Of the estimated 10–11 year life expectancy gap between Indigenous  and non-Indigenous Australians, approximately one quarter is attributable to cardiovascular disease (CVD). Risk prediction of CVD is imperfect, but particularly limited for Indigenous Australians. The BIRCH  project aims to identify and assess existing and novel markers of early disease and risk in Indigenous Australians to optimise health outcomes in this disadvantaged population. It further aims to determine whether these markers are relevant in non-Indigenous Australians.BIRCH is a cross-sectional and prospective cohort study of Indigenous and non-Indigenous Australian adults (≥ 18 years) living in remote, regional and urban locations. Participants will be assessed for CVD risk factors, left ventricular mass and strain via echocardiography, sleep disordered breathing and quality via home-based polysomnography or actigraphy respectively, and plasma lipidomic profiles via mass spectrometry. Outcome data will comprise CVD events and death over a period of five years.Results of BIRCH may increase understanding regarding the factors underlying the increased burden of CVD in Indigenous Australians in this setting. Further, it may identify novel markers of early disease and risk to inform the development of more accurate prediction equations. Better identification of at-risk individuals will promote more effective primary and secondary preventive initiatives to reduce Indigenous Australian health disadvantage. Aboriginal Australian and/or Torres Strait Islander peoples  have well-documented health disadvantage in comparison with non-Indigenous Australians . AddressThere is an estimated 10–11 year gap between Indigenous and non-Indigenous Australian life expectancy at birth and, of this, one quarter is attributable to cardiovascular disease (CVD) . ImportaThere is, therefore, a strong rationale to develop more culturally and contextually appropriate strategies to address the disparity in life expectancy between Indigenous and non-Indigenous Australians resulting from CVD. In particular, improvements in the primary and secondary prevention of CVDs in Indigenous Australians has the potential to significantly reduce this gap. This paper describes the BIRCH  project that aims to assess cardiovascular risk and explore the potential value of novel cardiovascular risk factors and early disease markers in Indigenous Australians. The objective is to more accurately identify Indigenous Australians at risk of overt CVD so that preventive treatments can be commenced sooner in those whom will receive the greatest benefit. More specifically, BIRCH will determine whether measures of left ventricular dysfunction, sleep quality and sleep disturbances, and detailed lipid profiles are useful in determining the risk of development of overt CVD in Indigenous Australians such that interventions can be commenced to prevent or retard its development. In addition, it will seek to determine whether there is any difference in the utility of such measures between rural and urban Indigenous populations and whether such measures may have relevance in non-Indigenous Australians.A number of CVD risk stratification scores have been developed to identify individuals at risk of CVD who may benefit from the early introduction of preventive therapies , 8. One Such findings suggest that additional factors not included in the Framingham functions may be important in adding to, or interacting with, traditional CVD risk factors to result in the increased and currently ‘unmeasured’ risk of CVD in Indigenous Australians. For example, Wang and colleagues demonstrated that waist circumference, body mass index (BMI), hip circumference, and high sensitivity C-reactive protein (hs-CRP) were significantly associated with cardiovascular risk independent of other cardiovascular risk factors , 15. SimThese earlier studies predominantly or exclusively involved Indigenous Australians living in remote or outer regional Australian centres. Whether their findings are generalizable to Indigenous Australians residing in major cities and inner regional centres, where 57% of Indigenous Australians live , remainsWhilst many existing and newer markers of early CVD disease or risk may help identify Indigenous Australians at increased risk of development of overt CVD, they often do not independently provide new or additional targets for primary or secondary prevention. Furthermore, such markers continue to focus predominantly on atherosclerosis-mediated CVD despite the increasing recognition of non-atherosclerotic causes of conditions such as heart failure , 23. We Fatigue and poor sleep quality have been found to be related to CVD risk factors including hypertension and CHD, attributable to increased sympathetic activity . Poor slSpecific sleep-related disorders such as obstructive sleep apnoea (OSA) are associated with an increased risk in the development and progression of CVD , 28. OneDespite the evidence of an association between sleep disorders/quality and CVD, little has been achieved in the management of sleep disruptions for Indigenous Australians. Indeed, one recent study demonstrated that regional and remote Australians residents accessed diagnostic sleep studies at approximately 1/20th of the overall Australian rate . DespiteMarkers of left ventricular function and morphology (including size and mass) may represent additional markers of either CVD risk or the earliest signs of pre-clinical CVD. Left ventricular ejection fraction (LVEF) is one of the most commonly used measures of left ventricular systolic function. Whilst it has been shown to be a strong predictor of mortality in patients with heart failure, deteriorating LVEF is often a late marker of disease. On the contrary, patients with signs and symptoms of heart failure or who have subsequent CVD events may still exhibit preserved left ventricular systolic function , 35. WhiGLS is a novel index of myocardial deformation that can be quantified via automated speckle tracking echocardiography (STE). GLS has been validated against tagged magnetic resonance imaging and is more accurate than standard echocardiographic measurements of LVEF , 38. GLSOther measures of left ventricular morphology, and particularly left ventricular mass and volume, may also represent a common endpoint for other cardiovascular risk factors including systolic blood pressure, body mass index, smoking and diabetes . In addiLipidomic analysis permits the simultaneous quantification of several hundred lipid species within plasma, thereby providing a global assessment of lipid metabolism –48. WhilCompared to conventional clinical assessments, lipidomic approaches not only enable the quantification of a far greater number of lipid classes and individual species within plasma (up to approximately 600 lipid species), but can also be applied to lipoprotein sub-fractions including LDL, HDL and very low-density lipoprotein (VLDL) giving greater mechanistic context. Given the pivotal role that lipids play in the pathophysiology of atherosclerotic CVD, new lipidomic technologies provide a way of identifying novel lipid species involved in the pathogenesis of cardiovascular disease and which have potential as biomarkers of future disease risk .Using lipidomic profiling Meikle and colleagues undertook a targeted lipidomic analysis of 305 lipid species in plasma from 220 individuals who were either healthy, had stable CHD or unstable CHD . ResultsGiven previous findings that traditional risk factors for CVD derived from non-Indigenous Australian population cohorts may only provide limited predictive power in Indigenous Australian and other similarly disadvantaged , 55 popuexplain more of the actual risk of the subsequent development of overt CVD in Indigenous Australian adults residing in remote Australia;have equivalent additional explanatory power in Indigenous Australians residing in urban locations; andcapture some, but less, of the currently unexplained CVD risk in non-Indigenous Australians. see Fig. Accordingly, in undertaking the BIRCH study, we hypothesise that markers of sleep quality/disorders, left ventricular function/morphology and select lipidomic species will:It is anticipated the findings of this study and inclusion of these novel risk factors will provide evidence to inform the ongoing development of CVD risk detection, particularly in Indigenous and other vulnerable populations, and provide potential targets for the evaluation of interventions to enhance primary and secondary prevention of CVD.BIRCH is a cross-sectional and prospective observational cohort study of Aboriginal and Torres Strait Islander (Indigenous) and non-Indigenous Australian adults ≥ 18 years) living in remote and urban Australia. It will adhere to STROBE guidelines . BIRCH w8 years lSample size estimations will reflect the three-phase recruitment outlined above and in Fig. The second phase will investigate whether the additional predictive benefit of these novel markers is equivalent in remote and urban residing Indigenous Australians. It will hypothesise the anticipated Framingham-based five-year absolute risk of CVD of 2.6% is comparable in remote and city-based Indigenous participants. It will assume any difference in study-determined absolute risk for development of overt CVD (utilising novel measures) in the remote as compared with urban Indigenous cohorts will be less than 3%. Based on equivalence testing this would then require 738 Indigenous Australian participants, 369 remote and urban. Taking account of loss to follow-up and exclusion of those with pre-existing CVD this will be increased to 1000 in total with 500 remote and 500 urban Indigenous Australian participants.The third and final phase of BIRCH will assume the additional predictive power of novel risk and disease markers will be less for non-Indigenous as compared with Indigenous Australian participants. It will assume the greater absolute risk of CVD explained by the addition of novel markers will be 2% in non-Indigenous Australian participants and, as for phase one, 5% for Indigenous Australian participants. To detect this difference will require 1176 participants, 588 Indigenous and 588 non-Indigenous participants. Given 1000 Indigenous Australian participants will be required for phase two of the study it is proposed a similar number of non-Indigenous Australian participants is recruited to address the highlighted issues above.In summary the BIRCH study will anticipate enrolling a total of 2000 participants across three study phases including 500 Indigenous Australians residing in remote Australia, 500 Indigenous Australians residing in urban Australia and 1000 non-Indigenous Australians.Potential participants will be recruited via a convenience sampling method. Whilst this will be an opportunistic community-based sample, we will aim to ensure that the gender and age distribution of the cohort will reflect that of the broader Indigenous and non-Indigenous Australian population .Recruitment will take place with the assistance of local key informants with input from local research assistants and primary health care centres. In order to contact and enroll potential participants, BIRCH staff may visit the homes of such potential participants in the company of a local health care staff member or, where appropriate, a community-based Indigenous Australian research assistant.Potential participants will be informed about the study using local media, community groups, Indigenous research staff and local language translators as required. Plain language patient information sheets will be provided and written informed consent obtained.Participant recruitment and data collection for BIRCH will not commence at any site until approval has been granted by the relevant local human ethics research committees. The process of seeking ethics approval will necessarily involve consultation with local community, health and other stakeholders as well as consideration of local cultural issues.CVD risk assessment of participants will entail collection of data outlined in the Box below.At the time of initial assessment, participants will be provided with general feedback regarding their CVD risk profile based on existing Australian guidelines –61 and iEchocardiography with specific reference to left ventricular mass, function and strain studies (STE). This will reflect the approach of earlier studies as undertaken by Marwick and others . Data wiScreening home-based polysomnography (PSG) and/or actigraphy  for asseLipidomic analysis of both whole plasma and lipoprotein sub-fractions will be undertaken at Baker Heart and Diabetes Institute with an emphasis on HDL composition using electrospray-ionisation tandem mass spectrometry and HDL particle size and distribution using nuclear magnetic resonance (NMR) spectroscopy , 64.In addition to collecting and communicating data regarding conventional clinical cardiovascular risk factors, the following investigations will be undertaken:If any of these specific investigations reveal a clinical relevant abnormality then research staff will make contact with participants and their local health care provider to facilitate appropriate follow-up.Cardiovascular event data will be collected based on data linkage using ICD-10  based hoAll cause and cardiovascular mortality will be ascertained using jurisdictional data and the National Death Index maintained by the Australian Institute of Health and Welfare.Once these data have been collected and analysed, any participant found to have a pre-existing CVD based on primary care or hospital separation data will be excluded from further analysis and follow-up. The remaining participants will be stratified according to exposure. Exposure will be defined as the presence of conventional and potentially novel cardiovascular risk factors. Outcome data relating to CVD events and mortality over a five year follow-up period will be collected:Researchers involved in collecting and analyzing participant data under the four separate components of the BIRCH project  will be blinded as to the participant’s assessment in other components of the study. Thus the researchers tasked with collecting data on conventional risk factors, the sonographers and cardiologist reporters involved in measuring and reporting left function and morphology via echocardiography, the technicians and sleep specialists responsible for reporting portable PSG/actigraphy, and the researchers tasked with undertaking lipidomics analysis will be blinded to results obtained by other researchers. Furthermore, the researchers involved in collection of outcome data will be blinded as to each participant’s baseline assessment.All data collected on paper-based forms will be forwarded to an independent data management unit at the MacKillop Institute for processing in REDCap (Research Electronic Data Capture) according to standard operating procedures. Reports from the echocardiogram, PSG, actigraphy and lipidomics investigations will be received in electronic format and integrated with the other personal and health related information. All electronic data will be stored in a password-protected folder on a network drive that is regularly archived.Records will be de-identified and these non-identifiable data will be analysed using STATA version 14 . No identifying information will be published or disseminated upon completion of the study.Data will be stored for five years following the publication of research results or the end of study whichever is the later as per recommended guidelines .Data analysis will comprise descriptive statistics relating to previously detected and undetected CVD, the presence of traditional and potentially novel CVD risk factors, and assessment of CVD risk . The relCVD risk assessment in disadvantaged populations, including Indigenous Australians, remains an ongoing challenge. Despite CVD being a major contributor to overall health disadvantage in Indigenous Australians, there exist significant deficiencies in currently used CVD prediction tools. This hinders the ability of health services to target and deliver primary prevention initiatives to those individuals at greatest need. Furthermore, the presence of increased CVD burden unexplained by traditional risk factor prediction also challenges the existing understanding regarding the mechanisms responsible for CVD development in this unique Indigenous Australian population.In the context of this background, BIRCH aims to enhance the understanding of factors responsible for the greater burden of CVD disease in Indigenous Australians. While many existing and newer markers of early CVD disease or risk may help identify Indigenous Australians at increased risk of development of overt CVD, the BIRCH project specifically focuses on three markers comprising: sleep quality and disorders, left ventricular function and morphology, and select lipidomic species. These markers were chosen as they are potential targets for primary and/or secondary prevention in this setting. If results of the BIRCH study do indeed indicate that these markers are able to explain more of the actual risk of the subsequent development of overt CVD in Indigenous Australian adults then interventions are available that can be employed sooner to prevent or delay the onset of disease.In turn, BIRCH will potentially provide information to inform the development of more accurate tools for CVD risk prediction as a means for improving the targeting of existing and emerging preventive therapies in those at greatest risk of heart disease, ensuring those who can gain the greatest benefit are identified and treated early. Finally, the BIRCH cohort will form a valuable future and ongoing resource to facilitate early evaluation of new treatments to prevent and manage CVD. Together these elements will serve to support the development of effective, appropriate and sustainable interventions to prevent progression and complications of CVD in Indigenous Australians, thereby addressing this significant contributor to health disparity.


The fusion of muscle precursor cells is a required event for proper skeletal muscle development and regeneration. Numerous proteins have been implicated to function in myoblast fusion; however, the majority are expressed in diverse tissues and regulate numerous cellular processes. How myoblast fusion is triggered and coordinated in a muscle-specific manner has remained a mystery for decades. Through the discovery of two muscle-specific fusion proteins, Myomaker and Myomerger–Minion, we are now primed to make significant advances in our knowledge of myoblast fusion. This article reviews the latest findings regarding the biology of Myomaker and Minion–Myomerger, places these findings in the context of known pathways in mammalian myoblast fusion, and highlights areas that require further investigation. As our understanding of myoblast fusion matures so does our potential ability to manipulate cell fusion for therapeutic purposes. Muscle formation occurs during several stages of an organism’s life, including embryonic development, and growth and regeneration in the adult. Proper myogenesis begins through specification of precursor cells to the myoblast lineage, followed by differentiation, both of which are accomplished by the muscle-specific transcription factors MyoD and Myogenin , 2. A crCaenorhabditis elegans [Plasma membrane fusion is a complex process that requires recognition, adhesion, cell signaling, cytoskeletal alterations, and membrane rearrangements. Current evidence, reviewed elsewhere, implicates numerous proteins and regulatory pathways in the recognition, adhesion, and the cell signaling phases of myoblast fusion . However elegans .Until very recently, skeletal muscle fusogens fulfilling these criteria had not been identified in any species. It was in fact a major conundrum whether muscle-specific proteins that directly governed myoblast fusion actually existed. The discovery of the multi-pass transmembrane protein Myomaker as the first muscle-specific fusion factor was the first step toward resolving the confusion surrounding the question of vertebrate myoblast fusion . NotablyDrosophila, zebrafish, and C2C12 myoblasts did not yield muscle-specific proteins directly governing myoblast fusion. One possibility for the elusiveness of these genes could be their inadequate annotation, particularly as Minion–Myomerger represents a small ORF (smORF) encoded microprotein which is below the 100 amino acid thresholds normally used to bioinformatically predict open reading frames [Both Myomaker and Myomerger–Minion were identified through bioinformatic in silico searches. It is surprising that prior to their discovery, the numerous genetic screens performed in g frames .In vitro, these genes are expressed in both myocytes (differentiated mono-nucleated myoblasts) and myotubes. This contrasts with what is found in vivo where Myomaker and Myomerger–Minion are expressed in the developing myotome beginning at embryonic (E) day 10 but no longer detectable after postnatal (P) day 21, when muscle development has ceased. In healthy adult myofibers, Myomaker and Minion–Myomerger transcripts or protein have not been detected. However, upon muscle injury, both genes are activated as muscle stem cells (MuSC) differentiate. Additionally, expression of Myomaker and Myomerger–Minion has not been detected in the other mammalian cell types that fuse such as osteoclasts, sperm-egg, trophoblasts, and multinucleated giant cells. This raises the question of whether homologous proteins perform a similar function in other mammalian fusogenic cells and to what extent completely distinct pathways  activate fusion in the various cell types.Drosophila and other invertebrates is currently unknown. It is unlikely but formally possible that sequence-diverged homologs of Myomaker and Minion–Myomerger remain to be discovered in invertebrates. The more likely scenario is that invertebrates rely on functional homologs that lack sequence similarity. In this case, functional screening of Drosophila cDNAs in mammalian cells (for instance using the reconstitution system described below) may allow identification of such proteins. Alternatively, the basic biophysical mechanisms for fusion could be different between the species and thus require different factors to accomplish membrane coalescence.Myomaker function is conserved to zebrafish where it is expressed only in the fusion competent fast myocytes but not in the fusion-incompetent slow myocytes, and this expression is functional as myomaker deletion results in a lack of fast myocyte fusion , 13. Myo+ cells were detected in both strains of mice during development, indicating an ability to differentiate and thus demonstrating that loss of these genes do not directly regulate differentiation. Cultured myoblasts from each genetically modified mouse line also displayed the ability to differentiate, but not fuse, confirming a highly specific role for these proteins in fusion.In the mouse, genetic loss-of-function experiments for Myomaker and Minion–Myomerger have revealed that these proteins are necessary for muscle formation. Indeed, loss of either gene results in death at birth due to a lack of skeletal muscle, and defects are seen in muscles of both somitic and non-somitic origin. Of note, mono-nucleated myosinAlthough such loss-of-function studies have identified proteins required for muscle cell fusion in a variety of systems, the ultimate demonstration of fusogenic function would come from gain-of-function studies, particularly reconstitution of fusogenic activity in otherwise non-fusogenic cells. The identification of Myomaker represented a significant step toward this goal, in that fibroblast cells expressing Myomaker were capable of robustly fusing to differentiating C2C12 myoblasts—a surprising and dramatic demonstration that Myomaker expression alone is sufficient to confer fusion competence to non-muscle cells. The observation that Myomaker-expressing fibroblasts never fused with one another led to the hypothesis of another activity required for cell fusion, one that could induce fusogenicity by driving pore formation and expansion. The search for this elusive fusion-inducing activity culminated in the identification of Minion–Myomerger. Data from several groups has now demonstrated that co-expression of Myomaker and Minion–Myomerger is sufficient to drive fusion of otherwise non-fusogenic cells –9. This Drosophila myogenesis, with cells expressing both Myomaker and Minion–Myomerger playing the role of “founder” myoblasts and those expressing only Myomaker serving as “fusion competent” myoblasts.From an experimental perspective, the minimal two-component nature of this program offers the chance to investigate mechanisms through gain of function and cell mixing studies. For instance, mixing of differentially labeled populations revealed an intriguing asymmetry: mouse fibroblasts expressing both Myomaker and Minion–Myomerger were capable of fusing to myoblasts expressing Myomaker alone , 8. StatAlthough such a mechanism would reconcile the otherwise disparate-appearing fusion mechanisms of flies and mammals, it must be emphasized that there is currently no in vivo experimental evidence to support the existence of such myoblast heterogeneity in mammals. On the contrary, it has been shown that both the Myomaker and Minion–Myomerger genes contain evolutionarily conserved E-box binding sites for canonical myogenic regulatory factors such as MyoD and Myogenin; these basic helix-loop-helix transcription factors orchestrate expression of a large number of genes required for myogenesis, suggesting that Myomaker and Minion–Myomerger are indeed coordinately expressed. Minion–Myomerger was also reported as a hit in a dropout screen where presumably only one cell in the fusion pair was lacking the protein , suggestWhile the induction of fusion in fibroblasts by expression of Myomaker and Minion–Myomerger demonstrates that these two proteins are sufficient for cellular fusion, additional factors may be involved. One such example is the reported physical association of Minion with cytoskeleton-associated proteins, and the requirement for cytoskeletal remodeling downstream of Myomaker–Minion–Myomerger induced cell fusion . Recent Our understanding of the structural characteristics of these myogenic fusion proteins is in its infancy. Given that the structural analyses of the specific fusogenic proteins that mediate viral and intracellular membrane fusion have provided powerful insights into those phenomena, we anticipate that understanding the structure of Myomaker (221 amino acids) and Myomerger–Minion (84 amino acids), and their respective functional domains, will illuminate the mechanisms underlying myoblast fusion.Overall, Myomaker exhibits minimal homology to other known proteins. Although its primary sequence has modest similarity to Tmem8a and Tmem8b, neither of these orphan membrane proteins possesses the fusogenic activity for fibroblast-myoblast fusion . TherefoDrosophila myoblast fusion. This does not preclude the potential for Myomaker to act across short distances after adhesion has been initiated by other traditional factors, to bring membranes even closer and promote coalescence. Additionally, regions of the protein not embedded in the membrane extend modestly into the extracellular and intracellular space, suggesting that it could drive fusion by acting in cis or trans. A requirement for the intracellular C-terminal domain for fusogenic activity of Myomaker suggests that a function in cis is required. Deletion of the final seven amino acids (amino acids 215–221) or mutation of these amino acids to alanine results in a complete lack of fusion as assessed by the ability of the mutant protein to fuse fibroblasts to myoblasts or rescue the fusogenic defect in Myomaker knockout myoblasts [C. elegans to control the fusogen Eff-1 [Although the current topology model for Myomaker is not yet complete, it remains informative with respect to potential functions. The majority of the protein is embedded in the membrane with only small regions available in the extracellular space, suggesting that it does not act as a traditional adhesion molecule akin to the Ig superfamily that functions in yoblasts . The C-tyoblasts ; howeveren Eff-1 .In contrast to Myomaker, much less is known regarding the structural characteristics of Myomerger–Minion. In silico analysis of secondary structure reveals multiple helical regions that could mediate membrane binding. Portions of this helical region have predicted amphipathic nature  which may influence function, as discussed below. Of these three helical structures, only the N-terminal hydrophobic region (amino acids 5–25) has the required length (18–20 amino acids) to span the membrane bilayer and potentially act as a membrane anchor. Disrupting the positively charged arginines within the first α-helix following the N-terminal hydrophobic domain resulted in a loss of fusogenic activity and perturbed interaction with FLAG–Myomaker as evidenced by co-immunoprecipitation .+ fibroblasts. In keeping with this model, no evidence was detected to support extracellular secretion of Minion/Myomerger [Of note, the N-terminal hydrophobic region has also been interpreted as a cleavable signal peptide; if true, this would suggest that Myomerger–Minion is secreted and may activate fusion through extracellular interactions. However, subcellular fractionation indicates that the vast majority of the protein sorts with membrane fractions, suggesting that it is tethered to a membrane. A potential longer isoform containing sequence upstream of the signal peptide is also predicted in the mouse. While this species is unlikely to be translated as endogenous Minion matches the molecular weight of the “short” 84 amino acid isoform , it is sufficient to fuse myomakeryomerger .A working model that Myomerger–Minion is membrane tethered highlights the need to decipher its membrane topology. Although further experimentation with different epitope-tagged variants is required, a FLAG epitope engineered on the C-terminus of the protein was detectable by immunostaining on live cells, suggesting this region is extracellular . FurtherPerhaps, the most important questions about Myomaker and Minion–Myomerger are their mechanisms of action, specifically whether they participate in the same or different aspects of the fusion process, and how their roles may be integrated into the known pathways that govern fusion. The two main pieces of data that argue for a shared function through forming complexes are the reported interaction between epitope-tagged versions of these proteins when co-overexpressed , and theWith regard to the second point, subtle differences exist between the phenotypes of Myomaker and Minion–Myomerger knockout mice, which could provide insight into distinct functions of each protein. For instance, Minion–Myomerger KO mice displayed more myocytes, with organized sarcromeres, at E17.5. Moreover, it was observed that myocytes within a particular muscle are more closely aligned in Myomerger–Minion KO embryos. While direct side-by-side comparisons are needed to confirm these, and identify new, phenotypic differences, the available functional data also suggest distinct functions. This includes reconstitution experiments demonstrating an asymmetric requirement for the two proteins in the fusion pair, the finding that Myomerger–Minion is not required for Myomaker function, and the fact that neither protein is required for expression or appropriate localization of the other. Given the current biochemical and functional data, our interpretation is that Myomaker and Myomerger–Minion participate in distinct aspects of the fusion program. Whether a physical interaction exists and if so how it is regulated and functions will no doubt be resolved through continued experimentation in additional labs and by the development of more sensitive and robust antibody reagents.Drosophila are well defined (Dumbfounded and Roughest on founder cells and Sticks and stones in fusion competent myoblasts), whereas the adhesion proteins necessary for vertebrate myoblast fusion are not clear [Drosophila and vertebrates.Current models of myoblast fusion in multiple systems indicate a three-step process in which cells first recognize and adhere to each other, followed by induction of close membrane apposition through F-actin protrusions, and ultimately fusion pore formation through lipid bilayer rearrangement. Some components of this process do not appear highly conserved between vertebrates and invertebrates. For instance, the Ig superfamily of adhesion proteins involved in myoblast fusion in ot clear . Jamb anot clear . CompensDrosophila embryos showed that invasion of F-actin protrusions are necessary for fusion, potentially to enhance membrane proximity [Drosophila flight muscles [In contrast, the role of the actin cytoskeleton appears more well conserved. Elegant work in roximity . While t muscles , many of muscles , the GTP muscles , and Rac muscles , 26. Mor muscles .Drosophila adhesion regulator rolling pebbles [IP-mass spectrometry studies of Minion–Myomerger in differentiating C2C12 myoblasts recovered many interacting proteins with annotated functions in cytoskeletal function . Among t pebbles , 31 and  pebbles .Drosophila founder cells [Drosophila fusion competent myoblast [The fusion induced by co-expression of Myomaker and Myomerger/Minion is strongly dependent on remodeling of the actin cytoskeleton and can be blocked by inhibitors of actin polymerization , 7. Whiler cells . These smyoblast . WhetherA potentially related pathway that has recently been implicated in mammalian myoblast fusion involves the role of phospholipid membrane constituents such as phosphatidylserine (PS). Under normal conditions, PS is asymmetrically distributed to the cytoplasmic face of the plasma membrane but can be translocated to the extracellular face in a regulated manner, for instance during apoptosis. It has long been known that myoblast differentiation leads to a non-apoptotic transient exposure of PS that could serve as a signal directly promoting the fusion process . WhetherUltimately, the fusion of membranes requires a lowering of the thermodynamic barriers to bring cell membranes in close proximity and rearrange phospholipids. This is classically achieved through proper expression and localization of proteins that remodel the membrane ; howeverGiven the complexity of mammalian myoblast fusion and the multitude of mechanisms involved, the minimal “two-component” system for fusion induction using Myomaker and Myomerger–Minion has value as a tool for understanding which pathways are core and which are secondary or regulatory. Viruses have likewise evolved simplified systems to drive cell fusion, and comparison to these proteins may help to illuminate the key features of mammalian myogenic fusion regulators.Viral fusogens can be separated into structural and non-structural proteins. The structural fusion proteins are large surface components of enveloped viruses, serve to enhance viral entry, and are not considered further here. In contrast, the non-enveloped viruses of the reovirus family encode a group of small membrane-associated proteins which primarily function to drive target cell syncytialization through cell-cell fusion. These fusion-associated small transmembrane (FAST) proteins do not function in viral entry but are thought to aid in lateral spread within tissue, with the degree of syncytialization correlating with virulence , 44. LikA typical FAST protein  contains an N-terminal myristoylation site, a transmembrane (TM) domain, and an endodomain containing a polybasic motif (PM) and hydrophobic patch (HP). The polybasic motif functions as a Golgi export signal for membrane targeting , whereasThis model is useful in that it makes testable predictions that may help elucidate the mechanism of Myomaker and Myomerger–Minion activity. For instance, the hydrophobic and amphipathic amino terminus of Minion/Myomerger may be analogous to the hydrophobic patch of FAST proteins and may function to lower the energy barrier for formation and expansion of fusion pores. Indeed, this possibility is supported by the observation that the p15 HP can be replaced by a heterologous FAST amphipathic helix . LikewisTaken together, the findings discussed above support a model in which Myomaker expression defines the fusion competence of differentiating muscle precursors, whereas Minion–Myomerger functions to activate fusogenicity potentially through fusion pore formation and/or expansion Fig. . The parDystrophin gene leads to progressive myofiber injury, impaired muscle regeneration, and eventual death. Early work in the mdx mouse model of DMD suggested that exogenously delivered cells arising from either the muscle or hematopoietic lineage could contribute to myofiber formation via cell fusion [The identification of a minimal system for induction of cell fusion opens the door for potential translational applications. For instance, cell fusion has long been suggested as a potential approach for the complementation of genetic deficiencies in vivo. This model has been most clearly articulated in the context of Duchenne muscular dystrophy (DMD), in which mutation in the l fusion , 52. ThiThe identification of Myomaker opened the door to increasing the fusogenic potential of cell-based therapies, thereby increasing their complementation potential. Studies utilizing non-muscle-derived cells, such as fibroblasts and mesenchymal stem cells, indeed demonstrated that overexpression of Myomaker could promote the fusion of these cells into the muscle in vivo . It remaFah, a preclinical model of hereditary tyrosinemia type I; this occurs via fusion of donor cells with Fah-deficient hepatocytes, thereby restoring hepatocyte viability and function [In addition to skeletal muscle development and repair, cell fusion has been reported to occur in a number of physiological and pathophysiological settings. Inflammation appears to be a critical factor in the development of a permissive environment for fusion, and it has been observed that exogenous bone marrow-derived cells extensively fuse to cerebellar Purkinje neurons in experimental models of multiple sclerosis . Moreovefunction . If, as Additional opportunities exist for the therapeutic application of targeted fusion to cancer. Cancer vaccines based on tumor-dendritic cell heterokaryons can potently induce immunity through the enhanced presentation of tumor antigens via both class I and class II MHC , and co-Finally, the heterologous fusion systems discussed here promise to be equally useful in basic research related to cell fate and reprogramming. Early studies on somatic cell nuclear reprogramming relied on chemical methods such as PEG treatment to induce heterokaryon formation , an inefThe discovery of the first two mammalian muscle-specific fusion factors, Myomaker and Minion–Myomerger, marks the beginning of a new chapter in the study of myoblast fusion. Tools now exist to dominantly induce fusion, allowing detailed mechanistic analysis as well as identification of additional novel players. While these studies are only beginning, the existing data have already shed new light on unexpected similarities between fusion in various systems and may lead to further unification of our understanding of fusion in vertebrates and invertebrates. Given the numerous opportunities for clinical application of cell fusion, as well as a recent report demonstrating human mutations in this pathway as a cause of congenital myopathy, we anticipate that our expanding knowledge of the fundamental mechanisms of cell fusion will translate into novel opportunities to positively impact human health.


The increased hunger is associated with elevation of the orexigenic hormone ghrelin and decrements in anorexigenic hormones. The lower total daily energy expenditure with diet-induced weight loss results from (1) a disproportionately greater decrease in circulating leptin and resting metabolic rate (RMR) than would be predicted based on the decline in body mass, (2) decreased thermic effect of food (TEF), and (3) increased energy efficiency at work intensities characteristic of activities of daily living. These metabolic adaptations can readily promote weight regain. While more experimental research is needed to identify effective strategies to narrow the energy gap and attenuate weight regain, some factors contributing to long-term weight loss maintenance have been identified. Less hunger and greater satiation have been associated with higher intakes of protein and dietary fiber, and lower glycemic load diets. High levels of physical activity are characteristic of most successful weight maintainers. A high energy flux state characterized by high daily energy expenditure and matching energy intake may attenuate the declines in RMR and TEF, and may also result in more accurate regulation of energy intake to match daily energy expenditure.Metabolic adaptations occur with weight loss that result in increased hunger with discordant simultaneous reductions in energy requirements—producing the so-called  Obesity, now classified as a disease , has becThe first law of thermodynamics dictates that energy ingested (Ein) and available for cellular metabolism in excess of energy expended (Eout) will result in increased energy stores, and that body energy stores will decrease if energy expenditure exceeds the amount of energy consumed and available for cellular metabolism. This first law has been inappropriately applied in nutrition counseling settings to inform patients/clients as to the magnitude of weight loss they will experience based solely on the degree to which they restrict their energy intake. Regrettably, this approach does not recognize that changing the intake/availability side of the energy balance equation will invariably result in metabolic and/or behavioral changes on the energy expenditure side. When dieting, the magnitude of the initial energy deficit is diminished over time as energy expenditure decreases due to adaptive thermogenesis (discussed later) and the loss of respiring body mass in the form of fat and some lean tissue as well ,9.The second law of thermodynamics dictates that the cellular reactions that generate ATP are less than perfectly efficient. That is, a portion of the energy released from the oxidation of macronutrients will be lost as heat rather than conserved in the phosphoanhydride bonds of ATP molecules. In human metabolism, more energy is lost as heat than is conserved and available for biologic work such as muscle contraction. Energy efficiency maximizes ATP production relative to heat loss or relative to mitochondrial oxygen consumption. In human studies, energy efficiency is often crudely determined based on changes in energy expenditure relative to changes in body mass or fat-free mass (FFM). However, FFM includes a variety of different tissues and organs with widely differing rates of energy expenditure, so attempting to quantify energy efficiency based on FFM rather than the components of FFM can provide less than complete information .There is significant inter-individual variability in energy efficiency in response to well-controlled, experimentally-induced energy excesses and deficits ,12,13, lIt is estimated that only about 20% of individuals who experience significant weight loss are able to maintain their lost weight . Data frMacLean et al. ,28 and oMany metabolic factors contribute to the energy gap following dietary restriction. Observed changes in adiposity-related signals (leptin and insulin), hypothalamic neuronal activity and neuropeptide expression, and gut peptide expression are thought to play a role in the increased hunger in response to weight loss. An exhaustive review of these factors is beyond the scope of this paper. The interested reader is referred to comprehensive reviews on this topic ,27,28,30The hypothalamus integrates many signals from the periphery, including liver, gut, and adipose tissue to regulate energy expenditure and the initiation, termination, and frequency of eating. Homeostatic regulation of food intake occurs such that severe caloric energy restriction leading to weight loss results in a strong internal drive to eat, whereas an overabundance of food intake and weight gain may be followed by a reduction in food intake. Insulin and leptin are putative players in this regard ,32,33,34Speakman ,37,38 haA host of other anorexigenic peptides originate in the gut and typically increase in circulation in response to feeding, which then communicate with the hypothalamus to terminate food intake, increase satiety, and increase satiation between meals ,43,44. TGhrelin is an orexigenic hormone, primarily produced by oxyntic cells of the stomach and is the endogenous ligand for the growth hormone secretagogues receptor type 1a (GHS-R1a) ,46. The 2) with no history of obesity—and “reduced-obese” participants, who were overweight or obese (BMI 27–32 kg/m2) but recently lost weight in a weight-loss program. The study was designed to analyze brain responses to food images in the overfed state versus eucaloric state. Among thin individuals, overfeeding attenuated neural activation compared to that observed during the eucaloric state. This response to overfeeding did not occur among reduced-weight overweight/obese individuals. In the baseline fasting state, thin individuals had a much more robust neuronal response to food-related visual cues than reduced-obese individuals. Overfeeding resulted in significant attenuation of the response to visual foods cues in thin but not reduced-obese individuals.Additionally, compared with normal-weight individuals with no history of obesity, individuals who are overweight or obese, but have lost weight, have a different neural response to overfeeding . In a raMuch of the research on body weight/composition regulation has been adipocentric—that is the control of energy intake and expenditure has focused on adiposity-related signals as discussed above. However, recent studies have demonstrated that fat-free mass (FFM) and resting metabolic rate (RMR) are positively associated with energy intake ,55. FFM Taken together, diet-induced weight loss appears to impact appetite via changes to both fat mass and FFM by altering known and unknown peripheral factors that communicate to the brain a state of nutrient deprivation. These changes result in increased hunger , which iTotal daily energy expenditure (TDEE) is a function of four components: resting metabolic rate , the thermic effect of food (TEF: the increase in energy expenditure above RMR in response to food ingestion), non-exercise activity thermogenesis (NEAT: energy expenditure above RMR required to support the activities of daily living as well as fidgeting), and exercise energy expenditure (ExEE: energy expended above RMR necessary for performing exercise). NEAT and ExEE together make up physical activity energy expenditure (PAEE). Diet-induced weight loss almost always causes significant decreases in TDEE, which can negatively impact the maintenance of lost weight. As far back as 1984, Leibel and Hirsch reported lower TDEE in post-obese compared to never-obese individuals—on the order of 25% lower than predicted by metabolic body size . In a laThe TEF is reduced with dieting because there is a reduction in the total caloric load that requires obligatory digestion and absorption. A portion of the normal TEF results from increased sympathetic nervous system activity that accompanies food ingestion , which aNumerous studies have shown that weight loss-induced declines in RMR contribute to the reduced TDEE. The reduction in RMR is partially the result of the loss of respiring body mass, but many studies ,63,64,65There is substantial evidence of AT in regard to RMR with weight loss. For example, Leibel et al. found that a 10–20% weight loss in obese patients caused a decrease in RMR of 3–4 Kcal/kg fat-free mass per day . In the The effect of diet-induced weight loss on ExEE and NEAT is quite variable. From a thermodynamic perspective, without changes in movement efficiency and economy, the loss of body mass will result in fewer calories expended to perform the same weight bearing movements. When considering the energy expenditure incurred from activities of daily living at the lower body weight, this could conceivably contribute substantially to the reduction in total daily energy expenditure. Indeed, following a 10% weight loss, individuals were found to exhibit greater skeletal muscle work efficiency at lower intensities, which Rosenbaum et al. estimated could account for one-third of the reduction in PAEE . Others To summarize the changes in energy expenditure, diet-induced weight loss can result in significant reductions in RMR, TEF, ExEE, and NEAT. These metabolic changes that result in lower energy expenditure would not obligatorily contribute to weight regain if there were a proportional decrease in food intake. However, as discussed earlier, body composition and hormonal changes occur with weight loss that are associated with increased rather than decreased appetite. This scenario coupled with the environmental pressures of modern society that favor excessive calorie consumption and minimize physical activity, could lead to a sense of futility among both patients and practitioners regarding lifestyle obesity treatment. However, as MacLean et al.  suggest,Thus, the purpose of this second section is to examine lifestyle approaches to attenuate the increase in hunger and the decrease in energy expenditure that accompany weight loss and to identify important areas for further research.While some individuals are genetically more susceptible to obesity and will likely find it difficult to maintain weight loss , there iThere currently are no clear-cut dietary recommendations for successful weight maintenance following weight loss. In experimental studies examining dietary approaches to curb appetite during weight maintenance, there is substantial inter-individual variability in response to the different diets, suggesting that a one-size-fits-all weight maintenance dietary approach is not likely to be effective for all. Clearly, there is need for more experimental research on dietary patterns and weight maintenance, with special focus on approaches tailored to the unique behavioral and metabolic characteristics of the individual. While much research has focused on understanding the biologic basis for changes in hunger and satiety associated with acute energy perturbations, fewer studies have addressed experimental approaches to reduce hunger over the long-term following weight loss. With this caveat in mind, the following sections provide a brief review of the possible role of diet composition in attenuating the increased hunger that occurs with weight loss. There are other dietary features that might also be useful in minimizing weight regain such as time restricted feeding to better match meal consumption with circadian rhythms ,89, alte−1 and 1.6 g·kg−1, with each meal providing at least 25–30 g protein. The greater satiety and better weight maintenance outcomes seen with higher protein intakes [Diets of varying macronutrient composition  for weight loss have been the focus of significant research in the past two decades. However, the possible effects of dietary macronutrient composition on weight maintenance following weight loss have received far less research attention, especially in regard to long-term (years) results. Theoretically, a dietary pattern that enhances weight loss maintenance by attenuating the energy gap would result in less hunger and greater satiation relative to energy intake and less reduction in energy expenditure relative to energy intake. Given the greater satiating value of protein compared to other macronutrients ,94,95 al intakes  may be r intakes ,100,101.Another aspect of dietary macronutrient composition that may have relevance to attenuation of the weight loss-induced energy gap is the glycemic load, a function of the glycemic index  multiplied by the total amount of carbohydrate in the diet. Observational studies described in a prior section have found weight loss maintenance to often be associated with consumption of low energy dense foods including vegetables, whole fruits, and legumes, and low carbohydrate foods such as lean meats, which together would constitute a low-to-moderate glycemic load eating pattern. Experimental studies have shown that meals with lower energy density result in lower energy intake . HoweverA number of epidemiological studies have found high fiber intakes to be associated with lower food intake and improved weight control ,108,109.While exercise is the most obvious volitional approach to increasing energy expenditure, the different constituents of TDEE each represent a potential target for interventions.Because RMR accounts for upwards of 75% of TDEE, attenuating its decline could have major implications for weight maintenance. Elevating total caloric turnover while maintaining energy balance may be one possible method for attenuating the fall in RMR. We have previously seen in young males , femalesMacronutrients differ in the thermic effect they elicit, with fat having the lowest thermic effect at approximately 3% of ingested fat calories, carbohydrates producing a thermic effect of 5–10% of ingested carbohydrate calories, and protein consumption eliciting a thermic response of 20–30% of protein calories . TherefoTotal daily energy expenditure varies widely, even among individuals of similar age and body size  and becaExercise typically produces less energy deficit and less weight loss compared to energy-restricted diets, and its impact on body weight regulation has been questioned . NeverthIn 1956, Mayer and colleagues  observedA physically active lifestyle may be important not only for its contribution to energy expenditure, but also for its role in matching the energy desired with the energy the individual requires. Regular exercise may heighten the brain’s sensitivity to leptin and insulin, and enhance satiety signals coming from the periphery . A substIn regard to exercise and physical activity, the transition to energy balance during weight maintenance can be approached in several ways. Weight can be maintained by pairing a low energy intake with a low energy expenditure (low flux state), resulting in energy balance with a low total throughput of energy. Alternatively, energy balance can be maintained with a significantly greater energy throughput by coupling a higher expenditure via exercise with a higher intake (high energy flux) . While b2 max with daily energy intake set at RMR  × 1.7 to maintain energy balance) and 5 days of low flux . Mean RMR was significantly higher during high flux compared to low flux, suggesting that higher energy flux may help attenuate the decline in RMR in in obese individuals attempting to maintain lost weight. Also, resting fat oxidation was higher in the high compared to the low flux condition, which may have implications for fat balance. Importantly, while in the high flux condition, study participants reported significantly lower levels of hunger and greater fullness in comparison to their days spent in low flux, despite being in energy balance across the two different flux conditions [One could question whether previously obese and/or sedentary individuals can achieve adequately high levels of ExEE to maintain a state of high energy flux. To answer that question, we recently conducted a study  in whichnditions . Given tnditions . Also innditions .Successfully maintaining lost weight is difficult for many obese and overweight individuals who diet to lose weight. Metabolic adaptations occur with weight loss that result in increased hunger, which is discordant with simultaneous reductions in energy requirements—producing the so-called energy gap in which more energy is desired than is required. These metabolic adaptations have been characterized, with the increased hunger and decreased satiety and satiation associated with elevation of the orexigenic hormone ghrelin and decrements in anorexigenic hormones including CCK, PYY, and GLP-1. The lower total daily energy expenditure that accompanies weight loss results from (1) a disproportionately greater decrease in circulating leptin and RMR than would be predicted based on the decline in body mass, (2) lower decreased thermogenesis owing to reduced food consumption, and (3) increased energy efficiency at work intensities characteristic of activities of daily living. These metabolic adaptations can readily promote weight gain, especially in the face of an obesogenic environment characterized by abundant access to highly palatable, energy dense food and low levels of physical activity energy expenditure. However, weight regain following weight loss should not be viewed as inevitable. While more experimental research is needed to identify effective strategies to narrow the energy gap and attenuate weight regain, factors contributing to long-term weight loss maintenance have been identified from observational studies and some experimental studies. Vigilant self-monitoring of body weight and reduced time in sedentary activities including television viewing appear especially important. Lower hunger and greater satiation have been associated with higher intakes of protein and dietary fiber, and lower glycemic load diets. High levels of PAEE are characteristic of the vast majority of successful weight maintainers. One promising approach to tackle the elevated hunger and reduced energy expenditure that occur with weight loss is the establishment of a high energy flux state in which a high throughput of calories occurs, owing to high daily energy expenditure and matching energy intake. A high flux state compared to a low flux, sedentary state has been found in both observational and experimental studies to result in higher RMR and TEF. Also, energy intake may be more accurately regulated to match energy expenditure in a high flux state. More research is required to determine the magnitude of PAEE required to achieve an effective high energy flux state, whether the intensity of exercise in a high flux state affects the energy gap, and what might be the most advantageous dietary macronutrient composition for a high flux state to narrow the energy gap.


Overall Abstract: This symposium will draw together state of the art findings on the lifelong cognitive trajectories, on key-predictors of cognitive functioning and the functional consequences of cognitive impairments in schizophrenia and related psychotic disorders from developmental epidemiological, prodromal, and clinical research. Four speakers will take the audience through new findings on the cognitive course of the lifespan, ranging from childhood to old age. Specifically, the talks will address four key-questions:1) Which areas of cognitive functioning are impaired and when does this impairment start?2) How well can cognitive functioning predict the development of a psychotic illness, as well as diagnostic and functional outcome?3) Does cognitive functioning remain stable after illness onset or are psychotic disorders characterized by continuing decline? When does decline occur and is it possible to predict it?4) And what is the functional sequelae of specific cognitive impairments in older adults with schizophrenia?Specifically, Dr. Mollon will present new data examining the origin of cognitive impairment across the psychosis spectrum using a population-based cohort followed prospectively from birth. Her findings demonstrate that while individuals with affective psychosis, subthreshold psychotic experiences and even depression experience some degree of cognitive impairment across the first two decades of life, only those who go on to develop non-affective psychosis exhibit large, widespread and increasing deficits.Most studies of neurocognitive functioning in Clinical High Risk (CHR) cohorts have examined group averages, likely concealing heterogeneous subgroups. The study of Dr. Velthorst therefore used two independent methods to identify neurocognitive subgroups in a large population at Clinical High Risk for developing psychosis. Her findings show that neurocognitive profiles vary substantially in their severity and are associated with diagnostic and functional outcome, underscoring neurocognition as a predictor of illness outcomes.Dr. Fett will present recent research on cognitive functioning in a large sample of patients at first hospitalization for a psychotic disorder who have been followed 20-years into the illness. Her findings indicate that cognitive functioning in psychotic disorders continues to decline after illness onset, that this decline is not specific to schizophrenia but present across psychotic disorders, and that, relative to never-psychotic individuals, impairments on some key-cognitive domains worsen with age. Decline could not reliably be predicted by key patient characteristics at baseline.Lastly, Dr. Harvey will share novel data on the course of cognitive functioning in middle aged and older patients with schizophrenia. His findings demonstrate that cognitive impairments are moderated in their impact on everyday outcomes by the presence of severe communication abnormalities. Interestingly, verbal under-productivity and disconnected speech had different functional correlates, with under-productivity impacting clinician rated social outcomes and performance on measures of interpersonal social competence.A lifetime focus on cognition is paramount in order pinpoint critical periods for prevention and intervention. This symposium seeks to present a comprehensive overview of the cognitive landscape of psychotic disorders by integrating findings on predictors and consequences of lifelong cognitive functioning of individuals diagnosed with a psychotic disorder.


The efforts toward individualized medicine have constantly increased in an attempt to improve treatment options. These efforts have led to the development of small molecules which target specific molecular pathways involved in cancer progression. We have reviewed preclinical studies of sunitinib that incorporate sex as a covariate to explore possible sex-based differences in pharmacokinetics and drug–drug interactions (DDI) to attempt a relationship with published clinical outputs. We observed that covariate sex is lacking in most clinical outcome reports and suggest a series of ethic-based proposals to improve research activities and identify relevant different sex outcomes. We propose a deeper integration of preclinical, clinical, and translational research addressing statistical and clinical significance jointly; to embed specific sex-divergent endpoints to evaluate possible gender differences objectively during all stages of research; to pay greater attention to sex-divergent outcomes in polypharmacy scenarios, DDI and bioequivalence studies; the clear reporting of preclinical and clinical findings regarding sex-divergent outcomes; as well as to encourage the active role of scientists and the pharmaceutical industry to foster a new scientific culture through their research programs, practice, and participation in editorial boards and Institutional Ethics Review Boards (IRBs) and Research Ethics Committees (RECs). We establish the IRB/REC as the centerpiece for the implementation of these proposals. We suggest the expansion of its competence to follow up clinical trials to ensure that sex differences are addressed and recognized; to engage in data monitoring committees to improve clinical research cooperation and ethically address those potential clinical outcome differences between male and female patients to analyze their social and clinical implications in research and healthcare policies. The research drive toward personalized medicine has grown exponentially during the last years in an attempt to improve therapeutic outcomes putting the patient at the center of the healthcare system. The shift toward precision medicine has deeply focused on diagnosis and treatment, particularly in the oncology field . The devFigure 1). Based on a deeper understanding of their nature, sex differences have been grouped in four types  quaur types . Althougur types  for diffur types , there hur types . In a laur types . Howevers fields  includins fields , 2017, ns fields , medicats fields , incidens fields  as well s fields . This evs fields .Tyrosine kinase receptors are a subset of protein kinases involved in numerous cellular processes . BecauseSunitinib is a small molecule, able to inhibit a multiplicity of TK receptors including the platelet-derived growth factors PDGFRα/β, the colony stimulating factor type I (CSF-1R), the vascular endothelium growth factor (VEGFR), the stem cell factor c-KIT, and the fetal liver TK receptor 3 (FLT3) amongst others . Owing tWe have carried out the analysis of the translational impact of covariate sex on an anticancer drug such as sunitinib and compiled various ethic proposals to better address the impact of sex on clinical outcomes and trigger scientific discussion to improve the design of future research.The translational relevance and clinical interest of covariate sex applied to sunitinib treatment has been assessed at different levels: (1) the possible sex-divergent pharmacokinetics of sunitinib and (2) the differential effect of DDI on male and female sexes.versus female mice (Table 1).The effect of sex differences on the pharmacokinetics and tissue distribution of TK inhibitor sunitinib was evaluated in a preclinical mouse model . In bothAn exploratory attempt to correlate these findings with clinical outcomes in male and female patients was previously done, although it was limited to individual cases and case series reports of patients treated with sunitinib . The indLast, sex-divergent differences in clearance and volume of distribution parameters between male and female patients have been shown  which suTable 1): diclofenac and paracetamol decreased plasma exposure in male mice which compares with a general reduction of sunitinib exposure in tissues of interest. However, female mice did not show any DDI effect in plasma and sunitinib exposure remained unchanged eventhough large effects did take place in tissues. Last, ibuprofen showed the opposite behavior: no effect was observed in sunitinib plasma exposure in male mice while significant changes took place in tissues. Similarly, after multiple dose administration to mice (Table 2).The covariate sex may also have a significant impact on DDI outcomes given the fact of cancer patient polymedication  includin to mice , sex-divLikewise, clinical observations identified numerous DDI with sunitinib , includiThere is an often forgotten translatability issue: TDM. Usually, TDM protocols are developed during the clinical stage and focused on a surrogate parameter (marker) that indicates treatment efficacy or toxicity. Generally, these surrogate parameters are either drug or biomarker concentrations in a measurable matrix  or genotyping able to relate to efficacy or toxicity .Table 1). Thus, plasma concentration based TDM would probably detect a DDI with paracetamol or diclofenac in a male patient and a possible DDI with ibuprofen in a female patient. Then, subsequent dosage adjustment may be carried out. However, it is likely that the DDI would remain undetected in female patients taking paracetamol or diclofenac and in male patients taking ibuprofen. In this case, their coadministration would mask changes in brain, liver or kidney sunitinib penetration and exposure as these do not correlate with the events occurring in plasma (Table 1). Thus, if monitored parameters remain unchanged in plasma , or cannot be measured in tissues, then, understanding male-female DDI differential effects becomes essential to carry out TDM successfully  is allowing the identification of patient oriented selection strategies , they affect the therapeutic response of the subject . These dsporters . Therefosporters . Amongstsporters . Becausesporters , the anasporters  or even sporters , 2016. Isporters .The significance of the covariate sex in crucial to identify possible differences. Women have gradually increased participation in clinical trials  and theip-value, e.g., p < 0.05  is clinically relevant. Factors such as higher medication use including non-prescribed and self-medication, comorbidities and age differences in female patient population  can indicate whether differential pharmacotherapy approaches in male and female patients are needed. If sex difference impact is inferred, then therapeutics may be adjusted and precision medicine realized , scientists and study coordinators for implementation of sex-differentiated data analysis in the results section of the study report, regardless of the realized .We would greatly encourage researchers and authors to implement sex-differentiated reporting of efficacy, toxicity and endpoint attainment, as well as editors to require this analysis in their scientific publications . FurtherThe IRB and/or the REC are the check point to ensure the clinical research proposal is sound and carried out ethically. This mission requires from the reviewing process to assess whether the goals and objectives of the proposal are rigorous, the pursued outcomes present a logical clinical expectation and are methodologically achievable . This as“bedside-to-bench-and-back-to-bedside” approach, the study proposals need to be sufficiently justified and supported with published preclinical information to demonstrate the rationale for the new clinical study/trial application. The IRB should request a preliminary assessment of sex-divergent outcomes based on the observations that lead to the new study application. In addition all clinically relevant evidence (“bedside”) of prior clinical trials submitted to the IRB/REC may include a posterior evaluation of possible sex-divergent outcomes and address how the endpoints may assess the difference between male and female patients. The supporting preclinical (“bench”) data should include details about the sex of the cells and animals used and an analysis of sex as a covariate in cell-based assays  as well as animal studies  carried out to anticipate its relevance in therapeutics  is withheld, new information is provided to the scientists conducting the clinical trial as well as the integrity of the study is preserved .“matters scientifically, it matters ethically and it matters socially” (The contribution of the IRBs and RECs to assess sex differences goes beyond the review process task given the nature of the findings. Their assessment ocially”  as it aiocially” . Althougocially” . Thus, iocially” .The translational perspective of covariate sex integrating preclinical and clinical evidence of TK sunitinib suggests the importance of addressing systematically sex-divergent therapeutic outcomes. This information should be available to foster and enhance future clinical research as well as to improve current treatment options. The IRBs and RECs, together with the pharmaceutical industry and other research sponsors play a key role to ensure that possible differences between male and female patients are identified, taken into account and an assessment of their clinical relevance performed. Empowering and expanding the role of IRB and REC to pursue this analysis may result in valuable inputs and improve future clinical trials outputs.Overall, although we have focused our evidence in a specific drug used in oncology, these proposals and views may be incorporated in other therapeutic areas. Furthermore, their review could serve to echo and foster scientific discussion at the technical and ethical level to emphasize the importance of developing studies addressing sex differences that would render the possibility to personalize treatments and benefit patients.IS conceptualized, designed the review, and wrote the manuscript, PM contributed to the conceptualization and writing of the manuscript. CF and EM contributed to the conceptualization of the manuscript. All authors participated and contributed critically to the review, discussion, and analysis of the information. All authors revised the manuscript and approved it.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Functional MRI is increasingly recognized for its potential as a powerful new tool in clinical neuropsychology. This is likely due to the fact that, with some degree of innovation, it is possible to convert practically any familiar cognitive test into one that can be performed in the MRI scanning environment. However, like any assessment approach, meaningful interpretation of fMRI data for the purpose of patient evaluation crucially requires normative data derived from a sample of unimpaired persons, against which individual patients may be compared. Currently, no such normative data are available for any fMRI-based cognitive testing protocol. In this paper, we report the first of a series of fMRI-compatible cognitive assessment protocols, a matrix reasoning test (f-MRT), for which normative samples of functional activation have been collected from unimpaired control subjects and structured in a manner that makes individual patient evaluation possible in terms of familiar z-score distributions. Practical application of the f-MRT is demonstrated via a contrastive case-study of two individuals with cognitive impairment in which fMRI data identifies subtleties in patient deficits otherwise missed by conventional measures of performance.


Self-regulated learning is the individual’s ability to effectively use various strategies to reach their learning goals. We conducted this scoping review to explore what has been found regarding self-regulated learning in the clinical environment and how this was measured.Using Arksey and O’Malley’s five-stage framework, we searched three medical and educational databases as well as Google Scholar for literature on the self-regulated learning of medical students in the clinical environment published between 1966 and February 2017. After results were screened and relevant studies were identified, the data was summarised and discursively reported.The search resulted in 911 articles, with 14 articles included in the scoping review after the inclusion criteria was applied. Self-regulated learning was explored in these studies in various ways including qualitative, quantitative and mixed methods. Three major findings were found: 1) levels of self-regulated learning change in the clinical environment, 2) self-regulated learning is associated with academic achievement, success in clinical skills and mental health and 3) various factors can support self-regulated learning levels in medical students.Most of articles exploring the self-regulated learning of medical students during the clinical years have been published in the last 5 years, suggesting a growing interest in the area. Future research could explore the self-regulated learning levels of medical students during the clinical years using a longitudinal approach or through the use of novel qualitative approaches. Self-regulated learning (SRL) is the process where one is ‘metacognitively, motivationally, and behaviourally proactive in the learning process’ . More sp.According to some theories, SRL can be categorized into four processes and four areas , 7. The Despite the importance of SRL in medicine, and the significant influence of the environment on student learning , SRL canAlthough scoping reviews are relatively new , they arWe used the five-stage framework proposed by Arksey and O’Malley in this scoping review which involves (1) identifying the research questions, (2) identifying relevant studies, (3) selecting the relevant studies, (4) charting the data, and (5) collating, summarising and reporting the results. As recommended by the five-stage framework, we did not use a quality appraisal tool for each study, but we followed the explicit process to ensure that our search was replicable, thus increasing the rigour of our findings .What has been found regarding the self-regulated learning of medical students in the clinical years of their program?How has self-regulated learning in medical students in the clinical learning environment been measured?The purpose of our review was to explore the SRL of medical students in the clinical environment. Our research questions were the following:A literature search was carried out to identify studies and reports between 1966 and February 2017 using the databases Medline (Ovid), ERIC, EBSCO and Google Scholar. The main search term “self-regulated learning” was combined with key terms and variants of “medical student”, “medical school” and “medical education”. The search identified 911 potential papers. The following criteria were used for the review and selection of the studies: available in English, focused on SRL in the medical clinical learning environment and relevant to the topic after review of abstracts. Articles investigating similar but different theories such as self-directed learning were not included due to conceptual differences , 17. TheThe full texts of the remaining 21 articles were read by KC and 7 articles were excluded as the studies focused solely on non-clinical medical students. The reference lists of all selected publications were then hand searched for any additional relevant studies. Authors of key papers were also contacted for relevant reports or publications. Fourteen articles were selected for inclusion in the review  changes in SRL occur in the clinical learning environment, 2) higher levels of SRL are associated with higher academic achievement, more success in clinical skills and better mental health outcomes and 3) certain factors can influence SRL levels. These are further explained below.Three studies qualitatively explored how students adapt to different clinical environments. White found that students from a PBL curriculum adapted more effectively and used more SRL skills when transitioning to the clinical environment than students from a traditional curriculum . In WoodFive studies reported correlations between levels of SRL and academic achievement during clerkship –24. SobrNguyen et al.  observedThis review found the SRL of medical students can be affected by a wide range of variables , 26–28. Methods used to study SRL were varied and included qualitative, quantitative and mixed-method approaches.Qualitative processes such as semi structured interviews and focus groups were used in five studies , 19, 27.Quantitative approaches included the Self-Regulated Learning Measure for Computer-based learning (SRMC) , the SelThe SRMC tests the use and frequency of 10 SRL subcategories  and two classes of non-self regulation behavior (will power and non-applicable statements). The tool has high internal consistency and high inter-rater reliability .The Self–Regulated Learning Perception Scale uses 41 items to measure 4 domains; 1) motivation and action to learning 2) planning and goalsetting 3) strategies for learning and assessment and 4) lack of self–directedness . The scaThree articles used the MSLQ , 24, 25,One study used Q-methodology, a method that uses features of both quantitative and qualitative measures . In the In this review, the literature on the SRL of medical students in the clinical environment was systematically searched and summarized. All but three studies , 26, 27 The four processes and four areas of SRL were well covered by the included studies (see Table Two studies that investigated clinical transition highlighted the importance of self-regulating learning to maximise experiential learning , 18. TheBeyond the transition period, Berkhout et al.’s study suggests SRL continues to develop in the clinical environment, with novice and experienced learners having different needs to support their SRL. The study notes that novice students require more support from others, specifically residents and peers, to help them formulate learning goals and navigate the new learning environment. In the broader medical education literature, the significance of resident teaching is mixed, with some studies highlighting the correlation between resident teaching and medical student learning  and acadThis review indicates that higher levels of SRL may be beneficial for medical students due to its positive correlation with academic achievement and clinical skills –24 as weThe positive correlation between SRL and diagnostic and clinical skills is important for students not just when they are in medical school but also as practising physicians. Furthermore, as doctors must maintain their competencies and skills throughout their careers in order to consistently meet the high standards of patient care –43, theyThe negative association between SRL and depression is significant because medical students as a whole face higher levels of psychological stress than the general population , 48, witIn their study, Turan et al. found that medical students in the clinical years appeared to intrinsically have higher levels of SRL than preclinical medical students . AlthougFinally it appears that at least some aspects of SRL can be targeted and improved, with one study suggesting that levels of reflection in students can be increased after an intervention. This finding that SRL can be improved by certain interventions is supported by research outside the transitions literature , 61–63, Several approaches have been used to measure SRL in clinical medical students and there appears to be no single best approach with each approach having its advantages and disadvantages. For example, studies using the qualitative approach were effective in identifying and exploring contextual factors related to SRL in the clinical environment whereas the quantitative approach could identify the strengths of associations to test hypotheses. Two of the three studies using quantitative methods developed their own instruments specifically for use in the clinical context , 26. In Studies which investigated SRL across multiple cohorts reported the cross-sectional design as a limitation. These authors suggest longitudinal studies may allow the transition to be better measured. Studies using questionnaires as their collection tool reported reliability and validity limitations, especially as they measured self-reported learning behaviours, which are subject to social desirability and recall bias. However each survey instrument had reasonable psychometric properties. Some studies also reported the possibility of selection bias either due to small sample sizes  or due tNot identified in our search were novel qualitative methods used in the broader SRL literature, such as think-aloud protocols , 67, vidThis review was restricted to articles published in the English language and those concerning the medical profession. We also did not include studies with similar terms to SRL such as self-directed learning in our review due to conceptual differences in perspectives and constructs. We did include medical and educational databases as well as Google Scholar to widen the search to where SRL research might be catalogued. Future research should consider including nursing and other allied health professionals as learning in the clinical environment is also a core part of health professional education and findings about effective learning could be transferrable across professions. As this article is a scoping review and not a systematic review, we did not critically appraise the selected studies. We did however locate findings which addressed our aims of examining the nature and extent of the literature, and to identify potential future research directions.We explored what is known about medical students in the clinical environment, specifically in relation with SRL. We found several relevant papers, with most published in the last 5 years, suggesting a growing interest in SRL in the clinical environment in medical education. Although most had a cross-sectional or qualitative design, quantitative approaches may yield complementary insights, with longitudinal research being needed to examine how SRL develops and is maintained in clinical learning environments. Additionally future studies could consider using novel qualitative methods to explore the SRL of medical students in the clinical environment.


Pain is the most common complaint in the emergency department (ED), and emergency physicians face unique challenges in making opioid-related treatment decisions. Medical students and residents experience significant variation in the quality of education they receive both about opioid prescribing as well as substance-use detection and intervention in the ED. To achieve a better standard of education, clinical educators will need to (a) develop a clearer understanding of the risk for aberrant opioid prescribing in the ED, (b) recognize prescribing bias and promote uptake of evidence-based opioid prescribing guidelines in their EDs, and (c) advocate for integrated opioid management and addiction medicine training formally into medical school curricula. At the start of training, medical students crave and cherish each patient encounter, are idealistic about the healing power of medicine, and have peak levels of empathy and benevolence.2The emergency department (ED), where the incidence of complex social situations is particularly high and providers are expected to make quick decisions with limited information, exposes students to many challenging patient interactions.For medical students in their early clinical training, this type of patient encounter is new and frustratingly equivocal. Voicing suspicion regarding aberrant medication-seeking behavior based on a patient’s medical history, specific requests, or behaviors can be uncomfortable for students, as it can feel like patient profiling and contradict their perceived role as benevolent caretaker. Compounding this discomfort is the very appropriate, overwhelming fear of unnecessarily and wrongly prolonging patient suffering. Conversely, as students develop a more pragmatic impression of medicine and work on building clinical acumen, they recognize the existence of patient dishonesty as a part of opioid addiction and are concerned about the dangers of inappropriate opioid prescribing.Students have not garnered enough wisdom to feel confident in their assessment of the “legitimacy” of patient analgesia requests, and thus rely on their clinical educators for guidance. However, students often experience greatly varying attending approaches to these patient encounters. EP prescribing behaviors vary along a spectrum, even within a single institution.Medical students’ varying experiences are not just anecdotal. In a national survey, at least 10% of EPs indicated they were less likely and 10% indicated they were more likely to prescribe opioids when they were presented with identical case scenarios.6Pain is subjective, but opioid prescribing decisions do not have to be altogether subjective and idiosyncratic. EPs have the potential to be role models and educators to the next generation of prescribers. While providing effective education on opioid prescribing is a responsibility that should be met by providers in all clinical settings, unique challenges make the ED a particularly difficult setting for making opioid prescribing decisions. In this paper, we advocate that EPs achieve a better standard of education on safe opioid prescribing by (a) developing a clearer understanding of the risk for aberrant opioid prescribing in the ED, (b) recognizing prescribing bias and promoting uptake of evidence-based opioid prescribing guidelines in their EDs, and (c) advocating for earlier integration of opioid management and addiction medicine training formally into medical school curricula.The ED is a setting where prescriptions for short-term opioids are frequently provided. Pain has been found to be the most common patient complaint in the ED with two-thirds of all visits being related to pain.5Opioid prescriptions offered in the ED tend to be aligned with short-term treatment goals. EPs most commonly prescribe immediate-release combinations and are significantly less likely to prescribe high doses or large quantities of opioids, which are more strongly associated with morbid outcomes such as overdoses.There are few studies available that evaluate opioid use behaviors among patients discharged with opioids from the ED. One study does demonstrate that a percentage of those who are prescribed opioids progress to more frequent use; among patients who presented with low back pain and were prescribed opioids on ED discharge, 46% were still using opioid analgesia three months post-discharge.13As EPs face the challenge of balancing their professional and moral duty to alleviate pain with their efforts to minimize opportunities for abuse of prescription opioids, more prospective studies on the degree to which short-term, low-dose opioid prescriptions lead to addiction are needed. Moreover, prospective studies could help identify risk factors that lead to misuse of or addiction to short-term, low-dose opioid prescriptions.Findings from prospective studies would help better inform ED providers about the actual risks of different prescribing patterns, potentially leading to more evidence-based prescribing habits. This more robust evidence should then be shared in medical school classrooms to equip students with a clearer understanding of the physician role in the opioid epidemic. Findings would also provide students with evidence-based opioid-misuse screening tools to incorporate into their clinical algorithms for pain management decisions.EPs should critically assess their individual prescribing patterns to limit individual biases and assess how well they reflect prescribing guidelines. There are multiple biases that impact ED prescribing patterns. ED opioid-prescribing decisions are known to vary by race: White patients who present to the ED are more likely to receive opioid analgesia than any other racial group.14EPs must limit perpetuation of long-standing racial and gender-related biases in clinical treatment to the next generation of opioid prescribers. EPs should reduce implicit bias by adapting their algorithms to emerging clinical guidelines and using evidence-based tools, including PDMPs to assess the risk of opioid misuse. Though recent studies have shown that PDMPs have not altered the average number of controlled substances prescribed per patient, providers perceived that PDMPs influenced their prescription decisions and felt more confident in their treatment decision after using these resources. 14There are many guidelines that have been produced for prescribing opioids for chronic pain, and many of these have been critically evaluated by authorities such as the Centers for Disease Control and Prevention, National Institute on Drug Abuse, and Substance Abuse and Mental Health Services Administration.19Hospital administrators should use strategies such as incorporating reminders and decision algorithms into electronic decision support systems or academic detailing to integrate clinical guidelines into everyday practice.Individual EPs have tremendous potential to improve quality of opioid prescribing education by role-modeling evidenced-based prescribing approaches in the ED. However, education on opioid management should begin early, before medical students enter the wards. Residents have expressed sentiments of under-preparedness in making opioid prescription decisions.The State of Massachusetts is beginning to recognize the need to aim interventions for managing the opioid crises at the roots. Massachusetts was the first state to develop a governmental initiative to reform opioid education for medical students and physicians in training, working with medical schools within the state to provide recommendations for and establish a commitment to medical school curriculum changes that would educate and train medical students on safe prescribing of opioids.22Educational strategies such as these will provide a structured, more evidenced-based foundation for medical students to begin developing their approach to preventing, diagnosing, and treating addiction. By incorporating such education into the formal curriculum, medical schools would be ensuring, rather than leaving to chance, that all students receive consistent, high-quality training on opioid use disorder prevention and treatment. National medical education accrediting bodies should develop similar initiatives to catalyze opioid education reform. As opioid use is a national problem, it is critical that the standard of education on safer opioid prescribing be raised uniformly across the United States.