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See detailHigh concentrations of Myeloperoxidase in the equine uterus as an indicator of endometritis
Parrilla Hernandez, Sonia ULg; Ponthier, Jérôme ULg; Franck, Thierry ULg et al

in Theriogenology (in press)

Intra-luminal fluid and excessive abnormal hyper-edema are regularly used for the diagnosis of endometritis in the mare, which is routinely confirmed by the presence of neutrophils on endometrial smears ... [more ▼]

Intra-luminal fluid and excessive abnormal hyper-edema are regularly used for the diagnosis of endometritis in the mare, which is routinely confirmed by the presence of neutrophils on endometrial smears. Studies show a relation between neutrophils and myeloperoxidase (MPO), an enzyme contained in and released by neutrophils during degranulation or after cell lysis. This enzyme has been found in many fluids and tissues and associated with different inflammatory pathologies in the horse. The aims of this study were to assess the presence and concentration of MPO in the equine uterus, and to investigate its relation with neutrophils, and other clinical signs of endometritis. Mares (n=51) were evaluated for presence of intra-luminal fluid and excessive endometrial edema before breeding, and a small volume lavage and cytology samples were obtained. From 69 cycles, supernatant of the uterine flushes was analysed with a specific equine MPO ELISA assay to measure MPO concentration. Cytology samples were used for the diagnosis of endometritis. MPO was present in the uterus of all estrus mares in highly variable concentrations. MPO concentrations were significantly (p<0.05) higher in samples with positive cytologies and in presence of intra-luminal fluid. Occasionally, some samples with negative cytologies showed high MPO concentration, but the opposite was never observed. Cycles presenting hyper-edema weren’t associated to high concentration of MPO, intra-luminal fluid or positive cytology making it a poor diagnostic tool of endometritis. [less ▲]

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See detailConcentration, Activity and Biochemical Characterization of Myeloperoxidase in Fresh and Post-thaw equine semen and their Implication on Freezability
Ponthier, Jérôme ULg; Franck, Thierry ULg; Parrilla Hernandez, Sonia ULg et al

in Reproduction in Domestic Animals (in press)

Myeloperoxidase (MPO) is a pro-oxidant enzyme associated with decreased motility in thawed equine semen. This study aimed to describe MPO concentration, activity and subunits in raw and thawed semen and ... [more ▼]

Myeloperoxidase (MPO) is a pro-oxidant enzyme associated with decreased motility in thawed equine semen. This study aimed to describe MPO concentration, activity and subunits in raw and thawed semen and to correlate these data with motilities in raw and thawed semen. Semen samples from five stallions were collected four times. Motilities were assessed in raw and thawed semen. MPO assays were performed in raw seminal plasma, raw sperm-rich pellet and thawed semen. Total and active MPO concentrations were, respectively, assayed by enzyme-linked immunosorbent assay and specific immunological extraction followed by enzymatic detection. MPO subunits present in semen were characterized by Western 3 blot. Purified active MPO was added in PBS and freezing extender to control its activity during freezing procedure. Differences between medians were determined using Kruskal– Wallis test, and correlations were determined using Spearman’s test for nonparametric data. Active MPO concentration was low in seminal plasma and thawed semen, but high in pellet (p = 0.0058), as the opposite relation was observed for total MPO concentration (p < 0.0001). In seminal plasma and post- thaw semen, inactive 86-kDa MPO precursor was mainly observed. Purified MPO activity was decreased in the extender (p = 0.0286). MPO activity in pellet was highly correlated with thawed progressive motility (r = 􏰑0.5576, p = 0.0086). Inac- tive MPO precursor and unknown low molecular weight inactive MPO precursor subunits explain low MPO activity in semen. Major MPO activity was observed in pellet, and post- thaw loss of activity is partially explained by MPO inactiva- tion in extender. Thawed semen motility was negatively correlated with MPO activity in pellet, becoming a potential freezability predictor. [less ▲]

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See detailÉtude des contraintes mécaniques locomotrices chez le cheval
Noble, Prisca ULg; Collin, Bernard ULg; Denoix, Jean-Marie et al

in Annales de Médecine Vétérinaire (in press)

In locomotor biomechanics, three high groups of contraints are commonly encountared : pressure, traction and torsion. In supra-maximal conditions, all of these contraints would be responsible of some ... [more ▼]

In locomotor biomechanics, three high groups of contraints are commonly encountared : pressure, traction and torsion. In supra-maximal conditions, all of these contraints would be responsible of some equine diseased locomotor systems. In order to understand better the contraints in the equine locomotor dynamics, some investigations have been carried out. Moreover, some measurement methods, based on the mechanics of Newton, have been performed. This review shows the different measurement techniques and introduces the mechanical basis that are compulsory for the understanding of the equine locomotor apparatus functioning. [less ▲]

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See detailEffect of inhaled hysrosoluble curcumin in lipopolysaccharide-induced lung neutrophilia in horses
Sandersen, Charlotte ULg; Cerri, Simona ULg; Franck, Thierry ULg et al

in Dorothy Russel Havemeyer Foundation IAD Workshop (2014)

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See detailEnhanced or Reduced Fetal Growth Induced by Embryo Transfer into Smaller or Larger Breeds Alters Post-Natal Growth and Metabolism in Pre-Weaning Horses
Peugnet, Pauline; Wimel, Laurence; Duchamp, Guy et al

in PLoS ONE (2014), 9(7), 102044

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See detailMyeloperoxidase and its products in synovial fluid of patients with treated or untreated rheumatoid arthritis.
Nzeusseu Toukap, A.; Delporte, C.; Noyon, C. et al

in Free radical research (2014), 48(4), 461-5

Abstract Objective. Plasma and synovial myeloperoxidase (MPO) and its products were strongly associated with osteoarthritis (OA) and rheumatoid arthritis (RA). In addition, it is well known that there is ... [more ▼]

Abstract Objective. Plasma and synovial myeloperoxidase (MPO) and its products were strongly associated with osteoarthritis (OA) and rheumatoid arthritis (RA). In addition, it is well known that there is a link between oxidative stress and cytokines. The present study aims at investigating the link between synovial MPO (and its products), interleukin (IL)-18, which is involved in the degradation of articular cartilage in RA, and IL-8, which is involved in recruitment and activation of neutrophils during inflammation. Effects of the treatment of RA on the biological parameters were also investigated. Methods. Patients (n = 105) were studied including 39 patients with OA, 33 with RA and 33 with RA receiving a specific treatment. Disease activity score (DAS-28) was calculated whereas MPO antigen/activity, neutrophils, chloro-tyrosine (Cl-Tyr), homocitrulline (Hcit), IL-8, and IL-18 were measured in synovial fluid (SF) and CRP was measured in serum. Results. DAS-28 and CRP levels were not significantly different between groups. MPO activity, and MPO, Cl-Tyr, and Hcit levels were significantly higher in SF of RA patients than OA patients. MPO specific activity (MPO activity/antigen ratio) was significantly lower in treated than in untreated RA patients as was IL-8. MPO activity and concentration were correlated with IL-8 and IL-18 in untreated but not in treated RA patients. Conclusions. MPO level is related to IL-8 and IL-18 levels in untreated RA patients. A link has been shown between treatment and decrease of IL-8, MPO specific activity and Hcit in SF. The causal role of MPO in SF inflammation and how treatment can affect MPO specific activity need further investigations. [less ▲]

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See detailEffect of different kinds of anoxia/reoxygenation on the mitochondrial function and the free radicals production of cultured primary equine skeletal myoblasts.
Ceusters, Justine ULg; Mouithys-Mickalad, Ange ULg; Franck, Thierry ULg et al

in Research in Veterinary Science (2013), 95

Horses are outstanding athletes, performing in many different disciplines involving different kinds of efforts and metabolic responses. Depending on exercise intensity, their skeletal muscle oxygenation ... [more ▼]

Horses are outstanding athletes, performing in many different disciplines involving different kinds of efforts and metabolic responses. Depending on exercise intensity, their skeletal muscle oxygenation decreases, and the reperfusion at cessation of the exercise can cause excessive production of free radicals. This study on cultured primary equine myoblasts investigated the effect of different kinds of anoxia/reoxygenation (A/R) on routine respiration, mitochondrial complex I specific activity and free radicals production. Our data revealed that short cycles of A/R caused a decrease of all the parameters, opposite to what a single long period of anoxia did. A preconditioning-like effect could explain our first pattern of results whereas mild uncoupling could be more appropriate for the second one. Anyway, it seems that mitochondrial complex I could play a major role in the regulation of the balance between metabolic and antioxidant protection of the muscular function of athletic horses. [less ▲]

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See detailEffect of non-sperm cells removal with single layer colloidal centrifugation on myeloperoxidase concentration in post-thaw equine semen
Ponthier, Jérôme ULg; Teague, Sheila; Franck, Thierry ULg et al

in Theriogenology (2013), 80(9), 1082-1087

Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in, and released by, neutrophils during degranulation or after lysis. Post-thaw semen contains MPO and its concentration is associated with ... [more ▼]

Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in, and released by, neutrophils during degranulation or after lysis. Post-thaw semen contains MPO and its concentration is associated with decreased sperm motility. Recently, MPO concentration in post-thaw semen was shown to be associated with presence of non-sperm cells. The aim of this study was to describe the effect of centrifugal fractionation of semen prior to cryopreservation on post-thaw concentrations of non-sperm cells and MPO. The experimental design consisted in freezing semen with or without previous centrifugation through two concentrations of single layer colloid media. Non-sperm cells and MPO concentrations were assessed in pellet and upper layer at each step of the procedure and MPO was detected in cells by immunocytochemistry. Single layer colloid centrifugation decreased non-sperm cells and MPO concentrations in post-thaw semen. The MPO concentration was correlated with concentration of non-sperm cells in the upper layer of the supernatant. In post-thaw semen, with or without previous single layer colloid centrifugation, MPO concentration was correlated with concentration of non-sperm cells. Overall, neutrophils were rarely observed and non-sperm cells were mainly epithelial cells or cellular debris, as demonstrated by MPO immunocytochemistry. Following single layer colloid centrifugation, MPO concentration was decreased, and correlations observed in different samples of the experiments revealed an association between concentrations of MPO and non-sperm cells. At all steps of the semen processing and cryopreservation, MPO immunostaining was clearly identified only on non-sperm cells. Our results demonstrate that non-sperm cells present in fresh semen release MPO during freezing. [less ▲]

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See detailMyeloperoxidase-Dependent LDL Modifications in Bloodstream Are Mainly Predicted by Angiotensin II, Adiponectin, and Myeloperoxidase Activity: A Cross-Sectional Study in Men
Zouaoui Boudjeltia, Karim; Delporte, Cédric; Van Antwerpen, Pierre et al

in Mediators of Inflammation (2013), 2013

The present paradigm of atherogenesis proposes that low density lipoproteins (LDLs) are trapped in subendothelial space of the vascular wall where they are oxidized. Previously, we showed that oxidation ... [more ▼]

The present paradigm of atherogenesis proposes that low density lipoproteins (LDLs) are trapped in subendothelial space of the vascular wall where they are oxidized. Previously, we showed that oxidation is not restricted to the subendothelial location. Myeloperoxidase (MPO), an enzyme secreted by neutrophils and macrophages, can modify LDL (Mox-LDL) at the surface of endothelial cells. In addition we observed that the activation of the endothelial cells by angiotensin II amplifies this process. We suggested that induction of the NADPH oxidase complex was a major step in the oxidative process. Based on these data, we asked whether there was an independent association, in 121 patients, between NADPH oxidase modulators, such as angiotensin II, adiponectin, and levels of circulating Mox-LDL. Our observations suggest that the combination of blood angiotensin II, MPO activity, and adiponectin explains, at least partially, serum Mox-LDL levels. [less ▲]

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See detailAN IMMUNOLOGICAL METHOD TO COMBINE THE MEASUREMENT OF ACTIVE AND TOTAL HUMAN MYELOPEROXIDASE ON THE SAME SAMPLE FROM A COMPLEX MEDIUM
Franck, Thierry ULg; MINGUET, Grégory ULg; Mossay, Pierre et al

Conference (2013, September 12)

Active neutrophil myeloperoxidase (MPO) is a powerful producer of oxidant molecules in acute or chronic inflammation, and it is essential to measure its activity in biological samples. We combined two ... [more ▼]

Active neutrophil myeloperoxidase (MPO) is a powerful producer of oxidant molecules in acute or chronic inflammation, and it is essential to measure its activity in biological samples. We combined two immunological techniques, the SIEFED (specific immunologic extraction followed by enzymatic detection) and an ELISA, to measure the active and total contents of human MPO on the same sample and with the same calibration curve, to define an accurate ratio between the active and total enzyme. After the extraction of MPO from aqueous or biological samples by immobilized anti-MPO antibodies followed by a washing to eliminate unbound material, the active and the total contents of the enzyme were sequentially measured without interferences (patent: EP2017351-B1, 2010). Compared to a classical sandwich ELISA, there is one additional step corresponding to the in situ measurement of MPO activity, but this step does not affect the following measurement of the total MPO content. After validation, the combined technique was applied to a whole blood model of in vitro stimulation with phorbol-myristate-acetate (PMA), cytochalasin B/N-formyl-methionyl-leucyl-phenyl-alanine (CB/fMLP) or lipopolysaccharide/ tumor necrosis factor-alpha (LPS/TNF-α) (n=9). The active/total MPO ratio in whole blood reached 0.267±0.126 for non-stimulated condition and was significantly (p<0.05) higher for PMA (0.360±0.106), CB/fMLP (0.380±0.113) and LPS/TNF-α (0.432±0.124) stimulated conditions. These different ratios highlight the real oxidant potential of MPO, which depends on the stimulating conditions, witness of what could happen in pathological situations with diagnostic purpose. The combined SIEFED/ELISA method also appeared as a powerful tool to screen potential inhibitors that could interact directly with the enzyme, either on its active site or on another key position. [less ▲]

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See detailA water-soluble salt of curcumin (NDS27) inhibits myeloperoxidase and NADPH oxidase activities, two major enzymes of neutrophils.
Derochette, Sandrine ULg; Mouithys-Mickalad, Ange ULg; Deby-Dupont, Ginette et al

Poster (2013, September 11)

Neutrophils (PMNs) produce reactive oxygen species (ROS) to kill pathogenic agents. After appropriate stimulation, leading to the activation of protein kinase C (PKC), the cytosolic subunits of the NADPH ... [more ▼]

Neutrophils (PMNs) produce reactive oxygen species (ROS) to kill pathogenic agents. After appropriate stimulation, leading to the activation of protein kinase C (PKC), the cytosolic subunits of the NADPH oxidase (Nox2) are phosphorylated and translocated to the membrane flavocytochrome b558, forming the active enzyme which produces superoxide anion (O2●-). From O2●- derives H2O2 used by the PMNs myeloperoxidase (MPO) to form strong oxidant species. Many human and animal pathologies with fatal issue are associated with uncontrolled activation of PMNs. The modulation of enzymes implied in ROS production is thus a primary target to manage excessive inflammatory events. For this purpose, we evaluated the effects of NDS27, a water-soluble salt of curcumin combined with hydroxypropyl-β-cyclodextrin, on the activities of PKC, Nox2 and MPO. PKC activation was determined by western blotting with specific antibodies against phosphorylated PKC in extracts from PMNs after their incubation or not with NDS27. A cell-free assay was used to evaluate the effect of NDS27 before or after the assembly of Nox2 subunits. MPO activity was tested by the SIEFED technique in which NDS27 was pre-incubated with the enzyme and discarded before its activity measurement. An inhibition of PKC phosphorylation and Nox2 activity were observed at respectively 10-4 and 10-5 M of NDS27. The Nox2 inhibition was more pronounced when NDS27 was added before the assembly stimulation, suggesting a direct action of NDS27 on the subunits translocation. NDS27 also dose-dependently decreased the activity of MPO (21 % at 10-5 M), indicating an interaction with the enzyme structure. Our results demonstrated that NDS27 is a potent inhibitor of the two major enzymes responsible for ROS production in PMNs, and also acts on the activation cascade of Nox2. The modulatory effect of NDS27 towards the oxidant activity of PMNs opens therapeutic perspectives to control pathologies with excessive inflammatory reactions. [less ▲]

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See detailNew 5-Aryl-1H-imidazoles display in vitro antitumor activity against apoptosis-resistant cancer models, including melanomas, through mitochondrial targeting.
Mathieu, Véronique; Van Den Brege, E; Ceusters, Justine ULg et al

in Journal of Medicinal Chemistry (2013), 56(17), 6626-6637

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See detailThe challenge of understanding myopathies in horses using permeabilized muscle cells
Votion, Dominique ULg; Mouithys-Mickalad, Ange ULg; Ceusters, Justine ULg et al

in In proceedings 9th Conference on Mitochondrial Physiology (2013, September)

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See detailSensitivity and specificity of blood leukocyte counts as an indicator of mortality in horses after colic surgery
Salciccia, Alexandra ULg; Sandersen, Charlotte ULg; Grulke, Sigrid ULg et al

in Veterinary Record : Journal of the British Veterinary Association (2013), 173(11),

The objectives of this study were to describe and relate perioperative changes in blood leukocyte counts to the outcome of surgical colic horses, determine a cut-off value in the early postoperative ... [more ▼]

The objectives of this study were to describe and relate perioperative changes in blood leukocyte counts to the outcome of surgical colic horses, determine a cut-off value in the early postoperative period to obtain an indicator of the outcome, and compare the obtained value to a validation population of horses. Fifty-three horses undergoing colic surgery were included in the descriptive part of the study. Total leukocyte counts were performed before, during and serially after surgery. A receiver operating characteristic analysis was performed on the leukocyte counts of 45 of these horses to determine a cut-off value for the outcome. The results obtained were validated on a second set of 50 horses that underwent colic surgery in similar conditions. The kinetics of blood leukocytes in survivors was higher than in non-survivors during the first days. Non-survivor horses were more likely to have at least one blood leukocyte count ≤3.9×103/mm3 between 28 and 60 hours after surgery than survivor horses. This cut-off value was confirmed in the validation population. These results suggest that routine values of blood leukocyte counts can be used as an additional prognostic indicator after colic surgery alongside other predictors previously associated with the outcome. [less ▲]

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See detailPrédiction des performances sportives et détection précoce des myopathies par respirométrie à haute résolution
Votion, Dominique ULg; Serteyn, Didier ULg

in cheval athlète, stress oxydant & inflammation (2013, June 07)

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See detailIdentification of methylenecyclopropyl acetic acid in serum of European horses with atypical myopathy
Votion, Dominique ULg; van Galen, G; Sweetan et al

in Equine Veterinary Journal (2013)

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See detailMyeloperoxidase activity decreases in equine semen freezing extenders
Ponthier, Jérôme ULg; Franck, Thierry ULg; Niesten, Ariane ULg et al

in Amir, Arav (Ed.) Proceedings of the 3rd Cryo Congress (2013, March 23)

Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutrophils, and associated with decreased post-thaw motility of equine semen. This study aimed to compare MPO activity in pure ... [more ▼]

Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutrophils, and associated with decreased post-thaw motility of equine semen. This study aimed to compare MPO activity in pure equine freezing extender, raw and post-thaw semen. Active MPO Concentration (AMC) was measured with Specific Immunologic Extraction Followed by Enzymatic Detection in 20 ejaculates. Raw semen intra cellular AMC was determined in the supernatant after membrane lysis, each pellet containing 100x106 spermatozoa. AMC was also assayed in supernatants of semen frozen following a conventional method using INRA FreezeTM (IMV, France). Effect of freezing procedure on AMC was tested by experimentally adding 500ng of purified active MPO (Calbiochem, Germany) in 4 samples either with 5ml of PBS or INRA FreezeTM before assay. AMC was higher in sperm-rich pellet (0.306ng/ml) than in post-thaw semen (0.002ng/ml) (p=0.058). After experimental MPO addition, no activity variation was observed during the freezing procedure (after dilution, 1, 2 hours of cooling and post-thawing) within the same medium. Purified MPO activity was decreased in INRA FreezeTM when compared to PBS at all timings of sampling (p=0.0286). When all samples were pooled, remaining activity in INRA FreezeTM was 23.93±13.13%. MPO fixation on large proteins contained in the extender experimentally reduces AMC, as previously observed in plasma. However, AMC decrease observed during semen freezing is more important than after experimental addition. That could be explained by a MPO interaction with seminal plasma, a partial MPO release or a MPO inactivation during equine semen freezing. [less ▲]

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See detailIntra- and extracellular antioxidant capacities of the new water-soluble form of curcumin (NDS27) on stimulated neutrophils and HL-60 cells
Derochette, Sandrine ULg; Franck, Thierry ULg; Mouithys-Mickalad, Ange ULg et al

in Chemico-Biological Interactions (2013), 201(1-3), 49-57

Phagocytic cells, especially neutrophils (PMNs) are specialized in the production of reactive oxygen species (ROS) to kill pathogenic agents, but an excessive ROS production is associated with tissue ... [more ▼]

Phagocytic cells, especially neutrophils (PMNs) are specialized in the production of reactive oxygen species (ROS) to kill pathogenic agents, but an excessive ROS production is associated with tissue damages and inflammatory diseases. Phagocytes are thus prime therapeutic targets to control inflammatory events associated to ROS production. Nowadays, there is a growing interest for the use of polyphenols to modulate the inflammatory response. The aim of this work was to study the antioxidant effect of NDS27, a highly water-soluble form of the polyphenolic molecule curcumin, on in vitro stimulated equine PMNs and human promyelocytic leukemia cells (HL-60). NDS27 was either pre-incubated with cells and eliminated before their activation (intracellular effect) or let in the medium (extracellular effect). Our results indicate that NDS27 significantly and dose-dependently (10 6 M–10 4 M) inhibited the ROS production in both cell types without affecting their viability. NDS27 was able to cross and interact with cell membrane, especially for HL-60 cells, while we observed a better intracellular antioxidant effect with PMNs. The activity of myeloperoxidase (MPO) released by PMNs and HL-60 cells, was decreased by NDS27, but more efficiently for PMNs. These results suggested that the greater efficiency of NDS27 in PMNs is due to an inhibitory effect on cells which are more mature for ROS production, probably by targeting the enzymes implied in respiratory burst like MPO. The modulatory effect of NDS27 on the oxidant activity of cells involved in immune and inflammatory responses opens perspectives for a therapeutic control of pathologies with excessive inflammatory reactions. [less ▲]

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See detailOsteochondosis in foals
Sandersen, Charlotte ULg; Vander Heyden, Laurent ULg; Detilleux, Johann ULg et al

in Veterinary Record : Journal of the British Veterinary Association (2013), 127(17), 456-467

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