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'name' => 'Saliva as a Blood Alternative for Genome-Wide DNA Methylation Profiling by Methylated DNA Immunoprecipitation (MeDIP) Sequencing',
'authors' => 'Staunstrup N.H. et al.',
'description' => '<p><b></b>Background: Interrogation of DNA methylation profiles hold promise for improved diagnostics, as well as the delineation of the aetiology for common human diseases. However, as the primary tissue of the disease is often inaccessible without complicated and inconvenient interventions, there is an increasing interest in peripheral surrogate tissues. Whereas most work has been conducted on blood, saliva is now becoming recognized as an interesting alternative due to the simple and non-invasive manner of collection allowing for self-sampling. Results: In this study we have evaluated if saliva samples are suitable for DNA methylation studies using methylated DNA immunoprecipitation coupled to next-generation sequencing (MeDIP-seq). This was done by comparing the DNA methylation profile in saliva against the benchmark profile of peripheral blood from three individuals. We show that the output, quality, and depth of paired-end 50 bp sequencing reads are comparable between saliva and peripheral blood and, moreover, that the distribution of reads along genomic regions are similar and follow canonical methylation patterns. Conclusion: In summary, we show that high-quality MeDIP-seq data can be generated using saliva, thus supporting the future use of saliva in the generation of DNA methylation information at annotated genes, non-RefSeq genes, and repetitive elements relevant to human disease.</p>',
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'name' => 'Emerging Role of One-Carbon Metabolism and DNA Methylation Enrichment on δ-Containing GABAA Receptor Expression in the Cerebellum of Subjects with Alcohol Use Disorders (AUD',
'authors' => 'Gatta E. et al.',
'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
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'authors' => 'Bak S.T. et al.',
'description' => '<p>We here characterize the usability of archival formalin-fixed paraffin-embedded (FFPE) brain tissue as a resource for genetic and DNA methylation analyses with potential relevance for brain-manifested diseases. We analyzed FFPE samples from The Brain Collection, Aarhus University Hospital Risskov, Denmark (AUBC), constituting 9479 formalin-fixated brains making it one of the largest collections worldwide. DNA extracted from brain FFPE tissue blocks was interrogated for quality and usability in genetic and DNA methylation analyses by different molecular techniques. Overall, we found that DNA quality was inversely correlated with storage time and DNA quality was insufficient for Illumina methylation arrays; data from methylated DNA immunoprecipitation, clonal bisulfite sequencing, and pyrosequencing of BDNF and ST6GALNAC1 suggested that the original methylation pattern is indeed preserved. Proof-of-principle experiments predicting sex based on the methylation status of the X-inactivated SLC9A7 gene, or genotype differences of the Y and X chromosomes, showed consistency between predicted and actual sex for a subset of FFPE samples. In conclusion, even though DNA from FFPE samples is of low quality and technically challenging, it is likely that a subset of samples can provide reliable data given that the methodology used is designed for small DNA fragments. We propose that simple PCR-based quality control experiments at the genetic and DNA methylation level, carried out at the beginning of any given project, can be used to enrich for the best-performing FFPE samples. The apparent preservation of genetic and DNA methylation patterns in archival FFPE samples may bring along new perspectives for the identification of genetic and epigenetic changes associated with brain-manifested diseases.</p>',
'date' => '2016-12-19',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/27995571',
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'authors' => 'Nicklas H. Staunstrup et al.',
'description' => '<p><strong>Background</strong> In utero and early-life experienced environmental exposures are suggested to play an important role in many multifactorial diseases potentially mediated through lasting effects on the epigenome. As the epigenome in addition remains modifiable throughout life, identifying specific disease-relevant biomarkers may prove challenging. This has led to an increased interest in epigenome-wide association studies using dried blood spots (DBS) routinely collected in perinatal screening programs. Such programs are in place in numerous countries around the world producing large and unique biobanks. However, availability of this biological material is highly limited as each DBS is made only from a few droplets of blood and storage conditions may be suboptimal for epigenetic studies. Furthermore, as relevant markers may reside outside gene bodies, epigenome-wide interrogation is needed.</p>
<p><strong>Results</strong> Here we demonstrate, as a proof of principle, that genome-wide interrogation of the methylome based on methylated DNA immunoprecipitation coupled with next-generation sequencing (MeDIP-seq) is feasible using a single 3.2 mm DBS punch (60 ng DNA) from filter cards archived for up to 16 years. The enrichment profile, sequence quality and distribution of reads across genetic regions were comparable between samples archived 16 years, 4 years and a freshly prepared control sample.</p>
<p><strong>Conclusions</strong> In summary, we show that high-quality MeDIP-seq data is achievable from neonatal screening filter cards stored at room temperature, thereby providing information on annotated as well as on non-RefSeq genes and repetitive elements. Moreover, the quantity of DNA from one DBS punch proved sufficient allowing for multiple epigenome studies using one single DBS.</p>',
'date' => '2016-07-26',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/27462375',
'doi' => '10.1186/s13148-016-0242-1',
'modified' => '2016-08-03 10:40:55',
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'name' => 'Epigenetic Modifications with DZNep, NaBu and SAHA in Luminal and Mesenchymal-like Breast Cancer Subtype Cells',
'authors' => 'Dagdemir A et al.',
'description' => '<h4>BACKGROUND/AIM:</h4>
<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
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<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
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'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
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<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
'date' => '2017-08-19',
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'description' => '<p>We here characterize the usability of archival formalin-fixed paraffin-embedded (FFPE) brain tissue as a resource for genetic and DNA methylation analyses with potential relevance for brain-manifested diseases. We analyzed FFPE samples from The Brain Collection, Aarhus University Hospital Risskov, Denmark (AUBC), constituting 9479 formalin-fixated brains making it one of the largest collections worldwide. DNA extracted from brain FFPE tissue blocks was interrogated for quality and usability in genetic and DNA methylation analyses by different molecular techniques. Overall, we found that DNA quality was inversely correlated with storage time and DNA quality was insufficient for Illumina methylation arrays; data from methylated DNA immunoprecipitation, clonal bisulfite sequencing, and pyrosequencing of BDNF and ST6GALNAC1 suggested that the original methylation pattern is indeed preserved. Proof-of-principle experiments predicting sex based on the methylation status of the X-inactivated SLC9A7 gene, or genotype differences of the Y and X chromosomes, showed consistency between predicted and actual sex for a subset of FFPE samples. In conclusion, even though DNA from FFPE samples is of low quality and technically challenging, it is likely that a subset of samples can provide reliable data given that the methodology used is designed for small DNA fragments. We propose that simple PCR-based quality control experiments at the genetic and DNA methylation level, carried out at the beginning of any given project, can be used to enrich for the best-performing FFPE samples. The apparent preservation of genetic and DNA methylation patterns in archival FFPE samples may bring along new perspectives for the identification of genetic and epigenetic changes associated with brain-manifested diseases.</p>',
'date' => '2016-12-19',
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'description' => '<p><strong>Background</strong> In utero and early-life experienced environmental exposures are suggested to play an important role in many multifactorial diseases potentially mediated through lasting effects on the epigenome. As the epigenome in addition remains modifiable throughout life, identifying specific disease-relevant biomarkers may prove challenging. This has led to an increased interest in epigenome-wide association studies using dried blood spots (DBS) routinely collected in perinatal screening programs. Such programs are in place in numerous countries around the world producing large and unique biobanks. However, availability of this biological material is highly limited as each DBS is made only from a few droplets of blood and storage conditions may be suboptimal for epigenetic studies. Furthermore, as relevant markers may reside outside gene bodies, epigenome-wide interrogation is needed.</p>
<p><strong>Results</strong> Here we demonstrate, as a proof of principle, that genome-wide interrogation of the methylome based on methylated DNA immunoprecipitation coupled with next-generation sequencing (MeDIP-seq) is feasible using a single 3.2 mm DBS punch (60 ng DNA) from filter cards archived for up to 16 years. The enrichment profile, sequence quality and distribution of reads across genetic regions were comparable between samples archived 16 years, 4 years and a freshly prepared control sample.</p>
<p><strong>Conclusions</strong> In summary, we show that high-quality MeDIP-seq data is achievable from neonatal screening filter cards stored at room temperature, thereby providing information on annotated as well as on non-RefSeq genes and repetitive elements. Moreover, the quantity of DNA from one DBS punch proved sufficient allowing for multiple epigenome studies using one single DBS.</p>',
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<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
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<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
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<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
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'description' => '<p><b></b>Background: Interrogation of DNA methylation profiles hold promise for improved diagnostics, as well as the delineation of the aetiology for common human diseases. However, as the primary tissue of the disease is often inaccessible without complicated and inconvenient interventions, there is an increasing interest in peripheral surrogate tissues. Whereas most work has been conducted on blood, saliva is now becoming recognized as an interesting alternative due to the simple and non-invasive manner of collection allowing for self-sampling. Results: In this study we have evaluated if saliva samples are suitable for DNA methylation studies using methylated DNA immunoprecipitation coupled to next-generation sequencing (MeDIP-seq). This was done by comparing the DNA methylation profile in saliva against the benchmark profile of peripheral blood from three individuals. We show that the output, quality, and depth of paired-end 50 bp sequencing reads are comparable between saliva and peripheral blood and, moreover, that the distribution of reads along genomic regions are similar and follow canonical methylation patterns. Conclusion: In summary, we show that high-quality MeDIP-seq data can be generated using saliva, thus supporting the future use of saliva in the generation of DNA methylation information at annotated genes, non-RefSeq genes, and repetitive elements relevant to human disease.</p>',
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<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
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<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
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<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
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<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
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<p><strong>Results</strong> Here we demonstrate, as a proof of principle, that genome-wide interrogation of the methylome based on methylated DNA immunoprecipitation coupled with next-generation sequencing (MeDIP-seq) is feasible using a single 3.2 mm DBS punch (60 ng DNA) from filter cards archived for up to 16 years. The enrichment profile, sequence quality and distribution of reads across genetic regions were comparable between samples archived 16 years, 4 years and a freshly prepared control sample.</p>
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<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
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<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
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'description' => '<p><b></b>Background: Interrogation of DNA methylation profiles hold promise for improved diagnostics, as well as the delineation of the aetiology for common human diseases. However, as the primary tissue of the disease is often inaccessible without complicated and inconvenient interventions, there is an increasing interest in peripheral surrogate tissues. Whereas most work has been conducted on blood, saliva is now becoming recognized as an interesting alternative due to the simple and non-invasive manner of collection allowing for self-sampling. Results: In this study we have evaluated if saliva samples are suitable for DNA methylation studies using methylated DNA immunoprecipitation coupled to next-generation sequencing (MeDIP-seq). This was done by comparing the DNA methylation profile in saliva against the benchmark profile of peripheral blood from three individuals. We show that the output, quality, and depth of paired-end 50 bp sequencing reads are comparable between saliva and peripheral blood and, moreover, that the distribution of reads along genomic regions are similar and follow canonical methylation patterns. Conclusion: In summary, we show that high-quality MeDIP-seq data can be generated using saliva, thus supporting the future use of saliva in the generation of DNA methylation information at annotated genes, non-RefSeq genes, and repetitive elements relevant to human disease.</p>',
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<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
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'description' => '<p>We here characterize the usability of archival formalin-fixed paraffin-embedded (FFPE) brain tissue as a resource for genetic and DNA methylation analyses with potential relevance for brain-manifested diseases. We analyzed FFPE samples from The Brain Collection, Aarhus University Hospital Risskov, Denmark (AUBC), constituting 9479 formalin-fixated brains making it one of the largest collections worldwide. DNA extracted from brain FFPE tissue blocks was interrogated for quality and usability in genetic and DNA methylation analyses by different molecular techniques. Overall, we found that DNA quality was inversely correlated with storage time and DNA quality was insufficient for Illumina methylation arrays; data from methylated DNA immunoprecipitation, clonal bisulfite sequencing, and pyrosequencing of BDNF and ST6GALNAC1 suggested that the original methylation pattern is indeed preserved. Proof-of-principle experiments predicting sex based on the methylation status of the X-inactivated SLC9A7 gene, or genotype differences of the Y and X chromosomes, showed consistency between predicted and actual sex for a subset of FFPE samples. In conclusion, even though DNA from FFPE samples is of low quality and technically challenging, it is likely that a subset of samples can provide reliable data given that the methodology used is designed for small DNA fragments. We propose that simple PCR-based quality control experiments at the genetic and DNA methylation level, carried out at the beginning of any given project, can be used to enrich for the best-performing FFPE samples. The apparent preservation of genetic and DNA methylation patterns in archival FFPE samples may bring along new perspectives for the identification of genetic and epigenetic changes associated with brain-manifested diseases.</p>',
'date' => '2016-12-19',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/27995571',
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'name' => 'Genome-wide DNA methylation profiling with MeDIP-seq using archived dried blood spots',
'authors' => 'Nicklas H. Staunstrup et al.',
'description' => '<p><strong>Background</strong> In utero and early-life experienced environmental exposures are suggested to play an important role in many multifactorial diseases potentially mediated through lasting effects on the epigenome. As the epigenome in addition remains modifiable throughout life, identifying specific disease-relevant biomarkers may prove challenging. This has led to an increased interest in epigenome-wide association studies using dried blood spots (DBS) routinely collected in perinatal screening programs. Such programs are in place in numerous countries around the world producing large and unique biobanks. However, availability of this biological material is highly limited as each DBS is made only from a few droplets of blood and storage conditions may be suboptimal for epigenetic studies. Furthermore, as relevant markers may reside outside gene bodies, epigenome-wide interrogation is needed.</p>
<p><strong>Results</strong> Here we demonstrate, as a proof of principle, that genome-wide interrogation of the methylome based on methylated DNA immunoprecipitation coupled with next-generation sequencing (MeDIP-seq) is feasible using a single 3.2 mm DBS punch (60 ng DNA) from filter cards archived for up to 16 years. The enrichment profile, sequence quality and distribution of reads across genetic regions were comparable between samples archived 16 years, 4 years and a freshly prepared control sample.</p>
<p><strong>Conclusions</strong> In summary, we show that high-quality MeDIP-seq data is achievable from neonatal screening filter cards stored at room temperature, thereby providing information on annotated as well as on non-RefSeq genes and repetitive elements. Moreover, the quantity of DNA from one DBS punch proved sufficient allowing for multiple epigenome studies using one single DBS.</p>',
'date' => '2016-07-26',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/27462375',
'doi' => '10.1186/s13148-016-0242-1',
'modified' => '2016-08-03 10:40:55',
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'name' => 'Epigenetic Modifications with DZNep, NaBu and SAHA in Luminal and Mesenchymal-like Breast Cancer Subtype Cells',
'authors' => 'Dagdemir A et al.',
'description' => '<h4>BACKGROUND/AIM:</h4>
<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
'date' => '2016-07-01',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/27365379',
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'description' => '<h4>BACKGROUND/AIM:</h4>
<p><abstracttext label="BACKGROUND/AIM" nlmcategory="OBJECTIVE">Numerous studies have shown that breast cancer and epigenetic mechanisms have a very powerful interactive relation. The MCF7 cell line, representative of luminal subtype and the MDA-MB 231 cell line representative of mesenchymal-like subtype were treated respectively with a Histone Methyl Transferase Inhibitors (HMTi), 3-Deazaneplanocin hydrochloride (DZNep), two histone deacetylase inhibitors (HDACi), sodium butyrate (NaBu), and suberoylanilide hydroxamic acid (SAHA) for 48 h.</abstracttext></p>
<h4>MATERIALS AND METHODS:</h4>
<p><abstracttext label="MATERIALS AND METHODS" nlmcategory="METHODS">Chromatin immunoprecipitation (ChIP) was used to observe HDACis (SAHA and NaBu) and HMTi (DZNep) impact on histones and more specifically on H3K27me3, H3K9ac and H3K4ac marks with Q-PCR analysis of BRCA1, SRC3 and P300 genes. Furthermore, the HDACi and HMTi effects on mRNA and protein expression of BRCA1, SRC3 and P300 genes were checked. In addition, statistical analyses were used.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">In the MCF7 luminal subtype with positive ER, H3k4ac was significantly increased on BRCA1 with SAHA. On the contrary, in the MDA-MB 231 breast cancer cell line, representative of mesenchymal-like subtype with negative estrogen receptor, HDACis had no effect. Also, DZNEP decreased significantly H3K27me3 on BRCA1 in MDA-MB 231. Besides, on SRC3, a significant increase for H3K4ac was obtained in MCF7 treated with SAHA. And DZNEP had no effect in MCF7. Also, in MDA-MB 231 treated with DZNEP, H3K27me3 significantly decreased on SRC3 while H3K4ac was significantly increased in MDA-MB-231 treated with SAHA or NaBu for P300.</abstracttext></p>
<h4>CONCLUSION:</h4>
<p><abstracttext label="CONCLUSION" nlmcategory="CONCLUSIONS">Luminal and mesenchymal-like breast cancer subtype cell lines seemed to act differently to HDACis (SAHA and NaBu) or HMTi (DZNEP) treatments.</abstracttext></p>',
'date' => '2016-07-01',
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$externalLink = ' <a href="http://www.ncbi.nlm.nih.gov/pubmed/27365379" target="_blank"><i class="fa fa-external-link"></i></a>'include - APP/View/Products/view.ctp, line 755
View::_evaluate() - CORE/Cake/View/View.php, line 971
View::_render() - CORE/Cake/View/View.php, line 933
View::render() - CORE/Cake/View/View.php, line 473
Controller::render() - CORE/Cake/Controller/Controller.php, line 963
ProductsController::slug() - APP/Controller/ProductsController.php, line 1052
ReflectionMethod::invokeArgs() - [internal], line ??
Controller::invokeAction() - CORE/Cake/Controller/Controller.php, line 491
Dispatcher::_invoke() - CORE/Cake/Routing/Dispatcher.php, line 193
Dispatcher::dispatch() - CORE/Cake/Routing/Dispatcher.php, line 167
[main] - APP/webroot/index.php, line 118
×
