Metabolomics and lipidomics are emerging methods for detailed phenotyping of small molecules in samples. It is hoped that such data will: (i) enhance baseline prediction of patient response to pharmacotherapies ; (ii) reveal changes in metabolites shortly after initiation of therapy that may predict patient response, including adverse effects, before routine biomarkers are altered; and( iii) give new insights into mechanisms of drug action, particularly where the results of a trial of a new agent were unexpected, and thus help future drug development. In these ways, metabolomics could enhance research findings from intervention studies. This narrative review provides an overview of metabolomics and lipidomics in early clinical intervention studies for investigation of mechanisms of drug action and prediction of drug response (both desired and undesired). We highlight early examples from drug intervention studies associated with cardiometabolic disease. Despite the strengths of such studies, particularly the use of state-of-the-art technologies and advanced statistical methods, currently published studies in the metabolomics arena are largely underpowered and should be considered as hypothesis-generating. In order for metabolomics to meaningfully improve stratified medicine approaches to patient treatment, there is a need for higher quality studies, with better exploitation of biobanks from randomized clinical trials i.e. with large sample size, adjudicated outcomes, standardized procedures, validation cohorts, comparison witth routine biochemistry and both active and control/placebo arms. On the basis of this review, and based on our research experience using clinically established biomarkers, we propose steps to more speedily advance this area of research towards potential clinical impact. Key MessagesMetabolomics applied to clinical trial samples can help elucidate drug mode of action, but relevant cardiometabolic examples are limited by small size and suboptimal study design.Clinical studies can also be used to identify novel predictors of important outcomes, in the same way as high-quality observational studies.Metabolomics in pre-treatment samples can be tested to predict response to therapy (both desired and undesired), but current data in cardiometabolic trials are limited.Early intervention metabolomics may identify changes in metabolites soon after initiation of therapy, which may predict later response to therapy or outcomes (both desired and undesired) but, again, current data from cardiometabolic trials are limited.Future studies of cardiometabolic trials would benefit from better design (with greater exploitation of placebo-controlled study samples and adjudicated outcomes), larger sample sizes, standardization of procedures, validation of biomarkers in new cohorts and better testing of predictive abilities against current benchmarks to demonstrate real clinical utility.,,1H-NMR) spectroscopy, utilizing targeted or untargeted methods, are the most commonly used techniques.,,Metabolomics is defined as the study of the metabolome:,,,et al., 2009.,Pharmacometabolomics, the application of metabolomics (and lipidomics) to the study and prediction of variation in drug response, is a relatively new direction.,,The advantage of randomized controlled trials (RCT) over observational studies is that they provide unconfounded estimates of the effect of an intervention. RCTs with appropriate clinical outcomes, for example cardiovascular disease (CVD) events linked to low-density lipoprotein–cholesterol (LDL-c) lowering, are described as having the highest level of evidence for evaluating causal pathways which integrate biomarkers.,post hoc observational analyses. It is with this background that researchers value stored biobanks from such large-scale clinical intervention studies to test novel biomarkers for clinical value.Another value of major trials is that they typically offer rich phenotyping of all participants including conventionally measured clinical biomarkers of interest. Therefore, the added information obtained through more costly or novel biomarker approaches, such as metabolomics, can be readily measured against conventional biomarkers. This provides a realistic benchmark to ensure that the novel biomarker(s) add value over and above routine measures and are cost effective.To anticipate where metabolomics might be impactful, and to make a useful comparison with existing metabolomics data, evidence for conventional biomarker use in clinical trials  ought to be considered.,,So far, the clinical impact of routine biomarkers to predict the degree of treatment response in many fields has been relatively modest.Regarding type 2 diabetes mellitus (T2DM), patients are heterogeneous in terms of weight, insulin sensitivity, beta cell function and renal and liver function. We have limited information on which baseline phenotypes predict treatment responses to a wide range of glucose-lowering drugs with very different mechanisms. Ongoing work, such as in the MASTERMIND study ,n = 946, 4.9 years follow-up), we noted that plasma levels of the liver enzyme alanine aminotransferase (ALT) and fasting triglyceride were the only significant independent predictors of the development of T2DM. As previous data had linked ALT levels to liver fat, these serial data were amongst the first to suggest that hepatic fat accumulation may increase before diabetes development.This article concentrates on metabolomic gains from trials (including uncontrolled intervention studies), but the utility of trial datasets or biobanks with the availability of serially recorded routine biochemistry measures from individuals with adjudicated events should not be underestimated. These have been used to inform on predictors of events or to identify patterns of disease. For example, by exploiting the availability of serial 6-monthly liver function tests from the West of Scotland Coronary Prevention Study (WOSCOPS)  than changes in the transcriptome or proteome.The fact that drugs are: (i) transported using metabolite transporters (particularly membrane transporters);,Measurement of only a few metabolites, for example routine or emerging biochemical biomarkers, is often inadequate to interrogate drug effects, since most drugs affect multiple interconnected metabolic pathways resulting in multiple metabolic changes.,,Genomics and pharmacogenomics studies have clearly demonstrated that genetics alone cannot explain all the variation in drug response.We now describe early examples of the use of metabolomics and/or lipidomics to investigate the effect of cardiometabolic disease-related pharmacotherapies on metabolite profiles in trials , or to predict variation in drug response, with some specific examples from the cardiometabolic arena . We excl,,,,All of the studies used state-of-the art analytical methods . Many ofpost hoc or hypothesis-generating studies , but little else . Only a ,Few studies compared metabolite-based biomarkers with routine measures .47–52,55post hoc analysis of interventional studies used for observational investigations (no longer placebo-controlled) or are based on analysis of single-arm interventional studies  and corresponding genetic instruments (HMGCR rs12916 variant known to mimic HMGCR inhibition).,,,,As is the case for many drugs, there is clinical variation in the effects of statins on LDL-cholesterol. In one study, decreases in LDL-c ranged from less than 5% to more than 60% on simvastatin 40 mg daily  even when concordance with therapy was apparently accounted for.Whereas these studies are of academic interest, our view is that the use of pharmacometabolomics in prediction of statin LDL-c-lowering response in clinical practice is likely to be impractical, given the huge numbers of patients eligible for statin therapy, the low cost of treatment, the safety of statins and the LDL-c lowering that is likely to occur in the vast majority. Patients routinely start on a standard statin dose (e.g. atorvastatin 20 mg/day for primary prevention and atorvastatin 80 mg/day for secondary prevention) unless there is potential for drug interaction or there appears a high risk of adverse events.,Certainly there is a variation in response to statins and this does correlate with efficacy in terms of CVD risk reduction.Most importantly, we note that there are currently no lipid-modifying therapies that can compete with statins in terms of both clinical and cost effectiveness. There is no evidence that a patient who does not respond well to a statin would respond better to ezetimibe or bile acid sequestrants. Withholding statin therapy based on pharmacometabolomics and treating with such alternatives would likely be unethical. New LDL-cholesterol lowering therapies such as PCSK9 inhibitors may be more widely prescribed in future .,,Despite our reservations concerning pharmacometabolomics for the purpose of predicting statin efficacy, we agree that pharmacometabolomics of statin therapy may have a role in identifying potential biomarkers of myopathy .9,75,84 ,1H-NMR metabolomics, narrowing the detectable dynamic range (ratio of highest versus lowest concentration: e.g. pM to mM). Problems with metabolite identification are common, partly due to limited availability of standards needed to confirm metabolite identity. Absolute quantitation, particularly in untargeted MS methods, is also problematic, again partially due to limited availability of stable isotope-labelled standards needed for quantification by MS due to variability in ion suppression and instrument stability. Inter-batch variation is an issue in MS metabolomics of studies with a large number of samples.1H-NMR metabolomics have been highlighted in a number of studies, and are generally described as complementary.,,There are many challenges in metabolomics and lipidomics .10,85 Mo,,,,,,,n).,13C-Labeled Extracts (MIRACLE)],,1H-NMR method described by Soininen et al., 2015, which has been used in a large number of observational studies.1H-NMR, the other metabolic measures being lipoprotein and lipid measures.,Some of these problems can be overcome, and considerable effort has gone into optimizing available methods. Primarily, the analytical platform that naturally best meets the requirements to answer a specific research question should be chosen. Decisions on which analytical technique to use will often be pragmatic, based on instrument availability, costs, sample volumes or the need for absolute quantitation, high sensitivity, high throughput or improved coverage.Accepting that every technique, including those in routine biochemistry, has its limitations, we expect that many of these issues will not necessarily be overcome but minimized. There will always be trade-off between sensitivity, metabolite coverage and precision.There are multiple challenges in the interpretation of metabolomics studies . There i,,,,False-positive results can be minimized by appropriate statistical correction for multiple comparisons and by validation of models.As clinical trial samples are very valuable and limited, careful use is essential. Most importantly, the studies need to be backed up by relevant clinical or mechanistic questions to avoid generating a series of results that, although of academic interest, do not advance clinical utility or provide mechanistic insights. This can be achieved by interdisciplinary work.,,Many of these obstacles  are not unique to metabolomics, and also apply to emerging chemistry-based markers, proteomics, transcriptomics and genomics.Analysis of existing larger trials as well as trials including a wide range of age groups, ethnicities and other characteristics, or at least representing the population the drugs are expected to be prescribed for, rather than only healthy volunteers.Use of data from existing trials with hard endpoints and not just surrogate endpoints. This allows testing whether pharmacometabolomics can predict the out-comes that really matter in clinical practice.Use of data from trials with a control arm: either placebo-controlled or a new therapy compared with the current gold-standard pharmacotherapy.Comparison of predictive abilities of potential new biomarkers versus gold-standard conventional biomarkers. Such an approach enables proper assessment of incremental utility gained by novel biomarkers over and above established tests.Standardization of methods, including collection and storage of samples, to lessen the effect of measurement artefacts on data.Implementation of fully validated methodology with absolute quantification, batch correction and confident identification where possible, to yield data that can be trusted, compared between studies and interpreted  more easily.Whenever possible, validation of any novel findings in an independent cohort to give added confidence that results are robust and generalizable.Economic analysis to estimate whether any incremental benefits justify the increase in costs or to determine realistic prices to enable clinical consideration.Dissemination, communication and working with a range of stakeholders: scientists, patients, clinicians, clinical scientists, regulatory bodies, expert committees, statisticians, health economists, pharmaceutical/biotechnology companies and biobanks. This ensures studies are relevant, well designed and conducted, and useful.The issues described above suggest that considerable work is required to better test the clinical utility of metabolomics in clinical trial datasets. We now provide a list of recommendations, in part mirroring a roadmap for implementation of proteomic biomarkers which is equally valid for metabolic biomarkers.,,Metabolomics has been applied to biobanked samples from clinical trials in recent years, but selected studies have typically been of a small scale and focused on surrogate outcomes, with the consequence that published evidence has indicated at best only modest potential. If such findings can be extended and properly validated, there remains a potential for metabolomics to aid better tailoring of drugs to patients . Accordingly, pharmacometabolomics has been described as a ‘potential gateway’ to stratified medicine.For real advances to be made, investigators with metabolomics expertise need to work with clinicians and statisticians to: (i) develop rigorous experimental study designs; (ii) to identify the best biobanks to exploit; and (iii) to carefully outline the key clinical questions that could be usefully addressed by metabolomics at the very beginning of the project.1H-NMR metabolomics is supported by: (i) the Chief Scientist Office, Scotland (CZB/4/613); (ii) the Wellcome Trust Institutional Strategic Support Fund (ISSF) (WT097821MF); (iii) the European Federation of Pharmaceutical Industries Associations (EFPIA) Innovative Medicines Initiative Joint Undertaking (EMIF) (grant number 115372); (iv) Chest, Heart & Stroke, Scotland (R13/A149); and (v) the European commission, under the Health Cooperation Work Programme of the 7th Framework Programme (grant number 305507) under Heart ‘OMics’ in AGEing (HOMAGE).N.R. is supported by the UPBEAT RCT mother-child study; stratifying and treating obese pregnant women to prevent adverse pregnancy, perinatal and longer term outcomes: Medical Research Council (MRC) (MR/L002477/1). Our recent work in Conflict of interest: N.S. has consulted for AstraZeneca, Bristol-Myers Squibb, Amgen, Sanofi and Boehringer.


Surveillance, Epidemiology, and End Results (SEER) public research database does not include chemotherapy data due to concerns for incomplete ascertainment. To compensate for perceived lack of data quality many researchers use SEER-Medicare linked data, limiting studies to persons over age 65. We sought to determine current SEER ascertainment of chemotherapy receipt in two relatively large SEER registries compared to patient-reported receipt and to assess patterns of under-ascertainment.In 2011–14, we surveyed patients with Stage III colorectal cancer reported to the Georgia and Metropolitan Detroit SEER registries. 1301/1909 eligible patients responded (68% response rate). Survey responses regarding treatment and sociodemographic factors were merged with SEER data. We compared patient-reported chemotherapy receipt with SEER recorded chemotherapy receipt. We estimated multivariable regression models to assess associations of under-ascertainment in SEER.n = 104) were under-ascertained in SEER (coded as not receiving chemotherapy). In unadjusted analyses, under-ascertainment was more common for older patients  and varied with SEER registries . On multivariable analyses, chemotherapy under-ascertainment did not vary significantly by any patient attributes.Eighty-five percent of patients reported chemotherapy receipt. Among those, 10% (We found a 10% rate of under-ascertainment of adjuvant chemotherapy for resected, stage III colorectal cancer in two SEER registries. Chemotherapy under-ascertainment did not disproportionately affect any patient subgroups. Use of SEER data from select registries is an important resource for researchers investigating contemporary chemotherapy receipt and outcomes. Since its inception in 1973, the National Cancer Institute’s Surveillance, Epidemiology, and End Results (SEER) program has collected data of critical importance to cancer epidemiology, policy, and health services research. Today, SEER data cover 30% of the US population, , 3 howevConsequently, most population-based studies of chemotherapy receipt have relied on claims linkages, specifically the SEER-Medicare linked dataset, and therefore have been restricted to patients aged 65 years and older. –10 WhileThe potential for under-ascertainment of chemotherapy by SEER registries is real, especially with the increasing use of oral oncolytics and other chemotherapies administered outside of the hospital setting that can be difficult for registrars to capture. If under-ascertainment is prevalent or varies by patient sociodemographic, clinical, or geographic factors, then studies that use SEER data alone to investigate chemotherapy risk inaccurate conclusions that could spur unnecessary changes in clinical practice or health policy. If under-ascertainment of chemotherapy is not a significant problem, however, SEER data could inform studies that would not be limited to elderly patients only.To further explore the current utility of SEER data as a stand-alone source for population-based studies of chemotherapy, we compared patient-reported to SEER Registry-reported receipt of adjuvant chemotherapy from two diverse population-based registries. We specifically surveyed patients with Stage III colorectal cancer, the third most common cancer diagnosis among men and women in the United States.  Among paAs previously detailed,  we identOur data collection procedures have been detailed previously.  Briefly,The study protocol was approved by the institutional review boards of the University of Michigan, Wayne State University, Emory University, the State of Michigan, and the State of Georgia Department of Public Health.n = 34), so that the self-reported measure of chemotherapy receipt in this study was considered positive only for patients who reported already receiving chemotherapy treatment.We defined the primary dependent variable, under-ascertainment of chemotherapy, as a Boolean variable where true was defined by positive patient-reported receipt of chemotherapy and SEER-coded non-receipt of chemotherapy. We defined a secondary outcome, over-ascertainment of chemotherapy, as a Boolean variable where true was defined as negative patient-reported receipt of chemotherapy and SEER-coded receipt of chemotherapy. We measured chemotherapy receipt by asking: “Did you or are you going to have chemotherapy to treat your colorectal cancer?” The accuracy of self-report of cancer treatments, including chemotherapy, has been validated in previous studies. –29 We alIndependent variables included age, gender, marital status, race, comorbid conditions, insurance status, annual household income, educational attainment, and area-level SES index . Among the 19% of cases missing patient-reported annual household income, we performed multiple imputation. Values for missing income were imputed using sequential multiple imputation. Five multiply imputed datasets were analyzed and the results combined to account for additional uncertainty due to imputation.  These reWe assessed primary disease site (colon versus rectum) via the SEER registry using ICD-O-3 Site codes  and excluding ICD-O-3 histology codes 9050–5, 9140, and 9590–9992. SEER registries used data from the American Hospital Association Database to identify the hospital where the colorectal cancer surgery was performed, and we categorized hospitals based on bed size collapsed into tertiles. We also ascertained the hospital nurse:bed ratio , Joint Commission on Accreditation of Healthcare Organizations accreditation, American College of Surgeons cancer program, Accreditation Council for Graduate Medical Education residency training program, medical school affiliation, and Council of Teaching Hospitals status  but did not include these variables in our final analyses as they were not significantly associated with under-ascertainment in either bivariate or multivariable analyses.To determine the chemotherapy receipt status in registry data, we used the “RX_SUMM_CHEMO” variable, which indicates any receipt of chemotherapy as part of initial therapy in SEER . SEER defines adjuvant chemotherapy as postoperative first-course chemotherapy which ends when the documented treatment plan is completed or at disease progression, recurrence, or treatment failure. Individuals coded as 00 (none) or 85–87 (chemotherapy was not administered) were categorized as not receiving chemotherapy; those assigned codes 01–03  were categorized as receiving chemotherapy. The 4% who were coded as 88  and 99 (unknown if administered) were excluded from analyses of under-ascertainment.We compared patient-reported and SEER registry-reported receipt of chemotherapy. We then described the frequency of chemotherapy under-ascertainment at each SEER site after grouping patients by clinical and sociodemographic characteristics, as well as by treatment and hospital characteristics.2 tests. We then regressed registry under-ascertainment on age, sex, race, marital status, number of comorbid conditions, insurance status, primary tumor site, and SEER region, adjusting for clustering by hospital. Because none of the hospital attributes were significantly associated with under-ascertainment, we omitted these from the final regression models. We evaluated all first-order interactions between significant variables; none were significant except as reported. Survey nonresponse was significantly greater among older, nonwhite, and rectal cancer patients. To adjust for this, response weights were created as inverse probability weights derived from a logistic regression of survey response. The weights were normalized to equal the observed sample size and were incorporated in all multivariable models. [Univariate analyses were performed using Pearson’s χ models.  Results We identified 2168 patients with Stage III colorectal cancer reported to the SEER registries of Georgia and Detroit, Michigan using rapid case ascertainment. Among these, 259 (12%) were later determined to be ineligible . Of the 1909 eligible patients included in the final sample, 608 could not be located or did not return the survey, leaving a sample of 1301 patients (68% survey response rate). Of these patients, 48 had missing data in SEER regarding chemotherapy receipt. Thus, our analytic sample was comprised of 1253 patients.Among the entire analytic sample, 1068 patients self-reported receipt of adjuvant chemotherapy (85% chemotherapy receipt rate). Among patients who self-reported receipt of chemotherapy, 104 (10%) were coded as not receiving chemotherapy in SEER (under-ascertained). Among patients who self-reported non-receipt of chemotherapy, 33 (18%) were coded as receiving chemotherapy in SEER. (Table P = 0.01) and in patients living in Detroit compared with the State of Georgia   Table . VariablWe have shown a 10% rate of under-ascertainment of adjuvant chemotherapy among patients with resected, stage III colorectal cancer in the SEER registries participating in this study. In multivariable analyses, under-ascertainment did not vary by patient sociodemographic factors.A recent study comparing rates of chemotherapy ascertainment in SEER to those in SEER-Medicare reported a relatively low sensitivity of chemotherapy ascertainment in SEER, and cautioned against the use of SEER for studies of chemotherapy use.  However,In contrast, 85% of our study respondents reported receipt of adjuvant chemotherapy for Stage III colorectal cancer, a cancer in which clear guidelines for the use of adjuvant chemotherapy exist. We found high sensitivity of SEER to identify patients who received chemotherapy (90% sensitivity) and a high positive predictive value (97%), indicating that among those identified in SEER as receiving chemotherapy, the vast majority also self-reported receipt of chemotherapy. Our study includes younger patients and is therefore more representative of the total population of colorectal cancer patients. Additionally, we were able to include patients with both Medicare and non-Medicare health insurance and found no significant differences in chemotherapy ascertainment by insurance type. We included patients diagnosed between 2011 and 2013, providing a view into modern use of adjuvant chemotherapy. In recent years adjuvant chemotherapy for stage III colon cancer has been one of the most widely accepted and utilized quality of care measures , 32 and Because the concern for under-ascertainment of chemotherapy in SEER has limited its use as a stand-alone database for population-based studies of chemotherapy, we focused our study on under-ascertainment as the primary outcome. To expand upon the understanding of the accuracy of chemotherapy ascertainment in SEER, however, we looked at possible causes of chemotherapy over-ascertainment in our study population as a secondary outcome. We found that most patients who self-reported that they did not receive chemotherapy were prescribed oral capecitabine. A few patients started chemotherapy but stopped after one dose, and it is possible that some patients had not yet started chemotherapy at the time they completed the survey, answered “no” to the survey item regarding receipt of chemotherapy, but then started chemotherapy shortly afterwards.There are several ways in which SEER registries can improve the accuracy of chemotherapy ascertainment. SEER registries collect chemotherapy data through both passive and active means where possible. Cancer is a reportable disease in all states and the foundation of the surveillance system in the United States was built upon hospital reporting. Hospitals directly collect data on patients receiving chemotherapy at their own facility and additionally attempt to capture chemotherapy on patients receiving some diagnostic and treatment services at their facility who go elsewhere for receipt of medical oncology care. These data are reported electronically to SEER Registries on an ongoing basis (passive collection by SEER). Delivery of chemotherapy outside the hospital setting is often not actively reported and in those situations an attempt is made by SEER staff to collect these data either through remote access to free-standing medical oncology practices, through direct abstracting at the practice, or through other means of follow-back to those facilities (active collection by SEER). This is a resource intensive process, however, that offers enormous challenges as the number of these practices has increased over time. Some registries do receive administrative claims or other electronic files from medical oncology practices as their means of meeting state reporting requirements and a greater emphasis on collecting these data through automated means is in place.Both of the registries included in our study represent a large proportion of American College of Surgeons Commission on Cancer (CoC) facilities. CoC facilities have invested sizable effort into trying to capture chemotherapy administered outside of the hospital to inform quality measures around standards of care, which may in part explain the low under-ascertainment. , 34 AddiIn our study, focused on a specific cancer for which well-established quality of care metrics exist, we have shown low rates of under-ascertainment of adjuvant chemotherapy by SEER registries. Perhaps, rather than an “all or none” approach to making chemotherapy data available to researchers, a more nuanced approach could be considered where specific cancers are individually evaluated with respect to treatment ascertainment and cancer-specific datasets are made available. Methods like these would allow for studies of chemotherapy receipt among a more complete population of patients, including those under age 65 and those with non-Medicare insurance.Another benefit to use of SEER registry data is that receipt of oral chemotherapy agents is recorded. In stage III colorectal cancer oral chemotherapy, namely capecitabine, has become a common component of adjuvant therapy. , 40 WhilOur study is subject to several limitations inherent to survey research. Analyses were limited by the sample of respondents. We note, however, that the population-based sampling achieved broad demographic representation and the 68% response rate is higher than any previous published cohort of patients with colorectal cancer.  The survWhile SEER data regarding chemotherapy are available to researchers upon request, it comes with a warning that it is not appropriate for studies of treatment patterns, disparities in receipt of chemotherapy, or comparisons of patient outcomes by receipt of chemotherapy. In a robust study of a diverse population of patients with Stage III colorectal cancer using self-reported receipt of chemotherapy, we have shown a low rate of chemotherapy under-ascertainment among two large SEER registries. In addition, no variation in under-ascertainment was identified by subgroups. Further exploration is needed to determine if the patterns observed in this study carry over to other cancers with defined quality metrics around use of chemotherapy. Use of population-based registry data for select cancers with high chemotherapy ascertainment could be an important resource to researchers investigating modern day chemotherapy receipt and outcomes, providing critically important insight to clinicians and policy makers and informing patient-centered quality improvement initiatives.


One of the major shortcomings in the current treatment of schizophrenia is that we have no valid criteria in clinical practice to decide which antipsychotic treatment should be chosen first. This is why we need to define a blood-based biological signature of treatment response that can be easily tested at patient bedside and would also help to identify molecular mechanisms of treatment response by determining biological changes associated with symptom improvements.Through a European consortium on Optimization of Treatment and Management of Schizophrenia in Europe , we conducted a clinical trial on treatment response with Amisulpride in 500 subjects with first episode psychosis. For each patient, biological samples  have been collected before treatment and during follow-up visits at weeks 4, 10 and 22, to measure biological changes associated with treatment initiation and with symptoms improvements. We combined multiple high-throughput technologies for transcriptome, genome, metabolome, proteome analyses before and after treatment.The transcriptome analysis conducted on 10,683 genes expressed in peripheral mononuclear cells identified significantly more genes differentially expressed after treatment in 112 patients who will be in remission after 4-weeks treatment than in 51 non-remitters. Using interaction network analysis, we identified biological pathways affected by Amisulpride. For some genes, the expression level was significantly correlated with symptom improvement. Moreover, some genes were already differentially expressed before treatment between remitters and non-remitters, suggesting they might be used to predict treatment outcome. In addition, we identified genetic variations associated with gene expression level and thus may explain individual difference in treatment response.In parallel, as recent biological data have suggested a preponderant role of innate and adaptive immune system in the vulnerability to schizophrenia or in antipsychotic treatment response , we paid a particular attention to the analysis of inflammatory markers and the presence of auto-antibodies in patients’ sera. Circulating autoantibodies against glutamatergic N-methyl-D-aspartate receptor (NMDAR-Ab) have been reported in up to 10% of patients with psychotic disorders. In our study, we demonstrated the advantage of using cutting-edge methods to ascertain the presence of NMDAR-Ab in seropositive patients that cannot be clinically identified . Indeed, the only clinical characteristics found in NMDAR-Ab seropositive patients, was the high frequency of female patients, the presence of mild neurologic symptoms and signs of antipsychotic intolerance. In addition, using an advanced statistical classification algorithm, we defined clinically-based subgroups of patients who had specific cytokine signature associated to remission after 4-weeks of treatment, suggesting that these markers may be used to predict treatment response.Altogether, our multilevel biological approach resulted in the identification of promising biomarkers, which may be used both to predict drug response and remission in first psychosis episode.


Treatment of cancer is frequently unsuccessful related to the loss of apoptotic signaling in malignant cells. This is a particular problem for high-grade gliomas, such as Glioblastoma Multiforme (GBM), which are almost universally fatal within a year or so of diagnosis. Novel therapies that capitalize on non-apoptotic cell death pathways may yield more effective outcomes, if their underlying mechanisms can be more completely deciphered. In a recent publication ref , the mec Due to its high malignancy, traditional treatments including surgical removal, chemotherapy or radiotherapy do not currently lead to cures . Since the term “apoptosis” was first coined in 1972 , an ever-growing field of interest in cancer therapy has been associated with pharmacologically stimulating apoptotic death in tumor cells. However, frequent gene alterations in cancer cells give rise to resistance to apoptosis, which results in the eventual failure of certain chemotherapeutic drugs , 5. Hence, deploying alternative cell death pathways to eradicate tumor cells could lead to new breakthroughs in cancer treatment. Cyclophilins are a group of chaperones that function as peptidyl-prolyl isomerases (PPIases), whose main function is to catalyze the conversion of proline from cis to trans isomers , in many subcellular compartments. Allowing proline conformational changes is thought to be critically important for proper protein folding to occur . Increased expression of cyclophilins has been found to support viability of many different types of cancer cells , while normal cells seem to be relatively independent of cyclophilins. Previous studies from our laboratory illustrated that cyclophilin B is overexpressed in many cases of GBM, and that genetic depletion of cyclophilin B lead to cell death and elevated ER stress . However, the death mechanism induced by cyclophilin inhibition in GBM cells remains elusive.Glioblastoma Multiforme (GBM) is one of the most frequent brain tumors in adults . We found that NIM811 treated GBM cells established a distinct cell morphology featuring the formation of huge cytoplasmic vacuoles surrounding an intact nucleus. Vacuoles were determined to be of endoplasmic reticulum (ER) origin, which fit the definition of paraptotic cell death , 12. In vivo experimentation further confirmed the potency of NIM811 in restricting tumor growth. To obtain a better understanding of cellular pathways that were perturbed by NIM811, we performed RNA-seq to compare the gene status of drug treated cells with controls. Importantly, genes that are responsible for maintaining ER homeostasis were greatly upregulated following NIM811 treatment. Moreover, autophagy related genes were also increased significantly. Western blotting at various time points revealed that the unfolded protein response (UPR) and autophagy were transiently activated but became compromised during prolonged NIM811 incubation. Since NIM811 did not impair proteasome activity, we utilized a bicistronic reporter plasmid to investigate the protein translation status of NIM811 treated cells. By separately quantitating Cap-dependent and Cap-independent translation, we found that the compound increased both types of protein synthesis after 48hrs of exposure, thus contributing to the protein burden in the ER. Remarkably, a brief pre-treatment with the autophagy activators rapamycin or torin-2 only caused significant down-regulation of Cap-dependent translation. In this context, UPR-favored Cap-independent translation was spared. Rapamycin and torin-2 not only reduced ER vacuolization caused by NIM811, but also provided cells with a substantial long-term survival benefit based on colony forming assays. On the contrary, cycloheximide, which has typically been considered a paraptosis inhibitor –14 only transiently delayed vacuoles formation but actually elicited dramatically more cell death following prolonged NIM811 treatment. Finally, shRNA mediated depletion of cyclophilin A or B in GBM cells cause them to become more sensitive to NIM811, thus reinforcing that NIM811 induced paraptosis was cyclophilin-dependent.In our recently published study, we applied the small molecule cyclophilin inhibitor NIM811 to GBM cells and elucidated a novel non-apoptotic cell death mechanism, which is dependent on inhibition of cyclophilins We propose that NIM811 triggers a series of cellular events, starting with the generation of misfolded or unfolded protein in ER. Initially, the cellular response was to activate the UPR and autophagy to facilitate unfolded protein clearance. Within a short time period, however, these two mechanisms become persistently inactivated and protein aggregates begin to accumulate in the ER. To minimize cellular damage, the ER appears to form compartments to isolate these improperly folded proteins, and this process is observed as cellular vacuolization. Since NIM811 fosters escalating protein translation, ER stress concordantly increases eventually leading to cell death. Cycloheximide unselectively suppresses both Cap-dependent and Cap-independent translation, and also inhibits pro-survival autophagy. Therefore, brief pre-treatment with cycloheximide exacerbated NIM811 induced paraptosis.These findings expand our understanding of paraptoticcell death and allow us to identify crucial cellular mechanisms that determine the fate of cells on whether to live or to die. Although in GBM cells, autophagy is not greatly enhanced above normal cells, it remains indispensible for cancer cells to circumvent ER stress associated paraptosis. These studies reiterate the importance of cyclophilins in aiding cancer cell survival under stress conditions, and support their potential as therapeutic targets.


Drosophila embryogenesis. We review the existing body of evidence concerning EGF signaling in Drosophila embryogenesis focusing on current uncertainties in the field and areas for future study. This review provides a foundation for utilizing the Drosophila model system for research into EGF effects on cancer.EGF signaling is a well-known oncogenic pathway in animals. It is also a key developmental pathway regulating terminal and dorsal-ventral patterning along with many other aspects of embryogenesis. In this review, we focus on the diverse roles for the EGF pathway in  Drosophila homolog of the epidermal growth factor receptor abbreviated as EGFR is known by several names—faint little ball and torpedo—with flb referring to the embryonic phenotype and torpedo referring to an oogenesis defect observed in eggs [DER and Ellipse for imaginal disc and adult eye phenotypes [Drosophila embryogenesis [Drosophila EGFR is highly conserved with vertebrate forms of EGFR, sharing 40.5% identity [The  in eggs , DER andenotypes ,3. It isogenesis ,5. The eogenesis . Followiidentity ,7,8.gurken (grk), spitz (spi), vein (vn), and keren (krn) [grk, spi, and krn show the highest homology to TGF-α whereas vn is homologous to neuregulin [vn can be substituted for by spi when expressed in the endogenous vn pattern, showing that in some cases the identity of the ligand is not as critical compared to the ability of the receptor to bind the signal [EGFR receives its signal from a series of different ligands, which can interact with the receptor through different molecular mechanisms . These len (krn) ,11,12. Turegulin . It was uregulin , howeveruregulin . For exae signal .krn is less well-characterized than spi. Emerging literature appears to suggest that, aside from its alternative mechanism of transport and regulation, it is a functional homologue to spi, which functions redundantly with spi in early development [krn in embryogenesis lends itself to a question with no immediately compelling answer: what selective pressure would cause krn to be maintained as a ligand in EGF signaling, and why would it develop an alternative cleavage mechanism to the primary ligand, spi.elopment ,15,16. Hvn also appears to be biochemically redundant with the TGF-α-like ligands. In fact, loss of vn in lateral embryonic stripes had no effect on downstream activation of ERK [vn expression is determined by the gradient of nuclear Dorsal established by the Toll pathway [vn’s unique function may be to activate the EGF signal in tissues where low levels of activation are required due to its weaker activation capacity than spi [spi is able to rescue vn mutants and modulates its activity to roughly match the activation by vn [n of ERK . vn expr pathway , but vn’than spi ,18. spi on by vn . Overallarg) serves as a ligand antagonist by forming a clamp-like structure around spi and thereby inhibiting EGF signal transduction [arg functions in the pathway through a negative feedback loop where it is produced as a result of high levels of EGFR activity [arg is required for normal function of the EGF pathway, and loss of Arg results in excessive EGFR pathway activation and corresponding expansion of ventral cell fates [arg therefore acts to limit long-range effects of the signal by restricting the spread of spi [In addition to these four extracellular ligands, Argos  was central to the function of the receptor prior to the evolutionary events where other ligands were co-opted [Some recent experiments suggest that the EGFR ligands are interchangeable as there is significant redundancy between ligands, similarity in mechanisms of activation, and minimal differences in regulation between the ligands ,16,18,21co-opted .rho-1/star-mediated trafficking, despite its otherwise identical function to SPI in terms of activating the EGF signaling pathway [EGF ligands undergo significant intracellular regulation in the process of their production. SPI is initially produced as a transmembrane pro-protein, which is then cleaved to release the extracellular signaling domain . SPI is  pathway .rho-1 deserves particular attention as a regulatory protein within the EGF signaling pathway. Not only is rho-1 very highly conserved [rho-1 mediated cleavage [rho-1 on EGF extracellular ligands is clear, it is still unclear what, if any, regulatory function RHO-1 has on STAR [star and spi are ubiquitously expressed, whereas rho-1 is precisely spatially and temporally regulated [spi rather than grk. RHO-1 is a key point of cross-pathway regulation in early embryogenesis, which will be discussed below.onserved , but it onserved . All foucleavage . While t on STAR . Both stegulated ,29, whicegulated . This meDrosophila was originally thought to have three homologs of vertebrate ras, a gene at 85D based on chromosome location (ras85D or ras1), the second gene at 64D (ras64D or ras2), and rap1 (ras3) (which was later found to be a separate GTPase and not in fact a ras homolog). We will focus on Ras1 as it is most applicable to embryonic patterning [ras1 has been studied extensively in EGF—mediated signaling in the Drosophila eye and in higher organisms, and we will apply many of the structural findings which have elaborated the molecular mechanisms through which ras1 transduces the EGF signal to the later components of the pathway in embryogenesis.tterning ,31. ras1trans-phosphorylates across the dimer. The protein Downstream of Receptor Kinase  binds through its SH2 domain to the phosphorylated tyrosines on EGFR localizing it to the plasma membrane. In turn, DRK binds Son of Sevenless (SOS) the guanine exchange factor which triggers activation of RAS by promoting GTP binding [The basic outcome of EGF signal transduction is the activation of RAS. The actual mechanism is more complex. First, upon ligand binding, EGFR forms a dimer and  binding ,32.sprouty (sty). sty was once thought to be an extracellular inhibitor of EGF signaling [sty functions as an intracellular inhibitor of the pathway modulating ras activity [sty regulates the pathway is not clear, evidence suggests that STY interacts with DRK and GAP1, at least in eye development [sty functions will lead to a full molecular mechanism of signal transduction from EGF to ras [RAS is therefore positively regulated by the EGF. It is also negatively regulated by ignaling , but recactivity . While telopment . This inF to ras .polehole in Drosophila or just RAF), a highly conserved serine-threonine protein kinase. The fundamental function of activated RAS is to localize RAF to the plasma membrane, where RAF undergoes further activation [Drosophila as downstream of raf-1 (dsor1), and was shown to have significant sequence similarity to murine MEK and Xenopus MAP Kinase activator. MEK was shown to be activated through direct phosphorylation by RAF, causing activation of kinase activity [Drosophila homolog of MAP kinase (MAPK)/extracellular signal regulated kinase (ERK), known as rolled [rolled phosphorylates the transcription factors such as pointed and yan which in turn lead to transcriptional responses [rolled, as expected, produce the same cell-death phenotype as seen in an EGF loss of function [The main function of active RAS is to activate D-RAF , a transcription factor responsible for inhibiting cell differentiation. In essence, the two transcription factors are in opposition with pointed as the activator of EGF pathway target genes and yan as the repressor.In embryogenesis, ntiation ,45. poinentation . A seconyan is destabilized and therefore repressed, and pointed is activated [yan derepression and pointed activation results in activation of EGF target genes such as orthodenticle (otd), arg, and tartan , ventral nervous system defective (vnd), and intermediate neuroblasts defective (ind) [toll, dpp, and EGF signaling, which together are responsible for establishing three separate Hox domains. Specifically, where the msh domain is patterned due to repression of dpp, ind is expressed after being activated both by Toll and EGF signaling, and the vnd domain is established by Toll signaling and maintained by EGF signaling [dpp signaling [dpp signaling and to preserve the existing dorsal-ventral axis established by early Toll signaling [ind repression of vnd), and that EGF over expression causes the expansion of both vnd and ind into the lateral domain  [gcm and repo were reduced in the absence of EGF or spi, concluding that EGF signaling is responsible both for initial formation and maintenance of lateral glia. Likewise, over expressing EGF through over activated spi caused an expansion of the medial and intermediate cell fates into the lateral cell column, causing a repression of Repo [dpp, recent work shows that EGF could also be repressed by dpp. While dpp primarily functions at a short range, bounded by interaction with EGF and other pathways, dpp can also function indirectly at long-range in neuroblast development by repressing EGF signaling, again demonstrating the significant versatility of EGF signaling as a regulatory mechanism for precise spatial and temporal differentiation of the neuroectoderm [Three homeobox genes control the development of the dorsal-ventral axis of the ve (ind) . This suignaling ,47,48. Tignaling , and shoignaling ,51,52. W domain) . Recentl of Repo . This inectoderm .ind through its downstream effector MAP kinase. Phosphorylation of ind results in repression of ind target genes, which allows EGF signaling to modify cell fate even in the presence of high levels of ind protein [Furthermore, we now know that EGF signaling has at least some degree of ability to pattern the neuroectoderm independently of its genetic effect on key neural development homeobox genes. EGF signaling has been shown to mediate the direct phosphorylation of  protein .argos-mediated repression of EGF ligand binding) allows EGF signaling to maintain precise gradients of cell fate determination in the lateral regions of the ventral neuroectoderm [This finding strengthens the conclusion that EGF signaling is necessary for neural development, as an active regulator of key homeobox genes and their protein products. This is further supported by the fact that the loss of EGF signaling results in widespread apoptosis of alternative neuroblast identities . Finallyectoderm . Figure wnt and hedgehog signaling in the patterning of denticles. There are two ways in which WNT and EGF signaling interact [shavenbaby [downstream of raf-1  has been shown to inhibit AXIN-mediated destruction of Armadillo (ARM) in an EGF signaling independent manner [wntD, which limits dorsal (dl) nuclear localization at the poles and along the dorsal-ventral axis of the developing embryo [toll in embryonic patterning (unpublished observations). Overall, recent work on EGF signaling in non-neuroectodermal ventral patterning shows that EGF signaling is capable of playing a wide-ranging and diverse set of roles in its pathway interactions.EGF signaling is also involved in ventral epidermal patterning, where it interacts with interact ,60,61. Favenbaby . WNT sigavenbaby , but in avenbaby ,64. Thisg embryo . Our recvein (vn) and spitz (spi) are involved in the development of muscle precursors and argued that vn is qualitatively different from spi to avoid arg-mediated negative feedback into the EGF signaling process [vn and spi permits regenerative proliferation of multipoint gastric stem cells after damage [vn differs qualitatively from spi in muscle precursor development [Drosophila EGF signaling: that most EGF mediated pathways in Drosophila embryogenesis are involved both in the initial establishment and subsequent maintenance of processes, as shown by the requirement for EGF in both repair and establishment of muscle tissue.Yarnitzky et al. showed in 1999 that  process . Unfortu process ; and (2)r damage . Taken telopment . More imThe multiple roles for EGF signaling necessitate the need for tight regulatory mechanisms ensuring the correct development of a complex organism, and at the same time, allowing flexibility in damage repair, and preventing aberrant growth. As many experiments in a variety of model organisms and in humans have shown, errors in EGF regulation result in highly tumorigenic growth, as the pathway’s excessive activation allows uncontrolled growth of many tissue types as well as metastasis .pointed one of the main transcription factors under the regulation of EGF [A new focus for embryonic research is the effect of EGF signaling in tracheal development, specifically its role in specification of tissue integrity. EGF signaling was found to be necessary for maintaining tissue integrity of epithelial cells during tracheal development through modulation of cell adhesion a function independent of n of EGF . Activatn of EGF , demonstn of EGF . EGF sign of EGF . The netn of EGF ,71. Addin of EGF . Studiesatonal (a transcription factor responsible for nerve cell development) triggers rhomboid expression, causing EGF activation, which initiates recruitment of attachment cells from the ectoderm and induce their specialization [Recent work has provided some additional insight into EGF’s influence on development of Chordotonal organs. Inbal et al. had previously shown that lization . Again, In 2004, Brodu et al.  showed tAntagonism between Dpp and EGF signaling is responsible for dorsal closure and corresponding patterning of cells near the dorsal midline . Dpp bloDrosophila pharynx by affecting planar cell polarity of cells in a square cell grid [EGF signaling is responsible for modulating square cell packing in the presumptive ell grid . This prell grid . This fi(1)EGF often functions in cell growth and proliferation and regulates apoptosis.(2)Drosophila is highly self-regulating, particularly through RHO-1 and STAR mediated processing of EGF ligands and post-transcriptional control of ARGOS expression.EGF signaling in (3)vein vs. spi). The availability of ligands with slightly different functionality allows EGF signaling to be precisely regulated based on ligand availability, rather than attempting to modulate the effects of the pathway after activation by a single ligand.EGF signaling also can differ qualitatively due to expression of different yet functionally similar ligands EGF signaling functions in cell adhesion and polarity. These four characteristics make EGF signaling highly effective at modulating tissue boundaries involving multiple alternative signaling pathways, as EGF signaling can self-regulate expression and interact with many other pathways through both negative and positive regulation.Drosophila embryogenesis has proven to be a valuable tool for understanding the complex mechanisms which give rise to the complexity of the various tissue systems of the Drosophila embryo. Careful study and analysis of EGF signaling mechanisms can reveal broader themes about embryonic control of cell fate, and are highly clinically applicable in understanding how aberrant EGF signaling can contribute to a wide range of human pathologies. This review did not cover other EGF pathways such as PI3K/PTEN, mTOR and Src as this has mainly been studied in larval and adult tissues. For EGF pathway studies, these tissues have provided a variety of useful tumor and drug screening models [Overall, EGF signaling is a crucial factor in promoting cell survival and differentiation. The study of EGF signaling in g models , aging mg models , colon sg models ,81, braig models ,83, and g models .Drosophila) through a CRY2 fusion and blue light, an approach that should work for EGFR [polehole and the faint little ball phenotype, new methods will allow much more precise study of intracellular components of the pathway, and will allow a deeper understanding of the complex interactions between the EGF pathway and more well-studied embryonic signaling pathways, helping to further unravel the complexities of the developing Drosophila embryo. The work we have presented here represents a broad foundation upon which a deeper understanding of the mechanisms through which EGF signaling affects cancer can be built.With the advent of incredibly powerful new gene editing technologies such as the CRISPR/Cas9 system, and new mutants revealed in the genetic screens , there efor EGFR ,86,87,88


After a prolonged warm-up period of basic research, several modalities of cell replacement therapies are under development for diseases with no available cure. Diabetic polyneuropathy (DPN) is one of the most prevalent chronic diabetes complications that causes sensorimotor dysfunction, subsequent high risks for lower limb amputations, and high mortality. Currently, no disease modifying therapy exists for DPN.Several types of well-documented stem/progenitor cells have been utilized for cell transplantation therapies in DPN model rodents: mesenchymal stromal cells (MSCs), endothelial progenitor cells (EPCs), and cells with similar characteristics of MSCs or EPCs derived from embryonic stem cells or induced pluripotent stem cells. Some recent experimental studies reported that these immature cells may have beneficial effects on DPN.Although the role of nerve regeneration in the pathology of DPN has not been sufficiently elucidated, many intervention studies attempting regenerative therapy of DPN have been reported. Further studies are needed to better evaluate the potential of regeneration in reversing the pathology of DPN Today, the field of regenerative medicine is undergoing great development.Traditionally, there have been several options for tissue regeneration, i.e., artificial scaffold, cell transplantation, and replacement of various cytoprotective or growth factors. However, as new cell sources including induced pluripotent stem cells (iPSCs) , 2•• werAn overview outlining proposed steps to develop this new therapeutic option for DPN is presented in Fig. Adult stem cells or tissue stem cells are cells distributed in various tissues and organs throughout our adult lives. It is generally accepted that hematopoietic stem cells, neural stem cells, and intestinal stem cells can be regarded as stem cells –12 becauCommon features of these MSCs are adherence to plastic dishes, and the ability to differentiate into mesodermal cells in, for example, bone, cartilage, and adipose tissues , 25. HowHowever, it is not yet established that MSCs are true stem cells because their proliferation is mostly self-limited and their self-replication ability is scarcely verified , 30, 31.As described above, MSCs are expected to behave as progenitor-like cells, i.e., to promote tissue reconstruction by providing an extracellular matrix, to exert cytoprotective actions through production of various growth factors, and to accelerate cell proliferation. These features imply possible usefulness in the whole body; therefore, clinical trials have been considered for a wide variety of diseases.As of June 2017, on the WHO’s International Clinical Trials Registry Platform, 651 trials were retrieved in response to the search query "mesenchymal stem cell(s)", and 89 trials with "mesenchymal stromal cell(s)".Although the tissue used for the isolation of MSCs has conventionally been bone marrow, most current studies utilize placentae, umbilical cords, or adipose tissues to obtain MSCs. This expansion of resource options has allowed for a rapid increase in clinical trials around the world. At the moment, as briefly mentioned above, MSCs are used in clinical settings against GVHD associated with bone marrow transplantation in Canada, New Zealand, and Japan. On the other hand, no application as regenerative medicine has been achieved except for a phase III trial for Crohn's disease, which is being carried out in the United States of America.Some basic research studies have shown that bone marrow-derived MSCs have improved nerve conduction velocities in streptozotocin-induced diabetic rats , 36. TheThe differentiation potential of MSCs into Schwann cells may be an additional argument for their potential benefit in the treatment of DPNb , 40, in In addition, MSCs also have the ability to differentiate into pericytes of capillaries, which suggests that MSCs might aid angiogenesis in numerous tissues and organs. However, as the potential for differentiation can decrease with aging of donor cells , the expClinicalTrials.gov database (NCT02387749), but although the trial has already concluded, the results have not yet been published.Although cell transplantation therapies are in full bloom in this decade, it should be noted that cytotrophic/growth factors also attracted attention in experimental medicine. Some growth factor replacement therapies in DPN using nerve growth factor  or VEGF Although a bright future is anticipated for the use of MSCs, some problems need to be addressed. First, the existence of MSCs is still questionable. In 2008, Crisan et al. proved that non-cultured human pericytes expressed MSC markers and exhibited the potentials of osteogenic, chondrogenic and adipogenic differentiation . On the Like MSCs, vascular endothelial progenitor cells (EPCs) have a wide target range of diseases in the field of regenerative medicine. EPCs were first reported as CD34 positive cells in the mononuclear cells derived from the bone marrow by Asahara and Murohara et al., in 1997 . Since tRegardless of the lack of validated EPCs, many basic studies utilizing EPCs have been conducted. Naruse et al., reported that the transplantation of umbilical cord blood-derived EPCs improved the capillary-muscle fiber ratio in soleus muscle and nerve conduction velocities in a rat DPN model . AnotherOwing to the engineering breakthrough of pluripotent cells including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), several types of immature cells that can differentiate into mature cells comprising the PNS were derived from pluripotent cells, and employed for DPN treatment in experimental studies. We previously reported that the transplantation of angioblast-like cells derived from murine ESCs increased capillary density in the soleus muscle and improved motor and sensory nerve conduction velocities (NCVs) in diabetic mice . As an aMany tissue engineering materials have been proposed for wound healing in diabetic animals, particularly on diabetic foot ulcers . FurtherThe potential roles of Schwann cells and vascular cells in the pathophysiology of DPN have not been taken sufficiently into consideration. It is assumed that these cells would promote regeneration of axons or blood vessels in the PNS.Therefore, although most glial or vascular progenitor/stem cells have been injected in intravenous or intramuscular fashion , the valAlthough regenerative therapies of DPN are hampered by an insufficient insight into the pathology of DPN, numerous trials that might be slightly speed-before-quality have been presented. After revealing the regeneration mechanism of the PNS in DPN, several types of cells including vascular, nervous, and/or mesenchymal cells could be considered for an application in regenerative medicine.


The influence of music on the human brain has been recently investigated innumerous studies. Several investigations have shown that structural andfunctional cerebral neuroplastic processes emerge as a result of long-termmusical training, which in turn may produce cognitive differences betweenmusicians and non-musicians. Musicians can be considered ideal cases for studieson brain adaptation, due to their unique and intensive training experiences.This article presents a review of recent findings showing positive effects ofmusical training on non-musical cognitive abilities, which probably reflectplastic changes in brains of musicians. Musicians may serve as a unique model for studying plastic changes in thehuman brain, due to the complexity of this singular stimulus, normally related to veryhigh levels of exposure during musical practice.15The interest in studying the effects of musical training on cerebral function has grownconsiderably in the last decade. A body of evidence indicates that musicians possessstructural and functional cerebral characteristics absent in non-musicians, whichgenerally correlate to the age of commencement of musical training.28 have shown positive associations between formal musical training inchildren and non-musical cognitive abilities. Moreover, other studies41 have demonstrated, directly or indirectly, enhanced cognitiveabilities in adult musicians compared to non-musicians.The structural and functional reorganization observed in musicians’ brains may lead tocognitive differences in comparison to non-musicians. Some studiesOur review initially presents evidence of neurobiological changes produced by long-termmusical training and then focuses on the cognitive differences between musicians andnon-musicians, considering visual and verbal abilities in more detail. The aim of thisstudy was to review some interesting and relevant studies which have provided a growingbody of evidence that intensive musical training may trigger neuroplastic processes, andconsequently, modifications in cognitive processing. The influence of musical trainingon the human brain, specifically on cognition, is a recent and intriguing field ofresearch.4 found that its anterior portion was significantlylarger in musicians compared to non-musicians. Moreover, a comparison betweensubgroups revealed that this region was larger in those who began musical trainingearlier. The authors pointed out that the observed anatomical difference has to betaken in the context of a need for increased interhemispheric communicationunderlying complex bimanual motor sequences in musicians.Several investigations have demonstrated structural and functional changes, by usingmagnetic resonance imaging, eletroencephalography or magnetoencephalography, inbrains of musicians as a result of many years of musical practice. In examining themorphometry of the corpus callosum, Schlaug et al.5 evaluated the size ofright and left motor cortices in right-handed musicians and non-musicians. Asexpected, a leftward asymmetry was observed in both groups, but musicians showed asmaller degree of asymmetry, since they presented a larger right motor cortex. Theinvestigators also found a negative correlation between the size of motor cortex ofboth hemispheres and the age of commencement of musical practice. These authorsinterpreted the structural differences in motor cortex as structural compliance inresponse to intense and early hand skill training.Amunts et al.7 measured the volume of cerebellum in musicians andnon-musicians, showing higher average relative cerebellar volume in male musicianswhen compared to male non-musicians, a finding interpreted as evidence formicrostructural adaptations in the cerebellum in response to early commencement andcontinual practice of complicated bimanual finger sequences.In a retrospective study, Schlaug et al.10 foundincreased gray matter volume in motor, auditory and visuospatial cerebral areas inmusicians, using voxel-based morphometry analysis, a method enabling structuraldifferences to be identified across the whole brain space. According to the authors,the results can be viewed as evidence of structural adaptations in response tolong-term skill acquisition and the repetitive rehearsal of these skills, a notionsupported by the strong association found by the authors between structuraldifferences and practice intensity.Gaser & Schlaug11 using diffusiontensor imaging, investigated effects of extensive piano practice on white matter inchildhood, adolescence and adulthood. They found more structured right posteriorinternal capsule, which carries corticospinal tracts, in pianists compared tonon-musicians, and also found positive correlations between practicing and fibertract organization in different regions for each age period. The authors noted thattraining can induce white matter plasticity if this occurs during periods when fibertracts involved are still undergoing maturation. Han et al.13 examined both gray matter andwhite matter in pianists and non-musicians and demonstrated, in the first group,higher gray matter density in the left primary sensory-motor cortex and rightcerebellum as well as higher white matter integrity in the right posterior internalcapsule. According to the researchers, these results indicate that long-term pianopracticing may lead to gray matter and white matter adaptation in movement-relatedregions, which may impact number of synapses, volume of glia or increasedmyelination and diameter of axons.Bengtsson et al.,3 investigated the somatosensory cortical representation ofthe fingers D1 (thumb) and D5 (little finger) of string players – violinists,cellists, guitarists – and non-musicians. After excitation of left-hand fingers D1and D5, the strength of cortical activation was higher in musicians than in controlsubjects. This effect was particularly pronounced for finger D5, while there was nodifference in cortical representations following right-hand stimulation. The authorsstated that cerebral representation is enhanced to the fingers of the hand that ismost intensively used in string players, and that the more a given finger isstimulated, the larger the increase in cortical response.Other studies have shown differences in functional cerebral characteristics betweenmusicians and non-musicians. Elbert et al.6 investigated auditorycortical representation in musicians and non-musicians. After acoustic stimulation,consisting of a pseudorandom sequence of four piano tones and four pure tonesmatched in frequency and loudness, the strength of cortical activation for pianotones was 25% greater in musicians. There was no difference between the groups inrelation to cortical representation of pure tones. The authors stressed that puretones, unlike piano tones, are not part of our natural acoustic environment and alsoare not commonly encountered in musical training and practice, which may account forthe observed results. Both Elbert et al.3 and Pantev et al.6 revealed a greater degree of cortical representation inmusicians who began musical training early.Pantev et al.8 and Rüsseleret al.9 compared musicians andnon-musicians by using mismatch negativity (MMN), a component of the evokedpotentials in auditory cortex, obtained by submitting subjects to hundreds ofidentical stimuli that are randomly replaced by distinct ones, in the absence ofattention to stimulus. Koelsch et al.8 observed MMN for slightly impure chords presented among perfectmajor chords, only in musicians, suggesting that they are superior inpre-attentively extracting more information out of musically relevant stimuli.Rüsseler et al.9demonstrated that while musicians showed MMN for regularly spaced tones mistimed by20 milliseconds, presence of MMN in non-musicians needed mistiming of longer than 50milliseconds, indicating that temporal integration may be more precise inmusicians.Koelsch et al.14 investigatedplastic capabilities of the hippocampus by evaluating brain responses induced bytemporal novelty in musicians and non-musicians, since the hippocampus structure, inaddition to its key role for memory and spatial navigation, has been suggested to becrucially involved in various forms of novelty detection. They observed enhancedneural responses to temporal novelty in the anterior left hippocampus inprofessional musicians  and in music students after one yearof intensive aural skills training . The authors also found acorrelation between hippocampal sensitivity to temporal novelty and musicalabilities. They assumed that the observed changes in hippocampal activity inmusicians represented a functional correlate of a tuning of aural skills, related totime interval perception, during the course of their studies.Herdener et al.12 showed thatchanges in functional organization also extend to subcortical sensory structures,demonstrating that musicians, compared to controls, had earlier and larger auditoryand audiovisual brainstem responses to speech and music stimuli. The data alsoshowed a positive correlation between years of musical practice and strength ofbrainstem response to speech stimulus, suggesting that musicians acquire an enhancedrepresentation of pitch through musical training. Musacchia et al.42demonstrated that cortical activation during music processing reflects the auditory“learning biography”, the personal experiences accumulated over time. The authorproposed a model to represent the relationship between auditory information andneural networks involved in music processing. According to his model, complexity ofneural networks enhances with the complexity of auditory information. Moreinterestingly, musical training can add mental representations of music, which mayinvolve different cerebral substrates. These representations can be auditory,sensory-motor, symbolic, visual, among others. Therefore, according to the author,for the same level of auditory information complexity, professional musicianspresumably use larger and more complex neural networks compared tonon-musicians.All of the previously addressed studies suggested the existence of neuroplasticprocesses as an effect of musical training. In a longitudinal study,Altenmüller28 andadults.41 The present study focuses on two cognitive domains– visual and verbal abilities – since studies comparing cognition in musicians andnon-musicians have demonstrated benefits of musical training mainly in thesedomains. These previous studies are summarized in The structural and functional neuroplastic processes verified in brains of musiciansmay influence their cognitive functioning, and yield differences between musiciansand non-musicians. Many investigators have studied the effects of extensive musicaltraining on non-musical cognitive abilities in children31investigated visuospatial abilities in musicians and non-musicians by using aneuropsychological task in which subjects had to detect the position of a targetdot relative to vertical or horizontal reference lines flashed on a screen. Inone condition (perception condition), the reference line remained on the screenuntil the dot was displayed, while in a second condition (imagery condition),the line disappeared before the target dot was presented, requiring subjects tokeep a mental image of the reference line. In both conditions musicianspresented shorter reaction times compared to controls, suggesting enhancedvisuospatial abilities in the former group. Another experiment in the same studycompared sensory-motor abilities between the two groups, showing that visualabilities of musicians could only be partially explained by better sensory-motorintegration. The investigators attributed their results to long-term practice ofmusical reading, which involves fine recognition of relative vertical andhorizontal positions of musical notes on the score. They also ascribed theobserved differences between the two groups to more efficient attentionalprocesses in musicians.Several studies have described positive effects of musical training on visualcognition. Brochard et al.29 the patternof saccadic eye movements can be used as an indicator of mental disabilities, aswell as a measure of mental processing speed. The same authors stated that dueto specific demands of music reading, it is reasonable to presume that an earlystart of musical practice may modify the way visual information is processed bythe nervous system of adult musicians. Kopiez & Galley29 and Gruhn et al.32 compared saccadic eyemovements during oculo-motor tasks in adult musicians and non-musicians andreported more efficient oculo-motor strategies in the former, which, accordingto the authors, could be associated with complex visual processes involved inlong-term practice of music reading. They highlighted that the differenceobserved between the two groups could also be related to more efficientattentional processes in musicians, although this aspect was not directlyinvestigated by the researchers. According to Gruhn et al.,32 some studies46 indicate the existence of a strong association betweenattention and saccadic eye movements. The voluntary control of eye movementsrequires highly complex mental processes, involving many cerebralareas.47 According toKimmig,48 allmodalities of attention have an impact on the oculo-motor system. Fixation,characterized by a voluntary suppression of saccadic movements, as well as therate of express saccadic movements, involve dorsolateral frontal lobe processes,which also participate in attentional mechanisms.32Important aspects of music reading practice are addressed by comparing saccadiceye movements in musicians and non-musicians. According to Kopiez &Galley,33 compared theperformance of musicians, members of a symphony orchestra and a symphony band,and non-musicians, in tasks involving visual attention ability. The mainneuropsychological test used in the study was the Multiple Choice Reaction Time(MCRT) test,49 which consistedof specific motor responses to various luminous stimuli. In order to evaluatedivided visual attention ability, the MCRT test was applied twice, the secondtime concomitantly with other continuously and randomly changing visual stimulipresented in video form. Subjects were asked to respond verbally to each change.Musicians showed a higher percentage of correct responses to MCRT when the testwas applied alone. Although no significant difference of accuracy was observedbetween the groups when MCRT was applied together with changing visual stimuli,musicians showed shorter reaction times for verbal responses to stimuli changes.The authors pointed out that this result may suggest augmented divided visualattention ability in musicians compared to non-musicians, which they ascribed toensemble musical practice. The professional routine of musicians ischaracterized by constant demands of divided visual attention, since it requiresdealing with several kinds of visual stimuli simultaneously such as the musicscore, conductor’s gestural instructions and body movements of other musicians,while playing the instrument.Rodrigues et al.34 comparedright-handed musicians and non-musicians in a line-bisection task,50 which entailed marking thecenter of 17 horizontal lines, varying in length from 10 to 26 cm, displayedrandomly on a page. As pointed by Hausmann et al.,50 on this task, neurologically intactright-handers show a slight yet reliable tendency to bisect about 2% to the leftof the true center, a phenomenon attributed to dominance of the right hemispherefor visuospatial attention51.Patston et al.34 demonstratedthat musicians showed a slight rightward bias, while non-musicians showedgreater deviation to the left, and that musicians bisected the lines moreaccurately and with smaller intermanual difference than the control group. Theresearchers suggested that musicians may develop an increased ability for theleft hemisphere to perform cognitive functions that are usually right-hemispheredominant, resulting in a more balanced spatial attention.Patston et al.35 investigated lateralization of visuospatial attention inmusicians and non-musicians by comparing reaction times and accuracy to stimulipresented to the left and right of a vertical line, a similar task to that usedby Brochard et al.31 in the“imagery condition”. While both groups performed more accurately with the leftsided-stimuli, musicians were significantly more accurate than controls for theright-sided stimuli and also had faster reaction times overall. According to theauthors, consistent with previous research,34 the resultsindicated a more balanced attentional capacity in musicians, as well as enhancedvisuomotor ability. It is possible that musicians have an advantage online-and-dot tasks, of the type used both in this study and that by Brochard etal.,31 due to theirfamiliarity with these components from music reading. In the task used byPatston et al.,35 however, theline was vertical, with dots occurring to either side, removing the directcomparability of the stimuli to musical notation.In another study, Patston et al.36 also studiedthe lateralization of visuospatial attention electrophysiologically using ameasure of callosal function – interhemispheric transfer time (IHTT) –calculated by comparing the latencies of occipital N1 components of evokedpotentials between hemispheres. Musicians and non-musicians responded to stimulipresented to the left and right visual fields while submitted toelectroencephalography. Non-musicians showed significantly faster IHTT in theright-to-left direction than in the opposite direction and a shorter N1 latencyin the left than in the right hemisphere. In contrast, musicians exhibited nodirectional difference between hemispheres in IHTT and no hemispheric differencein latency, indicating a more bilateral neural connectivity. The researchersproposed that bimanual training facilitates an unusual process of extramyelination that results in more balanced connections between hemispheres thannormally found in those without musical training.Patston et al.37 investigatedvisual processing of local details in musicians and non-musicians, by utilizingdisembedding and constructional tasks. In experiment 1, they used the GroupEmbedded Figures Test (GEFT),52 which consists of presenting a series of 25 complexfigures, each containing one of nine targets hidden in the design. The subjectis asked to examine each test figure and to outline the hidden target as soon asthey identify it. In experiment 2, the authors used two tests: the Block Designsubtest from the Wechsler Adult Intelligence Scale-Third Edition(WAIS-III),53 thatrequires the subject to replicate a geometric pattern presented on a card usingthe top surfaces of several colored blocks, and a task involving copyingpossible and impossible drawings of objects. Musicians outperformednon-musicians on the GEFT, on the Block Design subtest and also on the task ofcopying drawings of physically impossible objects, suggesting enhanced visualprocessing of local details. A correlation was found between the Block Designscores and the accuracy scores for the impossible figures, but not for thepossible figures, indicating that local processing ability did not correlatewith drawing ability per se. The authors concluded that arelative strength in local processing contributed to superior performance ofmusicians on the drawing task. They hypothesized that the increased visualprocessing of local details seen in musicians may reflect training-inducedchanges in a fronto-parietal system involved in controlling exploratory eyemovements and shifts in visual attention, skills that are important for musicscore reading, which also requires analysis of visual details.Stoesz et al.39 in a part oftheir work, studied visual memory in pianists and non-musicians by using the ReyVisual Design Learning Test (RVDLT).54 In this test, participants try to learn and remember asequentially presented set of 15 line drawings of simple geometric figures, eachcontaining two elements . Recall, registered byasking subjects to draw all figures they can remember, is tested after each offive learning trials and following a delay period. A test of delayed recognitionis also administered. The results suggested superior visual memory in musicians,since they outperformed non-musicians on the fourth and fifth learning trialsand on the delayed recall and delayed recognition tasks. After controllingstatistically for general intelligence, the group difference on the delayedrecall tasks persisted. The authors emphasized that drawing ability was not aconfounder in this investigation, given that all figures in the RVDLT areextremely simple. Stoesz et al.,37 cited above, showed that musicians and non-musicians wereequally accurate at drawing possible objects. According to the researchers, theobserved relationship between visual memory and musical training may be relatedto improvement in processes supporting attention to visual details, to increasedskill of musicians at holding and manipulating visual images in working memory,which confers an advantage during the encoding proces, or may reflect superioruse of high level, strategic memory processes.Jakobson et al.,40 have found no group differences. Brandler &Rammsayer30 used atask in which participants were asked to indicate, from memory, the locations ofbuildings on a city map they had previously studied. Chan et al.40 and Ho et al.24 used tests to assess memoryfor visual designs. Jakobson et al.39 identified some aspects in these studies that may havecontributed to an apparent discrepancy. Chan et al.40 and Ho et al.24 worked with Asian samples, that would havebeen trained in the use of an ideographic writing system. Some previousstudies56 showed that such training isassociated with better memory for abstracts designs. Brandler &Rammsayer,30 on theother hand, did not investigate an Asian sample, but the nature of the visualmemory task used by these researchers, requiring topographical skills, differedfrom that of Jakobson et al.39Since some studies58 have suggested that neuralsubstrates of topographical memory are separate from those supporting visualmemory for faces and designs, the authors hypothesized that piano training mayhave a greater effect on systems involved in storing simple design information,than on those involved in storing topographical information, since musicnotation shares many common features with simple designs.Curiously, other studies examining visual memory in musicians andnon-musicians59showed activation of an associative visual area (Brodmann area 19) in musiciansduring a pitch discrimination task. The authors suggested that musicians wouldbe able to imagine melodic lines in some kind of visual axis in order to detectpitch changes. Schmithorst & Holland60 investigated the relationship between musical practiceand specific cerebral processing of two musical elements: melody and harmony.The results demonstrated that musicians and non-musicians recruit differentneural networks to process these elements. Inferior parietal areas wereactivated only in musicians during melody perception  and harmony perception , which have beenidentified as involved in general visuospatial processing. Sluming etal.61 compared malemusicians, members of a symphony orchestra, with non-musicians and revealed anincreased density of gray matter in Broca’s area in the brains of the musicians.This area is an important neuroanatomical substrate for spoken language and,according to the authors, for several musically relevant abilities, includingvisuospatial localization. More recently, Sluming et al.62 showed enhanced performance ona visuospatial task by orchestral musicians, compared to non-musicians,associated with increased activation in Broca’s area.Some brain imaging studies have also suggested more efficient visual processes inmusicians. Platel et al.39. In fact, there is much evidence pointing to alink between musical practice and verbal memory ability. Most of the groups ofmusicians involved in such studies consisted of instrument players. Chan etal.40, by using alist-learning task, showed that adults who received musical training before theage of 12 for at least six years had a better memory for spoken words than thosewho did not. Kilgour et al.41showed that young adults with musical training, when compared to controls,performed better in recalling verbal materials under all conditions, includingimmediate and delayed recall trials. Brandler & Rammsayer30 compared some mental abilitiesin musicians and non-musicians by utilizing a comprehensive intelligence testbattery, and verified superior performance of musicians only on the verbalmemory test.The skills underlying the ability to learn a piece of music resemble thoseinvolved in memorizing a poem or a piece of prose24 used a hybridcross-sectional and longitudinal design and assessed verbal memory ability inchildren with and without musical training using the Hong Kong List LearningTest (HKLLT).63 In thecross-sectional part of the study, the results showed that children with musicaltraining demonstrated better verbal memory than those without such training. Thelongitudinal part of the study consisted of a one-year follow-up of thechildren, forming three groups: those with experience who continued lessons,those who discontinued their lessons, and those who were new to musicaltraining. The researchers verified that children who had begun or continuedmusical training demonstrated significant verbal memory improvement and thosewho had discontinued the training showed no improvement at all. They associatedtheir results with the neuroimaging findings of Schlaug et al.64 that revealed an enlargementin the left planum temporale in musicians. Ho et al.24 suggested that musical training duringchildhood may serve as a kind of sensory stimulation that contributes to thereorganization of the left temporal lobe in musicians, which in turn facilitatescognitive processing mediated by this specific brain area, as is the case ofverbal memory.Ho et al.39 in anotherpart of their study mentioned earlier, investigated verbal memory in pianistsand non-musicians using the California Verbal Learning Test-Second Edition(CVLT-II).65 This testincludes two aurally presented word lists, each consisting of 16 words from fourdifferent semantic categories. Recall is tested after each of five learningtrials, and free and cued recalls are assessed following both short and longdelay periods. The results suggested superior verbal memory in musicians, sincethey outperformed non-musicians on the short delay cued recall and on the longdelay with both free and cued recalls. After controlling statistically forgeneral intelligence, group difference persisted in long delay free recalltrials. In line with the part of this study investigating visual memory, theseresults were consistent with the idea that formal music training is specificallyassociated with superior memory functioning, beyond any effects it may have ongeneral intelligence. Moreover, musicians demonstrated greater use of a semanticclustering strategy during the learning phase, suggesting, according to theauthors, that the verbal memory advantage does not simply reflect bettermemorization skills. Rather, it is associated with a relative strength in theextraction of higher-order, semantic information during encoding. Theresearchers highlighted that the encoding and storage of information abouthierarchical relationships between words, as in the case of structuralrepresentations of music, reflects the use of strategic memory processes. Asargued by Patel,66 even ifthese two types of representations are stored separately, the processes involvedin activating them may depend on a common neural substrate. According toJakobson et al.,39 it seemslikely that changes in this neural circuit, induced by training, underlie theverbal memory advantage displayed by musicians in their study.Jakobson et al.,38 alsoinvestigated verbal memory in musicians and non-musicians. The study wasconducted in two phases: in phase 1, they tested long-term verbal memory usingthe Rey Auditory Verbal Learning Test (RAVLT),67 which uses a list-learning paradigm, and inphase 2, they introduced articulatory suppression within the same test in orderto interfere with verbal rehearsal processes. As described by the researchers,articulatory suppression, achieved by giving subjects the additional instructionof saying the word “the” between each word read from the list, is a techniquethat prevents participants from rehearsing verbal material to aid its storage inworking memory and after in long-term memory. They also tested verbal workingmemory using reading-span and operation-span tasks. The results showed thatmusicians outperformed non-musicians in phase 1, but not in phase 2. Accordingto the authors, the fact that the superior performance of musicians is lost withthe introduction of articulatory suppression may suggest that an overt rehearsalstrategy is being used by musicians to boost performance. Moreover, a greaterverbal working memory capacity for musicians was also found. Therefore, theresearchers suggested that musical training affects verbal long-term and workingmemory and that enhanced rehearsal mechanisms are likely responsible forsuperior performance of musicians in verbal memory tasks.Franklin et al.64 cited earlier, investigated the morphometry ofthe planum temporale, a brain area containing auditory association cortex andpreviously shown to be a marker of structural and functional asymmetry, andfound stronger leftward asymmetry in musicians compared to non-musicians. Theresearchers considered this asymmetry as potentially related to language andpitch processing skills. Ohnishi et al.68 assessed cortical activation during a passive listeningtask and found greater activation of the planum temporale and the leftdorso-lateral prefrontal cortex in musicians than in controls. The authors alsofound a negative correlation between the degree of activation in the left planumtemporale and the age at commencement of musical training. Somestudies71 have shown that musiciansoften demonstrate greater left-hemisphere lateralization than non-musicians whenpresented with musical stimuli. More recently, Fujioka et al.72 investigated brain activationin children while listening to violin tones and also observed a greaterleft-hemisphere lateralization in those children who received music lessonsthroughout a year.Results of some brain imaging studies have been consistent with more efficientverbal processes in musicians. Schlaug et al.,73 is the question of causation. It is not yet known whetherenhanced cognitive abilities verified in musicians are in fact a consequence oflong-term training or are innate. Most of the studies suggesting benefits of musicalpractice on cognition are of a correlational or quasiexperimental nature, hamperingthe determination of a clear causal link.An important issue related to music and cognition that remains unresolved, asemphasized by Schellenberg & Peretz,22 using a pre-testand post-test design, have demonstrated augmented spatial abilities in childrenafter periods of music lessons. It has also been reported, as mentioned previously,that the degree of structural and functional enhancement in specific brain areascorrelates with the age of onset of musical training,68 the number of years of musicalpractice61 and its intensity.11 Nortonet al.,74 in a study involvingchildren aged five to seven years, compared those who were about to begin musicstudies with those who were not, and found no pre-existing neural, cognitive, motoror musical differences between the groups. Part of an ongoing longitudinalinvestigation into the effects of musical practice on brain and cognitivedevelopment, initiated by Norton et al.,74 Hyde et al.75 demonstrated regional structural brain plasticity that occurredafter only 15 months of musical instrument training in early childhood. Structuralbrain changes in motor and auditory areas were correlated with behavioralimprovements on motor and auditory-musical tests. According to the researchers, thelack of brain and behavioral differences between music trainees and control childrenat baseline supported the view that differential brain development is induced byintensive musical training rather than by preexisting biological predictors ofmusicality.However, some studies have provided evidence supporting the idea of training-relatedeffects of music on brain and cognition. For instance, training studies,76 reported that taxidrivers have larger posterior hippocampal volume in comparison to the generalhealthy population, argued to be due to the intense visuospatial demands of theirjob. Draganski et al.77 observedthat adults trained for three months in juggling showed an increase of gray matterin particular brain areas, followed by a decrease after a three-month cessation ofjuggling. Pascual-Leone et al.78showed that the motor cortical output map to the reading hand in Braille readers waseven modifiable after only a few hours of training. Similarly, Karni etal.80 have shown enhanced activation of the primarymotor cortex following daily practice of a finger opposition task. Sims &Mayer81 demonstratedenhanced mental rotation skills in video game players. It has been shown thatchildren (Carlson & Meltzoff82), adults  and older adults  who are lifelong bilinguals anduse both languages on a regular basis exhibit higher levels of executive controlthan comparable monolinguals.It is also noteworthy that musical practice is just one sort of training that canlead to neural and cognitive changes on human brain. Other studies have demonstratedsuch changes after different forms of training. For example, Maguire etal.74 cognitive enhancement effects of musical training, theresult of neuroplastic processes, might be due to a combination of skills requiredby music study, such as decoding visual information into motor activity, memorizingextended passages of music, learning music structures and rules, learning to makefine auditory spectral and temporal discriminations and learning to perform skilledbimanual finger movements.Hence, several forms of intensive training have an impact on brain and cognition, butit is possible that musical training has specific effects that other forms oftraining do not have, or even produces a range of different effects, considering itsmultifaceted nature. According to Norton et al.,37studies demonstrating enhanced cognitive abilities in musicians can be of intereston both theoretical and applied grounds. At a theoretical level, such studies maylend a better understanding of the effects of musical practice on the developmentand functioning of multiple neural systems, which support a range of cognitiveabilities, whereas at an applied level, they may contribute to advances in thefields of education and cognitive rehabilitation.As emphasized by Stoesz et al.,