An objective reduction technique of proteomic mass spectra based on multi-scale fuzzy thresholding

A proteomic approach offers a powerful and complementary tool to genomics. It allows to index and characterize proteins, and, for example, to compare their levels of expression between healthy and pathological states. Proteomic analyses are mainly based on the separation of proteins by two-dimensional gel electrophoresis and their subsequent identification by comparing the data from Mass Spectrometry (SM) analyses to the theoretical ones contained in databases. In mass spectrometry, the detector noise, the electronic and chemical noise, sometimes the small amount of peptides that has to be treated and finally the spectrum reduction noise (due to bad filtering and/or thresholding), can induce Parasitic Mass Peaks (PMP) and/or hide some Useful Mass Peaks (UMP) of low intensities. The immediate consequence is that the presence of the PMP and the absence of the UMP will be detrimental to the protein identification quality. In this article, we propose an original algorithm eliminating the PMP, detecting and amplifying those which are useful. The preprocessing principle uses a multi-scale analysis technique coupled to a fuzzy thresholding (multi-scale fuzzy thresholding), a local amplification of the UMP, and finally an adaptive Base Line Correction. The associated frequencies with the PMP are distributed on all the spectrum pass bandwidth. This leads us to a dyadic tree structure subband decomposition. The algorithm principle consists of dividing the frequential pass bandwidth of each masses spectrum into two subbands, a Low and High Frequency (LF,HF) subband, then each subband is in turn divided into two subbands etc. The HF subbands are then thresholded according to the minimization criterion of the Shannon fuzzy entropy, and then amplified locally; the base line is calculated in an adaptive way and subtracted from reconstructed spectrum. To evaluate the quality of this algorithm, we present a comparison of the results obtained by our algorithm, and those obtained by the DataExplorer software. The latter is a reduction software provided within the MALDI-TOF spectrometer software package.La protéomique offre une approche puissante et complémentaire à la génomique. Elle permet de répertorier et caractériser les protéines, de comparer leur niveau d’expression entre un état physiologique sain et malade par exemple. L’analyse protéomique se fait essentiellement par l’utilisation de la technique d’électrophorèse bidimensionnelle couplée à la technique d’analyse par Spectrométrie de Masse (SM). La première, aidée par l’imagerie protéomique, conduit à la localisation des protéines candidates à une analyse par SM. La comparaison des spectres de masses obtenus à des bases de données protéiques, conduit à l’identification des protéines d’intérêt en terme de peptides. Le problème qui se pose souvent est que les spectres sont bruités et pauvres en masses. En effet, le bruit du détecteur, le bruit électronique et chimique, la présence de peu de matériel protéique et enfin le bruit de la réduction des spectres (mauvais filtrage et/ou seuillage), tous ces bruits peuvent induire des Pics de Masses Parasites (PMP) et/ou supprimer des Pics de Masses Utiles (PMU) de faible intensité. La conséquence immédiate est que la présence des PMP et l’absence des PMU seront utilisées au dépens de la qualité d’identification de la protéine. Dans cet article, nous proposons un algorithme original éliminant les PMP, détectant et amplifiant ceux utiles. Le principe du pré-traitement utilise une Analyse Multirésolution (AM) couplée à un seuillage basé sur la logique floue (seuillage flou multi-échelle), une amplification locale des PMU, et enfin une correction adaptative de la Ligne de Base (LB). Les fréquences associées aux PMP sont réparties sur toute la bande passante du spectre, ce qui nous conduit à une AM dite en arbre. Le principe consiste à découper la bande passante fréquentielle de chaque spectre de masses en deux sous-bandes, une Basse Fréquence (BF), l’autre Haute Fréquence (HF), ensuite chaque sous-bande est à son tour découpée en deux sous-bandes etc. Les sous-bandes HF sont seuillées selon le critère de minimisation de l’entropie floue de Shannon et amplifiées localement, la ligne de base est calculée automatiquement et soustraite du spectre reconstruit. Pour évaluer la qualité de cet algorithme, nous présentons une comparaison des résultats obtenus par notre algorithme, et ceux fournis par le spectromètre MALDI-TOF (Matrix Assisted Laser Desorption/Ionisation-Time Of Flight), qui utilise le logiciel « DataExplorer » comme logiciel de réduction

Variability of diagnostic criteria and treatment of idiopathic nephrotic syndrome across European countries

The aim of the surveys conducted by the Idiopathic Nephrotic Syndrome Working Group of the ESPN was to study the possible variability of treatment in Europe at different stages of the disease by means of questionnaires sent to members of the Working Group. Four surveys have been completed: treatment of the first flare, treatment of the first relapse and the issue of steroid dependency, use of rituximab, and the management of steroid-resistant patients. A uniform treatment of the first flare was applied in only three countries, and ten additional centers have adopted one of the three main protocols. Reported treatment of the first relapse was relatively uniform, whereas the use of additional immunosuppressants in steroid dependency was widely variable. Rituximab had already been used in hundreds of patients, although the formal evidence of efficiency in steroid dependency was relatively recent at the time of the survey. The definition of steroid resistance was variable in the European centers, but strikingly, the first-line treatment was uniform throughout the centers and included the combination of prednisone plus calcineurin antagonists. Conclusion: The variability in the approach of idiopathic nephrotic syndrome is unexpectedly large and affects treatment of the first flare, strategies in the case of steroid dependency, as well as the definitions of steroid resistance.What is Known:• Steroids and immunosuppressants are the universal treatment of idiopathic nephrotic syndrome.What is New:• The variability of treatments and strategy of treatment in European centers of pediatric nephrology

Guidelines for diagnosis and management of the cobalamin-related remethylation disorders cblC, cblD, cblE, cblF, cblG, cblJ and MTHFR deficiency

BACKGROUND: Remethylation defects are rare inherited disorders in which impaired remethylation of homocysteine to methionine leads to accumulation of homocysteine and perturbation of numerous methylation reactions. OBJECTIVE: To summarise clinical and biochemical characteristics of these severe disorders and to provide guidelines on diagnosis and management. DATA SOURCES: Review, evaluation and discussion of the medical literature (Medline, Cochrane databases) by a panel of experts on these rare diseases following the GRADE approach. KEY RECOMMENDATIONS: We strongly recommend measuring plasma total homocysteine in any patient presenting with the combination of neurological and/or visual and/or haematological symptoms, subacute spinal cord degeneration, atypical haemolytic uraemic syndrome or unexplained vascular thrombosis. We strongly recommend to initiate treatment with parenteral hydroxocobalamin without delay in any suspected remethylation disorder; it significantly improves survival and incidence of severe complications. We strongly recommend betaine treatment in individuals with MTHFR deficiency; it improves the outcome and prevents disease when given early

Further Insights into Brevetoxin Metabolism by de Novo Radiolabeling,

International audienceThe toxic dinoflagellate Karenia brevis, responsible for early harmful algal blooms in the Gulf of Mexico, produces many secondary metabolites, including potent neurotoxins called brevetoxins (PbTx). These compounds have been identified as toxic agents for humans, and they are also responsible for the deaths of several marine organisms. The overall biosynthesis of these highly complex metabolites has not been fully ascertained, even if there is little doubt on a polyketide origin. In addition to gaining some insights into the metabolic events involved in the biosynthesis of these compounds, feeding studies with labeled precursors helps to discriminate between the de novo biosynthesis of toxins and conversion of stored intermediates into final toxic products in the response to environmental stresses. In this context, the use of radiolabeled precursors is well suited as it allows working with the highest sensitive techniques and consequently with a minor amount of cultured dinoflagellates. We were then able to incorporate [U-14C]-acetate, the renowned precursor of the polyketide pathway, in several PbTx produced by K. brevis. The specific activities of PbTx-1, -2, -3, and -7, identified by High-Resolution Electrospray Ionization Mass Spectrometer (HRESIMS), were assessed by HPLC-UV and highly sensitive Radio-TLC counting. We demonstrated that working at close to natural concentrations of acetate is a requirement for biosynthetic studies, highlighting the importance of highly sensitive radiolabeling feeding experiments. Quantification of the specific activity of the four, targeted toxins led us to propose that PbTx-1 and PbTx-2 aldehydes originate from oxidation of the primary alcohols of PbTx-7 and PbTx-3, respectively. This approach will open the way for a better comprehension of the metabolic pathways leading to PbTx but also to a better understanding of their regulation by environmental factors

[Parapneumonic pleural effusion incidence in a French region before and during the antipneumococcal vaccine era].

International audienceBACKGROUND AND AIMS: The aim of the study was to compare the incidence of parapneumonic pleural effusion in the Limousin region of France, based on the comparison of pre- and postvaccination periods. METHODS: Subjects, 0-18-years-old, were retrospectively identified by searching in computerized databases of coded discharge diagnosis for patients with a diagnosis of pleural effusion and/or empyema and/or pulmonary infection in all the pediatric departments in Limousin hospitals. Medical records were reviewed by one of the authors and those with parapneumonic effusion and confirmed or suspected pneumococcal infection were included in the study. Data from the children hospitalized for parapneumonic pleural effusion were collected for two periods: period A, from July 2000 to July 2006, and period B, from July 2006 to July 2009 (before and after the generalization of the antipneumococcal vaccination). The main endpoint was the number of parapneumonic pleural effusion cases in each period in order to calculate the incidence within each period. RESULTS: A total of 35 children were included: nine during period A and 26 during period B. The incidence was 1 per 100,000 children for period A and 5.8 per 100,000 for period B. Bacteriological tests allowed us to serotype eight S. pneumoniae over the two periods. All serotypes were non-vaccine serotypes (1, 3, and 19A). CONCLUSION: This study demonstrates the increase in parapneumonic pleural effusion in the Limousin region

Urinary ketone body loss leads to degeneration of brain white matter in elderly SLC5A8-deficient mice.

SLC5A8 is a sodium-coupled monocarboxylate and ketone transporter expressed in various epithelial cells. A putative role of SLC5A8 in neuroenergetics has been also hypothesized. To clarify this issue, we studied the cerebral phenotype of SLC5A8-deficient mice during aging. Elderly SLC5A8-deficient mice presented diffuse leukoencephalopathy characterized by intramyelinic oedema without demyelination suggesting chronic energetic crisis. Hypo-metabolism in the white matter of elderly SLC5A8-deficient mice was found using <sup>99m</sup> Tc-hexamethylpropyleneamine oxime (HMPAO) single-photon emission CT (SPECT). Since the SLC5A8 protein could not be detected in the mouse brain, it was hypothesized that the leukoencephalopathy of aging SLC5A8-deficient mice was caused by the absence of slc5a8 expression in a peripheral organ, i.e. the kidney, where SLC5A8 is strongly expressed. A hyper-excretion of the ketone β-hydroxybutyrate (BHB) in the urine of SLC5A8-deficient mice was observed and showed that SLC5A8-deficient mice suffered a cerebral BHB insufficiency. Elderly SLC5A8-deficient mice also presented altered glucose metabolism. We propose that the continuous renal loss of BHB leads to a chronic energetic deficiency in the brain of elderly SLC5A8-deficient mice who are unable to counterbalance their glucose deficit. This study highlights the importance of alternative energetic substrates in neuroenergetics especially under conditions of restricted glucose availability

Relationship between the oxidative status and the tumor growth in transplanted triple-negative 4T1 breast tumor mice after oral administration of rhenium(I)-diselenoether.

Background: Selective inhibitory effects of rhenium(I)-diselenoether (Re-diSe) were observed in cultured breast malignant cells. They were attributed to a decrease in Reactive Oxygen Species (ROS) production. A concomitant decrease in the production of Transforming Growth Factor-beta (TGFβ1), Insulin Growth Factor 1 (IGF1), and Vascular Endothelial Growth Factor A (VEGFA) by the malignant cells was also observed. Aim: The study aimed to investigate the anti-tumor effects of Re-diSe on mice bearing 4T1 breast tumors, an experimental model of triple-negative breast cancer, and correlate them with several biomarkers. Material and methods: 4T1 mammary breast cancer cells were orthotopically inoculated into syngenic BALB/c Jack mice. Different doses of Re-diSe (1, 10, and 60 mg/kg) were administered orally for 23 consecutive days to assess the efficacy and toxicity. The oxidative status was evaluated by assaying Advanced Oxidative Protein Products (AOPP), and by the dinitrophenylhydrazone (DNPH) test in plasma of healthy mice, non-treated tumor-bearing mice (controls), treated tumor-bearing mice, and tumors in all tumor-bearing mice. Tumor necrosis factor (TNFα), VEGFA, VEGFB, TGFβ1, Interferon, and selenoprotein P (selenoP) were selected as biomarkers. Results: Doses of 1 and 10 mg/kg did not affect the tumor weights. There was a significant increase in the tumor weights in mice treated with the maximum dose of 60 mg/kg, concomitantly with a significant decrease in AOPP, TNFα, and TGFβ1 in the tumors. SelenoP concentrations increased in the plasma but not in the tumors. Conclusion: We did not confirm the anti-tumor activity of the Re-diSe compound in this experiment. However, the transplantation of the tumor cells did not induce an expected pro-oxidative status without any increase of the oxidative biomarkers in the plasma of controls compared to healthy mice. This condition could be essential to evaluate the effect of an antioxidant drug. The choice of the experimental model will be primordial to assess the effects of the Re-diSe compound in further studies

Rituximab for minimal-change nephrotic syndrome in adulthood: predictive factors for response, long-term outcomes and tolerance

BACKGROUND: Minimal-change nephrotic syndrome (MCNS) is a common cause of steroid sensitive nephrotic syndrome (NS) with frequent relapse. Although steroids and calcineurin inhibitors (CNIs) are the cornerstone treatments, the use of rituximab (RTX), a monoclonal antibody targeting B cells, is an efficient and safe alternative in childhood. METHODS: Because data from adults remain sparse, we conducted a large retrospective and multicentric study that included 41 adults with MCNS and receiving RTX. RESULTS: Complete (NS remission and withdrawal of all immunosuppressants) and partial (NS remission and withdrawal of at least one immunosuppressants) clinical responses were obtained for 25 and 7 patients, respectively (overall response 78%), including 3 patients that only received RTX and had a complete clinical response. After a follow-up time of 39 months (6-71), relapses occurred in 18 responder patients [56%, median time 18 months (3-36)]. Seventeen of these received a second course of RTX and then had a complete (n = 13) or partial (n = 4) clinical response. From multivariate analysis, on-going mycophenolate mofetil (MMF) therapy at the time of RTX was the only predictive factor for RTX failure [HR = 0.07 95% CI (0.01-0.04), P = 0.003]. Interestingly, nine patients were still in remission at 14 months (3-36) after B-cell recovery. No significant early or late adverse event occurred after RTX therapy. CONCLUSIONS: RTX is safe and effective in adult patients with MCNS and could be an alternative to steroids or CNIs in patients with a long history of relapsing MCNS