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Find video protocols related to scientific articles indexed in Pubmed.
DNA copy number analysis of fresh and formalin-fixed specimens by shallow whole-genome sequencing with identification and exclusion of problematic regions in the genome assembly.
Genome Res.
PUBLISHED: 09-18-2014
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Detection of DNA copy number aberrations by shallow whole-genome sequencing (WGS) faces many challenges, including lack of completion and errors in the human reference genome, repetitive sequences, polymorphisms, variable sample quality, and biases in the sequencing procedures. Formalin-fixed paraffin-embedded (FFPE) archival material, the analysis of which is important for studies of cancer, presents particular analytical difficulties due to degradation of the DNA and frequent lack of matched reference samples. We present a robust, cost-effective WGS method for DNA copy number analysis that addresses these challenges more successfully than currently available procedures. In practice, very useful profiles can be obtained with ?0.1× genome coverage. We improve on previous methods by first implementing a combined correction for sequence mappability and GC content, and second, by applying this procedure to sequence data from the 1000 Genomes Project in order to develop a blacklist of problematic genome regions. A small subset of these blacklisted regions was previously identified by ENCODE, but the vast majority are novel unappreciated problematic regions. Our procedures are implemented in a pipeline called QDNAseq. We have analyzed over 1000 samples, most of which were obtained from the fixed tissue archives of more than 25 institutions. We demonstrate that for most samples our sequencing and analysis procedures yield genome profiles with noise levels near the statistical limit imposed by read counting. The described procedures also provide better correction of artifacts introduced by low DNA quality than prior approaches and better copy number data than high-resolution microarrays at a substantially lower cost.
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Interacting chemokine signals regulate dendritic cells in acute brain injury.
PLoS ONE
PUBLISHED: 08-25-2014
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Brain trauma is known to activate inflammatory cells via various chemokine signals although their interactions remain to be characterized. Mice deficient in Ccl3, Ccr2 or Cxcl10 were compared with wildtype mice after controlled cortical impact injury. Expression of Ccl3 in wildtypes was rapidly upregulated in resident, regularly spaced reactive microglia. Ccl3-deficiency enhanced endothelial expression of platelet selectin and invasion of peripheral inflammatory cells. Appearance of Ccr2 transcripts, encoding the Ccl2 receptor, reflected invasion of lysozyme 2-expressing phagocytes and classical antigen-presenting dendritic cells expressing major histocompatibility complex class II. Ccr2 also directed clustered plasmacytoid dendritic cells positive for the T-cell attracting chemokine Cxcl10. A reduction in Ccr2 and dendritic cells was found in injured wildtype cortex after cyclophosphamide treatment resembling effects of Ccr2-deficiency. The findings demonstrate the feasibility to control inflammation in the injured brain by regulating chemokine-dependent pathways.
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Altered expression of myelin-associated inhibitors and their receptors after traumatic brain injury in the mouse.
Restor. Neurol. Neurosci.
PUBLISHED: 08-01-2014
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When central nervous system axons are injured, regeneration is partly inhibited by myelin-associated inhibitors (MAIs). Following traumatic brain injury (TBI) in the rat, pharmacological neutralisation of the MAIs Nogo-A and myelin-associated glycoprotein (MAG) resulted in improved functional outcome. In contrast, genetic or pharmacological neutralization of the MAI receptors Nogo-66 receptor 1 (NgR1) or paired-immunoglobulin like receptor-B (PirB) showed an unaltered or impaired outcome following TBI in mice. The aim of the present study was thus to evaluate the MAI expression levels following TBI in mice.
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Two distinct routes to oral cancer differing in genome instability and risk for cervical node metastasis.
Clin. Cancer Res.
PUBLISHED: 11-08-2011
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Problems in management of oral cancers or precancers include identification of patients at risk for metastasis, tumor recurrence, and second primary tumors or risk for progression of precancers (dysplasia) to cancer. Thus, the objective of this study was to clarify the role of genomic aberrations in oral cancer progression and metastasis.
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Effects of everyday life events on glucose, insulin, and glucagon dynamics in continuous subcutaneous insulin infusion-treated type 1 diabetes: collection of clinical data for glucose modeling.
Diabetes Technol. Ther.
PUBLISHED: 10-24-2011
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In the development of glucose control algorithms, mathematical models of glucose metabolism are useful for conducting simulation studies and making real-time predictions upon which control calculations can be based. To obtain type 1 diabetes (T1D) data for the modeling of glucose metabolism, we designed and conducted a clinical study.
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Subtype and pathway specific responses to anticancer compounds in breast cancer.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 10-14-2011
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Breast cancers are comprised of molecularly distinct subtypes that may respond differently to pathway-targeted therapies now under development. Collections of breast cancer cell lines mirror many of the molecular subtypes and pathways found in tumors, suggesting that treatment of cell lines with candidate therapeutic compounds can guide identification of associations between molecular subtypes, pathways, and drug response. In a test of 77 therapeutic compounds, nearly all drugs showed differential responses across these cell lines, and approximately one third showed subtype-, pathway-, and/or genomic aberration-specific responses. These observations suggest mechanisms of response and resistance and may inform efforts to develop molecular assays that predict clinical response.
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Parent-specific copy number in paired tumor-normal studies using circular binary segmentation.
Bioinformatics
PUBLISHED: 06-11-2011
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High-throughput techniques facilitate the simultaneous measurement of DNA copy number at hundreds of thousands of sites on a genome. Older techniques allow measurement only of total copy number, the sum of the copy number contributions from the two parental chromosomes. Newer single nucleotide polymorphism (SNP) techniques can in addition enable quantifying parent-specific copy number (PSCN). The raw data from such experiments are two-dimensional, but are unphased. Consequently, inference based on them necessitates development of new analytic methods.
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Copy number variation in patients with disorders of sex development due to 46,XY gonadal dysgenesis.
PLoS ONE
PUBLISHED: 02-14-2011
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Disorders of sex development (DSD), ranging in severity from mild genital abnormalities to complete sex reversal, represent a major concern for patients and their families. DSD are often due to disruption of the genetic programs that regulate gonad development. Although some genes have been identified in these developmental pathways, the causative mutations have not been identified in more than 50% 46,XY DSD cases. We used the Affymetrix Genome-Wide Human SNP Array 6.0 to analyse copy number variation in 23 individuals with unexplained 46,XY DSD due to gonadal dysgenesis (GD). Here we describe three discrete changes in copy number that are the likely cause of the GD. Firstly, we identified a large duplication on the X chromosome that included DAX1 (NR0B1). Secondly, we identified a rearrangement that appears to affect a novel gonad-specific regulatory region in a known testis gene, SOX9. Surprisingly this patient lacked any signs of campomelic dysplasia, suggesting that the deletion affected expression of SOX9 only in the gonad. Functional analysis of potential SRY binding sites within this deleted region identified five putative enhancers, suggesting that sequences additional to the known SRY-binding TES enhancer influence human testis-specific SOX9 expression. Thirdly, we identified a small deletion immediately downstream of GATA4, supporting a role for GATA4 in gonad development in humans. These CNV analyses give new insights into the pathways involved in human gonad development and dysfunction, and suggest that rearrangements of non-coding sequences disturbing gene regulation may account for significant proportion of DSD cases.
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ACNE: a summarization method to estimate allele-specific copy numbers for Affymetrix SNP arrays.
Bioinformatics
PUBLISHED: 06-06-2010
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Current algorithms for estimating DNA copy numbers (CNs) borrow concepts from gene expression analysis methods. However, single nucleotide polymorphism (SNP) arrays have special characteristics that, if taken into account, can improve the overall performance. For example, cross hybridization between alleles occurs in SNP probe pairs. In addition, most of the current CN methods are focused on total CNs, while it has been shown that allele-specific CNs are of paramount importance for some studies. Therefore, we have developed a summarization method that estimates high-quality allele-specific CNs.
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TumorBoost: normalization of allele-specific tumor copy numbers from a single pair of tumor-normal genotyping microarrays.
BMC Bioinformatics
PUBLISHED: 05-12-2010
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High-throughput genotyping microarrays assess both total DNA copy number and allelic composition, which makes them a tool of choice for copy number studies in cancer, including total copy number and loss of heterozygosity (LOH) analyses. Even after state of the art preprocessing methods, allelic signal estimates from genotyping arrays still suffer from systematic effects that make them difficult to use effectively for such downstream analyses.
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Appearance of Cxcl10-expressing cell clusters is common for traumatic brain injury and neurodegenerative disorders.
Eur. J. Neurosci.
PUBLISHED: 04-09-2010
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Traumatic brain injury (TBI) in the mouse results in the rapid appearance of scattered clusters of cells expressing the chemokine Cxcl10 in cortical and subcortical areas. To extend the observation of this unique pattern, we used neuropathological mouse models using quantitative reverse transcriptase-polymerase chain reaction, gene array analysis, in-situ hybridization and flow cytometry. As for TBI, cell clusters of 150-200 mum expressing Cxcl10 characterize the cerebral cortex of mice carrying a transgene encoding the Swedish mutation of amyloid precursor protein, a model of amyloid Alzheimer pathology. The same pattern was found in experimental autoimmune encephalomyelitis in mice modelling multiple sclerosis. In contrast, mice carrying a SOD1(G93A) mutant mimicking amyotrophic lateral sclerosis pathology lacked such cell clusters in the cerebral cortex, whereas clusters appeared in the brainstem and spinal cord. Mice homozygous for a null mutation of the Cxcl10 gene did not show detectable levels of Cxcl10 transcript after TBI, confirming the quantitative reverse transcriptase-polymerase chain reaction and in-situ hybridization signals. Moreover, unbiased microarray expression analysis showed that Cxcl10 was among 112 transcripts in the neocortex upregulated at least threefold in both TBI and ageing TgSwe mice, many of them involved in inflammation. The identity of the Cxcl10(+) cells remains unclear but flow cytometry showed increased numbers of activated microglia/macrophages as well as myeloid dendritic cells in the TBI and experimental autoimmune encephalomyelitis models. It is concluded that the Cxcl10(+) cells appear in the inflamed central nervous system and may represent a novel population of cells that it may be possible to target pharmacologically in a broad range of neurodegenerative conditions.
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Identification of a CpG island methylator phenotype that defines a distinct subgroup of glioma.
Cancer Cell
PUBLISHED: 02-18-2010
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We have profiled promoter DNA methylation alterations in 272 glioblastoma tumors in the context of The Cancer Genome Atlas (TCGA). We found that a distinct subset of samples displays concerted hypermethylation at a large number of loci, indicating the existence of a glioma-CpG island methylator phenotype (G-CIMP). We validated G-CIMP in a set of non-TCGA glioblastomas and low-grade gliomas. G-CIMP tumors belong to the proneural subgroup, are more prevalent among lower-grade gliomas, display distinct copy-number alterations, and are tightly associated with IDH1 somatic mutations. Patients with G-CIMP tumors are younger at the time of diagnosis and experience significantly improved outcome. These findings identify G-CIMP as a distinct subset of human gliomas on molecular and clinical grounds.
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Identification of SOX3 as an XX male sex reversal gene in mice and humans.
J. Clin. Invest.
PUBLISHED: 02-08-2010
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Sex in mammals is genetically determined and is defined at the cellular level by sex chromosome complement (XY males and XX females). The Y chromosome-linked gene sex-determining region Y (SRY) is believed to be the master initiator of male sex determination in almost all eutherian and metatherian mammals, functioning to upregulate expression of its direct target gene Sry-related HMG box-containing gene 9 (SOX9). Data suggest that SRY evolved from SOX3, although there is no direct functional evidence to support this hypothesis. Indeed, loss-of-function mutations in SOX3 do not affect sex determination in mice or humans. To further investigate Sox3 function in vivo, we generated transgenic mice overexpressing Sox3. Here, we report that in one of these transgenic lines, Sox3 was ectopically expressed in the bipotential gonad and that this led to frequent complete XX male sex reversal. Further analysis indicated that Sox3 induced testis differentiation in this particular line of mice by upregulating expression of Sox9 via a similar mechanism to Sry. Importantly, we also identified genomic rearrangements within the SOX3 regulatory region in three patients with XX male sex reversal. Together, these data suggest that SOX3 and SRY are functionally interchangeable in sex determination and support the notion that SRY evolved from SOX3 via a regulatory mutation that led to its de novo expression in the early gonad.
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A single-array preprocessing method for estimating full-resolution raw copy numbers from all Affymetrix genotyping arrays including GenomeWideSNP 5 & 6.
Bioinformatics
PUBLISHED: 06-17-2009
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High-resolution copy-number (CN) analysis has in recent years gained much attention, not only for the purpose of identifying CN aberrations associated with a certain phenotype, but also for identifying CN polymorphisms. In order for such studies to be successful and cost effective, the statistical methods have to be optimized. We propose a single-array preprocessing method for estimating full-resolution total CNs. It is applicable to all Affymetrix genotyping arrays, including the recent ones that also contain non-polymorphic probes. A reference signal is only needed at the last step when calculating relative CNs.
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A single-sample method for normalizing and combining full-resolution copy numbers from multiple platforms, labs and analysis methods.
Bioinformatics
PUBLISHED: 02-04-2009
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The rapid expansion of whole-genome copy number (CN) studies brings a demand for increased precision and resolution of CN estimates. Recent studies have obtained CN estimates from more than one platform for the same set of samples, and it is natural to want to combine the different estimates in order to meet this demand. Estimates from different platforms show different degrees of attenuation of the true CN changes. Similar differences can be observed in CNs from the same platform run in different labs, or in the same lab, with different analytical methods. This is the reason why it is not straightforward to combine CN estimates from different sources (platforms, labs and analysis methods).
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CalMaTe: a method and software to improve allele-specific copy number of SNP arrays for downstream segmentation.
Bioinformatics
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CalMaTe calibrates preprocessed allele-specific copy number estimates (ASCNs) from DNA microarrays by controlling for single-nucleotide polymorphism-specific allelic crosstalk. The resulting ASCNs are on average more accurate, which increases the power of segmentation methods for detecting changes between copy number states in tumor studies including copy neutral loss of heterozygosity. CalMaTe applies to any ASCNs regardless of preprocessing method and microarray technology, e.g. Affymetrix and Illumina.
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Querying genomic databases: refining the connectivity map.
Stat Appl Genet Mol Biol
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The advent of high-throughput biotechnologies, which can efficiently measure gene expression on a global basis, has led to the creation and population of correspondingly rich databases and compendia. Such repositories have the potential to add enormous scientific value beyond that provided by individual studies which, due largely to cost considerations, are typified by small sample sizes. Accordingly, substantial effort has been invested in devising analysis schemes for utilizing gene-expression repositories. Here, we focus on one such scheme, the Connectivity Map (cmap), that was developed with the express purpose of identifying drugs with putative efficacy against a given disease, where the disease in question is characterized by a (differential) gene-expression signature. Initial claims surrounding cmap intimated that such tools might lead to new, previously unanticipated applications of existing drugs. However, further application suggests that its primary utility is in connecting a disease condition whose biology is largely unknown to a drug whose mechanisms of action are well understood, making cmap a tool for enhancing biological knowledge.The success of the Connectivity Map is belied by its simplicity. The aforementioned signature serves as an unordered query which is applied to a customized database of (differential) gene-expression experiments designed to elicit response to a wide range of drugs, across of spectrum of concentrations, durations, and cell lines. Such application is effected by computing a per experiment score that measures "closeness" between the signature and the experiment. Top-scoring experiments, and the attendant drug(s), are then deemed relevant to the disease underlying the query. Inference supporting such elicitations is pursued via re-sampling. In this paper, we revisit two key aspects of the Connectivity Map implementation. Firstly, we develop new approaches to measuring closeness for the common scenario wherein the query constitutes an ordered list. These involve using metrics proposed for analyzing partially ranked data, these being of interest in their own right and not widely used. Secondly, we advance an alternate inferential approach based on generating empirical null distributions that exploit the scope, and capture dependencies, embodied by the database. Using these refinements we undertake a comprehensive re-evaluation of Connectivity Map findings that, in general terms, reveal that accommodating ordered queries is less critical than the mode of inference.
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Influence of hypodermic needle dimensions on subcutaneous injection delivery--a pig study of injection deposition evaluated by CT scanning, histology, and backflow.
Skin Res Technol
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Thinner and shorter needles for subcutaneous administration are continuously developed. Previous studies have shown that a thinner needle causes fewer occurrences of painful needle insertions and that a shorter needle decreases the occurrence of painful intramuscular injections. However, little is known about local drug delivery in relation to needle length and thickness. This study aimed to compare deposition depth and backflow from three hypodermic needles of 3 mm 34G (0.19 mm), 5 mm 32G (0.23 mm), and 8 mm 30G (0.30 mm) in length and thickness.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.