Inhibitory Effect of Curcuminoids and Tetrahydrocurcuminoids on Equine Activated Neutrophils and Myeloperoxidase Activity

in Physiological Research (2007), 57(4), 577-587

In the horse, the inflammation response to various pathologies (intestinal strangulations, laminitis, etc.) involves an excessive stimulation of the polymorphonuclear neutrophils releasing reactive oxygen ... [more ▼]

In the horse, the inflammation response to various pathologies (intestinal strangulations, laminitis, etc.) involves an excessive stimulation of the polymorphonuclear neutrophils releasing reactive oxygen species (ROS) and myeloperoxidase (MPO). The aim of the present work was to study the effect of natural polyphenols, curcuminoids and tetrahydrocurcuminoids (THC) on isolated stimulated equine neutrophils and on the activity of purified MPO. The ROS production and the release of MPO by activated neutrophils were measured by chemiluminescence and ELISA techniques, respectively. The activity of purified MPO was measured by studying its nitration, chlorination or oxidation capacity and by using an original method called SIEFED allowing the study of drug interaction with the enzyme without interferences of the medium. Curcuminoids and THC had dose-dependent inhibitory effects on ROS production and MPO release by activated neutrophils and on purified MPO activity. We suggest that the higher efficacy of curcuminoids versus THC could be explained, at least partially, by its chemical structure: the conjugated double bounds and the plane structure of curcuminoids made easier the neutralization of the radical species generated by activated neutrophils and the interaction of the drug with the active site of MPO. These inhibitory effects of curcuminoids on the oxidant activity of equine neutrophils and on MPO activity open therapeutic perspectives in equine pathologies with excessive inflammatory reactions. [less ▲]

Neutrophil myeloperoxidase measurements in plasma, laminar tissue, and skin of horses given black walnut extract

in American Journal of Veterinary Research (2007), 68(1), 81-86

Objective-To compare measurements of myeloperoxidase (MPO) in plasma, laminar tissues, and skin obtained from control horses and horses given black walnut heartwood extract (BWHE). Animals-22 healthy 5 ... [more ▼]

Objective-To compare measurements of myeloperoxidase (MPO) in plasma, laminar tissues, and skin obtained from control horses and horses given black walnut heartwood extract (BWHE). Animals-22 healthy 5- to 15-year-old horses. Procedures-Horses were randomly assigned to 4 groups as follows: a control group given water (n = 5) and 3 experimental groups given BWHE (17) via nasogastric intubation Experimental groups consisted of 5, 6, and 6 horses that received BWHE and were euthanatized at 1.5, 3, and 12 hours after intubation, respectively. Control horses were euthanatized at 12 hours after intubation. Plasma samples were obtained hourly for all horses. Laminar tissue and skin from the middle region of the neck were harvested at the time of euthanasia. Plasma and tissue MPO concentrations were determined via an ELISA; tissue MPO activity was measured by use of specific immunologic extraction followed by enzymatic detection. Results-Tissues and plasma of horses receiving BWHE contained significantly higher concentrations of MPO beginning at hour 3. Laminar tissue and skin from horses in experimental groups contained significantly higher MPO activity than tissues from control horses. Concentrations and activities of MPO in skin and laminar tissues were similar over time. Conclusions and Clinical Relevance-in horses, BWHE administration causes increases in MPO concentration and activity in laminar tissue and skin and the time of increased MPO concentration correlates with emigration of WBCs from the vasculature. These findings support the hypothesis that activation of peripheral WBCs is an early step in the pathogenesis of acute laminitis. [less ▲]

Method and kit for the measurement of neutrophil cell activation

Patent (2006)

The present invention is related to methods and kits (or devices) for the measurement of equine myeloperoxidase (MPO), a specific enzyme of equine neutrophils, either in total [first method], or ... [more ▼]

The present invention is related to methods and kits (or devices) for the measurement of equine myeloperoxidase (MPO), a specific enzyme of equine neutrophils, either in total [first method], or specifically in its active form [second method]. Said methods and kits (or devices), used independently or in combination, find improved applications in the veterinary field and can be adapted for application in human health care. The concept of the second method is applicable to any other enzyme. [less ▲]

An in vitro study of the modulating activity of indole derivatives on H2O2/hemic enzyme systems.

Poster (2006, May 12)

Myeloperoxidase concentration in bronchoalveolar lavage fluid from healthy horses and those with recurrent airway obstruction

in Canadian Journal of Veterinary Research = Revue Canadienne de Recherche Vétérinaire (2006), 70(4), 291-296

The aim of this work was to measure the myeloperoxidase (MPO) concentration in bronchoalveolar lavage (BAL) fluid collected from horses with recurrent airway obstruction (RAO), both in crisis and in ... [more ▼]

The aim of this work was to measure the myeloperoxidase (MPO) concentration in bronchoalveolar lavage (BAL) fluid collected from horses with recurrent airway obstruction (RAO), both in crisis and in remission, as well as from healthy horses. Seven horses with RAO were exposed to moldy hay until the maximum change in pleural pressure was greater than 1.5 kPa. At that point, BAL was performed, and the total cell counts and percentages in the fluid were immediately determined. To measure the MPO concentration in BAL-fluid supernatant, we used a specific enzyme-linked immunosorbent assay with polyclonal antibodies against equine MPO. The tests were repeated on the horses with RAO after they had spent 2 mo on pasture. Six healthy horses serving as controls underwent the same tests. The absolute and relative neutrophil counts and the MPO concentration in the BAL fluid were significantly greater in the horses with an RAO crisis than in the control horses. After 2 mo on pasture, the horses that had been in RAO crisis were clinically normal, and their neutrophil counts and MPO levels in BAL fluid had significantly decreased; during remission their neutrophil counts were not significantly different from those in the healthy horses, but their MPO concentration remained significantly higher. This study showed that determining the MPO concentration in a horse's BAL fluid is technically possible and that during remission from RAO the concentration remains higher than normal. Thus, MPO may be a marker of neutrophil presence and activation in the lower airways. [less ▲]

Development of an enzyme-linked immunosorbent assay for specific equine neutrophil myeloperoxidase measurement in blood

in Journal of Veterinary Diagnostic Investigation (2005), 17(5), 412-419

Equine inflammatory disease is accompanied by a neutrophil activation resulting in the release of granulocytic enzyme myeloperoxidase (MPO). To measure MPO in horse plasma as marker of neutrophil ... [more ▼]

Equine inflammatory disease is accompanied by a neutrophil activation resulting in the release of granulocytic enzyme myeloperoxidase (MPO). To measure MPO in horse plasma as marker of neutrophil activation, the authors purified equine neutrophil MPO and developed a specific enzyme immunoassay using 2 specific polyclonal antibodies obtained from rabbit (primary antibody) and guinea pig (secondary antibody). The sandwich complex "primary antibody-MPO-secondary antibody" was detected using a goat anti-guinea pig immunoglobulin antibody conjugated to alkaline phosphatase. The enzyme-linked immunosorbent assay (ELISA) showed good precision and accuracy, with intra- and interassay coefficients of variation below 10% for MPO concentrations ranging from 0.78 to 50 ng/ml. A stable plasma MPO value, unaffected by time elapsed between blood collection and centrifugation, was obtained with plasma from EDTA anticoagulated blood. The mean MPO value measured in 38 healthy horses was 181.80 +/- 64.74 ng/ml. In 20 horses suffering from obstruction of the large or small intestine, MPO concentrations measured at the time of arrival at the intensive care unit were significantly higher than mean normal value, ranging from 477.88 to 2,748.13 ng/ml. Work is in progress to apply this MPO ELISA technique to other biological fluids and other equine diseases. [less ▲]

Design of an ELISA specific for the measurement of equine neutrophil myeloperoxidase in biological fluids

in Clinical Chemistry (2005), 51(Suppl. 6), 211-212

Hyperhydricity of micropropagated shoots: a typically stress-induced change of physiological state

in Plant Cell, Tissue & Organ Culture (2004), 77(2), 181-191

Hyperhydricity of micropropagated shoots, formerly called vitrification, undoubtedly results from growth and culture conditions, subjectively reputated as stressing factors: wounding, infiltration of soft ... [more ▼]

Hyperhydricity of micropropagated shoots, formerly called vitrification, undoubtedly results from growth and culture conditions, subjectively reputated as stressing factors: wounding, infiltration of soft culture medium, generally of a high ionic strength, rich in nitrogen and in growth regulators in a special balance, in a humid and gaseous confined atmosphere. Stress is (objectively) defined as a disruption of homeostasis resulting from a constraint escaping the usual flexibility of metabolism. It induces another temporary (reversible) or definitive (irreversible) thermodynamic physiological state. The state-change concept developed by Strasser (1988) and Strasser and Tsimilli-Michael (2001) is applicable to the phenomenon of hyperhydricity. An appraisal of the redox capacities of hyperhydrated shoots together with a study of some enzymic activities that catalyse pentose phosphate and glycolytic pathways has indeed shown that such shoots have evolved towards a temporary state of lower differentiation or a juvenile state with a sufficient activity to survive and to defend themselves. [less ▲]

Equine trypsin: purification and development of a radio-immunoassay

in Veterinary Research (2003), 34(3, May-Jun), 317-330

Shock is accompanied by generalised splanchnic hypoperfusion, and splanchnic organs like the pancreas can be damaged, as shown in animal experimental models and in humans, by the presence of high plasma ... [more ▼]

Shock is accompanied by generalised splanchnic hypoperfusion, and splanchnic organs like the pancreas can be damaged, as shown in animal experimental models and in humans, by the presence of high plasma concentrations of trypsin and other pancreatic enzymes. In order to design a radioimmunoassay technique (RIA) for the measurement of equine trypsin-like immunoreactivity (TLI) in biological fluids, trypsin was purified (with purity greater than or equal to 96 %) from the equine pancreas by extraction in an acid medium, ammonium sulfate precipitations, gel filtration chromatography and, after activation of trypsinogen into trypsin, affinity chromatography. Gel polyacrylamide electrophoresis showed a monomeric enzyme with a molecular weight of 27 kDa. The purified equine trypsin served for the immunisation of rabbits in order to obtain a specific antiserum, and the labelled antigen was prepared by iodination of equine trypsin with I-125. The RIA was based on the binding of the antigen to the antibody followed by the separation of the antigen-antibody complex by immunoprecipitation in the presence of sheep anti-rabbit gammaglobulins and the assay of the radioactivity in the precipitate. The RIA showed good sensitivity, specificity, precision, accuracy and reproducibility. The reference mean value of TLI in the plasma of healthy horses (n = 20) was 30.01 +/- 6.84 ng/mL (upper confidence limit 50.52 ng/mL; p < 0.01). Three horses with non strangulating intestinal obstruction without shock showed TLI values within normal limits whereas 5 of 7 horses with strangulation obstruction showed TLI levels above the upper confidence limit. Further studies using the RIA and the enzymatic assay should be performed in order to confirm the role of the pancreas in equine intestinal obstruction. [less ▲]

Concepts in plant stress physiology. Application to plant tissue cultures

in Plant Growth Regulation (2002), 37(3), 263-285

Because the term stress is used, most often subjectively, with various meanings, this paper first attempts to clarify the physiological definition, and the appropriate terms as responses in different ... [more ▼]

Because the term stress is used, most often subjectively, with various meanings, this paper first attempts to clarify the physiological definition, and the appropriate terms as responses in different situations. The flexibility of normal metabolism allows the development of responses to environmental changes which fluctuate regularly and predictably over daily and seasonal cycles. Thus every deviation of a factor from its optimum does not necessarily result in stress. Stress begins with a constraint or with highly unpredictable fluctuations imposed on regular metabolic patterns that cause bodily injury, disease, or aberrant physiology. Stress is the altered physiological condition caused by factors that tend to alter an equilibrium. Strain is any physical and/or chemical change produced by a stress, i.e. every established condition, which forces a system away from its thermodynamic optimal state. The paper secondly summarises the Strasser's state-change concept which is precisely that suboptimality is the driving force for acclimation (genotype level) or adaptation (population level) to stress. The paper continues with the actual knowledge on the mechanisms of stress recognition and cell signalling. Briefly: plasma membranes are the sensors of environmental changes; phytohormones and second messengers are the transducers of information from membranes to metabolism; carbon balance is the master integrator of plant response; betwixt and between, some genes are expressed more strongly, whereas others are repressed. Reactive oxygen species play key roles in up- and down-regulation of metabolism and structure. The paper shows finally that the above concepts can be applied to plant tissue cultures where the accumulating physiological and genetical deviations (from a normal plant behaviour) are related to the stressing conditions of the in vitro culture media and of the confined environment. The hyperhydrated state of shoots and the cancerous state of cells, both induced under conditions of stress in in vitro cultures, are identified and detailed, because they perfectly illustrate the stress-induced state-change concept. It is concluded that stress responses include either pathologies or adaptive advantages. Stress may thus contain both destructive and constructive elements: it is a selection factor as well as a driving force for improved resistance and adaptive evolution. [less ▲]