Mitochondria are organelles found in the cells of horses that are responsible for producing energy through the process of oxidative phosphorylation. They play a role in various cellular functions, including energy metabolism, regulation of the cell cycle, and apoptosis. Mitochondria contain their own DNA, which is maternally inherited and can provide insights into genetic lineage and population studies in equines. Research on equine mitochondria encompasses topics such as mitochondrial function, genetic variation, and their implications for health and performance. This page compiles peer-reviewed research studies and scholarly articles that explore the structure, function, and genetic aspects of mitochondria in horses.
Ortgies F, Klewitz J, Görgens A, Martinsson G, Sieme H.Reasons for low in vitro fertilisation rates in the horse include the difficulties in inducing capacitation and/or hyperactivation of stallion spermatozoa. The aim of this study was to analyse the effect of noncapacitating and capacitating modified Whitten's (MW) and modified Tyrode's medium (MT) and treatment with procaine (5 mmol), pentoxifylline (3.5 mmol) and trolox (120 mmol) on motility (CASA), capacitation, acrosomal status, viability and mitochondrial membrane potential of stallion spermatozoa (n = 4). While there was no influence of MW and MT on sperm motility, a significant increase ...
Votion DM, Fraipont A, Goachet AG, Robert C, van Erck E, Amory H, Ceusters J, de la Rebière de Pouyade G, Franck T, Mouithys-Mickalad A, Niesten A....Limited information exists about the muscle mitochondrial respiratory function changes that occur in horses during an endurance season. Objective: To determine effects of training and racing on muscle oxidative phosphorylation (OXPHOS) and electron transport system (ETS) capacities in horses with high resolution respirometry (HRR). Methods: Mitochondrial respiration was measured in microbiopsies taken from the triceps brachii (tb) and gluteus medius (gm) muscles in 8 endurance horses (7 purebred Arabians and 1 crossbred Arabian) before training (T0), after two 10 week training periods (T1, T2)...
Kawai M, Kuwano A, Hiraga A, Miyata H.The myonuclear domain (MND) is the region of cytoplasm governed by a single myonucleus. Myonuclear domain size is an important factor for muscle fibre plasticity because each myonucleus has limitations in the capacity of protein synthesis. Previous studies have demonstrated that differences in MND size exist in different fibre types in several species, including horses. Objective: To understand the basic mechanism of muscle plasticity, the relationships between MND size, muscle fibre type population and metabolic properties of skeletal muscles throughout the whole body in Thoroughbred horses w...
Franck T, Votion DM, Ceusters J, De La Rebière de Pouyade G, Mouithys-Mickalad A, Niesten A, Fraipont A, VAN Erck E, Goachet AG, Robert C, Serteyn D.Intense exercise in horses induces an increase of creatine kinase (CK) and stimulation of neutrophils which release the strong oxidant enzyme, myeloperoxidase (MPO) into the blood. It is not known whether active MPO is found in equine muscles and whether oxidant activity of neutrophils could affect muscular tissues and mitochondrial activity. Objective: Specific immuno-extraction followed by enzymatic detection (SIEFED) methods will be employed for the first time to assess both the normal range of MPO and mitochondrial complex I (MCI) activities in equine muscular microbiopsies and to study th...
da Silva CM, Macías-García B, Miró-Morán A, González-Fernández L, Morillo-Rodriguez A, Ortega-Ferrusola C, Gallardo-Bolaños JM, Stilwell G....Lipid peroxidation (LPO) has been claimed as a major factor involved in stallion damage during storage or cryopreservation. Because melatonin is a well-known potent antioxidant, the aim of the present study was to investigate the effect of melatonin during in vitro incubation. Furthermore, we investigated the presence of specific melatonin receptors (MT1 and MT2) using specific polyclonal antibodies and western blotting. Stallion spermatozoa were incubated up to 3 hr at 37°C in the presence of different concentrations of melatonin (0, 50 pm, 100 pm, 200 pm, or 1 μm). At the beginning and at ...
García BM, Moran AM, Fernández LG, Ferrusola CO, Rodriguez AM, Bolaños JM, da Silva CM, Martínez HR, Tapia JA, Peña FJ.Cryopreservation introduces extreme temperature and osmolality changes that impart lethal and sublethal effects on spermatozoa. Additionally, there is evidence that the osmotic stress induced by cryopreservation causes oxidative stress to spermatozoa. The main sources of reactive oxygen species in mammalian sperm are the mitochondria. In view of this, the aim of our study was to test whether or not osmotic stress was able to induce mitochondrial damage and to explore the osmotic tolerance of the mitochondria of stallion spermatozoa. Ejaculates from 7 stallions were subjected to osmolalities ra...
Tang X, Sayre LM, Tochtrop GP.Cytochrome c is a key mitochondrial respiratory protein that is particularly susceptible to modification during oxidative stress. The nature of this susceptibility is linked to the mitochondrial membrane being rich in esterified linoleic acid, which predisposes this organelle to the formation of lipid peroxidation products such as 4-hydroxy-2-(E)-nonenal (4-HNE). To better understand the nature of cytochrome c modification by 4-HNE, we initiated an in vitro study utilizing a combination of MALDI-TOF mass spectrometry, LC-ESI-MS/MS and isotope labeling to monitor 4-HNE modification of cytochrom...
Cieslak M, Pruvost M, Benecke N, Hofreiter M, Morales A, Reissmann M, Ludwig A.Domestic horses represent a genetic paradox: although they have the greatest number of maternal lineages (mtDNA) of all domestic species, their paternal lineages are extremely homogeneous on the Y-chromosome. In order to address their huge mtDNA variation and the origin and history of maternal lineages in domestic horses, we analyzed 1961 partial d-loop sequences from 207 ancient remains and 1754 modern horses. The sample set ranged from Alaska and North East Siberia to the Iberian Peninsula and from the Late Pleistocene to modern times. We found a panmictic Late Pleistocene horse population r...
Macías García B, González Fernández L, Ortega Ferrusola C, Morillo Rodríguez A, Gallardo Bolaños JM, Rodríguez Martinez H, Tapia JA....Fatty acids and plasmalogens were extracted from the phospholipids of the plasma membrane of stallion spermatozoa, to determine their relation with sperm quality after freezing and thawing. Sperm quality was rated using a quality index that combined the results of the analysis of sperm motility and velocity (CASA analysis), membrane status and mitochondrial membrane potential (flow cytometry) post thaw. Receiving operating system (ROC) curves were used to evaluate the value of specific lipid components of the sperm membrane herein studied as forecast of potential freezeability. From all parame...
Nergadze SG, Lupotto M, Pellanda P, Santagostino M, Vitelli V, Giulotto E.The insertion of mitochondrial DNA in the nuclear genome generates numts, nuclear sequences of mitochondrial origin. In the horse reference genome, we identified 82 numts and showed that the entire horse mitochondrial DNA is represented as numts without gross bias. Numts were inserted in the horse nuclear genome at random sites and were probably generated during the repair of DNA double-strand breaks. We then analysed 12 numt loci in 20 unrelated horses and found that null alleles, lacking the mitochondrial DNA insertion, were present at six of these loci. At some loci, the null allele is prev...
Bower MA, Campana MG, Whitten M, Edwards CJ, Jones H, Barrett E, Cassidy R, Nisbet RE, Hill EW, Howe CJ, Binns M.The paternal origins of Thoroughbred racehorses trace back to a handful of Middle Eastern stallions, imported to the British Isles during the seventeenth century. Yet, few details of the foundation mares were recorded, in many cases not even their names (several different maternal lineages trace back to 'A Royal Mare'). This has fuelled intense speculation over their origins. We examined mitochondrial DNA from 1929 horses to determine the origin of Thoroughbred foundation mares. There is no evidence to support exclusive Arab maternal origins as some historical records have suggested, or a sign...
Albrizio M, Lacalandra GM, Micera E, Guaricci AC, Nicassio M, Zarrilli A.Opioid receptors and endogenous opioid peptides act not only in the control of nociceptive pathways, indeed several reports demonstrate the effects of opiates on sperm cell motility and morphology suggesting the importance of these receptors in the modulation of reproduction in mammals. In this study we investigated the expression of delta opioid receptors on equine spermatozoa by western blot/indirect immunofluorescence and its relationship with sperm cell physiology. Methods: We analyzed viability, motility, capacitation, acrosome reaction and mitochondrial activity in the presence of naltri...
McGivney BA, McGettigan PA, Browne JA, Evans AC, Fonseca RG, Loftus BJ, Lohan A, MacHugh DE, Murphy BA, Katz LM, Hill EW.Digital gene expression profiling was used to characterize the assembly of genes expressed in equine skeletal muscle and to identify the subset of genes that were differentially expressed following a ten-month period of exercise training. The study cohort comprised seven Thoroughbred racehorses from a single training yard. Skeletal muscle biopsies were collected at rest from the gluteus medius at two time points: T(1) - untrained, (9 +/- 0.5 months old) and T(2) - trained (20 +/- 0.7 months old). Results: The most abundant mRNA transcripts in the muscle transcriptome were those involved in mus...
García BM, Fernández LG, Ferrusola CO, Salazar-Sandoval C, Rodríguez AM, Martinez HR, Tapia JA, Morcuende D, Peña FJ.Lipids were extracted from ejaculated spermatozoa from seven individual stallions to distinguish neutral lipids (NL) and polar lipids (PL) and determine their variation among stallions and their relationship with sperm quality and sperm susceptibility to lipid peroxidation. The isolated fatty acids were correlated with sperm quality (membrane integrity, mitochondrial membrane potential (ΔΨm) and expression of active caspases) and the sensitivity of the sperm plasma membrane to LPO. The miristic (C14: 0), palmitic (C16: 0), stearic (C18: 0) and oleic (C18: 1n9) acids were predominant among th...
Ortega Ferrusola C, González Fernández L, Salazar Sandoval C, Macías García B, Rodríguez Martínez H, Tapia JA, Peña FJ.In order to evaluate to what extent the changes that occur during cryopreservation involve the mitochondrial permeability transition pore (PT-pore), a specific inhibitor was used during the cryopreservation process of stallion spermatozoa. Four ejaculates from each of 7 stallions were frozen using a standard protocol. Before freezing, each ejaculate was split into three subsamples. The first was supplemented with 2.5 microM bongkrekic acid (BA) and the second with 5 microM BA. The third subsample served as control. The BA significantly reduced the percentage of spermatozoa depicting active cas...
Jiang Q, Wei Y, Huang Y, Jiang H, Guo Y, Lan G, Liao J.The Debao pony (Equus caballus) is the most important local variety of domestic horses, and is strictly protected by the Chinese government. Their average adult withers height is 94.42±3.76 cm for males and 98.35±4.55 cm for females, respectively. In the present study, the complete sequence of the Debao pony mitochondrial genome was determined (GenBank Accession No. EU939445), and was found to be similar to other equine mitochondrial genomes. However, there were 85 nucleotide substitutions in the 13 protein-coding genes; the percentage of substitution was 0.8±0.1. Polymorphisms of mtDNA con...
Ning T, Xiao H, Li J, Hua S, Zhang YP.Mitochondria play a crucial role in energy metabolism through oxidative phosphorylation. Organisms living at high altitudes are potentially influenced by oxygen deficits and cold temperatures. The severe environmental conditions can impact on metabolism and direct selection of mitochondrial DNA. As a wide-ranging animal, the domestic horse (Equus caballus) has developed various morphological and physiological characteristics for adapting to different altitudes. Thus, this is a good species for studying adaption to high altitudes at a molecular level. We sequenced the complete NADH dehydrogenas...
Gamboa S, Rodrigues AS, Henriques L, Batista C, Ramalho-Santos J.A group of stallions with different reproductive indexes were used to study seasonal variations in sperm quality (Equus caballus). Semen samples were collected from late September to July and analyzed according to four seasonal periods: late September-December, January-March, late March-May, and June-July. Parameters monitored included sperm concentration, sperm motility, sperm morphology, sperm viability, acrosomal status, plasma membrane stability, and sperm mitochondrial membrane potential. Overall, seminal parameters monitored are affected mostly by time period, followed by animal and last...
Orlando L, Metcalf JL, Alberdi MT, Telles-Antunes M, Bonjean D, Otte M, Martin F, Eisenmann V, Mashkour M, Morello F, Prado JL, Salas-Gismondi R....The rich fossil record of the family Equidae (Mammalia: Perissodactyla) over the past 55 MY has made it an icon for the patterns and processes of macroevolution. Despite this, many aspects of equid phylogenetic relationships and taxonomy remain unresolved. Recent genetic analyses of extinct equids have revealed unexpected evolutionary patterns and a need for major revisions at the generic, subgeneric, and species levels. To investigate this issue we examine 35 ancient equid specimens from four geographic regions (South America, Europe, Southwest Asia, and South Africa), of which 22 delivered 8...
Lira J, Linderholm A, Olaria C, Brandström Durling M, Gilbert MT, Ellegren H, Willerslev E, Lidén K, Arsuaga JL, Götherström A.Multiple geographical regions have been proposed for the domestication of Equus caballus. It has been suggested, based on zooarchaeological and genetic analyses that wild horses from the Iberian Peninsula were involved in the process, and the overrepresentation of mitochondrial D1 cluster in modern Iberian horses supports this suggestion. To test this hypothesis, we analysed mitochondrial DNA from 22 ancient Iberian horse remains belonging to the Neolithic, the Bronze Age and the Middle Ages, against previously published sequences. Only the medieval Iberian sequence appeared in the D1 group. N...
Meyer W, Kacza J, Schnapper A, Verspohl J, Hornickel I, Seeger J.Based on cryo-SEM, standard and high resolution TEM, glycoconjugate histochemistry, and microbiological differentiation, the present study demonstrates the colonisation of the epithelium of the equine oesophagus with microorganisms. As particularly apparent using cryo-SEM to illustrate natural conditions, the present microbiota were clearly dominated by bacteria, forming a one-layer system, as attached to and embedded in concentrated mannose/mannan substances covering the outer stratum corneal cells. Bacterial numbers ranged from 5600 to 7200 per mm(2) in the central part of the oesophagus, th...
Priskin K, Szabó K, Tömöry G, Bogácsi-Szabó E, Csányi B, Eördögh R, Downes CS, Raskó I.Movements of human populations leave their traces in the genetic makeup of the areas affected; the same applies to the horses that move with their owners This study is concerned with the mitochondrial control region genotypes of 31 archaeological horse remains, excavated from pre-conquest Avar and post-conquest Hungarian burial sites in the Carpathian Basin dating from the sixth to the tenth century. To investigate relationships to other ancient and recent breeds, modern Hucul and Akhal Teke samples were also collected, and mtDNA control region (CR) sequences from 76 breeds representing 921 in...
Lei CZ, Su R, Bower MA, Edwards CJ, Wang XB, Weining S, Liu L, Xie WM, Li F, Liu RY, Zhang YS, Zhang CM, Chen H.To obtain more knowledge of the origin and genetic diversity of domestic horses in China, this study provides a comprehensive analysis of mitochondrial DNA (mtDNA) D-loop sequence diversity from nine horse breeds in China in conjunction with ancient DNA data and evidence from archaeological and historical records. A 247-bp mitochondrial D-loop sequence from 182 modern samples revealed a total of 70 haplotypes with a high level of genetic diversity. Seven major mtDNA haplogroups (A-G) and 16 clusters were identified for the 182 Chinese modern horses. In the present study, nine 247-bp mitochondr...
Barrey E, Mucher E, Jeansoule N, Larcher T, Guigand L, Herszberg B, Chaffaux S, Guérin G, Mata X, Benech P, Canale M, Alibert O, Maltere P, Gidrol X.Several cases of myopathies have been observed in the horse Norman Cob breed. Muscle histology examinations revealed that some families suffer from a polysaccharide storage myopathy (PSSM). It is assumed that a gene expression signature related to PSSM should be observed at the transcriptional level because the glycogen storage disease could also be linked to other dysfunctions in gene regulation. Thus, the functional genomic approach could be conducted in order to provide new knowledge about the metabolic disorders related to PSSM. We propose exploring the PSSM muscle fiber metabolic disorder...
Waller AP, Geor RJ, Spriet LL, Heigenhauser GJ, Lindinger MI.Oral acetate supplementation enhances glycogen synthesis in some mammals. However, while acetate is a significant energy source for skeletal muscle at rest in horses, its effects on glycogen resynthesis are unknown. We hypothesized that administration of an oral sodium acetate-acetic acid solution with a typical grain and hay meal after glycogen-depleting exercise would result in a rapid appearance of acetate in blood with rapid uptake by skeletal muscle. It was further hypothesized that acetate taken up by muscle would be converted to acetyl CoA (and acetylcarnitine), which would be metaboliz...
Hendricks KE, Hansen PJ.Apoptosis is common during spermatogenesis. Here, it was tested whether apoptosis could be induced in sperm after ejaculation. There were several lines of evidence to indicate that sperm are resistant to induction of apoptosis. First, incubation of bull sperm at temperatures characteristic of normothermia (38.5 degrees C) or heat shock (40 and 41 degrees C) for 4h did not increase the proportion of sperm positive for the TUNEL reaction. There was also no reduction in mitochondrial polarity caused by exposure to 40 or 41 degrees C. Incubation at 38.5 degrees C (least-squares mean+/-SEM=4.0+/-1....
Ambruosi B, Lacalandra GM, Iorga AI, De Santis T, Mugnier S, Matarrese R, Goudet G, Dell'aquila ME.Lipid droplets (LDs) and mitochondria in the ooplasm are essential for energy production required for maturation, fertilization and embryo development. This study investigates the correlations between cytoplasmic LDs polar aggregation and: (1) nuclear maturation (Experiment 1); (2) mitochondrial (mt) distribution pattern and localization (Experiment 2); (3) fertilization and embryonic development after intracytoplasmic sperm injection (ICSI; Experiment 3) in equine oocytes recovered from slaughtered mares and matured in vitro. Morphologically normal oocytes were selected after culture and cate...
Mari G, Rizzato G, Merlo B, Iacono E, Bucci D, Seren E, Tamanini C, Galeati G, Spinaci M.Little information is available on the quality of stallion spermatozoa after sex sorting. The objectives of the present study were to assess the quality of sex-sorted stallion spermatozoa and determine its fertilizing ability after hysteroscopic low dose insemination. Ejaculates from four stallions were collected and sorted by a MoFlo SX flow cytometer/sperm sorter. Before and after sorting, spermatozoa were evaluated for motility by Computer Assisted Sperm Analysis, viability (SYBR 14-propidium iodide), mitochondrial function (JC-1) and acrosomal status (fluorescein isothiocyanate Pisum sativ...
Lavoie JP, Thompson D, Hamilton E, Debrue M, David F, Hickey G.It has been suggested that many of the beneficial effects of corticosteroids are mediated through mitogen-activated protein kinase (MAPK) p38 inhibition. Objective: To investigate the efficacy of the MAPK p38 inhibitor compound MRL-EQ1 to either prevent (Phase 1) or treat (Phase 2) recurrent airway obstruction (RAO) in horses. Methods: MRL-EQ1 was administered i.v. at a dosage of 0.75-1.5 mg/kg bwt q. 12 h. In Phase 1, susceptible horses in clinical remission were divided into 2 groups (n = 5/group), based on historical values of respiratory mechanics. All horses were entered in the study in p...
Wilson JM, Caceci T, Potter GD, Kraemer DC.Embryos were recovered non-surgically at about Day 6 after ovulation from 15 Quarter horse-type mares and were evaluated for morphological changes which may occur because of exposure to the cryoprotectant and/or cryopreservation. Electron microscopy was used to elucidate the fine structure of intracellular organelles which, if damaged, could cause cellular death. The horse embryo does not totally re-expand in the 10% glycerol freezing medium, nor will it completely re-expand in the isotonic holding medium following glycerol removal whether or not the embryo has been frozen. Embryos in this stu...
Hedayat Evrigh N, Omri M, Boustan A, Seyedsharifi R, Vahedi V.The source and history of horse domestication have been studied for decades in biology and archeology. The aim of this study was to evaluate the genetic structure and the genetic relationship between six breeds of native horses using mitochondrial D-loop region. Blood samples were collected from 96 horses. Total DNA was extracted, and 430 bp of D-Loop region (hyper variable) was amplified and sequenced using Sanger sequencing methods. The analysis of data led to identify 48 polymorphic sites that create 52 haplotypes. The plotted phylogenic tree for haplotypes of Iranian native horses is plac...
Elsner J, Hofreiter M, Schibler J, Schlumbaum A.On large geographical scales, changes in animal population distribution and abundance are driven by environmental change due to climatic and anthropogenic processes. However, so far, little is known about population dynamics on a regional scale. We have investigated 92 archaeological horse remains from nine sites mainly adjacent to the Swiss Jura Mountains dating from c. 41,000-5,000 years BP. The time frame includes major environmental turning points such as the Last Glacial Maximum (LGM), followed by steppe vegetation, afforestation and initial re-opening of the landscape by human agricultur...
Meyer W, Kacza J, Hornickel IN, Schoennagel B.Using immunohistochemistry and transmission electron microscopy (TEM), the esophagus epithelia of seven domesticated mammals (horse, cattle, goat, pig, dog, laboratory rat, cat) of three nutrition groups (herbivorous, omnivorous, carnivorous) were studied to get first information about energy generation, as demonstrated by succinate dehydrogenase (SDH) activities. Distinct reaction intensities could be observed in all esophageal cell layers of the different species studied reflecting moderate to strong metabolic activities. The generally strong staining in the stratum basale indicated that new...
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...
Nikbakhsh M, Varkoohi S, Seyedabadi HR.Kurdish horse is one of the most valuable horse genetic resources in the Middle East. To assess the genetic diversity of Kurdish horses, Mitochondrial DNA D-loop hyper-variable region1 (HVR1) was sequenced in 29 non-related Kurdish horses which were sampled from diverse geographic regions of Iran. Total DNA was extracted from the collected blood samples by modified salting out method. The HVR1 was amplified by PCR and then sequenced using ABI PRISM BigDyeTM Terminator Cycle Sequencing Ready Reaction Kit. Consequently, the sequences were trimmed to 294 bp using BIOEDIT to become comparable wit...
Albornoz A, Morales N, Uberti B, Henriquez C, Burgos RA, Alarcon P, Moran G.Neutrophils participate in innate immunity as the first line of host defence against microorganisms. However, persistent neutrophil activity and delayed apoptosis can be harmful to surrounding tissues; this problem occurs in diverse inflammatory diseases, including asthma-affected horses. Previous studies in horses with acute lung inflammation indicated that treatment with tamoxifen (TX), a selective oestrogen receptor modulator, produces a significant decrease in bronchoalveolar lavage fluid (BALF) neutrophil content. The aim of this study was to investigate the effect of tamoxifen and its me...
de Vasconcelos Franco JS, Faheem M, Chaveiro A, Moreira da Silva F.The effects of supplementation of α-tocopherol and different freezing rates (FRs) on the ability of stallion sperm to fertilize bovine oocytes with intact zona pellucida were investigated, in an attempt to develop a model to assess cryopreserved sperm function. Semen was obtained from four purebred Lusitano stallions (n = 4). Each ejaculate was subjected to cryopreservation with a commercial extender (Ghent, Minitub Iberia, Spain), without any supplementation (control) or supplemented with 2-mM α-tocopherol. The semen was exposed to two different FRs between 5 °C and -15 °C: slow (5 °C...
Kruse CJ, Stern D, Mouithys-Mickalad A, Niesten A, Art T, Lemieux H, Votion DM.Equine atypical myopathy is a seasonal intoxication of grazing equids. In Europe, this poisoning is associated with the ingestion of toxins contained in the seeds and seedlings of the sycamore maple (Acer pseudoplatanus). The toxins involved in atypical myopathy are known to inhibit ß-oxidation of fatty acids and induce a general decrease in mitochondrial respiration, as determined by high-resolution respirometry applied to muscle samples taken from cases of atypical myopathy. The severe impairment of mitochondrial bioenergetics induced by the toxins may explain the high rate of mortality obs...
Johannisson A, Figueiredo MI, Al-Kass Z, Morrell JM.An improved fertility prediction for stallions is of importance for equine breeding. Here, we investigate the potential of a combined staining of stallion spermatozoa for superoxide and mitochondrial membrane potential (MMP) for this purpose. Semen samples were analysed immediately after arrival at the laboratory, as well as after 24 h. Superoxide was measured by MitoSOXRed, while MMP was measured with JC-1. Menadione was used to stimulate superoxide production. In addition, other parameters of sperm quality, namely motility, membrane integrity, chromatin integrity, sperm kinematics and Hoec...
Nürnberger S, Rentenberger C, Thiel K, Schädl B, Grunwald I, Ponomarev I, Marlovits S, Meyer C, Barnewitz D.The present study reports for the first time the presence of giant crystals in mitochondria of equine chondrocytes. These structures show dark contrast in TEM images as well as a granular substructure of regularly aligned 1-2 nm small units. Different zone axes of the crystalline structure were analysed by means of Fourier transformation of lattice-resolution TEM images proving the crystalline nature of the structure. Elemental analysis reveals a high content of nitrogen referring to protein. The outer shape of the crystals is geometrical with an up to hexagonal profile in cross sections. It ...
Cassart D, Fett T, Sarlet M, Baise E, Coignoul F, Desmecht D.The morphopathological picture of a subset of equine myopathies is compatible with a primary mitochondrial disease, but functional confirmation in vivo is still pending. The cationic dye JC-1 exhibits potential-dependent accumulation in mitochondria that is detectable by a fluorescence shift from green to orange. As a consequence, mitochondrial membrane potential can be optically measured by the orange/green fluorescence intensity ratio. A flow cytometric standardized analytic procedure of the mitochondrial function of equine peripheral blood mononuclear cells is proposed along with a critical...
Nam GH, Ahn K, Bae JH, Cho BW, Park KD, Lee HK, Yang YM, Kim TH, Seong HH, Han K, Kim HS.In the mitochondrial matrix, the OXCT1 gene catalyzes the reversible transfer of coenzyme A from succinyl-CoA to acetoacetate in a reaction related to energy production from ketone bodies. Here, horse OXCT1 gene containing coenzyme A transferase domain was identified in the transcriptome analysis of cDNAs derived from skeletal muscles. Horse OXCT1 gene consisted of 1761 [corrected] nucleotide sequences with an open reading frame of 1560 nucleotides encoding a protein of 520 putative amino acid residues.The number of non-synonymous substitutions was lower than the number of synonymous substitut...
Pérez-Gutiérrez LM, De la Peña A, Arana P.Hispano-Breton (HB) is a horse breed with a recent mixed ancestry. It was developed in the 1930s by crossing local mares with Breton draught horses imported from France. Nowadays it is considered to be in a vulnerable situation due to census decline. To genetically characterize the breed and to set up the basis for a conservation programme, we have employed two types of molecular markers: a 347-bp D-loop mitochondrial DNA (mtDNA) fragment and 13 microsatellite loci. A representative sample of 53 HB individuals was analysed together with a sample of 40 Pura Raza Española horses for comparison....
Giaretta E, Mislei B, Martínez-Pastor F, Nesci S, Spinaci M, Galeati G, Nerozzi C, Mari G, Tamanini C, Bucci D.Equine spermatozoa highly rely on oxidative phosphorylation for their energy management. The present work aimed to characterize the role of mitochondria on horse sperm motility and ROS production by incubating spermatozoa with specific inhibitors of the different mitochondrial complexes. Equine spermatozoa were incubated 1 h and 3 h at 37 °C with: complex I inhibitor rotenone (5 μM, ROT), complex II inhibitor dimethyl-malonate (10 mM, DMM), complex III inhibitor antimycin A (1.8 μM, ANTI), the uncoupling agent carbonyl cyanide m-chlorophenyl hydrazine (5 μM, CCCP), ATP synthase inhi...
Albrizio M, Lacalandra GM, Volpe S, Nicassio M, Cinone M.Heat Shock Proteins are chaperones primary involved in the repair of cellular damages induced by temperature. The harmful effect of temperature on the male gonad is well known, on the contrary knowledge on the effects of the environment on semen quality are still insufficient. The aim of this paper was to learn more about the role of HSPs and the environment in modulating the physiology of equine male gonads. We showed a detailed analysis of equine semen characteristic and the expression level of three HSPs (60-70-90) over a one-year period analyzing the effects of temperature and humidity and...
Milner PI, Smith HC, Robinson R, Wilkins RJ, Gibson JS.Hypoxia and acidosis are recognized features of inflammatory arthroses. This study describes the effects of IGF-1 and TGF-β(1) on pH regulatory mechanisms in articular cartilage under hypoxic conditions. Acid efflux, reactive oxygen species (ROS), and mitochondrial membrane potential were measured in equine articular chondrocytes isolated in the presence of serum (10% fetal calf serum), IGF-1 (1, 10, 50, 100 ng/ml) or TGF-β(1) (0.1, 1, 10 ng/ml) and then exposed to a short-term (3 h) hypoxic insult (1% O(2)). Serum and 100 ng/ml IGF-1 but not TGF-β(1) attenuated hypoxic regulation of pH hom...
Di Bernardo G, Del Gaudio S, Galderisi U, Cipollaro M.Ancient DNA extracted from 2000 year-old equine bones was examined in order to amplify mitochondrial and nuclear DNA fragments. A specific equine satellite-type sequence representing 3.7%-11% of the entire equine genome, proved to be a suitable target to address the question of the presence of aDNA in ancient bones. The PCR strategy designed to investigate this specific target also allowed us to calculate the molecular weight of amplifiable DNA fragments. Sequencing of a 370 bp DNA fragment of mitochondrial control region allowed the comparison of ancient DNA sequences with those of modern hor...
Cacciamali A, Pascucci L, Villa R, Dotti S.Nanoscience and nanotechnologies have recently gained importance in several fields, such as industry and medicine. A big issue of the increasing application of nanomaterials is the poor literature regarding their potential toxicity in humans and animals. Recently, adult stem cells have been proposed as putative targets of nanoparticles (NPs). This study aims to investigate the effects of zerovalent-metallic NPs on isolated and amplified equine Adipose tissue derived Mesenchymal Stem Cells (eAdMSCs). Cells were treated with Cobalt (Co-), Iron (Fe-), and Nickel (Ni-) nanoparticles (NPs) at diffe...
Di Bernardo G, Galderisi U, Del Gaudio S, D'Aniello A, Lanave C, De Robertis MT, Cascino A, Cipollaro M.DNA extracted from the skeletons of five equids discovered in a Pompeii stable and of a horse found in Herculaneum was investigated. Amino acid racemization level was consistent with the presence of DNA. Post-mortem base modifications were excluded by sequencing a 146 bp fragment of the 16S rRNA mitochondrial gene. Sequencing of a 370 bp fragment of mitochondrial (mt)DNA control region allowed the construction of a phylogenetic tree that, along with sequencing of nuclear genes (epsilon globin, gamma interferon, and p53) fragments, gave us the possibility to address some questions puzzling arch...
Aitken RJ, Gregoratos D, Kutzera L, Towney E, Lin M, Wilkins A, Gibb Z.MTT is widely used in biology as a probe for cell viability by virtue of its ability to generate deposits of insoluble formazan at sites of intense oxidoreductase activity. This response is generally held to reflect mitochondrial redox activity; however, extra-mitochondrial MTT reduction has also been recorded in certain cell types. Given this background, we set out to determine the major sites of formazan deposition in mammalian spermatozoa. In the mouse, most MTT reduction took place within the extensive mitochondrial gyres, with a single minor site of formazan deposition on the sperm head. ...
Sanny CG, Weiner H.The inhibition of mitochondrial (pI 5) horse liver aldehyde dehydrogenase by disulfiram (tetraethylthiuram disulphide) was investigated to determine if the drug was an active-site-directed inhibitor. Stoichiometry of inhibition was determined by using an analogue, [35S]tetramethylthiuram disulphide. A 50% loss of the dehydrogenase activity was observed when only one site per tetrameric enzyme was modified, and complete inactivation was not obtained even after seven sites per tetramer were modified. Modification of only two sites accounted for a loss of 75% of the initial catalytic activity. Th...
Valberg SJ, Iglewski H, Henry ML, Schultz AE, McKenzie EC.Selective breeding and discipline specific training has led to equine breeds adept at various athletic disciplines. Breed-specific skeletal muscle adaptations have been studied in many breeds but not Warmbloods (WB). We evaluated gluteal muscle contractile muscle fiber types and citrate synthase activity (CS), a marker for mitochondrial volume density, in WB trained for dressage (second level-Grand Prix) contrasted with Quarter Horses (QH). Gluteus medius muscle biopsies from 14 unfit/18 fit dressage-trained WB and 20 unfit/16 fit reining/working cow QH were analyzed fluorometrically and fiber...
Mollenhauer HH, Rowe LD, Witzel DA.Heart, diaphragm, and rear limb muscle from ponies and rats treated with monensin were examined by light and electron microscopy. In both species, mitochondrial aberrations were observed in selected muscle cells. The aberrations consisted of loss of matrix substance (vacuolization) usually accompanied by swelling. Vacuolated mitochondria were characterized by an almost total loss of matrix substance with retention of the rudimentary cristae form. In ponies, most vacuolated mitochondria were observed in heart muscle though some also were found in the diaphragm. In rats, most vacuolated mitochon...
Meng Q, Li S, Liu L, Xu J, Liu Y, Zhang Y, Zhang X, Shao Y.Equine Infectious Anemia Virus (EIAV), like other lentiviruses, has a transmembrane glycoprotein with an unusually long cytoplasmic tail (CT). Viral envelope (Env) proteins having CT truncations just downstream the putative membrane-spanning domain (PMSD) are assumed to exist among all wild-type budded virions, and also in some cell-adapted strains. To determine whether CT-truncated Env proteins can cause particularly deleterious effects on the Env expressing cells and/or their neighboring cells, plasmids encoding codon-optimized env gene including full-length (pE863) or CT-truncated (pE686* a...
Saad B, Corradin G, Bosshard HR.The antigenic determinants for two monoclonal antibodies directed against horse apo-cytochrome c, a protein of disordered structure, as judged by spectroscopic and hydrodynamic criteria, have been studied by a combination of methods: antigen competition in solution by radio immunoassay and enzyme-linked immunoassay, and differential acetylation of free and antibody-bound antigen. In the latter method the accessibility of lysine residues of the antigen in the antigen-antibody complex is compared to the accessibility in the free antigen. The two antibodies against the heme-free protein do not re...
Peña FJ, Martín-Cano FE, Becerro-Rey L, da Silva-Álvarez E, Gaitskell-Phillips G, Ortega-Ferrusola C, Aparicio IM, Gil MC.Although stallion spermatozoa are now recognized as highly dependent on oxidative phosphorylation for ATP production in the mitochondria, most extenders in use contain supraphysiological concentrations of glucose as the main energy source. While the toxicity of cryoprotectants has been well documented in the literature, the potential toxicity of excessive glucose in extenders is largely ignored. However, the toxicity of excess glucose, known as "carbotoxicity", is well-established in many areas of medicine. In this paper, we review the basic aspects of stallion spermatozoa metabolism, focusing...
Yordanov G, Palova N, Mehandjyiski I, Hristov P.From a historical perspective, horse breeding in Bulgaria has been very well developed since the time of the Thracians (early Bronze Age c. 3000 BCE). Archaeological discoveries from this era present us with an extremely rich type diversity, including wild and local primitive horses, the prototype of heavy draft horses, and fine riding horses.The objective of this study was to investigate the genetic structure of unexamined populations of three closely related horse breeds - the Danubian Nonius Hungarian Nonius and Serbian Nonius horses. A 608?bp long fragment of the mtDNA D-loop region was am...
Ishida N, Hasegawa T, Oyunsuren T, Mukoyama H.The mitochondrial DNA sequence of cytochrome b gene in a Thoroughbred horse was determined. By comparing DNA sequences between the Thoroughbred and published sequence data (two horses and one Grevyi zebra), polymerase chain reaction (PCR) primers were designed for amplification of a 590 bp DNA fragment in the cytochrome b gene, and PCR-restriction fragment length polymorphism (RFLP) analysis was studied in 140 horses of six breeds using three restriction enzymes (AciI, BamHI, RsaI). Two morphs were found using each of the three enzymes. By combining three enzymes morphs, the 140 horses examine...
Reynolds S, Hedberg M, Herrin B, Jesudoss Chelladurai JRJ.Here, we report two complete and three partial mitochondrial genome sequences of Dermacentor variabilis specimens collected from horses in the United States. The complete genomes are 14,837 bp long and contain 13 protein-coding genes, 2 rRNA genes, and 22 tRNA genes. The sequences have been deposited under GenBank accession numbers ON052120 to ON052124.
