The Boston Carpal Tunnel Questionnaire (BCTQ) is a disease-specific measure of self-reported symptom severity and functional status. It is frequently used in the reporting of outcomes from trials into interventions for carpal tunnel syndrome. We conducted a systematic review of published studies on the psychometric properties of the BCTQ to determine the level of evidence on the instrument's validity, reliability and responsiveness to date.A search of the databases Medline, CINAHL, AMED and PsychInfo was conducted to retrieve studies which investigated one or more of the psychometric properties of the BCTQ. Data abstraction was undertaken by the first two authors.Ten studies were retrieved which met the inclusion criteria. One study evaluated face and content validity (43 patients) eight studies assessed construct validity (932 patients), four studies tested reliability (126 patients) and nine studies assessed responsiveness (986 patients). Interpretability was evaluated in one study and acceptability in eight studies (978 patients).The BCTQ is a standardised, patient-based outcome measure of symptom severity and functional status in patients with carpal tunnel syndrome. The evidence base of the psychometric properties indicates that the BCTQ is a valid, reliable, responsive and acceptable instrument and should be included as a primary outcome measures in future CTS trials. The effectiveness of surgical and non-surgical interventions for carpal tunnel syndrome (CTS) has been investigated through randomised controlled trials and systematic reviews -4. The oThe Boston Carpal Tunnel Questionnaire (BCTQ), also referred to as the Levine scale, BrighamThe review considered all studies designed primarily to investigate an aspect of validity, reliability or responsiveness of the BCTQ in patients with carpal tunnel syndrome. We also considered any studies reporting on interpretability and patient acceptability of the BCTQ.The bibliographic databases Medline (1966–2005), CINAHL (1982–2005), AMED (1985–2005) and PsycINFO (1887–2005) were searched using the following MeSH terms: carpal tunnel syndrome, outcome assessment, questionnaires, psychometrics, validity, reliability, reproducibility, responsiveness. Keyword searches were also made using 'Boston Carpal Tunnel Questionnaire' and 'carpal tunnel instrument'. Bibliographies of the articles obtained were checked to identify any studies not retrieved through the electronic databases. The search was limited to English language articles only.The title and abstract of the articles retrieved were read and selected for inclusion if they fulfilled the following criteria: a prospective, observational study or clinical trial designed to evaluate validity, reliability, responsiveness, acceptability or interpretability of the BCTQ in patients with CTS. The full text was obtained for those articles which met the inclusion criteria.A data extraction form was developed and used to summarise information regarding the psychometric properties assessed including design, methods, sample and main results of those studies included see . Each arThe psychometric properties of outcome measures should be assessed by their face and content validity, construct validity, inter-tester and intra-tester reliability, responsiveness, interpretability, and acceptability and responder burden. A full The search yielded 21 hits. After reading the titles and abstracts, eleven studies were excluded because they did not include the BCTQ (n = 7); the BCTQ was used as a criterion measure for other instruments (n = 1); the BCTQ was applied as a measure to ascertain the incidence or severity of CTS (n = 2) or the BCTQ was compared against other diagnostic tests designed to detect CTS (n = 1).A total of ten studies which were primarily designed to evaluate one or several psychometric properties of the BCTQ were included -12,15-21Face and content validity were examined in a study which led to the original development of the BCTQ  construcOnly one study evaluated face and content validity of the BCTQ . The resThe BCTQ was compared with 12 other outcome measures for carpal tunnel syndrome to assess construct validity . The correlations between pinch and grip strength and the two subscales of the BCTQ were moderate with higher values for the Functional Status Scale than the Symptom Severity Scale, whereas sensibility measures showed a stronger association with the Symptom Severity Scale  were not significantly different from zero, and correlations were high . In a third study [Δ) were not significantly different from zero, and correlations were moderate . Test-retest reliability was assessed of the Spanish version of the BCTQ in a prospective study of 42 patients with confirmed CTS. Pearson correlation coefficients were reported as r = 0.87 for the Symptom Severity Scale and r = 0.85 for the Functional Status Scale [Test-retest reliability was reported in four studies. Levine et al  assessedrd study , the BCTus Scale .Responsiveness was assessed in prospective follow-up studies of surgical interventions only and reported as effect size (ES) or standard response mean (SRM) Table . The timAcceptability was examined in eight studies Table . The burInterpretability was assessed in a study including 196 subjects. Using tThe ten selected studies presented some strength. The studies sampled a wide age range of participants, which is desirable considering that items of relevance for the young and the elderly are incorporated ,22. SampLimitations must also be acknowledged. Firstly, none of the ten studies assessed all the psychometric properties, making comparisons difficult specially regarding face and content validity, acceptability, interpretability and reliability. Secondly, the factor structure of the BCTQ, an aspect of construct validity, has not been examined in the selected studies. Factor analysis is a method of assessing the construct validity of a questionnaire. In confirmatory factor analysis, the scores from each item in the scale would show high loadings, expressed as high Eigenvalues, on one of the predicted factors . It has been hypothesized that the BCTQ comprises a two-factor structure consistent with symptom severity and functional status . BecauseValidity of the BCTQ was assessed in terms of face, content and construct validity. Face and content validity were assessed through consultation with individuals with relevant expertise in order to generate the content of the questionnaire. The content of the BCTQ had been examined in one study, suggestResponsiveness to clinical change is another important feature of an outcome measure. The data on effect sizes and standard response means demonstrated that the Symptom Severity Scale and Functional Status Scales are able to detect clinically meaningful change resulting from the treatment for carpal tunnel syndrome and yielded large effect sizes over a 6 month interval. However in two of the studies the data on responsiveness were based on a subgroup of patients reporting greater satisfaction with surgery. Responsiveness indices in these are therefore likely to be larger than in the other studies. Using a responsive outcome measure will facilitate the detection of moderate treatment effects in clinical research.The BCTQ has shown good levels of acceptability with response rates of 90% and above and takes less than 10 minutes to complete. The interpretability has been assessed in relation to patient satisfaction with the outcomes of surgery and an overall difference of 0.74 has been designated as the minimally clinical important difference.This review considered English language publications only, however there are a number of published studies which have used translations of the BCTQ into other languages including Italian  Swedish Scale development is an ongoing process which may never be complete. The properties such as the validity, reliability and responsiveness investigated to date are not fixed properties but specific to the instrument used in a given situation and with a given population. The BCTQ was developed for use in heterogeneous samples of patients of a wide age range with CTS. Further research is needed to examine the consistency of its psychometric properties, with special attention to the factor structure, among specific populations, test-retest reliability using appropriate statistical measures and defining the MCID for each subscale against appropriate external criteria.Clinicians looking for a disease-specific measure for assessing pre- and post-operative symptom severity and functional status can be confident that the BCTQ is responsive to change, repeatable over time and that the scales measure what they purport to measure. The BCTQ is also acceptable and quick to administer and as it relies on self-report can be applied via postal methods.In summary, the BCTQ offers a standardised patient-based outcome measure of symptom severity and functional status for which there is good evidence on validity, reliability and responsiveness and it should be recommended for inclusion in future trials on carpal tunnel interventions.The author(s) declare that they have no competing interests.JCL carried out the searching and reviewing of studies and drafted the manuscript. CJH conceived of the original idea, obtained funding and participated in the search, review and drafting of the manuscript. FS provided expert advice on systematic reviewing and contributed to the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:Data extraction form (word document)Click here for fileDefinitions of psychometric properties (word document)Click here for file


Sudden cardiac death (SCD) is one of the leading causes of mortality in industrialized countries. Thus, identifying patients at high risk of SCD is an important goal. T wave alternans (TWA) is a new method for identifying patients with lethal ventricular tachyarrhythmias, and is dependent on heart rate. The maximal predictive accuracy is achieved at heart rates between 100 and 120 bpm, so that TWA is usually measured during exercise, phamacological stress, or atrial pacing. It has been shown that TWA has high sensitivity and negative predictive value for predicting SCD after myocardial infarction and is also useful for predicting SCD in patients with nonischemic cardiomyopathy. Although the implantable cardioverter defibrillator (ICD) is now the primary therapy for preventing SCD, it is difficult to identify those patients who are susceptible to lethal ventricular tachyarrhythmias for primary prevention. In the prediction of SCD, TWA can be used as a screening test of appropriate patients for further electrophysiological examination and therapy. Sudden cardiac death (SCD) is a leading cause of mortality and remains a major clinical challenge . ConcernRecently, assessment of repolarization alternans  in the electrocardiogram (ECG) has been suggested as a predictor of susceptibility to lethal ventricular tachyarrhythmias ,8. AlthoTWA was first described in 1908; it is the variation in vector and amplitude of the T wave that occurs on an every-other-beat basis . In a reThere is some evidence that TWA is linked to alternations in cellular calcium homeostasis, which significantly influence the action potential duration (APD) 1982 . In the Potassium channels may also play an important role in ischemia-induced TWA. The different sensitivity of KATP channel activation during ischemia between epicardium and endocardium may be linked to TWA at the cellular level 1982 -32.Currently, the hypothesis regarding the arrhythmogenic mechanisms associated with TWA is based on the concept that heterogeneous prolongation and increased dispersion of repolarization produce reentrant ventricular tachyarrhythmias 1982 . The hetDetection of microvolt TWA has been made possible by the use of advanced signal processing techniques and high-resolution electrodes to reduce noise. A number of beats, generally 128, are sampled and a time series of amplitudes of multiple corresponding points on the T wave are analyzed using a Fast Fourier Transform to generate a power spectrum . SeveralTWA is a rate-dependent phenomenon and microvolt TWA could develop in normal subjects at sufficiently high heart rate. It has been shown that the onset heart rate is relatively low in patients with structural heart disease and history of sustained ventricular tachyarrhythmia . Kavesh The original study of TWA was performed with atrial pacing to increase heart rate . Either The first large clinical study of TWA was performed by Rosenbaum and co-workers , who obsRegarding the prognostic utility of TWA in patients with a prior myocardial infarction (MI), only a few studies have been reported. TWA, SAECG, and LVEF were measured in 102 patients with recent MI (20±6 days after the onset of the MI) . A positThe sensitivity, NPV, and risk hazard of TWA for predicting SCD or VF were higher than LVEF, SAECG, and the presence of non-sustained ventricular tachycardia (VT); however, the specificity and PPV remained worse. The utility of TWA for prognosis in patients with recent MI needs to be further clarified.Adachi et al.  reportedKlingenheben et al.  studied  A study of 104 patients with DCM, and with 12 arrhythmic events during 21±14 months demonstrated that TWA in a group of patients with an onset heart rate less than 100 beats per minute was the most significant predictor of arrhythmia-free survival  .Studies of TWA in patients with other cardiomyopathies are more scarce. Momiyama et al.  studied Macroscopic TWA has been reported in patients with the long QT syndrome -22,34. PIn patients with the Brugada syndrome, some reports have revealed that intravenous administration of class Ic antiarrhythmic drugs induced macroscopic TWA and resulted in VF ,24. ThesThere are some technical and electrophysiologic limitations of TWA. The technical limitations are: 1) TWA cannot be measured in patients with atrial fibrillation, which is a common arrhythmia in patients with structural heart disease; and 2) the presence of frequent atrial or ventricular ectopy, excessive motion artifacts, and, in particular, the inability to achieve the target heart rate would render the results of the TWA test indeterminate. An incidence of indeterminate results of up to 25% is present in most of the published studies. The electrophysiologic limitation is that TWA testing may lose much of its predictive power within a few weeks after onset of MI.The clinical utility of TWA in evaluating arrhythmic risk stratification appears promising for patients with suggestive ventricular tachyarrhythmias, congestive heart failure or an LVEF of less than 40%, and a recent MI. The ultimate role of the TWA test as a noninvasive predictor of SCD awaits larger-scale prospective studies. In the near future, better definition of the clinical role of TWA test will be achieved with the results of the ongoing ABCD (Alternans Before Cardioverter Defibrillator) trial, in which ICD implantation will be performed in patients with ischemic cardiomyopathy and abnormal EPS and TWA results.


The occurrence of calcitonin-secreting primary carcinoid tumor of the urinary bladder is extremely rare.The case of a 68-year-old male with carcinoid tumor arising in the urinary bladder is presented. Transurethral resection of a polypoid small tumor 0.4 cm in diameter was performed. Immunohistochemical study using neuroendocrine markers allowed a straightforward diagnosis of a low-grade neuroendocrine carcinoma (carcinoid tumor) of the urinary bladder. Immunohistochemistry demonstrated calcitonin immunoreactivity in the most of the tumor cells.This tumor shows specific clinical, macroscopical and histological features and must be considered in the differential diagnosis of bladder neoplasms. The occurrence of primary carcinoid tumor of the urinary bladder is extremely rare. Two cases of these neoplasms have been reported recently by Martignoni and Eble . AccordiA 68-year-old man was admitted in January 2004 for a macroscopic hematuria. The patient had no relevant past medical or family history, and laboratory data were all within normal limits. A cystoscopic examination was performed and a transurethral resection of a tiny, sessile polypoid lesion of the bladder neck was carried out. Clinical evaluation included ultrasonography and computerized tomography (CT) scan. The patients was seen at last follow-up 14 months after diagnosis and he was completely negative.Grossly, the tumor was a well-circumscribed, small ovoid nodule of 0.4 cm in diameter. Histologically a polypoid tumor appeared to be covered by slightly attenuated normal urothelium. The tumor filled-up the lamina propria showing a predominantly glandular arrangement Fig , with scImmunohistochemically the tumor strongly reacted with synaptophysin Fig , chromogThe urinary bladder can be the site of tumors exhibiting various degrees of endocrine differentiation. It has become the most common site of extrapulmonary small cell undifferentiated carcinoma . The speThe present case shows clinically and microscopically the same features showed by two cases of carcinoid of urinary bladder reported recently by Martignoni and Eble . Their rAll 6 cases presented with hematuria, most of the patients aged of seven decade. All tumors were small, polypoid nodules at cystoscopy and most of them were localized in the neck of the bladder or trigone. All previously reported 6 tumors and the present case show a glandular architecture. Previous study, and our results have demonstrated the neuroendocrine differentiation. Strikingly the present case showed, immunohistochemically, a strongly reactivity with calcitonin. To the best of our knowledge this is the first case of a well-differentiated neuroendocrine carcinoma of the urinary bladder reported positive with this neuropeptide.Some tumors could simulate carcinoid of the bladder, in particular the nested variant of transitional cell carcinoma; differences in the cytologic features and immunohistochemistry should establish the diagnosis. The distinction between carcinoid tumor and inverted papilloma with glandular differentiation can be very difficult, however strong staining for neuroendocrine markers support the diagnosis of carcinoid tumor. Like the classic carcinoid tumors of the appendix, small bowel, or lung, all of the carcinoid tumors of the bladder are histologically and clinically not aggressive.In conclusion, we here add another case of carcinoid tumor of the urinary bladder to the existing literature, this tumor shows specific clinical, macroscopical and histological features and must be considered in the differential diagnosis of bladder neoplasms.The author(s) declare that they have no competing interests.MM participated to write the manuscriptVA operated the patient and participated to draft the manuscriptCM carried out imunohistochemical studyGN operated the patient and participated to draft the manuscriptGDR participated in the design of the studyLI participated in the design of the studyThe pre-publication history for this paper can be accessed here:


PLoS Medicine that is helpful in reconsidering the applicability of the WHO treatment guidelines.Heikens discusses a new study published in More than 10 million children under the age of five die each year, of whom 1.5 million are severely malnourished , represehttp://www.who.int/child-adolescent-health/integr.htm), have been developed to address the synergy between childhood malnutrition and infection, and these programs include nutritional rehabilitation  concerns possible missing pathophysiological elements in the concept of rehabilitation, as expressed in the WHO manual and the study by Maitland et al., which do not adequately address the sick child with multiple co-morbidity in sub-Saharan Africa and which we are still not able to comprehend. Adequate nursing  is critiI would like to suggest that five aggregated determinants characterise the unresolved problem in the case management of sick severely malnourished children in sub-Saharan Africa.Practitioners rehabilitating severely malnourished children should be aware of the important concept of reductive adaptation ,44 whichAs severely malnourished children often present with multi-organ failure, more intensive critical care might prove beneficial but would require more complex assessments  in order to establish the appropriateness of proposed new interventions.Detailed prospective clinical studies of co-morbidity are required with accurate documentation of both metabolic and clinical status, and response to different therapies, in the first few days following admission. These studies will form an essential basis for proposing changes in the early critical care of these children or in designing appropriate randomised controlled trials.Since the first draft of the WHO 1999 manual was circulated in the early 1990s, many specialists  have criAs argued by Collins  and BrieDuke et al. recently suggested that the process of WHO consultations be expanded through testing and piloting of the proposed guidelines in a variety of settings . Multi-c


A description of the diagnostic features of Miller Fisher syndrome.The clinical presentation, investigation, and subsequent progress of our patient with clinical unilateral external ophthalmoplegia.Our case demonstrates the presentation of clinical unilateral external ophthalmoplegia as part of the full triad of Miller Fisher syndrome. Miller Fisher syndrome was originally described in 1956 as a triad of total external ophthalmoplegia, ataxia and loss of tendon reflexes . Anti GQA 32-year old gentleman presented to the ophthalmic accident and emergency department with a four-day history of worsening horizontal and vertical diplopia. He denied any other ophthalmic or neurological symptoms at presentation. Other than a recent mild coryzal illness he was fit and well. He had no significant past ocular or medical history and took no regular medications.Ocular examination within the accident and emergency department showed a one-millimetre ptosis of the left upper lid, with limitation of left ocular movements in all directions of gaze. Right ocular movements were full. The remainder of the ocular and cranial nerve examination was normal. Eye movement testing showed a -2/-3 reduction in left ocular movements in all directions of gaze, with an associated +2 over-action in the corresponding right ocular movements. Figure A CT scan of his head with contrast showed no abnormality and the patient was transferred to the regional neurosciences centre for further investigation where the patient was found to be ataxic on heel to toe walk and the deep tendon reflexes were absent. Further investigation including an MRI, serology, lumbar puncture and Tensilon test were normal. Anti GQ1b IgG antibodies were positive with a significant titre by laboratory reference range, confirming the clinical diagnosis of Miller Fisher syndrome. With conservative management the patient's signs and symptoms resolved spontaneously within five weeks of presentation, leaving no permanent sequelae.The marked asymmetry of the ophthalmoplegia in this case is unusual, and to our knowledge only a single previous report of a clinical unilateral external ophthalmoplegia as part of the full triad of Miller Fisher syndrome has been documented within the literature . Acute pThis case demonstrates that uniocular external ophthalmoplegia can occur as part of the full triad in Miller Fisher syndrome. This reinforces the importance of considering systemic autoimmune diseases normally associated with bilateral ophthalmoparesis, in the differential diagnosis of a patient presenting with apparently unilateral signs.The author(s) declare that they have no competing interests.JS and LC drafted and referenced the case study.PS and RB revised the manuscript.All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


The completion of the sequencing of human, mouse and rat genomes and knowledge of cross-species gene homologies enables studies of differential gene expression in animal models. These types of studies have the potential to greatly enhance our understanding of diseases such as liver cancer in humans. Genes co-expressed across multiple species are most likely to have conserved functions. We have used various bioinformatics approaches to examine microarray expression profiles from liver neoplasms that arise in albumin-SV40 transgenic rats to elucidate genes, chromosome aberrations and pathways that might be associated with human liver cancer.In this study, we first identified 2223 differentially expressed genes by comparing gene expression profiles for two control, two adenoma and two carcinoma samples using an F-test. These genes were subsequently mapped to the rat chromosomes using a novel visualization tool, the Chromosome Plot. Using the same plot, we further mapped the significant genes to orthologous chromosomal locations in human and mouse. Many genes expressed in rat 1q that are amplified in rat liver cancer map to the human chromosomes 10, 11 and 19 and to the mouse chromosomes 7, 17 and 19, which have been implicated in studies of human and mouse liver cancer. Using Comparative Genomics Microarray Analysis (CGMA), we identified regions of potential aberrations in human. Lastly, a pathway analysis was conducted to predict altered human pathways based on statistical analysis and extrapolation from the rat data. All of the identified pathways have been known to be important in the etiology of human liver cancer, including cell cycle control, cell growth and differentiation, apoptosis, transcriptional regulation, and protein metabolism.The study demonstrates that the hepatic gene expression profiles from the albumin-SV40 transgenic rat model revealed genes, pathways and chromosome alterations consistent with experimental and clinical research in human liver cancer. The bioinformatics tools presented in this paper are essential for cross species extrapolation and mapping of microarray data, its analysis and interpretation. For decades, classical toxicology has used risk assessments based on animal studies for regulatory decisions. The underlying assumption is that important biological functions are often conserved across species. In continuation of this paradigm, the effort in toxicogenomics is placed on studying rodents and other surrogates using advanced genomics technologies, such as DNA microarrays. Microarray studies enable simultaneous measurement of the expression of large numbers of genes. Given the completion of the DNA sequence of the human, mouse and rat genomes -3, genesHepatocellular carcinoma (HCC) is a leading cause of death worldwide and, like most cancers, is a genetic disease caused by the accumulation of genetic and epigenetic cell alterations. The progression of hepatic neoplasia is characterized by increasing genetic instability, including duplication and deletion of parts of chromosomes and an increasing proliferative growth advantage of the affected cells. Molecular cytogenetic techniques, such as Comparative genomic hybridization (CGH) and Spectral karyotyping (SKY) -11, havea priori knowledge. Alternatively, significant pathways can be identified based on statistical analysis, potentially leading to new discoveries and a more complete interpretation of microarray data in the context of biological processes at the mechanistic level.Bioinformatics methods and tools are essential to analyze and interpret data from microarrays. The critical and urgent task is to associate altered patterns of gene expression with disease. Interpreting microarray data in the context of signaling and regulatory pathways is a particularly effective bioinformatics approach to transform data into biological meaning and to generate hypotheses for further research. Using pathways, disease mechanisms can be interpreted as disturbances of the intricate interconnections among genes, molecules and cells. Most reported pathway analysis of microarray data has examined the role of differentially expressed genes in pathways selected with The primary mechanism for the analysis of HCC is by the administration of carcinogenic agents. A number of model systems have been developed to understand the pathogenesis of primary liver cancer -15. AddiIn this manuscript, we describe a bioinformatics process where microarray data from the SV40 transgenic rat was examined for application to the study of HCC in human. We first used a novel visualization tool to investigate liver cancer by mapping chromosomal location of differentially expressed genes from the rat model to the chromosomal regions of human orthologs. Then, CGMA analysis was used to relate gene expression bias patterns to cytogenetic aberration profiles on human. Lastly, a statistical approach was used to identify several pathways involved in human HCC based on the rat microarray. The pathway analysis reveals that the expected involvement in apoptosis, cell cycle, growth and differentiation, genetic stability and methionine metabolism are important for cancer development, maintenance and progression. The results indicate that the gene expression profiles of the transgenic rat model may be useful in the study of human liver cancer.The details of the microarray experimental procedure is reported elsewhere. BrieflyThe log2 ratio-based mean global normalization was first applied and the normalized ratios of the swapped dye labels were then averaged. A total of 9150 genes remained for further analysis after removing non hybridized genes due to low intensity. Significantly differentially expressed genes were determined using an F-test with P < 0 .05.. ArrayTrack is bioinformatics software, where data management, analysis and interpretation are fully integrated [Most analyses were conducted using in-house software, ArrayTrack tegrated . ArrayTrtegrated . In this• Gene Orthology Analysis – The human and mouse orthologs to rat were obtained from the Orthologene Library in ArrayTrack. The content of the Orthologene Library are mainly derived from the NCBI HomoloGene database ,5. Array• Chromosome-based analysis – The cytogenetic locations of genes were exported directly from the Gene Library of ArrayTrack. A novel visualization tool, the Chromosome Plot, was developed to study the effect of a gene expression pattern on liver cancer through identifying the altered cytogenetic regions of each chromosome. Figure i:• Pathway analysis – The pathway data was obtained from the Pathway Library in ArrayTrack. The Pathway Library contains pathways from both the Kyoto Encyclopedia of Genes and Genomes (KEGG) and PathN is total number of genes on the chip , m is the number of differential expressed genes identified using the F-test , ni is the number of genes out of N that belong to pathway i while mi is the number of genes out of M differential expressed genes belong to pathway i. The two-sided Fisher's Exact Test p-value less than 0.05, suggest that the probability of significant genes in this pathway is not expected by chance alone.Where . Output human CGMA results contained 2728 genes from an input of 2925 Refseq genes. A Z-statistics of 1.96 corresponds to 95% confidence (that the expression bias in the chromosome was not due to chance) and 2.58 corresponds to 99% confidence.CGMA identifies cytogenetic regions containing unidirectional gene expression biases. The biased regions possibly indicate chromosomal gains and losses . Of the A total of 2223 differentially expressed genes was identified across three groups of samples  based on an F-test. The differentially expressed genes were first mapped to the rat chromosome. As depicted in Figure To investigate the cross-species extrapolation based on the results from the transgenic rat, the differentially expressed genes were first mapped to the orthologous chromosomal location of the mouse chromosomes. As depicted in Figure Table To further confirm the validity of cross-species extrapolation, we investigated chromosomal aberration in human based on the differentially expressed genes from the rat model using CGMA. Table We also investigated which pathways in human were significantly affected based on the differentially expressed genes identified in the transgenic rat model. Pathway analysis is a particular effective way to examine how the findings in the rat model relate to human in the context of biological functions. Table This study investigates the implications of using microarray results from the albumin-SV40 transgenic rat for the study of human liver cancer. We demonstrated the importance of bioinformatics to interpret microarray data for the cross-species comparisons. Specifically, two in-house bioinformatics tools are of importance for the analysis, the Chromosome Plot and ArrayTrack. The Chromosome Plot not only provides a visual presentation of the gene expression pattern at the level of gene order across chromosomes  human chromosomal regions that could be involved in liver cancer development, maintenance and progression. These chromosomal aberrations are consistent with the CGAP report for 10 out of 15 chromosome regions by karyotypes  and Integrin mediated signaling pathway have been reported to be over-expressed in human HCC, though the mechanism is not fully understood . In addiThe human-relevant liver cancer pathways based on SV 40 transgenic rat liver model are confirmed by reports on human liver cancer models. Therefore, the pathway analysis using Fisher's Exact Test is novel and efficient.We presented several bioinformatics approaches to extrapolate microarray data involving rat liver cancer to the human. Microarray has been widely used in many fields of medical and biological research. The current challenge of bioinformatics of microarray is no longer to identify a list of differentially expressed genes, but to develop effective bioinformatics processes and tools for data interpretation and knowledge discovery. In this study, we first developed a Chromosome Plot that provides a compact visual summary of gene expression data at the level of chromosomal location for identification of altered chromosomal regions. This tool facilitates cross-species comparison. The information available in ArrayTrack on gene ontology, gene orthologs and gene pathways was then used to interpret the microarray data. Finally, the CGMA bioinformatics tool was uesd to infer HCC chromosomal aberrations in the human based on microarray data from rat. The important lesson of this study is how to limit the information using bioinformatics resources and statistical means to present an unbiased  view to interpret microarray results with respect to genes, pathways, chromosomes and functions. Based on a thorough bioinformatics analysis, we found that the albumin-SV40 transgenic rat is a useful animal model for prediction of human liver cancer.HF did all calculations and analysis using bioinformatics tools, and wrote the first draft manuscript. WT guided the analysis and helped draft the manuscript. YD had the original idea for the liver cancer and liver toxicity study, provided original tissue, and designed the microarray analysis experiment; provided insightful information concerning liver cancer. HCP provided discussion and the transgenic rats. SHY provided the microarray data. JMY coordinated the collaboration and microarray experiment. WT, HH, QX, LS, XC and RP were involved in the discussion of the data analysis. RP and LS assisted with writing the manuscript. All authors read and approved the final manuscript.


Upper and lower limb robotic tools for neuro-rehabilitation are effective in reducing motor impairment but they are limited in their ability to improve real world function. There is a need to improve functional outcomes after robot-assisted therapy. Improvements in the effectiveness of these environments may be achieved by incorporating into their design and control strategies important elements key to inducing motor learning and cerebral plasticity such as mass-practice, feedback, task-engagement, and complex problem solving.This special issue presents nine articles. Novel strategies covered in this issue encourage more natural movements through the use of virtual reality and real objects and faster motor learning through the use of error feedback to guide acquisition of natural movements that are salient to real activities. In addition, several articles describe novel systems and techniques that use of custom and commercial games combined with new low-cost robot systems and a humanoid robot to embody the " supervisory presence" of the therapy as possible solutions to exercise compliance in under-supervised environments such as the home. Stroke is the leading cost of disability in the USA and rehabilitation is estimated to cost $60 billion annually for the 5.4 million living with disability. Neurological impairment after stroke frequently leads to hemiparesis or partial paralysis of one side of the body. This hemiparesis can profoundly impair functional performance of activities of daily living (ADLs) such as walking, running, and eating . For exaIncreasingly, robot-assisted therapy devices are used in stroke rehabilitation. Robotic tools provide opportunities to study functional adaptation after a stroke and can provide objective measurements of the time-course of changes in motor control of the affected limbs. Robot-assisted therapy permits semi-autonomous practice of therapeutic tasks -14.Early examples of upper limb robots such as the MIT-MANUS therapy robots  were desEarly examples of robotic lower limb robots are the GT I servo-controlled gait trainer developed and used for training in the 1990s in Germany ,9 and thPreliminary studies using these upper and lower limb robotic tools demonstrate their effectiveness and their limitations. The extent of motor impairment reduction seen after upper limb robot-assisted therapy environments has been shown to be dependent on lesion size and location, and the treatment has been shown to be target-area specific, e.g., training tasks emphasizing the shoulder will improve the shoulder but not the hand. In general, these upper arm systems have mixed impact on upper limb real-life function. They can reduce motor impairment after stroke, but still have mixed impact on functioning in real life use of the upper arm -4. New uThe mixed results from robot therapy environments, especially upper limb ones, suggest that there is still a need to optimize these treatment strategies and prove that rehabilitation robot systems are worth pursuing. If we believe this is true and that these systems have the potential to decrease long-term healthcare costs for patient, then we must clarify how best to design and use them. For answers rehabilitation engineers have begun to examine the neuroscience literature on cerebral plasticity to gain some insight into the next generation of robot therapy environments. The following briefly describes some of the relevant findings from neuro-rehabilitation and neuroscience and introduce nine articles that present new robots and new control models and feedback techniques to enrich robot-assisted therapy environments.The underlying neurological mechanisms and central nervous system recovery patterns after stroke therapy is poorly understood and this is true whether the intervention is mediated with robots or other strategies such as the Bobath method of Neuro-Development Therapy (NDT) . PrelimiSpecifically, functional imaging studies indicate that motor recovery is characterize by the following: 1) an increase in the size of the motor and sensory areas in the lesioned hemisphere that is dedicated to the impaired limb; 2) enhance activity and recruitment in preexisting motor networks in unaffected regions and those surrounding the lesion site and in the cerebellum, and 3) a reduction the amount of activity in primary and secondary motor regions over time, especially in areas in the hemisphere ipsilateral to the lesion ,28-32. SResearchers have begun to respond to the neurological evidence and have begun to create robot-assisted therapy environments that can better capitalize on these findings and improve the likelihood of use-dependent cortical reorganization and carryover to ADL function. In this special issue, we highlight several attempts to improve the effectiveness of robot therapy environments using several extrinsic motivational techniques including feedback. Figure The first set of two articles deals with lower limb robotics and demonstrate the use of biofeedback, virtual reality, and haptics to create more engaging gait training environments. The environments also provide opportunity for more complex and more functional gait training.The article by Lunenburger and colleagues  discuss The next set of four articles discuss new developments in upper limb robot-assisted stroke therapy from the point of view of using game- and social-oriented activities to define motivating training environments. The articles present strategies that seek to understand and improve the use of the impaired arm in daily activities in environments away from clinical supervision. In the past, robotic and computer-assisted systems such as JavaTherapy  and DrivJohnson, Feng, and colleagues  discuss In the article by Mataric, Eriksson, and colleagues  we gain Finally in this series, Amirabdollahian, Loureiro, and colleagues  discuss The next set of three articles describe solutions and ideas for improving the modeling and control of robot-assisted therapy systems to aide them in adapting patients' movements to natural and functional activities such as walking, drinking, and pinching. In the past other researchers have examined the use of error to improve motor adaptation for a point to point task after stroke . For theFinally, Wisneski and Johnson  suggest The special issue presented nine articles that seek to capitalize on new developments in neuro-rehabilitation after stroke to improve the effectiveness of robot-assisted stroke rehabilitation. Improvements may be achieved by providing robot training environments that incorporate into their design and control strategies important elements key to inducing motor learning and cerebral plasticity such as mass-practice, feedback, task-engagement, and complex problem solving. Novel design and control strategies covered in this issue provide new methods for training more natural movements, for inducing faster motor learning control of more complex movements salient to everyday activities, and for encouraging engagement and compliance in under-supervised environments such as the home and over-burdened clinics.The author(s) declare that they have no competing interests.MJJ was the primary composer of the manuscript and was responsible for the intellectual content of the manuscript and gave final approval of the version to be published.


One of the best examples is illustrated by Arvicolinae rodents , the most impressive mammalian radiation of the Northern Hemisphere which produced voles, lemmings and muskrats. Here, we compare the relative contribution of a nuclear marker – the exon 10 of the growth hormone receptor (GHR) gene – to the one of the mitochondrial CYB for inferring phylogenetic relationships among the major lineages of arvicoline rodents.Mitochondrial and nuclear genes have generally been employed for different purposes in molecular systematics, the former to resolve relationships within recently evolved groups and the latter to investigate phylogenies at a deeper level. In the case of rapid and recent evolutionary radiations, mitochondrial genes like cytochrome Prometheomys schaposnikowi) is one of the basalmost arvicolines, and confirm that true lemmings (Lemmus) and collared lemmings (Dicrostonyx) are not closely related as suggested by morphology. Red-backed voles (Myodini) are found as the sister-group of a clade encompassing water vole (Arvicola), snow vole (Chionomys), and meadow voles . Within the latter, no support is recovered for the generic recognition of Blanfordimys, Lasiopodomys, Neodon, and Phaiomys as suggested by morphology. Comparisons of parameter estimates for branch lengths, base composition, among sites rate heterogeneity, and GTR relative substitution rates indicate that CYB sequences consistently exhibit more heterogeneity among codon positions than GHR. By analyzing the contribution of each codon position to node resolution, we show that the apparent higher efficiency of GHR is due to their third positions. Although we focus on speciation events spanning the last 10 million years (Myr), CYB sequences display highly saturated codon positions contrary to the nuclear exon. Lastly, variable length bootstrap predicts a significant increase in resolution of arvicoline phylogeny through the sequencing of nuclear data in an order of magnitude three to five times greater than the size of GHR exon 10.The analysis of GHR sequences improves the overall resolution of the Arvicolinae phylogeny. Our results show that the Caucasian long-clawed vole  less heterogeneity among sites, and (ii) the presence of highly informative sites in third codon positions, that evolve rapidly enough to accumulate synapomorphies, but slow enough to avoid substitutional saturation. By proSeveral mtDNA-based studies of mammalian systematics have shed light on the difficulties experienced in using CYB to resolve phylogenies at lower taxonomic levels. Several works at the family level recovered multifurcations among genera or even species, which led to the conclusion of rapid, near-simultaneous divergences of multiple lineages (= star-phylogeny), without any time left for synapomorphies to accumulate in mtDNA. This interpretation has been put forward for some speciose clades of rodents, such as Ctenomyidae , EchimyiMicrotus and the inconsistency of their systematic treatment. Phylogenetic reconstructions based on mitochondrial sequences (CYB and NADH-dehydrogenase 4 [ND4]) provided unresolved topologies at two different levels: (1) among arvicoline genera, and (2) among species of Microtus 8 model p 3.6a2.1  of the PFollowing , sequencWe performed sensitivity analyses using PAUP* to assess the relative contribution of the various model parameters to the log-likelihood increase when more complex models are considered. To take the evolutionary properties of each codon position into account, nucleotide sites were categorized into each of the three codon positions. The procedure was conducted separately for CYB and GHR sequences. All sets of parameters – base composition (BC), substitution rate matrix (GTR), heterogeneity of substitution rate among sites (Γ), and branch lengths (BL) – were estimated independently for each codon partition. AIC values were compared to assess the significance of likelihood variation between global and partitioned models. For instance, to test the contribution of BC, only BC parameters were computed for each partition whereas other parameter values  were fixed for the whole gene.Lasiopodomys mandarinus was removed). The inferred number of substitutions was estimated from the ML tree as the sum of the branch lengths linking two terminals. The level of saturation was estimated by the slope (S) of the linear regression between the observed and inferred substitutions. Substitutions saturation is evidenced when the number of inferred substitutions increased, whereas the number of observed differences remained constant. We performed saturation analyses independently for each phylogenetic marker and for each codon position. ML branch lengths were obtained for each partition using PAUP* and by enforcing the topology as identical to the combined CYB + GHR tree.The nucleotide saturation of the phylogenetic markers was assessed graphically according to the procedure of , by plotTo compare the phylogenetic resolving power of GHR and CYB at varying taxonomic levels, we used the variable-length bootstrap (VLB). In this method, bootstrap support is estimated as a function of a variable number of resampled characters . For eacTG and EJPD initiated the study, assembled the data, designed and ran the calculations. TG contributed to collect specimens in the field, obtained DNA sequences, and wrote the manuscript under the supervision of EJPD. MKT intensively helped TG in DNA sequencing and sequence alignment. EP initiated the project on arvicoline phylogenetics and obtained financial support. He contributed with TG to collect specimens and assisted with drafting. PC and SS provided CYB sequences and helped to improve the manuscript. All authors read and approved the final manuscript.


The distribution of isoelectric point (pI) of proteins in a proteome is universal for all organisms. It is bimodal dividing the proteome into two sets of acidic and basic proteins. Different species however have different abundance of acidic and basic proteins that may be correlated with taxonomy, subcellular localization, ecological niche of organisms and proteome size.We have analysed 1784 proteomes encoded by chromosomes of Archaea, Bacteria, Eukaryota, and also mitochondria, plastids, prokaryotic plasmids, phages and viruses. We have found significant correlation in more than 95% of proteomes between the protein length and pI in proteomes – positive for acidic proteins and negative for the basic ones. Plastids, viruses and plasmids encode more basic proteomes while chromosomes of Archaea, Bacteria, Eukaryota, mitochondria and phages more acidic ones. Mitochondrial proteomes of Viridiplantae, Protista and Fungi are more basic than Metazoa. It results from the presence of basic proteins in the former proteomes and their absence from the latter ones and is related with reduction of metazoan genomes. Significant correlation was found between the pI bias of proteomes encoded by prokaryotic chromosomes and proteomes encoded by plasmids but there is no correlation between eukaryotic nuclear-coded proteomes and proteomes encoded by organelles. Detailed analyses of prokaryotic proteomes showed significant relationships between pI distribution and habitat, relation to the host cell and salinity of the environment, but no significant correlation with oxygen and temperature requirements. The salinity is positively correlated with acidicity of proteomes. Host-associated organisms and especially intracellular species have more basic proteomes than free-living ones. The higher rate of mutations accumulation in the intracellular parasites and endosymbionts is responsible for the basicity of their tiny proteomes that explains the observed positive correlation between the decrease of genome size and the increase of basicity of proteomes. The results indicate that even conserved proteins subjected to strong selectional constraints follow the global trend in the pI distribution.The distribution of pI of proteins in proteomes shows clear relationships with length of proteins, subcellular localization, taxonomy and ecology of organisms. The distribution is also strongly affected by mutational pressure especially in intracellular organisms. The abundance of genomic data allows for answering questions concerning the whole genomes' or proteomes' structure and evolution. One of the most intriguing recent findings is the distribution of isoelectric point (pI) of proteins in the whole proteomes. So called virtual 2D gels (i.e. plots where pI of proteins is plotted against their molecular weight) seem to be universal – in all proteomes they are usually bimodal with very low fractions of proteins with pI close to 7.4 -8. More Coxiella burnetti and Helicobacter pylori are connected with an adaptation to acidic environment [Different relationships between pI and other phenomena were discovered. Schwartz et al.  observedironment  and the ironment . On the ironment  have focironment  and Knigironment  found thironment  found thIn this paper we have broadened our analysis to proteomes of organelles, viruses and bacteriophages and applied different methods and parameters to describe quantitatively the pI distribution of proteomes. One of the aims of these studies is uncovering the relationship between the pI and the protein length. In the previous analyses, such correlations even if existing, were neglected. Furthermore, we have analysed the relations between the pI distributions of proteomes and the taxonomy and ecology of corresponding organisms considering different taxonomical levels and ecological signatures such as habitat, relation to the host cell, salinity, oxygen and temperature requirements. Moreover, we have tried to explain the observed relationship between the proteome size and the pI distribution of proteomes .Some examples of the distributions of isoelectric points of proteins coded by selected proteomes are shown in Fig. To simplify comparative analyses of the pI distribution of different proteomes, we have divided the whole sets of proteins into two sets called later the acidic and basic sets. The division point corresponds to the pI value for which the pI distribution reaches the minimum between acidic and basic sets (see the Methods section for details). For most of proteomes the division point was between 7.4 and 7.5. The distribution of the values of division points is very narrow – the range is 7.22 – 7.54 with the average and median 7.41. We have not found any correlation between the pH of these points and the taxonomical or ecological classification of organisms and genome sizes. The universality of the pH value of the minimum in pI distributions supports the conclusion of other authors that the bimodal distribution of pI results from intrinsic chemical properties of amino acids .For each proteome we have calculated the average pI separately for the basic and the acidic sets of proteins, the average length of proteins and the pI bias (see the Methods section). The pI bias simply describes the asymmetry of the bimodal distribution of pI Fig. . It rangThe 2D virtual distribution of proteins shows a characteristic pattern that may resemble a "butterfly"  or "lungE. coli proteins and the length of the real E. coli proteins; (ii) equal frequencies of amino acids and the length of the real E. coli proteins; (iii) the average amino acid composition of the E. coli proteins and the uniform length distribution in the range of the E. coli proteins.  the average amino acid composition of The results mean that the longer proteins can maintain the neutral or nearly neutral pI whereas the most extreme pI values are specific for shorter proteins. It probably results from purely statistical reasons. The shorter proteins show higher fluctuation of amino acid composition, which strongly influences their pI – incorporation of even one charged residue shifts significantly the protein pI to the lower or higher value. On the other hand, long proteins, usually composed of more charged amino acids much better buffer the effect of fluctuation in their composition and can keep their pI even close to 7.4. This 'statistical' explanation of the relationship between proteins size and pI does not exclude the possibility that this relation may have some biological consequences in a diversification of proteins' structure and function. For example, if there exists a selection constraint to generate a very acidic or very basic protein, the easiest way to accomplish that is "to make" it short. Actually, very basic proteins interacting with nucleic acids – ribosomal proteins – are usually very short. We think that more proteins subjected to such selection should be found. These proteins may belong to specific regulatory proteins, transcriptional factors, modulators, signalling proteins, small proteins interacting with other proteins etc.We have also noticed a difference in the length of acidic and basic proteins. The comparative analyses have shown that acidic proteins are significantly longer than basic ones  in more than 95% of Archaea, Bacteria and Eukaryota proteomes. The acidic proteins are on average 73 and 107 amino acid residues longer than the basic ones in the case of Prokaryota and Eukaryota, respectively. Probably it is connected with the presence of very short ribosomal proteins in the basic set and long aminoacyl-tRNA synthetases in the acidic one .Nandi et al.  observedIn the analysis of potential relationships between pI of proteomes and taxonomy we have used the pI bias that is quite concise but a very informative parameter emphasising the most distinct and basic differences in the pI distributions. Fig. Our analyses of prokaryotic proteomes performed on the lower taxonomical level did not show any relation between the pI bias and phylogeny, which is in agreement with the results of Knight et al. . Many moOn the other hand, we have found an interesting grouping related to a phylogenetic signal when analysing mitochondrial proteomes, which are much smaller and more conserved according to their protein content than large chromosome-encoded proteomes Fig. . The anaThe observed phylogenetic signal in the pI bias is probably the result of different composition of proteomes. Mitochondrial proteomes of Viridiplantae, Protista and Fungi are usually several times larger than Metazoa proteomes that contain usually 12–13 proteins. Actually, the most of proteins  that are absent from Metazoa but present in Viridiplantae, Protista and Fungi are very basic  responsible for their import into organelles. These peptides are cleaved off by organellar peptidases after the proteins are transported. Because these presequences are rich in basic amino acid residues, the pI of premature unprocessed proteins (i.e. the whole sequences) should be more basic than the pI of mature proteins (i.e. without transit peptide). In the Tab. Schwartz et al.  analyzed2 test was performed. Fig. To analyse the relationships between the pI of proteomes and ecology of organisms in details, all analysed prokaryotic proteomes were classified to different ecological subgroups (see the Methods section for details) and then compared in respect to the pI bias Tab. . The KruThe analyses showed that salinity is positively correlated with the acidity of proteomes – the more halophilic organisms have more acidic proteomes. It agrees with results of other authors who observed in halophiles predominance of acidic over basic residues -22 and lRickettsia conorii, Tropheryma, Wolbachia) reside and usually replicate in vacuoles or phagosomes. What is interesting, R. conorii has the least basic proteome within the Rickettsia genus and as the only representative of its genus was observed in a vacuole [Considering habitat preferences, host-associated organisms have the least acidic proteomes compared to other groups and aquatic bacteria possess the most acidic ones. In the 'acidic' class aquatic bacteria are the most overrepresented group and host-associated species the most underrepresented one Fig. . On the  vacuole . It woulEcological changes quite strongly and quickly influence the pI bias in the course of evolution of proteomes because the changes in the pI bias are seen even among closely related organisms. We have gathered all analysed species belonging to the same genus and having different ecological assignments  have changed their pI from acidic to basic during transition to intracellular way of life. Therefore, in contradiction to the mitochondrial proteomes, the observed shift of small bacterial proteomes towards basicity cannot be explained by the gain or loss of some proteins in the proteome but by the shift of pI of many proteins.We have considered if this hypothesis explains the basicity of small prokaryotic proteomes as well. We can exclude acquisition of basic proteins as the explanation because it is difficult to find any new genes gained by intracellular bacteria that would be absent from the closely related free-living relatives -34. UsuaE. coli example). Actually, according to the genetic code table, assuming equal frequencies of codons, representation of the acidic residues is lower than the basic ones. The relationship between the fraction of the basic or acidic amino acid and GC content is non-linear but any nucleotide composition favours basic residues especially for extreme GC content /(Nbasic+Nacidic), where Nacidic and Nbasic denote the numbers of proteins in the acidic and the basic sets, respectively.Proteomes were characterized by the average pI and the average length of proteins separately for the basic and the acidic sets of proteins and by the "pI bias" (b) describing the relation between the number of proteins in the basic set and the number of proteins in the acidic set: b = 100· (NThe different sets of proteomes were clustered by the UPGMA (Unweighted Pair-Group Method Arithmetic Averages) method based on the median of the pI bias. The clustering was performed with the neighbour program from the PHYLIP 3.6 package . To eval2 test.All analysed prokaryotic proteomes were classified according to five ecological signatures: habitat , relation to host cell , salinity , oxygen  and temperature requirements . The classification was based on the data published on the NCBI web site, papers related to the sequenced genomes and other sources. A given species was assigned only to one subgroup, the most typical for its ecological property. To analyse the relationship between the pI bias and ecological classification of proteomes the analysed proteomes of particular ecological subgroups were distributed among three classes of the pI bias . Then the observed numbers of proteomes in the given class were compared to the expected ones by χSets of orthologous proteins of prokaryotic organisms were downloaded from Microbial Genome Databse – MBGD .The non-parametric Kruskal-Wallis test and t-Student test were applied accordingly to determine statistical significance of tested hypotheses. The Benjamini-Hochberg multiple comparisons procedure for controlling the false discovery rate was used .JK started this work and generated preliminary results, carried out analyses, collected data about ecological classification of organisms and wrote scripts in Perl. MP supervised the project, wrote this manuscript, downloaded the data, carried out analyses and wrote scripts in Perl. DM wrote scripts in Perl and generated sequence data. MK wrote this manuscript and contributed substantially to the final manuscript. PB performed some statistical analyses and wrote scripts in the R environment. NP and KS collected data about ecological classification of organisms and generated sequence data. MRD wrote programs in C++ calculating isoelectric point and generating random amino acid sequences. SC drafted manuscript, provided overall guidance and interpretation of results. All authors read and approved the final manuscript.PI distribution for prokaryotic proteomes. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis). Black points represent the set of acidic proteins while grey ones – the set of basic proteins.Click here for filePI distribution for prokaryotic proteomes. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis). Black points represent the set of acidic proteins while grey ones – the set of basic proteins.Click here for filePI distribution for eukaryotic proteomes. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis). Black points represent the set of acidic proteins while grey ones – the set of basic proteins.Click here for fileRelationship between the average pI of proteins and the pI bias for acidic (A) and basic sets (B).Click here for fileEscherichia coli K12. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis).The real and artificial proteomes of Click here for filePI distribution of mitochondrial proteomes belonging to Fungi (A), Protista (B) and Viridiplantae (C) prepared separately for proteins common for Metazoa and the analyzed group (present in Metazoa) and absent from Metazoa.Click here for filePI distribution for nuclear-encoded proteins targeted to organelles: plastid and mitochondrion. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis). Black points represent the set of acidic proteins while grey ones – the set of basic proteins. Transit peptides, premature proteins (i.e. the whole sequence) and mature proteins (i.e. without transit peptide) were analysed separately. Mitochondrial proteomes were divided to various taxonomical groups.Click here for filePI distribution for proteomes of different subcellular localization. Left panel: histograms of pI values at 0.1 unit intervals ; right panel: relationships between the logarithm of length of proteins (Y axis) and their pI (Y axis). Black points represent the set of acidic proteins while grey ones – the set of basic proteins.Click here for fileEcological changes and their relationships with the pI bias of proteomes in closely related organisms.Click here for fileRelationship between the pI bias and logarithm of proteome size for archaeal, bacterial and eukaryotic organisms.Click here for fileCharacteristics of 39 sets of conserved orthologous proteins present in all selected 100 prokaryotic organisms. The correlation coefficient was calculated between the pI of proteins and the pI bias of the whole proteomes.Click here for fileDiscussion about relationships between pI and other phenomenaClick here for fileList of analysed proteomes.Click here for file


Rheumatic fever continues to put a significant burden on the health of low socio-economic populations in low and middle-income countries despite the near disappearance of the disease in the developed world over the past century. Antibiotics have long been thought of as an effective method for preventing the onset of acute rheumatic fever following a Group-A streptococcal (GAS) throat infection; however, their use has not been widely adopted in developing countries for the treatment of sore throats. We have used the tools of systematic review and meta-analysis to quantify the effectiveness of antibiotic treatment for sore throat, with symptoms suggestive of group A streptococcal (GAS) infection, for the primary prevention of acute rheumatic fever.Trials were identified through a systematic search of titles and abstracts found in the Cochrane Central Register of Controlled Trials , MEDLINE (1966–2003), EMBASE (1966–2003), and the reference lists of identified studies. The selection criteria included randomised or quasi-randomised controlled trials comparing the effectiveness of antibiotics versus no antibiotics for the prevention of rheumatic fever in patients presenting with a sore throat, with or without confirmation of GAS infection, and no history of rheumatic fever.Ten trials (n = 7665) were eligible for inclusion in this review. The methodological quality of the studies, in general, was poor. All of the included trials were conducted during the period of 1950 and 1961 and in 8 of the 10 trials the study population consisted of young adult males living on United States military bases. Fixed effects, meta-analysis revealed an overall protective effect for the use of antibiotics against acute rheumatic fever of 70% . The absolute risk reduction was 1.67% with an NNT of 53. When meta-analysis was restricted to include only trials evaluating penicillin, a protective effect of 80% was found  with an NNT of 60. The marginal cost of preventing one case of rheumatic fever by a single intramuscular injection of penicillin is approximately US$46 in South Africa.Antibiotics appear to be effective in reducing the incidence of acute rheumatic fever following an episode of suspected GAS pharyngitis. This effect may be achieved at relatively low cost if a single intramuscular penicillin injection is administered. Rheumatic fever is the most common cause of acquired heart disease in children and adults worldwide ,2. AcuteThe global burden of disease caused by rheumatic fever currently falls disproportionately on children living in the developing world. Rheumatic heart valve disease causes 400,000 deaths annually mainly among children and young adults living in developing countries. At least 12 million people are estimated to be currently affected by rheumatic heart disease with two million patients requiring repeated hospitalisation and one million requiring, often unaffordable, heart surgery in the next 5 to 20 years . In manyth century and was conducted primarily on adult populations living in the United States  or 1 major and 2 minor criteria  .Acceptable interventions included any antibiotic versus placebo or no treatment. Acceptable trials included the incidence of a first attack of acute rheumatic fever following throat infection as an outcome. Secondary outcomes that were noted but not required included adverse events to antibiotic use, adherence to antibiotic therapy and mortality associated with the first attack of rheumatic fever.Using a standardized data form, two reviewers  independently extracted data from each eligible trial. The methodological quality of the trials was assessed based on the method of randomisation, concealment of allocation, blinding and loss to follow-up. Additional data extracted from each trial included information on the study setting , on the participants , and on the details of the interventions.The results from each study were summarised using relative risk and a 95% confidence interval (CI). The chi-squared test for heterogeneity was used, along with the visual inspection of graphs to determine the level of between study variation. As the p-value associated with this test was large (>0.10), indicating low-levels of statistical heterogeneity between study results, a fixed-effect model was used to pool the data to estimate an overall effect. A subgroup analysis included only studies where penicillin was used as the intervention.Seventy seven potentially eligible studies were identified by the preliminary search. Based on their titles and abstracts (when available), 41 studies were retrieved for a thorough evaluation, performed independently by two reviewers  , not trials (5), no randomisation or method of randomisation not described (5), trial on secondary prevention of rheumatic fever (3), control group not used (3), retrospective study (2), patients included with a history of rheumatic fever (1), non-human subjects used (1).Ten studies were included in the review Table  16-25].-25.16-25All 10 studies conducted appropriately timed follow-up visits at 3–4 weeks following the initial onset of symptoms given the 3-week delay in the onset of clinical signs of rheumatic fever. Adherence to therapy was not measured in any of the studies, as all of the studies included were hospital-based and antibiotics were not self-administered. Approximately half of the studies mentioned side effects as an outcome; however, most were not quantified. Symptoms mentioned were pain at the site of injection, nausea and diarrhea.The methodological quality of the studies, in general, was poor. Nine out of the ten studies reviewed date back to the 1950's when randomised trial methods were still evolving and good guidelines for conducting trials were not yet available -24. RegaAll 10 studies evaluated the effectiveness of antibiotics in preventing acute rheumatic fever (n = 7665). Three thousand nine hundred and ninety six (3996) received antibiotic treatment and 3669 received either placebo or no specific treatment. As no statistical heterogeneity was present p = 0.28) we pooled the results of the individual studies using the fixed-effect model. Meta-analysis revealed a substantial protective effect against the onset of acute rheumatic fever in patients treated with antibiotics   infection is effective in reducing the attack rate of acute rheumatic fever by 70%. Intramuscular penicillin appears to reduce the attack rate by as much as 80%. There was one fewer case of acute rheumatic fever for every 50–60 patients treated with antibiotics. These findings suggest that antibiotic treatment can be effective for preventing acute rheumatic fever in a population with suspected GAS throat infection.The treatment of GAS pharyngitis is directed towards eradication of the bacteria from the upper respiratory tract. The infection can usually be eradicated by a single intramuscular injection of benzathine benzylpenicillin or by 10 days' treatment with oral penicillin . There iThe challenge for policymakers and clinicians is how best to apply these findings to a developing country setting where the risk of rheumatic fever persists. The most evident hindrance to the generalisability of these findings is the discrepancy in characteristics of the studied population and the population currently at risk for rheumatic fever. All of the included studies were conducted in a developed country setting a half century ago, and 8 out of 10 studies included only young adult males in their study population. Variables such as geographic location and age will affect the incidence of GAS infection as well as the attack rate of acute rheumatic fever within a population. These epidemiological differences across populations will in turn affect the number of cases needed to treat (NNT) with antibiotics in order to prevent one case of rheumatic fever. Unfortunately, trials similar to those included in this review have not been conducted in developing countries. However, the strong findings of this review do provide supportive evidence for the development of primary prevention programs to decrease the long-term sequelae associated with GAS pharyngitis in developing countries.The validity of applying the NNT found in this review to pediatric populations in developing countries is questionable due to probable differences in GAS infection rates. In populations with a high incidence of GAS infection, the proportion of patients with respiratory symptoms that are infected with GAS will be higher than in a setting of low GAS infection incidence. A higher GAS incidence will yield a lower NNT, under the assumption that antibiotics provide the same protective effect in both populations. In the population studied, the proportion of suspected GAS infections that were true positives was large given the high specificity of the presence of exudate in the oropharynx area, estimated at 70–90% . To imprIdentifying true cases of streptococcal pharyngitis is difficult in a developing country setting where laboratory confirmation of GAS infection is not readily available. Therefore there is a need to use a clinical prediction rule (CPR) for the diagnosis of GAS infection that takes into account these limitations while at the same time maximizing both sensitivity and specificity. Several CPR's have been developed and implemented worldwide for diagnosing streptococcal pharyngitis. A CPR put forth by the WHO Acute Respiratory Infection (ARI) Guideline suggests that both pharyngeal exudate and an enlarged and tender cervical node should be present . An evalIn developing countries, it is more important to detect and treat all possible cases of GAS infection than to prevent antibiotic treatment for sore throats not attributable to GAS infection. Therefore an appropriate CPR for this setting will have high sensitivity, despite the subsequent tradeoffs of a lowered specificity. It was found that a revised version of the WHO CPR that included either the presence of exudate or enlarged cervical nodes exhibited a sensitivity and specificity of 84% and 40% respectively when tested in a developing country setting . It shouIn the absence of accurate epidemiological data on streptococcal pharyngitis and rheumatic fever for the developing world, a formal cost-effectiveness analysis of the use of antibiotics for the primary prevention of rheumatic fever is not feasible. However, anecdotal evidence exists supporting the cost-effectiveness argument. The cost of one intramuscular injection of penicillin (1.2 million units) in South Africa is ~ R5.00. Using the NNT found in this review, one case of rheumatic fever will be prevented for every 60 patients receiving one intramuscular injection of penicillin for the treatment of suspected streptococcal pharyngitis. Therefore the marginal cost of preventing one case of rheumatic fever is R300.00, or US$46. A recent study in Sao Paulo, Brazil estimates the economic burden associated with rheumatic fever and its long-term sequelae in a low-income population to be over US$50 million. Furthermore, treatment costs for chronic rheumatic patients were estimated at US$319/patient/year [Intuitively, primary prevention is a more desirable outcome for the individual as well as the health sector. Primary prevention decreases the burden placed on patients and health facilities by decreasing reliance on time- and resource-intensive secondary prevention programs, the success of which is entirely dependent on patient compliance. However, secondary prevention programmes are currently thought to be a more cost-effective approach to RF/RHD prevention and therefore more deserving of scarce resources available to communities in poor countries. This study highlights the lack of reliable data required to accurately assess the cost-effectiveness of a primary prevention program. While there is some scattered data on the prevalence of RHD in developing countries, additional data on the incidence of RF in these settings as well as the incidence of GAS infections is still needed. Furthermore, the preliminary cost-analysis put forth by this study suggesting that primary prevention is likely to be cost-effective, when considered in light of the favorable outcome of primary prevention, calls for additional attention and evaluation. At this point, primary prevention should not be dismissed.It is important to note that the results of the included trials could have been distorted by either selection bias, detection bias, or both, arising from poor methodological quality. Few studies were adequately randomised, none used an appropriate method for concealing allocation, only four blinded the outcome assessors, and only five commented on subjects lost to follow-up. Empirical evidence suggests that trials with inadequate or unclear allocation concealment or lack of blinding, on average, lead to overestimation of the effects of interventions .Acute rheumatic fever is common among children living in poor socioeconomic conditions. The findings of this review support the notion that antibiotic treatment given to cases of suspected streptococcal pharyngitis is an effective and safe option for reducing the complication of acute rheumatic fever. In other parts of the world the incidence of acute rheumatic fever is so low that the risks of antibiotic use may outweigh the potential benefits. Evidence is needed concerning the effects of antibiotic therapy for preventing rheumatic fever and rheumatic heart disease in children living in developing countries. Given the overwhelmingly positive results in the trials included in this review, placebo-controlled trials could be considered unethical. Trials comparing antibiotic types, routes of administration, and treatment regimens within high-risk groups, however, should be considered. Additionally, the collection of baseline epidemiological data on GAS infections, rheumatic fever, and rheumatic heart disease, and compliance with existing guidelines  is needeThe author(s) declare that they have no competing interests.KAR and BMM contributed to all aspects of this study. JAV contributed to study design, data analysis and writing of the paper. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


The Chinese term "Hot Qi" is often used by parents to describe symptoms in their children. The current study was carried out to estimate the prevalence of using the Chinese term "Hot Qi" to describe symptoms in children by their parents and the symptomatology of "Hot Qi".A cross sectional survey by face-to-face interview with a semi-structured questionnaire was carried out in a public hospital and a private clinic in Hong Kong. The parental use of the term "Hot Qi", the symptoms of "Hot Qi" and the remedies used for "Hot Qi" were asked.Chrysanthemum morifolii, Lonicera japonica, Bombax malabaricum, Sophora japonica, and Plumeria rubra) (57.6%) or selfheal fruit spike (Prunella vulgaris) (42.4%).1060 pairs of children and parents were interviewed. 903 (85.1%) of parents claimed that they had employed the term "Hot Qi" to describe their children's symptoms. Age of children and place of birth of parents were the predictors of parents using the term "Hot Qi". Eye discharge (37.2%), sore throat (33.9%), halitosis(32.8%), constipation(31.0%), and irritable (21.2%) were the top five symptoms of "Hot Qi" in children. The top five remedies for "Hot Qi" were the increased consumption of water (86.8%), fruit (72.5%), soup (70.5%), and the use of herbal beverages "five-flower- tea" (a combination of several flowers such as "Hot Qi" is often used by Chinese parents to describe symptoms in their children in Hong Kong. Place of birth of parents and age of the children are main factors for parents to apply the term "Hot Qi" to describe symptoms of their children. The common symptoms of "Hot Qi" suggest infections or allergy. As an ex-British colony in China, Hong Kong sees a thriving practice in both Western and Traditional Chinese medicine (TCM). Chinese often adopt the jargons of TCM to describe the symptoms. It is the authors' observation that the Chinese term "Hot Qi" is often used by parents to describe symptoms in their children. The term "Hot" is used by TCM practitioners to describe a phenomenon but not a diagnosis per se. ,2 A bettThe current study was undertaken to find out the frequency of using the term "Hot Qi" in parents and the symptomatology of the term "Hot Qi". This included a questionnaire on the symptoms and treatments for "Hot Qi". We hypothesized that the term "Hot Qi" was used among the majority of Hong Kong Chinese parents to describe symptoms in their children and the usage was governed by the cultural background of parents and status of children, e.g. age and gender.All parents of patients who were in-patients or attending outpatient clinics of our department, a private paediatric clinic, as well as the nursing and secretarial staff of this hospital were recruited for the study. Parents were approached individually by one of the authors (FYK) and the purpose of the survey was explained. A semi-structured interview based on the questionnaire see  was condThe current study did not involve any medical intervention nor invasive intervention to the subjects and no ethical approval was deemed necessary under Hong Kong ethical framework.Parents were asked to express their view on the term "Hot Qi" as used on their children, one child per parent, based on a questionnaire (appendix 1). This questionnaire was developed by DKN and DC (a registered TCM practitioner). "Hot Qi" was investigated with the question: "Have you ever used the term "Hot Qi" to describe your child?" Parents who answered "yes" were asked to volunteer symptoms of "Hot Qi" that their children displayed and the remedies used.All analysis was done with statistical software . All continuous data were presented as mean and standard deviation. Age of children was compared between those who used the term "Hot Qi" and those who did not by Mann-Whitney U test. The categorical variables, included age group of parents, gender of parents, education level of parents, place of birth of parents, household income, gender of children were compared by Chi-squared test. Variables with significant difference between the two groups of parents were entered into a forward logistic regression model to predict the use of "Hot Qi" in parents. The demographical predictors of "Hot Qi" with adjusted odds ratio significantly larger than one were reported. Top five symptoms of "Hot Qi" and Top five remedies of "Hot Qi" were reported.Chrysanthemum morifolii, Lonicera japonica, Bombax malabaricum, Sophora japonica, and Plumeria rubra), "selfheal fruit spike" (Prunella vulgaris), appetite stimulant , Abrus herb (Abrus precatorius), Turtle Jelly, Mulberry leaf & Chrysanthemum flower tea, Bo Ying Compound , 24 taste herb tea, instant chrysanthemum tea and "Yin Chiao Chieh Tu Pien" . All significance tests were two sided, and a p < 0.05 was considered statistically significant.In those who reported use of the term "Hot Qi", we compared the mean age of children between those used and those who did not use a specific remedy by unpaired t-test. "Consumption of herbal products" was defined as any positive answer for "Five-flower-tea"  Nine hundred and three parents (85.2%) claimed that they had employed the term "Hot Qi" to describe their children's symptoms. Characteristics of the two groups of parents that used or did not use the term "Hot Qi' by parents were listed in Table There was no significant difference in gender, education level, and monthly household income distribution between the group of parents who employed the term "Hot Qi" and those who did not. Among children whose parents employed the term "Hot Qi" to describe them, there were 469 girls (44.2%) and mean age were 8.5 years (SD = 6.6). The children whose parents have used the term "Hot Qi" were significantly older. The frequency of the use of the term "Hot Qi" increased with older children. Figure In univariate analysis Table , age of Symptoms of "Hot Qi" were summarized in Fig. We studied the reasons for irritability in children Table  because Remedies used by parents for improving each of the aforementioned symptoms of "Hot Qi" were summarized in Table The types of remedies given by parents were affected by the ages of their children Table . Childre"Hot Qi" was commonly used by parents in Hong Kong to describe symptoms of their children. The current study shed light on the meanings of the term "Hot Qi" as used by the lay public in a Chinese society and the treatment employed for alleviation of "Hot Qi".Parents who were born in Hong Kong were more likely to use the term "Hot Qi" than those born in China and other part of the world. The association between "Hot Qi" and place of birth of the interviewed parent could be related to the intense advertisement of herbal beverages in the Hong Kong society. Herbal beverage companies promoted the concept of "Hot Qi" and advocated the effectiveness of herbal remedies in controlling "Hot Qi". The readily available Chinese herbal beverages in the market might have led to the widespread use of the term "Hot Qi". The other reason might be related to the fact that majority of adults in Hong Kong had first-hand experience with TCM ,5 and moThe age of the children was another parameter useful for predicting whether parents used the term "Hot Qi". Parents seem more inclined to use the term "Hot Qi" to describe older children, median age of 7.0-year vs. median age of 1.0-year. This could be because parents became more confident with their childcare experience as the children get older and would let their children consume herbal products at an older age.The most frequently mentioned symptoms of "Hot Qi" were eye discharge, sore throat, halitosis, constipation, and irritable mood. As allergic rhinitis with or without allergic rhinosinusitis is a common cause for nasal obstruction  / mouth For the fifth most frequently mentioned symptoms of "Hot Qi" – irritability – we looked at the associated symptoms. Irritability and its associated symptoms  suggested possible upper respiratory system infection or allergic rhinitis. Dry mucosa complaints may well be related to the anti-cholinergic drugs prescribed to control the nasal symptoms of upper respiratory tract or allergic rhinitis. An increased consumption of liquid and fruits may help relieve some of the symptoms related to dry mucosa, but may not be appropriate for curing the underlying diseases.Water, fruits, soups, and "five-flower-tea" and "selfheal fruit spike" were the top five remedies that parents used to alleviate the aforementioned symptoms of "Hot Qi". We found that these self-remedies used were generally appropriate for treating the symptoms (vide infra). Halitosis is caused by mouth breathing or poor dental hygiene or sinusitis. An increase in water consumption may help improve the dry mouth associated with mouth breathing and hence decrease the halitosis. Sore throat is usually caused by viral or bacterial infection, or by dry throat resulted from mouth breathing. Increased water and fruits consumption give the pharynx more fluid exposure to lessen the discomfort caused by sore throat. Constipation is associated with inadequate fluid intake and fibre intake. A greater intake of water and fruits would be effective to control constipation.Prunella vulgaris) could inhibit the production of nitric oxide from cultured murine macrophages, suggested anti-allergic and anti-inflammatory effects. [Chrysanthemum morifolium (a component of "five-flower-tea") was reported to have anti-inflammatory effects. [Lonicera japonica demonstrated potent suppression of nitric oxide and tumor necrosis factor-alpha production in an activated macrophage-like cell line. [Bombax malabaricum was reported to have analgesic, antioxidant [Helicobacter pylori effects. [Sophora Japonica had anti-inflammatory effect, anti-virus effect as well as protecting hepatocytes and anti-hepatic fibrosis in murine. [Plumeria rubra had molluscicidal, cytotoxic and antibacterial activity. [The Chinese herbal teas "five-flower-tea" and "selfheal fruit spike" were frequently used by parents for treating "Hot Qi". In TCM, both herbs are useful for clearing heat of the body and restoring the smooth flow of "Qi". Despite their popularity, we found no study that evaluated the beneficial effect of "self-heal fruit-spike" and "five-flower-tea" in human. In animal model study, the triterpenes extracted from selfheal fruit-spike  declare that they have no competing interests.FYP participated in interview and preparation of the manuscript; DKN participated in the conceptualization and design of the study and final approval of the manuscript to be published; CHC participated in statistical analysis of the data; WLY, DC, KLK and PYC participated in the design of the study.Brief description of the content: Questionnaire used in this studyClick here for file