine lysozyme are straight or curved; they can assemble into thicker threads, but they do not appear to undergo circularisation. At pH 2.0, ...
Zembowicz A, Knoepp SM, Bei T, Stergiopoulos S, Eng C, Mihm MC, Stratakis CA.Pigmented epithelioid melanocytoma (PEM) is a recently described entity comprising most cases previously described as "animal-type melanoma" and epithelioid blue nevus (EBN) occurring in patients with the multiple neoplasia syndrome Carney complex (CNC). Mutations of the protein kinase A regulatory subunit type 1alpha (R1alpha) (coded by the PRKAR1A gene) are found in more than half of CNC patients. In this study, we investigated whether PEM and EBN are related at the molecular level, and whether changes in the PRKAR1A gene status and the expression of the R1alpha protein may be involved in th...
McClain AK, McCarrel TM.Platelet-rich plasma (PRP) is a therapeutic biologic that is used for treatment of musculoskeletal pathologies in equine athletes. Due to the expense of PRP kits, and the volumes obtained, freezing aliquots for future dosing is common. Aliquots of PRP are also commonly frozen for later analysis of growth factor concentrations in in vitro research. A variety of freezing methods are used and storage duration until analysis is often not reported. The optimal frozen storage conditions and duration to maintain concentrations of commonly measured growth factors and enzymes in PRP are unknown. Our ob...
Malisauskas M, Ostman J, Darinskas A, Zamotin V, Liutkevicius E, Lundgren E, Morozova-Roche LA.In amyloid diseases, it is not evident which protein aggregates induce cell death via specific molecular mechanisms and which cause damage because of their mass accumulation and mechanical properties. We showed that equine lysozyme assembles into soluble amyloid oligomers and protofilaments at pH 2.0 and 4.5, 57 degrees C. They bind thioflavin-T and Congo red similar to common amyloid structures, and their morphology was monitored by atomic force microscopy. Molecular volume evaluation from microscopic measurements allowed us to identify distinct types of oligomers, ranging from tetramer to oc...
Merchant KA, Thompson DE, Xu QH, Williams RB, Loring RF, Fayer MD.Two-dimensional (2D) infrared vibrational echoes were performed on horse heart carbonmonoxymyoglobin (MbCO) in water over a range of temperatures. The A(1) and A(3) conformational substates of MbCO are found to have different dephasing rates with different temperature dependences. A frequency-frequency correlation function derived from molecular dynamics simulations on MbCO at 298 K is used to calculate the vibrational echo decay. The calculated decay shows substantial agreement with the experimentally measured decays. The 2D vibrational echo probes protein dynamics and provides an observable ...
Creutz C, Sutin N.The kinetics of the reduction of horseheart ferricytochrome c by sodium dithionite (phosphate buffer-sodium chloride; pH 6.5, mu = 1.0, 25 degrees ) features two reaction pathways; one with the rate constant k(3) = 1.17 x 10(4) M(-1) sec(-1), the other with the rate constant k(1)k(2)/k(-1) = 6.0 x 10(4) M(-1) sec(-1). These pathways are interpreted in terms of remote attack (possibly by way of the exposed edge of the porphyrin system) and adjacent attack (requiring the opening of the heme crevice). The limiting rate for the adjacent pathway (k(1) = 30 sec(-1)) is in good agreement with the rat...
MacLachlan NJ, Balasuriya UB, Hedges JF, Schweidler TM, McCollum WH, Timoney PJ, Hullinger PJ, Patton JF.Equine arteritis virus (EAV) is the causative agent of equine viral arteritis, an apparently emerging disease of equids. In this study, the antibody response of horses to the structural proteins of EAV was evaluated using gradient-purified EAV virions and baculovirus-expressed recombinant EAV structural proteins (G(L), G(S), M, N) as antigens in a Western immunoblotting assay. Thirty-three sera from horses that previously had been naturally or experimentally infected with EAV were evaluated, including samples from mares, geldings, and both persistently and nonpersistently infected stallions. S...
Hatanaka H, Iourin O, Rao Z, Fry E, Kingsman A, Stuart DI.The Gag polyprotein is key to the budding of retroviruses from host cells and is cleaved upon virion maturation, the N-terminal membrane-binding domain forming the matrix protein (MA). The 2.8-A resolution crystal structure of MA of equine infectious anemia virus (EIAV), a lentivirus, reveals that, despite showing no sequence similarity, more than half of the molecule can be superimposed on the MAs of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV). However, unlike the structures formed by HIV-1 and SIV MAs, the oligomerization state observed is not trimeric...
González-Fernández L, Ortega-Ferrusola C, Macias-Garcia B, Salido GM, Peña FJ, Tapia JA.Protein tyrosine kinases have important roles in spermatozoa; however, little is known about the presence and regulation in these cells of their counterparts in signaling, namely, protein tyrosine phosphatases (PTPs) and dual-specificity phosphatases (DSPs). The objectives of the present study were to identify PTPs and DSPs in boar, stallion, and dog spermatozoa; to characterize their subcellular distribution; and to investigate the roles of tyrosine phosphatases in maintenance of protein tyrosine phosphorylation level and in sperm motility. Using Western blotting with specific antibodies in b...
Luo J, Bruice TC.Molecular dynamics and cross-correlation analysis of the horse liver alcohol dehydrogenase HLADH.NAD(+).PhCH(2)O(-) complex has established anticorrelated motions between the NAD(+)-binding domain and other portions of the enzyme. Four pairs of anticorrelated interactions are (i and ii) cofactor-binding domain: C(alpha) of V292 and the CG1 of V203 with C7 of PhCH(2)O(-); (iii) cofactor-binding domain: amide carbonyl oxygen of I318 with amide N of H67; and (iv) cofactor domain: C(alpha) of T178 with carbonyl oxygen of L141. The average distances between pairs are 9.2 A for i, 8.2 A for ii, 14.7...
Goodrich LR, Grieger JC, Phillips JN, Khan N, Gray SJ, McIlwraith CW, Samulski RJ.A gene therapeutic approach to treat osteoarthritis (OA) appears to be on the horizon for millions of people who suffer from this disease. Previously we described optimization of a scAAVIL-1ra gene therapeutic vector and initially tested this in an equine model verifying long-term intrasynovial IL-1ra protein at therapeutic levels. Using this vector, we carried out a dosing trial in six horses to verify protein levels and establish a dose that would express relevant levels of therapeutic protein for extended periods of time (8 months). A novel arthroscopic procedure used to detect green fluore...
Wang X, Kikuchi T, Rikihisa Y.Anaplasma phagocytophilum is an obligatory intracellular bacterium that causes human granulocytic anaplasmosis. The polymorphic 44-kDa major outer membrane proteins of A. phagocytophilum are dominant antigens recognized by patients and infected animals. However, the ability of anti-P44 antibody to neutralize the infection has been unclear due to a mixture of P44 proteins with diverse hypervariable region amino acid sequences expressed by a given bacterial population and lack of epitope-defined antibodies. Monoclonal antibodies (MAbs) 5C11 and 3E65 are directed to different domains of P44 prote...
Corradin G, Chiller JM.Murine T-lymphocyte specificity was determined in a system of antigen driven in vitro T-cell proliferation using cytochrome c molecules from different species, their derived peptides and reconstituted hybrid proteins. It was observed that primed T cells could discriminate between peptide fragments which differed from each other at a single amino acid residue. These conclusions were substantiated by the pattern of cross-reactivity noted in the response of closely related cytochrome c proteins as well as when artificial hybrid molecules reconstituted by the covalent linkage of peptide fragments ...
Suire S, Stewart F, Beauchamp J, Kennedy MW.The equine conceptus is surrounded by a fibrous capsule that persists until about day 20 of pregnancy, whereupon the capsule is lost, the conceptus attaches to the endometrium and placentation proceeds. Before attachment, the endometrium secretes in abundance a protein of the lipocalin family, uterocalin. The cessation of secretion coincides with the end of the period during which the conceptus is enclosed in its capsule, suggesting that uterocalin is essential for the support of the embryo before direct contact between maternal and foetal tissues is established. Using recombinant protein and ...
Kleinheinz J, Jung S, Wermker K, Fischer C, Joos U.Current approaches in bone regeneration combine osteoconductive scaffolds with bioactive cytokines like BMP or VEGF. The idea of our in-vitro trial was to apply VEGF165 in gradient concentrations to an equine collagen carrier and to study pharmacological and morphological characteristics of the complex in a circulation model. Methods: Release kinetics of VEGF165 complexed in different quantities in a collagen matrix were determined in a circulation model by quantifying protein concentration with ELISA over a period of 5 days. The structural changes of the collagen matrix were assessed with lig...
Birch HL, Worboys S, Eissa S, Jackson B, Strassburg S, Clegg PD.Tendon matrix integrity is vital to ensure adequate mechanical properties for efficient function. Although historically tendon was considered to be relatively inert, recent studies have shown that tendon matrix turnover is active. During normal physiological activities some tendons are subjected to stress and strains much closer to their failure properties than others. Tendons with low safety margins are those which function as energy stores such as the equine superficial digital flexor tendon (SDFT) and human Achilles tendon (AT). We postulate therefore that energy storing tendons suffer a hi...
Rode HJ, Janssen W, Rösen-Wolff A, Bugert JJ, Thein P, Becker Y, Darai G.A gene was identified within the DNA sequences of the EcoRI DNA fragment N (4.3 kbp) of the genome of equine herpesvirus type 2 (EHV-2) coding for a protein (179 amino acid residues) homologous to the cytokine synthesis inhibitory factor (CSIF; interleukin 10) of the human and mouse, and to the Epstein-Barr virus (EBV) protein BCRF1. This finding is further significant evidence that the interleukin 10 (IL-10) and/or IL-10-like gene can indeed be present in the genomes of members of the herpesviral family.
Zwart L, Potgieter CA, Clift SJ, van Staden V.African horse sickness is a serious equid disease caused by the orbivirus African horse sickness virus (AHSV). The virus has ten double-stranded RNA genome segments encoding seven structural and three non-structural proteins. Recently, an additional protein was predicted to be encoded by genome segment 9 (Seg-9), which also encodes VP6, of most orbiviruses. This has since been confirmed in bluetongue virus and Great Island virus, and the non-structural protein was named NS4. In this study, in silico analysis of AHSV Seg-9 sequences revealed the existence of two main types of AHSV NS4, designat...
Boehm AK, Seth M, Mayr KG, Fortier LA.Many metabolic processes in chondrocytes thought to contribute to age-related changes in the extracellular matrix are influenced by known roles of Hsp90. Age-related decreases in the level of Hsp90 have been documented in numerous cell types and could contribute to cartilage degeneration. The aim of this study was to investigate the roles of age and Hsp90 in insulin-like growth factor 1 (IGF-1) and interleukin-1beta (IL-1beta) signaling in chondrocytes. Methods: Levels of Hsp90 messenger RNA (mRNA) and protein, with respect to age, were determined by quantitative real-time polymerase chain rea...
