Idiopathic pulmonary fibrosis (IPF) is a chronic, fibrosing interstitial lung disease that primarily affects older adults. Median survival after diagnosis is 2–3 years. The clinical course of IPF may include periods of acute deterioration in respiratory function, which are termed acute exacerbations of IPF (AEx-IPF) when a cause cannot be identified. AEx-IPF may represent a sudden acceleration of the underlying disease process of IPF, or a biologically distinct pathological process that is clinically undiagnosed. An AEx-IPF can occur at any time during the course of IPF and may be the presenting manifestation. The incidence of AEx-IPF is hard to establish due to variation in the methodology used to assess AEx-IPF in different studies, but AEx-IPF are believed to occur in between 5 and 10% of patients with IPF every year. Risk factors for AEx-IPF are unclear, but there is evidence that poorer lung function increases the risk of an AEx-IPF and reduces the chances of a patient surviving an AEx-IPF. The presence of comorbidities such as gastroesophageal reflux disease (GERD) and pulmonary hypertension may also increase the risk of an AEx-IPF. AEx-IPF are associated with high morbidity and mortality. Patients who experience an AEx-IPF show a worsened prognosis and AEx-IPF are believed to reflect disease progression in IPF. Current treatments for AEx-IPF have only limited data to support their effectiveness. The latest international treatment guidelines state that supportive care remains the mainstay of treatment for AEx-IPF, but also give a weak recommendation for the treatment of the majority of patients with AEx-IPF with corticosteroids. There is emerging evidence from clinical trials of investigational therapies that chronic treatment of IPF may reduce the incidence of AEx-IPF. Additional clinical trials investigating this are underway. Idiopathic pulmonary fibrosis (IPF) is a chronic, fibrosing interstitial pneumonia of unknown cause that occurs primarily in older adults . In the A universally agreed definition of an AEx-IPF has not been established. Clinical studies have used a variety of criteria to define AEx-IPF, which differ in factors such as whether chest X-ray, or high-resolution computed tomography (HRCT) is required, how hypoxemia is defined, and the conditions that must be ruled out -14. VariIn 2007, a consensus definition for AEx-IPF was proposed by an expert committee sponsored by the IPF Clinical Research Network and the National Heart Lung and Blood Institute (NHLBI), in an attempt to standardize the diagnostic criteria used across studies . This coThe true incidence of AEx-IPF remains unclear. Reported incidences vary widely across studies, due to the lack of a consistent definition and differences in study methodology and patient selection ,21. The A variety of patterns of acute lung injury have been observed in AEx-IPF . The mosSeveral hypotheses for the etiology of AEx-IPF have been proposed. AEx-IPF may represent a sudden acceleration of the underlying disease process due to unknown acute injury to the lung, or a biologically distinct pathological process due to a clinically occult condition, such as infection or gastroesophageal reflux disease (GERD) . As AEx-Although research is ongoing into gene expression patterns and the identification of biomarkers, the molecular mechanisms underlying AEx-IPF remain poorly understood. Activation of the immune system, disordered coagulation/fibrinolysis, and oxidative stress may all contribute to the pathophysiology of AEx-IPF. Immune cells  ,28,29, iEpithelial cell damage in patients with IPF is demonstrated by over-expression of matrix metalloproteinase (MMP)-7 , MMP-9 , and KreAn AEx-IPF can occur at any time during the course of IPF and, for some patients, may be the presenting manifestation ,11,23,39A higher degree of dyspnea   or of fiGERD, a common comorbidity in patients with IPF, may contribute to the pathogenesis of IPF through introduction of gastric acid into the respiratory tree ,47,48. I2, DLCO) prior to AEx-IPF are more likely to survive an AEx-IPF  vs. 7 [7.6%]), but the number of events was small and the difference was not statistically significant . In triaPirfenidone, an anti-fibrotic molecule that has been licensed for the treatment of IPF in Japan, India, China, Europe, and Canada, but was not approved in the United States, has shown inconsistent effects on AEx-IPF. A Phase II study in Japanese patients with IPF was terminated after 9 months of a planned 1-year follow-up based on a higher frequency of AEx-IPF in the placebo group than in the pirfenidone 1800 mg/day group . HoweverNintedanib (formerly known as BIBF 1120) is a tyrosine kinase inhibitor in clinical development for the treatment of IPF. In the Phase II, 12-month, randomized, placebo-controlled TOMORROW trial, a lower incidence of AEx-IPF was observed in patients treated with nintedanib 300 mg/day than placebo (2.4 vs. 15.7 AEx-IPF per 100 patient years) . HoweverFinally, a recent study using prospective data from 242 patients in the placebo groups of three randomized clinical trials showed that patients who used anti-acid treatments (proton-pump inhibitors or histamine-receptor-2 blockers) at baseline had fewer AEx-IPF than those not taking such treatments at baseline (zero vs. nine events after a mean follow-up of 30 weeks) . The autIPF is a progressive disease and all patients will deteriorate over time. However, IPF has a highly variable disease course. Some patients with IPF suffer AEx-IPF, which are associated with high morbidity and mortality and a worsened prognosis. The methodology used to define AEx-IPF has varied across studies, making it difficult to form an accurate understanding of the incidence and clinical implications of AEx-IPF. More data are needed on the impact of AEx-IPF on patients with IPF. Current treatments for AEx-IPF have only limited data to support their effectiveness and supportive therapy remains the mainstay of care. There is emerging evidence that chronic treatment of IPF may reduce the risk of AEx-IPF and clinical trials investigating this are underway.AEx-IPF: Acute exacerbation(s) of idiopathic pulmonary fibrosis; ALAT: Latin American Thoracic Association; ATS: American Thoracic Society; BAL: Bronchoalveolar lavage; DLco: Diffusing capacity of the lung for carbon monoxide; ERS: European Respiratory Society; FGFR: Fibroblast growth factor receptor; FVC: Forced vital capacity; GERD: Gastroesophageal reflux disease; HRCT: High-resolution computed tomography; IPF: Idiopathic pulmonary fibrosis; JRS: Japanese Respiratory Society; KL-6: Krebs von den Lungen-6; MMP: Matrix metalloproteinase; NHLBI: National Heart Lung and Blood Institute; PaO2: Arterial oxygen tension; PDGFR: Platelet-derived growth factor receptor; PMX: Polymyxin B-immobilized fiber column; TGF: Transforming growth factor; UIP: Usual interstitial pneumonia; VEGFR: Vascular endothelial growth factor receptor.DSK has acted as a consultant, steering committee member, and advisory board member for Boehringer Ingelheim, which is developing nintedanib as a treatment for IPF. The article processing charge for this article has been covered by Boehringer Ingelheim.The author was fully responsible the content of this review article.


Regular moderate intensity physical activity positively influences the immune system with a lower incidence of upper respiratory tract infections (URTI) and lower levels of pro-inflammatory markers. However, marathon running due to its strenuous and prolonged nature results in immune perturbations with a major increase in pro-inflammatory markers and subsequent increased incidence of URTI. Furthermore, marathon running results in muscle damage and changes in hemostasis that promote a pro-thrombotic state.Naturally occurring hydrolytic enzymes and flavonoids have antioxidant, anti-inflammatory and fibrinolytic effects, and may serve as countermeasures to exercise-induced inflammation, immune dysfunction and URTI.The aim of this study is to determine whether the ingestion of oral hydrolytic enzymes and flavonoids before and after a marathon attenuates post-race muscle damage and inflammation, counters pro-thrombotic changes in hemostasis and decreases URTI incidence.Enzymes, Marathon runninG, Inflammation, Coagulation) is a randomized, double-blind, placebo-controlled, monocenter phase I trial. 160 healthy males (age 20-65 years) will be randomized to receive either placebo or treatment  which contains the hydrolytic enzymes  and the flavonoid rutoside. One week before the marathon race, participants will begin daily ingestion of the investigational product (3×4 tablets). Intake will be continued for two weeks after the race (3×2 tablets per day). Clinical and laboratory measures will be collected 5-weeks and 1-week before the race, and immediately-, 24-h, 72-h, and 2 weeks after the race. The primary endpoint is the influence of the treatment on the pre-to-post marathon race plasma concentration change of the inflammatory marker interleukin-6 (IL-6). Secondary endpoints include the effect of treatment on salivary IgA concentration and the frequency of upper respiratory tract infections (URTI) for two weeks post-marathon as determined by the Wisconsin Upper Respiratory Symptom Survey (WURSS-24). Furthermore, changes of muscular and rheological parameters will be measured before and after the marathon race.The Enzy-MagIC-study .NCT01916408ClinicalTrials.gov Identifier:  Regular moderate intensity physical activity has been shown to positively influence the immune system ,2. CliniHowever, it is discussed equivocally whether IL-6 is primarily a pro- or anti-inflammatory cytokine. IL-6 activity appears to be essential for both the effective management of acute inflammation and to balance pro- and anti-inflammatory activity . Post-exBesides immune alterations, marathon running results in muscular damage which causes elevated levels of muscle enzymes (e.g. creatine kinase) and muscle soreness . AdditioConsumption of plant derived foods such as vegetables and fruits has been linked to a reduced incidence of chronic diseases such as cancer and atherosclerosis ,22. The Supplementation with oral hydrolytic enzymes has been reported to have analgetic, anti-edematous and anti-inflammatory influences ,34. BromWobenzym  is an orally applied formulation composed of hydrolytic enzymes  and the flavonoid rutoside. Clinical trials in both humans and animals suggest a beneficial effect of this combination on inflammatory diseases. These include immunologically mediated atherosclerosis in rat aortic allografts and rheumatologic disease ,40. FollTaken together, supplementation with polyphenols and hydrolytic enzymes appears to be a promising approach to mitigate marathon-induced changes in inflammation, muscular damage and soreness, and rheological status.Therefore, the aim of the study is to investigate the influence of hydrolytic enzyme and flavonoid supplementation on changes in inflammatory, muscular, and rheological status in healthy males participating in a marathon race.Enzy-MagIC is a randomized, double-blind, placebo-controlled, monocenter phase I trial. Participants will be randomized in a 1:1 manner to receive either Wobenzym (treatment) or placebo. An overview of timing of randomization, treatment schema and data collection is presented in Figure A total of 160 healthy male athletes between the ages of 20 and 65 years who intend to participate in the Munich Marathon 2013 will be recruited from August-September 2013. The inclusion and exclusion criteria are presented in Table The study will be conducted in accordance with Good Clinical Practice guidelines, the guiding principles of the Declaration of Helsinki 2008, and local laws and regulations. The study protocol has been approved by the ethics committee of the University Hospital Klinikum rechts der Isar, Munich, Germany  and the Bundesinstitut für Arzneimittel und Medizinprodukte (BfArM), Bonn, Germany .All participants will have to provide written informed consent. The trial is registered at ClinicalTrials.gov (NCT01916408).One week before the marathon race, the participants will begin daily ingestion  of the investigational product Wobenzym (3×4 tablets) and for the two weeks after the marathon race 3×2 tablets. The treatment schema can be seen in Figure The study product is Wobenzym, an orally applied enzyme-rutoside combination supplement. Each tablet contains 90 mg bromelain, 48 mg trypsin, and 100 mg rutoside. The current indications of the investigational product are swelling and inflammation caused by injuries/trauma, thrombophlebitis and inflammation of joints due to osteoarthritis. Pharmacological investigations in animals have proven no toxic, teratogenic or mutagenic characteristics after single, multiple or long-term intake.Wobenzym (formerly known as Phlogenzym®) has been on the German market since 1991. Typical adverse reactions of this oral enzyme therapy are a change of consistency, color and odor of the stool. At higher doses, sensations of fullness, flatulence and diarrhea may appear. These symptoms may also be related to the excipient lactose. Very few allergic reactions have been reported. All reported adverse reactions have been minor and disappeared after discontinuation. None of the cases required further treatment. The placebo supplement for the control group consists of lactose-monohydrate and has no active ingredient.Blood samples will be collected 5-weeks (visit 1) and 1-week (visit 2) before the race, and immediately (visit 3), 24-h (visit 4), 72-h (visit 5) and 2 weeks (visit 6) after the race. At these visits inflammation measures , leukocyte count) will be analyzed.Muscle damage parameters (creatine kinase (CK), myoglobin, glutamate oxalacetate (GOT), lactate dehydrogenase (LDH)) will be collected and measured at each visit (1-6). The short form of the McGill-questionnaire will be used to assess muscular strain .Rheological parameters , platelet aggregation) will be collected and measured at each visit (1-6).Baseline data of the participants will be collected at the first visit. This includes questionnaires assessing training history, history of cardiovascular risk factors, physical examination, anthropometry, clinical chemistry, resting ECG, and echocardiography. One week prior to the race URTI-rates will be assessed daily by the Wisconsin Upper Respiratory Symptom Survey (WURSS-24) and will be compared between treatment and placebo group during the 2-week period following the race. The WURSS-24 is a reliable and valid tool for assessing symptoms, functional impairments, and global severity and global change of common cold . Only coParticipants will be requested not to use medications influencing inflammation such as non-steroidal anti-inflammatory drugs. Furthermore, subjects will be asked to refrain from fatty food and from all polyphenol-containing foods, especially beverages such as wine, beer, and fruit juice, as well as fresh and dried fruits or vegetables. Nutritional intake will be recorded with a 3-days nutritional record before visit 2 and before the marathon race.The primary endpoint criterion is the effect of Wobenzym compared to placebo on plasma IL-6 concentration changes following the marathon race.Secondary endpoints include the effect of consuming Wobenzym on:1. the frequency of upper respiratory tract infections (URTI) during the 2-week post-marathon period as determined by the Wisconsin Upper Respiratory Symptom Survey (WURSS-24).2. pre-to-post marathon race changes in salivary IgA concentration.3. pre-to-post marathon race changes of muscular  and rheological  parameters, and muscular strain ( McGill Pain Questionnaire).4. pre-to-post marathon race changes of cytokine concentrations , oxidative markers (e.g. myeloperoxidase) and acute-phase-proteins (e.g. high-sensitivity C-reactive protein [hs-CRP]) as well as differential blood and leukocyte count (e.g. leukocytosis).Sample size calculation was based on the primary endpoint criterion, the difference in marathon-induced change of plasma IL-6 between the study groups. Due to the expected right-skewed distribution of IL-6, differences in changes of logarithmized IL-6 levels (relative group changes) were considered as the base for inferential statistics. From previous publications a coefficient of variation (CV) of 0.60 for IL-6 changes was expected for the proposed study population . AssuminParticipants will be randomized to Wobenzym or placebo in a 1:1 ratio using a pre-generated randomization list with variable block sizes.According to the randomization list the IMP (either Wobenzym or placebo) will be labelled with the participant’s ID by the manufacturer MUCOS GmbH and will be sent to the principal investigator.Adverse events are undesirable signs or symptoms that occur during the study and may or may not be causally related to the treatment. All adverse events considered at any level to be related to the IMP will be recorded on eCRFs. A common cold or URTI episode is not regarded as an AE in this study.Serious adverse events (SAEs) are defined as events that are fatal, life-threatening, disabling, incapacitating, or resulting in hospitalization, prolonged hospital stay, or malformation. All will be recorded in the eCRF, whether they are related to the IMP or not. According to preclinical data Wobenzym is a safe supplement. Any SAE considered at any level to be related to the IMP will be regarded as unexpected. All SAEs will be reported according to the GCP-guidelines.Participants have the right to withdraw from the clinical trial at any time and in any case without giving a reason. The investigator has the right to withdraw a subject for any reason which is in the best interests of the subject, including intercurrent illness or adverse events. Discontinued participants will not be replaced.The difference of logarithmized IL-6 values will be calculated for each participant as distribution of IL-6 differences within the groups is expected to be skewed. Differences of logarithmized values will be compared between both study groups using a two-sample t-test. A two-sided level of significance of 5% will be considered for this primary efficacy analysis. Quantitative secondary endpoints will be analyzed using ANOVA methods, and qualitative data using chi-squared tests. Analysis of the primary endpoint and the secondary efficacy endpoints will be performed following the intention-to-treat (ITT) principle. These endpoints will also be assessed in the per-protocol dataset in the sense of sensitivity analyses.The ITT dataset will consist of all trial subjects enrolled into the trial and randomized. For subjects with missing post marathon inflammation markers, a conservative replacement of the missing values will be conducted by imputing the lowest observed post marathon value for the placebo group and the highest observed value for the treatment group. The per-protocol dataset includes all trial subjects who were treated according to protocol. A tertiary dataset for analysis is the safety population. This population includes all trial subjects who received any IMP or other trial treatment.The trial database will be based on the MARVIN system, an electronic data base developed for clinical trials. The system is CDISC certified, well defined and represents a worldwide accepted data and documentation standard. This approach allows online plausibility checks and automated queries, and contains automatic validation rules. Every modification will be automatically tracked during the trial.The database is integrated in an IT infrastructure and safety system containing a backup and firewall system. The data are saved on a daily basis.The medical data in this trial are to be recorded in eCRFs. After completing the data input (including data cleaning) the database will be locked and can be exported for statistical analysis in SPSS or SAS format.The Enzy-MagIC trial will investigate the effects of oral hydrolytic enzymes and flavonoids (Wobenzym) on marathon-induced changes in inflammation, muscle impairment, and rheological status in healthy males. Previous studies indicate that Wobenzym has anti-edematous, anti-inflammatory, antioxidant, and fibrinolytic effects ,39,40. TPlasma IL-6 concentration is increased after strenuous prolonged exercise such as marathon running . PolypheSupplementation of a mixture of flavonoids has been reported to have anti-inflammatory and antioxidant effects after exercise ,28. ResuSupplementation with bromelain in children with sinusitis showed a significantly faster recovery from symptoms compared to standard therapy . TherefoMarathon running induces significant muscle soreness and damage, with large increases in creatine kinase and myoglobin . MusculaFollowing marathon race participation, athletes experience changes of hemostasis associated with a pro-thrombotic state . Two stuIn summary, ingestion of oral hydrolytic enzymes and flavonoids is hypothesized to function as a countermeasure strategy to marathon-induced inflammation, muscle damage and soreness, negative changes in rheological status, and increased URTI incidence. These negative physiologic effects after marathon competition prolong recovery. For athletes it is very important that the regeneration is as effective and as short as possible. A product that could support recovery from strenuous exercise would be beneficial not only for high-performance athletes but also for non-elite athletes.ANOVA: Analysis of variance; CDISC: Clinical data interchange standards consortium; CK: Creatine kinase; (hs-)CRP: (high-sensitivity) c-reactive protein; eCRF: Electronic case report form; GCP: Good clinical practice; GOT: Glutamate oxalacetate transaminase; IL: Interleukine; IMP: Investigational medical product; LDH: Lactate dehydrogenase; SPSS: Statistic package for social sciences, statistical program; SAS: Statistical data format; SAE: Serious adverse event; tPA: Tissue plasminogen activator; URTI: Upper respiratory tract infections.None of the authors had any personal or financial conflicts of interest.Funding for the study was partly received by MUCOS Pharma GmbH & Co. KG, Berlin. The funders had no direct role in the study’s design, conduct, analysis, interpretation of data and reporting beyond approval of the scientific protocol in peer review for funding. No other grants were received.All authors made substantial contributions on the design of the trial. VG wrote the first and final draft manuscript and contributed to the conception of the final draft of the trial protocol and was responsible for the trial conduct and acquisition of data. As principal investigator JS wrote the first and final draft of the trial protocol. JS supervised the conduction of the trial and coordinated the cooperation between different parties. All other authors provided parts of the manuscript and revised the manuscript critically for important intellectual content and approved the final version to be published.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/2052-1847/6/8/prepub


Neuropsychiatric manifestations are common in childhood-onset systemic lupus erythematosus (cSLE) and often include neurocognitive dysfunction (NCD). Functional magnetic resonance imaging (fMRI) can measure brain activation during tasks that invoke domains of cognitive function impaired by cSLE. This study investigates specific changes in brain function attributable to NCD in cSLE that have potential to serve as imaging biomarkers.n = 7) vs. without NCD  using voxel-wise and region of interest-based analyses. The relationship of brain activation during fMRI tasks and performance in formal neuropsychological testing was assessed.Formal neuropsychological testing was done to measure cognitive ability and to identify NCD. Participants performed fMRI tasks probing three cognitive domains impacted by cSLE: visuoconstructional ability (VCA), working memory, and attention. Imaging data, collected on 3-Tesla scanners, included a high-resolution T1-weighted anatomic reference image followed by a T2*-weighted whole-brain echo planar image series for each fMRI task. Brain activation using blood oxygenation level-dependent contrast was compared between cSLE patients with NCD (NCD-group, Greater brain activation was observed in the noNCD-group vs. NCD-group during VCA and working memory fMRI tasks. Conversely, compared to the noNCD-group, the NCD-group showed more brain activation during the attention fMRI task. In region of interest analysis, brain activity during VCA and working memory fMRI tasks was positively associated with the participants' neuropsychological test performance. In contrast, brain activation during the attention fMRI task was negatively correlated with neuropsychological test performance. While the NCD group performed worse than the noNCD group during VCA and working memory tasks, the attention task was performed equally well by both groups.NCD in patients with cSLE is characterized by differential activation of functional neuronal networks during fMRI tasks probing working memory, VCA, and attention. Results suggest a compensatory mechanism allows maintenance of attentional performance under NCD. This mechanism appears to break down for the VCA and working memory challenges presented in this study. The observation that neuronal network activation is related to the formal neuropsychological testing performance makes fMRI a candidate imaging biomarker for cSLE-associated NCD. Studies suggest that neuropsychiatric systemic lupus erythematosus (NPSLE) is present in as many as 80% of adults with SLE -3 and maBrain function can now be mapped using blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI) that utilizes deoxyhemoglobin as an endogenous contrast agent to identify areas of altered perfusion. The coupling of neuronal activity to hemodynamics allows the identification of neuronal networks whose activity changes during the performance of cognitive tasks . Our ownFor this cross-sectional study, participants were recruited from two study sites (Cincinnati and Chicago). The study was approved by the institutional review boards of Cincinnati Children's Hospital Medical Center, Ann and Robert H. Lurie Children's Hospital of Chicago, and Northwestern University. Assent and written parental consent were obtained prior to any study procedure.All participants fulfilled the American College of Rheumatology classification criteria for SLE prior to age 16 years  and wereSociodemographic status was assessed for each participant and medical histories were reviewed for information relevant to cSLE. Disease activity and damage were measured by the SLE Disease Activity Index and the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index , respectFormal neuropsychological testing was performed by a trained psychometrician, using a standardized neuropsychological battery for cSLE . Under cParticipants with at least one domain z-score ≤ -2 or at least two domain z-scores ≤ -1 were categorized as members of the NCD-group. Otherwise, participants were considered members of the noNCD-group .® software  running on a dedicated computer was used for stimulus display and response collection. Subjects responded using a handheld response box  that was connected by optical fiber to the computer. Each response was recorded and subsequently analyzed for correctness and response time.During a single imaging session, each participant completed three fMRI paradigms probing attention, working memory, and VCA. Each of the fMRI paradigms used a block periodic design, with active task intervals interleaved with control task intervals. Visual stimuli were projected onto a screen 50 cm behind the subject's head and viewed by a mirror attached to the head coil. PresentationAttention allows concentration on a specific target stimulus over a span of time, while avoiding distraction from extraneous stimuli. An identical pairs continuous performance task (CPT-IP), consisting of identifying the repetition of any item in a sequence, was utilized in this study to measure attention . The actWorking memory allows for information to be maintained and available for use for a brief period of time. An N-back paradigm was used in this study to invoke working memory . ParticiVisuoconstruction involves the ability to organize and manually manipulate spatial information to make a design. We adapted a previously developed fMRI paradigm to probe VCA in children ,13. ThisWhile the contrasts of matching vs. motor conditions and square completion vs. motor conditions both delineate brain activation involving visuoconstruction, the contrast between the square completion vs. matching conditions further controls for purely perceptual function to focus on constructional ability .Imaging was performed at two separate sites with matching protocols using a Philips Achieva 3 Tesla (3T) and a Siemens Trio 3T scanner, respectively. The same T2*-weighted gradient-echo echo-planar imaging sequence was used for all fMRI paradigms, with the following parameters: repetition time (TR) 3,000 ms (milliseconds), echo time (TE) 30 ms, field of view (FOV) 256 × 256 mm, matrix 64 × 64 pixels, 44 axial slices, slice thickness 3 mm. In addition, a high-resolution, T1-weighted, inversion-prepared three-dimensional magnetization-prepared rapid gradient echo (MPRAGE) whole brain scan was acquired for each study participant. Parameters for this scan were as follows at site 1: TR 6.8 ms, TE 2.9 ms, inversion recovery time 904 ms, FOV 176 × 256 × 256 mm, matrix 176 × 256 × 256 pixels . At site 2, the parameters were identical except for a TE of 3.1 ms and an inversion recovery time of 900 ms to account for differences in scanners. These scans served as the anatomic reference for co-registration and overlay of functional data. During each session, the MPRAGE volume was acquired first, followed by the functional imaging. A series of 148 images  was acquired for the VCA paradigm, while the CPT-IP and N-back tasks required 114 images each . Data from the initial four time points were discarded from the imaging series of each paradigm to allow for attainment of T1 relaxation equilibrium.In order to compare signal characteristics between sites and to monitor stability, a phantom was scanned each day a subject was imaged, as recommended for multisite studies by the Function Biomedical Informatics Research Network (FBIRN) . MatchinProcessing of three-dimensional anatomic and fMRI data was done using Statistical Parametric Mapping (SPM) software  in the MPhantom data were analyzed via dedicated software obtained from FBIRN . Site cot-tests between groups. For categorical variables, frequencies (in %) of correct responses per fMRI task were compared using Fisher's exact test.Continuous measures of sociodemographic and cSLE relevant information as well as numbers of correct responses during each of the fMRI tasks were summarized by means and SDs and compared using 2 sample The functional imaging data were analyzed using both voxel-wise- and region of interest (ROI)-based approaches. Voxel-based analyses are able to examine activations across the entire brain, but are limited by the need for more stringent correction for multiple comparisons, and thus, are less sensitive. ROI-based analyses examine neuronal activation only in pre-specified regions of the brain linked to a given fMRI task. ROI-based analyses sacrifice examining the entire brain for enhanced sensitivity to capture group differences at the selected brain areas using aggregate measures.P = 0.37), SNR differed significantly . In light of this, the analysis was adjusted for a site effect by adding a site covariate to the model. Differences in mean activation between groups were assessed for each task contrast separately using the two-sample t-test. The resulting T-score maps were thresholded at a nominal voxel P-value of 0.005, uncorrected for multiple comparisons, with resulting clusters of voxels assessed in SPM8 for significance at a corrected P-value < 0.05.Voxel-wise brain activation was compared between the NCD and noNCD-groups using the following two-level analytic approach. At the first level, brain activation was estimated for each fMRI task at each voxel under the general linear model framework. The design matrix included the block periodic time series for each condition of the task. The time series was adjusted for the known delay of BOLD responses using the canonical hemodynamic response function in SPM8. Motion parameters were included as nuisance covariates in the design. Contrasts of interest between task conditions were assessed as differences in corresponding estimated model parameters. In the second level, a random- (or mixed-) effect model was used to compare means between groups at each voxel, after accounting for within-person (or between-voxel) relationships using a random effect. Phantom data suggested differences between scanners used in this study. While SFNR was markedly similar between scanners  the voxels had to be part of a cluster of at least 10 activated (or deactivated) adjacent voxels. This threshold and cluster size within a given ROI was chosen to help reduce noise, hence avoid the detection of spurious activations in isolated voxels. Use of aggregate T-scores provides a measure that reflects a combination of activation and variance.For each ROI, its activation (or deactivation) level was measured by aggregating T-scores ,22 from For each task-specific ROI, the association between brain activation and NCD status was determined using an analysis of variance (ANOVA) model, after controlling for imaging site. Post hoc means of activation were compared between the NCD-group and the no-NCD-group under the ANOVA model framework.P-values < 0.05 were considered statistically significant.The relationship of each domain z-score from formal neuropsychological testing and the ROI activation level was determined by calculation of a partial correlation coefficient, after controlling for imaging site. All analyses were repeated after adding more controlling covariates, such as family income levels and the current dose of systemic steroids to the models. Because the results of models with these additional covariates were found no different from those adjusting only for imaging site, they are not reported. All ROI analyses were performed using SAS 9.3 software . Among the twenty-two study participants, fourteen had normal cognition (noNCD-group) and eight were found to have NCD (NCD-group) based on their performance during formal neuropsychological testing. One participant with NCD failed to complete the fMRI session and was excluded, leaving a total of twenty-one participants, including seven with NCD, with complete data. Key demographic and cSLE-related data from the study population are summarized in Table P < 0.0003) and the square completion condition that is part of the VCA fMRI task . The correct response rate for the CPT-IP attention fMRI task, however, did not differ significantly between groups . Control tasks for both N-back and VCA showed improved performance with insignificant group differences: 0-back , VCA matching condition .The correct response rates and response times required for correct responses during the CPT-IP attention, 2-back working memory, and VCA fMRI tasks were compared between groups. The mean response times for all three fMRI tasks were not significantly different between groups. Conversely, compared to the noNCD-group, the NCD-group had a significantly lower correct response rate for the 2-back working memory fMRI task . The VCA paradigm also produced stronger activation in the noNCD-group compared to the NCD-group, with significant clusters  in the precuneus and right occipital regions for the matching vs. motor contrast of the VCA paradigm. There were also trends in differential brain activation in the basal ganglia between groups for the square completion vs. motor contrast. The square completion vs. matching contrast failed to show activation at the uncorrected P < 0.005 threshold for individual voxels. While the NCD-group exhibited lower brain activation during the working memory and VCA fMRI tasks than the noNCD-group in relevant brain regions, there was a trend toward greater brain activity in the frontal lobe regions in the NCD-group than in the noNCD-group during the CPT-IP attention task.T-maps of differences in activation between the NCD-group and the noNCD-group for the three paradigms are displayed in Figure In line with the voxel-wise analysis, ROI-based analysis suggested significant differences between groups in neuronal network activation in select ROIs that are considered relevant for the N-back, VCA and CPT-IP fMRI tasks Table . The noNWe then assessed the relationship between participant performance during formal neuropsychological testing and neuronal network activation patterns. Table We found differential neuronal activation in several brain regions during fMRI tasks exercising VCA, working memory, and attention in children with cSLE who have NCD, compared to those with normal cognition based on formal neuropsychological testing. Furthermore, we newly report details about the relationship between the level of cognitive performance and task-driven regional brain activation in cSLE.Previous investigations have reported various cognitive deficits in both adult SLE and cSLE, most commonly including impairment of attention, working memory, and VCA -25. EarlThe results of our study support the use of a similar compensatory mechanism for NCD for the attention task CPT-IP. We demonstrated greater activation in an insular/superior temporal ROI in the presence of clinically overt NCD compared to no NCD while CPT-IP performance remained unaffected. Notably, a negative relationship between CPT-IP brain activation in several ROI and patients' working memory abilities, as measured by formal neuropsychological testing, was observed. A possible explanation for this finding is that CPT-IP is, in effect, a 1-back working memory task. It represents a simpler version of the N-back (2-back) task. Lack of negative association of CPT-IP activation with attention domain scores suggests that attention deficits may not be the principal drivers of compensatory activation for this task. The current study in young patients with cSLE directly supports the notion that there is a relationship between the presence of NCD and alterations in brain activation.et al. reported that longer SLE disease durations are associated with diminished neuronal activation during an fMRI task of working memory among adults with SLE [Mackay with SLE . The patwith SLE  with lonet al. suggest that compensatory augmentation of task activation can break down with sufficient levels of disease damage. It is plausible that the threshold for breakdown also depends on task difficulty. Unlike the CPT-IP task, we observed diminished activation in select brain regions pertaining to the N-back and VCA tasks in cSLE patients with NCD compared to those with no NCD. Thus, some neuronal activity changes we observed in children with cSLE-related NCD parallel those in adults with SLE of extended duration. Intriguingly, this decrease of activation associated with NCD occurred in the precuneus and inferior parietal areas, which are regions where cSLE patients have previously showed more brain activation than healthy controls under a working memory fMRI task [Nonetheless, the observations by Mackay MRI task .In aggregate, our results are consistent with a model in which intact cognitive performance can be maintained in children and adults with SLE via compensatory increased neuronal activation. However, under sufficient disease burden or cognitive challenge, that compensatory pattern breaks down, resulting in diminished activation and clinically apparent cognitive dysfunction.As a group, the children with NCD in this study had a significantly lower correct response rate than the noNCD-group during fMRI tasks probing working memory and VCA, while performance on the CPT-IP attention fMRI task showed no significant group differences. Correspondingly, while activation decreased for working memory and VCA in those with NCD, the attention task (CPT-IP) elicited stronger activations for the NCD-group on both voxel-based and ROI-based analyses. It is possible that the CPT-IP task did not challenge the study participants as much as the other fMRI tasks, allowing the NCD-group to maintain performance by means of the compensatory strategy of greater activation described above. Alternatively, the lack of CPT-IP performance differences between the NCD-group and the noNCD-group may be due to relatively well-matched attention domain z-scores in formal neuropsychological testing, or to a more robust mechanism for maintaining attentional ability compared to working memory function or VCA.For all three fMRI tasks, there is a common thread of positive correlation between activation and attention domain formal neuropsychological testing z-scores. Thus attention deficits in the NCD-group may play a role in diminishing performance during fMRI tasks that probe working memory and VCA, and the corresponding decrease in brain activation in patients with NCD.This study offers a framework for viewing NPSLE as a burden on brain function that elicits compensatory mechanisms, relying on neuronal plasticity to maintain cognitive performance. Plasticity has limits which, when exceeded result in clinical manifestations of cognitive deficit. In the context of brain plasticity, the course of NPSLE may have a more profound impact on children with cSLE, given their dramatic, ongoing, brain development. While children possess a greater capacity for brain plasticity in response to an assault such as NPSLE , there rA distinct strength of our study is that its participants were well-phenotyped with respect to their clinical cSLE status, and their cognitive abilities were accurately assessed using the cSLE Cognitive Battery of Standardized Tests . HoweverIn summary, we found differences in brain activation patterns that are related to distinct cognitive deficits in cSLE. Building on the results of our previous study , we postAAL: automated anatomical labeling; ANOVA: analysis of variance; BOLD: blood oxygenation level-dependent; CPT-IP: continuous performance task-identical pairs; cSLE: childhood-onset SLE; FBIRN: Function Biomedical Informatics Research Network; fMRI: functional magnetic resonance imaging; FOV: field of view; FWHM: full width at half maximum; IQ: intelligence quotient; MNI: Montreal Neurological Institute; MPRAGE: magnetization-prepared rapid gradient echo; NCD: neurocognitive dysfunction; NPSLE: neuropsychiatric systemic lupus erythematosus; ROI: region of interest; SES: socioeconomic status; SFNR: signal-to-fluctuation noise ratio; SLE: systemic lupus erythematosus; SNR: signal-to-noise ratio; SPM: Statistical Parametric Mapping;TE: echo time; TR: repetition time; VCA: visuoconstructional ability.The authors declare that they have no competing interests.HB, MD, and DG participated in study design. MD, DG, TP, and AS-G took part in imaging data acquisition. MK-G, FZ, DB, HB, and AS-G carried out neuropsychological data acquisition. MD and JY performed data analysis. MD, DG, JY, and HB participated in interpretation of results. MD, HB, and JY prepared the manuscript. All authors participated in review and approval of the final manuscript.Summary of brain activity vs. NCD status for all regions of interest examined. A table listing mean region of interest (ROI) activation, defined as (sum of T-scores within the ROI among voxels with T-scores > 1.66 that are part of clusters of at least 10 voxels)/total voxels in the ROI, for systemic lupus erythematosus (SLE) patients with and without neurocognitive dysfunction (NCD).Click here for file


Introduced in the early 1960s, diazepam remains among the most frequently prescribed benzodiazepine-type sedatives and hypnotics. Patients with chronic use of short-acting benzodiazepines are frequently switched to diazepam because the accumulating, long-acting metabolite, N-desmethyl-diazepam, prevents benzodiazepine-associated withdrawal symptoms, which can occur during trough plasma levels of short-acting benzodiazepines. Although mild to moderate withdrawal symptoms are frequently observed during benzodiazepine switching to diazepam, severe medical complications associated with this treatment approach have thus far not been reported.A 64-year-old female Caucasian with major depression, alcohol dependence and benzodiazepine dependence was successfully treated for depression and, after lorazepam-assisted alcohol detoxification, was switched from lorazepam to diazepam to facilitate benzodiazepine discontinuation. Subsequent to the benzodiazepine switch, our patient unexpectedly developed an acute delirious state, which quickly remitted after re-administration of lorazepam. A newly diagnosed early form of mixed dementia, combining both vascular and Alzheimer-type lesions, was found as a likely contributing factor for the observed vulnerability to benzodiazepine-induced withdrawal symptoms.Chronic use of benzodiazepines is common in the elderly and a switch to diazepam often precedes benzodiazepine discontinuation trials. However, contrary to common clinical practice, benzodiazepine switching to diazepam may require cross-titration with slow tapering of the first benzodiazepine to allow for the build-up of N-desmethyl-diazepam, in order to safely prevent severe withdrawal symptoms. Alternatively, long-term treatment with low doses of benzodiazepines may be considered, especially in elderly patients with chronic use of benzodiazepines and proven vulnerability to benzodiazepine-associated withdrawal symptoms. Diazepam was the most widely prescribed drug in the United States and Europe for nearly two decades. Even with the subsequent introduction of numerous other BZDs, diazepam remained one of the first-choice BZDs among prescribers and in 1985 was added to the World Health Organization's essential medicines list [et al. nearly one decade ago [et al. [The benzodiazepine (BZD)-type sedatives and hypnotics, introduced in the 1960s, marked a major advance in the treatment of anxiety, depression, insomnia and seizures, not least because of their improved therapeutic index. The first BZDs marketed by F. Hoffmann-La Roche, chlordiazepoxide  BZD discontinuation or (ii) low-dose BZD maintenance. Where BZD discontinuation is favored, a frequently employed BZD detoxification strategy is to switch to diazepam, or to other BZDs with long elimination half-lives of the parent drug or its active metabolites, followed by slow tapering of the drug. In individuals with chronic BZD use, a BZD switch to diazepam  frequentet al. . The autet al. .In this report we describe the case of an elderly woman with a number of mental and physical conditions who developed an acute delirious state after being switched from lorazepam to diazepam. Drug pharmacokinetics and psychiatric vulnerabilities are considered as possible causal factors.A 64-year-old Caucasian woman with major depression : 296.33), alcohol dependence (DSM IV-TR: 303.90) and benzodiazepine (lorazepam) dependence (DSM IV-TR: 304.10) was referred to a specialist ward for the treatment of substance use and co-occurring mental disorders. Daily use of alcohol started ten years prior to admission and daily use of lorazepam one year prior. In addition, our patient had a history of medical conditions for which she had been receiving medication. Levothyroxine was used to treat hypothyroidism , standard female hormone replacement therapy consisting of 1 mg 17β-estradiol and 5 mg dydrogesterone was used to alleviate and prevent symptoms associated with menopause, and atorvastatin was prescribed to control hyperlipidemia. Felodipine, metoprolol and lisinopril were used to treat hypertension. Salicylate was used for the prevention of cardiac infarction. Pantoprazole was used to alleviate symptoms related to gastro-esophageal reflux. All prescriptions, including a previous trial with the antidepressant mianserin, were well tolerated without significant side effects and, except for mianserin, were continued after admission without dose changes.On admission, lorazepam, at 5-10 mg daily, was begun to prevent alcohol- and BZD-associated withdrawal symptoms. Antidepressant treatment with mianserin (60 mg daily) resulted in a partial response. A three-week add-on trial with venlafaxine (maximum dose 450 mg daily) resulted in further improvement of depressive symptoms. However, residual depressive symptoms persisted and mianserin was switched to mirtazapine (maximum dose 75 mg daily). Complete remission was achieved two months later and venlafaxine maintained at 150 mg daily and mirtazapine at 60 mg daily (relapse prevention).Lorazepam was tapered and discontinued after six weeks of treatment. One day after the last lorazepam dose, our patient exhibited difficulties in maintaining attention and showed signs of psychomotor agitation, anxiety and disturbed thought processes, such as loosened, illogical and tangential associations. Dissociative symptoms were also observed. These signs were consistent with an ensuing BZD withdrawal syndrome (DSM IV-TR: 292.0) and suggested that an even more protracted discontinuation phase may have been necessary. Delirium was not diagnosed at this time because orientation remained intact and the severity of the symptoms was not in excess of those usually associated with a BZD withdrawal syndrome. Lorazepam was resumed at 3 mg per day. Symptoms of withdrawal resolved and lorazepam was maintained at 3 mg daily for one month. Lorazepam was then discontinued without tapering, and diazepam was started at an equivalent dose of 15 mg daily. Less than one day after the BZD switch, our patient became increasingly irritable and agitated and eventually exhibited fluctuating levels of consciousness with reduced clarity of awareness and reduced ability to maintain attention. Disorientation, incomprehensible speech and memory deficits were also noted. The diagnosis of a BZD intoxication delirium was ruled out based on the absence of slurred speech, incoordination, unsteady gait, nystagmus or stupor. Our patient was diagnosed with BZD withdrawal delirium (DSM IV-TR: 292.81) and received 4 mg of lorazepam combined with 4 mg of haloperidol. The acute delirious state resolved within hours and our patient remained well. Haloperidol was stopped after two weeks. Lorazepam was continued at 4 mg daily. Our patient was dismissed one month later with a lorazepam maintenance dose of 4 mg per day.An initial review of the prescriptions and the diagnostic tests performed in the course of treatment failed to reveal our patient's vulnerabilities to delirious states. Except for a slight increase of plasma diazepam levels in the presence of metoprolol, no relevant drug interactions were identified. A physical examination, laboratory workup and an analysis of cerebrospinal fluid were unremarkable. In particular, there was no evidence of hepatic dysfunction. Serum levels of alanine and aspartate aminotransferase, alkaline phosphatase, lactate dehydrogenase, and bilirubin were normal. Prothrombin time was within the normal range. Serum γ-glutamyltransferase was slightly elevated . However, serum concentrations of O-desmethyl-venlafaxine (a liver-derived metabolite of venlafaxine) were seven-fold higher than those of venlafaxine. Taken together, these results suggested normal liver function with intact hepatic drug metabolism and ruled out the differential diagnosis of a diazepam-exacerbated hepatic encephalopathy.Electroencephalogram recordings showed the rhythm of alpha waves (11-12 Hz) and beta waves (18-22 Hz) to be as expected during treatment with lorazepam. Epileptiform discharges or focal abnormalities were not present. Electrocardiogram recordings showed a normal sinus rhythm with a frequency of around 90 per minute. Neuropsychological tests revealed only mild cognitive deficits. At the time these test were performed our patient was receiving the medication to treat her physical conditions as described above. In addition, she received mirtazapine (60 mg daily), venlafaxine (150 mg daily) and lorazepam (4 mg daily). The Mini-Mental State Examination score was 27. The Consortium to Establish a Registry for Alzheimer's Disease test revealed minimal cognitive impairment, mainly affecting her verbal memory (left hippocampus system), executive functions  and mental rotation . On the Hamburg Wechsler Intelligence test for adults, she achieved 90 points, a score slightly below average.18F-fluorodeoxyglucose (FDG) showed diminished FDG uptake in temporal and parietal cortical regions. Lower FDG accumulation was also found in parts of the visual cortex and in both her basal ganglia and thalamus. The absence of hallucinations, Parkinsonian or extra-pyramidal symptoms, together with the previously well-tolerated trial with haloperidol, argued against Lewy body dementia [Computed tomography scans showed no signs of brain atrophy. However, using T2 pulses, magnetic resonance imaging showed scattered sub-cortical signal disturbances in her frontal, parietal and occipital regions. Additional signal disturbances were found in her pontine regions and brain stem. Together, these findings were consistent with sub-cortical atherosclerotic encephalopathy, in other words, Binswanger's disease. Positron emission tomography scanning of the brain using the radio-labeled glucose analog dementia .In agreement with the present case, evidence now suggests that (i) both short-and long-term BZD use is associated with old age, female sex, psychological stress and physical disease , (ii) loet al. who found that, despite its longer half-life, unbound diazepam distributes more extensively into tissue than lorazepam does [Since chronic BZD use constitutes a risk for cognitive decline  and sincpam does  and thuspam does . Lorazeppam does . ConsequWhile our case demonstrates a particular vulnerability to BZD-induced withdrawal symptoms in a patient with discrete vascular and Alzheimer-type lesions, the molecular mechanisms responsible for this vulnerability remain unexplained. However, irrespective of the presence of vascular or Alzheimer-type lesions, which may be associated with such vulnerability, the complex nature of the γ-amino-butyric acid (GABA) system, through which BZDs mediate their anxiolytic, sedative, anti-convulsant and muscle relaxant effects, may provide other clues.GABA, the major inhibitory neurotransmitter in the mammalian central nervous system, mediates fast post-synaptic inhibition through binding to the GABA-A receptor, a hetero-pentameric chloride-selective ligand-gated ion channel . To dateThe observed vulnerability to BZD-induced withdrawal symptoms could be explained by the expression patterns of GABA-A pentamers in our patient's brain, because different BZD effects are mediated by different GABA-A pentamers . FurtherThis case report describes a 64-year-old female Caucasian with several pre-existing medical conditions and psychiatric disorders common for her age. These were treated successfully with one notable exception, BZD dependence, for which discontinuation was the favored treatment goal. Unexpectedly, our patient developed a severe adverse reaction (delirium) associated with a switch from lorazepam to diazepam.Since BZD prescriptions in the elderly are common to almost all medical subspecialties, severe adverse events associated with BZD use must be reported quickly to alert prescribers and to improve treatment safety and quality. The present case suggests that elderly polymorbid patients with chronic BZD use may benefit from cross-tapering when switched to diazepam. Finally, long-term treatment with low doses of BZDs may be considered in these patients when BZD discontinuation trials fail.Written informed consent was obtained from the patient for publication of this case report. A copy of the written consent is available for review by the Editor-in-Chief of this journal.The author declares that they have no competing interests.The author is a board-certified psychiatrist and head of a private outpatient facility for the treatment of substance use and co-occurring disorders. The author has a background in molecular and cell biology research.


Purpose. The diagnostic performance of positron emission tomography using 18F-fluorodeoxyglucose (FDG-PET) in detecting nodal involvement in patients with anal canal cancer (ACC) has been investigated by several studies with conflicting results. The aim of our study is to systematically review and meta-analyze published data about this topic. Methods. A comprehensive computer literature search of PubMed/MEDLINE, Scopus, and Embase databases was carried out on July 10 to find relevant articles concerning the diagnostic performance of FDG-PET in detecting locoregional nodal involvement in patients with ACC. No language restriction was used. Pooled diagnostic performance on a lesion-based analysis was calculated. Results. Seven retrospective and five prospective studies have been reviewed. Six studies allowed assessing pooled sensitivity; five studies allowed assessing pooled specificity. Sensitivity and specificity values of FDG-PET/CT on a lesion-based analysis ranged from 31 to 100% and from 53 to 98%, with pooled estimates of 56% (95% CI: 45–67%) and 90% (95% CI: 86–93%), respectively. Conclusions. Our meta-analysis demonstrates that FDG-PET is a specific diagnostic tool in detecting locoregional lymph node involvement in patients with ACC. Low sensitivity is a major concern; however, higher sensitivity could be reached combining FDG-PET with MR scan.  Anal canal cancer (ACC) is a relatively uncommon neoplasm, with an incidence of about 1–3 cases per 100,000 and accounting for about 2% of gastrointestinal malignancies –6.Rectal bleeding, usually of small entity, eventually accompanied with anal itching, tenesmus, or change in bowel habits, is the most frequently occurring sign; swollen inguinal or perineal lymph nodes, mucous anal leakage, and ribbon-like stools are far less frequently observed. Fecal incontinence and frank pain while seated are common in locally advanced ACC. However, such signs and symptoms of ACC are not specific for this disease, since more common benign conditions, like hemorrhoids, rectal solitary ulcer, or anal fissure, may arise the same way. Moreover, ACC may be completely asymptomatic in about 20% of patients and incidentally discovered during rectal exam for prostate screening colonoscopy in patients at risk for colon-rectum cancer or even during surgical intervention for hemorrhoids.Early disease detection and treatment may have a significant positive impact on the overall survival of patients with ACC . HistoloCurrently, magnetic resonance imaging (MRI) performed on high-magnetic-field scanners is the imaging modality of choice to investigate the anal region for both staging and treatment evaluation purposes, due to its high-detail visualization of the anal canal and nearby anatomical structures, reliable soft tissue differentiation, noninvasiveness, and not needing special patient preparation –11. ExtrComputed tomography (CT) performed with multi-detector technique allows a good visualization of the primary lesion, although with a lower contrast resolution than MRI, being a reliable alternative in patients with contraindications to MRI. Nevertheless, CT may be helpful in detecting eventual spread of neoplastic cells in the liver or other organs: indeed, dissemination, though uncommon, is associated with a higher risk of recurrent disease after treatment .The role of positron emission tomography/computed tomography (PET/CT) using fluorine-18 fluorodeoxyglucose (FDG) in staging, response evaluation, and followup of ACC is becoming increasingly important because of its higher reported sensitivity for the detection of primary tumor, regional lymph node involvement, and distant metastases than conventional imaging . MoreoveSentinel lymph node biopsy procedure has proved to be a more accurate method than clinical or radiological techniques in staging the disease of patients with ACC, with detection rates up to 95–100% in the inguinal region , 21. ConA comprehensive computer literature search of PubMed/MEDLINE, Scopus, and Embase databases was carried out to find relevant published articles concerning the diagnostic performance of FDG-PET in detecting locoregional nodal involvement in patients with biopsy-proven ACC. We used a search algorithm based on a combination of the terms “ AND (PET OR positron emission tomography).” No language restriction was used. The search was performed from inception to July 10, 2013. To expand our search, references of the retrieved articles were also screened for additional studies.Studies or subsets in studies investigating the role of positron emission tomography/computed tomography using FDG-PET in detecting locoregional nodal involvement in patients with biopsy-proven ACC were eligible for inclusion. Case reports, small case series, review articles, letters, editorials, and conference proceedings were excluded.FDG-PET performed in patients with biopsy-proven ACC,a sample size of at least ten patients with ACC who performed FDG-PET in the course of their diagnostic workup,sufficient data to reassess sensitivity and specificity of FDG-PET in patients with ACC, andno data overlap .The following inclusion criteria were applied to select studies for this meta-analysis:Two researchers  independently reviewed titles and abstracts of the retrieved articles, applying the above-mentioned selection criteria. Articles were rejected if clearly ineligible. The same two researchers then independently evaluated the full-text version of the included articles to determine their eligibility for inclusion.Information about basic study , study design (prospective or retrospective), patients' characteristics , technical aspects , and reference standard results  was collected.Each study was analyzed to retrieve the number of true-positive (TP), true-negative (TN), false-positive (FP), and false-negative (FN) findings of FDG-PET in patients with ACC, according to the reference standard. Only studies providing such complete information were finally included in the meta-analysis.http://www.cebm.net/index.aspx?o=5653) assessment tool for diagnostic performance evaluation. For each included paper, this tool takes into account several parameters: spectrum of the studied cases, recruitment of patients (consecutive/not consecutive), reference standard, ascertainment of the gold standard regardless of the index test results (Yes/No), blind comparison of the index test and reference standard (Yes/No), enough explanation of the index test to ensure reproducibility (Yes/No), and study design (prospective/retrospective). A level-of-evidence score is obtained, ranging from 1 to 5.Two independent reviewers (R. Sadeghi and N. Ayati) evaluated the methodology of the selected studies using the “2011 Oxford Center for Evidence-Based Medicine Level of Evidence”  curve was obtained for selected studies and area under curve (AUC) was calculated to assess the overall accuracy of FDG-PET. Statistical analyses were performed using Meta-DiSc statistical software version 1.4 (available at: http://www.hrc.es/investigacion/metadisc_en.htm).Sensitivity was determined according to the following formula: TP/(TP + FN); specificity was determined according to this formula: TN/(TN + FP). Statistical pooling of the data was performed by means of a random effects model. Pooled data are presented with 95% confidence intervals (95% CI). Heterogeneity between studies was assessed by a The comprehensive computer literature search from PubMed/MEDLINE, Embase, and Scopus databases revealed 97 articles . Most paOverall, the studies included in this meta-analysis have shown moderate methodological quality according to the “2011 Oxford Center for Evidence-Based Medicine Level of Evidence” assessment tool for diagnostic performance evaluation, with level-of-evidence score ranging from 3 to 4 for most papers. Use of poor/nonindependent reference standard was the main drawback of the included studies. Results of methodology assessment are summarized in Tables In this review, seven retrospective studies about the diagnostic performance of PET/CT using FDG in detecting locoregional lymph node involvement in patients with biopsy-proven ACC have been included.Cotter et al.  evaluateDe Winton et al.  have detI2: 84.6%) and specificity (I2: 90.5%). The area under the ROC curve was 0.83.The diagnostic performance values of FDG-PET/CT in the studies included in this meta-analysis are presented in Figures ACC is a not common neoplasm, with an incidence of about 1–3 cases per 100,000 and accounting for about 2% of gastrointestinal malignancies –6.To date, MRI performed is a good tool to investigate the anal region for both primary tumour and pelvic node involvement –12. CT iFDG PET/CT is not routinely used in the staging of patients with ACC but can significantly alter the initial stage in comparison with conventional diagnostic techniques in at least 20–25% of evaluated patients –18. It iOur meta-analysis aimed to assess the diagnostic performance of PET/CT using FDG in detecting locoregional lymph node involvement in patients with biopsy-proven ACC, in order to provide evidence-based data in this setting. Sensitivity and specificity values of FDG-PET/CT on a per patient-based analysis were of 56% (95% CI: 45–67%) and 90% (95% CI: 86–93%), respectively. False-negative findings reduce sensitivity, probably caused by high uptake of primary anal tumour and small size of node lesions. Although specificity was good, false-positive findings occur, especially when the extent of inguinal involvement is assessed, as inflammatory disease of pelvic region or lower extremities should be taken into account.Limits of the Study. A systematic review process was adopted in ascertaining studies: to avoid selection bias, we adopted rigid inclusion criteria. Moreover, a methodological quality evaluation according to the “2011 Oxford Center for Evidence-Based Medicine Level of Evidence” has been performed: the index test and the reference standard were often interpreted without blinding, therefore limiting the methodological quality of the included studies.Heterogeneity between studies may represent a potential source of bias; the included studies were statistically heterogeneous in their estimates of sensitivity and specificity. Since systematic reviews bring together studies that are different both clinically and methodologically, heterogeneity in their results should be expected. For example, heterogeneity is likely to arise through diversity in technical aspects, sample size, study quality, and inclusion criteria.Publication bias is a major concern in all forms of pooled analyses since studies reporting significant findings are more likely to be published than those reporting nonsignificant results. Indeed, it is not unusual for small-sized early studies to report a positive relationship that subsequent larger studies fail to replicate. We cannot exclude a publication bias in our analysis, but we tried to minimize it by selecting only articles that included at least ten patients who performed FDG-PET with biopsy-proven anal canal cancer.Our meta-analysis demonstrates that FDG-PET is a specific diagnostic tool in detecting locoregional lymph node involvement in patients with biopsy-proven ACC. Low sensitivity is a major concern, especially when considering small size lymph node involvement; however, a higher sensitivity could be reached by combining FDG-PET with MRI scan in the diagnostic management of patients with ACC, to reduce the risk of false negative results. Upcoming hybrid PET-MRI hybrid tomographs are expected to improve the diagnostic performance of FDG-PET in these patients.


Objective: Infants of diabetic mothers (IDM) are considered as a risk group for atherosclerosis. Increased aortic intima-media thickness has been reported in IDM. The purpose of this study was to assess carotid artery intima-media thickness (CA-IMT), left ventricular mass index (LVMI)  and atherosclerotic risk factors in IDM.   Methods: Thirty IDM and 25 healthy controls were included in the study. Of these infants, 14 were appropriate-for-gestational age (AGA) and 16 were large-for-gestational age (LGA). CA-IMT and LVMI were obtained by M-mode  echocardiographic examination.  The relationship between parameters of atherosclerosis and echocardiographic measurements was assessed by Pearson’s correlation analysis. Results: LVMI was higher in LGA IDM when compared to AGA IDM and controls. CA-IMT was not significantly different between the groups and was also not related to atherosclerotic risk factors. Serum lipid and insulin levels were higher in LGA IDM when compared with AGA IDM and controls. There were no correlations between CA-IMT, LVMI and atherosclerotic risk factors. Conclusions: In contrast to previous reports indicating an increase in CA-IMT in IDM, no differences were found between IDM and controls in this study. Our results indicate that  macrosomic IDM are prone to hypertrophic cardiomyopathy but not to atherosclerotic changes in the blood vessels.    Conflict of interest:None declared. Diabetes mellitus during pregnancy affects fetal development adversely and causes metabolic disorders in neonates. The most important effect of diabetes on the fetus is macrosomia. The fetus will increase its insulin levels in response to the mother’s hyperglycemia  and this results in macrosomia . Alterat In this study, we aimed to assess the risk of  subclinical  atherosclerosis by measuring carotid artery intima-media  thickness (CA-IMT) and left ventricular mass index (LVMI) in infants of diabetic mothers (IDM) by B-mode ultrasonography. Also, we investigated the relationship between CA-IMT and  cardiovascular risk factors.This study was performed prospectively between December 2005 and December 2007 at Selcuk University, Meram School of Medicine, Pediatric Endocrinology Department. The study was approved by the local ethics committee. Signed informed consent was obtained from the parents.  Thirty  IDM and 25 healthy controls were included in the study. Fourteen of the IDM were appropriate-for-gestational age (AGA) and sixteen were large-for-gestational age (LGA). Gestational ages were determined by one or a combination of information based on mother’s last menstrual date, obstetrical ultrasound and Ballard score. Infants of mothers with a history of cardiovascular disease, high cholesterol level, hypertension, smoking, pre-eclampsia were excluded. Also, infants of mothers who had a history of taking drugs affecting lipid metabolism, such as steroids and ritodrine, were excluded. Infants with asphyxia, prematurity, respiratory distress, dysmorphic features, a stressful birth history and intrauterine infection were not included in the study. Neonates whose birth weights were above the 90th  percentile for gestational age and sex according to Lubchenco curves were accepted as macrosomic. IDM were divided into two groups as LGA and AGA. Weight was measured at birth by using digital scale and recorded in grams. Length and head circumference were measured using a standard board and tape and were recorded in centimeters. The ponderal index (PI) was calculated by the formula “weight (g) x 100/cm3.  A PI value over 2.85 was accepted as an index of LGA. Cord blood samples were taken from the study and  control groups. Cord blood samples received from Rh negative mother’s babies for blood group determination were used as the control group. Plasma triglyceride, total cholesterol,  high-density lipoprotein (HDL), low-density lipoprotein (LDL), very low-density lipoprotein (VLDL) and glucose levels were analyzed by colorimetric method (Beckman Coulter).  Plasma insulin and C-peptide levels were determined by chemiluminescence method . Reference ranges accepted for insulin and C-peptide were 1.9-23 mIU/mL and 0.9-7.1 ng/dL, respectively. Ultrasound Examination Ultrasound assessment of all infants was performed by the same clinician who was uninformed about the patient’s  laboratory values and risk factor levels. The subjects were  laid quietly for 10 minutes before the first scan in a dark,  temperature-controlled room. Scanning was performed in the supine position while the head was slightly extended and turned to the left. High-resolution B-mode ultrasonography of the right common carotid artery was performed by using a Philips Sonos 5500 Doppler  ultrasound with  3.5 MHz probe. Longitudinal images of the common carotid artery were obtained by combination of B-mode and color Doppler  ultrasound examinations. The IMT of the far wall of the  common carotid artery was measured with the electronic calipers of the machines, as described by Pignoli et al .  On a longitudinal two-dimensional ultrasound image of the carotid artery, the posterior  wall of the carotid artery  was shown as two echogenic lines. The external line was considered as the limit of medial adventitia and the inner  line - as the limit of the luminal intima. The distance between these two parallel lines shows the intimal medial thickness.  CA-IMT was calculated by taking the mean value of three measurements of maximum far wall thickness obtained from the common carotid artery, 10 mm below the carotid bulb.  Echocardiography Echocardiographic examination was conducted in all  infants in the postnatal 48-72 hours by the same clinician. Two-dimensional guided M-mode echocardiography (Philips Sonos 5500) was performed to determine LVM.  The ventricular  septal (IVSd) and posterior wall thicknesses at end-diastole (LVPWd), and the left ventricular end-diastolic (LVEDd) dimensions were determined according to the American Society of Echocardiography recommendations :  0.8 * StatisticsAll statistical analyses were performed using SPSS 13.0. Descriptive data were shown as mean±SD. Compliance with normal distribution was analyzed. Kruskal-Wallis variance analysis was performed for data which did not conform to a normal distribution. Bonferroni-corrected Mann-Whitney U test was applied for dual comparisons. For the relationship between CA-IMT and other parameters, Pearson’s correlation test was used.   A p-value lower than 0.05 was considered statistically significant.The clinical characteristics and laboratory findings of the IDM and control neonates are shown in Fetal macrosomia and obesity are common complications occurring in IDM. Maternal hyperglycemia leads to fetal hyperglycemia which causes hyperinsulinism by stimulating the pancreatic islet cells in the fetus . The hyp   Along with fetal hyperinsulinemia, macrosomia and  hypoglycemia, maternal hyperglycemia can also cause  asymmetric septal hypertrophy in the infant. Hyperinsulinemia induces increased synthesis of protein, glycogen and fat leads to hyperplasia and hypertrophy of the myocardium . Atheros  Serum lipid, insulin, C-peptide and leptin levels were all reported to be elevated in IDM . We founMehta et al  observed


Understanding of immune response mechanisms of pathogen-infected host requires multi-scale analysis of genome-wide data. Data integration methods have proved useful to the study of biological processes in model organisms, but their systematic application to the study of host immune system response to a pathogen and human disease is still in the initial stage.To study host-pathogen interaction on the systems biology level, an extension to the previously described BiologicalNetworks system is proposed. The developed methods and data integration and querying tools allow simplifying and streamlining the process of integration of diverse experimental data types, including molecular interactions and phylogenetic classifications, genomic sequences and protein structure information, gene expression and virulence data for pathogen-related studies. The data can be integrated from the databases and user's files for both public and private use.http://www.biologicalnetworks.org) and web interface (at http://flu.sdsc.edu).The developed system can be used for the systems-level analysis of host-pathogen interactions, including host molecular pathways that are induced/repressed during the infections, co-expressed genes, and conserved transcription factor binding sites. Previously unknown to be associated with the influenza infection genes were identified and suggested for further investigation as potential drug targets. Developed methods and data are available through the Java application (from BiologicalNetworks program at  Public health initiatives increasingly recognize the importance of the cross-scale data integration, such as mounting a data-driven risk assessment of potential pandemic outbreak in specific geographical locations or discovering novel therapeutic approaches -6. For eExperimental data on host-pathogen interaction are distributed throughout many heterogeneous data sources. Among the integration systems enabling studying host-pathogen interactions at multi-level scale are PHI-base , PHIDIASAt the same time, existing link-based integration systems, such as Entrez , EnsemblInfluenza infection. The proposed system is an extension of the previously developed BiologicalNetworks [http://www.biologicalnetworks.org and web interface http://flu.sdsc.edu; Demo page: http://flu.sdsc.edu/examples.jsp.In this paper, the approach at cross-scale data integration to study host-pathogen interactions is proposed and demonstrated on a study of the Networks ,41 and INetworks . It reprhttp://www.w3.org/TR/owl-ref. The IntegromeDB [http://www.w3.org/RDF/); that is, it stores biological data in an RDF-compatible format, the standard format of the Semantic Web [http://www.BiologicalNetworks.net/Database/tut0.php. The ontology is available as an OWL file at http://flu.sdsc.edu/bionetsonto.jsp.The architecture of the system, data integration and mapping procedures, database schema, ontology model and data query engine are described in detail elsewhere . TherefoegromeDB  internalntic Web . The dathttp://www.biologicalnetworks.net/Database/tut5.php[The full list of integrated databases and statistics are provided at /tut5.php. To enab/tut5.php, the folhylogenetic trees that connect host and pathogen proteins/genes with orthologs/homologs in model organisms . These data were obtained from PhyloFacts database [• Pdatabase .Literature-curated information on physiological effects of pathogen infection in experimental systems (including cell cultures and in vivo models)• irulence data about mortality/morbidity information related to isolate and incidence, isolated organism's sequence data (from WHO statistics)• VEpidemiological data on infection occurrence, pathogen culture sites and dates, migration data of vectors, past movement data of infected individuals, etc. (from NCBI Influenza Virus Resource).• Additional experimental data sets on host-pathogen interactions integrated into our system include human interactomes used by Influenza virus, HIV, HCV, dengue virus and West Nile virus (WNV) .http://flu.sdsc.edu  is provided. The data will become public, but unless curated by the data administrators, will remain "tagged" as 'uncurated' under the contributor's name. The user's data integration procedure consists of 3 simple steps: 1) User registration, 2) Data Mapping and 3) Data integration (see Section 'Integration of user's data' below). To be integrated, the data needs to be in the table format.To integrate the user's data into the system, the web page e.g. sequences) into interval trees; two- and three-dimensional data (e.g. images) into R-trees. To keep the number of the index structures small, a single interval tree is created per chromosome instead of per annotated DNA sequence regions, and the images of the same resolution are referenced with respect to the same coordinate system and placed in a single R-tree. The examples of operations on RI-trees that are used in BiologicalNetworks for Navigation and Annotation of multiply overlapping gene regulatory regions, protein binding regions, disease and geographical maps are provided in the Additional File The internal schema of the BiologicalNetworks database is shown in Figure http://flu.sdsc.edu/bionetsonto.jsp  query language that provides constructs (operators and functions) to search, manipulate and query the data. The developed for this purpose the  project . The synTo enable systems-level study of host-pathogen interaction, in addition to the query capabilities described previously , a numbeTo address the first question, the database can be searched for evolutionary distances between two specified genome sequences of different species/strains/isolates, executing the following three queries Figure : (1) recmax from a given genome X?--the following queries need to be executed  in the ancestor A rooted sub-tree, find the nodes at the distance Lmax from X.To find the answer to the second question--Which genome sequences are within the evolutionary distance Ld Figure : (1) recTo predict the virulence of a species/strain/isolate by its gene/protein sequence, the published method for virulence evaluation of low and high pathogenic avian influenza LPAI and HPAI  was implSimilarly, the probability of a protein of a given 3D structure to be virulent can be estimated, using the phylogenetic tree built based on 3D structure multiple sequence alignment , proteolysis and peptidolysis , lipid transport , and complement activation .Second, six sub-networks were combined into a single fully-connected network Figure ; that isTo study individual genes, we used the constructed in the previous section network of significantly induced/repressed genes in the influenza infection Figure . For exaHomo sapiens, Mus musculus and Rattus norvegicus. Such sites were found in 7 genes: SFRS11, SFRS1, FMNl2, LEPROT, NICN1, FHOD2, f3-contactin; if one is interested, the binding sites can be searched by sequence , signal transduction and transcriptional modulation of multiple signaling pathways, including BMP (GO:0030509) and TGF-beta receptor-signaling (GO:0030512). GRAP is involved in activation of the T-cell antigen receptor (TCR) signal transduction pathway , Ras proundantly .AMPD, or AMP deaminase, is an enzyme that converts adenosine monophosphate (AMP) to inosine monophosphate (IMP), freeing an ammonia molecule in the process. Deficiency of this enzyme is a common cause of myopathy and rhabdomyolysis . The influenza infections are known to be a cause of rhabdomyolysis, including seasonal  and receNFE2L2 gene codes the transcription factor NRF2 that is a known master regulator of the antioxidant response . By induhttp://flu.sdsc.edu/examples.jsp. Together with capability to integrate new phylogeographic data, these methods make BiologicalNetworks a unique among other integration systems. BiologicalNetworks allows visualizing and comparing host-pathogen networks in respect to the pathogens phylogenetic distance and geographic origin  strains/isolates. In particular, host-pathogen interaction networks for two or more pathogen genomes can be compared, using both phylogenetic and geographical data Figure . Phylogen Figure .http://flu.sdsc.edu/integration.jsp , Cell journal, for example, now asks the authors to accompany their publications with graphical abstracts. Presenting the data in the form of a digital integrated model  instead of the graphical picture would be much more useful for researchers and our future work will be towards that direction.The ability to integrate Supplementary data for host-pathogen studies and to represent it in the digital integrated form is an extremely important feature for reproducible and integrated research. Several journals including We chose for comparison six resources: Cytoscape  and fiveNo two resources were similar by all 15 properties considered Table . Most ofDue to the graph-based data integration model and the Semantic Web technologies implemented in BiologicalNetworks , it is sBiologicalNetworks extensions for the host-pathogen studies enable diverse data in major human-disease systems to be subjected to efficient integrated analysis. The results show the utility of multi-scale data integration from large-scale human molecular-interaction, sequence and expression data to epidemiological and virulence data. The approach described allows information to be extracted that is not restricted to any one data type. Moreover, our analyses suggest how various host pathways act in response to viral infection, and serve as a large-scale window into the genomic response to Influenza and other respiratory infections. The pathways identified should provide insights into the mechanisms by which the host interacts with different pathogens, useful information about stage of disease, and selection of suitable targets for early diagnosis and treatments.BiologicalNetworks has general purpose graph architecture and is data-type-neutral. Therefore, there is the prospect of further integration of data such as detailed clinical data that will enable clinical variables to be associated quantitatively with the activities of molecular pathways and processes. Capacity for integration gives our system a unique capability, the full potential of which will be realizable when a multitude of host-virus interaction data are available, so that similarities and differences between the interaction networks can be interrogated across the phylogenetic distance for more accurate prediction of the potential virulence of a newly isolated virus identified only by its sequence. We believe that the methods and tools we have implemented and described here will allow for the efficient dynamic integration and analysis of diverse data in other disease systems.SK, MS, AG and MB contributed to system concept. SK, MS, YD and MB implemented the system and performed major programming work. MB, JP and AnR contributed to data analysis. This work was coordinated by AG, AnR, JP and MB. MB and JP wrote the manuscript. All authors read and approved the final manuscript.Methods. Detailed description of the methods and data types used in the BiologicalNetworks system for host-pathogen studies.Click here for file


Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic synovitis that progresses to destruction of cartilage and bone. Bone marrow (BM) cells have been shown to contribute to this pathogenesis. In this study, we compared differentially expressed molecules in BM cells from RA and osteoarthritis (OA) patients and analyzed abnormal regulatory networks to identify the role of BM cells in RA.Gene expression profiles (GEPs) in BM-derived mononuclear cells from 9 RA and 10 OA patients were obtained by DNA microarray. Up- and down-regulated genes were identified by comparing the GEPs from the two patient groups. Bioinformatics was performed by Expression Analysis Systemic Explorer (EASE) 2.0 based on gene ontology, followed by network pathway analysis with Ingenuity Pathways Analysis (IPA) 7.5.The BM mononuclear cells showed 764 up-regulated and 1,910 down-regulated genes in RA patients relative to the OA group. EASE revealed that the gene category response to external stimulus, which included the gene category immune response, was overrepresented by the up-regulated genes. So too were the gene categories signal transduction and phosphate metabolism. Down-regulated genes were dominantly classified in three gene categories: cell proliferation, which included mitotic cell cycle, DNA replication and chromosome cycle, and DNA metabolism. Most genes in these categories overlapped with each other. IPA analysis showed that the up-regulated genes in immune response were highly relevant to the antigen presentation pathway and to interferon signaling. The major histocompatibility complex (MHC) class I molecules, HLA-E, HLA-F, and HLA-G, tapasin (TAP) and TAP binding protein, both of which are involved in peptide antigen binding and presentation via MHC class I molecules, are depicted in the immune response molecule networks. Interferon gamma and interleukin 8 were overexpressed and found to play central roles in these networks.Abnormal regulatory networks in the immune response and cell cycle categories were identified in BM mononuclear cells from RA patients, indicating that the BM is pathologically involved in RA.


BiDaS server enables users to analyze their data and generate large amounts of: (i) Simulated DNA/RNA and aminoacid (AA) sequences following practically identical sequence and/or extracted feature distributions with the original data. (ii) Simulated numerical features, presenting identical distributions, while preserving the exact 2D or 3D between-feature correlations observed in the original data sets. The server can project the provided sequences to multidimensional feature spaces based on: (i) 38 DNA/RNA features describing conformational and physicochemical nucleotide sequence features from the B-DNA-VIDEO database, (ii) 122 DNA/RNA features based on conformational and thermodynamic dinucleotide properties from the DiProDB database and (iii) Pseudo-aminoacid composition of the initial sequences. To the best of our knowledge, this is the first available web-server that allows users to generate vast numbers of biological data sets with realistic characteristics, while keeping between-feature associations. These data sets can be used for a wide variety of current biological problems, such as the in-depth study of gene, transcript, peptide and protein groups/families; the creation of large data sets from just a few available members and the strengthening of machine learning classifiers. All simulations use advanced Monte Carlo sampling techniques. The BiDaS web-application is available at  The advent of new and powerful workstations and supercomputers has enabled systems biologists, computational biologists and bioinformaticians to design and implement complex biological models . NeverthRandSeq web server -generating web servers have been developed. Namely, in the b server , users ab server , allows b server  is a webb server  and rMotb server  that genBiDaS webserver represents a next generation of webservers, incorporating advanced simulation methods, such as hidden Markov models (HMMs) and Monte Carlo (MC) techniques. The implementation of a state-of-the-art simulation pipeline can be a resource- and time-demanding task. BiDaS simulator provides a user-friendly interface helping users to accurately generate data that follow evident, as well as hidden properties of the original data sets. Furthermore, the implemented 2D and 3D Monte Carlo Rejection techniques and methodologies, such as Cholesky decomposition, can provide an adapted framework for data generation that can keep high order correlations potentially present in the initial data.De novo simulation of Sequences, (ii) De novo simulation of Numerical Features and (iii) Sequence Driven simulation of Numerical Features. In addition, BiDaS offers a personalized user workspace, where uploaded and generated data are stored. The application front-end was implemented using html and PHP, whereas all computations and simulations have been developed in R (http://www.r-project.org/).The BiDaS web application is divided into three main sections: (i) De novo simulation of Sequences’: in this section, researchers are able to upload a file to the workspace that contains either DNA/RNA, or AA sequences in text or FASTA format. Following the file upload, the users can analyze the provided sequences and generate MC-simulated sequences having identical characteristics with the original samples, such as length distribution and nucleotide/AA compositional probabilities. To this end, the server analyzes and simulates original distributions using functions belonging to the logspline family. These distributions are subsequently sampled, to generate characteristics for each simulated sequence. The resulting MC-simulated sequences are created by using each feature instance (e.g. length and nucleotide/AA probability) as a blueprint. Moreover, this section enables the user to realize de novo sequence generation based on Hidden Markov models. The input sequences are initially aligned with the Clustal Omega algorithm  and not sequence data. In this section, users can directly upload numerical data sets, such as amino acid composition, hydrophobicity, polarizability, charge, Van der Waals volume, polarity, composition, transition and distribution properties, GC content or any other feature and sequence property. The web server can analyze and simulate data following identical feature distributions with the original data set (1D simulation), while safeguarding all 2D and 3D between-feature correlations observed in the provided data. Correlated features are generated using Cholesky decomposition. The web server in this section, as in all available modules, can help users analyze the original and simulated data, identify correlated features, detect normally distributed characteristics , and visualize the original data sets as well as all MC-simulated data. User uploaded data sets and generated data are stored into the personalized user workspace.‘‘Sequence Driven simulation of Numerical Features’: In this section, users having DNA/RNA and AA sequences can calculate up to ten 10 numerical features using pseudo-AA composition , 38 DNA/BiDaS can be used in various bioinformatics and computational biology studies in biomolecule level  or in a systems biology setting. Whenever the number of available samples is smaller than what is required to derive biologically meaningful results, to perform complex simulations, to formulate systems biology models or to implement classifiers, BiDaS is able to use the initially available group of gene, transcript and peptide sequences or any feature characteristics, and simulate vast numbers of samples with identical properties.For instance, a researcher is studying an experimentally identified small set of non-coding transcripts, presenting an enhancer-like function for protein-coding genes. However, this data set can be substantially smaller than what is required to train, test and validate a machine learning classifier, able to detect this transcript subclass. In this case, BiDaS can be used to derive an arbitrarily larger set of RNA molecules following similar composition, conformational features and any other user-defined properties, enabling the implementation of the transcript classifier model.MC AA sequences. Subsequently, MC-simulated sequences were differentiated as antimicrobial or non-antimicrobial, using specifically implemented machine learning classifiers. More precisely, two sets, comprising 1464 proteins annotated as antimicrobial and 3888 proteins annotated as non–anti-microbial, were collected from SwissProt database (Anti-Train) and 480 (SwissP-Anti-Test) sequences, respectively. The latter set was separated into two subsets, training and test each including 2000 (SwissP-NON-Anti-Train) and 1888 (SwissP-NON-Anti-Test) sequences, respectively. Fourth degree (4th) pseudo-AA composition features were calculated for all sets using the BiDaS online interface . Subsequently, three classification algorithms  with Gaussian and Exponential kernel) were implemented in R. All algorithms were trained using the same training data set (SwissP-Anti-Train) and validated against the SwissP-Anti-Test data set. For each classification scheme, Accuracy, F-Score and Matthews Correlation Coefficient (MCC) were estimated for all possible combinations of the selected four features.In a practical application scenario, BiDaS performance has been evaluated in real numerical and sequence data sets. In a first case study, the web server was used to generate Anti-Train and SwissP-NON-Anti-Train training data sets. Subsequently, 4th degree pseudo-AA composition were calculated. BiDaS MC-rejection method was used, rejecting all sequences not having identical pseudo-AA distributions and inter-feature correlations with the original data. Specifically, 154 563 MC-simulated anti-microbial (MC-Anti-Train) and 155 742 MC-simulated non-antimicrobial sequences (MCNON-Anti-Train) were finally generated that conformed to the selected criteria. Histograms of AAs within the initial and the MC-generated training data sets are presented in BiDaS web server was used to generate 400 000 antimicrobial and non-antimicrobial sequences (200 000 for each data set) following length distributions and AA compositional probabilities of the initial SwissP-The resulting features, derived from the positive  and negative  MC-simulated sequences, were used to train again a Bayesian and the two PNN classifiers (MC-Anti-Train and MC-NON-AntiTrain). The classifiers were trained using solely MC-generated features, without the inclusion of any of the original SwissProt-derived anti-microbial sequences. The performance of the classification algorithms was validated on the same test sets (SwissP-Anti-Test & SwissP-NON-Anti-Test) and with the same metrics used in the previous experiment. It is noteworthy that the performance of the classifiers being trained on real or simulated data was comparable. Detailed results are presented on De novo simulation of Numerical Features’ has been selected. The final set of the simulated data was composed of 4000 instances; half of them were generated with Cholesky Decomposition based on 2D and the rest on 3D correlations of the original features. Graphical representations of the simulated 2D, 3D correlated versus the original data are presented in In a second case study, BiDaS web server was used to simulate properties/features for a set of 153 3′untranslated regions (3′UTRs) of ribosomal protein-coding transcripts. The computations were performed on the transcript variants with the longest 3′UTR sequence. BiDaS has generated MC-simulated features based on an initial set of 22 conformational and thermodynamic dinucleotide properties derived from DiProDB database. In this case study, ‘BiDaS web server is designed to support Monte Carlo AA or DNA/RNA sequence generation, as well as numerical feature simulations. To our knowledge, there are no existing web applications with that extent of functionalities, as the currently available tools restrict their simulations to either sequence or feature-specific properties. We strongly believe that BiDaS will empower researchers with state-of-the-art sequence- and feature-generation tools, as well as simulation methodologies without the necessity to develop and maintain advanced computational pipelines. BiDaS will be a major tool when building complex biological models or developing computational intelligence schemes especially in cases with small or rare experimental/clinical data sets.Biomedical Research Foundation of the Academy of Athens. Emmanouil Athanasiadis and George Spyrou are supported by the NSRF 2007–2013, co-funded by the European Regional Development Fund and national resources, under grant “Cooperation” [No. 09ΣYN-11-675].Funding for open access charge: Conflict of interest statement. None declared.