Concentration, Activity and Biochemical Characterization of Myeloperoxidase in Fresh and Post-thaw equine semen and their Implication on Freezability
Ponthier, Jérôme ; Franck, Thierry ; Parrilla Hernandez, Sonia 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 ▲]Detailed reference viewed: 35 (12 ULg)
Use of automatic stapling device during castration to prevent (re)occurrence of inguinal hernia in horses with large vaginal rings.
Salciccia, Alexandra ; de la Rebière de Pouyade, Geoffroy ; Ponthier, Jérôme et al
in Journal of Equine Veterinary Science (2014, January), 34(1), 99-100
(Re)ocurrence of inguinal hernia in case of very large vaginal rings may simply be prevented by closure of the vaginal tunic by TA staples during castration by inguinal approach. This method provides a ... [more ▼]
(Re)ocurrence of inguinal hernia in case of very large vaginal rings may simply be prevented by closure of the vaginal tunic by TA staples during castration by inguinal approach. This method provides a good resistance to internal pressure, appears to be safe, fast and easy to perform and may therefore be an interesting alternative to laparoscopic techniques when castration is considered. [less ▲]Detailed reference viewed: 21 (4 ULg)
Effect of non-sperm cells removal with single layer colloidal centrifugation on myeloperoxidase concentration in post-thaw equine semen
Ponthier, Jérôme ; ; Franck, Thierry 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 ▲]Detailed reference viewed: 37 (13 ULg)
Sensitivity and specificity of blood leukocyte counts as an indicator of mortality in horses after colic surgery
Salciccia, Alexandra ; Sandersen, Charlotte ; Grulke, Sigrid 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 ▲]Detailed reference viewed: 14 (2 ULg)
Sevoflurane inhibits equine myeloperoxidase release and activity in vitro.
MINGUET, Grégory ; de la Rebière de Pouyade, Geoffroy ; Franck, Thierry et al
in Veterinary Anaesthesia & Analgesia (2013), 40
Objective To investigate the effects of the volatile anaesthetic sevoflurane on the release of total and active myeloperoxidase (MPO) by non-stimulated and stimulated polymorphonuclear neutrophils (PMNs ... [more ▼]
Objective To investigate the effects of the volatile anaesthetic sevoflurane on the release of total and active myeloperoxidase (MPO) by non-stimulated and stimulated polymorphonuclear neutrophils (PMNs) in whole blood from healthy horses. Study design In vitro experimental study. Animals Adult healthy horses. Methods Samples of whole venous blood were collected and incubated in air or in air plus 2.3% or 4.6% sevoflurane for 1 hour. PMNs were stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP), with a combination of cytochalasin B (CB) and fMLP or with phorbol myristate acetate (PMA). Total and active MPO contents released by PMNs in blood were measured by enzyme-linked immunosorbent assay (ELISA) and specific immunological extraction followed by enzymatic detection (SIEFED) respectively. Additional experiments were performed to assess the effect of sevoflurane on the peroxidase and chlorination cycles of purified equine MPO using Amplex Red and 3'-(p-aminophenyl) fluorescein as fluorogenic substrates respectively. Results As compared with air alone, 1 hour exposure of whole blood to 4.6% sevoflurane in air significantly inhibited the release of total and active MPO by unstimulated and both fMLP- and CB + fMLP-stimulated PMNs but not by PMA-stimulated PMNs. Although 2.3% sevoflurane had no effect on total MPO release by unstimulated and stimulated PMNs, it significantly reduced the release of active MPO by unstimulated and fMLP-stimulated PMNs. Additionally, sevoflurane reversibly inhibited the activity of MPO, especially the peroxidase cycle of the enzyme. Conclusions and clinical relevance Although our experimental study was not designed to assess the effects of sevoflurane in vivo, this inhibition of MPO release and activity may have relevance for anaesthetized horses and deserves further studies to examine the clinical importance of these findings. [less ▲]Detailed reference viewed: 24 (9 ULg)
An in vitro whole blood model to test the effects of different stimuli conditions on the release of myeloperoxidase and elastase by equine neutrophils.
Ceusters, Justine ; Serteyn, Didier ; MINGUET, Grégory et al
in Veterinary Immunology and Immunopathology (2012), 150(3-4), 221-7
Horses are particularly sensitive and exposed to excessive inflammatory responses evolving toward an important stimulation of polymorphonuclear neutrophils (PMNs). The aim of this work was to stimulate ... [more ▼]
Horses are particularly sensitive and exposed to excessive inflammatory responses evolving toward an important stimulation of polymorphonuclear neutrophils (PMNs). The aim of this work was to stimulate equine neutrophils in whole blood and to evaluate their response by measuring the release of total and active myeloperoxidase (MPO) and total elastase, considered as markers of neutrophil stimulation and degranulation. Because of the critical importance of the concomitant presence of LPS and TNF-alpha in equine pathological situations, we combined these two natural mediators to stimulate PMN and compared the response with those obtained after the PMN stimulation with each mediator used alone and well-known artificial stimulation systems such as 12-phorbol 13-myristate acetate (PMA) and the combination of cytochalasin B (CB) and N-formyl-methionyl-leucyl-phenylalanine (fMLP). All the activation systems, PMA, CB/fMLP, TNF-alpha, LPS and LPS/TNF-alpha, induced a significant release of total MPO in whole blood but only the combinations CB/fMLP and LPS/TNF-alpha significantly favored the release of active MPO. Regarding the total elastase, we did not observe a significant release in all the stimulated conditions except with PMA. It appears clearly that the choice of the neutrophil stimulation model is fundamental for the selection of potentially active pharmacological agents, especially on MPO activity. [less ▲]Detailed reference viewed: 32 (17 ULg)
Surgical treatment of open joint injuries: a retrospective study of 22 horses.
Salciccia, Alexandra ; de la Rebière de Pouyade, Geoffroy ; Evrard, Laurence et al
Conference (2012, October 19)
Aims: To evaluate the prognosis of horses treated surgically for open joint injuries. Methods: The medical records of horses admitted for surgical treatment of a wound in communication with a joint were ... [more ▼]
Aims: To evaluate the prognosis of horses treated surgically for open joint injuries. Methods: The medical records of horses admitted for surgical treatment of a wound in communication with a joint were reviewed. A telephone questionnaire was used for the long term evaluation. Fisher’s exact tests were used for the statistical analyses. Results: Twenty two horses were included in the study with the following distribution of lesions: 6 carpi, 6 fetlocks, 4 tibiotarsal joints, 2 proximal and 3 distal interphalangeal joints, 1 elbow. The duration of the injury before referral ranged from 3 hours to 10 days. Surgical treatment consisted of 1-3 joint lavages. Of the 22 horses, 4 were euthanatized during hospitalization and 18 were discharged. After discharge, 3 horses died due to colic, 2 were lost and 13 were still alive. The survival was not influenced by the duration of the wound. All horses with cutaneous defects less than 5 cm and all horses affected in the lower limb (below the level of the canon) were discharged. Having an affected joint proximal to the canon was significantly associated to the need of multiple surgeries (OR: 17.5; p= 0.024). Conclusions: Even if the prognosis remains guarded for open joint injuries, a long delay between injury and treatment should not be systematically associated with a bad prognosis for survival. Open joint injuries of the lower limb were associated with survival. They required less often multiple articular lavages than open joint injuries of the upper limb, warranting thus a better prognosis. [less ▲]Detailed reference viewed: 59 (14 ULg)
Les kératomes: lésions et traitements chez 19 chevaux
Salciccia, Alexandra ; de la Rebière de Pouyade, Geoffroy ; Bolen, Géraldine et al
Conference (2012, October 12)
Introduction: Les kératomes, masses hyperplasiques de kératine, croissent entre la phalange distale (P3) et la paroi du sabot et provoquent des boiteries chez le cheval. La littérature comporte de ... [more ▼]
Introduction: Les kératomes, masses hyperplasiques de kératine, croissent entre la phalange distale (P3) et la paroi du sabot et provoquent des boiteries chez le cheval. La littérature comporte de nombreux rapports de cas isolés mais peu de séries cliniques. Matériel et méthodes: Les dossiers cliniques des cas de kératomes présentés depuis 2004 à la faculté de médecine vétérinaire de Liège ont été revus. Résultats: Vingt-trois kératomes ont été diagnostiqués sur 19 chevaux boiteux. Un cheval présentait 2 kératomes sur le même pied et 3 chevaux présentaient 2 kératomes sur des pieds différents. Sur les 22 pieds atteints, les lésions observées étaient: un abcès, souvent récidivant (18 cas), une déformation de la muraille (6 cas), une déviation de la ligne blanche (6 cas) et une seime (3 cas). Un kératome était situé sous la sole et 22 sous la muraille. La durée des symptômes variait de 2 semaines à 15 ans (plus d'un an sur 12 pieds). Sur les radiographies, une lyse par compression de P3 était visible en regard de 20 kératomes. Le cheval présentant 2 kératomes sur le même pied a été euthanasié. Trois kératomes ont été traités de manière conservative. Dix-huit kératomes (chez 16 chevaux) ont été excisés chirurgicalement dont 16 par avulsion complète de muraille, 1 par avulsion partielle de muraille et 1 par curetage d'une portion de sole. Aucune complication postopératoire n'a été observée dans 7/18 cas (39%). Les complications rencontrées étaient: une granulation excessive (10 cas), une douleur importante pendant plusieurs jours (5 cas), une légère infection (4 cas), un enfoncement de P3 en regard de la zone avulsée (2 cas) et une récidive (2 cas, dont 1 a été réopéré avec succès ultérieurement). Un cheval est encore en convalescence au moment de cette étude. Le suivi à long terme d’un cheval a été perdu. Des 14 chevaux repris dans l'évaluation postopératoire, 1 a été euthanasié pour une autre raison que le kératome, 3 chevaux ont gardé une boiterie résiduelle au trot et 10 chevaux ont récupéré leur niveau d'activité. Discussion: La littérature décrit les kératomes comme rares1. Or, 21% des chevaux de cette étude (4 chevaux sur 19) présentaient plusieurs kératomes. De plus, dans 54 % des cas, les symptômes duraient depuis plus d’un an. Dès lors, il apparait que les kératomes sont sous diagnostiqués ou le sont souvent tardivement. La chronicité et l’étendue des kératomes de cette étude n’a que très rarement permis leur résection par avulsion partielle de muraille, technique de choix dont les complications postopératoires sont moindres². Un diagnostic précoce par inspection minutieuse des sabots, particulièrement en cas d’abcès de pied récidivant permettrait une excision par une technique moins invasive, qui diminuerait la convalescence et les complications postopératoires. Bibliographie: 1. Sundberg, J.P. et al. Neoplasms of Equidae. Journal of American veterinary medical association, 1977, 170: 150-152. 2. Boys Smith SJ. et al. Complete and partial hoof wall resection for keratoma removal: postoperative complications and final outcome in 26 horses (1994-2004). Equine Veterinary Journal, 2006, 38 (2): 127-133. [less ▲]Detailed reference viewed: 280 (30 ULg)
Intérêt d’un système de traitement des plaies par pression négative chez le cheval : Etat des lieux et cas cliniques
de la Rebière de Pouyade, Geoffroy ; Salciccia, Alexandra ; Gougnard, Alexandra et al
Conference (2012, October 12)
Les plaies traumatiques ou chirurgicales nécessitent souvent une cicatrisation par seconde intention lorsque la fermeture primaire n’est pas possible ou échoue. La fermeture assistée par le vide (Vacuum ... [more ▼]
Les plaies traumatiques ou chirurgicales nécessitent souvent une cicatrisation par seconde intention lorsque la fermeture primaire n’est pas possible ou échoue. La fermeture assistée par le vide (Vacuum Assisted Closure : VAC) est une méthode active et non invasive qui expose le lit de la plaie à une pression sub-atmosphérique locale afin de promouvoir sa cicatrisation. Elle stimule la granulation et la perfusion sanguine locale et réduit significativement l’œdème et la charge bactérienne.1 Cette technique s’est révélée efficace dans le traitement des plaies aiguës et chroniques chez l’homme. Chez le cheval, la méthode est encore peu décrite.2,3 Nous rapportons ici l’application et les effets de la thérapie des plaies par pression négative à travers les cas traités à la clinique vétérinaire universitaire de Liège depuis 2010. Les plaies étaient chirurgicalement débridées avant l’application du système VAC. Une mousse en polyuréthane était appliquée après avoir été adaptée à la géométrie de la plaie. Un champ plastique adhésif était alors apposé sur la plaie pour créer un environnement hermétique et un orifice d’1 cm2 était créé au centre du système pour appliquer la ventouse du tuyau d’aspiration. Cette ligne était alors raccordée à l’appareil d’aspiration contrôlée pour maintenir une pression continue de -125 mmHg dans la plaie. Quinze cas ont été recensés: 3 plaies intra-articulaires (au carpe, au coude et au grasset), 3 sur la face dorsale du canon postérieur, 2 sur la face dorsale du jarret, 1 sur la pointe du calcanéum, 2 sur l’encolure et 4 plaies de laparotomie infectées. Le traitement a accéléré la croissance du tissu de granulation, stimulé la contraction des plaies et favorisé la cicatrisation dans la majorité des cas (12/15). Les plaies apparaissaient aussi plus saines et présentaient moins de sécrétions lors des changements de bandages. Des résultats plus modérés ont été notés chez les 3 autres cas. Quelques complications mineures comme des irritations cutanées ont été observées. Le traitement de plaies contaminées nous a orientés vers l’usage d’une mousse de polyuréthane argenté mais d’autres types de matrice sont disponibles et permettent de modifier le comportement de la plaie. L’usage d’une mousse de polyvinyl Alcool s’est ainsi avéré efficace pour améliorer l’adhésion des greffons lors d’une greffe de peau chez le cheval.3 La principale limite du traitement réside dans la difficulté technique de maintenir la plaie sous vide lors d’atteinte de régions très mobiles comme les articulations. En conclusion, le système VAC semble très prometteur pour optimiser la qualité de la cicatrisation et réduire la durée de l’hospitalisation. 1 - Morykwas, M.J., Argenta, L.C., et al. Vacuum-assisted closure: a new method for wound control and treatment: animal studies and basic foundation. Ann. Plast. Surg. 1997, 38, 553-562 2 - Gemeinhardt K.D., Molnar J.A. Vacuum-assisted closure for management of a traumatic neck wound in a horse. Equine Vet. Educ. 2005, 17, 27-33 3 - Jordana M., Pint E., et al. The use of vacuum-assisted wound closure to enhance skin graft acceptance in a horse. Vlaams Diergeneeskundig Tijdschrift 2011, 80, 343-350 [less ▲]Detailed reference viewed: 117 (9 ULg)
Myeloperoxidase in Equine Semen: Concentration and Localization during Freezing Processing
Ponthier, Jérôme ; ; Franck, Thierry et al
in Journal of Equine Veterinary Science (2012), (32), 32-37
Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutro- phils during degranulation or after lysis. Post-thaw semen contains MPO, and concen- tration of this enzyme is associated ... [more ▼]
Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutro- phils during degranulation or after lysis. Post-thaw semen contains MPO, and concen- tration of this enzyme is associated with decreased motility. The aim of this study was to determine kinetics of MPO concentration during freezing, its origin, and its impact on frozenethawed semen. Forty ejaculates were used. Semen was frozen using the classical freezing procedure. MPO concentrations were assayed in fresh semen, after centrifuga- tion, and after cooling down to 4 C. Post-thaw MPO assay results and spermogram characteristics were determined. MPO immunocytochemistry was performed on 4 different ejaculates at each step of freezing procedure. MPO concentration increased after cooling down to 4 C and thawing compared with fresh semen. As temperature decreased, MPO was higher or tended to be higher in post-thaw poor quality samples. Nonsperm cells showed various degrees of MPO immunostaining and were observed as epithelial cells with nuclear pyknosis and keratinization. MPO immunostaining increased in medium and decreased in nonsperm cells during freezing. Our study shows that MPO concentration in equine semen increases when temperature decreases. We hypothesize that nonsperm cells present in fresh semen could release MPO. [less ▲]Detailed reference viewed: 60 (15 ULg)
Assessment of reactive oxygen species production in cultured equine skeletal myoblasts in response to conditions of anoxia followed by reoxygenation with or without exposure to peroxidases.
Ceusters, Justine ; Mouithys-Mickalad, Ange ; de la Rebière de Pouyade, Geoffroy et al
in American Journal of Veterinary Research (2012), 73(3), 426-434
Objective—To culture equine myoblasts from muscle microbiopsy specimens, examine myoblast production of reactive oxygen species (ROS) in conditions of anoxia followed by reoxygenation, and assess the ... [more ▼]
Objective—To culture equine myoblasts from muscle microbiopsy specimens, examine myoblast production of reactive oxygen species (ROS) in conditions of anoxia followed by reoxygenation, and assess the effects of horseradish peroxidase (HRP) and myeloperoxidase (MPO) on ROS production. Animals—5 healthy horses (5 to 15 years old). Procedures—Equine skeletal myoblast cultures were derived from 1 or 2 microbiopsy specimens obtained from a triceps brachii muscle of each horse. Cultured myoblasts were exposed to conditions of anoxia followed by reoxygenation or to conditions of normoxia (control cells). Cell production of ROS in the presence or absence of HRP or MPO was assessed by use of a gas chromatography method, after which cells were treated with a 3,3′-diaminobenzidine chromogen solution to detect peroxidase binding. Results—Equine skeletal myoblasts were successfully cultured from microbiopsy specimens. In response to anoxia and reoxygenation, ROS production of myoblasts increased by 71%, compared with that of control cells. When experiments were performed in the presence of HRP or MPO, ROS production in myoblasts exposed to anoxia and reoxygenation was increased by 228% and 183%, respectively, compared with findings for control cells. Chromogen reaction revealed a close adherence of peroxidases to cells, even after several washes. Conclusions and Clinical Relevance—Results indicated that equine skeletal myoblast cultures can be generated from muscle microbiopsy specimens. Anoxia-reoxygenation– treated myoblasts produced ROS, and production was enhanced in the presence of peroxidases. This experimental model could be used to study the damaging effect of exercise on muscles in athletic horses. [less ▲]Detailed reference viewed: 54 (27 ULg)
Dystocies équines: Données descriptives et approche de la gestion des soins intensifs
Deleuze, Stefan ; de la Rebière de Pouyade, Geoffroy ; Sandersen, Charlotte et al
Conference (2011, December 02)
La fréquence de fourbures rencontrées après une dystocie semble faible par rapport à la littérature. L’ajout d’héparines fractionnées aux traitements habituels de la métrite du post-partum (ocytocine ... [more ▼]
La fréquence de fourbures rencontrées après une dystocie semble faible par rapport à la littérature. L’ajout d’héparines fractionnées aux traitements habituels de la métrite du post-partum (ocytocine, lavage utérins, antibiotiques et anti-inflammatoires) semble intéressant pour réduire le risque de fourbure. [less ▲]Detailed reference viewed: 68 (10 ULg)
Time trends of blood leucocytes, neutrophils and plasmatic myeloperoxidase in the perioperative period of horses undergoing colic surgery.
Salciccia, Alexandra ; Grulke, Sigrid ; Detilleux, Johann et al
Poster (2011, July)
Background: Despite the recent advances in this area, colic remains a major cause of morbidity and death in horses. Neutrophilic activation and degranulation may play a key role in the postoperative ... [more ▼]
Background: Despite the recent advances in this area, colic remains a major cause of morbidity and death in horses. Neutrophilic activation and degranulation may play a key role in the postoperative complications. Activated neutrophils release enzymes like proteases and myeloperoxidase (MPO). MPO concentrations in plasma and tissue are considered as a marker of neutrophil activation. (McConnico et al. 1999; Hoy et al. 2002). When freed in the tissue, active MPO is able to oxidize, nitrate and chlorate most organic molecules (Klebanoff 2005). Objectives: The aim of this study was 1) to determine the time trends of blood leukocyte and neutrophil counts as well as of plasmatic MPO concentrations in the perioperative period of horses undergoing colic surgery and 2) to relate these time trends to the location of the pathology, the severity of postoperative complications and to the outcome of the patients. Methods: Fifty two horses undergoing colic surgery at the Equine Teaching Hospital of the University of Liege were included in this study. The location of the predominant lesion of the intestine, the severity of the postoperative complications and the outcome were recorded for each horse. Total leukocyte and neutrophil counts were performed in all of the horses while plasmatic myeloperoxidase levels were determined in 16 of them. The blood samplings were realized before and during the surgery (after correction of the intestinal lesion), during the recovery and every 4 hours during the first 4 days (from day 0 until day 4) and then every 12 hours until day 6 (150th hour after the first blood sampling) or until euthanasia. Hematologic analyses were performed at the time of sampling by use of the Medonic CA 530 (Menarini, Zaventem, Belgium). The blood was then centrifuged and the plasma was aliquoted and frozen at -20° C until assayed. MPO was assayed with a specific enzyme-linked immunosorbent assay (Equine MPO-ELISA kit, BiopTis, Liège, Belgium). A mixed model was used to analyze the time trends of leukocytes, neutrophils and MPO. All computations were done with the SAS (Statistical Analysis System) procedure Proc Mixed, with Satterthwaite degrees of freedom. The significance level was set at p=0.05. Results: The main pathology was found in the large intestine in 30 horses (58%) and the small intestine in 22 horses (42%). Forty horses (77%) survived to discharge from the clinic. Twelve horses were euthanized during the postoperative period. Their survival time varied from 0.5 day to 20 days with a mean of 7.8 days. Twenty-two horses (42%) showed none or mild complications, 12 horses (23%) showed moderate complications and 18 horses (35%) suffered from severe complications. Time trends for leukocytes and neutrophils were similar to each other (p= 0.7205) and significantly different (p< 0.0001) from the MPO time trend, which increased during the first hours, while the neutrophil time trend decreased immediately after the admission. The time trend of neutrophils was higher in large intestinal than in small intestinal pathologies and the time trend of MPO was lower in large intestinal than in small intestinal pathologies. The time trends of neutrophils were significantly different between the degrees of complications (no/mild vs moderate vs severe). For the first part of the curve, the more severe the complication, the lower is the time trend. The time trend of MPO was lower in survivors. The time trend of neutrophils in survivors was higher during the first 4 days thereafter it becomes lower than in non survivors. Conclusions: These results confirm that neutrophil counts and MPO levels undergo timely changes and that they are related to the severity of the inflammatory reaction in surgical colic cases. Knowing the kinetics of these parameters is an essential step to further determine cut-off values (with a larger group of horses) for the prognosis of horses after colic surgery. References Hoy, A., Leininger-Muller, B., Kutter, D., Siest, G. and Visvikis, S. (2002) Growing significance of myeloperoxidase in non-infectious diseases. Clin. Chem. Lab. Med. 40, 2-8. Klebanoff, S.J. (2005) Myeloperoxidase: friend and foe. J. Leukoc. Biol. 77, 598-625. McConnico, R.S., Weinstock, D., Poston, M.E. and Roberts, M.C. (1999) Myeloperoxidase activity of the large intestine in an equine model of acute colitis. Am. J. Vet. Res. 60, 807-813. [less ▲]Detailed reference viewed: 26 (5 ULg)
Evaluation of the effect of general anaesthesia on ultrasonographic images of the small intestine in horses.
Salciccia, Alexandra ; Gougnard, Alexandra ; de la Rebière de Pouyade, Geoffroy et al
Poster (2011, July)
ultrasonography is currently more and more used in equine acute abdominal disease as well as in the follow-up of surgical colic patients because of its sensibility for the detection of small intestinal ... [more ▼]
ultrasonography is currently more and more used in equine acute abdominal disease as well as in the follow-up of surgical colic patients because of its sensibility for the detection of small intestinal distension. General anaesthesia is known to diminish gastrointestinal motility even if there are no clinical signs associated with it. The aim of this study was to evaluate the effect of general anaesthesia on transabdominal ultrasonographic images of the small intestine in horses undergoing routine non abdominal surgeries. The ultrasonographic exams were performed in 19 horses before and after the general anaesthesia (immediately after the recovery and then 2h, 12h and 24h after it). Each ultrasonographic exam focused on the duodenum, the jejunum and the presence of peritoneal fluid. For the jejunum, 5 locations were used: on the ventral midline just caudally to the sternum, on the left and right cranial ventral parts of the abdomen, on the left and right inguinal regions. Other parameters such as the gut sounds and the postoperative fecal output were also recorded. Anova and Chi-square tests were used for the statistical analysis. No horse showed colic signs. No significant difference was found between the pre and post anaesthetic period considering the maximal diameter of the duodenum, the maximal diameter of the jejunum on the 5 locations and the peritoneal fluid. The contractions of the duodenum were increased at the recovery compared to before the anaesthesia (p= 0,0299). The small intestine was most visible at the recovery (57,5%) and then at 2 hours after it (38,3%). It seemed that the ventral midline just caudally to the sternum and the right and left inguinal regions were the best locations to observe the jejunum (with a mean of respectively 44,6%, 39,8% and 38,5% of visualization of the jejunum).The gut sounds were very significantly decreased at the recovery (p < 0,0001) and at 2h after it (p = 0,0006). The postoperative fecal output was not decreased. In conclusion, even if general anaesthesia seems to reduce temporarily the intestinal activity (decreased gut sounds in the early post anaesthetic period), it does not cause significant distension of the small intestine (almost the small intestinal diameters were in the normal range). If an increased diameter of small intestine is observed by ultrasonography after surgery it should therefore be attributed to a pathological process and not to the anaesthesia. [less ▲]Detailed reference viewed: 13 (1 ULg)
Modulating effects of acepromazine on the reactive oxygen species production by stimulated equine neutrophils
Sandersen, Charlotte ; Mouithys-Mickalad, Ange ; de la Rebière de Pouyade, Geoffroy et al
in Veterinary Anaesthesia & Analgesia (2011), 38
To investigate the effect of acepromazine (ACP) on reactive oxygen species (ROS) production by stimulated equine neutrophils.Detailed reference viewed: 23 (5 ULg)
Production of free radicals and oxygen consumption by primary equine endothelial cells during anoxia-reoxygenation.
de la Rebière de Pouyade, Geoffroy ; Salciccia, Alexandra ; Ceusters, Justine et al
in Open Biochemistry Journal (The) (2011), 5
The endothelium plays an active role in ischemia/reperfusion injuries. Herein, we report the effect of a single or successive cycles of anoxia/reoxygenation (A/R) on the mitochondrial respiratory function ... [more ▼]
The endothelium plays an active role in ischemia/reperfusion injuries. Herein, we report the effect of a single or successive cycles of anoxia/reoxygenation (A/R) on the mitochondrial respiratory function of equine endothelial cells (cultured from carotids) monitored by high resolution oxymetry, and on their production of reactive oxygen species (ROS). ROS were measured by electron paramagnetic resonance (ESR) using POBN and DMPO spin traps, and by gas chromatography (GC) of ethylene released by ROS-induced alpha-keto-gamma-(methylthio)butyric acid (KMB) oxidation. The oxygen consumption significantly decreased with the number of A/R cycles, and POBN-ESR spectra were specific of adducts formed in the cells from superoxide anion. After a one-hour A/R cycle, high intensity DMPO-ESR spectra were observed and assigned to superoxide anion trapping; the GC results confirmed an important production of ROS compared to normoxic cells. These results show that A/R induces mitochondrial alterations in endothelial cells, and strongly stimulates their oxidative activity as demonstrated by ESR and GC methods. [less ▲]Detailed reference viewed: 29 (6 ULg)
The role of activated neutrophils in the early stage of equine laminitis.
de la Rebière de Pouyade, Geoffroy ; Serteyn, Didier
in Veterinary Journal (2011), 189(1), 27-33
Despite ongoing research and a widening range of treatment options, laminitis remains a severely damaging condition with poorly understood pathophysiology. Results obtained from cytokine regulation ... [more ▼]
Despite ongoing research and a widening range of treatment options, laminitis remains a severely damaging condition with poorly understood pathophysiology. Results obtained from cytokine regulation studies during the last decade have highlighted the inflammatory nature of laminitis. This review will describe the role of systemic activation and local infiltration of neutrophils in laminar tissues in the induction of laminitis. Particular emphasis is placed on the role of neutrophil activation in subsequent vascular dysfunction and oxidative and proteolysis imbalances that are pathways previously implicated in laminitis. Neutrophils, by the way of their interdependent relationship with endothelial cells and keratinocytes, dramatically increase the inflammatory response culminating in the failure of the laminar dermal-epidermal interface. [less ▲]Detailed reference viewed: 22 (3 ULg)
Development of an enzyme-linked immunosorbent assay for equine neutrophil elastase measurement in blood: Preliminary application to colic cases.
de la Rebière de Pouyade, Geoffroy ; Franck, Thierry ; Salciccia, Alexandra et al
in Veterinary immunology and immunopathology (2010)
Equine neutrophil elastase (NE) is a protease released in inflammatory diseases and participating in tissue destruction. To measure NE in horse plasma to assess its role in pathological conditions, we ... [more ▼]
Equine neutrophil elastase (NE) is a protease released in inflammatory diseases and participating in tissue destruction. To measure NE in horse plasma to assess its role in pathological conditions, we purified elastase from equine neutrophils by a double step chromatography and obtained a pure protein of 27kDa, 4kDa smaller than the NE 2A previously purified (Scudamore et al., 1993; Dagleish et al., 1999), which was likely to be NE 2B. We developed an ELISA by using two specific polyclonal antibodies obtained from rabbit and guinea pig. The sandwich complex was detected using a secondary antibody conjugated to alkaline phosphatase. The ELISA showed good precision and accuracy, with intra- and inter-assay coefficients of variation below 10% for equine NE concentrations ranging from 1.875 to 60ng/ml. A stable plasma NE value, unaffected by the delay of centrifugation (over 4h), was obtained with plasma from EDTA anticoagulated blood. The mean value (+/-SEM) measured in 37 healthy horses was 32.53+/-4.6ng/ml. NE level in plasma of horses with colic at the time of admission was significantly higher than in healthy horses. Our results indicate that the ELISA technique we developed to measure plasmatic NE is a powerful tool for studying the role of elastase in equine inflammatory disease. In future, the application will be extended to other equine biological fluids. [less ▲]Detailed reference viewed: 51 (8 ULg)
Is neutrophil elastase associated with myeloperoxidase concentration and post-thawing parameters in equine frozen semen?
Ponthier, Jérôme ; de la Rebière de Pouyade, Geoffroy ; et al
in Animal Reproduction Science (2010), 121S
Non sperm cells concentration is higher in unfreezable semen. A relation between non-sperm cells in fresh semen and MPO and post thawing quality has been observed. Neutrophil Elastase seems to have no ... [more ▼]
Non sperm cells concentration is higher in unfreezable semen. A relation between non-sperm cells in fresh semen and MPO and post thawing quality has been observed. Neutrophil Elastase seems to have no effect on semen characteristics and to be not associated with on-sperm cells and freezability. [less ▲]Detailed reference viewed: 81 (28 ULg)
EFFECT OF DIFFERENT ANOXIA/REOXYGENATION MODELS ON MITOCHONDRIAL COMPLEX I ACTIVITY OF CULTURED EQUINE SKELETAL MUSCLE CELLS: ESR AND OXYGRAPHIC STUDIES.
Ceusters, Justine ; Mouithys-Mickalad, Ange ; Niesten, Ariane et al
Poster (2010)Detailed reference viewed: 17 (9 ULg)