Identification of protein biomarkers associated with cardiac ischemia by a proteomic approach.

Angina is chest pain induced by ischemia of the heart muscle, generally due to obstruction or spasm of the coronary arteries. People that suffer from average to severe cases of angina have an increased percentage of death before the age of 55, usually around 60%. Therefore, prevention of major complications, optimizing diagnosis, prognosis and therapeutics are of primary importance. The main objective of this study was to uncover biomarkers by comparing serum protein profiles of patients suffering from stable or unstable angina and controls. We identified by non-targeted proteomic approach and confirmed by the means of independent techniques, the differential expression of several proteins indicating significantly increased vascular inflammation response, disturbance in the lipid metabolism and in atherogenic plaques stability.

[Rhupus: when rheumatoid arthritis meets lupus]. / Le rhupus: à la frontière entre polyarthrite rhumatoïde et lupus érythémateux disséminé.

There exists diseases in rheumatology fulfilling classification criteria for either rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). They are called "rhupus". We retrospectively analyzed the data base "GLIMS" of the CHU de Liège from the starting date of november 2005 until april 2011 to identified those patients that were positive for the anti-sDNA antibody marker of SLE and for the anti-CCP antibody, marker of RA. Fourteen patients were identified and two other patients were added, one suffering from SLE, and the other from RA, and likely to be rhupus. Of the 16 patients analyzed, 9 were real RA with anti-dsDNA antibodies induced by anti-TNF-alpha therapies. Seven were candidates to be rhupus and 6 were retained. They were all women, with a median age of 51 years and in addition were all anti-SS-A antibody positive.

[BNP and NT-proBNP: reference values and cutoff limits]. / BNP et NT-proBNP: valeurs de référence et seuils décisionnels.

Natriuretic peptides, particularly BNP and NT-proBNP, are increasingly used as screening test in patients with symptoms suggestive of heart failure (HF). Due to their high negative predictive values, natriuretic peptide determinations allow to exclude chronic HF with great certainty and to identify patients for whom echography is not necessary. These biomarkers are also useful for diagnostic purposes, high plasma levels being related to an increased risk of cardiovascular hospitalisation and death. Risk stratification in patients with HF symptoms is based on "low" and "high" cut-off limits, for which different values have been proposed. The aim of this paper is to discuss the delineation of the decision limits and the intermediate grey zone in comparison to NT-proBNP reference values obtained in a representative group of subjects living in the Liège area (Belgium). Data were analysed in relation to age and gender, two of the main parameters influencing the natriuretic peptide plasma levels.

Seasonal variations in vitamin D levels in melanoma patients: a single-centre prospective pilot comparative study.

BACKGROUND: More than 90% of vitamin D synthesis is dependent on UV exposure. Photosensitive disorders such as lupus erythematosus, protoporphyria and xeroderma require strict sun avoidance, and vitamin D deficiency has been demonstrated in these patients. Melanoma patients are also instructed to avoid sun exposure and may hence be expected to be vitamin D deficient. MATERIALS AND METHODS: Winter and summer vitamin D levels were compared in a group of melanoma patients (n =61) and age- and phototype-matched controls (n = 53) without photosensitive disorders. RESULTS: Oral supplementary vitamin D intake was reported in 32.7% of the melanoma patients and in 15.1% in the control group. Despite oral supplementation, only 25% of the melanoma patients and the controls presented with vitamin D levels of 30 ng/mL or higher. In non-supplemented subjects in the melanoma and control groups, respectively, mean winter vitamin D levels were below the recommended threshold at 12.6 ng/mL vs. 13.2 ng/mL, respectively, but not statistically different. These values increased significantly in both groups during the summer to 24.6 and 23.8 ng/mL respectively. CONCLUSION: Unexpected, significant increases in vitamin D levels were seen in melanoma patients during summer, suggesting non-adherence with photoprotective measures and reflecting a heliophilic behaviour. Vitamin D supplementation is recommended in melanoma patients during both winter and summer.

Estimate of total salt intake in two regions of Belgium through analysis of sodium in 24-h urine samples.

OBJECTIVES: To evaluate total salt intake in the adult population through an analysis of sodium in 24-h urine samples in two regions of Belgium. METHODS: Urine samples were collected over 24 h from participants and they had to complete a specific questionnaire about salt intake afterwards. Sodium and creatinine concentrations were analysed in these samples. SUBJECTS: The target population comprised adults aged 45-65 years in the region of Ghent and Liege. A total of 123 and 157 volunteers from Ghent and Liege, respectively, were included in the study. RESULTS: The mean creatinine level in Flanders (n=114) amounted to 0.173±0.035 mmol/kg/day, whereas in the Walloon region (n=135) it amounted to 0.161±0.036 mmol/kg/day, after the exclusion of subjects with incomplete urine collection. Intake of sodium in Flanders (n=114) was 4.29±1.29 g/day, whereas in the Walloon region (n=135) it was 3.94±1.44 g/day. In both regions, sodium intake in men was higher than in women. CONCLUSION: Salt intake was more or less twice as high as the recommended intake. Salt intake as estimated from 24-h urine collections is substantially higher than that previously calculated on the basis of food consumption data. A salt reduction programme for Belgium is primordial.

Measurement uncertainty of 25-OH vitamin D determination with different commercially available kits: impact on the clinical cut offs.

SUMMARY: Due to "measurement uncertainty", the "true" 25-OH vitamin D (25(OH)D) of a patient (whatever the commercially available assay tested) will be >80 nmol/L if its measured concentration is >100 nmol/L. Thus, if a physician considers that a normal VTD status is a 25(OH)D level >or=80 nmol/L, he should ensure that the patient's results are >or=100 nmol/L. INTRODUCTION: Many experts recommend that serum levels of 25(OH)D should be above a lower normal limit of 75-80 nmol/L. However, the value delivered by laboratories is only an estimation of the "true" value due to "measurement uncertainty." When using a cut off, measurement uncertainty around the cut off is important because therapeutic actions may differ if the measured value is below or above the limit. We aimed to establish the "measurement uncertainty" at different levels of concentration for several commercially available 25(OH)D analytical techniques. METHODS: We constituted three pools of serum with different 25(OH)D concentrations. Each pool was assayed in triplicate during 5 days with the DiaSorin RIA, Liaison, Elecsys, and Chromsystems-HPLC assays. RESULTS: We report a relatively high "measurement uncertainty" for the measurement of 25(OH)D for the four different techniques: the mean relative uncertainties, all techniques confounded were 19.4%, 16.0%, and 11.3% for pool 1 (35.3 nmol/L), pool 2 (79.5 nmol/L), and pool 3 (126.1 nmol/L), respectively. CONCLUSIONS: Our results show that, whatever the assay, the "true" 25(OH)D of a patient will be >80 nmol/L if its measured concentration is >100 nmol/L. In other words, if a physician considers that a normal VTD status is defined by a 25(OH)D level >or=80 nmol/L, he should ensure that the patients present a 25(OH)D >or=100 nmol/L.

[Propositions for the standardized expression of HbA 1c results]. / Propositions pour l'expression standardisée des résultats d'HbA 1c.

The 2007 international consensus about the standardization of HbA(1c) determination and expression of results is progressively implemented in most countries. In France, a common working group of the Société française de biologie clinique (SFBC) and the Société francophone de diabétologie (SFD) has expressed the following recommendations. HbA(1c) results are expressed in percentage of total hemoglobin and in mmol HbA(1c)/mol Hb, but are not converted into estimated average glucose. A table indicating the correspondence between HbA(1c) and estimated average glucose may be given with the results, subject to precautions of interpretation at the individual level.

[Production of monoclonal antibodies]. / Production des anticorps monoclonaux.

In contrast to a polyclonal antiserum, a monoclonal antibody is specific to a single epitope on the surface of a complex antigen. In 1975, Kohler and Milstein produced the first monoclonal antibodies by using a method which rapidly became a key technology in immunology. By fusing activated antibody-forming cells (B cells) with myeloma cells, they obtained hybrid cells--the so-called hydridomas--which combine the ability of the activated B cells to secrete a single species of antibody and the immortality of the myeloma cell. The selected hybridomas proliferate continuously, their clonal progeny providing an unending supply of antibody with a single specificity. These antibodies have found many applications in basic research and in vitro diagnosis. In the clinical laboratory, monoclonal antibodies are used as reagents in immunoassays, often replacing traditional antisera. Many years of development and innovation were needed to humanize monoclonal antibodies in order to make them usable in human therapy.

[Interest of monoclonal antibodies in biomedical laboratory analysis]. / Intérêt des anticorps monoclonaux dans le laboratoire d'analyses biomédicales.

Immunoassays, or assays with antibodies as reagents, are widely used in medical laboratories. These assays are used to identify and quantify various substances in biological fluids, such as specific proteins (various tissue markers, markers of inflammation, hormones, coagulation factors...) or immunoglobulins (viral or bacterial antibodies, auto-antibodies...) and even both viral antigens and antibodies (HIV virology). The use of monoclonal antibodies allowed, through their specificity for a single epitope of the target molecule, the development of increasingly sophisticated immunoassays. In particular, the use of monoclonal antibodies with microarrays permits the simultaneous determination of various proteins (inflammatory profile, cardiac profile, specifics IgE...) quickly and accurately. Very important tools in the clinical laboratory, immunoassays techniques are, however, subject to various analytical interferences which may be responsible for significant changes in the test results.

Estimation of GFR by different creatinine- and cystatin-C-based equations in anorexia nervosa.

BACKGROUND: Patients with anorexia nervosa (AN) are at high risk of renal failure. Glomerular filtration rate (GFR) is overestimated when estimated by the creatinine-based equations. We have studied the accuracy and precision of cystatin C-based equations. METHOD: 27 AN patients were included. GFR was measured with the chromium-51-ethylenediaminetetraacetate (51Cr-EDTA) method. We have compared the accuracy and precision of creatinine-based equations (MDRD and Cockcroft) with those of different new cystatin C-based equations. RESULTS: The creatinine-based equations overestimate measured GFR, especially the MDRD study equation. All the cystatin C-based equations also overestimate measured GFR. The Cockcroft and Gault formula and the cystatin C-based equation published by Rule have the best accuracy and precision, but these last performances remain unsatisfactory. CONCLUSION: Both creatinine and cystatin C-based equations strongly overestimate measured in patients with AN.

Matrix Metalloproteinase-9 gene induction by a truncated oncogenic NF-kappaB2 protein involves the recruitment of MLL1 and MLL2 H3K4 histone methyltransferase complexes.

Constitutive nuclear factor (NF)-kappaB activation in haematological malignancies is caused in several cases by loss of function mutations within the coding sequence of NF-kappaB inhibitory molecules such as IkappaBalpha or p100. Hut-78, a truncated form of p100, constitutively generates p52 and contributes to the development of T-cell lymphomas but the molecular mechanism underlying this oncogenic potential remains unclear. We show here that MMP9 gene expression is induced through the alternative NF-kappaB-activating pathway in fibroblasts and also on Hut-78 or p52 overexpression in fibroblasts as well as in lymphoma cells. p52 is critical for Hut-78-mediated MMP9 gene induction as a Hut-78 mutant as well as other truncated NF-kappaB2 proteins that are not processed into p52 failed to induce the expression of this metalloproteinase. Conversely, MMP9 gene expression is impaired in p52-depleted HUT-78 cells. Interestingly, MLL1 and MLL2 H3K4 methyltransferase complexes are tethered by p52 on the MMP9 but not on the IkappaBalpha promoter, and the H3K4 trimethyltransferase activity recruited on the MMP9 promoter is impaired in p52-depleted HUT-78 cells. Moreover, MLL1 and MLL2 are associated with Hut-78 in a native chromatin-enriched extract. Thus, we identified a molecular mechanism by which the recruitment of a H3K4 histone methyltransferase complex on the promoter of a NF-kappaB-dependent gene induces its expression and potentially the invasive potential of lymphoma cells harbouring constitutive activity of the alternative NF-kappaB-activating pathway.

[Hymenoptera venom anaphylaxis and in-vitro diagnostic tools: new approaches to manage patients at high risk of a severe reaction]. / Allergie au venin d'hyménoptère et tests de laboratoires: de nouvelles approches pour la prise en charge des patients a haut risque de réaction sévère.

The consequences of Hymenoptera venom anaphylaxis are very severe but it is not obvious to predict which reactions will occur in one single patient when he is stung for the second time. For a couple of years, many new laboratory tests have been experimented and many studies published. CAST, BAT, WB, tryptase, sIgE and sIgG4 are the new valuable additional diagnostic tools that can help the decision to perform an immunotherapy or to discontinue this therapy after 3 years. The aim of our study was to determine which could be the profile of a desensitized patient and to screen for good candidates for venom immunotherapy.

[Plasma calcium in haemodialysis patients: total calcium or ionized calcium? Should we systematically provide a value of total corrected calcium on our protocols?]. / Calcium chez les patients hémodialysés : calcémie totale ou calcium ionisé ? Le laboratoire doit-il systématiquement fournir au clinicien une valeur de calcémie totale corrigée obtenue par calcul ?

Ionized calcium is the only physiologically active form of calcium. Because of the variation of albumin, pH and haemoconcentration observed during haemodialysis session in patients with chronic renal failure, measure of total calcium does not reflect the real variation of ionized calcium. However, many formulae to correct total calcium by albumin have been proposed but none of them has been validated in dialysis patients. At present time, computing progress permit laboratory to systematically provide a value of corrected total calcium on protocols but is it really indicated? Our results showed that any of those formulae allows obtaining a value of total calcium that possesses a significant critical difference in relation to total calcium. Thus, correction formulae must be abandoned in aid of ionized calcium in haemodialysis patients.

[Vitamin D deficiency in recently pregnant women]. / La carence en vitamine d chez les femmes enceintes en région liégeoise: un problème méconnu.

We have evaluated the prevalence of the 25-hydroxy vitamin D (25VTD) deficiency in recently pregnant women and new mothers in the area of Liege, Belgium. The study took place in November 2006. Twenty four women who underwent a positive pregnancy test and 65 new mothers were enrolled. The level of 25VTD did not differ between the two groups. Only 12% of the pregnant women and 14% of the new mothers (>12 ng/ml) had an optimal level of 25VTD (>30 ng/ ml). We also observed a severe 25VTD deficiency in 21% of pregnant women and 32% of new mothers. Our results showed that more than 80% of pregnant women and new mothers in the area of Liege presented a deficiency in 25VTD. In Belgium, daily vitamin supplementation of pregnant women is common, but the level of vitamin D3 concentration range from 10 microg (400 UI) to zero microg. In our area, vitamin D production in the skin is not always important enough to achieve optimal levels. Our data show that vitamin D supplementation of pregnant women is not enough and that 25VTD deficiency is not diagnosed in this high-risk population. Children born from deficient mothers will present a higher risk of suffering from bone mineral diseases as well as other pathologies, as type 1 diabetes or neurological disorders. Of course, this insufficiency will also have an impact on mother's bone reserve, but these mothers will also be at higher risk for preeclampsia.

[Vitamin D2 or vitamin D3?]. / Vitamine D2 ou vitamine D3?

PURPOSE: Nearly one billion people around the world are deficient in vitamin D and need to be supplemented. Vitamin D is available in medicines and fortified foods. It is available in two forms: vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). KEY POINTS: The pharmacopeiae consider these steroid hormones as equivalent and interchangeable. However, several studies have showed that serum level of 25(OH)D is increased more effectively with vitamin D3 than vitamin D2. Vitamin D2 has shorter plasma half-life and a lower affinity for the vitamin D binding protein, the hepatic vitamin D hydroxylase and the vitamin D receptor. CONCLUSION: Vitamin D2 should not be regarded anymore as suitable for supplementation or fortification. Currently though, it is still the most used in some countries such as Portugal and Australia.

[How I explore ... review of the principals of antibodies]. / Comment j'explore ... revue des principaux auto-anticorps.

Auto-immune diseases represent the 3rd cause of morbidity after cardiovascular and oncologic diseases. They often occur in young subjects. Their presence is not synonymous of disease and must be associated to clinical signs to be pathological. However, their discovery can require a complement of investigations and the possibility of a follow-up because some auto-antibodies are predictive of disease. This paper is concerned with the main autoantibodies that can be picked out at the laboratory of immunology. Some technical explanations and INAMI rules are explained too.

[Oxidative stress]. / Le stress oxydant.

Oxidative stress is defined as an imbalance between the production of reactive oxygen species (ROS) and the antioxidant network, in favour of the former. Our lifestyle (smoking, alcoholism, obesity, intense physical exercise), but also our inadequate diet, contributes to significantly increase the production of ROS in our organism. This is potentially associated with an increased risk of developing ageing-related pathologies such as cardiovascular diseases and cancer. As a matter of prevention, it is necessary to have in hands a high technology allowing to correctly evidence the oxidative stress status of an individual in order to render optimal our antioxidant defences and to decrease the oxidative damages in DNA, proteins and lipids.

[Multiparametric evaluation of cardiac risk in acute coronary syndromes: value of biological blood markers]. / Evaluation multiparamétrique du risque cardiaque dans les syndromes coronariens aigus: intérêt des marqueurs biologiques sanguins.

Appropriate evaluation of short- and medium-term cardiovascular risk in acute coronary syndromes (ACS) patients should allow to individualize and improve treatment in the future. Natriuretic peptides can be measured in plasma using reliable and fast methods. B-type natriuretic peptide (BNP) is released into the circulation by the cardiac ventricle in response to increases in wall stress. Both BNP and NT-proBNP have been shown to aid in the diagnosis of heart failure. In addition, these peptides correlate with left ventricular dilatation, remodelling and dysfunction, and have the capacity of improving significantly the early risk stratification in patients with ST- or non-ST-elevation ACS. BNP determinations can be associated to cardiac troponins (for example, cTnT) and C-reactive protein (CRP). Plasma concentrations of these biomarkers are related to different physiopathological mechanisms: tissue necrosis (cTnT), neuro-hormonal activation (BNP, NT-proBNP) or inflammatory process and plaque instability (CRP). Combining two or three of these biomarkers for better risk stratification has the potential to improve substantially the outcomes in patients with ACS.

[Comparison of five techniques to detect anti-Saccharomyces cerevisiae antibodies (ASCA) in serum for diagnosing Crohn's disease]. / Comparaison de cinq techniques de détection des anticorps anti-Saccharomyces cerevisiae (ASCA) dans le sérum pour le diagnostic de la maladie de Crohn.

OBJECTIVE: the anti-Saccharomyces cerevisiae antibodies (ASCA) are diagnostic markers found in Crohn's disease patients. The aim of this study was to compare three Elisa (enzyme linked immunosorbent assay) kits with the indirect immunofluorescence (IFI) technique and an immunodot for ASCA detection. MATERIALS AND METHODS: we compared the results obtained using IFI (IgA and IgG) and Elisa (IgA and IgG) in 139 patients (37 Crohn's disease). An immunodot (IgA+IgG) was tested in a sub-group of 24 patients (18 Crohn's disease). RESULTS AND DISCUSSION: for the different techniques by Elisa (IgA or IgG), the sensitivity ranged from 65% to 76%, the specificity from 88% to 98%, the positive predictive value (PPV) from 84% to 94% and the negative predictive value (NPV) from 88% to 93%. For IFI, the sensitivity was 81%, the specificity 100%, the PPV 100% and the NPV 93%. The immunodot showed a specificity and PPV of 100% and NPV of 33%. CONCLUSION: the detection of the ASCA is useful in the diagnosis of Crohn's disease. IFI appears as the method of choice for its excellent sensitivity and specificity, and affordable costs.