Considering the proposed simulation-estimation processes as common mechanisms for interaction between motor and perceptual systems, we outline how mental imagery (of speech) relates to perception and production, and how these hypothesized mechanisms might underpin certain neural disorders.The neural basis of mental imagery has been investigated by localizing the underlying neural networks, mostly in motor and perceptual systems, separately. However, how modality-specific representations are top-down induced and how the action and perception systems interact in the context of mental imagery is not well understood. Imagined speech production (“articulation imagery”), which induces the kinesthetic feeling of articulator movement and its auditory consequences, provides a new angle because of the concurrent involvement of motor and perceptual systems. On the basis of previous findings in mental imagery of speech, we argue for the following regarding the induction mechanisms of mental imagery and the interaction between motor and perceptual systems: (1) Two distinct top-down mechanisms,  Mental imagery can be characterized as a quasi-perceptual experience, induced in the absence of external stimulation. Neuroimaging studies have shown that common neural substrates mediate mental imagery and the corresponding perceptual processes, such as in visual  mechanism to Kosslyn's memory-attention-based account Kosslyn, : plannedMental imagery has been proposed to be essentially a memory retrieval process. That is, perceptual experience is simulated by reconstructing stored perceptual information in modality-specific cortices Kosslyn, . In partdirect simulation.Mental imagery is further hypothesized to be a predictive process , in which the dynamics of perceptual experience can be retrieved/calculated and reconstructed internally  account in that it requires a transformation between motor and somatosensory systems. Our question here, though, extends beyond this: can a motor simulation deliver perceptual consequences that extend to other sensory domains  as well? If so, internal simulation and estimation processes would serve as an additional path to induce modality-specific neural representations similar to the ones induced on the basis of memory retrieval. In the next section, we discuss this possibility in the framework of internal forward models and propose a sequential simulation and estimation account. We will use the interaction of motor, somatosensory, and auditory systems in speech production as an example to illustrate such internal cascaded processes, which can generalize to other sensory domains.The Perception and production systems are functionally connected: perceptual systems analyze the sensory input generated by self actions; the motor system is also regulated by perceptual feedback to perform updates on actions in the future. For example, when people talk, they move their articulators, feel the movement, and hear the self-produced speech that can be used to detect and correct any pronunciation errors. The temporal sequence of physical articulation, proprioception of the articulators, and auditory perception of one's own vocalization makes it possible—on the basis of co-occurrence and associative learning during development—to create internal connections among the neural processes that mediate motor action, somatosensory feedback, and auditory perception. After establishing the connections, motor commands can cycle internally through somatosensory regions and “reach” auditory regions. That is, the estimation in the somatosensory system can serve as a link between motor and auditory systems. Theoretically, such a cascaded estimation architecture has been hypothesized by Hesslow . Anatomiovert speech processing [Figure auditory prediction is the result of a coordinate transformation from the somatosensory to the auditory domain. This sequential estimation mechanism (motor plan → somatosensory estimation → auditory prediction/estimation) can derive detailed auditory predictions that are then compared with auditory feedback for self-monitoring and online control.On the basis of recent neurophysiological (MEG) studies, we proposed that a process of auditory inference after somatosensory estimation occurs during mental imagery of speech, we propose that the quasi-perceptual experience of articulator movement and the subsequent auditory percept are induced by the same sequential estimation mechanism. However, the “cancellation” deriving from somatosensory and auditory feedback, which is generated by the overt outputs during production, is absent in the imagery case  neural representations can be generated either by internal estimation or by memory retrieval, based on contextual variation and task demands.On the basis of sequential estimation account, particular neural activity patterns for the two sequential estimates are predicted to occur in a temporal order. Specifically, an auditory pattern should follow a somatosensory one during mental imagery of speech. Applying a novel multivariate technique constructed first in higher level associative areas and conveyed to a perceptual-sensory representation in lower areas. The observation of neural activity in the posterior superior temporal sulcus (pSTS) during silent speaking . Therefore, motor simulation to estimate perceptual consequences is only modulatory and not necessary for perceptual tasks.The major evidence supporting a modulatory role of motor simulation in perception  comes from lesion studies. For example, lesions in the frontal lobe only caused deficits in action production, whereas lesions in the parietal lobe caused deficits both during production and perception of movement  as follows: when patients prepare to articulate speech covertly or subvocally (either consciously or unconsciously), the internal motor simulation leads to perceptual estimation (intact efference copy). But the source monitoring process malfunctions (broken corollary discharge). Therefore, the internal prediction of a perceptual consequence, which has the same neural representation as an external perception, is erroneously interpreted as the result of external sources, resulting in an auditory hallucination.The comparison between internal estimation and external feedback provides information to fine-tune motor control. However, if the internal estimate from motor simulation malfunctions and generates imprecise perceptual predictions, an inaccurate or incorrect feedback control signal would be conveyed. Stuttering could be an example of such erroneous correction. We suggest, along the lines of similar theories . Stutterers showed speed and latency deficits when required to sequentially update articulator movement  can enhance speech fluency in people who stutter  accompanied with chronic and sometimes acute pain. We hypothesize that the apparent awareness and control of a lost limb occurs as follows: the missing somatosensory feedback is “replaced” by the results of internal estimation  are the focus of motor control research from an engineering perspective. Our atypical pairing of internal models as an additional source for mental imagery yields, in our view, some provocative new angles on mental imagery in both basic research and applied contexts.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.


My laboratory studies how chromosomes are organized in three dimensions. The three-dimensional organization of the genome is critical for regulating gene expression by bringing genes in close spatial proximity to distal regulatory elements such as enhancers. We have developed powerful molecular approaches, based on our Chromosome Conformation Capture technology, to determine the folding of genomes at unprecedented resolution (Kb) and scale (genome-wide).We have applied these methods to determine the spatial folding of 1% of the human genome (the ENCODE pilot regions) across a panel of cell lines. We discovered that chromosomes fold into extensive long-range interaction networks in which genes are interacting with distal gene regulatory elements. These results start to place genes and regulatory elements, that are often separated by large genomic distances, in three-dimensional context to reveal their functional relationships.Our analysis of chromosome folding also revealed that chromosomes are compartmentalized in a series of "Topological Association Domains" (TADs) that are hundreds of Kb in size. Loci located within a TAD mingle freely, but interact far less frequently with loci located outside their TAD. TADs appear involved in gene expression, as we found that genes located within the same TAD tend to be co-expressed, but the mechanism(s) by which these domains affect gene regulation is still unknown. TADs represent novel universal and genetically encoded building blocks of chromosomes.


Here we propose to take advantage of these resources to develop a tool, TARGETgene, for efficiently identifying mutation drivers, possible therapeutic targets, and drug candidates in cancer. The simple graphical user interface enables rapid, intuitive mapping and analysis at the systems level. Users can find, select, and explore identified target genes and compounds of interest , and validate predictions using user-defined benchmark genes  and curated cancer genes via TARGETgene. The high-level capabilities of TARGETgene are also demonstrated through two applications in this paper. The predictions in these two applications were then satisfactorily validated by several ways, including known cancer genes, results of RNAi screens, gene function annotations, and target genes of drugs that have been used or in clinical trial in cancer treatments. TARGETgene is freely available from the Biomedical Simulations Resource web site ( Intensive use of cytotoxic agents in multimodal therapies has improved five-year disease-free survival and even resulted in cure for some cancer patients. This success can be associated with severe toxicities and an increased occurrence of secondary cancers. The emergence of targeted therapies directed against dysregulated or mutated genes/proteins in malignant cells represents a paradigm shift in cancer therapy, with less reliance on drugs that kill normal cells as well as tumor cells. Examples include therapies against HER2 overexpressed breast cancers (such as Trastuzumab and Lapatinib), c-Kit-targeted therapy in BCR-ABL defective leukemias (Gleevec), and VEGF/VEGF-R-targeted compounds for inhibiting cancerous angiogenesis (such as Bevacizumab). While high throughput technologies such as microarray and next generation sequencing can now be used to identify hundreds or thousands of candidate genes that are differentially expressed or mutated in cancerous versus normal tissues, it is difficult to prioritize potential cancer therapeutic targets from such a large number of candidate genes.A systematic studying of the complex regulatory pathways is required to understand the mechanisms of oncognesis to discover mutation drivers or develop effective therapies. Several pathways have been found to be deregulated in cancer cells due to the over-expression or repression of some control elements Several concepts also have led to the development of network-based approaches to predict novel disease genes in molecular networks Broader use of such constructed molecular networks and network-based approaches would be promoted by the availability of tools that allow easy identification of potential therapeutic targets for specific cancers. This report thus introduces the software tool TARGETgene that utilizes a constructed gene network that integrates multiple genomic and proteomic data using the RVM-based model Seventeen heterogeneous genomic and proteomic data were integrated using the RVM-based ensemble model reported in Based on the constructed gene network, TARGETgene identifies potential therapeutic targets using one of two network-based metrics: 1) hub score or 2) seed gene association score (as detailed in the section 2 of ftp://ftp.ncbi.nih.gov/gene/). Through this panel, the user also can find, select identified target genes , and explore their functions, cited literature as well as enriched biological processes. In addition, TARGETgene enables users to validate their predictions using user-defined benchmark genes  and curated cancer genes via this panel. Finally, drug and their target information compiled from several public databases, such as DrugBank The graphical user interface of TARGETgene consists of four main working panels, including Input, Implementation, Gene, and Drug panels . The InpTo illustrate the use of TARGETgene, we have applied it to two examples: 1) identification of potential therapeutic targets from thousands of differentially expressed genes identified by exon array; 2) identification of driver mutated genes from sequencing and copy number data.in vitro and in vivo study to evaluate their importance these cancer types. Several of these also have been identified by RNAi screens (as detailed in the following section).In this example, TARGETgene was applied in turn to each of three cancer types: Her2-positive breast cancer, colon cancer, and Lung Adenocarcinoma. Human Exon datasets in the Affymetrix platform for the three cancer types were collected from the National Center for Biotechnology Information Gene Expression Omnibus (GEO) The resulting ranked genes from TARGETgene are also validated using gene functional annotations and several benchmark gene sets, including the set of curated cancer genes, the set of genes cited in cancer literature, and the set of target genes detected by RNAi screens. Receiver Operating Characteristic (ROC) Curves and AUC are used for these benchmark evaluations. In each evaluation, the benchmark gene sets are treated as positive instances while others genes are treated as negative instance.The curated cancer genes downloaded from the CancerGenes database High-throughput RNAi screens have recently been shown to be a promising tool to discover new targets for the treatment of several cancers Gorilla After mapping the information of drugs/compounds and their targets to the ranked gene lists from TARGETgene, the Drug Panel helps to identify compounds that either have been approved or are currently in clinical trials for the treatment of each of the three cancers. Other drugs and compounds identified by TARGETgene that have not as yet been used in clinical trials, have also shown anti-cancer effect and could thus be considered as potential novel drug for these cancers. Large numbers of gene mutations have been discovered from next generation sequencing Mutations of hub genes in molecular networks are capable of dyregulating the regular functions of many genes and their pathways, due to the ability of hub genes to directly or indirectly alter other components of the cell during their extensive interactions. Accordingly, mutated hub genes may be drivers of cancer progression. In this example, we applied TARGETgene to identify possible driver mutated genes from the approximately 500 mutated genes in the genome of Glioblastoma Multiforme (GBM) Based on the results in the two examples presented, most well studied cancer genes, including those that have shown clinical benefit  phenotypes without using any prior biological information. RNAi screens thus can be expected to be a powerful tool for identifying and validating novel targets in the drug discovery process In principle, RNAi screens could be combined with the gene network-based approach in TARGETgene to arrive at a more refined list of accurate cancer targets without the need for extensive validation of RNAi screens, and with lower false negative rates. The abundant biological information embed in the constructed gene network can provide biological interpretation for the novel targets through their connected genes. In addition, by taking advantage of the gene network-based approach that can identify potential targets using clinical data, one could provide clinical relevance to the novel targets detected by RNAi screens. The gene network-based screen in combination with RNAi screens could be persuasive and provide a complementary mechanism for the identification of therapeutic targets, and thus accelerate drug discovery process.While the primary purpose of TARGETgene is to identify potential therapeutic targets using integration of heterogeneous biological data, TARGETgene also lists existing drugs and other compounds that may have possible action on the identified targets, as illustrated in the examples presented. These results provide some direct confirmation of abilities of TARGETgene to identify potential drugs. However, these identified drugs may not be effective in the treatment of the indicated cancer for a number of reasons, including: 1). the drug binding affinities are target dependent; 2) the mechanisms of actions of some drugs are unclear; 3). most drugs act against multiple targets, of which some are up-regulated while others are down-regulated in cancers. Therefore, it is difficult to evaluate any possible therapeutic effect of the identified drugs in the predictions.The results presented in these two applications, however, suggest that TARGETgene could be a tool for initial screening of potential new drugs for further evaluation. Novel drugs whose target genes are highly ranked in TARGETgene's prediction could be considered as potential new drugs for these cancers. These drugs can then be further validated using preclinical testing, such as testing in cell lines or animal models. Most importantly, if targets of some FDA-approved drugs or compounds are highly ranked in the predictions, it is possible to reuse these drugs in the treatment of other cancers or diseases. The results for each cancer type also identify several naturally occurring compounds. Two examples are melatonin and vitamin D whose targets are highly ranked in the case of breast cancer .There is a vast and diverse amount of public genomic and proteomic resources in the life sciences that may aid in the understanding of disease mechanisms and in the drug discovery process. TARGETgene integrates these resources and provides a platform that enables people to efficiently identify mutation drivers, possible therapeutic targets, and drug candidates in cancer. TARGETgene can rapidly extract gene functional interactions from a precompiled database that is stored as a MATLAB MAT-file without the need to interrogate remote SQL databases. Millions of interactions of thousands of candidate genes can be extracted from the gene network within minutes. While TARGETgene is currently based on the gene network reported in One study successfully applied a single gene network to accurately predict tissue-specific phenotypic effects of gene perturbation in Caenorhabditis elegans File S1Enriched GO terms that appear densely at the top of TARGETgene's ranked gene lists for each of the three cancer types.(XLS)Click here for additional data file.Text S1Supporting information text.(DOC)Click here for additional data file.


Background. Native Americans disproportionately experience adverse childhood experiences (ACEs) as well as health disparities, including high rates of posttraumatic stress, depression, and substance abuse. Many ACEs have been linked to methylation changes in genes that regulate the stress response, suggesting that these molecular changes may underlie the risk for psychiatric disorders related to ACEs. Methods. We reviewed published studies to provide evidence that ACE-related methylation changes contribute to health disparities in Native Americans. This framework may be adapted to understand how ACEs may result in health disparities in other racial/ethnic groups. Findings. Here we provide evidence that links ACEs to methylation differences in genes that regulate the stress response. Psychiatric disorders are also associated with methylation differences in endocrine, immune, and neurotransmitter genes that serve to regulate the stress response and are linked to psychiatric symptoms and medical morbidity. We provide evidence linking ACEs to these epigenetic modifications, suggesting that ACEs contribute to the vulnerability for developing psychiatric disorders in Native Americans. Conclusion. Additional studies are needed to better understand how ACEs contribute to health and well-being. These studies may inform future interventions to address these serious risks and promote the health and well-being of Native Americans. Reservation-based Native Americans live in pervasively adverse social and physical environments that place them at increased risk of exposure to a myriad of stressors during childhood which impact their psychological and physical health over their lifetimes . About 1A neighborhood's safety and access to quality health care, economic opportunities, social connections, and social capital are all key determinants of the health of its residents over time –12. ReseUnique to Native Americans is the race-based stress associated with historical trauma , as wellNative Americans are disproportionally affected by trauma in childhood, including abuse, neglect, and exposure to intimate partner violence (IPV) –29. ApprAssaultive trauma in childhood is linked to the highest risk for PTSD, suggesting that this ACE is specifically linked to this high risk for psychiatric disorders . SpecifiACEs have also been linked to increased risk of depression onset –38. ThesReservation-based Native Americans die at higher rates than other Americans from tuberculosis (750% higher), alcoholism (524% higher), diabetes (293% higher), unintentional injuries (153% higher), homicide (103.3% higher), and suicide (66% higher)  . Not onlThe adverse childhood experiences (ACE) study suggests that certain adversities are major risk factors for morbidity and mortality . The stuIn some cases, genetic predisposition may explain some of the enduring effects of ACEs; however, the evidence for this link remains poorly understood. Genetic inheritance provides information encoded in DNA which is transcribed to various types of RNA molecules which likely shape the response of the individual to stressors such as ACEs. One important concept related to phenotypic variation is heritability, which estimates the extent of which genetic inheritance contributes to the phenotypic variance in a population . HeritabPolymorphisms in Native Americans have been linked to a greater vulnerability for alcohol abuse , as wellThe stress-response system provides the individual protection from acute stressors through an activation of interactive biological systems . One bioEvidence is accumulating that environmental influences early in development remain pervasive into adulthood, a relationship that is attributed to an interaction of gene function and environment. Both genetic and environmental factors are critical to developmental processes and even minor changes in either type of factor can result in trajectories of resilience or vulnerability ; howeverEpigenetics refers to changes in an individual's phenotype independent of genotype. These changes occur through mechanisms such as histone modification, methylation, acetylation, and noncoding ribonucleic acids which alter the accessibility of genes for transcription. The resulting transcription modifications and protein production result from factors such as environmental challenges including, but not limited to, ACEs , 69. An Preclinical models illustrate how ACEs result in epigenetic modifications in neurons, thereby increasing the risk for psychiatric symptoms. To illustrate this link, a study reports that the offspring of high-licking canine mothers exhibit reduced methylation of the glucocorticoid receptor gene  and endoF of the promoter region of the glucocorticoid receptor gene [The ACE most linked to epigenetic differences and vulnerability for health disparities is that of child abuse. To illustrate this link, in a hallmark study by Labonté in suicide completers, ACEs were linked to increased DNA methylation of the glucocorticoid receptor in the hippocampus, and this differential methylation was particularly linked to childhood abuse . This sttor gene . These stor gene , 80. ThuF of the promoter region of the glucocorticoid receptor gene in leukocytes [Epigenetic changes resulting from ACEs can also be observed in studies that use peripheral blood in living participants, which show that HPA-regulating genes are often impacted. A study by Klengel et al. linked ACEs to reduced methylation of the FKBP5 gene, an essential regulator of the stress response, as well as to changes in the function of the HPA axis under stress, and to reduced cognitive ability . Direct ukocytes . Similarukocytes . AlthougOther studies link ACEs to hypomethylation of inflammatory genes, suggesting that these experiences result in a greater inflammation later in life. In a recent study of children who were removed from their parents due to abuse or neglect, a reduction in methylation of NR3C1, an inflammatory regulation gene, as well as differential methylation of cancer related pathways was found in children with ACEs compared to controls . A studyAltered serotonergic neurotransmission is also postulated to result from ACEs and provides a mechanistic link to increased vulnerability for psychiatric disorders. The Iowa adoption study demonstrated a link between hypermethylation of the serotonin gene SLC64A to childhood sex abuse , and thiClinical studies are restricted to examining differential methylation in samples of peripheral fluids, but these studies do provide some key insights into how these molecular changes relate to PTSD, depression, and drug abuse risk. For instance, PTSD is associated with changes in the methylation of inflammatory  and endocrine genes FK506 binding protein-5 (FKBP5) . AnotherStudies in individuals with depression have also shown differential DNA methylation. Sabunciyan et al. carried out the first genome-wide DNA methylation scan in major depressive disorder patients. The study pinpointed 224 candidate regions, primarily involved in neuronal growth and development genes, which showed differential methylation; PRIMA1 showed the greatest differences . SpecifiIn addition to observing differential DNA methylation in PTSD and depression, many studies have observed methylation differences in those that suffer from drug abuse as compared to healthy controls. Increases in DNA methylation of the OPRM1 gene that codes for opioid receptors have been reported in individuals with chronic opioid use –102 and Reservation-based Native Americans disproportionately experience ACEs and health disparities, significantly impacting long-term physical and psychological health. In addition to these experiences, the persistence of stress associated with discrimination and historical trauma converges to add immeasurably to these challenges. Here we provide evidence to suggest that ACEs result in methylation differences in genes that regulate the stress response and that these changes may contribute to an increased vulnerability for developing psychiatric disorders, as depicted in


Children exposed to alcohol prenatally can experience significant deficits in cognitive and psychosocial functioning as well as alterations in brain structure and function related to alcohol’s teratogenic effects. These impairments are present both in children with fetal alcohol syndrome (FAS) and in children with heavy in utero alcohol exposure who do not have facial dysmorphology required for the FAS diagnosis. Neuropsychological and behavioral studies have revealed deficits in most cognitive domains measured, including overall intellectual functioning, attention/working memory, executive skills, speed of processing, and academic skills in children and adolescents across the range of fetal alcohol spectrum disorders (FASD). As with neuropsychological studies, brain-imaging studies have detected differences in brain structure related to alcohol exposure in multiple brain systems and abnormalities in the white matter that connects these brain regions. Several studies have found relationships between these morphological differences and cognitive function, suggesting some clinical significance to the structural brain abnormalities. Concentrations of neurotransmitter metabolites within the brains of prenatally exposed children also appear to be altered, and functional imaging studies have identified significant differences in brain activation related to working memory, learning, and inhibitory control in children and adolescents with FASD. In the last four decades, much has been learned about the significant and permanent changes caused by the damaging effects of alcohol on the developing fetus . Fetal alcohol syndrome (FAS), for example, is the leading known and preventable cause of intellectual disabilities . One rec1–14) for a thorough review of FAS diagnostic criteria. These children have been found to have differences in brain structure  is a non-invasive imaging technique that allows for in vivo measurement of brain structures. Individuals are exposed to a high-intensity magnet, and, using computational techniques, researchers can capitalize on physical changes specific to different tissue types, resulting in contrast between gray and white matter within the brain . Measures such as regional volume, cortical thickness, and other measures of shape can be analyzed and compared across groups. It is not currently possible to diagnose FASD based on MRI findings; all findings described below are based on group-average differences in which measures for individual subjects with FASD overlap considerably with individuals in unexposed control groups. Postmortem studies of severely affected children with heavy prenatal alcohol exposure have consistently shown smaller head sizes as well as smaller brains . NeuroimResearchers also have reported differences in the volume and thickness of the frontal lobes, which is consistent with neuropsychological studies that have found impairments in attention/working memory and executive functioning typically associated with frontal regions (Fuster 2003). An early study of frontal lobe volumes found them to be relatively spared when controlling for whole brain size . In contThe parietal lobes, which generally are associated with visuospatial functioning and attention (Fuster 2003), appear to be more affected by PEA than other cortical areas. The temporal lobes usually are associated with memory formations, auditory processing, and language comprehension (Fuster 2003). Temporal-lobe volume appeared to be spared relative to other brain structures when controlling for overall brain size in children with FASD , but thiAs noted previously, one study reported group differences in cortical thickness between individuals affected with FASD and control subjects see  Sowell b. This sOn a test of visuospatial functioning, the FASD group also performed more poorly than control subjects. Within the control group, negative correlations between cortical thickness and visuospatial skills were observed in bilateral parietal, left occipital, right dorsal and ventral frontal, and temporal cortices see . DecreasSubcortical structures have been associated historically with basic cognitive processes such as emotion, movement, arousal, and memory formation, but, through white-matter connections with cortical regions, they are part of higher-level networks related to more complex cognitive processes such as attention and executive functions  . ResearcThe cerebellum now is known to play a role in attention, executive functions, and other complex tasks as well as movement . O’Hare The superior edge of the anterior vermis showed the strongest negative correlation with verbal learning performance in the alcohol-exposed group see . This reIn general, there is a higher incidence of midline brain anomalies in FASD compared with other brain regions , and midOther researchers found structural abnormalities in the location and size of the CCs of children, adolescents, and young adults with FASD and attempted to identify cognitive correlates of callosal dysmorphology by evaluating verbal learning abilities and visuospatial skills in these subjects see  Sowell a. In theSubjects with FASD performed more poorly than control subjects on a test of verbal learning. The researchers reported relationships between verbal learning and the location of the CC in the FASD group a. The moAnother study  of childThese results are consistent with previous studies that have documented abnormalities in callosal morphology in children with FASD. For example, N-acetyl aspartate (NAA), a neuronal or axonal marker, and creatine (Cr), a marker of metabolic activity, can be measured either in absolute concentration values or as ratios relative to each other.Compared with other neuroimaging techniques, MR spectroscopy (MRS) has been relatively underutilized in studying FASD, and the few existing studies have had somewhat inconclusive results. Using MRI technology, MRS capitalizes on different physical properties of chemicals in the body when they are exposed to a high magnetic field, and in brain research, measures of brain chemicals that are important to specific processes can be detected . For exaOne study that examined metabolite ratios in multiple brain structures of children with FASD and control subjects found lower NAA/Cho and NAA/Cr ratios in parietal and frontal cortices, frontal white matter, the CC, thalamus, and cerebellar dentate nucleus of children with FASD . The invOther researchers  detectedA more recent MRS analysis observed significantly lower concentrations of choline in frontal/parietal white matter regions lateral to the medial corpus callosum b. As froDiffusion Tensor Imaging (DTI) also uses MRI technology and allows measurement of the diffusion of water molecules within brain tissues, specifically white matter, in order to produce images of white matter tracts and measures of white matter integrity . FractioResearch with young adults with FAS has found lower FA values in the genu, splenium , and istResearchers using a combination of DTI and T1-weighted MRI to evaluate white matter integrity in people with FASD attempted to relate these findings to neurocognitive deficits, in particular visuomotor integration and reading skills a. FA refAlthough correlations between reading scores and FA were not significant in any region tested, it appeared that lower FA values in the lateral splenium and parietal white matter bilaterally were associated with worse performance on a test of visuomotor integration, but only within the FASD group a. This iWhen reviewing sMRI studies of children with FASD, it appears that many brain systems are affected by the teratogenic consequences of prenatal alcohol exposure see , and 4. Although the focus of many sMRI studies has been on morphometric differences between people with and without FASD, a handful of studies have successfully integrated what is known about function from neuropsychological studies with brain structure. Differences in specific brain structures do not necessarily translate directly to knowledge about differences in cognitive function given the complex and interrelated anatomical systems within the brain. For example, it is possible that relationships between cognitive function and frontal lobe brain structure actually are mediated by striatal dysmorphology, and relationships between brain and behavior are thus likely indirect in some instances. Most studies have attempted to investigate brain–behavior relationships through correlational methods on a structure-by-structure basis, and although these findings are important, interpretations are limited by the nature of correlational research. Future studies may benefit from investigating connectivity , a noninvasive method that allows researchers to localize functional brain activation with an accuracy of millimeters and a temporal resolution of seconds. This technique measures the dynamic regulation of blood flow in the brain  related to neural activity. Blood releases oxygen to firing neurons at a greater rate than to inactive neurons. The MR signal of oxygenated hemoglobin is different from the MR signal of deoxygenated hemoglobin. These transient changes in MR signal, which accompany hemodynamic events, can be observed when nuclear magnetic resonance images of the brain are acquired in rapid succession. The brain regions showing an increase in the intensity of the MR signal correspond to focal areas of activation . Only fiThe first fMRI study that examined brain function in adults and children affected by FASD evaluated brain activation during a spatial working memory (WM) “N-back” taskfMRI results indicated that both children and adults with FASD showed greater activity than control subjects in the inferior and middle frontal cortex . This sufMRI results strongly supported this hypothesis. Despite equivalent behavioral performance, study participants with FASD showed increased functional activation relative to control subjects in the prefrontal cortex and decreased activation in the caudate nucleus during trials that required response inhibition . This fiGiven that neuropsychological studies have shown verbal learning deficits, and because brain structures involved in verbal learning show structural abnormalities in children with heavy prenatal alcohol exposure, research has examined fMRI activation patterns corresponding to verbal paired associate learning (PAL) in children with heavy prenatal alcohol exposure see  Sowell . fMRI reThis pattern of increased frontal and decreased medial temporal activation may be consistent with the interpretation that people with FASD rely more heavily on frontal memory systems to encode and retrieve verbal information, perhaps compensating for dysfunctional medial temporal memory systems, which have been reported in previous studies to show structural abnormalities such as increased gray matter volume . In addiAn fMRI study of verbal working memory (WM) function in children and adolescents affected by FASD see  O’Hare  also wasIn addition, individuals with FASD showed increased dorsal frontal activation relative to control subjects see , which iIn the most recent study to report abnormal functional brain activation patterns in individuals with FASD, investigators used fMRI to study brain activation during performance on a visual N-back WM task a. One ofn = 2) the FAS/pFAS group had significantly lower activation than control subjects in various areas that are part of the neurocognitive network involved in WM, such as the right posterior parietal lobe, right dorso-lateral prefrontal cortex (DLPFC), and right middle frontal regions. Progressing across the four groups from FAS/pFAS to control subjects, activation levels increased in these brain regions. It appeared that higher activation in the DLPFC and middle frontal gyrus in both hemispheres was related to lower numbers of errors on the 2-back task. These results suggest that children across the full spectrum of FASD show significant WM deficits, which are correlated with abnormalities in activation in brain regions known to be involved in WM.fMRI results indicated that, when considering 2-back task activation patterns  and to understand how these functional brain abnormalities relate to cognitive performance and behavioral outcomes.Neuropsychological studies have demonstrated that nearly every cognitive domain evaluated is affected by prenatal exposure to alcohol and that these deficits are present in children with FAS and PEA. sMRI studies of children with FASD show that many brain systems are affected by the teratogenic effects of PEA, including structures previously believed to be relatively spared. Several studies have attempted to evaluate relationships between brain structure and cognitive functioning and found that these relationships were altered as a function of PEA. Consistent with the neuropsychological and structural MRI literature, fMRI findings demonstrate significant differences in brain activation between FASD and control groups on a variety of cognitive tasks. However, all these studies have limitations, and it is important that future investigators replicate the results presented above in larger samples of participants in order to increase power. Most studies reviewed utilized case–control methodology, which has both advantages and disadvantages. Although this methodology allows the study of relatively rare conditions such as FASD and may be the only practical method, exposure and other history are obtained by interview, which is subject to recall bias. It is important to note that in some cases validation of exposure may be incomplete or impossible to obtain. An important goal for future studies will be to continue integrating our understanding of structure and function, and it also will be essential to continue using multimodal methods that combine neuropsychological tools and structural and functional neuroimaging.


Exploration of chronobiological systems emerges as a growing research field within bioinformatics focusing on various applications in medicine, agriculture, and material sciences. From a systems biological perspective, the question arises whether biological control systems for regulation of oscillatory signals and their technical counterparts utilise similar mechanisms. If so, modelling approaches and parameterisation adopted from building blocks can help to identify general components for frequency control in circadian clocks along with gaining insight into mechanisms of clock synchronisation to external stimuli like the daily rhythm of sunlight and darkness. Phase-locked loops could be an interesting candidate in this context. Both, biology and engineering, can benefit from a unified view resulting from systems modularisation. In a first experimental study, we analyse a model of coupled repressilators. We demonstrate its ability to synchronise clock signals in a monofrequential manner. Several oscillators initially deviate in phase difference and frequency with respect to explicit reaction and diffusion rates. Accordingly, the duration of the synchronisation process depends on dedicated reaction and diffusion parameters whose settings still lack to be sufficiently captured analytically. In both spheres, biological and technical systems, oscillatory signals play a major role in order to trigger and control time-dependent processes. Core oscillators are the simplest devices for generation of continuously running clock signals. To this end, signal processing units consisting of at least one feedback loop can suffice . So, it modularisation inspired by engineering.The situation becomes more complicated if several of those core oscillators start to interact. Resulting biological systems are commonly driven to achieve a synchronous behaviour towards an evolutionary advantage. Correspondingly, clock synchronisation in technical systems is frequently inspired by the need to follow a global time. Interestingly, the formalisation of clock synchronisation processes is quite distant from each other. While, in distributed computer systems, stepwise algorithmic approaches  predomi“structure follows function.” Although there is a plethora of different strategies and implementations to achieve a certain complex network function, the pool of sufficient network candidates can be divided into compositions of a low number of elementary functional units called modules. This term is not new in systems biology when considering recurrent motifs conserved in metabolic, cell signalling, and gene regulatory networks. We extend the notion of modules in terms of information processing. In this context, a module is able to fulfill an elementary computational task. Here, the spatiotemporal course of substrate concentrations along with molecular and compartmental structures acts as data carrier. Beyond logic and arithmetic functions carried out by the module's steady-state behaviour, simple buffer and delay elements contribute to a collection of biochemical modules, each of which comprising few molecular species and a maximum number of reactions within the same magnitude. When combining those modules towards reaction networks capable of a more complex functionality, we permit so-called shared molecular species among distinct modules. This way of module coupling enables compact network topologies in accordance with evident observations from in-vivo studies. Moreover, there is no need for further separate interface specifications. In most cases, the behaviour of a module can be captured by chemical counterparts of transfer functions in conjunction with characteristic curves which exhibit an established practice in engineering. Utilisation of transfer functions for modules significantly reduces the number of distinct parameters to be considered by keeping the relevant characteristics of the entire network.Our concept is based on the assumption that Within a case study, we exemplify oscillatory signal synchronisation by a biological system composed of bidirectionally coupled repressilators. To this end, we model the entire gene regulatory networks using reaction-diffusion kinetics. Afterwards, we conduct two comprehensive simulation studies. The first one discloses the time to synchronisation subject to initial phase shift between the elementary repressilators. Its balanced diffusion rate acts as coupling strength. It appears that synchronisation of initially antiphasic signals is most time consuming for weak coupling while it has a negligible effect for strong coupling. A second simulation study investigates the synchronisation behaviour with respect to different initial frequencies of the single repressilators.phase-locked loops. Corresponding circuits comprise three essential modules:Coupled repressilators represent a prototypic example of core oscillators embedded into a complex reaction-diffusion network constituting a circadian clock as an entire system able to be adjusted by external stimuli. From a systems point of view, circadian clocks form biochemical frequency control circuits whose functionality resembles technical counterparts utilising so-called a core oscillator whose frequency has been controlled to adapt to an external stimulus,a signal comparator (phase detector) responsible for determining the deviation between the signal produced by the core oscillator on the one hand and the external stimulus on the other one, a biochemical low-pass filter.Finally, we illustrate a corresponding scheme by the identification of feasible network candidates (modules) composing a pure biochemical frequency control by a phase-locked loop.Beyond frequency control circuits, there are some examples on how biological and technical systems utilise equal or similar mechanisms to achieve a certain function. In the context of fascinating approaches within biomimicry , those rsynchronous to each other if and only if they meet three conditions.We define different temporally oscillating signals to be undamped to avoid signal weakening.The oscillatory signal must run Asymptotical or total harmonisation of the oscillatory signals meaning that after a finite amount of time called tsync (time to synchronisation), both temporal signal courses converge within an arbitrarily small ε-neighbourhood.tsync has to be monofrequential to ensure chronoscopy (constant progression of time measure).The resulting oscillatory signal after Note that synchronicity is stricter than obtaining entrainment. While an adaptation of oscillatory signal frequency and phase suffices for entrainment, synchronicity additionally requires harmonisation of the entire signal shape (waveform) over time.forcing, by resetting, or by specific selective perturbations affecting the oscillating signal. Without any external influences, core oscillators resume their individual free-running oscillatory behaviour, mostly by loosing their synchronicity. The central prerequisite of a core oscillator to be capable of synchronisation to others is its ability to vary its oscillation frequency within a specified range . This vaexternal and internal .Bidirectionally coupled repressilators synchronise their oscillatory signals by forcing. Arbitrary initial phase shifts become compensated while adaptation to different initial frequencies spans a synchronisation window. Coupled repressilators can be seen as a part of a biological control system based on the concept of phase-locked loops. Further research has been directed to finalise the entire frequency control system by integration of additional components for signal comparison and damping, demonstrated by low-pass filters biologically implemented as specific signal transduction cascades. The simulations described in this paper were carried out using Copasi , statist


Since the advent of techniques for the expression of recombinant peptide antigens, the availability of human vaccines for parasitic diseases has been ‘imminent’. Yet vaccines based on recombinant proteins are still largely aspirations, not realities. It is now apparent that vaccine development needs additional knowledge about host protective immune response(s), antigen characteristics, and the delivery required to induce those responses. The most successful immune protection against parasites has been generated by infection and treatment, the induction of protective immunity by truncating the course of an infection with drug treatment. Here, we consider the characteristics of an effective, protective anti-parasite vaccine and propose a conceptual framework to aid parasite vaccine development using malaria and schistosomiasis as examples. Theileria) of livestock and companion animals are based on inoculation with attenuated or drug treated parasites. In humans, the most widely used ‘vaccination’ for a parasitic infection is the practice of leishmanization Leishmania sore in a location typically covered by clothing. The resulting, self-limiting lesion provides protection against subsequent infections that might otherwise form a disfiguring ulceration on an exposed area. However, no vaccines against parasitic infections are licensed for human use. This is at least in part attributable to the antigenic complexity of parasites, arising from multiple life cycle stages, immune evasion strategies, and use of intermediate and reservoir hosts. Unfortunately, obtaining adequate numbers of parasites, attenuated or otherwise, of consistent and acceptable quality to use in vaccinations is highly challenging, as demonstrated by recent studies of the Plasmodium falciparum attenuated sporozoite vaccine (PfSPZ vaccine) in humans P. falciparum sporozoites can be used in controlled human malaria infections will greatly facilitate this research in the future Exposure to pathogens allows vertebrate hosts to mount pathogen-specific acquired immune responses that sometimes protect against subsequent infection, forming the basis of vaccinology P. falciparum. Recently, an I&T trial for malaria was performed by exposing volunteers who were receiving chloroquine prophylaxis to P. falciparum sporozoites. The chemoprophylaxis with sporozoites (CPS) protocol succeeded in inducing sterile immunity in all immunized participants and was maintained in four of six participants for >2 years Schistosoma haematobium or Schistosoma mansoni can induce partially protective immunity against subsequent infections An alternative to infection with attenuated parasites is the infection and treatment (I&T) approach where immunity is induced by the release of antigens from parasitic infections that are treated or naturally die in the host . One of Ancylostoma secreted protein (ASP-2) was discontinued when vaccination induced urticarial reactions in people with pre-existing IgE responses to ASP-2 Another outcome of I&T is that individuals from areas where they are likely to have been exposed to malaria or schistosome antigens early in life tend to have a lower risk of developing severe pathological consequences such as cerebral malaria or hepatosplenic schistosomiasis, respectively. Protective mechanisms against pathology are poorly understood but are hypothesized to involve induction of different regulatory or memory immune responses. In addition to modulation of pathology in subsequent infections, I&T effects on host immune responses are also instructive with respect to development of defined antigen vaccines. Most vaccine recipients in endemic areas are likely to have had some exposure to the parasite, leading to reactions during immunization that may differ from those of parasite naïve vaccine trial participants. For example, the Phase I clinical trial evaluating the vaccine against human hookworm using Similar to inoculation with attenuated parasites, I&T has limitations that may preclude it from being a feasible public health tool; for some parasite species, it may not be possible to generate sufficient quantities of infectious stage parasites to vaccinate the millions of people exposed to these infections. Nevertheless, I&T approaches provide key answers to some fundamental intellectual and practical questions for successful vaccine development. By concentrating on the principles of classical vaccination, we describe how I&T protocols have overcome some of the challenges of using recombinant protein immunizations.Parasites causing the greatest morbidity and disease typically induce a more or less protective immunity very slowly. Reasons for this include poor immunogenicity of individual antigens, poor protective immunity of major antigens, antigenic variation (protozoa), antigen polymorphism, immune evasion, immunomodulation of effector responses, and/or the requirement for a threshold amount of antigen which is released more easily upon treatment than from natural parasite death Echinococcus granulosus onchospheres provoke a high degree of protective immunity, which is the basis of a highly effective vaccine in lambs  or through gene conversion  However, protozoan parasites can vary the antigens seen by host immune systems through mechanisms including transcriptional and epigenetic control  that prevent development of memory T cell effector functions of cytotoxic lymphocytes E. granulosus that interferes with antigen presentation and cell activation Not all immune responses result in parasite killing or resistance to re-infection. Indeed, some parasite evasive mechanisms may divert the immune system to respond against ‘decoy’ antigens. In Plasmodium suggests that blood stage parasites induce immunosuppression. This was confirmed in humans as blood stages of P. falciparum suppress T lymphocyte reactivity to malarial and unrelated antigens Plasmodium parasites include utilization of T cell mimotopes to inhibit T cell activation, induce anergy, or shift the T cell phenotype P. falciparum I&T trial in humans resulted in equivalent or better protection than that produced by irradiated sporozoite immunizations Plasmodium from a single mosquito bite and treated based on clinical symptoms or diagnosis, at which time the blood stage parasites may be suppressing pre-erythrocytic immunity. In successful Plasmodium CPS, the drug was administered prophylactically, including during inoculation of sporozoites from 15 mosquitoes Plasmodium has achieved this by minimizing immune exposure to immunosuppressive blood stage parasites. How or even if this is achieved in people who develop natural resistance to malaria is unclear.Amelioration of autoimmunity in rodents by + T cells towards a T helper 2 (Th2) phenotype , and immunosuppress worm-specific + T cell subsets  also means that effector cytokines [interferon (IFN)-γ, IL-4, and IL-21) are required to maintain a balanced acquired immune response Experimental and natural helminth infections are associated with immunoregulatory responses that polarize CD4Plasmodium infection for sufficient amounts of antigens from different circulating strains of the parasite to stimulate development of partially protective immunity Plasmodium and schistosome infection prevalence rates peak and decline at an earlier age. This pattern, first described in Plasmodium in 1949 Quantitative studies in human schistosomiasis show that immunosuppression alone does not explain the effects of age on infection intensity observed in schistosome endemic areas P. falciparum sporozoites or irradiated cercariae suggest that high antigen doses are required to stimulate protective responses + regulatory responses in humans and promote the development of tolerance Immunization with irradiated TheileriaAlmost all current vaccines work through induction of serum or mucosal antibodies that block infection or interfere with invasion and proliferation. Although antibody half-life is ∼30 days, effective vaccine-induced protection persists beyond the time when antibodies should have disappeared. Possible reasons for this include boosting from natural infection, antigen retention in peripheral lymph nodes by follicular dendritic cells, presence of sequestered memory B cells in bone marrow sanctuaries, or maintenance of antigen-specific memory B cells by nonspecific B cell activators or idiotypic networks. The importance of natural boosting has been demonstrated in animal I&T protocols for protozoa infections  lack of vaccine candidates providing reproducible protection in experimental models and humans; (ii) limitations in current understanding of the nature, development, and maintenance of protective immune responses, particularly in people already exposed to schistosomes; and (iii) how to induce protective immune responses better and faster than what occurs naturally while at the same time avoiding pathology or tolerance The conclusions in this paper are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.


Aberrant CpG island promoter DNA hypermethylation is frequently observed in cancer and is believed to contribute to tumor progression by silencing the expression of tumor suppressor genes. Previously, we observed that promoter hypermethylation in breast cancer reflects cell lineage rather than tumor progression and occurs at genes that are already repressed in a lineage-specific manner. To investigate the generality of our observation we analyzed the methylation profiles of 1,154 cancers from 7 different tissue types.We find that 1,009 genes are prone to hypermethylation in these 7 types of cancer. Nearly half of these genes varied in their susceptibility to hypermethylation between different cancer types. We show that the expression status of hypermethylation prone genes in the originator tissue determines their propensity to become hypermethylated in cancer; specifically, genes that are normally repressed in a tissue are prone to hypermethylation in cancers derived from that tissue. We also show that the promoter regions of hypermethylation-prone genes are depleted of repetitive elements and that DNA sequence around the same promoters is evolutionarily conserved. We propose that these two characteristics reflect tissue-specific gene promoter architecture regulating the expression of these hypermethylation prone genes in normal tissues.As aberrantly hypermethylated genes are already repressed in pre-cancerous tissue, we suggest that their hypermethylation does not directly contribute to cancer development via silencing. Instead aberrant hypermethylation reflects developmental history and the perturbation of epigenetic mechanisms maintaining these repressed promoters in a hypomethylated state in normal cells. Aberrant DNA hypermethylation of CpG island (CGI) promoters (promoter hypermethylation) occurs in many cancers. This epigenetic reprogramming is associated with the absence of transcription and can occur at a number of known tumor suppressor genes, suggesting that it contributes to tumor progression by silencing the expression of affected genes . AlthougDnmt1, in mice can lead to reactivation of the Xi later in development suggesting that in this case CGI hypermethylation acts as a stabilizing factor that maintains silencing [Hypermethylation also plays a role in the regulation of some genes during normal development, particularly at imprinted loci and at CGI promoters on the inactive X-chromosome (Xi) in female mammalian cells ,12. Duriilencing . Where tilencing .APC frequently occurs in gastric cancer, but at a promoter that is not utilized in normal gastric tissue [RUNX3, whose tumor suppressor gene status is largely based on the fact that it is frequently methylated in gastric cancer, is never expressed in the gastrointestinal epithelial cells that give rise to these tumors [We have recently shown that genes whose promoters are hypermethylated in breast cancer cell lines and tumors are already repressed in the putative lineage of origin and that when methylation is removed in cancer cell lines, either pharmacologically or genetically, most hypermethylated genes do not become re-activated . This imc tissue  and thate tumors .Here, we explore the generality of our observations in breast cancer by analyzing data derived from 1,154 tumors arising in 7 different human tissues. We show that variability in promoter CGI hypermethylation patterns between tumors is explained by variability in gene expression patterns between normal tissues and it is genes that are repressed in the pre-cancerous tissue that become preferentially hypermethylated in tumors. Our study represents the first comprehensive analysis of promoter CGI hypermethylation in different human cancers and we propose that the hypermethylation of repressed CGI promoters is a common feature of most cancers.P < 2 × 10-16, Fisher's exact test), demonstrating the reproducibility of our methodology. In total, 1,009 genes were prone to hypermethylation by this analysis in at least one type of cancer, including a number reported to be frequently hypermethylated in cancer  [We have previously shown that cell lineage determines promoter hypermethylation patterns in breast cancer . To examX genes) -31. NoneP < 2 × 10-16). The next three components of the data accounted for 10.4% of the variance in the data and clearly separated out the samples into the seven different tissue types . The hypermethylation-prone and -resistant gene sets were associated with different Gene Ontology (GO) terms Figure . In partWe used a method based on information theory to directly quantify the degree of tissue-specificity in a given gene's expression pattern across nine normal tissues that were profiled by high-throughput mRNA sequencing  -35, withBased on genes hypermethylated in multiple cancer cell lines, one study has suggested that the transcriptional start sites (TSSs) of genes prone to hypermethylation are depleted of repetitive elements . We inveMany of the bioinformatic techniques used to discover functional elements in the human genome use comparisons of the genomes of multiple species to infer their presence through evolutionary conservation . Therefocis regulatory role at some tissue specific genes [Recently, long intergenic non-coding RNAs (lincRNAs) have been proposed to play a ic genes . Their pic genes . Thus, hAlthough repeat occupancy and conservation differ between hypermethylation-prone and -resistant genes, these factors displayed overlapping distributions for the two gene sets . We have previously shown a similar preference for genes specifically repressed in luminal epithelial cells to be hypermethylated in breast tumors [We tested if normal expression patterns determined hypermethylation susceptibility by considering VM genes with differential susceptibility in individual cancer types. VM genes that were frequently hypermethylated in breast tumors (67 genes) were repressed in the cells of origin of most breast tumors, luminal epithelial cells , as compt tumors . TogetheThe aberrant hypermethylation of CGI promoters is of interest because it correlates with gene silencing and can occur at tumor suppressor genes . Here, wRUNX3 is frequently hypermethylated in gastric cancers but is never expressed in normal gastric epithelia [de novo aberrant mechanism.During normal development, DNA methylation accumulates at loci that are already repressed and may facilitate stable transcriptional repression rather than directly cause silencing . We havepithelia . TherefoBRCA1 only occurs in 12% of ovarian cancers and 2% of breast cancers. We find that APC is hypermethylated more frequently  but it has multiple TSSs and a promoter that is repressed in normal gastric tissue has been shown to be the site of hypermethylation in gastric cancers [APC hypermethylation depends on whether it occurs at the major promoter in these tissues. It is unclear whether the hypermethylation of these potential driver genes occurs as a by-product of the process that results in the hypermethylation of repressed genes or by an alternative mechanism . It is known that MLH1 is frequently hypermethylated in colorectal tumors that possess a CIMP phenotype [Our analyses show that this model applies to the majority of hypermethylated genes found in tumors, but it has been proposed that within each tumor a few 'driver' genes are directly repressed by hypermethylation . Under t cancers . Therefohenotype . Howeverhenotype ,8,58.Our results confirm a previous observation that the promoters of genes prone to aberrant hypermethylation in cancer are depleted of repetitive elements . HoweverDNMT3B promotes tumorigenesis in a mouse model of colorectal cancer and is associated with the hypermethylation of specific genes [RUNX3 was the earliest CGI promoter to show significant change [We find that the aberrant hypermethylation of repressed genes occurs in all cancer types analyzed implying that a common mechanism might be responsible for promoter hypermethylation in all cancers. Although the exact mechanism remains unknown, our results mean that proposed mechanisms must account for the specificity of hypermethylation for repressed genes Figure . For exaic genes . Higher ic genes ,70. One t change ,69, suggBRAF mutations and CIMP-low mutations associated with KRAS mutations [Here we have shown that differences exist in the aberrant hypermethylation profiles of cancers arising in different tissue contexts. However, our results also make it clear that there is heterogeneity in the methylation profiles within particular types of cancer Figure . It is uutations ,71. Inteutations . Our ownutations . Taken tRecently, hydroxymethylated cytosine (hmC) has been re-discovered as a DNA modification present at significant levels in mammalian cells . The IllIn summary, our results argue that the bulk of aberrant promoter hypermethylation in cancer occurs predominantly at genes that are repressed in pre-cancerous tissue and therefore does not directly contribute to tumor progression by silencing tumor suppressor genes. This epigenetic alteration is common to all the cancer types we have analyzed implying that a common mechanism is responsible for promoter hypermethylation at repressed genes in all cancers. Future research in this field should, therefore, focus on confirming whether aberrant hypermethylation does directly suppress rare driver genes and if the mechanism responsible for driver gene suppression is the same as that acting at repressed genes. Finally, we would suggest that researchers must exercise caution in assigning a tumor suppressor status to a gene based on its propensity to become hypermethylated in cancer.All statistical analyses were performed using the R statistical software (version 2.12.1) . AdditioGene expression and methylation data used in this study were taken from previously published studies. The sources of the data are indicated in Table In order to apply a consistent annotation to the data used in this study, all data were re-annotated to Ensembl 54 gene IDs (NCBI36). CpG probes from the Illumina Infinium arrays were mapped to the closest Ensembl gene based on TSS location using custom Perl and R scripts. CpGs that ambiguously mapped to more than one gene ID were removed from the analysis. CGI locations were taken from those biologically defined in a recent study . SimilarmethI, and unmethylated, unmethI, allele probes: meth I/(meth I+ unmethI) ). We have previously shown that these are a reliable estimate of the level of methylation at a locus [P-value from the Infinium arrays (threshold 0.01). Methylation data originating from other techniques (MDB-seq or whole-genome bisulfite sequencing) were either downloaded as processed data provided by the authors [coverageBED tool to quantify the number of reads at each CGI [For data originating from Infinium methylation arrays, beta values were used as a measure of the methylation level at a given CpG probe . Previously, we have shown that probes with beta values <0.3 represent unmethylated areas of the genome  and we tTo ensure that the method of gene selection did not bias our results, we also carried out analyses in which parameters were varied . CGIs were assigned to genes if their TSS was present in the CGI. Using this methodology, we were unable to define a set of hypermethylation resistant genes.To define genes which were hypermethylated in colorectal tumors from the Xu seq data , we usedet al. whole- genome bisulfite sequencing data [We defined genes prone to and resistant to hypermethylation from the Berman ing data  as genesTo process RNA-seq data, raw sequence data for nine human tissues  were dowaffy package and an updated annotation [affy package.Processed Illumina gene expression data were downloaded from GEO and summarized to individual Ensembl IDs by taking the mean value of all probes mapped to that gene . Reportenotation . DetectiTo define CGI genes that were differentially expressed between normal colon and normal liver, we assayed for differential expression in processed expression microarray data using t-tests and Benjamini-Hochberg correction for multiple testing  of 5%). We then refined these lists to only those that had CGI promoters and had probes on the Infinium array within 200bp of their TSS that were unmethylated in normal colon  before comparing methylation levels in colorectal tumors between these groups.To analyze functional terms, Ensembl Biomart was used to map gene identifiers to GO biological process terms (Ensembl 54). Enrichment of specific terms in each gene list was then assessed using Fisher's exact test as compared to all genes present on the Infinium array. Terms that were associated with less than 10 genes on the Infinium arrays were excluded from the analysis. Data were presented as change in the percent of genes in each set as compared to the control.The specificity of a gene's expression pattern in normal tissues was measured using a method based on information theory . A low sGenes reported as mutated in breast cancer were defined using the Catalogue of Somatic Mutations in Cancer  databasTo define the density of repetitive elements around TSSs, repetitive element positions were downloaded from the Repeat Masker track of the University of California, Santa Cruz (UCSC) genome browser (hg18) . Custom coverageBed tool from the BEDtools suite of bioinformatics tools [We defined the level of conservation around gene promoters using two different measurements. The first was based upon the measurement of base substitutions between 17 vertebrate species . To defics tools  to calcuWe used a recent survey of lincRNAs in the human genome to define genes that had a neighboring lincRNA . The neaAML: acute myeloid leukemia; bp: base pair; CGI: CpG island; CM: consistently methylated; COSMIC: Catalogue of Somatic Mutations in Cancer; DNMT: DNA methyltransferases; ES cell: embryonic stem cell; GEO: Gene Expression Omnibus; GO: gene ontology; hmC: hydroxymethylated cytosine; lincRNA: long intergenic non-coding RNA; LINE: long interspersed element; LTR: long terminal repeat; MDB: methyl-binding domain; mC: methylated cytosine; PCA: principal component analysis; RPKM: reads per kb per million mapped reads; seq: high-throughput sequencing; SINE: short interspersed element; SRA: sequence read archive; TCGA: The Cancer Genome Atlas; TSS: transcription start site; VM: variably methylated.The authors declare that they have no competing interests.DS and RRK performed the research and analyzed data. DS, CEN, JMD, AHS, DJH, BHR and RRM helped design the research and review the manuscript. DS and RRM wrote the paper. All authors read and approved the final version of the manuscript for publication.1,009 hypermethylation prone genes. Excel file containing details of the 1,009 hypermethylation prone genes in the 7 cancer types.Click here for fileSupplementary data. PDF file containing four supplementary figures, one table and their legends.Click here for file446 variably methylated genes. Excel file containing details of the 446 variably hypermethylated genes along with their susceptibility in the different cancer types.Click here for file220 consistently methylated genes. Excel file containing details of the 220 consistently hypermethylated genes in the 7 different types of cancer.Click here for file


Numerous screening methods have been developed to detect hazardous and harmful drinking in a range of health settings. Recent research has focused on developing briefer screening tools to maximize implementation in busy practice settings, particularly emergency departments (EDs) and primary care. However the relative utility of these tools is not fully understood. Further, there is a need to identify the utility of universal screening, in which all patients approaching primary care are screened, compared with targeted screening, which includes only patients with certain “red flag” conditions or presentations. The Screening and Intervention Program for Sensible Drinking (SIPS) program compared the relative utility of different screening tools  and approaches  in primary care. In addition, the utility of the Paddington Alcohol Test (PAT), a targeted screening tool, was compared with SASQ and FAST in EDs. Compared with the Alcohol Use Disorders Identification Test (AUDIT), the FAST had a higher sensitivity than the SASQ in primary care. Although targeted screening in primary care is a more efficient screening method, it misses a large proportion of patients who could benefit from brief interventions. The SASQ performed better in EDs than either the FAST or PAT. These results have important implications for the choice of screening tools in different settings.


Decades of research have deepened our understanding of how the brain forms memories and uses them to build our mental past and future. But how does it determine whether an evoked memory refers to the present and can be acted upon? The study of patients who confuse reality, as evident from confabulation and disorientation, has opened ways to explore this vital capacity. Results indicate that the brain recurs to a phylogenetically old faculty of the orbitofrontal cortex – extinction – and structures of the reward system to keep thought and behavior in phase with reality. Since the early studies of patients who lost the ability to acquire new memories after damage to the hippocampal area  or a previous run; it requires reality filtering Figure B. DespitWhen healthy subjects performed repeated runs of this task, they had activation of the posterior medial orbitofrontal cortex, area 13 Figures , which cThe task structurally resembles well-known source memory tasks requiring attribution of stimuli to previous task stages, such as, the exclusion condition of the process dissociation procedure Jacoby, , in thatOur reality-filtering task, as easy as it may be for healthy subjects, proved an insurmountable challenge for reality-confusing patients, even at much longer intervals between the runs (30–60 min) than in healthy subjects (1 min). While healthy subjects and non-confabulating amnesics maintained performance over repeated runs, reality-confusing patients had a sharp increase of false positive responses: they believed increasingly more often that they had already seen pictures within the ongoing run, which in reality appeared for the first time within the run , as depicted in Figure Reality filtering is not limited to visual information: we observed similar orbitofrontal activation in functional imaging with visually presented verbal or non-verbal visual material . Thus, reality filtering, similar to reward processing, is under dopaminergic modulation. Of note, a hyper-dopaminergic state has also been suspected to underlie schizophrenia, another disorder characterized by reality confusion  Intrusions in memory tests (simple provoked confabulations). These dissociate from all other forms of confabulation and are independent of reality confusion  behaviorally spontaneous confabulation, as defined above – the topic of this review. It can be conceived as a specific subform of momentary confabulations, namely, the form caused by reality confusion, as evident from inappropriate acts in accordance with the confabulations and disorientation. (4) Fantastic confabulations, which defy any sense of plausibility. These are rare and occur in severe confusion, dementia, or psychosis  Confabulations emanate from a combination of amnesia with frontal executive failures. The hypothesis stems from the observation that executive functions may recover in parallel with the cessation of confabulations  LeDoux,  extinctiThe author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Momordica charantia, commonly known as bitter melon has been traditionally used in Ayurvedic and Chinese medicine to treat diabetes and its complications. The aim of this study was to develop bitter melon-containing recipes and test their palatability and acceptability in healthy individuals for future clinical studies.Although beneficial to health, dietary phytonutrients are bitter, acid and/or astringent in taste and therefore reduce consumer choice and acceptance during food selection. 2) analyses.A cross-sectional sensory evaluation of bitter melon-containing ethnic recipes was conducted among 50 healthy individuals. The primary endpoints assessed in this analysis were current consumption information and future intentions to consume bitter melon, before and after provision of attribute- and health-specific information. A convenience sample of 50, self-reported non-diabetic adults were recruited from the University of Hawaii. Sensory evaluations were compared using two-way ANOVA, while differences in stage of change (SOC) before and after receiving health information were analyzed by Chi-square (χOur studies indicate that tomato-based recipes were acceptable to most of the participants and readily acceptable, as compared with recipes containing spices such as curry powder. Health information did not have a significant effect on willingness to consume bitter melon, but positively affected the classification of SOC.This study suggests that incorporating bitter foods in commonly consumed food dishes can mask bitter taste of bitter melon. Furthermore, providing positive health information can elicit a change in the intent to consume bitter melon-containing dishes despite mixed palatability results. Prevalence of obesity and associated metabolic syndromes, such as type 2 diabetes mellitus (T2DM) and cardiovascular diseases (CVD) are escalating worldwide. In the United States, an estimated 20.8 million people have T2DM with an estimated two to four-fold increased risk of CVD . TreatmeMomordica charantia (commonly known as bitter melon) has been widely used throughout the world for centuries, to manage diabetes and its complications  in one box only. Participants were requested to refrain from discussing their opinions with others and compliance was monitored by study coordinators. During data analysis, numerical scores were generated by authors based on the above mentioned hedonic scale. Participants also listed factors that would improve acceptability and increase consumption frequency for each dish. Sensory evaluations took approximately 10 minutes to complete. Upon completion, participants answered questions related to their current consumption, knowledge, and attitudes, as well as to their intentions to consume bitter melon dishes as a meal in the future, over a period of two weeks.precontemplation, defined as a stage when an individual has no intentions of changing behavior within six months and remains resistant to change, contemplation, defines an individual who has limited awareness of potential problems or risks and contemplates making changes, preparation, defines an individual who is planning a change through specific action within the next month, action, defines an individual who conducts the specific behavior, maintenance, classified as the point in which the behavior has been occurring for at least six months and termination, the point at which a previous behavior is completely eliminated and is no longer a possibility when temptation arises. In the last part of the survey, participants received "attribute-specific" and "consequence-specific" health information in writing as described by Wansink, et al.,[The second part of the survey form was designed to determine previous bitter melon consumption as well as future intentions to consume bitter melon-containing dishes after receiving bitter melon-associated health information. Initially, individuals were placed in one of the Stages of Change (SOC) applicable to dietary studies based on their answers to questions provided in an algorithm format as shown in Table , et al., and adapKnowledge about bitter melon traits or characteristics of action was provided by attribute-specific information. For example, "research with animals and humans have shown that bitter melon can lower blood sugar levels" provides information about bitter melon attribute or quality. The consequences of such attributes were provided separately. An example of consequence-specific information included in the survey was "eating bitter melon may help lower your risk for type 2 diabetes or heart disease". Surveys were randomized such that half of the participants read the attribute-specific information first while the other half read the consequence-specific information first.After receiving each set of information, participants were asked to again list how many times they would be willing to consume each dish as a part of meal over a two-week period. They were asked to check only one box for each sample that included zero times, one time, two times, three times, four times or five times per week. This was designed to assess changes in willingness to consume each dish given varying levels of knowledge about bitter melon. Additionally, individuals initially classified in the pre-action stages  were asked after each set of information to check one of three boxes to indicate that they were either "not thinking," "thinking," or "planning" on consuming bitter melon in the future. This was included to determine if providing health information affected stage movement for these participants despite palatability of bitter melon. SOC classification was reevaluated for individuals in the pre-action stages after each set of health information to determine if the information had an effect on stage movement.Post hoc pair-wise multiple comparisons were evaluated using the Bonferroni test after ANOVA. Similarly, a two-way ANOVA model with Bonferroni post test was used to determine if participant consumption intentions for each sample were affected by participants receiving "attribute-specific," "consequence-specific," or both types of information about bitter melon compared with those receiving no information. Chi-square (χ2) analyses were conducted to determine if there were statistically significant differences in participants' stage distribution before and after receiving health information as well as the order of providing information. Results were considered significant at p < 0·05.Survey data were analyzed using SAS  and GraphPad Prism, Prism 5 for Windows, version 5.01. Frequency distributions among the hedonic scale categories were determined for each of the three sample attributes  as well as the overall sample rating. Statistically significant differences for each attribute were assessed using a two-way ANOVA. + 12.5 (SD).All 50 participants completed all or part of the sensory evaluation and survey. One participant survey was excluded from the analysis due to unreliable demographic responses. Table Overall acceptability of specific attributes varied among the four food samples. Mean hedonic scores for attributes and overall liking are presented in Table Figure Table Of the 49 participants, 69% n = 34) indicated they had tried bitter melon before, while 31% (n = 15) had never eaten bitter melon. However, only 13 of these 34 participants (38%) indicated that they previously consumed BM at least once every two months. Consumption at least once every 2 months was used as the means to classify an individual in the "action stages" according to the SOC algorithm shown in Table 4 indicatThe number of individuals in the precontemplation stage significantly decreased by more than half from 53% to 25% and 26% after individuals received "attribute- and consequence-specific" information, respectively , respectively . These vAlthough spices in the curry were unable to mask the bitter taste, its low total energy content makes it an attractive therapeutic alternative in Asian countries such as India wherein curry preparation may be culturally preferred and more acceptable. The other three dishes consisted of several other ingredients such as tomatoes, garlic and basil that masked their bitterness. Some traditional Asian recipes remove the bitter flavor from the fruit by adding salt, which extract the bitter juices and is then discarded. Alternatively, a combination of salt and vinegar can also be used to mask the bitter flavor. Besides salt, natural and artificial sweeteners as well as fat or cooking techniques such as pickling, caramelizing, or braising can be effective in reducing bitter taste ,31,32. ISince this was a palatability testing, the variation in calories of different food dishes is not expected to affect the taste. Similarly, bland starches such as rice and pasta are not expected to significantly affect palatability of bitter melon-containing food dishes. Although participants commented on "more rice" or "less rice" to increase palatability, there was no preference for one starch over the other. Moreover, these bitter melon-containing dishes could have been consumed without the starches. However, that may have resulted in a false positive or negative palatability score as these food dishes are generally consumed with starches.Besides taste, health information also may influence the gradual acceptance and consumption of functional foods with bitter taste . In our Most of the information provided in this study focused on attributes and consequences of bitter melon consumption that would benefit individuals at higher risk for diabetes or CVD. Although diabetes is considered as an old age disease, the prevalence of T2DM is increasing in children and young adults, especially in the Pacific region . TherefoMajor strength of our study was identifying ingredients from western diet that are able to effectively mask the bitterness of functional food such as bitter melon. Although the order of samples was not randomized, this study limitation may not necessarily affect the taste of subsequent samples since water and saltine crackers were provided between dishes to cleanse their palates. However, the first food sample was randomized so that each dish would not be influenced by the perceived bitterness of the other dishes. This is exemplified by the fact that the participants at the soup station did not all dislike the chili (the last to be sampled by these participants). Randomizing the first dish was an effort to eliminate any potential cumulative effect on taste and therefore may be perceived as "structured randomization".Overall, our study demonstrated that incorporating bitter melon in commonly consumed dishes increased acceptability among a mixed-ethnic and variable age group. However, previous exposure and knowledge of its health benefits had no effect on consumption intentions. Although our study was limited in the number of bitter melon-containing dishes that were tested and the age of the population, acceptability of foods with masked bitter taste merits future studies with large number of bitter melon-containing food dishes.Bitter melon is routinely consumed as a part of many Asian diets and should therefore be expected to have minimal side effects. Well-designed clinical trials are also warranted to verify the dosage, safety, and pharmacokinetics of bitter melon. Results obtained from such studies will be significant since the information can help to develop nutritional regimens that can be incorporated into a daily diet. Developing such dietary alternatives to increase palatability and ultimately consumption of bitter tasting functional foods is expected to positively impact diabetes outcomes. Such intervention strategies would also be valuable to populations where conventional therapy is inaccessible and/or unaffordable, thereby improving diabetes care and management.Overall, sour flavor of tomato based recipes were able to mask the bitterness of bitter melon and these food dishes were considered acceptable by most participants. Taste rather than health information was a major determinant of willingness to consume bitter melon in future. The potential benefits associated with many bitter fruits and vegetables, such as bitter melon, may warrant the development of recipes to use in nutrition interventions aimed at increasing consumption of these bitter foods. Future studies are warranted to test the palatability of additional bitter melon containing food choices among participants of varying ages.CVD: cardiovascular diseases; SOC: stages of changes; T2DM: type 2 diabetes mellitus.The authors declare that they have no competing interests.SSL conducted the experiments and designed the questionnaire and participated in developing ideas and writing the manuscript. VRN provided critical insights in conceptualization and development of this project and in the process of revising draft versions of the manuscript. DAD and ACS provided critical insights in the designing of questionnaire and reviewing the manuscript. JTF and his team performed the statistical analysis. PVN, developed ideas, designed experiments and questionnaire, secured funding and wrote the manuscript. All authors have read and approved the final manuscript.


Rabies is rapidly progressive, an almost universally fatal encephalitis caused by lyssaviruses, most notably rabies virus. Canine rabies is estimated to kill about 55,000 persons annually in Africa and Asia aloneThe article in this issue by Gilbert and colleagues3If complete or relative resistance to rabies were to occur in humans, it should be detected sporadically, such as in the recent well-characterized reports of rabies survivors in Texas and California.6If complete or relative resistance to rabies were to exist in distinct populations, that would then likely require prolonged selection in a rabies-endemic region of genetically isolated human populations. Where would one look? One might postulate remote areas with small indigenous populations such as the Arctic (where fox rabies virus is endemic) or the Amazon, Africa, or Asia where bat or carnivore rabies viruses, and probably other bat-associated lyssaviruses, are endemic. The article in this issue by Gilbert brings us much closer to this possibility. It is notable that subjects in the Peruvian Amazon had antibodies to either the rabies virus nucleoprotein (as did survivors in the United States) or neutralizing antibodies to the membrane glycoprotein, but rarely to both. It is also notable that all seropositive subjects were adults, suggesting a fitness advantage to rabies resistance in a region where child survival is poor and 80% of subjects suffered depredation by vampire bats.Where does this leave the rest of humankind who are highly susceptible to rabies? Rabies has been vaccine-preventable for over a century; the economic and geographic maldistributions of rabies vaccines and immunoglobulins are indefensible. Technology transfer will hopefully improve this with time. Gilbert has identified an opportunity for novel therapeutics, discoverable through whole genome sequencing. Careful, respectful genetic study of these genetically unique populations may provide information on which pathways in human biochemistry and physiology promote resistance to human rabies. Equally important, knowing that there is a continuum of disease, even for infectious diseases like rabies, should push us harder to try for cures when confronted by so-called untreatable infectious diseases or intoxications. Modern therapeutics can move us along a continuum toward greater survival, even when specific cures or antidotes remain undiscovered.