Genome-wide analysis of aberrant methylation in human breast cancer cells using methyl-DNA immunoprecipitation combined with high-throughput sequencing

<p>Abstract</p> <p>Background</p> <p>Cancer cells undergo massive alterations to their DNA methylation patterns that result in aberrant gene expression and malignant phenotypes. However, the mechanisms that underlie methylome changes are not well understood nor is the genomic distribution of DNA methylation changes well characterized.</p> <p>Results</p> <p>Here, we performed methylated DNA immunoprecipitation combined with high-throughput sequencing (MeDIP-seq) to obtain whole-genome DNA methylation profiles for eight human breast cancer cell (BCC) lines and for normal human mammary epithelial cells (HMEC). The MeDIP-seq analysis generated non-biased DNA methylation maps by covering almost the entire genome with sufficient depth and resolution. The most prominent feature of the BCC lines compared to HMEC was a massively reduced methylation level particularly in CpG-poor regions. While hypomethylation did not appear to be associated with particular genomic features, hypermethylation preferentially occurred at CpG-rich gene-related regions independently of the distance from transcription start sites. We also investigated methylome alterations during epithelial-to-mesenchymal transition (EMT) in MCF7 cells. EMT induction was associated with specific alterations to the methylation patterns of gene-related CpG-rich regions, although overall methylation levels were not significantly altered. Moreover, approximately 40% of the epithelial cell-specific methylation patterns in gene-related regions were altered to those typical of mesenchymal cells, suggesting a cell-type specific regulation of DNA methylation.</p> <p>Conclusions</p> <p>This study provides the most comprehensive analysis to date of the methylome of human mammary cell lines and has produced novel insights into the mechanisms of methylome alteration during tumorigenesis and the interdependence between DNA methylome alterations and morphological changes.</p

統合失調症初発エピソードの認知機能障害の経過と発症5 年経過群との比較

OBJECTIVE: The course of neurocognitive deficits in schizophrenia has not yet been established. Therefore, we followed patients with first-episode schizophrenia to verify the course of these deficits. METHODS:In Study 1, tests of neurocognitive functioning were administered to patients with first-episode schizophrenia (FE group) every 6 months. Of the 26 patients who completed the baseline assessment, 19 completed a 6-month follow-up, and 13 completed a 1-year follow-up. In Study 2, 19 patients in FE group at 6-months when the neuropsychological measures was less influenced by psychotic symptoms and other patients who experienced schizophrenia 5-years earlier (5-year group) were compared. RESULTS:In Study 1, verbal memory, motor speed, and executive functions significantly improved at the 1-year follow-up. In Study 2, patients in 5-year group performed worse in verbal memory and executive functions than patients in FE at 6-month group, but marginally but significantly better in verbal fluency. CONCLUSIONS:Verbal memory, executive functions, and verbal fluency were significantly different between 5-year group and FE at 6-month group, and may indicate progression of schizophrenia. Executive functions may reflect the state of psychosis. Working memory and processing speed which did not change significantly from onset are needed to verify the course in further research.博士（医学）・甲610号・平成26年3月17

Novel large deletion involving EVC and EVC2 in Ellis–van Creveld syndrome

Ellis–van Creveld syndrome is an autosomal recessive skeletal dysplasia that is characterized by thoracic hypoplasia, polydactyly, oral abnormalities, and congenital heart disease. It is caused by pathogenic variants in the EVC or EVC2 genes. We report a case of a newborn with a compound heterozygous variant comprising NM_147127.5: c.1991dup:[p.Lys665Glufs*10] in the EVC2 gene and a novel large deletion involving exon 1 in EVC and exons 1–7 in EVC2

The Possible Role of TASK Channels in Rank-Ordered Recruitment of Motoneurons in the Dorsolateral Part of the Trigeminal Motor Nucleus.

Because a rank-ordered recruitment of motor units occurs during isometric contraction of jaw-closing muscles, jaw-closing motoneurons (MNs) may be recruited in a manner dependent on their soma sizes or input resistances (IRs). In the dorsolateral part of the trigeminal motor nucleus (dl-TMN) in rats, MNs abundantly express TWIK (two-pore domain weak inwardly rectifying K channel)-related acid-sensitive-K(+) channel (TASK)-1 and TASK3 channels, which determine the IR and resting membrane potential. Here we examined how TASK channels are involved in IR-dependent activation/recruitment of MNs in the rat dl-TMN by using multiple methods. The real-time PCR study revealed that single large MNs (&gt;35 μm) expressed TASK1 and TASK3 mRNAs more abundantly compared with single small MNs (15-20 μm). The immunohistochemistry revealed that TASK1 and TASK3 channels were complementarily distributed in somata and dendrites of MNs, respectively. The density of TASK1 channels seemed to increase with a decrease in soma diameter while there were inverse relationships between the soma size of MNs and IR, resting membrane potential, or spike threshold. Dual whole-cell recordings obtained from smaller and larger MNs revealed that the recruitment of MNs depends on their IRs in response to repetitive stimulation of the presumed Ia afferents. 8-Bromoguanosine-cGMP decreased IRs in small MNs, while it hardly changed those in large MNs, and subsequently decreased the difference in spike-onset latency between the smaller and larger MNs, causing a synchronous activation of MNs. These results suggest that TASK channels play critical roles in rank-ordered recruitment of MNs in the dl-TMN

Multidendritic sensory neurons in the adult Drosophila abdomen: origins, dendritic morphology, and segment- and age-dependent programmed cell death

<p>Abstract</p> <p>Background</p> <p>For the establishment of functional neural circuits that support a wide range of animal behaviors, initial circuits formed in early development have to be reorganized. One way to achieve this is local remodeling of the circuitry hardwiring. To genetically investigate the underlying mechanisms of this remodeling, one model system employs a major group of <it>Drosophila </it>multidendritic sensory neurons - the dendritic arborization (da) neurons - which exhibit dramatic dendritic pruning and subsequent growth during metamorphosis. The 15 da neurons are identified in each larval abdominal hemisegment and are classified into four categories - classes I to IV - in order of increasing size of their receptive fields and/or arbor complexity at the mature larval stage. Our knowledge regarding the anatomy and developmental basis of adult da neurons is still fragmentary.</p> <p>Results</p> <p>We identified multidendritic neurons in the adult <it>Drosophila </it>abdomen, visualized the dendritic arbors of the individual neurons, and traced the origins of those cells back to the larval stage. There were six da neurons in abdominal hemisegment 3 or 4 (A3/4) of the pharate adult and the adult just after eclosion, five of which were persistent larval da neurons. We quantitatively analyzed dendritic arbors of three of the six adult neurons and examined expression in the pharate adult of key transcription factors that result in the larval class-selective dendritic morphologies. The 'baseline design' of A3/4 in the adult was further modified in a segment-dependent and age-dependent manner. One of our notable findings is that a larval class I neuron, ddaE, completed dendritic remodeling in A2 to A4 and then underwent caspase-dependent cell death within 1 week after eclosion, while homologous neurons in A5 and in more posterior segments degenerated at pupal stages. Another finding is that the dendritic arbor of a class IV neuron, v'ada, was immediately reshaped during post-eclosion growth. It exhibited prominent radial-to-lattice transformation in 1-day-old adults, and the resultant lattice-shaped arbor persisted throughout adult life.</p> <p>Conclusion</p> <p>Our study provides the basis on which we can investigate the genetic programs controlling dendritic remodeling and programmed cell death of adult neurons, and the life-long maintenance of dendritic arbors.</p

高齢者福祉施設における入所者とデイサービス部門通所者に対して提供された給食の残食実態

　高齢者の栄養問題，特に低栄養の問題を検討するための基礎資料とするために，デイサービス部門を併設しているA特別養護老人ホームにおける残食量調査を行った。対象者は2週間の調査期間中に昼食を摂った同ホーム入所者と，同ホームのデイサービス部門への通所者である。日によっては通所者のみに対して追加献立が提供された場合もあったが，提供された給食の各人1回当たりの平均エネルギー量及びたんぱく質，脂質，炭水化物の各平均重量は，入所者及び通所者の間に有意差を認めなかった。同様なことは，平均給食提供量にも言えたが，平均残食量については，入所者のそれよりも通所者における方が少なく，両者の間に有意差を認めた。また，残食量に基づいて摂取エネルギー量を計算したところ，入所者の摂取エネルギー量は通所者におけるよりも有意に少ないことが分かった。以上のことは，高齢者福祉施設の入所者は，同じ施設のデイサービス部門への通所者におけるよりも低栄養状態にあることを示唆しているように思われる

A robust culture method for maintaining tumorigenic cancer stem cells in the hepatocellular carcinoma cell line Li‐7

Cancer tissues contain small populations of highly tumorigenic cells termed cancer stem cells (CSCs). Immortalized cell lines containing CSCs are valuable and powerful experimental tools for research into the characteristics of these stem cells. We previously reported that the hepatocellular carcinoma cell line Li‐7 includes abundant CD13+CD166− CSCs; however, the number of these cells decreases after long‐term culture as a result of differentiation to non‐CSC populations. To ensure consistent and reproducible results in experiments using Li‐7 cells, it is important that the CSC population is maintained stably regardless of culture duration and passage. In the present study, we found that a commercially available culture medium for maintenance of embryonic stem cells and induced pluripotent stem cells, mTeSR1, effectively prevented spontaneous differentiation by CD13+CD166− cells to CD13−CD166+ cells and therefore maintained the CSC population in Li‐7 cell cultures. CD13+CD166− CSCs maintained using this culture medium retained high tumorigenicity after transplantation into mice; they also showed the ability to differentiate in vitro into non‐CSC populations in RPMI‐1640 with 10% FBS medium. We analyzed gene expression profiles of CSC and non‐CSC populations in Li‐7 cultures using an RNA sequencing method. Genes such as FGFR, NOTCH1, and JAG1, that are associated with tumorigenicity and stemness, were upregulated in the CSC population. Our results suggest that CSCs can be maintained in immortalized cancer cell lines cultured over an extended period using a medium developed for culture of embryonic/induced pluripotent stem cells