Myeloperoxidase and Prostate volume: A preliminary study.

OBJECTIVES: Oxidative stress is associated with the development of BPH and might be modulated by several factors. Myeloperoxidase (MPO) has recently been observed in prostate tissue. Our goal was to investigate the correlation between MPO and the prostate volume. MATERIAL AND METHODS: Hundred and twenty-one patients (48-70 years) with a filled IPSS were prospectively included. Blood sampling (PSA, testosterone, Angiotensin II (AngII), MPO, Mox-LDL) and transrectal ultrasound of the prostate were performed with total volume (TV) and transitional zone volume (TZ) measurements. For correlation, univariate analyses were depicted by Pearson's coefficient. Multilinear regression analysis used a stepwise backward selection of the explicative variables. RESULTS: In multivariate analysis, the TV was positively correlated to the combination of age and Ang II but negatively to MPO specific activity (Std Coef=-0.272, P=0.004). Significant correlations were confirmed between TZ, age and MPO specific activity but not with Ang II. A negative correlation between TZ and MPO specific activity was also observed (Std Coef=-0.21, P=0.016). No correlation was found with Mox-LDL. CONCLUSIONS: Negative correlation between MPO and prostate volume was observed but careful interpretations may be endorsed and longitudinal study is necessary. It seems relevant to focus on the potential contribution of MPO in the development of prostatic diseases as this enzyme can also promote DNA oxidation. LEVEL OF EVIDENCE: 4.

An immunological method to combine the measurement of active and total myeloperoxidase on the same biological fluid, and its application in finding inhibitors which interact directly with the enzyme.

Myeloperoxidase (MPO) is a pro-oxidant enzyme involved in inflammation, and the measurement of its activity in biological samples has emerged essential for laboratory and clinical investigations. We will describe a new method which combines the SIEFED (specific immunological extraction followed by enzymatic detection) and ELISA (ELISAcb) techniques to measure the active and total amounts of MPO on the same human sample and with the same calibration curve, as well as to define an accurate ratio between both the active and total forms of the enzyme. The SIEFED/ELISAcb method consists of the MPO extraction from aqueous or biological samples by immobilized anti-MPO antibodies coated onto microplate wells. After a washing step to eliminate unbound material, the activity of MPO is measured in situ by adding a reaction solution (SIEFED). Following aspiration of the reaction solution, a secondary anti-MPO antibody is added into the wells and the ELISAcb test is carried out in order to measure the total MPO content. To validate the combined method, a comparison was made with SIEFED and ELISA experiments performed separately on plasma samples isolated from human whole blood, after a neutrophil stimulation. The SIEFED/ELISAcb provides a suitable tool for the measurement of specific MPO activity in biological fluids and for the estimation of the inhibitory potential of a fluid. The method can also be used as a pharmacological tool to make the distinction between a catalytic inhibitor, which binds to MPO and inhibits its activity, and a steric inhibitor, which hinders the enzyme and prevents its immunodetection.

Rational drug design applied to myeloperoxidase inhibition.

Rational drug design is a general approach using protein-structure technique in which the discovery of a ligand can be driven either by chance, screening, or rational theory. Myeloperoxidase (MPO) was rapidly identified as a therapeutical target because of its involvement in chronic inflammatory syndromes. In this context, the research of MPO inhibitors was intensified and development of new chemical entities was rationally driven by the research of ligands that enter into the MPO catalytic pocket. Actually, as soon as crystallography data of MPO have become available and its structure was virtually designed, the rational drug design has been applied to this peroxidase. Pharmaceutical industries and academic laboratories apply rational drug design on MPO by either optimizing known inhibitors or searching new molecules by high-throughput virtual screening. By these ways, they were able to find efficient MPO inhibitors and understand their interactions with the enzyme. During this quest of MPO inhibition, it appears that Glu268 is a crucial residue in order to optimize ligand-target interaction. This amino acid should be carefully considered by medicinal chemist when they design inhibitors interfering with MPO activity.

Myeloperoxidase and its products in synovial fluid of patients with treated or untreated rheumatoid arthritis.

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.

Intriguing location of myeloperoxidase in the prostate: a preliminary immunohistochemical study.

BACKGROUND: Myeloperoxidase (MPO) is a member of the peroxidase-cyclooxygenase superfamily, which is secreted from stimulated leucocytes at inflammatory sites. It is well known that MPO catalyses oxidation reactions via the release of reactive halogenating and nitrating species and thus induces tissue damage. Several studies have already implicated MPO in the development of neoplasia. Chronic or recurrent prostatic inflammation has long been recognized as having the potential to initiate and promote the development of prostate cancer. The objective was to investigate whether MPO is present in the prostate. METHODS: Human prostate material was obtained from biopsies, transurethral resections of the prostate (TURP), prostatic adenomectomies, and retropubic radical prostatectomies. Twenty-nine slides of normal prostate tissue, benign prostatic hyperplasia (BPH), and prostate cancer were reviewed by a pathologist. Immunohistochemical analysis using MPO-specific human antibody was performed to detect MPO in the prostate tissue. RESULTS: Immunocytohistochemistry showed cellular colocalization of MPO in the secretory epithelial cells of the prostate with staining varying from light to strong intensity. Staining in the glandular apical snouts was often reinforced although staining of basal as well as of luminal glandular cells was also present. CONCLUSIONS: We identified, for the first time, the presence of MPO at the surface of prostatic epithelial cells. In view of the pro-oxidant properties of this enzyme, further research is needed to define whether MPO contributes to the development of prostatic lesions.

A new easy method for specific measurement of active myeloperoxidase in human biological fluids and tissue extracts.

The SIEFED ("Specific Immunological Extraction Followed by Enzymatic Detection") method already developed for the specific detection of the activity of equine myeloperoxidase (MPO) was adapted for the specific measurement of active human MPO in biological fluids or tissue extracts. The method consists of the extraction of MPO from aqueous solutions by immobilized anti-MPO antibodies followed by a washing (to eliminate the extraction medium and the biological fluid with their possible interfering molecules) and the measurement of the activity of MPO with a detection system containing a fluorogenic substrate, H(2)O(2) and nitrite ions as reaction enhancer. The SIEFED was applied to study active MPO in human biological fluids (plasma, bronchoalveolar lavage fluid and supernatant from carotids extracts). The SIEFED for human MPO has a sensitivity limit of 0.080 mU/mL and showed good precision with intra- and inter-assay coefficients of variation below 10 and 20% respectively within a broad range of MPO activities establish from 0.156 to 473 mU/mL. The SIEFED for human MPO will be useful for the specific detection of active MPO in complex fluids and can be complementary to an ELISA to determine an active/total MPO ratio in healthy volunteers and patients especially in case of chronic or acute inflammatory diseases.

The reactions of oxicam and sulfoanilide non steroidal anti-inflammatory drugs with hypochlorous acid: determination of the rate constants with an assay based on the competition with para-aminobenzoic acid chlorination and identification of some oxidation products.

Hypochlorous acid (HOCl) is an oxygen-derived species involved in physiological processes related to the defence of the organism that may cause adverse effects when its production is insufficiently controlled. In order to examine its reactivity with potential scavenging molecules from the non steroidal anti-inflammatory drugs (NSAIDs) family, a competition assay based on para-aminobenzoic acid (PABA) chlorination was developed. The original optimised in vitro fluorimetric procedure offered the possibility to determine rate constants (ks) for the reaction with HOCl in physiologically relevant conditions. The specificity of the system was improved by a liquid chromatography (LC) which allows the separation of the drugs and their oxidation products. After determination of the rate constant for PABA chlorination by HOCl (mean +/- SD in M(-1) s(-1): 4.3 +/- 0.3 x 10(3)), the applied mathematical model for a chemical competition permits to obtain linear curves from competition studies between several NSAIDs and PABA. Their slopes provided the following rate constants for the different studied drugs: tenoxicam: 4.0 +/- 0.7 x 10(3), piroxicam: 3.6 +/- 0.7 x 10(3), lornoxicam: 4.3 +/- 0.7 x 10(3), meloxicam: 1.7 +/- 0.3 x 10(4), nimesulide: 2.3 +/- 0.6 x 10(2). Meloxicam therefore reacted significantly faster than the other oxicams and nimesulide, which is the weakest scavenger of the studied series. The identification of some of the oxidation products by NMR or MS permitted to explore the reaction mechanism and to examine some aspects of the structure/activity relationships for the molecules of the same chemical family.