Global Mapping of Cell Type–Specific Open Chromatin by FAIRE-seq Reveals the Regulatory Role of the NFI Family in Adipocyte Differentiation

Identification of regulatory elements within the genome is crucial for understanding the mechanisms that govern cell type–specific gene expression. We generated genome-wide maps of open chromatin sites in 3T3-L1 adipocytes (on day 0 and day 8 of differentiation) and NIH-3T3 fibroblasts using formaldehyde-assisted isolation of regulatory elements coupled with high-throughput sequencing (FAIRE-seq). FAIRE peaks at the promoter were associated with active transcription and histone modifications of H3K4me3 and H3K27ac. Non-promoter FAIRE peaks were characterized by H3K4me1+/me3-, the signature of enhancers, and were largely located in distal regions. The non-promoter FAIRE peaks showed dynamic change during differentiation, while the promoter FAIRE peaks were relatively constant. Functionally, the adipocyte- and preadipocyte-specific non-promoter FAIRE peaks were, respectively, associated with genes up-regulated and down-regulated by differentiation. Genes highly up-regulated during differentiation were associated with multiple clustered adipocyte-specific FAIRE peaks. Among the adipocyte-specific FAIRE peaks, 45.3% and 11.7% overlapped binding sites for, respectively, PPARγ and C/EBPα, the master regulators of adipocyte differentiation. Computational motif analyses of the adipocyte-specific FAIRE peaks revealed enrichment of a binding motif for nuclear family I (NFI) transcription factors. Indeed, ChIP assay showed that NFI occupy the adipocyte-specific FAIRE peaks and/or the PPARγ binding sites near PPARγ, C/EBPα, and aP2 genes. Overexpression of NFIA in 3T3-L1 cells resulted in robust induction of these genes and lipid droplet formation without differentiation stimulus. Overexpression of dominant-negative NFIA or siRNA–mediated knockdown of NFIA or NFIB significantly suppressed both induction of genes and lipid accumulation during differentiation, suggesting a physiological function of these factors in the adipogenic program. Together, our study demonstrates the utility of FAIRE-seq in providing a global view of cell type–specific regulatory elements in the genome and in identifying transcriptional regulators of adipocyte differentiation

Development of new optical imaging systems of oxygen metabolism and simultaneous measurement in hemodynamic changes using awake mice.

Background:PET allows the measurement of CBF, CBV and CMRO2 in human and plays an important role in the diagnosis of pathologic conditions and clinical research. On the other hand, in animal studies, there is no optical imaging system for evaluating changes in CBF and CBV, and oxygen metabolism, from the same brain area under awake condition.New method:In the present study, we developed a simultaneous measurement system of LSI and IOSI, which was verified by LDF. Moreover, to evaluate oxygen metabolism, FAI was performed from the same brain area as LSI and IOSI measurements.Results:The change in CBF according to LSI was correlated with that by LDF. Similarly, the change in CBV obtained by IOSI was also correlated with RBC concentration change measured by LDF. The change in oxygen metabolism by FAI was associated with that in CBF obtained by LSI, although the change in CBF was greater than that in oxygen metabolism.Comparison with existing method(s): We revealed that the relationship between oxygen metabolism and CBF as measured by our system was in good agreement with the relationship between CMRO2 and CBF in human PET studies.Conclusions:Our measurement system of CBF, CBV and oxygen metabolism is not only useful for studying neurovascular coupling, but also easily corroborates human PET studies

Afatinib-induced severe esophagitis in a lung cancer patient with an activated epidermal growth factor receptor mutation: A case report

A 58-year-old woman with lung cancer complaint odynophagia by sour food. Endoscopic examination revealed severe erosion strictly limited to the esophagus. Drug-induced esophagitis was suspected. She was taking afatinib, loxoprofen, pregabalin, lorazepam, a formulation of butyric acid bacteria, and amino acid supplements for more than 1 month at the time of developing esophagitis. Her appetite was very poor for several days before developing the esophagitis. Although not definitive, the most probable cause of her esophagitis is an increased blood concentration of afatinib due to prolonged starved condition

Inhibitory effects of caffeine on gustatory plasticity in the nematode Caenorhabditis elegans.

The effects of caffeine on salt chemotaxis learning were investigated using the nematode Caenorhabditis elegans. To estimate the degree of salt chemotaxis learning, nematodes were placed in a mixed solution of NaCl and caffeine, and then the chemotaxis index of NaCl was obtained from the nematodes placed on agar medium after pre-exposure to caffeine concentrations of 0.01, 0.1, 0.3, and 1.0%. Locomotor activity and preference behavior for caffeine were also estimated under these caffeine conditions. Nematodes pre-exposed to 0.3% caffeine showed inhibition of salt chemotaxis learning. Additional experiments indicated that nematodes showed a preference response to the middle concentration of caffeine (0.1%), with preference behavior declining in the 0.3% caffeine condition. Stable locomotor activity was observed under 0.01-0.3% caffeine conditions. These results suggest that salt chemotaxis learning with 0.3% caffeine is useful for investigating the effects of caffeine on learning in nematodes

Genetic dissection of intratumor heterogeneity of PD‐L1 expression in EGFR‐mutated lung adenocarcinoma

Abstract In this study, we investigated the association between PD‐L1 expression in tumor cells and underlying genetic mutations, which was analyzed in detail using laser microdissection and next‐generation sequencing analysis. To investigate whether driver mutations are involved in the background of PD‐L1 expression, the EGFR major activating mutation was selected as the most frequent driver mutation. Surgical resection specimens were used to extract sufficient amounts of nucleic acids for analysis, and the high tumor proportion score (TPS:100%) and low (TPS: 0%) PD‐L1‐expressing parts of the tumor were each laser microdissected to examine the association between PD‐L1 expression heterogeneity and genetic mutations within the same tumor. The association between PD‐L1 heterogeneity and gene mutations within the same tumor was investigated. Analysis showed no association between PD‐L1 expression heterogeneity and genetic variants, which were found to be almost identical. However, PD‐L1 expression was found to be associated with the number of tumor infiltrating lymphocytes (TILs) present in the tumor, which may be related to whether or not lymphocytes can infiltrate into the tumor depending on the tumor histological type (solid pattern, lepidic pattern, etc.) and other factors