Anaerobic Mercury Methylation and Demethylation by Geobacter bemidjiensis Bem

DOE PAGES

Lu, Xia; Liu, Yurong; Johs, Alexander; ...

2016-03-28

Two competing processes controlling the net production and bioaccumulation of neurotoxic methylmercury (MeHg) in natural ecosystems are microbial methylation and demethylation. Though mercury (Hg) methylation by anaerobic microorganisms and demethylation by aerobic Hg-resistant bacteria have both been extensively studied, little attention has been given to MeHg degradation by anaerobic bacteria, particularly the iron-reducing bacterium Geobacter bemidjensis Bem. Here we report, for the first time, that the strain G. bemidjensis Bem can methylate inorganic Hg and degrade MeHg concurrently under anoxic conditions. Our results suggest that G. bemidjensis cells utilize a reductive demethylation pathway to degrade MeHg, with elemental Hg(0) asmore » the major reaction product, possibly due to the presence of homologs encoding both organo-mercurial lyase (MerB) and mercuric reductase (MerA) in this organism. In addition, the cells can mediate multiple reactions including Hg/MeHg sorption, Hg reduction and oxidation, resulting in both time and concentration dependent Hg species transformations. Moderate concentrations (10 500 M) of Hg-binding ligands such as cysteine enhance Hg(II) methylation but inhibit MeHg degradation. These findings indicate a cycle of methylation and demethylation among anaerobic bacteria and suggest that mer-mediated demethylation may play a role in the net balance of MeHg production in anoxic water and sediments.« less

High glucose induces podocyte epithelial-to-mesenchymal transition by demethylation-mediated enhancement of MMP9 expression

PubMed Central

Ling, Li; Chen, Libo; Zhang, Changning; Gui, Shuyan; Zhao, Haiyan; Li, Zhengzhang

2018-01-01

Abnormal expression of matrix metalloproteinase 9 (MMP9) is correlated with podocyte epithelial-to-mesenchymal transition (EMT) in diabetic nephropathy (DN). However, the mechanisms underlying this process are not well defined. Site-specific demethylation may sustain high expression levels of target genes. In the present study, in order to investigate the association between DNA demethylation of MMP9 promoter and podocyte EMT in DN, human podocytes were cultured in high-glucose (HG) medium and a rat model of DN was established by intraperitoneal injection of streptozotocin (STZ) to determine whether site-specific demethylation of the MMP9 promoter was involved in regulating podocyte EMT in DN. The MTT assay was used to assess the effects of HG culture on the growth of podocytes, and the demethylation status of the MMP9 promoter was assessed by bisulfite sequencing polymerase chain reaction. mRNA and protein expression levels of MMP9, α-smooth muscle actin (α-SMA), podocalyxin and fibronectin-1 in podocytes were assessed by reverse transcription-quantitative PCR (RT-qPCR) and western blot analyses. The results demonstrated that HG treatment up regulated the expression of MMP9, α-SMA and fibronectin-1, but down regulated the expression of podocalyxin in podocytes. The MMP9 promoter region was revealed to contain a variety of demethylated CpG sites, and HG treatment reduced the rate of MMP9 promotermethylation, which, in turn, enhanced its promoter activity. In summary, these data suggested that demethylation of the MMP9 promoter may serve an important role in podocyte EMT in DN. The demethylation status of the MMP9 promoter maybe used as an important prognostic marker of DN in clinic. PMID:29436620

NF-κB mediates Gadd45β expression and DNA demethylation in the hippocampus during fear memory formation.

PubMed

Jarome, Timothy J; Butler, Anderson A; Nichols, Jessica N; Pacheco, Natasha L; Lubin, Farah D

2015-01-01

Gadd45-mediated DNA demethylation mechanisms have been implicated in the process of memory formation. However, the transcriptional mechanisms involved in the regulation of Gadd45 gene expression during memory formation remain unexplored. NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) controls transcription of genes in neurons and is a critical regulator of synaptic plasticity and memory formation. In silico analysis revealed several NF-κB (p65/RelA and cRel) consensus sequences within the Gadd45β gene promoter. Whether NF-κB activity regulates Gadd45 expression and associated DNA demethylation in neurons during memory formation is unknown. Here, we found that learning in a fear conditioning paradigm increased Gadd45β gene expression and brain-derivedneurotrophic factor (BDNF) DNA demethylation in area CA1 of the hippocampus, both of which were prevented with pharmacological inhibition of NF-κB activity. Further experiments found that conditional mutations in p65/RelA impaired fear memory formation but did not alter changes in Gadd45β expression. The learning-induced increases in Gadd45β mRNA levels, Gadd45β binding at the BDNF gene and BDNF DNA demethylation were blocked in area CA1 of the c-rel knockout mice. Additionally, local siRNA-mediated knockdown of c-rel in area CA1 prevented fear conditioning-induced increases in Gadd45β expression and BDNF DNA demethylation, suggesting that c-Rel containing NF-κB transcription factor complex is responsible for Gadd45β regulation during memory formation. Together, these results support a novel transcriptional role for NF-κB in regulation of Gadd45β expression and DNA demethylation in hippocampal neurons during fear memory.

Protein arginine methylation/demethylation and cancer

PubMed Central

Poulard, Coralie; Corbo, Laura; Le Romancer, Muriel

2016-01-01

Protein arginine methylation is a common post-translational modification involved in numerous cellular processes including transcription, DNA repair, mRNA splicing and signal transduction. Currently, there are nine known members of the protein arginine methyltransferase (PRMT) family, but only one arginine demethylase has been identified, namely the Jumonji domain-containing 6 (JMJD6). Although its demethylase activity was initially challenged, its dual activity as an arginine demethylase and a lysine hydroxylase is now recognized. Interestingly, a growing number of substrates for arginine methylation and demethylation play key roles in tumorigenesis. Though alterations in the sequence of these enzymes have not been identified in cancer, their overexpression is associated with various cancers, suggesting that they could constitute targets for therapeutic strategies. In this review, we present the recent knowledge of the involvement of PRMTs and JMJD6 in tumorigenesis. PMID:27556302

Anaerobic Mercury Methylation and Demethylation by Geobacter bemidjiensis Bem.

PubMed

Lu, Xia; Liu, Yurong; Johs, Alexander; Zhao, Linduo; Wang, Tieshan; Yang, Ziming; Lin, Hui; Elias, Dwayne A; Pierce, Eric M; Liang, Liyuan; Barkay, Tamar; Gu, Baohua

2016-04-19

Microbial methylation and demethylation are two competing processes controlling the net production and bioaccumulation of neurotoxic methylmercury (MeHg) in natural ecosystems. Although mercury (Hg) methylation by anaerobic microorganisms and demethylation by aerobic Hg-resistant bacteria have both been extensively studied, little attention has been given to MeHg degradation by anaerobic bacteria, particularly the iron-reducing bacterium Geobacter bemidjiensis Bem. Here we report, for the first time, that the strain G. bemidjiensis Bem can mediate a suite of Hg transformations, including Hg(II) reduction, Hg(0) oxidation, MeHg production and degradation under anoxic conditions. Results suggest that G. bemidjiensis utilizes a reductive demethylation pathway to degrade MeHg, with elemental Hg(0) as the major reaction product, possibly due to the presence of genes encoding homologues of an organomercurial lyase (MerB) and a mercuric reductase (MerA). In addition, the cells can strongly sorb Hg(II) and MeHg, reduce or oxidize Hg, resulting in both time and concentration-dependent Hg species transformations. Moderate concentrations (10-500 μM) of Hg-binding ligands such as cysteine enhance Hg(II) methylation but inhibit MeHg degradation. These findings indicate a cycle of Hg methylation and demethylation among anaerobic bacteria, thereby influencing net MeHg production in anoxic water and sediments.

MET18 Connects the Cytosolic Iron-Sulfur Cluster Assembly Pathway to Active DNA Demethylation in Arabidopsis

PubMed Central

Tang, Kai; Zhang, Huiming; Mangrauthia, Satendra K.; Lei, Mingguang; Hsu, Chuan-Chih; Hou, Yueh-Ju; Wang, Chunguo; Li, Yan; Tao, W. Andy; Zhu, Jian-Kang

2015-01-01

DNA demethylation mediated by the DNA glycosylase ROS1 helps determine genomic DNA methylation patterns and protects active genes from being silenced. However, little is known about the mechanism of regulation of ROS1 enzymatic activity. Using a forward genetic screen, we identified an anti-silencing (ASI) factor, ASI3, the dysfunction of which causes transgene promoter hyper-methylation and silencing. Map-based cloning identified ASI3 as MET18, a component of the cytosolic iron-sulfur cluster assembly (CIA) pathway. Mutation in MET18 leads to hyper-methylation at thousands of genomic loci, the majority of which overlap with hypermethylated loci identified in ros1 and ros1dml2dml3 mutants. Affinity purification followed by mass spectrometry indicated that ROS1 physically associates with MET18 and other CIA components. Yeast two-hybrid and split luciferase assays showed that ROS1 can directly interact with MET18 and another CIA component, AE7. Site-directed mutagenesis of ROS1 indicated that the conserved iron-sulfur motif is indispensable for ROS1 enzymatic activity. Our results suggest that ROS1-mediated active DNA demethylation requires MET18-dependent transfer of the iron-sulfur cluster, highlighting an important role of the CIA pathway in epigenetic regulation. PMID:26492035

Stereoselectivity of the demethylation of nicotine piperidine homologues by Nicotiana plumbaginifolia cell suspension cultures.

PubMed

Bartholomeusz, Trixie Ann; Molinié, Roland; Roscher, Albrecht; Felpin, François-Xavier; Gillet, Françoise; Lebreton, Jacques; Mesnard, François; Robins, Richard J

2005-08-01

The metabolism of (R,S)-N-methylanabasine and (R,S)-N-methylanatabine has been studied in a cell suspension culture of Nicotiana plumbaginifolia. Both substrates are effectively demethylated, anabasine or anatabine, respectively, accumulating in the medium. Similarly, there is strong stereoselectivity for the (R)-isomers of both substrates. The kinetics of metabolism of (R,S)-N-methylanabasine differ significantly from those of nicotine in that no further degradation of the initial demethylation product occurs. (R,S)-N-Methylanatabine, however, shows kinetics closer to those of nicotine, with loss of alkaloid from the system. Further more, (R,S)-N-methylanabasine does not diminish (S)-nicotine demethylation, indicating a lack of competition. However, the metabolism of (S)-nicotine is affected by the presence of (R,S)-N-methylanabasine. Hence, the demethylation of the piperidine homologues of nicotine is seen to be similar but not identical to that of the pyridine analogues. The implications of these different metabolic profiles in relation to the demethylation activity are discussed.

Enrichment of Desulfitobacterium spp. from forest and grassland soil using the O-demethylation of phenyl methyl ethers as a growth-selective process.

PubMed

Mingo, Felix Sebastian; Diekert, Gabriele; Studenik, Sandra

2016-02-01

The O-demethylation of phenyl methyl ethers under anaerobic conditions is a metabolic feature of acetogens and Desulfitobacterium spp. Desulfitobacteria as well as most acetogens are Gram-positive bacteria with a low GC content and belong to the phylum Firmicutes. The consumption of the phenyl methyl ether syringate was studied in enrichment cultures originating from five different topsoils. Desulfitobacterium spp. were detected in all topsoils via quantitative PCR. Desulfitobacteria could be enriched using the O-demethylation of syringate as a growth-selective process. The enrichment was significantly favoured by an external electron acceptor such as 3-chloro-4-hydroxyphenylacetate or thiosulfate. Upon cultivation in the presence of syringate and thiosulfate, which naturally occur in soil, a maximum number of 16S rRNA gene copies of Desulfitobacterium spp. was reached within the first three subcultivation steps and accounted for 3-10% of the total microbial community depending on the soil type. Afterwards, a loss of Desulfitobacterium gene copies was observed. Community analyses revealed that Proteobacteria, Acidobacteria, Actinobacteria and Bacteroidetes were the main phyla in the initial soil samples. Upon addition of syringate and thiosulfate as growth substrates, these phyla were rapidly outcompeted by Firmicutes, which were under-represented in soil. The main Firmicutes genera identified were Alkalibaculum, Clostridium, Sporobacterium, Sporomusa and Tissierella, which might be responsible for outcompeting the desulfitobacteria. Most of these organisms belong to the acetogens, which have previously been described to demethylate phenyl methyl ethers. The shift of the native community structure to almost exclusively Firmicutes supports the participation of members of this phylum in environmental demethylation processes.

Demethylation and cleavage of dimethylsulfoniopropionate in marine intertidal sediments

USGS Publications Warehouse

Visscher, P.T.; Kiene, R.P.; Taylor, B.F.

1994-01-01

Demethylation and cleavage of dimethylsulfoniopropionate (DMSP) was measured in three different types of,intertidal marine sediments: a cyanobacterial mat, a diatom-covered tidal flat and a carbonate sediment. Consumption rates of added DMSP were highest in cyanobacterial mat slurries (59 ?? mol DMSP l-1 slurry h-1) and lower in slurries from a diatom mat and a carbonate tidal sediment (24 and 9 ??mol DMSP l-1 h-1, respectively). Dimethyl sulfide (DMS) and 3-mercaptopropionate (MPA) were produced simultaneously during DMSP consumption, indicating that cleavage and demethylation occurred at the same time. Viable counts of DMSP-utilizing bacteria revealed a population of 2 x 107 cells cm-3 sediment (90% of these cleaved DMSP to DMS, 10% demethylated DMSP to MPA) in the cyanobacterial mat, 7 x 105 cells cm-3 in the diatom mat (23% cleavers, 77% demethylators), and 9 x 104 cells cm-3 (20% cleavers and 80% demethylators) in the carbonate sediment. In slurries of the diatom mat, the rate of MPA production from added 3-methiolpropionate (MMPA) was 50% of the rate of MPA formation from DMSP. The presence of a large population of demethylating bacteria and the production of MPA from DMSP suggest that the demethylation pathway, in addition to cleavage, contributes significantly to DMSP consumption in coastal sediments.

Stimulation of ribosomal RNA gene promoter by transcription factor Sp1 involves active DNA demethylation by Gadd45-NER pathway.

PubMed

Rajput, Pallavi; Pandey, Vijaya; Kumar, Vijay

2016-08-01

The well-studied Pol II transcription factor Sp1 has not been investigated for its regulatory role in rDNA transcription. Here, we show that Sp1 bound to specific sites on rDNA and localized into the nucleoli during the G1 phase of cell cycle to activate rDNA transcription. It facilitated the recruitment of Pol I pre-initiation complex and impeded the binding of nucleolar remodeling complex (NoRC) to rDNA resulting in the formation of euchromatin active state. More importantly, Sp1 also orchestrated the site-specific binding of Gadd45a-nucleotide excision repair (NER) complex resulting in active demethylation and transcriptional activation of rDNA. Interestingly, knockdown of Sp1 impaired rDNA transcription due to reduced engagement of the Gadd45a-NER complex and hypermethylation of rDNA. Thus, the present study unveils a novel role of Sp1 in rDNA transcription involving promoter demethylation. Copyright © 2016 Elsevier B.V. All rights reserved.

Method for recovering and using lignin in adhesive resins by extracting demethylated lignin

DOEpatents

Schroeder, Herbert A.

1991-01-01

Lignin, or a lignin derived material, which has been significantly demethylated (e.g., the demethylated lignin found in the raffinate produced as a by-product of dimethyl sulfide production which can be carried out using the spent liquor from wood pulping operations) can be isolated by a process wherein an organic solvent is added to a lignin-containing aqueous solution. The organic solvent is typically a polar, and at least a partially water-immiscible substance such as, for example, ethyl acetate. The resulting lignin-containing aqueous solution/organic solvent mixture is acidified to produce a water layer which is discarded and an organic solvent layer which contains the demethylated lignin. Upon its recovery, the demethylated lignin is preferably dried and stored until it is used (along with an alkali, an aldehyde and an adhesive filler) in compounding an adhesive of the type generally used in the manufacture of plywood.

Method for recovering and using lignin in adhesive resins by extracting demethylated lignin

DOEpatents

Schroeder, Herbert A.

1991-01-01

Lignin, or a lignin derived material, which has been significantly demethylated (e.g., the demethylated lignin found in the raffinate produced as a by-product of dimethyl sulfide production which can be carried out using the spent liquor from wood pulping operations) can be isolated by a process wherein an organic solvent is added to a lignin-containing aqueous solution. The organic solvent is typically a polar, and at least a partially water-immiscible substance such as, for example, ethyl acetate. The resulting lignin-containing aqueous solution/organic solvent mixture is acidified to produce a water layer which is discarded and an organic solvent layer which contains the demethylated lignin. Upon its recovery, the demethylated lignin is dissolved in an alkaline solution to which an aldehyde source is added to produce a resol-type resin. The aldehyde source may be formaldehyde in solution, paraformaldehyde, hexamethylenetetramine, or other aldehydes including acetaldehyde, furfural, and their derivatives.

Early demethylation of non-CpG, CpC-rich, elements in the myogenin 5′-flanking region

PubMed Central

Fuso, Andrea; Ferraguti, Giampiero; Grandoni, Francesco; Ruggeri, Raffaella; Scarpa, Sigfrido; Strom, Roberto

2010-01-01

The dynamic changes and structural patterns of DNA methylation of genes without CpG islands are poorly characterized. The relevance of CpG to the non-CpG methylation equilibrium in transcriptional repression is unknown. In this work, we analyzed the DNA methylation pattern of the 5′-flanking of the myogenin gene, a positive regulator of muscle differentiation with no CpG island and low CpG density, in both C2C12 muscle satellite cells and embryonic muscle. Embryonic brain was studied as a non-expressing tissue. High levels of both CpG and non-CpG methylation were observed in non-expressing experimental conditions. Both CpG and non-CpG methylation rapidly dropped during muscle differentiation and myogenin transcriptional activation with active demethylation dynamics. Non-CpG demethylation occurred more rapidly than CpG demethylation. Demethylation spread from initially highly methylated short CpC-rich elements to a virtually unmethylated status. These short elements have a high CpC content and density, share some motifs and largely coincide with putative recognition sequences of some differentiation-related transcription factors. Our findings point to a dynamically controlled equilibrium between CpG and non-CpG active demethylation in the transcriptional control of tissue-specific genes. The short CpC-rich elements are new structural features of the methylation machinery, whose functions may include priming the complete demethylation of a transcriptionally crucial DNA region. PMID:20935518

Mercury methylation and demethylation in Hg-contaminated lagoon sediments (Marano and Grado Lagoon, Italy)

NASA Astrophysics Data System (ADS)

Hines, Mark E.; Poitras, Erin N.; Covelli, Stefano; Faganeli, Jadran; Emili, Andrea; Žižek, Suzana; Horvat, Milena

2012-11-01

Mercury (Hg) transformation activities and sulfate (SO42-) reduction were studied in sediments of the Marano and Grado Lagoons in the Northern Adriatic Sea region as part of the "MIRACLE" project. The lagoons, which are sites of clam (Tapes philippinarum) farming, have been receiving excess Hg from the Isonzo River for centuries. Marano Lagoon is also contaminated from a chlor-alkali plant. Radiotracer methods were used to measure mercury methylation (230Hg, 197Hg), methylmercury (MeHg) demethylation (14C-MeHg) and SO42- reduction (35S) in sediment cores collected in autumn, winter and summer. Mercury methylation rate constants ranged from near zero to 0.054 day-1, generally decreased with depth, and were highest in summer. Demethylation rate constants were much higher than methylation reaching values of ˜0.6 day-1 in summer. Demethylation occurred via the oxidative pathway, except in winter when the reductive pathway increased in importance in surficial sediments. Sulfate reduction was also most active in summer (up to 1600 nmol mL-1 day-1) and depth profiles reflected seasonally changing redox conditions near the surface. Methylation and demethylation rate constants correlated positively with SO42- reduction and pore-water Hg concentrations, and inversely with Hg sediment-water partition coefficients indicating the importance of SO42- reduction and Hg dissolution on Hg cycling. Hg transformation rates were calculated using rate constants and concentrations of Hg species. In laboratory experiments, methylation was inhibited by amendments of the SO42--reduction inhibitor molybdate and by nitrate. Lagoon sediments displayed a dynamic seasonal cycle in which Hg dissolution in spring/summer stimulated Hg methylation, which was followed by a net loss of MeHg in autumn from demethylation. Sulfate-reducing bacteria (SRB) tended to be responsible for methylation of Hg and the oxidative demethylation of MeHg. However, during winter in surficial sediments, iron

ERRα induces H3K9 demethylation by LSD1 to promote cell invasion

PubMed Central

Carnesecchi, Julie; Forcet, Christelle; Zhang, Ling; Tribollet, Violaine; Barenton, Bruno; Boudra, Rafik; Cerutti, Catherine; Billas, Isabelle M. L.; Sérandour, Aurélien A.; Carroll, Jason S.; Beaudoin, Claude; Vanacker, Jean-Marc

2017-01-01

Lysine Specific Demethylase 1 (LSD1) removes mono- and dimethyl groups from lysine 4 of histone H3 (H3K4) or H3K9, resulting in repressive or activating (respectively) transcriptional histone marks. The mechanisms that control the balance between these two antagonist activities are not understood. We here show that LSD1 and the orphan nuclear receptor estrogen-related receptor α (ERRα) display commonly activated genes. Transcriptional activation by LSD1 and ERRα involves H3K9 demethylation at the transcriptional start site (TSS). Strikingly, ERRα is sufficient to induce LSD1 to demethylate H3K9 in vitro. The relevance of this mechanism is highlighted by functional data. LSD1 and ERRα coregulate several target genes involved in cell migration, including the MMP1 matrix metallo-protease, also activated through H3K9 demethylation at the TSS. Depletion of LSD1 or ERRα reduces the cellular capacity to invade the extracellular matrix, a phenomenon that is rescued by MMP1 reexpression. Altogether our results identify a regulatory network involving a direct switch in the biochemical activities of a histone demethylase, leading to increased cell invasion. PMID:28348226

ERRα induces H3K9 demethylation by LSD1 to promote cell invasion.

PubMed

Carnesecchi, Julie; Forcet, Christelle; Zhang, Ling; Tribollet, Violaine; Barenton, Bruno; Boudra, Rafik; Cerutti, Catherine; Billas, Isabelle M L; Sérandour, Aurélien A; Carroll, Jason S; Beaudoin, Claude; Vanacker, Jean-Marc

2017-04-11

Lysine Specific Demethylase 1 (LSD1) removes mono- and dimethyl groups from lysine 4 of histone H3 (H3K4) or H3K9, resulting in repressive or activating (respectively) transcriptional histone marks. The mechanisms that control the balance between these two antagonist activities are not understood. We here show that LSD1 and the orphan nuclear receptor estrogen-related receptor α (ERRα) display commonly activated genes. Transcriptional activation by LSD1 and ERRα involves H3K9 demethylation at the transcriptional start site (TSS). Strikingly, ERRα is sufficient to induce LSD1 to demethylate H3K9 in vitro. The relevance of this mechanism is highlighted by functional data. LSD1 and ERRα coregulate several target genes involved in cell migration, including the MMP1 matrix metallo-protease, also activated through H3K9 demethylation at the TSS. Depletion of LSD1 or ERRα reduces the cellular capacity to invade the extracellular matrix, a phenomenon that is rescued by MMP1 reexpression. Altogether our results identify a regulatory network involving a direct switch in the biochemical activities of a histone demethylase, leading to increased cell invasion.

Integrated genomic analysis of colorectal cancer progression reveals activation of EGFR through demethylation of the EREG promoter

PubMed Central

Qu, X; Sandmann, T; Frierson, H; Fu, L; Fuentes, E; Walter, K; Okrah, K; Rumpel, C; Moskaluk, C; Lu, S; Wang, Y; Bourgon, R; Penuel, E; Pirzkall, A; Amler, L; Lackner, M R; Tabernero, J; Hampton, G M; Kabbarah, O

2016-01-01

Key molecular drivers that underlie transformation of colonic epithelium into colorectal adenocarcinoma (CRC) are well described. However, the mechanisms through which clinically targeted pathways are activated during CRC progression have yet to be elucidated. Here, we used an integrative genomics approach to examine CRC progression. We used laser capture microdissection to isolate colonic crypt cells, differentiated surface epithelium, adenomas, carcinomas and metastases, and used gene expression profiling to identify pathways that were differentially expressed between the different cell types. We identified a number of potentially important transcriptional changes in developmental and oncogenic pathways, and noted a marked upregulation of EREG in primary and metastatic cancer cells. We confirmed this pattern of gene expression by in situ hybridization and observed staining consistent with autocrine expression in the tumor cells. Upregulation of EREG during the adenoma–carcinoma transition was associated with demethylation of two key sites within its promoter, and this was accompanied by an increase in the levels of epidermal growth factor receptor (EGFR) phosphorylation, as assessed by reverse-phase protein analysis. In CRC cell lines, we demonstrated that EREG demethylation led to its transcriptional upregulation, higher levels of EGFR phosphorylation, and sensitization to EGFR inhibitors. Low levels of EREG methylation in patients who received cetuximab as part of a phase II study were associated with high expression of the ligand and a favorable response to therapy. Conversely, high levels of promoter methylation and low levels of EREG expression were observed in tumors that progressed after treatment. We also noted an inverse correlation between EREG methylation and expression levels in several other cancers, including those of the head and neck, lung and bladder. Therefore, we propose that upregulation of EREG expression through promoter demethylation

DNA demethylation of inflammasome-associated genes is enhanced in patients with cryopyrin-associated periodic syndromes.

PubMed

Vento-Tormo, Roser; Álvarez-Errico, Damiana; Garcia-Gomez, Antonio; Hernández-Rodríguez, José; Buján, Segundo; Basagaña, Maria; Méndez, Maria; Yagüe, Jordi; Juan, Manel; Aróstegui, Juan I; Ballestar, Esteban

2017-01-01

Inflammasomes are cytosolic multiprotein complexes in macrophages. They assemble after infection- or stress-associated stimuli, activating both caspase-1-mediated inflammatory cytokine secretion and pyroptosis. Increased inflammasome activity resulting from gene mutations is related to monogenic autoinflammatory syndromes. However, variable penetrance among patients with the same gene mutations suggests involvement of additional mechanisms associated with inflammasome gene regulation. We sought to investigate the role of DNA demethylation in activating inflammasome genes during macrophage differentiation and monocyte activation in healthy control subjects and patients with autoinflammatory syndrome. Inflammasome-related genes were tested for DNA methylation and mRNA levels by using bisulfite pyrosequencing and quantitative RT-PCR in monocytes in vitro differentiated to macrophages and exposed to inflammatory conditions. The contribution of Tet methylcytosine dioxygenase 2 (TET2) and nuclear factor κB to DNA demethylation was tested by using chromatin immunoprecipitation, small interfering RNA-mediated downregulation, and pharmacologic inhibition. We observed that inflammasome-related genes are rapidly demethylated in both monocyte-to-macrophage differentiation and on monocyte activation. Demethylation associates with increased gene expression, and both mechanisms are impaired when TET2 and nuclear factor κB are downregulated. We analyzed DNA methylation levels of inflammasome-related genes in patients with cryopyrin-associated periodic syndromes (CAPS) and familial Mediterranean fever, 2 archetypical monogenic autoinflammatory syndromes. Under the above conditions, monocytes from untreated patients with CAPS undergo more efficient DNA demethylation than those of healthy subjects. Interestingly, patients with CAPS treated with anti-IL-1 drugs display methylation levels similar to those of healthy control subjects. Our study is the first to demonstrate the

Eicosapentaenoic acid induces DNA demethylation in carcinoma cells through a TET1-dependent mechanism.

PubMed

Ceccarelli, Veronica; Valentini, Virginia; Ronchetti, Simona; Cannarile, Lorenza; Billi, Monia; Riccardi, Carlo; Ottini, Laura; Talesa, Vincenzo Nicola; Grignani, Francesco; Vecchini, Alba

2018-05-14

In cancer cells, global genomic hypomethylation is found together with localized hypermethylation of CpG islands within the promoters and regulatory regions of silenced tumor suppressor genes. Demethylating agents may reverse hypermethylation, thus promoting gene re-expression. Unfortunately, demethylating strategies are not efficient in solid tumor cells. DNA demethylation is mediated by ten-eleven translocation enzymes (TETs). They sequentially convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which is associated with active transcription; 5-formylcytosine; and finally, 5-carboxylcytosine. Although α-linolenic acid, eicosapentaenoic acid (EPA), and docosahexaenoic acid, the major n-3 polyunsaturated fatty acids, have anti-cancer effects, their action, as DNA-demethylating agents, has never been investigated in solid tumor cells. Here, we report that EPA demethylates DNA in hepatocarcinoma cells. EPA rapidly increases 5hmC on DNA, inducing p21 Waf1/Cip1 gene expression, which slows cancer cell-cycle progression. We show that the underlying molecular mechanism involves TET1. EPA simultaneously binds peroxisome proliferator-activated receptor γ (PPARγ) and retinoid X receptor α (RXRα), thus promoting their heterodimer and inducing a PPARγ-TET1 interaction. They generate a TET1-PPARγ-RXRα protein complex, which binds to a hypermethylated CpG island on the p21 gene, where TET1 converts 5mC to 5hmC. In an apparent shuttling motion, PPARγ and RXRα leave the DNA, whereas TET1 associates stably. Overall, EPA directly regulates DNA methylation levels, permitting TET1 to exert its anti-tumoral function.-Ceccarelli, V., Valentini, V., Ronchetti, S., Cannarile, L., Billi, M., Riccardi, C., Ottini, L., Talesa, V. N., Grignani, F., Vecchini, A., Eicosapentaenoic acid induces DNA demethylation in carcinoma cells through a TET1-dependent mechanism.

Vitamin C induces specific demethylation of H3K9me2 in mouse embryonic stem cells via Kdm3a/b.

PubMed

Ebata, Kevin T; Mesh, Kathryn; Liu, Shichong; Bilenky, Misha; Fekete, Alexander; Acker, Michael G; Hirst, Martin; Garcia, Benjamin A; Ramalho-Santos, Miguel

2017-01-01

Histone methylation patterns regulate gene expression and are highly dynamic during development. The erasure of histone methylation is carried out by histone demethylase enzymes. We had previously shown that vitamin C enhances the activity of Tet enzymes in embryonic stem (ES) cells, leading to DNA demethylation and activation of germline genes. We report here that vitamin C induces a remarkably specific demethylation of histone H3 lysine 9 dimethylation (H3K9me2) in naïve ES cells. Vitamin C treatment reduces global levels of H3K9me2, but not other histone methylation marks analyzed, as measured by western blot, immunofluorescence and mass spectrometry. Vitamin C leads to widespread loss of H3K9me2 at large chromosomal domains as well as gene promoters and repeat elements. Vitamin C-induced loss of H3K9me2 occurs rapidly within 24 h and is reversible. Importantly, we found that the histone demethylases Kdm3a and Kdm3b are required for vitamin C-induced demethylation of H3K9me2. Moreover, we show that vitamin C-induced Kdm3a/b-mediated H3K9me2 demethylation and Tet-mediated DNA demethylation are independent processes at specific loci. Lastly, we document Kdm3a/b are partially required for the upregulation of germline genes by vitamin C. These results reveal a specific role for vitamin C in histone demethylation in ES cells and document that DNA methylation and H3K9me2 cooperate to silence germline genes in pluripotent cells.

The Mechanism of Autoinduction of Methadone N-demethylation in Human Hepatocytes

PubMed Central

Campbell, Scott D.; Crafford, Amanda; Williamson, Brian L.; Kharasch, Evan D.

2013-01-01

Background There is considerable inter-and intraindividual variability in methadone metabolism and clearance. Methadone dosing is particularly challenging during initiation of therapy, due to time-dependent increases in hepatic clearance (autoinduction). Although methadone N-demethylation is catalyzed in vitro by cytochrome P4502B6 (CYP2B6) and CYP3A4, and clearance in vivo depends on CYP2B6, mechanism(s) of autoinduction are incompletely understood. In this investigation we determined mechanism(s) of methadone autoinduction using human hepatocytes. Methods Fresh human hepatocytes were exposed to 0.1-10 μM methadone for 72 hr. Cells were washed and methadone N-demethylation assessed. CYP2B6, CYP3A4, and CYP3A5 mRNA, protein expression (by gel-free high performance liquid chromatography-mass spectrometry) and catalytic activity (bupropion hydroxylation and alfentanil dealkylation for CYP2B6 and CYP3A4/5, respectively) were measured. Mechanisms of CYP induction were characterized using pregnane X receptor and constitutive androstane receptor reporter gene assays. Results Methadone (10 μM) increased methadone N-demethylation 2-fold, CYP2B6 and CYP3A4 mRNA 3-fold, and protein expression 2-fold. CYP3A5 mRNA was unchanged. CYP2B6 and CYP3A4/5 activities increased 2-fold. Induction by methadone enantiomers (R- vs S-methadone) did not differ. Induction was relatively weak compared with maximum induction by phenobarbital and rifampin. Lower methadone concentrations had smaller effects. Methadone was an agonist for the pregnane X receptor but not the constitutive androstane receptor. Conclusions Methadone caused concentration-dependent autoinduction of methadone N-demethylation in human hepatocytes, related to induction of CYP2B6 and CYP3A4 mRNA expression, protein expression, and catalytic activity. Induction was related to pregnane X receptor but not constitutive androstane receptor activation. These in vitro findings provide mechanistic insights into clinical

DNA demethylation activates genes in seed maternal integument development in rice (Oryza sativa L.).

PubMed

Wang, Yifeng; Lin, Haiyan; Tong, Xiaohong; Hou, Yuxuan; Chang, Yuxiao; Zhang, Jian

2017-11-01

DNA methylation is an important epigenetic modification that regulates various plant developmental processes. Rice seed integument determines the seed size. However, the role of DNA methylation in its development remains largely unknown. Here, we report the first dynamic DNA methylomic profiling of rice maternal integument before and after pollination by using a whole-genome bisulfite deep sequencing approach. Analysis of DNA methylation patterns identified 4238 differentially methylated regions underpin 4112 differentially methylated genes, including GW2, DEP1, RGB1 and numerous other regulators participated in maternal integument development. Bisulfite sanger sequencing and qRT-PCR of six differentially methylated genes revealed extensive occurrence of DNA hypomethylation triggered by double fertilization at IAP compared with IBP, suggesting that DNA demethylation might be a key mechanism to activate numerous maternal controlling genes. These results presented here not only greatly expanded the rice methylome dataset, but also shed novel insight into the regulatory roles of DNA methylation in rice seed maternal integument development. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

EG-13GENOME-WIDE METHYLATION ANALYSIS IDENTIFIES GENOMIC DNA DEMETHYLATION DURING MALIGNANT PROGRESSION OF GLIOMAS

PubMed Central

Saito, Kuniaki; Mukasa, Akitake; Nagae, Genta; Aihara, Koki; Otani, Ryohei; Takayanagi, Shunsaku; Omata, Mayu; Tanaka, Shota; Shibahara, Junji; Takahashi, Miwako; Momose, Toshimitsu; Shimamura, Teppei; Miyano, Satoru; Narita, Yoshitaka; Ueki, Keisuke; Nishikawa, Ryo; Nagane, Motoo; Aburatani, Hiroyuki; Saito, Nobuhito

2014-01-01

Low-grade gliomas often undergo malignant progression, and these transformations are a leading cause of death in patients with low-grade gliomas. However, the molecular mechanisms underlying malignant tumor progression are still not well understood. Recent evidence indicates that epigenetic deregulation is an important cause of gliomagenesis; therefore, we examined the impact of epigenetic changes during malignant progression of low-grade gliomas. Specifically, we used the Illumina Infinium Human Methylation 450K BeadChip to perform genome-wide DNA methylation analysis of 120 gliomas and four normal brains. This study sample included 25 matched-pairs of initial low-grade gliomas and recurrent tumors (temporal heterogeneity) and 20 of the 25 recurring tumors recurred as malignant progressions, and one matched-pair of newly emerging malignant lesions and pre-existing lesions (spatial heterogeneity). Analyses of methylation profiles demonstrated that most low-grade gliomas in our sample (43/51; 84%) had a CpG island methylator phenotype (G-CIMP). Remarkably, approximately 50% of secondary glioblastomas that had progressed from low-grade tumors with the G-CIMP status exhibited a characteristic partial demethylation of genomic DNA during malignant progression, but other recurrent gliomas showed no apparent change in DNA methylation pattern. Interestingly, we found that most loci that were demethylated during malignant progression were located outside of CpG islands. The information of histone modifications patterns in normal human astrocytes and embryonal stem cells also showed that the ratio of active marks at the site corresponding to DNA demethylated loci in G-CIMP-demethylated tumors was significantly lower; this finding indicated that most demethylated loci in G-CIMP-demethylated tumors were likely transcriptionally inactive. A small number of the genes that were upregulated and had demethylated CpG islands were associated with cell cycle-related pathway. In

Induction of anti-aging gene klotho with a small chemical compound that demethylates CpG islands

PubMed Central

Jung, Dongju; Xu, Yuechi; Sun, Zhongjie

2017-01-01

Klotho (KL) is described as an anti-aging gene because mutation of Kl gene leads to multiple pre-mature aging phenotypes and shortens lifespan in mice. Growing evidence suggests that an increase in KL expression may be beneficial for age-related diseases such as arteriosclerosis and diabetes. It remains largely unknown, however, how Kl expression could be induced. Here we discovered novel molecular mechanism for induction of Kl expression with a small molecule ‘Compound H’, N-(2-chlorophenyl)-1H-indole-3-caboxamide. Compound H was originally identified through a high-throughput screening of small molecules for identifying Kl inducers. However, how Compound H induces Kl expression has never been investigated. We found that Compound H increased Kl expression via demethylation in CpG islands of the Kl gene. The demethylation was accomplished by activating demethylases rather than inhibiting methylases. Due to demethylation, Compound H enhanced binding of transcription factors, Pax4 and Kid3, to the promoter of the Kl gene. Pax4 and Kid3 regulated Kl promoter activity positively and negatively, respectively. Thus, our results show that demethylation is an important molecular mechanism that mediates Compound H-induced Kl expression. Further investigation is warranted to determine whether Compound H demethylates the Kl gene in vivo and whether it can serve as a therapeutic agent for repressing or delaying the onset of age-related diseases. PMID:28657902

Simulated vibrational spectra of aflatoxins and their demethylated products and the estimation of the energies of the demethylation reactions.

PubMed

Billes, Ferenc; Móricz, Agnes M; Tyihák, Erno; Mikosch, Hans

2006-06-01

The structure of four natural mycotoxins, the aflatoxin B1, B2, G1 and G2 and their demethylated products were optimized with quantum chemical method. The energies and the thermodynamic functions of the molecules were calculated and applied to calculation of the reaction energies of the demethylations. Further results of the calculations are the vibrational force constants, the infrared spectra of the molecules and the assignments of the spectral bands.

TET1 promotes cisplatin-resistance via demethylating the vimentin promoter in ovarian cancer.

PubMed

Han, Xi; Zhou, Yuanyuan; You, Yuanyi; Lu, Jiaojiao; Wang, Lijie; Hou, Huilian; Li, Jing; Chen, Wei; Zhao, Le; Li, Xu

2017-04-01

The development of chemo-resistance impairs the outcome of the first line platinum-based chemotherapies for ovarian cancer. Deregulation of DNA methylation/demethylation provides a critical mechanism for the occurrence of chemo-resistance. The ten-eleven translocation (TET) family of dioxygenases including TET1/2/3 plays an important part in DNA demethylation, but their roles in cisplatin resistance have not been elucidated. Using cisplatin-sensitive and cisplatin-resistant ovarian cancer cell models, we found that TET1 was significantly upregulated in cisplatin-resistant CP70 cells compared with that in cisplatin-sensitive A2780 cells. Ectopic expression of TET1 in A2780 cells promoted cisplatin resistance and decreased cytotoxicity induced by cisplatin, while inhibition of TET1 by siRNA transfection in CP70 cells attenuated cisplatin resistance and enhanced cytotoxicity of cisplatin. Increased TET1 induced re-expression of vimentin through active DNA demethylation, and cause partial epithelial-to-mesenchymal (EMT) in A2780 cells. Contrarily, knocking down of TET1 in CP70 cells reduced vimentin expression and reversed EMT process. Immunohistochemical analysis of TET1 in human ovarian cancer tissues revealed that TET1 existed in nucleus and cytoplasm in ovarian cancer tissues. And the expression of nuclear TET1 was positively correlated with residual tumor and chemotherapeutic response. Thus, TET1 expression causes resistance to cisplatin and one of the targets of TET1 action is vimentin in ovarian cancer. © 2017 International Federation for Cell Biology.

Dynamic changes in DNA demethylation in the tree shrew (Tupaia belangeri chinensis) brain during postnatal development and aging.

PubMed

Wei, Shu; Hua, Hai-Rong; Chen, Qian-Quan; Zhang, Ying; Chen, Fei; Li, Shu-Qing; Li, Fan; Li, Jia-Li

2017-03-18

Brain development and aging are associated with alterations in multiple epigenetic systems, including DNA methylation and demethylation patterns. Here, we observed that the levels of the 5-hydroxymethylcytosine (5hmC) ten-eleven translocation (TET) enzyme-mediated active DNA demethylation products were dynamically changed and involved in postnatal brain development and aging in tree shrews ( Tupaia belangeri chinensis ). The levels of 5hmC in multiple anatomic structures showed a gradual increase throughout postnatal development, whereas a significant decrease in 5hmC was found in several brain regions in aged tree shrews, including in the prefrontal cortex and hippocampus, but not the cerebellum. Active changes in Tet mRNA levels indicated that TET2 and TET3 predominantly contributed to the changes in 5hmC levels. Our findings provide new insight into the dynamic changes in 5hmC levels in tree shrew brains during postnatal development and aging processes.

Arsenic Demethylation by a C·As Lyase in Cyanobacterium Nostoc sp. PCC 7120.

PubMed

Yan, Yu; Ye, Jun; Xue, Xi-Mei; Zhu, Yong-Guan

2015-12-15

Arsenic, a ubiquitous toxic substance, exists mainly as inorganic forms in the environment. It is perceived that organoarsenicals can be demethylated and degraded into inorganic arsenic by microorganisms. Few studies have focused on the mechanism of arsenic demethylation in bacteria. Here, we investigated arsenic demethylation in a typical freshwater cyanobacterium Nostoc sp. PCC 7120. This bacterium was able to demethylate monomethylarsenite [MAs(III)] rapidly to arsenite [As(III)] and also had the ability to demethylate monomethylarsenate [MAs(V)] to As(III). The NsarsI encoding a C·As lyase responsible for MAs(III) demethylation was cloned from Nostoc sp. PCC 7120 and heterologously expressed in an As-hypersensitive strain Escherichia coli AW3110 (ΔarsRBC). Expression of NsarsI was shown to confer MAs(III) resistance through arsenic demethylation. The purified NsArsI was further identified and functionally characterized in vitro. NsArsI existed mainly as the trimeric state, and the kinetic data were well-fit to the Hill equation with K0.5 = 7.55 ± 0.33 μM for MAs(III), Vmax = 0.79 ± 0.02 μM min(-1), and h = 2.7. Both of the NsArsI truncated derivatives lacking the C-terminal 10 residues (ArsI10) or 23 residues (ArsI23) had a reduced ability of MAs(III) demethylation. These results provide new insights for understanding the important role of cyanobacteria in arsenic biogeochemical cycling in the environment.

Two-step iron(0)-mediated N-demethylation of N-methyl alkaloids.

PubMed

Kok, Gaik B; Pye, Cory C; Singer, Robert D; Scammells, Peter J

2010-07-16

A mild and simple two-step Fe(0)-mediated N-demethylation of a number of tertiary N-methyl alkaloids is described. The tertiary N-methylamine is first oxidized to the corresponding N-oxide, which is isolated as the hydrochloride salt. Subsequent treatment of the N-oxide hydrochloride with iron powder readily provides the N-demethylated amine. Representative substrates include a number of opiate and tropane alkaloids. Key intermediates in the synthesis of semisynthetic 14-hydroxy pharmaceutical opiates such as oxycodone and oxymorphone are also readily N-demethylated using this method.

A non-heme iron-mediated chemical demethylation in DNA and RNA.

PubMed

Yi, Chengqi; Yang, Cai-Guang; He, Chuan

2009-04-21

DNA methylation is arguably one of the most important chemical signals in biology. However, aberrant DNA methylation can lead to cytotoxic or mutagenic consequences. A DNA repair protein in Escherichia coli, AlkB, corrects some of the unwanted methylations of DNA bases by a unique oxidative demethylation in which the methyl carbon is liberated as formaldehyde. The enzyme also repairs exocyclic DNA lesions--that is, derivatives in which the base is augmented with an additional heterocyclic subunit--by a similar mechanism. Two proteins in humans that are homologous to AlkB, ABH2 and ABH3, repair the same spectrum of lesions; another human homologue of AlkB, FTO, is linked to obesity. In this Account, we describe our studies of AlkB, ABH2, and ABH3, including our development of a general strategy to trap homogeneous protein-DNA complexes through active-site disulfide cross-linking. AlkB uses a non-heme mononuclear iron(II) and the cofactors 2-ketoglutarate (2KG) and dioxygen to effect oxidative demethylation of the DNA base lesions 1-methyladenine (1-meA), 3-methylcytosine (3-meC), 1-methylguanine (1-meG), and 3-methylthymine (3-meT). ABH3, like AlkB, works better on single-stranded DNA (ssDNA) and is capable of repairing damaged bases in RNA. Conversely, ABH2 primarily repairs lesions in double-stranded DNA (dsDNA); it is the main housekeeping enzyme that protects the mammalian genome from 1-meA base damage. The AlkB-family proteins have moderate affinities for their substrates and bind DNA in a non-sequence-specific manner. Knowing that these proteins flip the damaged base out from the duplex DNA and insert it into the active site for further processing, we first engineered a disulfide cross-link in the active site to stabilize the Michaelis complex. Based on the detailed structural information afforded by the active-site cross-linked structures, we can readily install a cross-link away from the active site to obtain the native-like structures of these complexes

Early de novo DNA methylation and prolonged demethylation in the muscle lineage.

PubMed

Tsumagari, Koji; Baribault, Carl; Terragni, Jolyon; Varley, Katherine E; Gertz, Jason; Pradhan, Sirharsa; Badoo, Melody; Crain, Charlene M; Song, Lingyun; Crawford, Gregory E; Myers, Richard M; Lacey, Michelle; Ehrlich, Melanie

2013-03-01

Myogenic cell cultures derived from muscle biopsies are excellent models for human cell differentiation. We report the first comprehensive analysis of myogenesis-specific DNA hyper- and hypo-methylation throughout the genome for human muscle progenitor cells (both myoblasts and myotubes) and skeletal muscle tissue vs. 30 non-muscle samples using reduced representation bisulfite sequencing. We also focused on four genes with extensive hyper- or hypo-methylation in the muscle lineage (PAX3, TBX1, MYH7B/MIR499 and OBSCN) to compare DNA methylation, DNaseI hypersensitivity, histone modification, and CTCF binding profiles. We found that myogenic hypermethylation was strongly associated with homeobox or T-box genes and muscle hypomethylation with contractile fiber genes. Nonetheless, there was no simple relationship between differential gene expression and myogenic differential methylation, rather only for subsets of these genes, such as contractile fiber genes. Skeletal muscle retained ~30% of the hypomethylated sites but only ~3% of hypermethylated sites seen in myogenic progenitor cells. By enzymatic assays, skeletal muscle was 2-fold enriched globally in genomic 5-hydroxymethylcytosine (5-hmC) vs. myoblasts or myotubes and was the only sample type enriched in 5-hmC at tested myogenic hypermethylated sites in PAX3/CCDC140 andTBX1. TET1 and TET2 RNAs, which are involved in generation of 5-hmC and DNA demethylation, were strongly upregulated in myoblasts and myotubes. Our findings implicate de novo methylation predominantly before the myoblast stage and demethylation before and after the myotube stage in control of transcription and co-transcriptional RNA processing. They also suggest that, in muscle, TET1 or TET2 are involved in active demethylation and in formation of stable 5-hmC residues.

Early de novo DNA methylation and prolonged demethylation in the muscle lineage

PubMed Central

Tsumagari, Koji; Baribault, Carl; Terragni, Jolyon; Varley, Katherine E.; Gertz, Jason; Pradhan, Sirharsa; Badoo, Melody; Crain, Charlene M.; Song, Lingyun; Crawford, Gregory E.; Myers, Richard M.; Lacey, Michelle; Ehrlich, Melanie

2013-01-01

Myogenic cell cultures derived from muscle biopsies are excellent models for human cell differentiation. We report the first comprehensive analysis of myogenesis-specific DNA hyper- and hypo-methylation throughout the genome for human muscle progenitor cells (both myoblasts and myotubes) and skeletal muscle tissue vs. 30 non-muscle samples using reduced representation bisulfite sequencing. We also focused on four genes with extensive hyper- or hypo-methylation in the muscle lineage (PAX3, TBX1, MYH7B/MIR499 and OBSCN) to compare DNA methylation, DNaseI hypersensitivity, histone modification, and CTCF binding profiles. We found that myogenic hypermethylation was strongly associated with homeobox or T-box genes and muscle hypomethylation with contractile fiber genes. Nonetheless, there was no simple relationship between differential gene expression and myogenic differential methylation, rather only for subsets of these genes, such as contractile fiber genes. Skeletal muscle retained ~30% of the hypomethylated sites but only ~3% of hypermethylated sites seen in myogenic progenitor cells. By enzymatic assays, skeletal muscle was 2-fold enriched globally in genomic 5-hydroxymethylcytosine (5-hmC) vs. myoblasts or myotubes and was the only sample type enriched in 5-hmC at tested myogenic hypermethylated sites in PAX3/CCDC140 andTBX1. TET1 and TET2 RNAs, which are involved in generation of 5-hmC and DNA demethylation, were strongly upregulated in myoblasts and myotubes. Our findings implicate de novo methylation predominantly before the myoblast stage and demethylation before and after the myotube stage in control of transcription and co-transcriptional RNA processing. They also suggest that, in muscle, TET1 or TET2 are involved in active demethylation and in formation of stable 5-hmC residues. PMID:23417056

Impairment of DNA Methylation Maintenance Is the Main Cause of Global Demethylation in Naive Embryonic Stem Cells.

PubMed

von Meyenn, Ferdinand; Iurlaro, Mario; Habibi, Ehsan; Liu, Ning Qing; Salehzadeh-Yazdi, Ali; Santos, Fátima; Petrini, Edoardo; Milagre, Inês; Yu, Miao; Xie, Zhenqing; Kroeze, Leonie I; Nesterova, Tatyana B; Jansen, Joop H; Xie, Hehuang; He, Chuan; Reik, Wolf; Stunnenberg, Hendrik G

2016-06-16

Global demethylation is part of a conserved program of epigenetic reprogramming to naive pluripotency. The transition from primed hypermethylated embryonic stem cells (ESCs) to naive hypomethylated ones (serum-to-2i) is a valuable model system for epigenetic reprogramming. We present a mathematical model, which accurately predicts global DNA demethylation kinetics. Experimentally, we show that the main drivers of global demethylation are neither active mechanisms (Aicda, Tdg, and Tet1-3) nor the reduction of de novo methylation. UHRF1 protein, the essential targeting factor for DNMT1, is reduced upon transition to 2i, and so is recruitment of the maintenance methylation machinery to replication foci. Concurrently, there is global loss of H3K9me2, which is needed for chromatin binding of UHRF1. These mechanisms synergistically enforce global DNA hypomethylation in a replication-coupled fashion. Our observations establish the molecular mechanism for global demethylation in naive ESCs, which has key parallels with those operating in primordial germ cells and early embryos. Crown Copyright © 2016. Published by Elsevier Inc. All rights reserved.

Decaffeination and measurement of caffeine content by addicted Escherichia coli with a refactored N-demethylation operon from Pseudomonas putida CBB5.

PubMed

Quandt, Erik M; Hammerling, Michael J; Summers, Ryan M; Otoupal, Peter B; Slater, Ben; Alnahhas, Razan N; Dasgupta, Aurko; Bachman, James L; Subramanian, Mani V; Barrick, Jeffrey E

2013-06-21

The widespread use of caffeine (1,3,7-trimethylxanthine) and other methylxanthines in beverages and pharmaceuticals has led to significant environmental pollution. We have developed a portable caffeine degradation operon by refactoring the alkylxanthine degradation (Alx) gene cluster from Pseudomonas putida CBB5 to function in Escherichia coli. In the process, we discovered that adding a glutathione S-transferase from Janthinobacterium sp. Marseille was necessary to achieve N 7 -demethylation activity. E. coli cells with the synthetic operon degrade caffeine to the guanine precursor, xanthine. Cells deficient in de novo guanine biosynthesis that contain the refactored operon are ″addicted″ to caffeine: their growth density is limited by the availability of caffeine or other xanthines. We show that the addicted strain can be used as a biosensor to measure the caffeine content of common beverages. The synthetic N-demethylation operon could be useful for reclaiming nutrient-rich byproducts of coffee bean processing and for the cost-effective bioproduction of methylxanthine drugs.

Lidocaine and ropivacaine, but not bupivacaine, demethylate deoxyribonucleic acid in breast cancer cells in vitro.

PubMed

Lirk, P; Hollmann, M W; Fleischer, M; Weber, N C; Fiegl, H

2014-07-01

Lidocaine demethylates deoxyribonucleic acid (DNA) in breast cancer cells. This modification of epigenetic information may be of therapeutic relevance in the perioperative period, because a decrease in methylation can reactivate tumour suppressor genes and inhibit tumour growth. The objectives of this study were to determine the effect of two amide local anaesthetics, ropivacaine and bupivacaine, on methylation in two breast cancer cell lines and to detect whether the combination of lidocaine with the chemotherapy agent 5-aza-2'-deoxycytidine (DAC) would result in additive demethylating effects. Breast cancer cell lines BT-20 [oestrogen receptor (ER)-negative] and MCF-7 (ER-positive) were incubated with lidocaine, bupivacaine, and ropivacaine to assess demethylating properties. Then, we tested varying concentrations of lidocaine and DAC to assess whether their demethylating effects were additive. Cell numbers and global methylation status were analysed. Lidocaine decreased methylation in BT-20 and MCF-7 cells, ropivacaine decreased methylation in BT-20 cells, and bupivacaine had no demethylating effect. When combined, lidocaine and DAC had additive demethylating effects. At clinically relevant doses, lidocaine and ropivacaine exert demethylating effects on specific breast cancer cell lines, but bupivacaine does not. The demethylating effects of lidocaine and DAC are indeed additive. © The Author [2014]. Published by Oxford University Press on behalf of the British Journal of Anaesthesia. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Chemical groups and structural characterization of lignin via thiol-mediated demethylation

Treesearch

Lihong Hu; Hui Pan; Yonghong Zhou; Chung-Yun Hse; Chengguo Liu; Baofang Zhang; Bin Xu

2014-01-01

A new approach to increase the reactivity of lignin by thiol-mediated demethylation was investigated in this study. Demethylated lignin was characterized by the changes in its hydroxyl and methoxyl groups, molecular weight, and other properties using titration and spectroscopy methods including FT-IR, 1H NMR, UV,and GPC. The total...

Demethylating Agents in the Treatment of Cancer

PubMed Central

Howell, Paul M.; Liu, Zixing; Khong, Hung T.

2010-01-01

Gene silencing resulting from aberrant DNA methylation can lead to tumorigenesis. Therefore, drugs that inhibit or interfere with DNA methylation have been used to reactivate and induce silenced gene re-expression in malignancies. Two demethylating agents, azacitidine and decitabine, are approved for the treatment of myelodysplastic syndromes (MDS) by the U.S. Food and Drug Administration (FDA), and are now considered the standard of care in MDS. In this review, we discuss clinical data, including clinical benefits and toxicities, which led to the approval of azacitidine and decitabine. We also summarize findings from clinical trials that used these two demethylating agents in the treatment of solid tumors. Lastly, we discuss some limitations in the use of azacitidine and decitabine in cancer therapy. PMID:27713340

κ chain monoallelic demethylation and the establishment of allelic exclusion

PubMed Central

Mostoslavsky, Raul; Singh, Nandita; Kirillov, Andrei; Pelanda, Roberta; Cedar, Howard; Chess, Andrew; Bergman, Yehudit

1998-01-01

Allelic exclusion in κ light-chain synthesis is thought to result from a feedback mechanism by which the expression of a functional κ light chain on the surface of the B cell leads to an intracellular signal that down-regulates the V(D)J recombinase, thus precluding rearrangement of the other allele. Whereas such a feedback mechanism clearly plays a role in the maintenance of allelic exclusion, here we provide evidence suggesting that the initial establishment of allelic exclusion involves differential availability of the two κ alleles for rearrangement. Analysis of κ+ B-cell populations and of individual κ+ B cells that have rearranged only one allele demonstrates that in these cells, critical sites on the rearranged allele are unmethylated, whereas the nonrearranged allele remains methylated. This pattern is apparently generated by demethylation that is initiated at the small pre-B cell stage, on a single allele, in a process that occurs prior to rearrangement and requires the presence in cis of both the intronic and 3′ κ enhancers. Taken together with data demonstrating that undermethylation is required for rearrangement, these results indicate that demethylation may actually underly the process of allelic exclusion by directing the initial choice of a single κ allele for rearrangement. PMID:9637682

Enhanced photo(geno)toxicity of demethylated chlorpromazine metabolites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Palumbo, Fabrizio

Chlorpromazine (CPZ) is an anti-psychotic drug widely used to treat disorders such as schizophrenia or manic-depression. Unfortunately, CPZ exhibits undesirable side effects such as phototoxic and photoallergic reactions in humans. In general, the influence of drug metabolism on this type of reactions has not been previously considered in photosafety testing. Thus, the present work aims to investigate the possible photo(geno)toxic potential of drug metabolites, using CPZ as an established reference compound. In this case, the metabolites selected for the study are demethylchlorpromazine (DMCPZ), didemethylchlorpromazine (DDMCPZ) and chlorpromazine sulfoxide (CPZSO). The demethylated CPZ metabolites DMCPZ and DDMCPZ maintain identical chromophore tomore » the parent drug. In this work, it has been found that the nature of the aminoalkyl side chain modulates the hydrophobicity and the photochemical properties (for instance, the excited state lifetimes), but it does not change the photoreactivity pattern, which is characterized by reductive photodehalogenation, triggered by homolytic carbon-chlorine bond cleavage with formation of highly reactive aryl radical intermediates. Accordingly, these metabolites are phototoxic to cells, as revealed by the 3T3 NRU assay; their photo-irritation factors are even higher than that of CPZ. The same trend is observed in photogenotoxicity studies, both with isolated and with cellular DNA, where DMCPZ and DDMCPZ are more active than CPZ itself. In summary, side-chain demethylation of CPZ, as a consequence of Phase I biotransformation, does not result a photodetoxification. Instead, it leads to metabolites that exhibit in an even enhanced photo(geno)toxicity. - Highlights: • Demethylated CPZ metabolites are phototoxic to cells, as revealed by the NRU assay. • Single cell electrophoresis (Comet Assay) confirms the photodamage to cellular DNA. • DNA single strand breaks formation is observed on agarose gel electrophoresis. �

CNS germinomas are characterized by global demethylation, chromosomal instability and mutational activation of the Kit-, Ras/Raf/Erk- and Akt-pathways

PubMed Central

Schulte, Simone Laura; Waha, Andreas; Steiger, Barbara; Denkhaus, Dorota; Dörner, Evelyn; Calaminus, Gabriele; Leuschner, Ivo; Pietsch, Torsten

2016-01-01

CNS germinomas represent a unique germ cell tumor entity characterized by undifferentiated tumor cells and a high response rate to current treatment protocols. Limited information is available on their underlying genomic, epigenetic and biological alterations. We performed a genome-wide analysis of genomic copy number alterations in 49 CNS germinomas by molecular inversion profiling. In addition, CpG dinucleotide methylation was studied by immunohistochemistry for methylated cytosine residues. Mutational analysis was performed by resequencing of candidate genes including KIT and RAS family members. Ras/Erk and Akt pathway activation was analyzed by immunostaining with antibodies against phospho-Erk, phosho-Akt, phospho-mTOR and phospho-S6. All germinomas coexpressed Oct4 and Kit but showed an extensive global DNA demethylation compared to other tumors and normal tissues. Molecular inversion profiling showed predominant genomic instability in all tumors with a high frequency of regional gains and losses including high level gene amplifications. Activating mutations of KIT exons 11, 13, and 17 as well as a case with genomic KIT amplification and activating mutations or amplifications of RAS gene family members including KRAS, NRAS and RRAS2 indicated mutational activation of crucial signaling pathways. Co-activation of Ras/Erk and Akt pathways was present in 83% of germinomas. These data suggest that CNS germinoma cells display a demethylated nuclear DNA similar to primordial germ cells in early development. This finding has a striking coincidence with extensive genomic instability. In addition, mutational activation of Kit-, Ras/Raf/Erk- and Akt- pathways indicate the biological importance of these pathways and their components as potential targets for therapy. PMID:27391150

Hepatic microsomal N-oxidation and N-demethylation of N,N-dimethylaniline in red-winged blackbird compared with rat and other birds

USGS Publications Warehouse

Pan, H.P.; Fouts, J.R.; Devereux, T.R.

1975-01-01

Hepatic microsomes prepared from red-winged blackbirds and albino rats were incubated with N,N-dimethylaniline (DMA)_in complete incubation mixtures at pH 7.9 and 37?C for 10 min. Formaldehyde and N,N-dimethylaniline--oxide produced from DMA were measured. Redwings were found to have significantly lower N-demethylation activities than rats, and redwings had only marginal or no N-oxidation activities. Hepatic microsomes from redwings did not further metabolize the N-oxide. The N-oxidation and N-demethylation activities of brown-headed cowbirds, common grackles, and starlings were similar to those of redwings.

Mammalian Gcm genes induce Hes5 expression by active DNA demethylation and induce neural stem cells.

PubMed

Hitoshi, Seiji; Ishino, Yugo; Kumar, Akhilesh; Jasmine, Salma; Tanaka, Kenji F; Kondo, Takeshi; Kato, Shigeaki; Hosoya, Toshihiko; Hotta, Yoshiki; Ikenaka, Kazuhiro

2011-07-17

Signaling mediated by Notch receptors is crucial for the development of many organs and the maintenance of various stem cell populations. The activation of Notch signaling is first detectable by the expression of an effector gene, Hes5, in the neuroepithelium of mouse embryos at embryonic day (E) 8.0-8.5, and this activation is indispensable for the generation of neural stem cells. However, the molecular mechanism by which Hes5 expression is initiated in stem-producing cells remains unknown. We found that mammalian Gcm1 and Gcm2 (glial cells missing 1 and 2) are involved in the epigenetic regulation of Hes5 transcription by DNA demethylation independently of DNA replication. Loss of both Gcm genes and subsequent lack of Hes5 upregulation in the neuroepithelium of E7.5-8.5 Gcm1(-/-); Gcm2(-/-) mice resulted in the impaired induction of neural stem cells. Our data suggest that Hes5 expression is serially activated first by Gcms and later by the canonical Notch pathway.

Human in vitro induced T regulatory cells and memory T cells share common demethylation of specific FOXP3 promoter region.

PubMed

Bégin, Philippe; Schulze, Janika; Baron, Udo; Olek, Sven; Bauer, Rebecca N; Passerini, Laura; Baccheta, Rosa; Nadeau, Kari C

2015-01-01

The FOXP3 gene is the master regulator for T regulatory cells and is under tight DNA methylation control at the Treg specific demethylated region (TSDR) in its first intron. This said, methylation of its promoter region, the significance of which is unknown, has also been associated with various immune-related disease states such as asthma, food allergy, auto-immunity and cancer. Here, we used induced T regulatory cells (iTreg) as a target cell population to identify candidate hypomethylated CpG sites in the FOXP3 gene promoter to design a DNA methylation quantitative assay for this region. Three CpG sites at the promoter region showed clear demethylation pattern associated with high FOXP3 expression after activation in presence of TGFβ and were selected as primary targets to design methylation-dependent RT-PCR primers and probes. We then examined the methylation of this 'inducible-promoter-demethylated-region' (IPDR) in various FOXP3+ T cell subsets. Both naïve and memory thymic-derived Treg cells were found to be fully demethylated at both the IPDR and TSDR. Interestingly, in addition to iTregs, both CD25- and CD25(lo) conventional memory CD4+CD45RA- T cells displayed a high fraction of IPDR demethylated cells in absence of TSDR demethylation. This implies that the fraction of memory T cells should be taken in account when interpreting FOXP3 promoter methylation results from clinical studies. This approach, which is available for testing in clinical samples could have diagnostic and prognostic value in patients with immune or auto-inflammatory diseases.

Activation of HIV Transcription by the Viral Tat Protein Requires a Demethylation Step Mediated by Lysine-specific Demethylase 1 (LSD1/KDM1)

PubMed Central

Sakane, Naoki; Kwon, Hye-Sook; Pagans, Sara; Kaehlcke, Katrin; Mizusawa, Yasuhiro; Kamada, Masafumi; Lassen, Kara G.; Chan, Jonathan; Greene, Warner C.; Schnoelzer, Martina; Ott, Melanie

2011-01-01

The essential transactivator function of the HIV Tat protein is regulated by multiple posttranslational modifications. Although individual modifications are well characterized, their crosstalk and dynamics of occurrence during the HIV transcription cycle remain unclear. We examine interactions between two critical modifications within the RNA-binding domain of Tat: monomethylation of lysine 51 (K51) mediated by Set7/9/KMT7, an early event in the Tat transactivation cycle that strengthens the interaction of Tat with TAR RNA, and acetylation of lysine 50 (K50) mediated by p300/KAT3B, a later process that dissociates the complex formed by Tat, TAR RNA and the cyclin T1 subunit of the positive transcription elongation factor b (P-TEFb). We find K51 monomethylation inhibited in synthetic Tat peptides carrying an acetyl group at K50 while acetylation can occur in methylated peptides, albeit at a reduced rate. To examine whether Tat is subject to sequential monomethylation and acetylation in cells, we performed mass spectrometry on immunoprecipitated Tat proteins and generated new modification-specific Tat antibodies against monomethylated/acetylated Tat. No bimodified Tat protein was detected in cells pointing to a demethylation step during the Tat transactivation cycle. We identify lysine-specific demethylase 1 (LSD1/KDM1) as a Tat K51-specific demethylase, which is required for the activation of HIV transcription in latently infected T cells. LSD1/KDM1 and its cofactor CoREST associates with the HIV promoter in vivo and activate Tat transcriptional activity in a K51-dependent manner. In addition, small hairpin RNAs directed against LSD1/KDM1 or inhibition of its activity with the monoamine oxidase inhibitor phenelzine suppresses the activation of HIV transcription in latently infected T cells. Our data support the model that a LSD1/KDM1/CoREST complex, normally known as a transcriptional suppressor, acts as a novel activator of HIV transcription through demethylation

Clinical and biological effects of demethylating agents on solid tumours - A systematic review.

PubMed

Linnekamp, J F; Butter, R; Spijker, R; Medema, J P; van Laarhoven, H W M

2017-03-01

It is assumed that DNA methylation plays a key role in both tumour development and therapy resistance. Demethylating agents have been shown to be effective in the treatment of haematological malignancies. Based on encouraging preclinical results, demethylating agents may also be effective in solid tumours. This systematic review summarizes the evidence of the effect of demethylating agents on clinical response, methylation and the immune system in solid tumours. We conducted a systematic literature search from 1949 to December 2016, according to the PRISMA guidelines. Studies which evaluated treatment with azacitidine, decitabine, guadecitabine, hydralazine, procaine, MG98 and/or zebularine in patients with solid tumours were included. Data on clinical response, effects on methylation and immune response were extracted. Fifty-eight studies were included: in 13 studies complete responses (CR) were observed, 35 studies showed partial responses (PR), 47 studies stable disease (SD) and all studies except two showed progressive disease (PD). Effects on global methylation were observed in 11/15 studies and demethylation/re-expression of tumour specific genes was seen in 15/17 studies. No clear correlation between (de)methylation and clinical response was observed. In 14 studies immune-related responses were reported, such as re-expression of cancer-testis antigens and upregulation of interferon genes. Demethylating agents are able to improve clinical outcome and alter methylation status in patients with solid tumours. Although beneficial effect has been shown in individual patients, overall response is limited. Further research on biomarker predicting therapy efficacy is indicated, particularly in earlier stage and highly methylated tumours. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Evidence for tangeretin O-demethylation by rat and human liver microsomes.

PubMed

Canivenc-Lavier, M C; Brunold, C; Siess, M H; Suschetet, M

1993-03-01

1. Tangeretin, a polymethoxylated flavone, was studied as a substrate for cytochrome P450-catalysed demethylation reactions by rat and human liver microsomes. Evidence has been presented for the production of formaldehyde in the presence of tangeretin and NAD(P)H. Kinetic studies showed a Km value for tangeretin of about 18 microM in both species. 2. The reaction was inhibited by CO, piperonyl butoxide, 7,8-benzoflavone, propyl gallate, aminobenzothiazole and metyrapone. 3. Rats pretreated with classical cytochrome P450 inducers (Aroclor 1254, 3-methylcholanthrene, phenobarbital, dexamethasone and ciprofibrate) or with flavonoids (flavone, flavanone, quercetin and tangeretin) resulted in increased microsomal demethylation of tangeretin after 3-methylcholanthrene and flavone only. Tangeretin did not enhance its own metabolism. 4. Tangeretin interacted with the oxidized form of cytochrome P450 to produce a reverse type I spectrum. 5. Results indicate that tangeretin is metabolized in liver microsomes by an O-demethylation reaction involving cytochrome P450.

FACT complex is required for DNA demethylation at heterochromatin during reproduction in Arabidopsis.

PubMed

Frost, Jennifer M; Kim, M Yvonne; Park, Guen Tae; Hsieh, Ping-Hung; Nakamura, Miyuki; Lin, Samuel J H; Yoo, Hyunjin; Choi, Jaemyung; Ikeda, Yoko; Kinoshita, Tetsu; Choi, Yeonhee; Zilberman, Daniel; Fischer, Robert L

2018-05-15

The DEMETER (DME) DNA glycosylase catalyzes genome-wide DNA demethylation and is required for endosperm genomic imprinting and embryo viability. Targets of DME-mediated DNA demethylation reside in small, euchromatic, AT-rich transposons and at the boundaries of large transposons, but how DME interacts with these diverse chromatin states is unknown. The STRUCTURE SPECIFIC RECOGNITION PROTEIN 1 (SSRP1) subunit of the chromatin remodeler FACT (facilitates chromatin transactions), was previously shown to be involved in the DME-dependent regulation of genomic imprinting in Arabidopsis endosperm. Therefore, to investigate the interaction between DME and chromatin, we focused on the activity of the two FACT subunits, SSRP1 and SUPPRESSOR of TY16 (SPT16), during reproduction in Arabidopsis We found that FACT colocalizes with nuclear DME in vivo, and that DME has two classes of target sites, the first being euchromatic and accessible to DME, but the second, representing over half of DME targets, requiring the action of FACT for DME-mediated DNA demethylation genome-wide. Our results show that the FACT-dependent DME targets are GC-rich heterochromatin domains with high nucleosome occupancy enriched with H3K9me2 and H3K27me1. Further, we demonstrate that heterochromatin-associated linker histone H1 specifically mediates the requirement for FACT at a subset of DME-target loci. Overall, our results demonstrate that FACT is required for DME targeting by facilitating its access to heterochromatin. Copyright © 2018 the Author(s). Published by PNAS.

MES16, a Member of the Methylesterase Protein Family, Specifically Demethylates Fluorescent Chlorophyll Catabolites during Chlorophyll Breakdown in Arabidopsis12[W][OA

PubMed Central

Christ, Bastien; Schelbert, Silvia; Aubry, Sylvain; Süssenbacher, Iris; Müller, Thomas; Kräutler, Bernhard; Hörtensteiner, Stefan

2012-01-01

During leaf senescence, chlorophyll (Chl) is broken down to nonfluorescent chlorophyll catabolites (NCCs). These arise from intermediary fluorescent chlorophyll catabolites (FCCs) by an acid-catalyzed isomerization inside the vacuole. The chemical structures of NCCs from Arabidopsis (Arabidopsis thaliana) indicate the presence of an enzyme activity that demethylates the C132-carboxymethyl group present at the isocyclic ring of Chl. Here, we identified this activity as methylesterase family member 16 (MES16; At4g16690). During senescence, mes16 leaves exhibited a strong ultraviolet-excitable fluorescence, which resulted from large amounts of different FCCs accumulating in the mutants. As confirmed by mass spectrometry, these FCCs had an intact carboxymethyl group, which slowed down their isomerization to respective NCCs. Like a homologous protein cloned from radish (Raphanus sativus) and named pheophorbidase, MES16 catalyzed the demethylation of pheophorbide, an early intermediate of Chl breakdown, in vitro, but MES16 also demethylated an FCC. To determine the in vivo substrate of MES16, we analyzed pheophorbide a oxygenase1 (pao1), which is deficient in pheophorbide catabolism and accumulates pheophorbide in the chloroplast, and a mes16pao1 double mutant. In the pao1 background, we additionally mistargeted MES16 to the chloroplast. Normally, MES16 localizes to the cytosol, as shown by analysis of a MES16-green fluorescent protein fusion. Analysis of the accumulating pigments in these lines revealed that pheophorbide is only accessible for demethylation when MES16 is targeted to the chloroplast. Together, these data demonstrate that MES16 is an integral component of Chl breakdown in Arabidopsis and specifically demethylates Chl catabolites at the level of FCCs in the cytosol. PMID:22147518

Systemic analysis of genome-wide expression profiles identified potential therapeutic targets of demethylation drugs for glioblastoma.

PubMed

Ning, Tongbo; Cui, Hao; Sun, Feng; Zou, Jidian

2017-09-05

Glioblastoma represents one of the most aggressive malignant brain tumors with high morbidity and motility. Demethylation drugs have been developed for its treatment with little efficacy has been observed. The purpose of this study was to screen therapeutic targets of demethylation drugs or bioactive molecules for glioblastoma through systemic bioinformatics analysis. We firstly downloaded genome-wide expression profiles from the Gene Expression Omnibus (GEO) and conducted the primary analysis through R software, mainly including preprocessing of raw microarray data, transformation between probe ID and gene symbol and identification of differential expression genes (DEGs). Secondly, functional enrichment analysis was conducted via the Database for Annotation, Visualization and Integrated Discovery (DAVID) to explore biological processes involved in the development of glioblastoma. Thirdly, we constructed protein-protein interaction (PPI) network of interested genes and conducted cross analysis for multi datasets to obtain potential therapeutic targets for glioblastoma. Finally, we further confirmed the therapeutic targets through real-time RT-PCR. As a result, biological processes that related to cancer development, amino metabolism, immune response and etc. were found to be significantly enriched in genes that differential expression in glioblastoma and regulated by 5'aza-dC. Besides, network and cross analysis identified ACAT2, UFC1 and CYB5R1 as novel therapeutic targets of demethylation drugs which also confirmed by real time RT-PCR. In conclusions, our study identified several biological processes and genes that involved in the development of glioblastoma and regulated by 5'aza-dC, which would be helpful for the treatment of glioblastoma. Copyright © 2017 Elsevier B.V. All rights reserved.

Genistein promotes DNA demethylation of the steroidogenic factor 1 (SF-1) promoter in endometrial stromal cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Matsukura, Hiroshi, E-mail: hmatsukura.epi@mri.tmd.ac.jp; Aisaki, Ken-ichi; Igarashi, Katsuhide

2011-08-26

Highlights: {yields} Genistein (GEN) is a phytoestrogen found in soy products. {yields} GEN demethylated/unsilenced the steroidogenic factor 1 gene in endometrial tissue. {yields} GEN thus altered mRNA expression in uteri of ovariectomized (OVX) mice. {yields} A high-resolution melting assay was used to screen for epigenetic change. {yields} We isolated an endometrial cell clone that was epigenetically modulated by GEN. -- Abstract: It has recently been demonstrated that genistein (GEN), a phytoestrogen in soy products, is an epigenetic modulator in various types of cells; but its effect on endometrium has not yet been determined. We investigated the effects of GEN onmore » mouse uterine cells, in vivo and in vitro. Oral administration of GEN for 1 week induced mild proliferation of the endometrium in ovariectomized (OVX) mice, which was accompanied by the induction of steroidogenic factor 1 (SF-1) gene expression. GEN administration induced demethylation of multiple CpG sites in the SF-1 promoter; these sites are extensively methylated and thus silenced in normal endometrium. The GEN-mediated promoter demethylation occurred predominantly on the luminal side, as opposed to myometrium side, indicating that the epigenetic change was mainly shown in regenerated cells. Primary cultures of endometrial stromal cell colonies were screened for GEN-mediated alterations of DNA methylation by a high-resolution melting (HRM) method. One out of 20 colony-forming cell clones showed GEN-induced demethylation of SF-1. This clone exhibited a high proliferation capacity with continuous colony formation activity through multiple serial clonings. We propose that only a portion of endometrial cells are capable of receiving epigenetic modulation by GEN.« less

Emodin enhances the demethylation by 5-Aza-CdR of pancreatic cancer cell tumor-suppressor genes P16, RASSF1A and ppENK.

PubMed

Pan, Feng-Ping; Zhou, Hong-Kun; Bu, He-Qi; Chen, Zi-Qiang; Zhang, Hao; Xu, Lu-Ping; Tang, Jian; Yu, Qing-Jiang; Chu, Yong-Quan; Pan, Jie; Fei, Yong; Lin, Sheng-Zhang; Liu, Dian-Lei; Chen, Liang

2016-04-01

5-Aza-2'-deoxycytidine (5-Aza-CdR) is currently acknowledged as a demethylation drug, and causes a certain degree of demethylation in a variety of cancer cells, including pancreatic cancer cells. Emodin, a traditional Chinese medicine (TCM), is an effective monomer extracted from rhubarb and has been reported to exhibit antitumor activity in different manners in pancreatic cancer. In the present study, we examined whether emodin caused demethylation and increased the demethylation of three tumor-suppressor genes P16, RASSF1A and ppENK with a high degree of methylation in pancreatic cancer when combined with 5-Aza-CdR. Our research showed that emodin inhibited the growth of pancreatic cancer Panc-1 cells in a dose- and time-dependent manner. Dot-blot results showed that emodin combined with 5-Aza-CdR significantly suppressed the expression of genome 5mC in PANC-1 cells. In order to verify the effect of methylation, methylation-specific PCR (MSP) and bisulfite genomic sequencing PCR (BSP) combined with TA were selected for the cloning and sequencing. Results of MSP and BSP confirmed that emodin caused faint demethylation, and 5-Aza-CdR had a certain degree of demethylation. When emodin was combined with 5-Aza-CdR, the demethylation was more significant. At the same time, fluorescent quantitative PCR and western blot analysis results confirmed that when emodin was combined with 5-Aza-CdR, the expression levels of P16, RASSF1A and ppENK were increased more significantly compared to either treatment alone. In contrast, the expression levels of DNA methyltransferase 1 (DNMT1) and DNMT3a were more significantly reduced with the combination treatment than the control or either agent alone, further proving that emodin in combination with 5-Aza-CdR enhanced the demethylation effect of 5-Aza-CdR by reducing the expression of methyltransferases. In conclusion, the present study confirmed that emodin in combination with 5-Aza-CdR enhanced the demethylation by 5-Aza-CdR of tumor

Emodin enhances the demethylation by 5-Aza-CdR of pancreatic cancer cell tumor-suppressor genes P16, RASSF1A and ppENK

PubMed Central

PAN, FENG-PING; ZHOU, HONG-KUN; BU, HE-QI; CHEN, ZI-QIANG; ZHANG, HAO; XU, LU-PING; TANG, JIAN; YU, QING-JIANG; CHU, YONG-QUAN; PAN, JIE; FEI, YONG; LIN, SHENG-ZHANG; LIU, DIAN-LEI; CHEN, LIANG

2016-01-01

5-Aza-2′-deoxycytidine (5-Aza-CdR) is currently acknowledged as a demethylation drug, and causes a certain degree of demethylation in a variety of cancer cells, including pancreatic cancer cells. Emodin, a traditional Chinese medicine (TCM), is an effective monomer extracted from rhubarb and has been reported to exhibit antitumor activity in different manners in pancreatic cancer. In the present study, we examined whether emodin caused demethylation and increased the demethylation of three tumor-suppressor genes P16, RASSF1A and ppENK with a high degree of methylation in pancreatic cancer when combined with 5-Aza-CdR. Our research showed that emodin inhibited the growth of pancreatic cancer Panc-1 cells in a dose- and time-dependent manner. Dot-blot results showed that emodin combined with 5-Aza-CdR significantly suppressed the expression of genome 5mC in PANC-1 cells. In order to verify the effect of methylation, methylation-specific PCR (MSP) and bisulfite genomic sequencing PCR (BSP) combined with TA were selected for the cloning and sequencing. Results of MSP and BSP confirmed that emodin caused faint demethylation, and 5-Aza-CdR had a certain degree of demethylation. When emodin was combined with 5-Aza-CdR, the demethylation was more significant. At the same time, fluorescent quantitative PCR and western blot analysis results confirmed that when emodin was combined with 5-Aza-CdR, the expression levels of P16, RASSF1A and ppENK were increased more significantly compared to either treatment alone. In contrast, the expression levels of DNA methyltransferase 1 (DNMT1) and DNMT3a were more significantly reduced with the combination treatment than the control or either agent alone, further proving that emodin in combination with 5-Aza-CdR enhanced the demethylation effect of 5-Aza-CdR by reducing the expression of meth-yltransferases. In conclusion, the present study confirmed that emodin in combination with 5-Aza-CdR enhanced the demethylation by 5-Aza-CdR of

Quaternary ammonium oxidative demethylation: X-ray crystallographic, resonance Raman, and UV-visible spectroscopic analysis of a Rieske-type demethylase.

PubMed

Daughtry, Kelly D; Xiao, Youli; Stoner-Ma, Deborah; Cho, Eunsun; Orville, Allen M; Liu, Pinghua; Allen, Karen N

2012-02-08

Herein, the structure resulting from in situ turnover in a chemically challenging quaternary ammonium oxidative demethylation reaction was captured via crystallographic analysis and analyzed via single-crystal spectroscopy. Crystal structures were determined for the Rieske-type monooxygenase, stachydrine demethylase, in the unliganded state (at 1.6 Å resolution) and in the product complex (at 2.2 Å resolution). The ligand complex was obtained from enzyme aerobically cocrystallized with the substrate stachydrine (N,N-dimethylproline). The ligand electron density in the complex was interpreted as proline, generated within the active site at 100 K by the absorption of X-ray photon energy and two consecutive demethylation cycles. The oxidation state of the Rieske iron-sulfur cluster was characterized by UV-visible spectroscopy throughout X-ray data collection in conjunction with resonance Raman spectra collected before and after diffraction data. Shifts in the absorption band wavelength and intensity as a function of absorbed X-ray dose demonstrated that the Rieske center was reduced by solvated electrons generated by X-ray photons; the kinetics of the reduction process differed dramatically for the liganded complex compared to unliganded demethylase, which may correspond to the observed turnover in the crystal.

Dioxin induces Ahr-dependent robust DNA demethylation of the Cyp1a1 promoter via Tdg in the mouse liver.

PubMed

Amenya, Hesbon Z; Tohyama, Chiharu; Ohsako, Seiichiroh

2016-10-07

The aryl hydrocarbon receptor (Ahr) is a highly conserved nuclear receptor that plays an important role in the manifestation of toxicity induced by polycyclic aromatic hydrocarbons. As a xenobiotic sensor, Ahr is involved in chemical biotransformation through activation of drug metabolizing enzymes. The activated Ahr cooperates with coactivator complexes to induce epigenetic modifications at target genes. Thus, it is conceivable that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a potent Ahr ligand, may elicit robust epigenetic changes in vivo at the Ahr target gene cytochrome P450 1a1 (Cyp1a1). A single dose of TCDD administered to adult mice induced Ahr-dependent CpG hypomethylation, changes in histone modifications, and thymine DNA glycosylase (Tdg) recruitment at the Cyp1a1 promoter in the liver within 24 hrs. These epigenetic changes persisted until 40 days post-TCDD treatment and there was Cyp1a1 mRNA hyperinduction upon repeat administration of TCDD at this time-point. Our demethylation assay using siRNA knockdown and an in vitro methylated plasmid showed that Ahr, Tdg, and the ten-eleven translocation methyldioxygenases Tet2 and Tet3 are required for the TCDD-induced DNA demethylation. These results provide novel evidence of Ahr-driven active DNA demethylation and epigenetic memory. The epigenetic alterations influence response to subsequent chemical exposure and imply an adaptive mechanism to xenobiotic stress.

Dioxin induces Ahr-dependent robust DNA demethylation of the Cyp1a1 promoter via Tdg in the mouse liver

NASA Astrophysics Data System (ADS)

Amenya, Hesbon Z.; Tohyama, Chiharu; Ohsako, Seiichiroh

2016-10-01

The aryl hydrocarbon receptor (Ahr) is a highly conserved nuclear receptor that plays an important role in the manifestation of toxicity induced by polycyclic aromatic hydrocarbons. As a xenobiotic sensor, Ahr is involved in chemical biotransformation through activation of drug metabolizing enzymes. The activated Ahr cooperates with coactivator complexes to induce epigenetic modifications at target genes. Thus, it is conceivable that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a potent Ahr ligand, may elicit robust epigenetic changes in vivo at the Ahr target gene cytochrome P450 1a1 (Cyp1a1). A single dose of TCDD administered to adult mice induced Ahr-dependent CpG hypomethylation, changes in histone modifications, and thymine DNA glycosylase (Tdg) recruitment at the Cyp1a1 promoter in the liver within 24 hrs. These epigenetic changes persisted until 40 days post-TCDD treatment and there was Cyp1a1 mRNA hyperinduction upon repeat administration of TCDD at this time-point. Our demethylation assay using siRNA knockdown and an in vitro methylated plasmid showed that Ahr, Tdg, and the ten-eleven translocation methyldioxygenases Tet2 and Tet3 are required for the TCDD-induced DNA demethylation. These results provide novel evidence of Ahr-driven active DNA demethylation and epigenetic memory. The epigenetic alterations influence response to subsequent chemical exposure and imply an adaptive mechanism to xenobiotic stress.

Uptake and Metabolism of Antibiotics Roseoflavin and 8-Demethyl-8-Aminoriboflavin in Riboflavin-Auxotrophic Listeria monocytogenes.

PubMed

Matern, Andreas; Pedrolli, Danielle; Großhennig, Stephanie; Johansson, Jörgen; Mack, Matthias

2016-12-01

The riboflavin analogs roseoflavin (RoF) and 8-demethyl-8-aminoriboflavin (AF) are produced by the bacteria Streptomyces davawensis and Streptomyces cinnabarinus Riboflavin analogs have the potential to be used as broad-spectrum antibiotics, and we therefore studied the metabolism of riboflavin (vitamin B 2 ), RoF, and AF in the human pathogen Listeria monocytogenes, a bacterium which is a riboflavin auxotroph. We show that the L. monocytogenes protein Lmo1945 is responsible for the uptake of riboflavin, RoF, and AF. Following import, these flavins are phosphorylated/adenylylated by the bifunctional flavokinase/flavin adenine dinucleotide (FAD) synthetase Lmo1329 and adenylylated by the unique FAD synthetase Lmo0728, the first monofunctional FAD synthetase to be described in bacteria. Lmo1329 generates the cofactors flavin mononucleotide (FMN) and FAD, whereas Lmo0728 produces FAD only. The combined activities of Lmo1329 and Lmo0728 are responsible for the intracellular formation of the toxic cofactor analogs roseoflavin mononucleotide (RoFMN), roseoflavin adenine dinucleotide (RoFAD), 8-demethyl-8-aminoriboflavin mononucleotide (AFMN), and 8-demethyl-8-aminoriboflavin adenine dinucleotide (AFAD). In vivo reporter gene assays and in vitro transcription/translation experiments show that the L. monocytogenes FMN riboswitch Rli96, which controls expression of the riboflavin transport gene lmo1945, is negatively affected by riboflavin/FMN and RoF/RoFMN but not by AF/AFMN. Treatment of L. monocytogenes with RoF or AF leads to drastically reduced FMN/FAD levels. We suggest that the reduced flavin cofactor levels in combination with concomitant synthesis of inactive cofactor analogs (RoFMN, RoFAD, AFMN, and AFAD) explain why RoF and AF contribute to antibiotic activity in L. monocytogenes IMPORTANCE: The riboflavin analogs roseoflavin (RoF) and 8-demethyl-8-aminoriboflavin (AF) are small molecules which are produced by Streptomyces davawensis and Streptomyces cinnabarinus

Alterations in microsomal electron transport, oxidative N-demethylation and azo-dye cleavage in carbon tetrachloride and dimethylnitrosamine-induced liver injury

PubMed Central

Smuckler, E. A.; Arrhenius, E.; Hultin, T.

1967-01-01

The effect of administration of carbon tetrachloride and dimethylnitrosamine in vivo on hepatic microsomal function related to drug metabolism was measured. It was found that the capacity of isolated microsomes to demethylate dimethylaniline was diminished during the first hour after carbon tetrachloride poisoning and during the second hour after dimethylnitrosamine poisoning. Thereafter the microsomes from carbon tetrachloride-poisoned livers showed a continuous decline in activity so that at 24hr. there was little residual capacity to undertake demethylation. Microsomes from dimethylnitrosamine-poisoned animals were not different from controls at 24hr. During the first 3hr. there was a transient rise in the accumulation of the N-oxide intermediate in carbon tetrachloride-poisoned livers, with a subsequent fall to below control values. In dimethylnitrosamine poisoning there was a parallel decrease in N-oxide accumulation with decreased demethylation. In the latter part of the first 24hr. the ratio of N-oxide accumulation to demethylation was increased in both instances. At 2hr. after poisoning with either compound there was no evidence of altered NADPH2-dependent neotetrazolium reduction or lipid peroxidation. NADPH2-dependent azo-dye cleavage was decreased. There was no difference in microsomal cytochrome b5 content, but there was a decrease in the amount of cytochrome P-450. This latter change was correlated with the decreased capacity for NADPH2-dependent oxidative demethylation. It is suggested that dimethylnitrosamine is associated with a defect in microsomal NADPH2-dependent electron transport at the level of cytochrome P-450. In addition to affecting cytochrome P-450, carbon tetrachloride is associated with a second severe block involving the release of formaldehyde from the N-oxide intermediate. PMID:6040018

Green synthesis of low-toxicity graphene-fulvic acid with an open band gap enhances demethylation of methylmercury.

PubMed

Hu, Xiangang; Mu, Li; Lu, Kaicheng; Kang, Jia; Zhou, Qixing

2014-06-25

The demethylation of methylmercury has received substantial attention. Here, a novel chemical method for the demethylation of methylmercury is proposed. The low-toxicity graphene-fulvic acid (FA, a ubiquitous material in the environment) was synthesized without the use of a chemical reagent. The hybridized graphene-FA presented an indirect open band gap of 2.25-2.87 eV as well as adequate aqueous dispersion. More importantly, the hybridized graphene-FA exhibited 6- and 10-fold higher photocatalytic efficiencies for the demethylation of methylmercury than FA and free FA with graphene, respectively. This result implies that immobilized, rather than free, FA accelerated the catalysis. Furthermore, inorganic mercuric ion, elemental mercury, and mercuric oxide were identified as the primary demethylation products. For free FA with graphene, graphene quenches the excited-state FA, inhibiting the demethylation by electron transfer. In contrast, the graphene of the self-assembled graphene-FA serves as an electron reservoir, causing electron-hole pair separation. Graphene-FA showed a negligible toxicity toward microalgae compared to graphene. The above results reveal that the green synthesis of graphene and organic molecules is a convenient strategy for obtaining effective cocatalysts.

α7 nAChR mediated Fas demethylation contributes to prenatal nicotine exposure-induced programmed thymocyte apoptosis in mice.

PubMed

Liu, Han-Xiao; Liu, Sha; Qu, Wen; Yan, Hui-Yi; Wen, Xiao; Chen, Ting; Hou, Li-Fang; Ping, Jie

2017-11-07

This study aimed to investigate the effects of prenatal nicotine exposure (PNE) on thymocyte apoptosis and postnatal immune impairments in vivo and further explore the epigenetic mechanisms of the pro-apoptotic effect of nicotine in vitro . The results showed that PNE caused immune impairments in offspring on postnatal day 49, manifested as increased IL-4 production and an increased IgG1/IgG2a ratio in serum. Enhanced apoptosis of total and CD4+SP thymocytes was observed both in fetus and in offspring. Further, by exposing thymocytes to 0-100 μM of nicotine in vitro for 48 h, we found that nicotine increased α7 nicotinic acetylcholine receptor (nAChR) expression, activated the Fas apoptotic pathway, and promoted thymocyte apoptosis in concentration-dependent manners. In addition, nicotine could induce Tet methylcytosine dioxygenase (TET) 2 expression and Fas promoter demethylation, which can be abolished by TET2 siRNA transfection. Moreover, the α7 nAChR specific antagonist α-bungarotoxin can abrogate nicotine-induced TET2 increase, and the following Fas demethylation and Fas-mediated apoptosis. In conclusion, our findings showed, for the first time, that α7 nAChR activation could induce TET2-mediated Fas demethylation in thymocytes and results in the upregulation of Fas apoptotic pathway, which provide evidence for elucidating the PNE-induced programmed thymocyte apoptosis.

Selective DNA demethylation by fusion of TDG with a sequence-specific DNA-binding domain

PubMed Central

Gregory, David J.; Mikhaylova, Lyudmila; Fedulov, Alexey V.

2012-01-01

Our ability to selectively manipulate gene expression by epigenetic means is limited, as there is no approach for targeted reactivation of epigenetically silenced genes, in contrast to what is available for selective gene silencing. We aimed to develop a tool for selective transcriptional activation by DNA demethylation. Here we present evidence that direct targeting of thymine-DNA-glycosylase (TDG) to specific sequences in the DNA can result in local DNA demethylation at potential regulatory sequences and lead to enhanced gene induction. When TDG was fused to a well-characterized DNA-binding domain [the Rel-homology domain (RHD) of NFκB], we observed decreased DNA methylation and increased transcriptional response to unrelated stimulus of inducible nitric oxide synthase (NOS2). The effect was not seen for control genes lacking either RHD-binding sites or high levels of methylation, nor in control mock-transduced cells. Specific reactivation of epigenetically silenced genes may thus be achievable by this approach, which provides a broadly useful strategy to further our exploration of biological mechanisms and to improve control over the epigenome. PMID:22419066

Inhibition of CYP2D6-mediated tramadol O-demethylation in methadone but not buprenorphine maintenance patients.

PubMed

Coller, Janet K; Michalakas, Jennifer R; James, Heather M; Farquharson, Aaron L; Colvill, Joel; White, Jason M; Somogyi, Andrew A

2012-11-01

Management of pain in opioid dependent individuals is problematic due to numerous issues including cross-tolerance to opioids. Hence there is a need to find alternative analgesics to classical opioids and tramadol is potentially one such alternative. Methadone inhibits CYP2D6 in vivo and in vitro. We aimed to investigate the effect of methadone on the pathways of tramadol metabolism: O-demethylation (CYP2D6) to the opioid-active metabolite M1 and N-demethylation (CYP3A4) to M2 in subjects maintained on methadone or buprenorphine as a control. Compared with subjects on buprenorphine, methadone reduced the clearance of tramadol to active O-desmethyl-tramadol (M1) but had no effect on N-desmethyltramadol (M2) formation. Similar to other analgesics whose active metabolites are formed by CYP2D6 such as codeine, reduced formation of O-desmethyltramadol (M1) is likely to result in reduced analgesia for subjects maintained on methadone. Hence alternative analgesics whose metabolism is independent of CYP2D6 should be utilized in this patient population. To compare the O- (CYP2D6 mediated) and N- (CYP3A4 mediated) demethylation metabolism of tramadol between methadone and buprenorphine maintained CYP2D6 extensive metabolizer subjects. METHODS Nine methadone and seven buprenorphine maintained subjects received a single 100 mg dose of tramadol hydrochloride. Blood was collected at 4 h and assayed for tramadol, methadone, buprenorphine and norbuprenorphine (where appropriate) and all urine over 4 h was assayed for tramadol and its M1 and M2 metabolites. The urinary metabolic ratio [median (range)] for O-demethylation (M1) was significantly lower (P= 0.0002, probability score 1.0) in the subjects taking methadone [0.071 (0.012-0.103)] compared with those taking buprenorphine [0.192 (0.108-0.392)], but there was no significant difference (P= 0.21, probability score 0.69) in N-demethylation (M2). The percentage of dose [median (range)] recovered as M1 was significantly lower

Targeted DNA demethylation in human cells by fusion of a plant 5-methylcytosine DNA glycosylase to a sequence-specific DNA binding domain

PubMed Central

Parrilla-Doblas, Jara Teresa; Ariza, Rafael R.; Roldán-Arjona, Teresa

2017-01-01

ABSTRACT DNA methylation is a crucial epigenetic mark associated to gene silencing, and its targeted removal is a major goal of epigenetic editing. In animal cells, DNA demethylation involves iterative 5mC oxidation by TET enzymes followed by replication-dependent dilution and/or replication-independent DNA repair of its oxidized derivatives. In contrast, plants use specific DNA glycosylases that directly excise 5mC and initiate its substitution for unmethylated C in a base excision repair process. In this work, we have fused the catalytic domain of Arabidopsis ROS1 5mC DNA glycosylase (ROS1_CD) to the DNA binding domain of yeast GAL4 (GBD). We show that the resultant GBD-ROS1_CD fusion protein binds specifically a GBD-targeted DNA sequence in vitro. We also found that transient in vivo expression of GBD-ROS1_CD in human cells specifically reactivates transcription of a methylation-silenced reporter gene, and that such reactivation requires both ROS1_CD catalytic activity and GBD binding capacity. Finally, we show that reactivation induced by GBD-ROS1_CD is accompanied by decreased methylation levels at several CpG sites of the targeted promoter. All together, these results show that plant 5mC DNA glycosylases can be used for targeted active DNA demethylation in human cells. PMID:28277978

Redistribution of demethylated RNA helicase A during foot-and-mouth disease virus infection: role of jumonji C-domain containing protein 6 in RHA demethylation

USDA-ARS?s Scientific Manuscript database

We previously reported that RNA Helicase A (RHA) re-localized from the nucleus to the cytoplasm in foot-and-mouth disease virus (FMDV) infected cells, coincident with a reduction in methylation of arginine residues in the RHA C-terminus. To further define the mechanism of RHA demethylation in FMDV-...

Methylation and demethylation of intermediates selenide and methylselenol in the metabolism of selenium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohta, Yuki; Suzuki, Kazuo T.

2008-01-15

All nutritional selenium sources are transformed into the assumed common intermediate selenide for the syntheses of selenoproteins for utilization and/or of selenosugar for excretion. Methylselenol [monomethylselenide, MMSe] is the assumed intermediate leading to other methylated metabolites, dimethylselenide (DMSe) and trimethylselenonium (TMSe) for excretion, and also to the intermediate selenide from methylselenocysteine and methylseleninic acid (MSA). Here, related methylation and demethylation reactions were studied in vitro by providing chemically reactive starting substrates ({sup 76}Se-selenide, {sup 77}Se-MMSe and {sup 82}Se-DMSe) which were prepared in situ by the reduction of the corresponding labeled proximate precursors ({sup 76}Se-selenite, {sup 77}Se-MSA and {sup 82}Se-dimethylselenoxide (DMSeO),more » respectively) with glutathione, the three substrates being incubated simultaneously in rat organ supernatants and homogenates. The resulting chemically labile reaction products were detected simultaneously by speciation analysis with HPLC-ICP-MS after converting the products and un-reacted substrates to the corresponding oxidized derivatives (selenite, MSA and DMSeO). The time-related changes in selenium isotope profiles showed that demethylation of MMSe to selenide was efficient but that of DMSe to MMSe was negligible, whereas methylation of selenide to MMSe, and MMSe to DMSe were efficient, and that of DMSe to TMSe occurred less efficiently. The present methylation and demethylation reactions on equilibrium between selenide, MMSe and DMSe without producing selenosugar and selenoproteins indicated that DMSe rather than TMSe is produced as the end product, suggesting that DMSe is to be excreted more abundantly than TMSe. Organ-dependent differences in the methylation and demethylation reactions were characterized for the liver, kidney and lung.« less

Lidocaine Sensitizes the Cytotoxicity of Cisplatin in Breast Cancer Cells via Up-Regulation of RARβ2 and RASSF1A Demethylation

PubMed Central

Li, Kehan; Yang, Jianxue; Han, Xuechang

2014-01-01

It has been reported that lidocaine is toxic to various types of cells. And a recent study has confirmed that lidocaine exerts a demethylation effect and regulates the proliferation of human breast cancer cell lines. To recognize a potential anti-tumor effect of lidocaine, we evaluated the DNA demethylation by lidocaine in human breast cancer lines, MCF-7 and MDA-MB-231 cells, and determined the influence of demethylation on the toxicity to these cells of cisplatin, which is a commonly utilized anti-tumor agent for breast cancer. Results demonstrated that lidocaine promoted a significant global genomic demethylation, and particularly in the promoters of tumor suppressive genes (TSGs), RARβ2 and RASSF1A. Further, the lidocaine treatment increased cisplatin-induced apoptosis and enhanced cisplatin-induced cytotoxicity. The combined treatment with both lidocaine and cisplatin promoted a significantly higher level of MCF-7 cell apoptosis than singular lidocaine or cisplatin treatment. Moreover, the abrogation of RARβ2 or RASSF1A expression inhibited such apoptosis. In conclusion, the present study confirms the demethylation effect of lidocaine in breast cancer cells, and found that the demethylation of RARβ2 and RASSF1A sensitized the cytotoxicity of cisplatin in breast cancer cells. PMID:25526566

Dietary compound isoliquiritigenin prevents mammary carcinogenesis by inhibiting breast cancer stem cells through WIF1 demethylation.

PubMed

Wang, Neng; Wang, Zhiyu; Wang, Yu; Xie, Xiaoming; Shen, Jiangang; Peng, Cheng; You, Jieshu; Peng, Fu; Tang, Hailin; Guan, Xinyuan; Chen, Jianping

2015-01-01

Breast cancer stem cells (CSCs) are considered as the root of mammary tumorigenesis. Previous studies have demonstrated that ISL efficiently limited the activities of breast CSCs. However, the cancer prevention activities of ISL and its precise molecular mechanisms remain largely unknown. Here, we report a novel function of ISL as a natural demethylation agent targeting WIF1 to prevent breast cancer. ISL administration suppressed in vivo breast cancer initiation and progression, accompanied by reduced CSC-like populations. A global gene expression profile assay further identified WIF1 as the main response gene of ISL treatment, accompanied by the simultaneous downregulation of β-catenin signaling and G0/G1 phase arrest in breast CSCs. In addition, WIF1 inhibition significantly relieved the CSC-limiting effects of ISL and methylation analysis further revealed that ISL enhanced WIF1 gene expression via promoting the demethylation of its promoter, which was closely correlated with the inhibition of DNMT1 methyltransferase. Molecular docking analysis finally revealed that ISL could stably dock into the catalytic domain of DNMT1. Taken together, our findings not only provide preclinical evidence to demonstrate the use of ISL as a dietary supplement to inhibit mammary carcinogenesis but also shed novel light on WIF1 as an epigenetic target for breast cancer prevention.

RNA-dependent chromatin localization of KDM4D lysine demethylase promotes H3K9me3 demethylation

PubMed Central

Zoabi, Muhammad; Nadar-Ponniah, Prathamesh T.; Khoury-Haddad, Hanan; Usaj, Marko; Budowski-Tal, Inbal; Haran, Tali; Henn, Arnon; Mandel-Gutfreund, Yael; Ayoub, Nabieh

2014-01-01

The JmjC-containing lysine demethylase, KDM4D, demethylates di-and tri-methylation of histone H3 on lysine 9 (H3K9me3). How KDM4D is recruited to chromatin and recognizes its histone substrates remains unknown. Here, we show that KDM4D binds RNA independently of its demethylase activity. We mapped two non-canonical RNA binding domains: the first is within the N-terminal spanning amino acids 115 to 236, and the second is within the C-terminal spanning amino acids 348 to 523 of KDM4D. We also demonstrate that RNA interactions with KDM4D N-terminal region are critical for its association with chromatin and subsequently for demethylating H3K9me3 in cells. This study implicates, for the first time, RNA molecules in regulating the levels of H3K9 methylation by affecting KDM4D association with chromatin. PMID:25378304

Genome-wide CpG island methylation and intergenic demethylation propensities vary among different tumor sites.

PubMed

Lee, Seung-Tae; Wiemels, Joseph L

2016-02-18

The epigenetic landscape of cancer includes both focal hypermethylation and broader hypomethylation in a genome-wide manner. By means of a comprehensive genomic analysis on 6637 tissues of 21 tumor types, we here show that the degrees of overall methylation in CpG island (CGI) and demethylation in intergenic regions, defined as 'backbone', largely vary among different tumors. Depending on tumor type, both CGI methylation and backbone demethylation are often associated with clinical, epidemiological and biological features such as age, sex, smoking history, anatomic location, histological type and grade, stage, molecular subtype and biological pathways. We found connections between CGI methylation and hypermutability, microsatellite instability, IDH1 mutation, 19p gain and polycomb features, and backbone demethylation with chromosomal instability, NSD1 and TP53 mutations, 5q and 19p loss and long repressive domains. These broad epigenetic patterns add a new dimension to our understanding of tumor biology and its clinical implications. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Dependence of microsomal methoxyflurane O-demethylation on cytochrome P-450 reductase and the stoichiometry of fluoride ion and formaldehyde release.

PubMed

Waskell, L; Gonzales, J

1982-07-01

In order to characterize further the in vitro liver microsomal O-demethylation and defluorination of the volatile anesthetic methoxyflurane, and obtain additional information regarding the participation of cytochrome P-450 in the oxidation, the stoichiometry of the reaction was determined and the effect of antibody to cytochrome P-450 reductase on this unique biotransformation was examined. Liver microsomes were isolated from rabbits and rats in which enzyme induction had previously been produced by phenobarbital. The O-demethylation of methoxyflurane by phenobarbital-induced microsomes results in the production of 1 mol of formaldehyde for every 2 mol of fluoride ion produced. Dichloroacetic acid is also a product of methoxyflurane O-demethylation. Antibody to cytochrome P-450 reductase inhibits by 85% the amount of fluoride ion produced by the microsomal metabolism of methoxyflurane. Thus critical indirect supportive data are contributed to the hypothesis that at least one, but perhaps more, cytochrome P-450 is indeed responsible for methoxyflurane O-demethylation and defluorination.

Base Excision Repair Facilitates a Functional Relationship Between Guanine Oxidation and Histone Demethylation

PubMed Central

Li, Jianfeng; Braganza, Andrea

2013-01-01

Abstract Significance: Appropriately controlled epigenetic regulation is critical for the normal development and health of an organism. Misregulation of epigenetic control via deoxyribonucleic acid (DNA) methylation or histone methylation has been associated with cancer and chromosomal instability syndromes. Recent Advances: The main function of the proteins in the base excision repair (BER) pathway is to repair DNA single-strand breaks and deamination, oxidation, and alkylation-induced DNA base damage that may result from chemotherapy, environmental exposure, or byproducts of cellular metabolism. Recent studies have suggested that one or more BER proteins may also participate in epigenetic regulation to facilitate gene expression modulation via alteration of the state of DNA methylation or via a reaction coupled to histone modification. BER proteins have also been reported to play an essential role in pluripotent stem cell reprogramming. Critical Issues: One emerging function for BER in epigenetic regulation is the repair of base lesions induced by hydrogen peroxide as a byproduct of lysine-specific demethylase 1 (LSD1) enzymatic activity (LSD1/LSD2-coupled BER) for transcriptional regulation. Future Directions: To shed light on this novel role of BER, this review focuses on the repair of oxidative lesions in nuclear DNA that are induced during LSD1-mediated histone demethylation. Further, we highlight current studies suggesting a role for BER proteins in transcriptional regulation of gene expression via BER-coupled active DNA demethylation in mammalian cells. Such efforts to address the role of BER proteins in epigenetic regulation could broaden cancer therapeutic strategies to include epigenetic modifiers combined with BER inhibitors. Antioxid. Redox Signal. 18, 2429–2443. PMID:23311711

Evaluation of mercury methylation and methylmercury demethylation rates in vegetated and non-vegetated saltmarsh sediments from two Portuguese estuaries.

PubMed

Cesário, Rute; Hintelmann, Holger; Mendes, Ricardo; Eckey, Kevin; Dimock, Brian; Araújo, Beatriz; Mota, Ana Maria; Canário, João

2017-07-01

Neurotoxic methylmercury (MMHg) is formed from inorganic divalent mercury (Hg 2+ ). However, it is poorly understood to what extent different mercury (Hg) pools contribute to existent MMHg levels. In this study, ambient concentrations of total Hg (THg) and MMHg as well as rates of methylation and demethylation were measured simultaneously in sediments with and without salt-marsh plant vegetation, which were collected in Guadiana and Tagus estuaries, Portugal. Concurrent processes of Hg methylation and MMHg demethylation were directly monitored and compared by spiking sediments cores with stable isotope tracers of 199 Hg 2+ and CH 3 201 Hg + followed by gas chromatographic separation and isotope-specific detection using inductively coupled plasma mass spectrometry. Compared to the Guadiana estuary, where concentrations were comparatively low, THg and MMHg levels varied between vegetated and non-vegetated sediments collected at the Rosário site (ROS) of the Tagus estuary. Methylation (K M ) and demethylation rates (K D ) were also different between estuaries being dependent on the presence of vegetation. In addition, the type of macrophyte species influenced K M and K D values. In fact, the highest K M value was found in Sarcocornia fruticosa vegetated sediments at the Castro Marim site in Guadiana (CM, 0.160 day -1 ) and the lowest K M was observed in non-vegetated sediments at the Alcochete site in Tagus (ALC, 0.009 day -1 ). K D varied by a factor of three among sites with highest rates of demethylation observed in non-vegetated sediments in Guadiana (12 ± 1.3 day -1 , corresponding to a half-life of 1.4 ± 0.2 h). This study clearly shows that the presence of vegetation in sediments favors the formation of MMHg. Moreover, this effect might be site specific and further studies are needed to confirm the findings reported here. Copyright © 2017 Elsevier Ltd. All rights reserved.

REPRESSOR OF SILENCING5 Encodes a Member of the Small Heat Shock Protein Family and Is Required for DNA Demethylation in Arabidopsis[C][W

PubMed Central

Zhao, Yusheng; Xie, Shaojun; Li, Xiaojie; Wang, Chunlei; Chen, Zhongzhou; Lai, Jinsheng; Gong, Zhizhong

2014-01-01

In Arabidopsis thaliana, active DNA demethylation is initiated by the DNA glycosylase REPRESSOR OF SILENCING1 (ROS1) and its paralogs DEMETER, DEMETER-LIKE2 (DML2), and DML3. How these demethylation enzymes are regulated, however, is poorly understood. Here, using a transgenic Arabidopsis line harboring the stress-inducible RESPONSIVE TO DEHYDRATION29A (RD29A) promoter–LUCIFERASE (LUC) reporter gene and the cauliflower mosaic virus 35S promoter (35S)–NEOMYCIN PHOSPHOTRANSFERASE II (NPTII) antibiotic resistance marker gene, we characterize a ROS locus, ROS5, that encodes a protein in the small heat shock protein family. ROS5 mutations lead to the silencing of the 35S-NPTII transgene due to DNA hypermethylation but do not affect the expression of the RD29A-LUC transgene. ROS5 physically interacts with the histone acetyltransferase ROS4/INCREASED DNA METHYLATION1 (IDM1) and is required to prevent the DNA hypermethylation of some genes that are also regulated by ROS1 and IDM1. We propose that ROS5 regulates DNA demethylation by interacting with IDM1, thereby creating a chromatin environment that facilitates the binding of ROS1 to erase DNA methylation. PMID:24920332

ERRα protein is stabilized by LSD1 in a demethylation-independent manner.

PubMed

Carnesecchi, Julie; Cerutti, Catherine; Vanacker, Jean-Marc; Forcet, Christelle

2017-01-01

The LSD1 histone demethylase is highly expressed in breast tumors where it constitutes a factor of poor prognosis and promotes traits of cancer aggressiveness such as cell invasiveness. Recent work has shown that the Estrogen-Related Receptor α (ERRα) induces LSD1 to demethylate the Lys 9 of histone H3. This results in the transcriptional activation of a number of common target genes, several of which being involved in cellular invasion. High expression of ERRα protein is also a factor of poor prognosis in breast tumors. Here we show that, independently of its demethylase activities, LSD1 protects ERRα from ubiquitination, resulting in overexpression of the latter protein. Our data also suggests that the elevation of LSD1 mRNA and protein in breast cancer (as compared to normal tissue) may be a key event to increase ERRα protein, independently of its corresponding mRNA.

ERRα protein is stabilized by LSD1 in a demethylation-independent manner

PubMed Central

Carnesecchi, Julie; Cerutti, Catherine; Vanacker, Jean-Marc

2017-01-01

The LSD1 histone demethylase is highly expressed in breast tumors where it constitutes a factor of poor prognosis and promotes traits of cancer aggressiveness such as cell invasiveness. Recent work has shown that the Estrogen-Related Receptor α (ERRα) induces LSD1 to demethylate the Lys 9 of histone H3. This results in the transcriptional activation of a number of common target genes, several of which being involved in cellular invasion. High expression of ERRα protein is also a factor of poor prognosis in breast tumors. Here we show that, independently of its demethylase activities, LSD1 protects ERRα from ubiquitination, resulting in overexpression of the latter protein. Our data also suggests that the elevation of LSD1 mRNA and protein in breast cancer (as compared to normal tissue) may be a key event to increase ERRα protein, independently of its corresponding mRNA. PMID:29190800

SET oncoprotein accumulation regulates transcription through DNA demethylation and histone hypoacetylation.

PubMed

Almeida, Luciana O; Neto, Marinaldo P C; Sousa, Lucas O; Tannous, Maryna A; Curti, Carlos; Leopoldino, Andreia M

2017-04-18

Epigenetic modifications are essential in the control of normal cellular processes and cancer development. DNA methylation and histone acetylation are major epigenetic modifications involved in gene transcription and abnormal events driving the oncogenic process. SET protein accumulates in many cancer types, including head and neck squamous cell carcinoma (HNSCC); SET is a member of the INHAT complex that inhibits gene transcription associating with histones and preventing their acetylation. We explored how SET protein accumulation impacts on the regulation of gene expression, focusing on DNA methylation and histone acetylation. DNA methylation profile of 24 tumour suppressors evidenced that SET accumulation decreased DNA methylation in association with loss of 5-methylcytidine, formation of 5-hydroxymethylcytosine and increased TET1 levels, indicating an active DNA demethylation mechanism. However, the expression of some suppressor genes was lowered in cells with high SET levels, suggesting that loss of methylation is not the main mechanism modulating gene expression. SET accumulation also downregulated the expression of 32 genes of a panel of 84 transcription factors, and SET directly interacted with chromatin at the promoter of the downregulated genes, decreasing histone acetylation. Gene expression analysis after cell treatment with 5-aza-2'-deoxycytidine (5-AZA) and Trichostatin A (TSA) revealed that histone acetylation reversed transcription repression promoted by SET. These results suggest a new function for SET in the regulation of chromatin dynamics. In addition, TSA diminished both SET protein levels and SET capability to bind to gene promoter, suggesting that administration of epigenetic modifier agents could be efficient to reverse SET phenotype in cancer.

Black Raspberry-Derived Anthocyanins Demethylate Tumor Suppressor Genes Through the Inhibition of DNMT1 and DNMT3B in Colon Cancer Cells

PubMed Central

Wang, Li-Shu; Kuo, Chieh-Ti; Cho, Seung-Ju; Seguin, Claire; Siddiqui, Jibran; Stoner, Kristen; Weng, Yu-I; Huang, Tim H.-M.; Tichelaar, Jay; Yearsley, Martha; Stoner, Gary D.; Huang, Yi-Wen

2013-01-01

We previously reported that oral administration of black raspberry powder decreased promoter methylation of tumor suppressor genes in tumors from patients with colorectal cancer. The anthocyanins (ACs) in black raspberries are responsible, at least in part, for their cancer-inhibitory effects. In the present study, we asked if ACs are responsible for the demethylation effects observed in colorectal cancers. Three days of treatment of ACs at 0.5, 5, and 25 μg/ml suppressed activity and protein expression of DNMT1 and DNMT3B in HCT116, Caco2 and SW480 cells. Promoters of CDKN2A, and SFRP2, SFRP5, and WIF1, upstream of Wnt pathway, were demethylated by ACs. mRNA expression of some of these genes was increased. mRNA expression of β-catenin and c-Myc, downstream of Wnt pathway, and cell proliferation were decreased; apoptosis was increased. ACs were taken up into HCT116 cells and were differentially localized with DNMT1 and DNMT3B in the same cells visualized using confocal laser scanning microscopy. Although it was reported that DNMT3B is regulated by c-Myc in mouse lymphoma, DNMT3B did not bind with c-Myc in HCT116 cells. In conclusion, our results suggest that ACs are responsible, at least in part, for the demethylation effects of whole black raspberries in colorectal cancers. PMID:23368921

A plasmaless, photochemical etch process for porous organosilicate glass films

NASA Astrophysics Data System (ADS)

Ryan, E. Todd; Molis, Steven E.

2017-12-01

A plasmaless, photochemical etch process using ultraviolet (UV) light in the presence of NH3 or O2 etched porous organosilicate glass films, also called pSiCOH films, in a two-step process. First, a UV/NH3 or UV/O2 treatment removed carbon (mostly methyl groups bonded to silicon) from a pSiCOH film by demethylation to a depth determined by the treatment exposure time. Second, aqueous HF was used to selectively remove the demethylated layer of the pSiCOH film leaving the methylated layer below. UV in the presence of inert gas or H2 did not demethylate the pSiCOH film. The depth of UV/NH3 demethylation followed diffusion limited kinetics and possible mechanisms of demethylation are presented. Unlike reactive plasma processes, which contain ions that can damage surrounding structures during nanofabrication, the photochemical etch contains no damaging ions. Feasibility of the photochemical etching was shown by comparing it to a plasma-based process to remove the pSiCOH dielectric from between Cu interconnect lines, which is a critical step during air gap fabrication. The findings also expand our understanding of UV photon interactions in pSiCOH films that may contribute to plasma-induced damage to pSiCOH films.

Global identification of genes regulated by estrogen signaling and demethylation in MCF-7 breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Putnik, Milica, E-mail: milica.putnik@ki.se; Zhao, Chunyan, E-mail: chunyan.zhao@ki.se; Gustafsson, Jan-Ake, E-mail: jan-ake.gustafsson@ki.se

Highlights: Black-Right-Pointing-Pointer Estrogen signaling and demethylation can both control gene expression in breast cancers. Black-Right-Pointing-Pointer Cross-talk between these mechanisms is investigated in human MCF-7 breast cancer cells. Black-Right-Pointing-Pointer 137 genes are influenced by both 17{beta}-estradiol and demethylating agent 5-aza-2 Prime -deoxycytidine. Black-Right-Pointing-Pointer A set of genes is identified as targets of both estrogen signaling and demethylation. Black-Right-Pointing-Pointer There is no direct molecular interplay of mediators of estrogen and epigenetic signaling. -- Abstract: Estrogen signaling and epigenetic modifications, in particular DNA methylation, are involved in regulation of gene expression in breast cancers. Here we investigated a potential regulatory cross-talk between thesemore » two pathways by identifying their common target genes and exploring underlying molecular mechanisms in human MCF-7 breast cancer cells. Gene expression profiling revealed that the expression of approximately 140 genes was influenced by both 17{beta}-estradiol (E2) and a demethylating agent 5-aza-2 Prime -deoxycytidine (DAC). Gene ontology (GO) analysis suggests that these genes are involved in intracellular signaling cascades, regulation of cell proliferation and apoptosis. Based on previously reported association with breast cancer, estrogen signaling and/or DNA methylation, CpG island prediction and GO analysis, we selected six genes (BTG3, FHL2, PMAIP1, BTG2, CDKN1A and TGFB2) for further analysis. Tamoxifen reverses the effect of E2 on the expression of all selected genes, suggesting that they are direct targets of estrogen receptor. Furthermore, DAC treatment reactivates the expression of all selected genes in a dose-dependent manner. Promoter CpG island methylation status analysis revealed that only the promoters of BTG3 and FHL2 genes are methylated, with DAC inducing demethylation, suggesting DNA methylation directs

Stable knockdown of PASG enhances DNA demethylation but does not accelerate cellular senescence in TIG-7 human fibroblasts

PubMed Central

Suzuki, Toshikazu; Farrar, Jason E.; Yegnasubramanian, Srinivasan; Zahed, Muhammed; Suzuki, Nobuo; Arceci, Robert J.

2009-01-01

Demethylation of 5-methylcytosine in genomic DNA is believed to be one of the mechanisms underlying replicative life-span of mammalian cells. Both proliferation associated SNF2-like gene (PASG, also termed Lsh) and DNA methyltransferase 3B (Dnmt3b) knockout mice result in embryonic genomic hypomethylation and a replicative senescent phenotype. However, it is unclear whether gradual demethylation of DNA during somatic cell division is directly involved in senescence. In this study, we retrovirally transduced TIG-7 human fibroblasts with a shRNA against PASG and compared the rate of change in DNA methylation as well as the replicative life-span to control cells under low (3%) and ambient (20%) oxygen. Expression of PASG protein was decreased by approximately 80% compared to control cells following transduction of PASG shRNA gene. The rate of cell growth was the same in both control and PASG-suppressed cells. The rate of demethylation of DNA was significantly increased in PASG-suppressed cells as compared control cells. However, decreased PASG expression did not shorten the replicative life-span of TIG-7 cells. Culture under low oxygen extended the life-span of TIG-7 cells but did not alter the rate of DNA demethylation. While knockout of PASG during development results in genomic hypomethylation and premature senescence, our results show that while downregulation of PASG expression in a somatic cell also leads to DNA hypomethylation, there is no associated senescent phenotype. These results suggest differences in cellular consequences of hypomethylation mediated by PASG during development compared to that in somatic cells. PMID:18948754

Stable knockdown of PASG enhances DNA demethylation but does not accelerate cellular senescence in TIG-7 human fibroblasts.

PubMed

Suzuki, Toshikazu; Farrar, Jason E; Yegnasubramanian, Srinivasan; Zahed, Muhammed; Suzuki, Nobuo; Arceci, Robert J

2008-09-01

Demethylation of 5-methylcytosine in genomic DNA is believed to be one of the mechanisms underlying replicative life-span of mammalian cells. Both proliferation associated SNF2-like gene (PASG, also termed Lsh) and DNA methyltransferase 3B (Dnmt3b) knockout mice result in embryonic genomic hypomethylation and a replicative senescent phenotype. However, it is unclear whether gradual demethylation of DNA during somatic cell division is directly involved in senescence. In this study, we retrovirally transduced TIG-7 human fibroblasts with a shRNA against PASG and compared the rate of change in DNA methylation as well as the replicative life-span to control cells under low (3%) and ambient (20%) oxygen. Expression of PASG protein was decreased by approximately 80% compared to control cells following transduction of PASG shRNA gene. The rate of cell growth was the same in both control and PASG-suppressed cells. The rate of demethylation of DNA was significantly increased in PASG-suppressed cells as compared control cells. However, decreased PASG expression did not shorten the replicative life-span of TIG-7 cells. Culture under low oxygen extended the life-span of TIG-7 cells but did not alter the rate of DNA demethylation. While knockout of PASG during development results in genomic hypomethylation and premature senescence, our results show that while downregulation of PASG expression in a somatic cell also leads to DNA hypomethylation, there is no associated senescent phenotype. These results suggest differences in cellular consequences of hypomethylation mediated by PASG during development compared to that in somatic cells.

Nicotine demethylation in Nicotiana cell suspension cultures: N'-formylnornicotine is not involved.

PubMed

Bartholomeusz, Trixie Ann; Bhogal, Ramneek K; Molinié, Roland; Felpin, François-Xavier; Mathé-Allainmat, Monique; Meier, Anna-Carolin; Dräger, Birgit; Lebreton, Jacques; Roscher, Albrecht; Robins, Richard J; Mesnard, François

2005-10-01

Nicotine or nornicotine enriched with stable isotopes in either the N'-methyl group or the pyrrolidine-N were fed to Nicotiana plumbaginifolia suspension cell cultures that do not form endogenous nicotine. The metabolism of these compounds was investigated by analysing the incorporation of isotope into other alkaloids using gas chromatography-mass spectroscopy (GC-MS). Nicotine metabolism primarily resulted in the accumulation of nornicotine, the N'-demethylation product. In addition, six minor metabolites appeared during the course of nicotine metabolism, four of which were identified as cotinine, myosmine, N'-formylnornicotine and N'-carboethoxynornicotine. While cotinine was formed from [(13)C,(2)H(3)-methyl]nicotine without dilution of label, N'-formylnornicotine was labelled at only about 6% of the level of nicotine and N'-carboethoxynornicotine was unlabelled. Feeding with [1'-(15)N]nornicotine resulted in incorporation without dilution of label into both N'-formylnornicotine and N'-carboethoxynornicotine. This pattern strongly indicates that, while nornicotine and cotinine are derived directly from nicotine, N'-formylnornicotine and N'-carboethoxynornicotine are metabolites of nornicotine. Thus, it is directly demonstrated that N'-formylnornicotine is not an intermediate in nicotine demethylation.

C-H activation in Ir(III) and N-demethylation in Pt(II) complexes with mesoionic carbene ligands: examples of monometallic, homobimetallic and heterobimetallic complexes.

PubMed

Maity, Ramananda; Tichter, Tim; van der Meer, Margarethe; Sarkar, Biprajit

2015-11-14

Mononuclear Pt(II) and the first dinuclear Pt(II) complexes along with a cyclometalated heterobimetallic Ir(III)/Pd(II) complex bearing mesoionic carbene donor ligands are presented starting from the same bis-triazolium salt. The mononuclear Pt(II) complex possesses a free triazole moiety which is generated from the corresponding triazolium salt through an N-demethylation reaction, whereas the mononuclear Ir(III) complex features an unreacted triazolium unit.

Sodium-dependent Vitamin C transporter 2 deficiency impairs myelination and remyelination after injury: Roles of collagen and demethylation.

PubMed

Röhr, Dominik; Halfter, Hartmut; Schulz, Jörg B; Young, Peter; Gess, Burkhard

2017-07-01

Peripheral nerve myelination involves rapid production of tightly bound lipid layers requiring cholesterol biosynthesis and myelin protein expression, but also a collagen-containing extracellular matrix providing mechanical stability. In previous studies, we showed a function of ascorbic acid in peripheral nerve myelination and extracellular matrix formation in adult mice. Here, we sought the mechanism of action of ascorbic acid in peripheral nerve myelination using different paradigms of myelination in vivo and in vitro. We found impaired myelination and reduced collagen expression in Sodium-dependent Vitamin C Transporter 2 heterozygous mice (SVCT2 +/- ) during peripheral nerve development and after peripheral nerve injury. In dorsal root ganglion (DRG) explant cultures, hypo-myelination could be rescued by precoating with different collagen types. The activity of the ascorbic acid-dependent demethylating Ten-eleven-translocation (Tet) enzymes was reduced in ascorbic acid deprived and SVCT2 +/- DRG cultures. Further, in ascorbic acid-deprived DRG cultures, methylation of a CpG island in the collagen alpha1 (IV) and alpha2 (IV) bidirectional promoter region was increased compared to wild-type and ascorbic acid treated controls. Taken together, these results provide further evidence for the function of ascorbic acid in myelination and extracellular matrix formation in peripheral nerves and suggest a putative molecular mechanism of ascorbic acid function in Tet-dependent demethylation of collagen promoters. © 2017 Wiley Periodicals, Inc.

Targeted inhibition of histone H3K27 demethylation is effective in high-risk neuroblastoma.

PubMed

Lochmann, Timothy L; Powell, Krista M; Ham, Jungoh; Floros, Konstantinos V; Heisey, Daniel A R; Kurupi, Richard I J; Calbert, Marissa L; Ghotra, Maninderjit S; Greninger, Patricia; Dozmorov, Mikhail; Gowda, Madhu; Souers, Andrew J; Reynolds, C Patrick; Benes, Cyril H; Faber, Anthony C

2018-05-16

High-risk neuroblastoma is often distinguished by amplification of MYCN and loss of differentiation potential. We performed high-throughput drug screening of epigenetic-targeted therapies across a large and diverse tumor cell line panel and uncovered the hypersensitivity of neuroblastoma cells to GSK-J4, a small-molecule dual inhibitor of lysine 27 of histone 3 (H3K27) demethylases ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX), and histone demethylase Jumonji D3 (JMJD3). Mechanistically, GSK-J4 induced neuroblastoma differentiation and endoplasmic reticulum (ER) stress, with accompanying up-regulation of p53 up-regulated modulator of apoptosis (PUMA) and induction of cell death. Retinoic acid (RA)-resistant neuroblastoma cells were sensitive to GSK-J4. In addition, GSK-J4 was effective at blocking the growth of chemorefractory and patient-derived xenograft models of high-risk neuroblastoma in vivo. Furthermore, GSK-J4 and RA combination increased differentiation and ER stress over GSK-J4 effects and limited the growth of neuroblastomas resistant to either drug alone. In MYCN -amplified neuroblastoma, PUMA induction by GSK-J4 sensitized tumors to the B cell lymphoma 2 (BCL-2) inhibitor venetoclax, demonstrating that epigenetic-targeted therapies and BCL-2 homology domain 3 mimetics can be rationally combined to treat this high-risk subset of neuroblastoma. Therefore, H3K27 demethylation inhibition is a promising therapeutic target to treat high-risk neuroblastoma, and H3K27 demethylation can be part of rational combination therapies to induce robust antineuroblastoma activity. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Tet1 and Tet2 maintain mesenchymal stem cell homeostasis via demethylation of the P2rX7 promoter.

PubMed

Yang, Ruili; Yu, Tingting; Kou, Xiaoxing; Gao, Xiang; Chen, Chider; Liu, Dawei; Zhou, Yanheng; Shi, Songtao

2018-06-01

Ten-eleven translocation (Tet) family-mediated DNA oxidation represents an epigenetic modification capable of converting 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC), which regulates various biological processes. However, it is unknown whether Tet family affects mesenchymal stem cells (MSCs) or the skeletal system. Here we show that depletion of Tet1 and Tet2 results in impaired self-renewal and differentiation of bone marrow MSCs (BMMSCs) and a significant osteopenia phenotype. Tet1 and Tet2 deficiency reduces demethylation of the P2rX7 promoter and downregulates exosome release, leading to intracellular accumulation of miR-297a-5p, miR-297b-5p, and miR-297c-5p. These miRNAs inhibit Runx2 signaling to impair BMMSC function. We show that overexpression of P2rX7 rescues the impaired BMMSCs and osteoporotic phenotype in Tet1 and Tet2 double knockout mice. These results indicate that Tet1 and Tet2 play a critical role in maintaining BMMSC and bone homeostasis through demethylation of P2rX7 to control exosome and miRNA release. This Tet/P2rX7/Runx2 cascade may serve as a target for the development of novel therapies for osteopenia disorders.

FOXP3 Expression in GARP-Transduced Helper T Cells Is Not Associated with FOXP3 TSDR Demethylation

PubMed Central

Kehrmann, Jan; Zeschnigk, Michael; Buer, Jan; Probst-Kepper, Michael

2011-01-01

Aim: Glycoprotein A repetitions predominant (GARP or LRRC32) represents a human regulatory CD4+ CD25hi FOXP3+ T (Treg) cell-specific receptor that controls FOXP3. Ectopic expression of GARP in helper T (Th) cells has been shown to be sufficient for the induction of FOXP3 and generation of a stable regulatory phenotype. Since expression of FOXP3 in Treg cells is epigenetically controlled by a conserved motif, the so-called Treg-specific demethylated region (TSDR), we asked whether GARP-mediated upregulation of FOXP3 in Th cells is similarly accompanied by demethylation of the TSDR. Methods: DNA methylation of the FOXP3 TSDR was analyzed by direct sequencing of polymerase chain reaction (PCR) products from bisulfite-treated genomic DNA. Results: Although GARP-transduced Th cells exhibit constitutive FOXP3 expression and a regulatory phenotype, the FOXP3 TSDR is completely methylated as in naive Th cells. GARP-mediated FOXP3 upregulation in Th cells is not associated with Treg-specific demethylation of the FOXP3 TSDR. Conclusion: Although GARP-engineered Th cells exhibit stable FOXP3 expression and a phenotypic reprogramming towards Treg cells in vitro, these cells do not completely mimic the epigenotype of natural Treg cells. Thus, concepts based on the genetic modification of Th cells as cellular therapies to treat autoimmune diseases or to control transplantation tolerance should be critically tested before any clinical application. PMID:22670117

FOXP3 Expression in GARP-Transduced Helper T Cells Is Not Associated with FOXP3 TSDR Demethylation.

PubMed

Kehrmann, Jan; Zeschnigk, Michael; Buer, Jan; Probst-Kepper, Michael

2011-10-01

AIM: Glycoprotein A repetitions predominant (GARP or LRRC32) represents a human regulatory CD4+ CD25(hi) FOXP3+ T (T(reg)) cell-specific receptor that controls FOXP3. Ectopic expression of GARP in helper T (T(h)) cells has been shown to be sufficient for the induction of FOXP3 and generation of a stable regulatory phenotype. Since expression of FOXP3 in Treg cells is epigenetically controlled by a conserved motif, the so-called T(reg)-specific demethylated region (TSDR), we asked whether GARP-mediated upregulation of FOXP3 in Th cells is similarly accompanied by demethylation of the TSDR. METHODS: DNA methylation of the FOXP3 TSDR was analyzed by direct sequencing of polymerase chain reaction (PCR) products from bisulfite-treated genomic DNA. RESULTS: Although GARP-transduced T(h) cells exhibit constitutive FOXP3 expression and a regulatory phenotype, the FOXP3 TSDR is completely methylated as in naive T(h) cells. GARP-mediated FOXP3 upregulation in T(h) cells is not associated with T(reg)-specific demethylation of the FOXP3 TSDR. CONCLUSION: Although GARP-engineered T(h) cells exhibit stable FOXP3 expression and a phenotypic reprogramming towards T(reg) cells in vitro, these cells do not completely mimic the epigenotype of natural T(reg) cells. Thus, concepts based on the genetic modification of T(h) cells as cellular therapies to treat autoimmune diseases or to control transplantation tolerance should be critically tested before any clinical application.

KDM1A triggers androgen-induced miRNA transcription via H3K4me2 demethylation and DNA oxidation.

PubMed

Yang, Shu; Zhang, Jiyuan; Zhang, Yalong; Wan, Xuechao; Zhang, Congzhe; Huang, Xiaohui; Huang, Wenhua; Pu, Honglei; Pei, Chaohan; Wu, Hai; Huang, Yan; Huang, Shengdong; Li, Yao

2015-06-15

Androgen receptor (AR) is a ligand dependent transcription factor that regulates the transcription of target genes. AR activity is closely involved in the maintenance and progression of prostate cancer. After the binding with androgen, AR moves into nucleus and binds to DNA sequence containing androgen response elements (ARE). Flavin-dependent monoamine oxidase KDM1A is necessary for AR driven transcription while the mechanism remains unclear. The association between androgen-dependent transcription and oxidation was tested through pharmaceutical inhibitions and siRNA knockdown of DNA oxidation repair components in prostate cancer cells. The recruitment of involved proteins and the histone methylation dynamics on ARE region was explored by chromatin immunoprecipitation (ChIP). Oxidation inhibition reduced AR dependent expression of KLK3, TMPRSS2, hsa-miR-125b2, and hsa-miR-133b. And such reduction could be restored by H2 O2 treatment. KDM1A recruitment and H3K4me2 demethylation on ARE regions, which produce H2 O2 , are associated with AR targets transcription. AR targets transcription and coupled oxidation recruit 8-oxoguanine-DNA glycosylase (OGG1) and the nuclease APEX1 to ARE regions. Such recruitment depends on KDM1A, and is necessary for AR targets transcription. Our work underlined the importance of histone demethylation and DNA oxidation/repairing machinery in androgen-dependent transcription. The present finds have implications for research into new druggable targets for prostate cancer relying on the cascade of AR activity regulation. © 2015 Wiley Periodicals, Inc.

TSA-induced DNMT1 down-regulation represses hTERT expression via recruiting CTCF into demethylated core promoter region of hTERT in HCT116.

PubMed

Choi, Jee-Hye; Min, Na Young; Park, Jina; Kim, Jin Hong; Park, Soo Hyun; Ko, Young Jong; Kang, Yoonsung; Moon, Young Joon; Rhee, Sangmyung; Ham, Seung Wook; Park, Ae Ja; Lee, Kwang-Ho

2010-01-01

Trichostatin A (TSA), an inhibitor of histone deacetylase, is a well-known antitumor agent that effectively and selectively induces tumor growth arrest and apoptosis. Recently, it was reported that hTERT is one of the primary targets for TSA-induced apoptosis in cancer cells but the mechanism of which has not yet been elucidated. In the present study, to better understand the epigenetic regulation mechanism responsible for the repression of hTERT by TSA, we examined expression of hTERT in the HCT116 colon cancer cell line after treatment with TSA and performed site-specific CpG methylation analysis of the hTERT promoter. We found that TSA-induced the demethylation of site-specific CpGs on the promoter of hTERT, which was caused by down-regulation of DNA methyltransferase 1 (DNMT1). Among the demethylated region, the 31st-33rd CpGs contained a binding site for CTCF, an inhibitor of hTERT transcription. ChIP analysis revealed that TSA-induced demethylation of the 31st-33rd CpGs promoted CTCF binding on hTERT promoter, leading to repression of hTERT. Taken together, down-regulation of DNMT1 by TSA caused demethylation of a CTCF binding site on the hTERT promoter, the result of which was repression of hTERT via recruitment of CTCF to the promoter. Copyright 2009 Elsevier Inc. All rights reserved.

A DEMETER-like DNA demethylase governs tomato fruit ripening.

PubMed

Liu, Ruie; How-Kit, Alexandre; Stammitti, Linda; Teyssier, Emeline; Rolin, Dominique; Mortain-Bertrand, Anne; Halle, Stefanie; Liu, Mingchun; Kong, Junhua; Wu, Chaoqun; Degraeve-Guibault, Charlotte; Chapman, Natalie H; Maucourt, Mickael; Hodgman, T Charlie; Tost, Jörg; Bouzayen, Mondher; Hong, Yiguo; Seymour, Graham B; Giovannoni, James J; Gallusci, Philippe

2015-08-25

In plants, genomic DNA methylation which contributes to development and stress responses can be actively removed by DEMETER-like DNA demethylases (DMLs). Indeed, in Arabidopsis DMLs are important for maternal imprinting and endosperm demethylation, but only a few studies demonstrate the developmental roles of active DNA demethylation conclusively in this plant. Here, we show a direct cause and effect relationship between active DNA demethylation mainly mediated by the tomato DML, SlDML2, and fruit ripening- an important developmental process unique to plants. RNAi SlDML2 knockdown results in ripening inhibition via hypermethylation and repression of the expression of genes encoding ripening transcription factors and rate-limiting enzymes of key biochemical processes such as carotenoid synthesis. Our data demonstrate that active DNA demethylation is central to the control of ripening in tomato.

A DEMETER-like DNA demethylase governs tomato fruit ripening

PubMed Central

Liu, Ruie; How-Kit, Alexandre; Stammitti, Linda; Teyssier, Emeline; Rolin, Dominique; Mortain-Bertrand, Anne; Halle, Stefanie; Liu, Mingchun; Kong, Junhua; Wu, Chaoqun; Degraeve-Guibault, Charlotte; Chapman, Natalie H.; Maucourt, Mickael; Hodgman, T. Charlie; Tost, Jörg; Bouzayen, Mondher; Hong, Yiguo; Seymour, Graham B.; Giovannoni, James J.; Gallusci, Philippe

2015-01-01

In plants, genomic DNA methylation which contributes to development and stress responses can be actively removed by DEMETER-like DNA demethylases (DMLs). Indeed, in Arabidopsis DMLs are important for maternal imprinting and endosperm demethylation, but only a few studies demonstrate the developmental roles of active DNA demethylation conclusively in this plant. Here, we show a direct cause and effect relationship between active DNA demethylation mainly mediated by the tomato DML, SlDML2, and fruit ripening— an important developmental process unique to plants. RNAi SlDML2 knockdown results in ripening inhibition via hypermethylation and repression of the expression of genes encoding ripening transcription factors and rate-limiting enzymes of key biochemical processes such as carotenoid synthesis. Our data demonstrate that active DNA demethylation is central to the control of ripening in tomato. PMID:26261318

The Influence of Hydroxylation on Maintaining CpG Methylation Patterns: A Hidden Markov Model Approach.

PubMed

Giehr, Pascal; Kyriakopoulos, Charalampos; Ficz, Gabriella; Wolf, Verena; Walter, Jörn

2016-05-01

DNA methylation and demethylation are opposing processes that when in balance create stable patterns of epigenetic memory. The control of DNA methylation pattern formation by replication dependent and independent demethylation processes has been suggested to be influenced by Tet mediated oxidation of 5mC. Several alternative mechanisms have been proposed suggesting that 5hmC influences either replication dependent maintenance of DNA methylation or replication independent processes of active demethylation. Using high resolution hairpin oxidative bisulfite sequencing data, we precisely determine the amount of 5mC and 5hmC and model the contribution of 5hmC to processes of demethylation in mouse ESCs. We develop an extended hidden Markov model capable of accurately describing the regional contribution of 5hmC to demethylation dynamics. Our analysis shows that 5hmC has a strong impact on replication dependent demethylation, mainly by impairing methylation maintenance.

Mitochondrial control through nutritionally regulated global histone H3 lysine-4 demethylation

PubMed Central

Soloveychik, Maria; Xu, Mengshu; Zaslaver, Olga; Lee, Kwanyin; Narula, Ashrut; Jiang, River; Rosebrock, Adam P.; Caudy, Amy A.; Meneghini, Marc D.

2016-01-01

Histone demethylation by Jumonji-family proteins is coupled with the decarboxylation of α-ketoglutarate (αKG) to yield succinate, prompting hypotheses that their activities are responsive to levels of these metabolites in the cell. Consistent with this paradigm we show here that the Saccharomyces cerevisiae Jumonji demethylase Jhd2 opposes the accumulation of H3K4me3 in fermenting cells only when they are nutritionally manipulated to contain an elevated αKG/succinate ratio. We also find that Jhd2 opposes H3K4me3 in respiratory cells that do not exhibit such an elevated αKG/succinate ratio. While jhd2∆ caused only limited gene expression defects in fermenting cells, transcript profiling and physiological measurements show that JHD2 restricts mitochondrial respiratory capacity in cells grown in non-fermentable carbon in an H3K4me-dependent manner. In association with these phenotypes, we find that JHD2 limits yeast proliferative capacity under physiologically challenging conditions as measured by both replicative lifespan and colony growth on non-fermentable carbon. JHD2’s impact on nutrient response may reflect an ancestral role of its gene family in mediating mitochondrial regulation. PMID:27897198

Inhibition of CYP2D6-mediated tramadol O-demethylation in methadone but not buprenorphine maintenance patients

PubMed Central

Coller, Janet K; Michalakas, Jennifer R; James, Heather M; Farquharson, Aaron L; Colvill, Joel; White, Jason M; Somogyi, Andrew A

2012-01-01

AIMS To compare the O- (CYP2D6 mediated) and N- (CYP3A4 mediated) demethylation metabolism of tramadol between methadone and buprenorphine maintained CYP2D6 extensive metabolizer subjects. METHODS Nine methadone and seven buprenorphine maintained subjects received a single 100 mg dose of tramadol hydrochloride. Blood was collected at 4 h and assayed for tramadol, methadone, buprenorphine and norbuprenorphine (where appropriate) and all urine over 4 h was assayed for tramadol and its M1 and M2 metabolites. RESULTS The urinary metabolic ratio [median (range)] for O-demethylation (M1) was significantly lower (P= 0.0002, probability score 1.0) in the subjects taking methadone [0.071 (0.012–0.103)] compared with those taking buprenorphine [0.192 (0.108–0.392)], but there was no significant difference (P= 0.21, probability score 0.69) in N-demethylation (M2). The percentage of dose [median (range)] recovered as M1 was significantly lower in subjects taking methadone compared with buprenorphine (0.069 (0.044–0.093) and 0.126 (0.069–0.187), respectively, P= 0.04, probability score 0.19), M2 was significantly higher in subjects taking methadone compared with buprenorphine (0.048 (0.033–0.085) and 0.033 (0.014–0.049), respectively, P= 0.04, probability score 0.81). Tramadol was similar (0.901 (0.635–1.30) and 0.685 (0.347–1.04), respectively, P= 0.35, probability score 0.65). CONCLUSIONS Methadone inhibited the CYP2D6-mediated metabolism of tramadol to M1. Hence, as the degree of opioid analgesia is largely dependent on M1 formation, methadone maintenance patients may not receive adequate analgesia from oral tramadol. PMID:22369095

Icariin may benefit the mesenchymal stem cells of patients with steroid-associated osteonecrosis by ABCB1-promoter demethylation: a preliminary study.

PubMed

Sun, Z-B; Wang, J-W; Xiao, H; Zhang, Q-S; Kan, W-S; Mo, F-B; Hu, S; Ye, S-N

2015-01-01

In this study, we found out a previously undefined function of icariin which restored the dynamic balance between osteogenic and adipogenic differentiation of mesenchymal stem cells (MSCs) in patients with osteonecrosis of femoral head (ONFH) via ABCB1-promoter demethylation. These findings provided important information regarding potential implication of icariin targeting epigenetic changes for the treatment of steroid -associated ONFH. Here, we investigated whether icariin can also exert a beneficial role in the reactivation of MSCs in the patients with steroid-associated ONFH via ABCB1-promoter demethylation. Bone marrow was collected from the proximal femur in patients with steroid-associated ONFH (n = 20) and patients with new femoral neck fractures (n = 22), and then MSCs were isolated. We investigated cell viability, intracellular reactive oxygen species (ROS) level, mitochondrial membrane potential (MMP), P-glycoprotein (P-gp) activity, the transcript levels of ABCB1 and oxidative stress-related genes, methylation extent at CpG islands of ABCB1 promoter, and osteogenic and adipogenic differentiation ability of MSCs from the femoral neck fractures group and from the steroid-associated ONFH group treated with or without icariin. We observed that MSCs from the steroid-associated ONFH group showed reduced proliferation ability, elevated ROS level, depressed MMP, weakened osteogenesis, and enhanced adipogenesis while low P-gp activity, transcription level of ABCB1, and oxidative stress-related genes as well as aberrant CpG islands hypermethylation of ABCB1 were also noted in steroid-associated ONFH group. Treatment with icariin obviously induced de novo P-gp expression, decreased oxidative stress, and promoted osteogenesis. Icariin may be a potential drug targeting epigenetic changes for the treatment of steroid-associated ONFH.

Induction of Cyclooxygenase-2 Expression by Hepatitis B Virus Depends on Demethylation-associated Recruitment of Transcription Factors to the Promoter

PubMed Central

2011-01-01

Background The hepatitis B virus (HBV) is a major etiological factor of inflammation and damage to the liver resulting in hepatocellular carcinoma. Transcription factors play important roles in the disordered gene expression and liver injury caused by HBV. However, the molecular mechanisms behind this observation have not been defined. Results In this study, we observed that circulating prostaglandin (PGE) 2 synthesis was increased in patients with chronic hepatitis B infection, and detected elevated cyclooxygenase (COX)-2 expression in HBV- and HBx-expressing liver cells. Likewise, the association of HBx with C/EBPβ contributed to the induction of COX-2. The COX-2 promoter was hypomethylated in HBV-positive cells, and specific demethylation of CpG dinucleotides within each of the two NF-AT sites in the COX-2 promoter resulted in the increased binding affinity of NF-AT to the cognate sites in the promoter, followed by increased COX-2 expression and PGE2 accumulation. The DNA methylatransferase DNMT3B played a key role in the methylation of the COX-2 promoter, and its decreased binding to the promoter was responsible for the regional demethylation of CpG sites, and for the increased binding of transcription factors in HBV-positive cells. Conclusion Our results indicate that upregulation of COX-2 by HBV and HBx is mediated by both demethylation events and recruitment of multiple transcription factors binding to the promoter. PMID:21401943

Tissue Inhibitor of Metalloproteinase 1 Expression Associated with Gene Demethylation Confers Anoikis Resistance in Early Phases of Melanocyte Malignant Transformation1

PubMed Central

Ricca, Tatiana I; Liang, Gangning; Suenaga, Ana Paula M; Han, Sang W; Jones, Peter A; Jasiulionis, Miriam G

2009-01-01

Although anoikis resistance has been considered a hallmark of malignant phenotype, the causal relation between neoplastic transformation and anchorage-independent growth remains undefined. We developed an experimental model of murine melanocyte malignant transformation, where a melanocyte lineage (melan-a) was submitted to sequential cycles of anchorage blockade, resulting in progressive morphologic alterations, and malignant transformation. Throughout this process, cells corresponding to premalignant melanocytes and melanoma cell lines were established and show progressive anoikis resistance and increased expression of Timp1. In melan-a melanocytes, Timp1 expression is suppressed by DNA methylation as indicated by its reexpression after 5-aza-2′-deoxycytidine treatment. Methylation-sensitive single-nucleotide primer extension analysis showed increased demethylation in Timp1 in parallel with its expression along malignant transformation. Interestingly, TIMP1 expression has already been related with negative prognosis in some human cancers. Although described as a MMP inhibitor, this protein has been associated with apoptosis resistance in different cell types. Melan-a cells overexpressing Timp1 showed increased survival in suspension but were unable to form tumors in vivo, whereas Timp1-overexpressing melanoma cells showed reduced latency time for tumor appearance and increased metastatic potential. Here, we demonstrated for the first time an increment in Timp1 expression since the early phases of melanocyte malignant transformation, associated to a progressive gene demethylation, which confers anoikis resistance. In this way, Timp1 might be considered as a valued marker for melanocyte malignant transformation. PMID:19956395

Identification of human cytochrome P450 2D6 as major enzyme involved in the O-demethylation of the designer drug p-methoxymethamphetamine.

PubMed

Staack, Roland F; Theobald, Denis S; Paul, Liane D; Springer, Dietmar; Kraemer, Thomas; Maurer, Hans H

2004-04-01

p-Methoxymethamphetamine (PMMA) is a new designer drug, listed in many countries as a controlled substance. Several fatalities have been attributed to the abuse of this designer drug. Previous in vivo studies using Wistar rats had shown that PMMA was metabolized mainly by O-demethylation. The aim of the study presented here was to identify the human hepatic cytochrome P450 (P450) enzymes involved in the biotransformation of PMMA to p-hydroxymethamphetamine. Baculovirus-infected insect cell microsomes, pooled human liver microsomes (pHLMs), and CYP2D6 poor-metabolizer genotype human liver microsomes (PM HLMs) were used for this purpose. Only CYP2D6 catalyzed O-demethylation. The apparent K(m) and V(max) values in baculovirus-infected insect cell microsomes were 4.6 +/- 1.0 microM and 92.0 +/- 3.7 pmol/min/pmol P450, respectively, and 42.0 +/- 4.0 microM and 412.5 +/- 10.8 pmol/min/mg protein in pHLMs. Inhibition studies with 1 microM quinidine showed significant inhibition of the metabolite formation (67.2 +/- 0.6%; p < 0.0001), and comparison of the metabolite formation between pHLMs and PM HLMs revealed significantly lower metabolite formation in the incubations with PM HLMs (87.3 +/- 1.1%; p < 0.0001). According to these studies, CYP2D6 is the major P450 involved in O-demethylation of PMMA.

Analysis of the machinery and intermediates of the 5hmC-mediated DNA demethylation pathway in aging on samples from the MARK-AGE Study.

PubMed

Valentini, Elisabetta; Zampieri, Michele; Malavolta, Marco; Bacalini, Maria Giulia; Calabrese, Roberta; Guastafierro, Tiziana; Reale, Anna; Franceschi, Claudio; Hervonen, Antti; Koller, Bernhard; Bernhardt, Jürgen; Slagboom, P Eline; Toussaint, Olivier; Sikora, Ewa; Gonos, Efstathios S; Breusing, Nicolle; Grune, Tilman; Jansen, Eugène; Dollé, Martijn E T; Moreno-Villanueva, María; Sindlinger, Thilo; Bürkle, Alexander; Ciccarone, Fabio; Caiafa, Paola

2016-08-29

Gradual changes in the DNA methylation landscape occur throughout aging virtually in all human tissues. A widespread reduction of 5-methylcytosine (5mC), associated with highly reproducible site-specific hypermethylation, characterizes the genome in aging. Therefore, an equilibrium seems to exist between general and directional deregulating events concerning DNA methylation controllers, which may underpin the age-related epigenetic changes. In this context, 5mC-hydroxylases (TET enzymes) are new potential players. In fact, TETs catalyze the stepwise oxidation of 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), driving the DNA demethylation process based on thymine DNA glycosylase (TDG)-mediated DNA repair pathway. The present paper reports the expression of DNA hydroxymethylation components, the levels of 5hmC and of its derivatives in peripheral blood mononuclear cells of age-stratified donors recruited in several European countries in the context of the EU Project 'MARK-AGE'. The results provide evidence for an age-related decline of TET1 , TET3 and TDG gene expression along with a decrease of 5hmC and an accumulation of 5caC. These associations were independent of confounding variables, including recruitment center, gender and leukocyte composition. The observed impairment of 5hmC-mediated DNA demethylation pathway in blood cells may lead to aberrant transcriptional programs in the elderly.

Inhibition of H3K27me3 Histone Demethylase Activity Prevents the Proliferative Regeneration of Zebrafish Lateral Line Neuromasts

PubMed Central

Bao, Beier; He, Yingzi; Tang, Dongmei; Li, Wenyan; Li, Huawei

2017-01-01

The H3K27 demethylases are involved in a variety of biological processes, including cell differentiation, proliferation, and cell death by regulating transcriptional activity. However, the function of H3K27 demethylation in the field of hearing research is poorly understood. Here, we investigated the role of H3K27me3 histone demethylase activity in hair cell regeneration using an in vivo animal model. Our data showed that pharmacologic inhibition of H3K27 demethylase activity with the specific small-molecule inhibitor GSK-J4 decreased the number of regenerated hair cells in response to neomycin damage. Furthermore, inhibition of H3K27me3 histone demethylase activity dramatically suppressed cell proliferation and activated caspase-3 levels in the regenerating neuromasts of the zebrafish lateral line. GSK-J4 administration also increased the expression of p21 and p27 in neuromast cells and inhibited the ERK signaling pathway. Collectively, our findings indicate that H3K27me3 demethylation is a key epigenetic regulator in the process of hair cell regeneration in zebrafish and suggest that H3K27me3 histone demethylase activity might be a novel therapeutic target for the treatment of hearing loss. PMID:28348517

The DNA methylation profile of activated human natural killer cells.

PubMed

Wiencke, John K; Butler, Rondi; Hsuang, George; Eliot, Melissa; Kim, Stephanie; Sepulveda, Manuel A; Siegel, Derick; Houseman, E Andres; Kelsey, Karl T

2016-05-03

Natural killer (NK) cells are now recognized to exhibit characteristics akin to cells of the adaptive immune system. The generation of adaptive memory is linked to epigenetic reprogramming including alterations in DNA methylation. The study herein found reproducible genome wide DNA methylation changes associated with human NK cell activation. Activation led predominately to CpG hypomethylation (81% of significant loci). Bioinformatics analysis confirmed that non-coding and gene-associated differentially methylated sites (DMS) are enriched for immune related functions (i.e., immune cell activation). Known DNA methylation-regulated immune loci were also identified in activated NK cells (e.g., TNFA, LTA, IL13, CSF2). Twenty-one loci were designated high priority and further investigated as potential markers of NK activation. BHLHE40 was identified as a viable candidate for which a droplet digital PCR assay for demethylation was developed. The assay revealed high demethylation in activated NK cells and low demethylation in naïve NK, T- and B-cells. We conclude the NK cell methylome is plastic with potential for remodeling. The differentially methylated region signature of activated NKs revealed similarities with T cell activation, but also provided unique biomarker candidates of NK activation, which could be useful in epigenome-wide association studies to interrogate the role of NK subtypes in global methylation changes associated with exposures and/or disease states.

Quantitative liquid chromatographic determination of bromadoline and its N-demethylated metabolites in blood, plasma, serum, and urine samples.

PubMed

Peng, G W; Sood, V K; Rykert, U M

1985-03-01

Bromadoline and its two N-demethylated metabolites were extracted into ether:butyl chloride after the addition of internal standard and basification of the various biological fluids (blood, plasma, serum, and urine). These compounds were then extracted into dilute phosphoric acid from the organic phase and separated on a reversed-phase chromatographic system using a mobile phase containing acetonitrile and a buffer of 1,4-dimethylpiperazine and perchloric acid. The overall absolute extraction recoveries of these compounds were approximately 50-80%. The background interferences from the biological fluids were negligible and allowed quantitative determination of bromadoline and the metabolites at levels as low as 2-5 ng/mL. At mobile phase flow rate of 1 mL/min, the sample components and the internal standard were eluted at the retention times within approximately 7-12 min. The drug- and metabolite-to-internal standard peak height ratios showed excellent linear relationships with their corresponding concentrations. The analytical method showed satisfactory within- and between-run assay precision and accuracy, and has been utilized in the simultaneous determination of bromadoline and its two N-demethylated metabolites in biological fluids collected from humans and from dogs after administration of bromadoline maleate.

DNA methylation and the potential role of demethylating agents in prevention of progressive chronic kidney disease.

PubMed

Larkin, Benjamin P; Glastras, Sarah J; Chen, Hui; Pollock, Carol A; Saad, Sonia

2018-04-24

Chronic kidney disease (CKD) is a global epidemic, and its major risk factors include obesity and type 2 diabetes. Obesity not only promotes metabolic dysregulation and the development of diabetic kidney disease but also may independently lead to CKD by a variety of mechanisms, including endocrine and metabolic dysfunction, inflammation, oxidative stress, altered renal hemodynamics, and lipotoxicity. Deleterious renal effects of obesity can also be transmitted from one generation to the next, and it is increasingly recognized that offspring of obese mothers are predisposed to CKD. Epigenetic modifications are changes that regulate gene expression without altering the DNA sequence. Of these, DNA methylation is the most studied. Epigenetic imprints, particularly DNA methylation, are laid down during critical periods of fetal development, and they may provide a mechanism by which maternal-fetal transmission of chronic disease occurs. Our current review explores the evidence for the role of DNA methylation in the development of CKD, diabetic kidney disease, diabetes, and obesity. DNA methylation has been implicated in renal fibrosis-the final pathophysiologic pathway in the development of end-stage kidney disease-which supports the notion that demethylating agents may play a potential therapeutic role in preventing development and progression of CKD.-Larkin, B. P., Glastras, S. J., Chen, H., Pollock, C. A., Saad, S. DNA methylation and the potential role of demethylating agents in prevention of progressive chronic kidney disease.

Design, synthesis, and biological activity of second-generation synthetic oleanane triterpenoids.

PubMed

Fu, Liangfeng; Lin, Qi-Xian; Onyango, Evans O; Liby, Karen T; Sporn, Michael B; Gribble, Gordon W

2017-07-19

We report the synthesis and biological activity of C-24 demethyl CDDO-Me 2 and the C-28 amide derivatives 3 and 4, which are analogues of the anti-inflammatory synthetic triterpenoid bardoxolone methyl (CDDO-Me) 1. Demethylation of the C-24 methyl group was accomplished via "abnormal Beckmann" rearrangement and subsequent ring A reformation. Amides 3 and 4 were found to be potent inhibitors of the production of the inflammatory mediator NO in vitro.

Molecular basis of the activity of the phytopathogen pectin methylesterase

PubMed Central

Fries, Markus; Ihrig, Jessica; Brocklehurst, Keith; Shevchik, Vladimir E; Pickersgill, Richard W

2007-01-01

We provide a mechanism for the activity of pectin methylesterase (PME), the enzyme that catalyses the essential first step in bacterial invasion of plant tissues. The complexes formed in the crystal using specifically methylated pectins, together with kinetic measurements of directed mutants, provide clear insights at atomic resolution into the specificity and the processive action of the Erwinia chrysanthemi enzyme. Product complexes provide additional snapshots along the reaction coordinate. We previously revealed that PME is a novel aspartic-esterase possessing parallel β-helix architecture and now show that the two conserved aspartates are the nucleophile and general acid-base in the mechanism, respectively. Other conserved residues at the catalytic centre are shown to be essential for substrate binding or transition state stabilisation. The preferential binding of methylated sugar residues upstream of the catalytic site, and demethylated residues downstream, drives the enzyme along the pectin molecule and accounts for the sequential pattern of demethylation produced by both bacterial and plant PMEs. PMID:17717531

Arsenic induces functional re-expression of estrogen receptor α by demethylation of DNA in estrogen receptor-negative human breast cancer.

PubMed

Du, Juan; Zhou, Nannan; Liu, Hongxia; Jiang, Fei; Wang, Yubang; Hu, Chunyan; Qi, Hong; Zhong, Caiyun; Wang, Xinru; Li, Zhong

2012-01-01

Estrogen receptor α (ERα) is a marker predictive for response of breast cancers to endocrine therapy. About 30% of breast cancers, however, are hormone- independent because of lack of ERα expression. New strategies are needed for re-expression of ERα and sensitization of ER-negative breast cancer cells to selective ER modulators. The present report shows that arsenic trioxide induces reactivated ERα, providing a target for therapy with ER antagonists. Exposure of ER-negative breast cancer cells to arsenic trioxide leads to re-expression of ERα mRNA and functional ERα protein in in vitro and in vivo. Luciferase reporter gene assays and 3-(4,5-dimethylthiazol-2-yl)- 5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assays show that, upon exposure to arsenic trioxide, formerly unresponsive, ER-negative MDA-MB-231 breast cancer cells become responsive to ER antagonists, 4-hydroxytamoxifen and ICI 182,780. Furthermore, methylation- specific PCR and bisulfite-sequencing PCR assays show that arsenic trioxide induces partial demethylation of the ERα promoter. A methyl donor, S-adenosylmethionine (SAM), reduces the degree of arsenic trioxide-induced re-expression of ERα and demethylation. Moreover, Western blot and ChIP assays show that arsenic trioxide represses expression of DNMT1 and DNMT3a along with partial dissociation of DNMT1 from the ERα promoter. Thus, arsenic trioxide exhibits a previously undefined function which induces re-expression ERα in ER-negative breast cancer cells through demethylation of the ERα promoter. These findings could provide important information regarding the application of therapeutic agents targeting epigenetic changes in breast cancers and potential implication of arsenic trioxide as a new drug for the treatment of ER-negative human breast cancer.

Growth-inducing effects of argon plasma on soybean sprouts via the regulation of demethylation levels of energy metabolism-related genes.

PubMed

Zhang, Jiao Jiao; Jo, Jin Oh; Huynh, Do Luong; Mongre, Raj Kumar; Ghosh, Mrinmoy; Singh, Amit Kumar; Lee, Sang Baek; Mok, Young Sun; Hyuk, Park; Jeong, Dong Kee

2017-02-07

This study was conducted to determine the effects of argon plasma on the growth of soybean [Glycine max (L.) Merr.] sprouts and investigate the regulation mechanism of energy metabolism. The germination and growth characteristics were modified by argon plasma at different potentials and exposure durations. Upon investigation, plasma treatment at 22.1 kV for 12 s maximized the germination and seedling growth of soybean, increasing the concentrations of soluble protein, antioxidant enzymes, and adenosine triphosphate (ATP) as well as up-regulating ATP a1, ATP a2, ATP b1, ATP b2, ATP b3, target of rapamycin (TOR), growth-regulating factor (GRF) 1-6, down-regulating ATP MI25 mRNA expression, and increasing the demethylation levels of the sequenced region of ATP a1, ATP b1, TOR, GRF 5, and GRF 6 of 6-day-old soybean sprouts. These observations indicate that argon plasma promotes soybean seed germination and sprout growth by regulating the demethylation levels of ATP, TOR, and GRF.

Growth-inducing effects of argon plasma on soybean sprouts via the regulation of demethylation levels of energy metabolism-related genes

NASA Astrophysics Data System (ADS)

Zhang, Jiao Jiao; Jo, Jin Oh; Huynh, Do Luong; Mongre, Raj Kumar; Ghosh, Mrinmoy; Singh, Amit Kumar; Lee, Sang Baek; Mok, Young Sun; Hyuk, Park; Jeong, Dong Kee

2017-02-01

This study was conducted to determine the effects of argon plasma on the growth of soybean [Glycine max (L.) Merr.] sprouts and investigate the regulation mechanism of energy metabolism. The germination and growth characteristics were modified by argon plasma at different potentials and exposure durations. Upon investigation, plasma treatment at 22.1 kV for 12 s maximized the germination and seedling growth of soybean, increasing the concentrations of soluble protein, antioxidant enzymes, and adenosine triphosphate (ATP) as well as up-regulating ATP a1, ATP a2, ATP b1, ATP b2, ATP b3, target of rapamycin (TOR), growth-regulating factor (GRF) 1-6, down-regulating ATP MI25 mRNA expression, and increasing the demethylation levels of the sequenced region of ATP a1, ATP b1, TOR, GRF 5, and GRF 6 of 6-day-old soybean sprouts. These observations indicate that argon plasma promotes soybean seed germination and sprout growth by regulating the demethylation levels of ATP, TOR, and GRF.

A DEMETER-like DNA demethylase governs tomato fruit ripening

USDA-ARS?s Scientific Manuscript database

This work shows that active DNA demethylation governs ripening, an important plant developmental process. Our work defines a molecular mechanism, which has until now been missing, to explain the correlation between genomic DNA demethylation and fruit ripening. It demonstrates a direct cause-and-effe...

Analysis of the machinery and intermediates of the 5hmC-mediated DNA demethylation pathway in aging on samples from the MARK-AGE Study

PubMed Central

Valentini, Elisabetta; Zampieri, Michele; Malavolta, Marco; Bacalini, Maria Giulia; Calabrese, Roberta; Guastafierro, Tiziana; Reale, Anna; Franceschi, Claudio; Hervonen, Antti; Koller, Bernhard; Bernhardt, Jürgen; Slagboom, P. Eline; Toussaint, Olivier; Sikora, Ewa; Gonos, Efstathios S.; Breusing, Nicolle; Grune, Tilman; Jansen, Eugène; Dollé, Martijn E.T.; Moreno-Villanueva, María; Sindlinger, Thilo; Bürkle, Alexander; Ciccarone, Fabio; Caiafa, Paola

2016-01-01

Gradual changes in the DNA methylation landscape occur throughout aging virtually in all human tissues. A widespread reduction of 5-methylcytosine (5mC), associated with highly reproducible site-specific hypermethylation, characterizes the genome in aging. Therefore, an equilibrium seems to exist between general and directional deregulating events concerning DNA methylation controllers, which may underpin the age-related epigenetic changes. In this context, 5mC-hydroxylases (TET enzymes) are new potential players. In fact, TETs catalyze the stepwise oxidation of 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), driving the DNA demethylation process based on thymine DNA glycosylase (TDG)-mediated DNA repair pathway. The present paper reports the expression of DNA hydroxymethylation components, the levels of 5hmC and of its derivatives in peripheral blood mononuclear cells of age-stratified donors recruited in several European countries in the context of the EU Project ‘MARK-AGE’. The results provide evidence for an age-related decline of TET1, TET3 and TDG gene expression along with a decrease of 5hmC and an accumulation of 5caC. These associations were independent of confounding variables, including recruitment center, gender and leukocyte composition. The observed impairment of 5hmC-mediated DNA demethylation pathway in blood cells may lead to aberrant transcriptional programs in the elderly. PMID:27587280

An examination of the factors influencing mercury and methylmercury particulate distributions, methylation and demethylation rates in laboratory-generated marine snow.

PubMed

Ortiz, Veronica L; Mason, Robert P; Ward, J Evan

2015-12-20

In the marine environment, settling particulates have been widely studied for their role as effective vertical transporters of nutrients and metals scavenged from the euphotic zone to the benthos. These particulates are composed of transparent exopolymers, plankton and bacterial cells, detritus and organic matter, and form various size fractions from colloids (<0.2μm) to aggregates, and finally marine snow (>300 μm). As marine snow forms in the water column, anoxic layers form around and within the aggregation potentially creating a prime environment for the methylation of mercury (Hg), which occurs primarily in low oxygen environments. To examine this process, marine aggregates were produced from sieved estuarine seawater (100 μm) in 1-L glass bottles spiked with stable isotope enriched methylmercury (CH 3 199 Hg) and inorganic mercury ( 200 Hg(II)) at 18° C using a roller-table. After the rolling period, different particle-size fractions were collected and analyzed, including: visible marine snow (>300μm), particulates 8 to 300 μm, and particulates 0.2 to 8μm. Particulate analysis indicated higher incorporation of both forms of Hg into marine snow compared to unrolled treatments, with greater incorporation of 200 Hg(II) than CH 3 199 Hg. In addition, inorganic Hg was methylated and CH 3 Hg was demethylated in the larger particulate fractions (>8μm). Methylation and demethylation rates were assessed based on changes in isotopic composition of Hg(II) and CH 3 Hg, and found to be comparable to methylation rates found in sediments. These results indicate that net Hg methylation can occur in marine snow and smaller aggregates in oxic coastal waters, and that this net formation of CH 3 Hg may be an important source of CH 3 Hg in both coastal and open ocean surface environments.

The Roles of a Flavone-6-Hydroxylase and 7-O-Demethylation in the Flavone Biosynthetic Network of Sweet Basil*

PubMed Central

Berim, Anna; Gang, David R.

2013-01-01

Lipophilic flavonoids found in the Lamiaceae exhibit unusual 6- and 8-hydroxylations whose enzymatic basis is unknown. We show that crude protein extracts from peltate trichomes of sweet basil (Ocimum basilicum L.) cultivars readily hydroxylate position 6 of 7-O-methylated apigenin but not apigenin itself. The responsible protein was identified as a P450 monooxygenase from the CYP82 family, a family not previously reported to be involved in flavonoid metabolism. This enzyme prefers flavones but also accepts flavanones in vitro and requires a 5-hydroxyl in addition to a 7-methoxyl residue on the substrate. A peppermint (Mentha × piperita L.) homolog displayed identical substrate requirements, suggesting that early 7-O-methylation of flavones might be common in the Lamiaceae. This hypothesis is further substantiated by the pioneering discovery of 2-oxoglutarate-dependent flavone demethylase activity in basil, which explains the accumulation of 7-O-demethylated flavone nevadensin. PMID:23184958

Automated Quantification of DNA Demethylation Effects in Cells via 3D Mapping of Nuclear Signatures and Population Homogeneity Assessment1

PubMed Central

Gertych, Arkadiusz; Wawrowsky, Kolja A.; Lindsley, Erik; Vishnevsky, Eugene; Farkas, Daniel L.; Tajbakhsh, Jian

2009-01-01

Background Today’s advanced microscopic imaging applies to the preclinical stages of drug discovery that employ high-throughput and high-content three-dimensional (3D) analysis of cells to more efficiently screen candidate compounds. Drug efficacy can be assessed by measuring response homogeneity to treatment within a cell population. In this study topologically quantified nuclear patterns of methylated cytosine and global nuclear DNA are utilized as signatures of cellular response to the treatment of cultured cells with the demethylating anti-cancer agents: 5-azacytidine (5-AZA) and octreotide (OCT). Methods Mouse pituitary folliculostellate TtT-GF cells treated with 5-AZA and OCT for 48 hours, and untreated populations, were studied by immunofluorescence with a specific antibody against 5-methylcytosine (MeC), and 4,6-diamidino-2-phenylindole (DAPI) for delineation of methylated sites and global DNA in nuclei (n=163). Cell images were processed utilizing an automated 3D analysis software that we developed by combining seeded watershed segmentation to extract nuclear shells with measurements of Kullback-Leibler’s (K-L) divergence to analyze cell population homogeneity in the relative nuclear distribution patterns of MeC versus DAPI stained sites. Each cell was assigned to one of the four classes: similar, likely similar, unlikely similar and dissimilar. Results Evaluation of the different cell groups revealed a significantly higher number of cells with similar or likely similar MeC/DAPI patterns among untreated cells (~100%), 5-AZA-treated cells (90%), and a lower degree of same type of cells (64%) in the OCT-treated population. The latter group contained (28%) of unlikely similar or dissimilar (7%) cells. Conclusion Our approach was successful in the assessment of cellular behavior relevant to the biological impact of the applied drugs, i.e. the reorganization of MeC/DAPI distribution by demethylation. In a comparison with other metrics, K-L divergence has

Activation-induced cytidine deaminase (AID) is necessary for the epithelial–mesenchymal transition in mammary epithelial cells

PubMed Central

Muñoz, Denise P.; Lee, Elbert L.; Takayama, Sachiko; Coppé, Jean-Philippe; Heo, Seok-Jin; Boffelli, Dario; Di Noia, Javier M.; Martin, David I. K.

2013-01-01

Activation-induced cytidine deaminase (AID), which functions in antibody diversification, is also expressed in a variety of germ and somatic cells. Evidence that AID promotes DNA demethylation in epigenetic reprogramming phenomena, and that it is induced by inflammatory signals, led us to investigate its role in the epithelial–mesenchymal transition (EMT), a critical process in normal morphogenesis and tumor metastasis. We find that expression of AID is induced by inflammatory signals that induce the EMT in nontransformed mammary epithelial cells and in ZR75.1 breast cancer cells. shRNA–mediated knockdown of AID blocks induction of the EMT and prevents cells from acquiring invasive properties. Knockdown of AID suppresses expression of several key EMT transcriptional regulators and is associated with increased methylation of CpG islands proximal to the promoters of these genes; furthermore, the DNA demethylating agent 5 aza-2'deoxycytidine (5-Aza-dC) antagonizes the effects of AID knockdown on the expression of EMT factors. We conclude that AID is necessary for the EMT in this breast cancer cell model and in nontransformed mammary epithelial cells. Our results suggest that AID may act near the apex of a hierarchy of regulatory steps that drive the EMT, and are consistent with this effect being mediated by cytosine demethylation. This evidence links our findings to other reports of a role for AID in epigenetic reprogramming and control of gene expression. PMID:23882083

Mercury methylation and demethylation by periphyton biofilms and their host in a fluvial wetland of the St. Lawrence River (QC, Canada).

PubMed

Hamelin, Stéphanie; Planas, Dolors; Amyot, Marc

2015-04-15

Wetlands in large rivers are important sites of production of the neurotoxin methylmercury (MeHg), and the periphyton growing on wetland macrophytes are increasingly recognized as key players in this production and transfer in food webs. Information is lacking about mercury methylation (Km) and demethylation (Kd) rates in periphytic biofilms from the Northern Hemisphere, as well as about the drivers of net MeHg production, hampering ecosystem modeling of Hg cycling. Mercury methylation and demethylation rates were measured in periphytic biofilms growing on submerged plants in a shallow fluvial lake located in a temperate cold region (St. Lawrence River, Quebec, Canada). Incubations were performed in situ within macrophyte beds using low-level spikes of (199)HgO and Me(200)Hg stable isotopes as tracers. A direct relationship was observed between Km (0.002 to 0.137 d(-1)) and [MeHg] in periphyton. A similar relationship was found between Kd (0.096 to 0.334 d(-1)) and [inorganic Hg]. Periphyton of Lake St. Pierre reached high levels of net MeHg production that were two orders of magnitude higher than those found in local sediment. This production varied through the plant growing season and was mainly driven by environmental variables such as depth of growth, available light, dissolved oxygen, temperature, plant community structure, and productivity of the habitat. Copyright © 2015 Elsevier B.V. All rights reserved.

Automated extraction of lysergic acid diethylamide (LSD) and N-demethyl-LSD from blood, serum, plasma, and urine samples using the Zymark RapidTrace with LC/MS/MS confirmation.

PubMed

de Kanel, J; Vickery, W E; Waldner, B; Monahan, R M; Diamond, F X

1998-05-01

A forensic procedure for the quantitative confirmation of lysergic acid diethylamide (LSD) and the qualitative confirmation of its metabolite, N-demethyl-LSD, in blood, serum, plasma, and urine samples is presented. The Zymark RapidTrace was used to perform fully automated solid-phase extractions of all specimen types. After extract evaporation, confirmations were performed using liquid chromatography (LC) followed by positive electrospray ionization (ESI+) mass spectrometry/mass spectrometry (MS/MS) without derivatization. Quantitation of LSD was accomplished using LSD-d3 as an internal standard. The limit of quantitation (LOQ) for LSD was 0.05 ng/mL. The limit of detection (LOD) for both LSD and N-demethyl-LSD was 0.025 ng/mL. The recovery of LSD was greater than 95% at levels of 0.1 ng/mL and 2.0 ng/mL. For LSD at 1.0 ng/mL, the within-run and between-run (different day) relative standard deviation (RSD) was 2.2% and 4.4%, respectively.

Inhibitors of enzymes catalyzing modifications to histone lysine residues: structure, function and activity.

PubMed

Lillico, Ryan; Stesco, Nicholas; Khorshid Amhad, Tina; Cortes, Claudia; Namaka, Mike P; Lakowski, Ted M

2016-05-01

Gene expression is partly controlled by epigenetic mechanisms including histone-modifying enzymes. Some diseases are caused by changes in gene expression that can be mitigated by inhibiting histone-modifying enzymes. This review covers the enzyme inhibitors targeting histone lysine modifications. We summarize the enzymatic mechanisms of histone lysine acetylation, deacetylation, methylation and demethylation and discuss the biochemical roles of these modifications in gene expression and in disease. We discuss inhibitors of lysine acetylation, deacetylation, methylation and demethylation defining their structure-activity relationships and their potential mechanisms. We show that there are potentially indiscriminant off-target effects on gene expression even with the use of selective epigenetic enzyme inhibitors.

Hydrogen Peroxide-Induced Secreted Frizzled-Related Protein 1 Gene Demethylation Contributes to Hydrogen Peroxide-Induced Apoptosis in Human U251 Glioma Cells.

PubMed

Xing, Zhiguo; Ni, Yaping; Zhao, Junjie; Ma, Xudong

2017-05-01

Glioblastoma multiforme is a type of central nervous system tumor with extremely poor prognosis. Previously, hydrogen peroxide (H 2 O 2 ), which promotes the oxidative stress response, has been reported to induce the apoptosis of glioma cells. Recently, secreted frizzled-related protein 1 (SFRP1) has been shown to be associated with various types of malignant tumors and with H 2 O 2 -induced oxidative stress in cardiomyocytes by negatively regulating the Wnt signaling pathway. This study aimed to explore SFRP1 expression and its roles in H 2 O 2 -induced apoptosis in human glioma cells. We found that the SFRP1 level was decreased in several human glioma cell lines, including U87, U251, and SW1783 cells. In U251 cells, SFRP1 could function as a cancer suppressor gene, and the growth of U251 cells could be inhibited not only by H 2 O 2 but also by the overexpression of SFRP1. Furthermore, we demonstrated that H 2 O 2 -induced SFRP1 gene demethylation partially contributed to H 2 O 2 -induced U251 cell apoptosis, which was verified by studies using an SFRP inhibitor (WAY-316606). Our research identified that H 2 O 2 -induced SFRP1 gene demethylation contributes to H 2 O 2 -induced apoptosis in human U251 glioma cells.

Analysis of acylcarnitines as their N-demethylated ester derivatives by gas chromatography-chemical ionization mass spectrometry.

PubMed

Huang, Z H; Gage, D A; Bieber, L L; Sweeley, C C

1991-11-15

A novel approach to the analysis of acylcarnitines has been developed. It involves a direct esterification using propyl chloroformate in aqueous propanol followed by ion-pair extraction with potassium iodide into chloroform and subsequent on-column N-demethylation of the resulting acylcarnitine propyl ester iodides. The products, acyl N-demethylcarnitine propyl esters, are volatile and are easily analyzed by gas chromatography-chemical ionization mass spectrometry. For medium-chain-length (C4-C12) acylcarnitine standards, detection limits are demonstrated to be well below 1 ng starting material using selected ion monitoring. Well-separated gas chromatographic peaks and structure-specific mass spectra are obtained with samples of synthetic and biological origin. Seven acylcarnitines have been characterized in the urine of a patient suffering from medium-chain acyl-CoA dehydrogenase deficiency.

Methylation of the chicken vitellogenin gene: influence of estradiol administration.

PubMed Central

Meijlink, F C; Philipsen, J N; Gruber, M; Ab, G

1983-01-01

The degree of methylation of the chicken vitellogenin gene has been investigated. Upon induction by administration of estradiol to a rooster, methyl groups at specific sites near the 5'-end of the gene are eliminated. The process of demethylation is slower than the activation of the gene. Demethylation is therefore probably not a prerequisite to gene transcription. At least two other sites in the coding region of the gene are methylated in the liver of estrogenized roosters, but not in the liver of a laying hen, where the gene is naturally active. Images PMID:6298743

Cancer-linked satellite 2 DNA hypomethylation does not regulate Sat2 non-coding RNA expression and is initiated by heat shock pathway activation.

PubMed

Tilman, Gaëlle; Arnoult, Nausica; Lenglez, Sandrine; Van Beneden, Amandine; Loriot, Axelle; De Smet, Charles; Decottignies, Anabelle

2012-08-01

Epigenetic dysfunctions, including DNA methylation alterations, play major roles in cancer initiation and progression. Although it is well established that gene promoter demethylation activates transcription, it remains unclear whether hypomethylation of repetitive heterochromatin similarly affects expression of non-coding RNA from these loci. Understanding how repetitive non-coding RNAs are transcriptionally regulated is important given that their established upregulation by the heat shock (HS) pathway suggests important functions in cellular response to stress, possibly by promoting heterochromatin reconstruction. We found that, although pericentromeric satellite 2 (Sat2) DNA hypomethylation is detected in a majority of cancer cell lines of various origins, DNA methylation loss does not constitutively hyperactivate Sat2 expression, and also does not facilitate Sat2 transcriptional induction upon heat shock. In melanoma tumor samples, our analysis revealed that the HS response, frequently upregulated in tumors, is probably the main determinant of Sat2 RNA expression in vivo. Next, we tested whether HS pathway hyperactivation may drive Sat2 demethylation. Strikingly, we found that both hyperthermia and hyperactivated RasV12 oncogene, another potent inducer of the HS pathway, reduced Sat2 methylation levels by up to 27% in human fibroblasts recovering from stress. Demethylation occurred locally on Sat2 repeats, resulting in a demethylation signature that was also detected in cancer cell lines with moderate genome-wide hypomethylation. We therefore propose that upregulation of Sat2 transcription in response to HS pathway hyperactivation during tumorigenesis may promote localized demethylation of the locus. This, in turn, may contribute to tumorigenesis, as demethylation of Sat2 was previously reported to favor chromosomal rearrangements.

Redistribution of cell cycle by arsenic trioxide is associated with demethylation and expression changes of cell cycle related genes in acute promyelocytic leukemia cell line (NB4).

PubMed

Hassani, Saeed; Khaleghian, Ali; Ahmadian, Shahin; Alizadeh, Shaban; Alimoghaddam, Kamran; Ghavamzadeh, Ardeshir; Ghaffari, Seyed H

2018-01-01

PML-RARα perturbs the normal epigenetic setting, which is essential to oncogenic transformation in acute promyelocytic leukemia (APL). Transcription induction and recruitment of DNA methyltransferases (DNMTs) by PML-RARα and subsequent hypermethylation are components of this perturbation. Arsenic trioxide (ATO), an important drug in APL therapy, concurrent with degradation of PML-RARα induces cell cycle change and apoptosis. How ATO causes cell cycle alteration has remained largely unexplained. Here, we investigated DNA methylation patterns of cell cycle regulatory genes promoters, the effects of ATO on the methylated genes and cell cycle distribution in an APL cell line, NB4. Analysis of promoter methylation status of 22 cell cycle related genes in NB4 revealed that CCND1, CCNE1, CCNF, CDKN1A, GADD45α, and RBL1 genes were methylated 60.7, 84.6, 58.6, 8.7, 33.4, and 73.7%, respectively, that after treatment with 2 μM ATO for 48 h, turn into 0.6, 13.8, 0.1, 6.6, 10.7, and 54.5% methylated. ATO significantly reduced the expression of DNMT1, 3A, and 3B. ATO induced the expression of CCND1, CCNE1, and GADD45α genes, suppressed the expression of CCNF and CDKN1A genes, which were consistent with decreased number of cells in G1 and S phases and increased number of cells in G2/M phase. In conclusion, demethylation and alteration in the expression level of the cell cycle related genes may be possible mechanisms in ATO-induced cell cycle arrest in APL cells. It may suggest that ATO by demethylation of CCND1 and CCNE1 and their transcriptional activation accelerates G1 and S transition into the G2/M cell cycle arrest.

Demethylation-mediated miR-129-5p up-regulation inhibits malignant phenotype of osteogenic osteosarcoma by targeting Homo sapiens valosin-containing protein (VCP).

PubMed

Long, Xin Hua; Zhou, Yun Fei; Peng, Ai Fen; Zhang, Zhi Hong; Chen, Xuan Yin; Chen, Wen Zhao; Liu, Jia Ming; Huang, Shan Hu; Liu, Zhi Li

2015-05-01

Previous studies demonstrated that increased Homo sapiens valosin-containing protein (VCP) may be involved in osteosarcoma (OS) metastasis. However, the underlying mechanism of VCP over-expression in OS remains unknown. In the present study, we found a significantly negative correlation between miR-129-5p and VCP protein expression in OS tissues with pulmonary metastasis (Spearman's rho, rs = -0.948). Bioinformatical prediction, Luciferase reporter assay, Western blot, and RT-PCR assays performed on OS cells indicated that VCP is a target of miR-129-5p. In addition, three CPG islands in the region of miR-129-5p promoter were detected by bioinformatical prediction, and significantly higher expression of miR-129-5p and lower methylation level of miR-129-2 gene in OS cells treated with 5-Aza-2'-deoxycytidine (a potent DNA demethylating agent) than in those untreated cells were observed. Furthermore, lower migratory and invasive ability was found in cells with elevated miR-129-5p than in those with decreased miR-129-5p. These findings indicated that increased miR-129-5p may be mediated by demethylation and inhibit OS cell migration and invasion by targeting VCP in OS, and targeting miR-129-5p/VCP signaling pathway may serve as a therapeutic strategy for OS management, although further studies will be necessary.

New indole, aminoindole and pyranoindole derivatives with anti-inflammatory activity.

PubMed

Nakkady, S S; Fathy, M M; Hishmat, O H; Mahmond, S S; Ebeid, M Y

2000-01-01

6-Methoxy-1-methyl-2,3-diphenyl indol-5-carboxaldehyde (2) was demethylated to give the 6-hydroxy derivative (3) which was cyclized to the pyrano[3,2-f]indole derivatives (4a-d) by the action of ethyl acetoacetate, diethyl malonate, malononitrile, ethyl cyanoacetate. When 4c was boiled in acetic acid, it gave 4d. Reduction of 4c by sodium borohydride yielded the orthoaminonitrile (5). Friedel Craft's acetylation of 1b yielded the 5-acetyl derivative (6), which reacted with hydrazine hydrate, o-toluidine and o-aminophenol to afford (7a-c). Demethylation of (1b) yielded the hydroxyl derivative (8), which differs from compound (9) obtained by demethylation of 6-methoxy-2,3-diphenyl-indole (1a). Friedel Craft's acetylation of 9 gave the 7-acetyl compound (10) which yielded the hydrazone (11). The reaction of primary aromatic amines, (i.e. p-nitroaniline, p-anisidine and p-bromo aniline) with 6-methoxy-1-methyl-2,3-diphenyl-indol-5-carboxaldehyde (2) gave the Schiff bases (12a-c). The latter compounds were reduced by sodium borohydride to yield the corresponding Mannich bases (13a-c). Treatment of 12a-c with thioglycolic acid led to the thiazolidin-4-one-derivatives (14a-c). When (12a-c) reacted with cyanoacetamide, the amino group was replaced by the active methylene to form the cyano compound (15). The structure was confirmed by reacting the carboxaldehyde (2) with cyanoacetamide to yield (15). Pharmacological screening was has been carried out to test the anti-inflammatory activity, ulcerogenecity, effect on the isolated rabbit intestine and the antispasmodic activity.

Inhibitors of histone demethylation and histone deacetylation cooperate in regulating gene expression and inhibiting growth in human breast cancer cells

PubMed Central

Vasilatos, Shauna N.; Boric, Lamia; Shaw, Patrick G.; Davidson, Nancy E.

2013-01-01

Abnormal activities of histone lysine demethylases (KDMs) and lysine deacetylases (HDACs) are associated with aberrant gene expression in breast cancer development. However, the precise molecular mechanisms underlying the crosstalk between KDMs and HDACs in chromatin remodeling and regulation of gene transcription are still elusive. In this study, we showed that treatment of human breast cancer cells with inhibitors targeting the zinc cofactor dependent class I/II HDAC, but not NAD+ dependent class III HDAC, led to significant increase of H3K4me2 which is a specific substrate of histone lysine-specific demethylase 1 (LSD1) and a key chromatin mark promoting transcriptional activation. We also demonstrated that inhibition of LSD1 activity by a pharmacological inhibitor, pargyline, or siRNA resulted in increased acetylation of H3K9 (AcH3K9). However, siRNA knockdown of LSD2, a homolog of LSD1, failed to alter the level of AcH3K9, suggesting that LSD2 activity may not be functionally connected with HDAC activity. Combined treatment with LSD1 and HDAC inhibitors resulted in enhanced levels of H3K4me2 and AcH3K9, and exhibited synergistic growth inhibition of breast cancer cells. Finally, microarray screening identified a unique subset of genes whose expression was significantly changed by combination treatment with inhibitors of LSD1 and HDAC. Our study suggests that LSD1 intimately interacts with histone deacetylases in human breast cancer cells. Inhibition of histone demethylation and deacetylation exhibits cooperation and synergy in regulating gene expression and growth inhibition, and may represent a promising and novel approach for epigenetic therapy of breast cancer. PMID:21452019

Meclofenamic acid selectively inhibits FTO demethylation of m6A over ALKBH5

PubMed Central

Huang, Yue; Yan, Jingli; Li, Qi; Li, Jiafei; Gong, Shouzhe; Zhou, Hu; Gan, Jianhua; Jiang, Hualiang; Jia, Gui-Fang; Luo, Cheng; Yang, Cai-Guang

2015-01-01

Two human demethylases, the fat mass and obesity-associated (FTO) enzyme and ALKBH5, oxidatively demethylate abundant N6-methyladenosine (m6A) residues in mRNA. Achieving a method for selective inhibition of FTO over ALKBH5 remains a challenge, however. Here, we have identified meclofenamic acid (MA) as a highly selective inhibitor of FTO. MA is a non-steroidal, anti-inflammatory drug that mechanistic studies indicate competes with FTO binding for the m6A-containing nucleic acid. The structure of FTO/MA has revealed much about the inhibitory function of FTO. Our newfound understanding, revealed herein, of the part of the nucleotide recognition lid (NRL) in FTO, for example, has helped elucidate the principles behind the selectivity of FTO over ALKBH5. Treatment of HeLa cells with the ethyl ester form of MA (MA2) has led to elevated levels of m6A modification in mRNA. Our collective results highlight the development of functional probes of the FTO enzyme that will (i) enable future biological studies and (ii) pave the way for the rational design of potent and specific inhibitors of FTO for use in medicine. PMID:25452335

Influence of very low doses of mediators on fungal laccase activity - nonlinearity beyond imagination

PubMed Central

Malarczyk, Elzbieta; Kochmanska-Rdest, Janina; Jarosz-Wilkolazka, Anna

2009-01-01

Laccase, an enzyme responsible for aerobic transformations of natural phenolics, in industrial applications requires the presence of low-molecular substances known as mediators, which accelerate oxidation processes. However, the use of mediators is limited by their toxicity and the high costs of exploitation. The activation of extracellular laccase in growing fungal culture with highly diluted mediators, ABTS and HBT is described. Two high laccase-producing fungal strains, Trametes versicolor and Cerrena unicolor, were used in this study as a source of enzyme. Selected dilutions of the mediators significantly increased the activity of extracellular laccase during 14 days of cultivation what was distinctly visible in PAGE technique and in colorimetric tests. The same mediator dilutions increased demethylation properties of laccase, which was demonstrated during incubation of enzyme with veratric acid. It was established that the activation effect was assigned to specific dilutions of mediators. Our dose-response dilution process smoothly passes into the range of action of homeopathic dilutions and is of interest for homeopaths. PMID:19732425

MINA controls proliferation and tumorigenesis of glioblastoma by epigenetically regulating cyclins and CDKs via H3K9me3 demethylation.

PubMed

Huang, M-Y; Xuan, F; Liu, W; Cui, H-J

2017-01-19

It is generally known that histone demethylases regulate gene transcription by altering the methylate status on histones, but their roles in cancers and the underlying molecular mechanisms still remain unclear. MYC-induced nuclear antigen (MINA) is reported to be a histone demethylase and highly expressed in many cancers. Here, for the first time, we show that MINA is involved in glioblastoma carcinogenesis and reveal the probable mechanisms of it in cell-cycle control. Kaplan-Meier analysis of progression-free survival showed that high MINA expression was strongly correlated with poor outcome and advancing tumor stage. MINA knockdown significantly repressed the cell proliferation and tumorigenesis abilities of glioblastoma cells in vitro and in vivo that were rescued by overexpressing the full-length MINA afterwards. Microarray analysis after knockdown of MINA revealed that MINA probably regulated glioblastoma carcinogenesis through the predominant cell-cycle pathways. Further investigation showed that MINA deficiency led to a cell-cycle arrest in G1 and G2 phases. And among the downstream genes, we found that cyclins and cyclin-dependent kinases were directly activated by MINA via the demethylation of H3K9me3.

Dioxygenases Catalyze O-Demethylation and O,O-Demethylenation with Widespread Roles in Benzylisoquinoline Alkaloid Metabolism in Opium Poppy*

PubMed Central

Farrow, Scott C.; Facchini, Peter J.

2013-01-01

In opium poppy, the antepenultimate and final steps in morphine biosynthesis are catalyzed by the 2-oxoglutarate/Fe(II)-dependent dioxygenases, thebaine 6-O-demethylase (T6ODM) and codeine O-demethylase (CODM). Further investigation into the biochemical functions of CODM and T6ODM revealed extensive and unexpected roles for such enzymes in the metabolism of protopine, benzo[c]phenanthridine, and rhoeadine alkaloids. When assayed with a wide range of benzylisoquinoline alkaloids, CODM, T6ODM, and the functionally unassigned paralog DIOX2, renamed protopine O-dealkylase, showed novel and efficient dealkylation activities, including regio- and substrate-specific O-demethylation and O,O-demethylenation. Enzymes catalyzing O,O-demethylenation, which cleave a methylenedioxy bridge leaving two hydroxyl groups, have previously not been reported in plants. Similar cleavage of methylenedioxy bridges on substituted amphetamines is catalyzed by heme-dependent cytochromes P450 in mammals. Preferred substrates for O,O-demethylenation by CODM and protopine O-dealkylase were protopine alkaloids that serve as intermediates in the biosynthesis of benzo[c]phenanthridine and rhoeadine derivatives. Virus-induced gene silencing used to suppress the abundance of CODM and/or T6ODM transcripts indicated a direct physiological role for these enzymes in the metabolism of protopine alkaloids, and they revealed their indirect involvement in the formation of the antimicrobial benzo[c]phenanthridine sanguinarine and certain rhoeadine alkaloids in opium poppy. PMID:23928311

Dioxygenases catalyze O-demethylation and O,O-demethylenation with widespread roles in benzylisoquinoline alkaloid metabolism in opium poppy.

PubMed

Farrow, Scott C; Facchini, Peter J

2013-10-04

In opium poppy, the antepenultimate and final steps in morphine biosynthesis are catalyzed by the 2-oxoglutarate/Fe(II)-dependent dioxygenases, thebaine 6-O-demethylase (T6ODM) and codeine O-demethylase (CODM). Further investigation into the biochemical functions of CODM and T6ODM revealed extensive and unexpected roles for such enzymes in the metabolism of protopine, benzo[c]phenanthridine, and rhoeadine alkaloids. When assayed with a wide range of benzylisoquinoline alkaloids, CODM, T6ODM, and the functionally unassigned paralog DIOX2, renamed protopine O-dealkylase, showed novel and efficient dealkylation activities, including regio- and substrate-specific O-demethylation and O,O-demethylenation. Enzymes catalyzing O,O-demethylenation, which cleave a methylenedioxy bridge leaving two hydroxyl groups, have previously not been reported in plants. Similar cleavage of methylenedioxy bridges on substituted amphetamines is catalyzed by heme-dependent cytochromes P450 in mammals. Preferred substrates for O,O-demethylenation by CODM and protopine O-dealkylase were protopine alkaloids that serve as intermediates in the biosynthesis of benzo[c]phenanthridine and rhoeadine derivatives. Virus-induced gene silencing used to suppress the abundance of CODM and/or T6ODM transcripts indicated a direct physiological role for these enzymes in the metabolism of protopine alkaloids, and they revealed their indirect involvement in the formation of the antimicrobial benzo[c]phenanthridine sanguinarine and certain rhoeadine alkaloids in opium poppy.

A Tetrahydrofolate-Dependent Methyltransferase Catalyzing the Demethylation of Dicamba in Sphingomonas sp. Strain Ndbn-20

PubMed Central

Yao, Li; Yu, Lin-Lu; Zhang, Jun-Jie; Xie, Xiang-Ting; Tao, Qing; Yan, Xin; Hong, Qing; Qiu, Ji-Guo

2016-01-01

ABSTRACT Sphingomonas sp. strain Ndbn-20 degrades and utilizes the herbicide dicamba as its sole carbon and energy source. In the present study, a tetrahydrofolate (THF)-dependent dicamba methyltransferase gene, dmt, was cloned from the strain, and three other genes, metF, dhc, and purU, which are involved in THF metabolism, were found to be located downstream of dmt. A transcriptional study revealed that the four genes constituted one transcriptional unit that was constitutively transcribed. Lysates of cells grown with glucose or dicamba exhibited almost the same activities, which further suggested that the dmt gene is constitutively expressed in the strain. Dmt shared 46% and 45% identities with the methyltransferases DesA and LigM from Sphingomonas paucimobilis SYK-6, respectively. The purified Dmt catalyzed the transfer of methyl from dicamba to THF to form the herbicidally inactive metabolite 3,6-dichlorosalicylic acid (DCSA) and 5-methyl-THF. The activity of Dmt was inhibited by 5-methyl-THF but not by DCSA. The introduction of a codon-optimized dmt gene into Arabidopsis thaliana enhanced resistance against dicamba. In conclusion, this study identified a THF-dependent dicamba methyltransferase, Dmt, with potential applications for the genetic engineering of dicamba-resistant crops. IMPORTANCE Dicamba is a very important herbicide that is widely used to control more than 200 types of broadleaf weeds and is a suitable target herbicide for the engineering of herbicide-resistant transgenic crops. A study of the mechanism of dicamba metabolism by soil microorganisms will benefit studies of its dissipation, transformation, and migration in the environment. This study identified a THF-dependent methyltransferase, Dmt, capable of catalyzing dicamba demethylation in Sphingomonas sp. Ndbn-20, and a preliminary study of its enzymatic characteristics was performed. Introduction of a codon-optimized dmt gene into Arabidopsis thaliana enhanced resistance against dicamba

Induction of the mesenchymal to epithelial transition by demethylation-activated microRNA-125b is involved in the anti-migration/invasion effects of arsenic trioxide on human chondrosarcoma.

PubMed

Bao, Xing; Ren, Tingting; Huang, Yi; Wang, Shidong; Zhang, Fan; Liu, Kuisheng; Zheng, Bingxin; Guo, Wei

2016-08-30

In addition to treating acute promyelocytic leukemia, arsenic trioxide (ATO) suppresses other solid tumors, including chondrosarcoma. However, the effects of ATO on metastasis in chondrosarcoma cells, and the underlying molecular mechanisms remain unclear. The effects of ATO on the migratory and invasive capacities of chondrosarcoma cells were investigated by Wound healing, Transwell and EMT assays. The expression of miR-125b in human chondrosarcoma tissues and cell lines was detected by real-time PCR analysis. Bisulfite sequencing analysis (BSP) was used to detect the effects of ATO on the expression of miR-125b. The gain-of-function and loss-of-function experiments were performed on chondrosarcoma cell lines to investigate the effects of miR-125b on chondrosarcoma invasion, and to determine whether signal transducer and activator of transcription 3(Stat3) mediates these effects. Dual-luciferase reporter assay was used to identify whether Stat3 is a direct target of miR-125b. MiR-125b was significantly downregulated in human metastatic chondrosarcoma tissues and cell lines but not in non-metastatic chondrosarcoma tissues. ATO up-regulates the expression of miR-125b by the demethylation of DNA. ATO induces MET and attenuates the invasive capacities of chondrosarcoma cells through miR-125b. Stat3 was verified as a direct target of miR-125b, which is involved in ATO regulating EMT-associated traits. These findings, for the first time, provides evidence that the miR-125b-mediated inhibition of Stat3 is involved in the ATO-induced attenuation of metastasis in chondrosarcoma cells.

Demethylation regulation of BDNF gene expression in dorsal root ganglion neurons is implicated in opioid-induced pain hypersensitivity in rats.

PubMed

Chao, Yu-Chieh; Xie, Fang; Li, Xueyang; Guo, Ruijuan; Yang, Ning; Zhang, Chen; Shi, Rong; Guan, Yun; Yue, Yun; Wang, Yun

2016-07-01

Repeated administration of morphine may result in opioid-induced hypersensitivity (OIH), which involves altered expression of numerous genes, including brain-derived neurotrophic factor (BDNF) in dorsal root ganglion (DRG) neurons. Yet, it remains unclear how BDNF expression is increased in DRG neurons after repeated morphine treatment. DNA methylation is an important mechanism of epigenetic control of gene expression. In the current study, we hypothesized that the demethylation regulation of certain BDNF gene promoters in DRG neurons may contribute to the development of OIH. Real-time RT-PCR was used to assess changes in the mRNA transcription levels of major BDNF exons including exon I, II, IV, VI, as well as total BDNF mRNA in DRGs from rats after repeated morphine administration. The levels of exon IV and total BDNF mRNA were significantly upregulated by repeated morphine administration, as compared to that in saline control group. Further, ELISA array and immunocytochemistry study revealed a robust upregulation of BDNF protein expression in DRG neurons after repeated morphine exposure. Correspondingly, the methylation levels of BDNF exon IV promoter showed a significant downregulation by morphine treatment. Importantly, intrathecal administration of a BDNF antibody, but not control IgG, significantly inhibited mechanical hypersensitivity that developed in rats after repeated morphine treatment. Conversely, intrathecal administration of an inhibitor of DNA methylation, 5-aza-2'-deoxycytidine (5-aza-dC) markedly upregulated the BDNF protein expression in DRG neurons and enhanced the mechanical allodynia after repeated morphine exposure. Together, our findings suggest that demethylation regulation of BDNF gene promoter may be implicated in the development of OIH through epigenetic control of BDNF expression in DRG neurons. Copyright © 2016 Elsevier Ltd. All rights reserved.

Characterization of the active site properties of CYP4F12.

PubMed

Eksterowicz, John; Rock, Dan A; Rock, Brooke M; Wienkers, Larry C; Foti, Robert S

2014-10-01

Cytochrome P450 4F12 is a drug-metabolizing enzyme that is primarily expressed in the liver, kidney, colon, small intestine, and heart. The properties of CYP4F12 that may impart an increased catalytic selectivity (decreased promiscuity) were explored through in vitro metabolite elucidation, kinetic isotope effect experiments, and computational modeling of the CYP4F12 active site. By using astemizole as a probe substrate for CYP4F12 and CYP3A4, it was observed that although CYP4F12 favored astemizole O-demethylation as the primary route of metabolism, CYP3A4 was capable of metabolizing astemizole at multiple sites on the molecule. Deuteration of astemizole at the site of O-demethylation resulted in an isotope effect of 7.1 as well as an 8.3-fold decrease in the rate of clearance for astemizole by CYP4F12. Conversely, although an isotope effect of 3.8 was observed for the formation of the O-desmethyl metabolite when deuterated astemizole was metabolized by CYP3A4, there was no decrease in the clearance of astemizole. Development of a homology model of CYP4F12 based on the crystal structure of cytochrome P450 BM3 predicted an active site volume for CYP4F12 that was approximately 76% of the active site volume of CYP3A4. As predicted, multiple favorable binding orientations were available for astemizole docked into the active site of CYP3A4, but only a single binding orientation with the site of O-demethylation oriented toward the heme was identified for CYP4F12. Overall, it appears that although CYP4F12 may be capable of binding similar ligands to other cytochrome P450 enzymes such as CYP3A4, the ability to achieve catalytically favorable orientations may be inherently more difficult because of the increased steric constraints of the CYP4F12 active site. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Immunomediated Pan-cancer Regulation Networks are Dominant Fingerprints After Treatment of Cell Lines with Demethylation.

PubMed

El Baroudi, Mariama; Cinti, Caterina; Capobianco, Enrico

2016-01-01

Pan-cancer studies are particularly relevant not only for addressing the complexity of the inherently observed heterogeneity but also for identifying clinically relevant features that may be common to the cancer types. Immune system regulations usually reveal synergistic modulation with other cancer mechanisms and in combination provide insights on possible advances in cancer immunotherapies. Network inference is a powerful approach to decipher pan-cancer systems dynamics. The methodology proposed in this study elucidates the impacts of epigenetic treatment on the drivers of complex pan-cancer regulation circuits involving cell lines of five cancer types. These patterns were observed from differential gene expression measurements following demethylation with 5-azacytidine. Networks were built to establish associations of phenotypes at molecular level with cancer hallmarks through both transcriptional and post-transcriptional regulation mechanisms. The most prominent feature that emerges from our integrative network maps, linking pathway landscapes to disease and drug-target associations, refers primarily to a mosaic of immune-system crosslinked influences. Therefore, characteristics initially evidenced in single cancer maps become motifs well summarized by network cores and fingerprints.

Characterization of human liver cytochrome P-450 enzymes involved in the O-demethylation of a new P-glycoprotein inhibitor HM-30181.

PubMed

Paek, In Bok; Kim, Sung Yeon; Kim, Maeng Sup; Kim, John; Lee, Gwansun; Lee, Hye Suk

2007-08-01

HM-30181, 4-oxo-4H-chromene-2-carboxylic acid [2-(2-{4-[2-(6,7-dimethoxy-3,4-dihydro-1H-isoquinolin-2-yl)-ethyl]-phenyl}-2H-tetrazol-5-yl)-4,5-dimethoxy-phenyl]-amide, is a new P-glycoprotein inhibitor with the potential to increase the cytotoxic activity of orally coadministered paclitaxel. This study was performed to characterize human cytochrome P-450 (CYP) enzymes involved in the metabolism of HM-30181 to 4- or 5-O-desmethyl-HM-30181 (M2) and 6- or 7-O-desmethyl-HM-30181 (M3) and to investigate the inhibitory potential of HM-30181 on CYP enzymes in human liver microsomes. CYP3A4 was identified as the major isozyme responsible for the O-demethylation of HM-30181 to M2 and M3 based on the correlation analysis, chemical inhibition and immuno-inhibition study and metabolism in cDNA-expressed human CYP isozymes. HM-30181 itself had no inhibitory effects on CYPs 1A2, 2A6, 2C8, 2C9, 2C19, 2D6, and 3A4 in human liver microsomes, suggesting the possibility that the pharmacokinetics of HM-30181 could be changed with coadministration of known CYP3A4 inducers or inhibitors.

Parallel Activation in Bilingual Phonological Processing

ERIC Educational Resources Information Center

Lee, Su-Yeon

2011-01-01

In bilingual language processing, the parallel activation hypothesis suggests that bilinguals activate their two languages simultaneously during language processing. Support for the parallel activation mainly comes from studies of lexical (word-form) processing, with relatively less attention to phonological (sound) processing. According to…

Phlpp1 facilitates post-traumatic osteoarthritis and is induced by inflammation and promoter demethylation in human osteoarthritis

PubMed Central

Bradley, Elizabeth W.; Carpio, Lomeli R.; McGee-Lawrence, Meghan E.; Becerra, Clara Castillejo; Amanatullah, Derek F.; Ta, Lauren E.; Otero, Miguel; Goldring, Mary B.; Kakar, Sanjeev; Westendorf, Jennifer J.

2016-01-01

OBJECTIVE Osteoarthritis (OA) is the most common form of arthritis and a leading cause of disability. OA is characterized by articular chondrocyte deterioration, subchondral bone changes and debilitating pain. One strategy to promote cartilage regeneration and repair is to accelerate proliferation and matrix production of articular chondrocytes. We previously reported that the protein phosphatase Phlpp1 controls chondrocyte differentiation by regulating the activities of anabolic kinases. Here we examined the role of Phlpp1 in osteoarthritis progression in a murine model. We also assessed PHLPP1 expression and promoter methylation. DESIGN Knee joints of WT and Phlpp1−/− mice were surgically destabilized by transection of the medial meniscal ligament (DMM). Mice were assessed for signs of OA progression via radiographic and histological analyses, and pain assessment for mechanical hypersensitivity using the von Frey assay. Methylation of the PHLPP1 promoter and PHLPP1 expression was evaluated in human articular cartilage and chondrocyte cell lines. RESULTS Following DMM surgeries, Phlpp1 deficient mice showed fewer signs of OA and cartilage degeneration. Mechanical allodynia associated with DMM surgeries was also attenuated in Phlpp1−/− mice. PHLPP1 was highly expressed in human articular cartilage from OA patients, but was undetectable in cartilage specimens from femoral neck fractures. Higher PHLPP1 levels correlated with less PHLPP1 promoter CpG methylation in cartilage from OA patients. Blocking cytosine methylation or treatment with inflammatory mediators enhanced PHLPP1 expression in human chondrocyte cell lines. CONCLUSION Phlpp1 deficiency protects against OA progression while CpG demethylation and inflammatory responses promote PHLPP1 expression. PMID:26746148

Interaction of Microbial and Abiotic Processes in Soil Leading to the (Bio)Conversion and Ultimate Attenuation of New Insensitive Munitions Compounds

DTIC Science & Technology

2016-12-30

Toxicity is expressed as percentage of toxicant- free activity 125 Figure 4.12-1. Panel A: (Bio)transformation pathways of DNAN in anaerobic incubations...O-demethylation of the methoxy group was confirmed by formation of formaldehye. Cell free extracts of the Bacillus culture yielded formation of 2...periodically until the production of methane became constant in the toxicant- free controls. The maximum specific methanogenic activity of the

Application of homology modeling to generate CYP1A1 mutants with enhanced activation of the cancer chemotherapeutic prodrug dacarbazine.

PubMed

Lewis, Benjamin C; Mackenzie, Peter I; Miners, John O

2011-11-01

The chemotherapeutic prodrug dacarbazine (DTIC) has limited efficacy in human malignancies and exhibits numerous adverse effects that arise from systemic exposure to the cytotoxic metabolite. DTIC is activated by CYP1A1 and CYP1A2 catalyzed N-demethylation. However, structural features of these enzymes that confer DTIC N-demethylation have not been characterized. A validated homology model of CYP1A1 was employed to elucidate structure-activity relationships and to engineer CYP1A1 enzymes with altered DTIC activation. In silico docking demonstrated that DTIC orientates proximally to Ser122, Phe123, Asp313, Ala317, Ile386, Tyr259, and Leu496 of human CYP1A1. The site of metabolism is positioned 5.6 Å from the heme iron at an angle of 105.3°. Binding in the active site is stabilized by H-bonding between Tyr259 and the N(2) position of the imidazole ring. Twenty-seven CYP1A1 mutants were generated and expressed in Escherichia coli in yields ranging from 9 to 225 pmol P450/mg. DTIC N-demethylation by the E161K, E256K, and I458V mutants exhibited Michaelis-Menten kinetics, with decreases in K(m) (183-249 μM) that doubled the catalytic efficiency (p < 0.05) relative to wild-type CYP1A1 (K(m), 408 ± 43 μM; V(max), 28 ± 4 pmol · min(-1) · pmol of P450(-1)). The generation of enzymes with catalytically enhanced DTIC activation highlights the potential use of mutant CYP1A1 proteins in P450-based gene-directed enzyme prodrug therapy for the treatment of metastatic malignant melanoma.

Mercury methylation in rice paddies and its possible controlling factors in the Hg mining area, Guizhou province, Southwest China.

PubMed

Zhao, Lei; Qiu, Guangle; Anderson, Christopher W N; Meng, Bo; Wang, Dingyong; Shang, Lihai; Yan, Haiyu; Feng, Xinbin

2016-08-01

Understanding mercury (Hg) methylation/demethylation processes and the factors controlling methylmercury (MeHg) production within the rice paddy ecosystem of Hg mining areas is critical to assess the risk of MeHg contamination in rice grain. Two typical Hg-contaminated mining sites, a current-day artisanal site (Gouxi) and an abandoned site (Wukeng), were chosen in this study. We qualified the in situ specific methylation/demethylation rate constants in rice paddy soil during a complete rice-growing season. Our results demonstrate that MeHg levels in rice paddy soil were a function of both methylation and demethylation processes and the net methylation potential in the rice paddy soil reflected the measured MeHg production at any time point. Sulfate stimulating the activity of sulfate-reducing bacteria was a potentially important metabolic pathway for Hg methylation in rice paddies. We suggest that bioavailable Hg derived from new atmospheric deposition appears to be the primary factor regulating net MeHg production in rice paddies. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dioxygen Binding in the Active Site of Histone Demethylase JMJD2A and the Role of the Protein Environment.

PubMed

Cortopassi, Wilian A; Simion, Robert; Honsby, Charles E; França, Tanos C C; Paton, Robert S

2015-12-21

JMJD2A catalyses the demethylation of di- and trimethylated lysine residues in histone tails and is a target for the development of new anticancer medicines. Mechanistic details of demethylation are yet to be elucidated and are important for the understanding of epigenetic processes. We have evaluated the initial step of histone demethylation by JMJD2A and demonstrate the dramatic effect of the protein environment upon oxygen binding using quantum mechanics/molecular mechanics (QM/MM) calculations. The changes in electronic structure have been studied for possible spin states and different conformations of O2 , using a combination of quantum and classical simulations. O2 binding to this histone demethylase is computed to occur preferentially as an end-on superoxo radical bound to a high-spin ferric centre, yielding an overall quintet ground state. The favourability of binding is strongly influenced by the surrounding protein: we have quantified this effect using an energy decomposition scheme into electrostatic and dispersion contributions. His182 and the methylated lysine assist while Glu184 and the oxoglutarate cofactor are deleterious for O2 binding. Charge separation in the superoxo-intermediate benefits from the electrostatic stabilization provided by the surrounding residues, stabilizing the binding process significantly. This work demonstrates the importance of the extended protein environment in oxygen binding, and the role of energy decomposition in understanding the physical origin of binding/recognition. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Demethylation of neferine in human liver microsomes and formation of quinone methide metabolites mediated by CYP3A4 accentuates its cytotoxicity.

PubMed

Shen, Qi; Zuo, Minjuan; Ma, Li; Tian, Ye; Wang, Lu; Jiang, Huidi; Zhou, Quan; Zhou, Hui; Yu, Lushan; Zeng, Su

2014-12-05

Neferine is a bisbenzylisoquinoline alkaloid isolated from the seed embryos of Nelumbonucifera Gaertn (Lotus) with various potent pharmacological effects. Recently, neferine has attracted attention for its anti-tumor activities. Our study explored its metabolism and cytotoxicity mechanism. Approaches using chemical inhibitors and recombinant human enzymes to characterize the involved enzymes and kinetic studies indicated that the demethylation of neferine by cytochrome P450 (CYP) 2D6 and CYP3A4 fitted a biphasic kinetic profile. Glutathione (GSH) was used as a trapping agent to identify reactive metabolites of neferine, and four novel GSH conjugates were detected with [M+H](+) ions at m/z 902.4, 916.2, 916.1, and 930.4. Based on its structure containing para-methylene phenol and results from a product ion scan, GSH tends to conjugate with C9' after undergoing oxidative metabolism to form the binding site predominated by CYP3A4. Furthermore, the addition of recombinant human GSTA1, GSTT1, and GSTP1 had little effect on the production of the GSH conjugates. In a 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide assay, combined with the GSH modulators l-buthionine sulfoximine or N-acetyl-l-cysteine, neferine treatment of MDCK-hCYP3A4 and HepG2 cells revealed that CYP3A4 expression and cellular GSH content could cause an EC50 shift. Metabolic activation mediated by CYP3A4 and GSH depletion significantly enhanced neferine-induced cytotoxicity. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Methylmercury oxidative degradation potentials in contaminated and pristine sediments of the Carson River, Nevada

USGS Publications Warehouse

Oremland, R.S.; Miller, L.G.; Dowdle, P.; Connell, T.; Barkay, T.

1995-01-01

Sediments from mercury-contaminated and uncontaminated reaches of the Carson River, Nevada, were assayed for sulfate reduction, methanogenesis, denitrification, and monomethylmercury (MeHg) degradation. Demethylation of [14C]MeHg was detected at all sites as indicated by the formation of 14CO2 and 14CH4. Oxidative demethylation was indicated by the formation of 14CO2 and was present at significant levels in all samples. Oxidized/reduced demethylation product ratios (i.e., 14CO2/14CH4 ratios) generally ranged from 4.0 in surface layers to as low as 0.5 at depth. Production of 14CO2 was most pronounced at sediment surfaces which were zones of active denitrification and sulfate reduction but was also significant within zones of methanogenesis. In a core taken from an uncontaminated site having a high proportion of oxidized, coarse-grain sediments, sulfate reduction and methanogenic activity levels were very low and 14CO2 accounted for 98% of the product formed from [14C]MeHg. There was no apparent relationship between the degree of mercury contamination of the sediments and the occurrence of oxidative demethylation. However, sediments from Fort Churchill, the most contaminated site, were most active in terms of demethylation potentials. Inhibition of sulfate reduction with molybdate resulted in significantly depressed oxidized/reduced demethylation product ratios, but overall demethylation rates of inhibited and uninhibited samples were comparable. Addition of sulfate to sediment slurries stimulated production of 14CO2 from [14C]MeHg, while 2-bromoethanesulfonic acid blocked production of 14CH4. These results reveal the importance of sulfate-reducing and methanogenic bacteria in oxidative demethylation of MeHg in anoxic environments.

Combination of PDT and a DNA demethylating agent produces anti-tumor immune response in a mouse tumor model

NASA Astrophysics Data System (ADS)

Mroz, Pawel; Hamblin, Michael R.

2009-06-01

Epigenetic mechanisms, which involve DNA methylation and histone modifications, result in the heritable silencing of genes without a change in their coding sequence. However, these changes must be actively maintained after each cell division rendering them a promising target for pharmacologic inhibition. DNA methyltransferase inhibitors like 5-aza-deoxycytidine (5-aza-dC) induce and/or up-regulate the expression of MAGE-type antigens in human and mice cancer cells. Photodynamic therapy (PDT) has been shown to be an effective locally ablative anti-cancer treatment that has the additional advantage of stimulating tumor-directed immune response. We studied the effects of a new therapy that combined the demethylating agent 5-aza-dC with PDT in the breast cancer model 4T1 syngenic to immunocompetent BALB/c mice. PDT was used as a locally ablating tumor treatment that is capable of eliciting strong and tumor directed immune response while 5-aza-dC pretreatment was used promote de novo induction of the expression of P1A.protein. This is the mouse homolog of human MAGE family antigens and is reported to function as a tumor rejection antigen in certain mouse tumors. This strategy led to an increase in PDT-mediated immune response and better treatment outcome. These results strongly suggest that the MAGE family antigens are important target for PDT mediated immune response but that their expression can be silenced by epigenetic mechanisms. Therefore the possibility that PDT can be combined with epigenetic strategies to elicit anti-tumor immunity in MAGE-positive tumor models is highly clinically significant and should be studied in detail.

Heterogeneous Antibody-Based Activity Assay for Lysine Specific Demethylase 1 (LSD1) on a Histone Peptide Substrate.

PubMed

Schmitt, Martin L; Ladwein, Kathrin I; Carlino, Luca; Schulz-Fincke, Johannes; Willmann, Dominica; Metzger, Eric; Schilcher, Pierre; Imhof, Axel; Schüle, Roland; Sippl, Wolfgang; Jung, Manfred

2014-07-01

Posttranslational modifications of histone tails are very important for epigenetic gene regulation. The lysine-specific demethylase LSD1 (KDM1A/AOF2) demethylates in vitro predominantly mono- and dimethylated lysine 4 on histone 3 (H3K4) and is a promising target for drug discovery. We report a heterogeneous antibody-based assay, using dissociation-enhanced lanthanide fluorescent immunoassay (DELFIA) for the detection of LSD1 activity. We used a biotinylated histone 3 peptide (amino acids 1-21) with monomethylated lysine 4 (H3K4me) as the substrate for the detection of LSD1 activity with antibody-mediated quantitation of the demethylated product. We have successfully used the assay to measure the potency of reference inhibitors. The advantage of the heterogeneous format is shown with cumarin-based LSD1 inhibitor candidates that we have identified using virtual screening. They had shown good potency in an established LSD1 screening assay. The new heterogeneous assay identified them as false positives, which was verified using mass spectrometry. © 2014 Society for Laboratory Automation and Screening.

Activating clinical trials: a process improvement approach.

PubMed

Martinez, Diego A; Tsalatsanis, Athanasios; Yalcin, Ali; Zayas-Castro, José L; Djulbegovic, Benjamin

2016-02-24

The administrative process associated with clinical trial activation has been criticized as costly, complex, and time-consuming. Prior research has concentrated on identifying administrative barriers and proposing various solutions to reduce activation time, and consequently associated costs. Here, we expand on previous research by incorporating social network analysis and discrete-event simulation to support process improvement decision-making. We searched for all operational data associated with the administrative process of activating industry-sponsored clinical trials at the Office of Clinical Research of the University of South Florida in Tampa, Florida. We limited the search to those trials initiated and activated between July 2011 and June 2012. We described the process using value stream mapping, studied the interactions of the various process participants using social network analysis, and modeled potential process modifications using discrete-event simulation. The administrative process comprised 5 sub-processes, 30 activities, 11 decision points, 5 loops, and 8 participants. The mean activation time was 76.6 days. Rate-limiting sub-processes were those of contract and budget development. Key participants during contract and budget development were the Office of Clinical Research, sponsors, and the principal investigator. Simulation results indicate that slight increments on the number of trials, arriving to the Office of Clinical Research, would increase activation time by 11 %. Also, incrementing the efficiency of contract and budget development would reduce the activation time by 28 %. Finally, better synchronization between contract and budget development would reduce time spent on batching documentation; however, no improvements would be attained in total activation time. The presented process improvement analytic framework not only identifies administrative barriers, but also helps to devise and evaluate potential improvement scenarios. The strength

Methylation-Sensitive Expression of a DNA Demethylase Gene Serves As an Epigenetic Rheostat

PubMed Central

Williams, Ben P.; Pignatta, Daniela; Henikoff, Steven; Gehring, Mary

2015-01-01

Genomes must balance active suppression of transposable elements (TEs) with the need to maintain gene expression. In Arabidopsis, euchromatic TEs are targeted by RNA-directed DNA methylation (RdDM). Conversely, active DNA demethylation prevents accumulation of methylation at genes proximal to these TEs. It is unknown how a cellular balance between methylation and demethylation activities is achieved. Here we show that both RdDM and DNA demethylation are highly active at a TE proximal to the major DNA demethylase gene ROS1. Unexpectedly, and in contrast to most other genomic targets, expression of ROS1 is promoted by DNA methylation and antagonized by DNA demethylation. We demonstrate that inducing methylation in the ROS1 proximal region is sufficient to restore ROS1 expression in an RdDM mutant. Additionally, methylation-sensitive expression of ROS1 is conserved in other species, suggesting it is adaptive. We propose that the ROS1 locus functions as an epigenetic rheostat, tuning the level of demethylase activity in response to methylation alterations, thus ensuring epigenomic stability. PMID:25826366

Relative contribution of rat cytochrome P450 isoforms to the metabolism of caffeine: the pathway and concentration dependence.

PubMed

Kot, Marta; Daniel, Władysława A

2008-04-01

The aim of the present study was to estimate the relative contribution of rat P450 isoforms to the metabolism of caffeine and to assess the usefulness of caffeine as a marker substance for estimating the activity of P450 in rat liver and its potential for pharmacokinetic interactions in pharmacological experiments. The results obtained using rat cDNA-expressed P450s indicated that 8-hydroxylation was the main oxidation pathway of caffeine (70%) in the rat. CYP1A2 was found to be a key enzyme catalyzing 8-hydroxylation (72%) and substantially contributing to 3-N-demethylation (47%) and 1-N-demethylation (37.5%) at a caffeine concentration of 0.1mM (relevant to "the maximum therapeutic concentration in humans"). Furthermore, CYP2C11 considerably contributed to 3-N-demethylation (31%). The CYP2C subfamily (66%) - mainly CYP2C6 (27%) and CYP2C11 (29%) - played a major role in catalyzing 7-N-demethylation. At higher substrate concentrations, the contribution of CYP1A2 to the metabolism of caffeine decreased in favor of CYP2C11 (N-demethylations) and CYP3A2 (mainly 8-hydroxylation). The obtained results were confirmed with liver microsomes (inhibition and correlation studies). Therefore, caffeine may be used as a marker substance for assessing the activity of CYP1A2 in rats, using 8-hydroxylation (but not 3-N-demethylation-like in humans); moreover, caffeine may also be used to simultaneously, preliminarily estimate the activity of CYP2C using 7-N-demethylation as a marker reaction. Hence caffeine pharmacokinetics in rats may be changed by drugs affecting the activity of CYP1A2 and/or CYP2C, e.g. by some antidepressants.

Dynamic changes in methylome and transcriptome patterns in response to methyltransferase inhibitor 5-azacytidine treatment in citrus

PubMed Central

Xu, Jidi; Wang, Xia; Cao, Hongbo; Xu, Haidan; Xu, Qiang

2017-01-01

Abstract DNA methylation is known to play an important role in various developmental processes in plants. However, there is a general lack of understanding about the possible functions of DNA methylation in fruit trees. Using callus as a model, methylome, transcriptome and metabolite changes were assessed after treatment with the DNA methyltransferase inhibitor 5-azacytidine (5azaC). Genome-wide methylome analysis revealed the demethylation of a diverse of genes, including many genes encoding transcription factors (TFs), genes involved in biological processes, and the up-regulation of a wide range of transposable elements (TEs). Combined with the RNA-seq data, we observed no obvious genome-wide correlation between the changes in methylation status and expression levels. Furthermore, 5azaC treatment induced carotenoid degradation along with strong activation of carotenoid cleavage dioxygenases 1 (CpCCD1). Functional complementation analysis in bacterial system showed that CpCCD1 exhibited strong catalytic activities toward zeaxanthin, β-carotene and lycopene. In summary, 5azaC treatments induced carotenoid degradation by CpCCD1 activation and led to a genome-wide demethylation effect. PMID:28575160

Effects of emodin on the demethylation of tumor-suppressor genes in pancreatic cancer PANC-1 cells.

PubMed

Zhang, Hao; Chen, Liang; Bu, He-Qi; Yu, Qing-Jiang; Jiang, Dan-Dan; Pan, Feng-Ping; Wang, Yu; Liu, Dian-Lei; Lin, Sheng-Zhang

2015-06-01

Emodin, a natural anthraquinone derivative isolated from Rheum palmatum, has been reported to inhibit the growth of pancreatic cancer cells through different modes of action; yet, the detailed mechanism remains unclear. In the present study, we hypothesized that emodin exerts its antitumor effect by participating in the regulation of the DNA methylation level. Our research showed that emodin inhibited the growth of pancreatic cancer PANC-1 cells in a dose- and time-dependent manner. Dot-blot results showed that 40 µM emodin significantly inhibited genomic 5 mC expression in the PANC-1 cells, and mRNA-Seq showed that different concentrations of emodin could alter the gene expression profile in the PANC-1 cells. BSP confirmed that the methylation levels of P16, RASSF1A and ppENK were decreased, while concomitantly the unmethylated status was increased. RT-PCR and western blotting results confirmed that the low expression or absence of expression of mRNA and protein in the PANC-1 cells was re-expressed following treatment with emodin. In conclusion, our study for the first time suggests that emodin inhibits pancreatic cancer cell growth, which may be related to the demethylation of tumor-suppressor genes. The related mechanism may be through the inhibition of methyltransferase expression.

Monozygotic twins discordant for common variable immunodeficiency reveal impaired DNA demethylation during naïve-to-memory B-cell transition

PubMed Central

Rodríguez-Cortez, Virginia C.; del Pino-Molina, Lucia; Rodríguez-Ubreva, Javier; Ciudad, Laura; Gómez-Cabrero, David; Company, Carlos; Urquiza, José M.; Tegnér, Jesper; Rodríguez-Gallego, Carlos; López-Granados, Eduardo; Ballestar, Esteban

2015-01-01

Common variable immunodeficiency (CVID), the most frequent primary immunodeficiency characterized by loss of B-cell function, depends partly on genetic defects, and epigenetic changes are thought to contribute to its aetiology. Here we perform a high-throughput DNA methylation analysis of this disorder using a pair of CVID-discordant MZ twins and show predominant gain of DNA methylation in CVID B cells with respect to those from the healthy sibling in critical B lymphocyte genes, such as PIK3CD, BCL2L1, RPS6KB2, TCF3 and KCNN4. Individual analysis confirms hypermethylation of these genes. Analysis in naive, unswitched and switched memory B cells in a CVID patient cohort shows impaired ability to demethylate and upregulate these genes in transitioning from naive to memory cells in CVID. Our results not only indicate a role for epigenetic alterations in CVID but also identify relevant DNA methylation changes in B cells that could explain the clinical manifestations of CVID individuals. PMID:26081581

Intelligent Signal Processing for Active Control

DTIC Science & Technology

1992-06-17

FUNDING NUMSI Intelligent Signal Processing for Active Control C-NO001489-J-1633 G. AUTHOR(S) P.A. Ramamoorthy 7. P2RFORMING ORGANIZATION NAME(S) AND...unclassified .unclassified unclassified L . I mu-. W UNIVERSITY OF CINCINNATI COLLEGE OF ENGINEERING Intelligent Signal Processing For Rctiue Control...NAURI RESEARCH Conkact No: NO1489-J-1633 P.L: P.A.imoodh Intelligent Signal Processing For Active Control 1 Executive Summary The thrust of this

Discovery of tanshinone derivatives with anti-MRSA activity via targeted bio-transformation.

PubMed

He, Wenni; Liu, Miaomiao; Huang, Pei; Abdel-Mageed, Wael M; Han, Jianying; Watrous, Jeramie D; Nguyen, Don D; Wang, Wenzhao; Song, Fuhang; Dai, Huanqin; Zhang, Jingyu; Quinn, Ronald J; Grkovi, Tanja; Luo, Houwei; Zhang, Lixin; Liu, Xueting

2016-09-01

Two potent anti-MRSA tanshinone glycosides ( 1 and 2 ) were discovered by targeted microbial biotransformation, along with rapid identification via MS/MS networking. Serial reactions including dehydrogenation, demethylations, reduction, glycosylation and methylation have been observed after incubation of tanshinone IIA and fungus Mucor rouxianus AS 3.3447. In addition, tanshinosides B ( 2 ) showed potent activities against serial clinical isolates of oxacillin-resistant Staphylococcus aureus with MIC values of 0.78 μg/mL. This is the first study that shows a significant increase in the level and activities of tanshinone glycosides relative to the substrate tanshinone IIA.

Cascading activation from lexical processing to letter-level processing in written word production.

PubMed

Buchwald, Adam; Falconer, Carolyn

2014-01-01

Descriptions of language production have identified processes involved in producing language and the presence and type of interaction among those processes. In the case of spoken language production, consensus has emerged that there is interaction among lexical selection processes and phoneme-level processing. This issue has received less attention in written language production. In this paper, we present a novel analysis of the writing-to-dictation performance of an individual with acquired dysgraphia revealing cascading activation from lexical processing to letter-level processing. The individual produced frequent lexical-semantic errors (e.g., chipmunk → SQUIRREL) as well as letter errors (e.g., inhibit → INBHITI) and had a profile consistent with impairment affecting both lexical processing and letter-level processing. The presence of cascading activation is suggested by lower letter accuracy on words that are more weakly activated during lexical selection than on those that are more strongly activated. We operationalize weakly activated lexemes as those lexemes that are produced as lexical-semantic errors (e.g., lethal in deadly → LETAHL) compared to strongly activated lexemes where the intended target word (e.g., lethal) is the lexeme selected for production.

Exploring the Active Center of the LSD1/CoREST Complex by Molecular Dynamics Simulation Utilizing Its Co-crystallized Co-factor Tetrahydrofolate as a Probe.

PubMed

Zalloum, Waleed A; Zalloum, Hiba M

2017-12-26

Epigenetic targeting of cancer is a recent effort to manipulate the gene without destroying the genetic material. Lysine-specific demethylase 1 (LSD1) is one of the enzymes associated with the chromatin for post-translational modifications, where it demethylates lysine amino acid in the chromatin H3 tail. Many studies showed that inhibiting LSD1 could potentially be used to treat cancer epigenetically. LSD1 is associated with its corepressor protein CoREST, and it uses tetrahydrofolate as a co-factor to accept CH 2 from the demethylation process. In this study, the co-crystallized co-factor tetrahydrofolate was utilized to determine possible binding regions in the active center of the LSD1/CoREST complex. Also, the flexibility of the complex has been investigated by molecular dynamics simulation and subsequent analysis by clustering and principal component analysis. This research supported other studies and showed that LSD1/CoREST complex exists in two main conformational structures: open and closed. Furthermore, this study showed that tetrahydrofolate stably binds to the LSD1/CoREST complex, in its open conformation, at its entrance. It then binds to the core of the complex, inducing the closed conformation. Furthermore, the interactions of tetrahydrofolate to these two binding regions and the corresponding binding mode of tetrahydrofolate were investigated to be used in structure-based drug design.

Regulation and function of DNA methylation in plants and animals

PubMed Central

He, Xin-Jian; Chen, Taiping; Zhu, Jian-Kang

2011-01-01

DNA methylation is an important epigenetic mark involved in diverse biological processes. In plants, DNA methylation can be established through the RNA-directed DNA methylation pathway, an RNA interference pathway for transcriptional gene silencing (TGS), which requires 24-nt small interfering RNAs. In mammals, de novo DNA methylation occurs primarily at two developmental stages: during early embryogenesis and during gametogenesis. While it is not clear whether establishment of DNA methylation patterns in mammals involves RNA interference in general, de novo DNA methylation and suppression of transposons in germ cells require 24-32-nt piwi-interacting small RNAs. DNA methylation status is dynamically regulated by DNA methylation and demethylation reactions. In plants, active DNA demethylation relies on the repressor of silencing 1 family of bifunctional DNA glycosylases, which remove the 5-methylcytosine base and then cleave the DNA backbone at the abasic site, initiating a base excision repair (BER) pathway. In animals, multiple mechanisms of active DNA demethylation have been proposed, including a deaminase- and DNA glycosylase-initiated BER pathway. New information concerning the effects of various histone modifications on the establishment and maintenance of DNA methylation has broadened our understanding of the regulation of DNA methylation. The function of DNA methylation in plants and animals is also discussed in this review. PMID:21321601

Generation and Characterization of Induced Pluripotent Stem Cells from Aid-Deficient Mice

PubMed Central

Shimamoto, Ren; Amano, Naoki; Ichisaka, Tomoko; Watanabe, Akira; Yamanaka, Shinya; Okita, Keisuke

2014-01-01

It has been shown that DNA demethylation plays a pivotal role in the generation of induced pluripotent stem (iPS) cells. However, the underlying mechanism of this action is still unclear. Previous reports indicated that activation-induced cytidine deaminase (Aid, also known as Aicda) is involved in DNA demethylation in several developmental processes, as well as cell fusion-mediated reprogramming. Based on these reports, we hypothesized that Aid may be involved in the DNA demethylation that occurs during the generation of iPS cells. In this study, we examined the function of Aid in iPS cell generation using Aid knockout (Aid −/−) mice expressing a GFP reporter under the control of a pluripotent stem cell marker, Nanog. By introducing Oct3/4, Sox2, Klf4 and c-Myc, Nanog-GFP-positive iPS cells could be generated from the fibroblasts and primary B cells of Aid −/− mice. Their induction efficiency was similar to that of wild-type (Aid +/+) iPS cells. The Aid −/− iPS cells showed normal proliferation and gave rise to chimeras, indicating their capacity for self-renewal and pluripotency. A comprehensive DNA methylation analysis showed only a few differences between Aid +/+ and Aid −/− iPS cells. These data suggest that Aid does not have crucial functions in DNA demethylation during iPS cell generation. PMID:24718089

Studies of metabolic pathways of trimebutine by simultaneous administration of trimebutine and its deuterium-labeled metabolite

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miura, Y.; Chishima, S.; Takeyama, S.

1989-07-01

Trimebutine maleate (I), (+-)-2-dimethylamino-2-phenylbutyl 3,4,5-trimethoxybenzoate hydrogen maleate, and a deuterium-labeled sample of its hydrolyzed metabolite, 2-dimethylamino-2-phenylbutanol-d3 (II-d3), were simultaneously administered to experimental animals at an oral dose of 10 or 50 mumol/kg, and distribution ratios of the two alternative initial metabolic steps, i.e., ester hydrolysis and N-demethylation, were estimated by determining the composition of the urinary alcohol-moiety metabolites, II, and its mono- and di-demethylated metabolites, III and IV, by GC/MS. In dogs, the order of quantities of the metabolites from II-d3 was II much greater than III much greater than IV, showing predominance of conjugation over N-demethylation. However, this ordermore » was reversed when the amounts of the metabolites from I were compared, indicating that I was preferentially metabolized by N-demethylation followed by ester hydrolysis and conjugation in this order. In rats, a considerable proportion of I was presumed to be metabolized by ester hydrolysis before N-demethylation. In in vitro experiments employing the liver microsomes and homogenates of liver and small intestine from rats and dogs, it was found that both ester-hydrolizing and N-demethylating activities were higher in rats than in dogs, and the conjugating activity was higher in dogs than in rats. It was also found that I, having a high lipophilicity, was more susceptible to N-demethylation than less lipophilic II. These results from the in vitro experiments could account for the species differences in the distribution ratio of the metabolic pathways of I in vivo.« less

Epigenetic features of FoxP3 in children with cow's milk allergy.

PubMed

Paparo, Lorella; Nocerino, Rita; Cosenza, Linda; Aitoro, Rosita; D'Argenio, Valeria; Del Monaco, Valentina; Di Scala, Carmen; Amoroso, Antonio; Di Costanzo, Margherita; Salvatore, Francesco; Berni Canani, Roberto

2016-01-01

DNA methylation of the Th1 and Th2 cytokine genes is altered during cow's milk allergy (CMA). Forkhead box transcription factor 3 (FoxP3) is essential for the development and function of regulatory T cells (Tregs) and is involved in oral tolerance acquisition. We assessed whether tolerance acquisition in children with IgE-mediated CMA is associated with DNA demethylation of the Treg-specific demethylated region (TSDR) of FoxP3. Forty children (aged 3-18 months) were enrolled: 10 children with active IgE-mediated CMA (group 1), 10 children who outgrew CMA after dietary treatment with an extensively hydrolyzed casein formula containing the probiotic Lactobacillus rhamnosus GG (group 2), 10 children who outgrew CMA after treatment with other formulas (group 3), and 10 healthy controls (group 4). FoxP3 TSDR demethylation and expression were measured in mononuclear cells purified from peripheral blood of the four groups of children. FoxP3 TSDR demethylation was significantly lower in children with active IgE-mediated CMA than in either children who outgrew CMA or in healthy children. Formula selection influenced the FoxP3 TSDR demethylation profile. The FoxP3 TSDR demethylation rate and expression level were correlated. Tolerance acquisition in children with IgE-mediated CMA involves epigenetic regulation of the FoxP3 gene. This feature could be a new target for preventive and therapeutic strategies against CMA.

Control of DEMETER DNA demethylase gene transcription in male and female gamete companion cells in Arabidopsis thaliana.

PubMed

Park, Jin-Sup; Frost, Jennifer M; Park, Kyunghyuk; Ohr, Hyonhwa; Park, Guen Tae; Kim, Seohyun; Eom, Hyunjoo; Lee, Ilha; Brooks, Janie S; Fischer, Robert L; Choi, Yeonhee

2017-02-21

The DEMETER (DME) DNA glycosylase initiates active DNA demethylation via the base-excision repair pathway and is vital for reproduction in Arabidopsis thaliana DME-mediated DNA demethylation is preferentially targeted to small, AT-rich, and nucleosome-depleted euchromatic transposable elements, influencing expression of adjacent genes and leading to imprinting in the endosperm. In the female gametophyte, DME expression and subsequent genome-wide DNA demethylation are confined to the companion cell of the egg, the central cell. Here, we show that, in the male gametophyte, DME expression is limited to the companion cell of sperm, the vegetative cell, and to a narrow window of time: immediately after separation of the companion cell lineage from the germline. We define transcriptional regulatory elements of DME using reporter genes, showing that a small region, which surprisingly lies within the DME gene, controls its expression in male and female companion cells. DME expression from this minimal promoter is sufficient to rescue seed abortion and the aberrant DNA methylome associated with the null dme-2 mutation. Within this minimal promoter, we found short, conserved enhancer sequences necessary for the transcriptional activities of DME and combined predicted binding motifs with published transcription factor binding coordinates to produce a list of candidate upstream pathway members in the genetic circuitry controlling DNA demethylation in gamete companion cells. These data show how DNA demethylation is regulated to facilitate endosperm gene imprinting and potential transgenerational epigenetic regulation, without subjecting the germline to potentially deleterious transposable element demethylation.

Impaired dacarbazine activation and 7-ethoxyresorufin deethylation in vitro by polymorphic variants of CYP1A1 and CYP1A2: implications for cancer therapy.

PubMed

Lewis, Benjamin C; Korprasertthaworn, Porntipa; Miners, John O

2016-10-01

To extend our understanding of how interindividual variability mediates the efficacy of cancer treatment. The kinetics of dacarbazine (DTIC) N-demethylation by the most frequent polymorphic variants of CYP1A1 (T461N, I462V) and CYP1A2 (F186L, D348N, I386F, R431W, R456H) were characterized, along with kinetic parameters for the O-deethylation of the prototypic CYP1A substrate 7-ethoxyresorufin, using recombinant protein expression and high-performance liquid chromatographic techniques. A reduction of ∼30% in the catalytic efficiencies (measured as in-vitro intrinsic clearance, CLint) was observed for DTIC N-demethylation by the two CYP1A1 variants relative to wild type. Although a modest increase in the CLint value for DTIC N-demethylation was observed for the CYP1A2 D348N variant relative to the wild type, the CLint for the F186L variant was reduced and the I386F, R431W, and R456H variants all showed loss of catalytic function. Comparison of the kinetic data for DTIC N-demethylation and 7-ethoxyresorufin O-deethylation indicated that alterations in the kinetic parameters (Km, Vmax, CLint) observed with each of the CYP1A1 and CYP1A2 polymorphic variants were substrate dependent. These data indicate that cancer patients treated with DTIC who possess any of the CYP1A1-T461N and I462V variants or the CYP1A2-F186L, D348N, I386F, R431W, and R456H variants are likely to have decreased prodrug activation, and hence may respond less favorably to DTIC treatment compared with individuals with wild-type CYP1A alleles.

Demethylation of induced pluripotent stem cells from type 1 diabetic patients enhances differentiation into functional pancreatic β cells.

PubMed

Manzar, Gohar S; Kim, Eun-Mi; Zavazava, Nicholas

2017-08-25

Type 1 diabetes (T1D) can be managed by transplanting either the whole pancreas or isolated pancreatic islets. However, cadaveric pancreas is scarcely available for clinical use, limiting this approach. As such, there is a great need to identify alternative sources of clinically usable pancreatic tissues. Here, we used induced pluripotent stem (iPS) cells derived from patients with T1D to generate glucose-responsive, insulin-producing cells (IPCs) via 3D culture. Initially, T1D iPS cells were resistant to differentiation, but transient demethylation treatment significantly enhanced IPC yield. The cells responded to high-glucose stimulation by secreting insulin in vitro The shape, size, and number of their granules, as observed by transmission electron microscopy, were identical to those found in cadaveric β cells. When the IPCs were transplanted into immunodeficient mice that had developed streptozotocin-induced diabetes, they promoted a dramatic decrease in hyperglycemia, causing the mice to become normoglycemic within 28 days. None of the mice died or developed teratomas. Because the cells are derived from "self," immunosuppression is not required, providing a much safer and reliable treatment option for T1D patients. Moreover, these cells can be used for drug screening, thereby accelerating drug discovery. In conclusion, our approach eliminates the need for cadaveric pancreatic tissue.

Chemoprevention of colorectal cancer by black raspberry anthocyanins involved the modulation of gut microbiota and SFRP2 demethylation.

PubMed

Chen, Lili; Jiang, Bowen; Zhong, Chunge; Guo, Jun; Zhang, Lihao; Mu, Teng; Zhang, Qiuhua; Bi, Xiuli

2018-03-08

Freeze-dried black raspberry (BRB) powder is considered as a potential cancer chemopreventive agent. In this study, we fed azoxymethane (AOM)/dextran sodium sulfate (DSS)-treated C57BL/6J mice with a diet containing BRB anthocyanins for 12 weeks, and this led to a reduction in colon carcinogenesis. These animals had consistently lower tumor multiplicity compared with AOM/DSS-treated mice not receiving BRB anthocyanins. In AOM/DSS-treated mice, the number of pathogenic bacteria, including Desulfovibrio sp. and Enterococcus spp., was increased significantly, whereas probiotics such as Eubacterium rectale, Faecalibacterium prausnitzii and Lactobacillus were dramatically decreased, but BRB anthocyanins supplement could reverse this imbalance in gut microbiota. BRB anthocyanins also caused the demethylation of the SFRP2 gene promoter, resulting in increased expression of SFRP2, both at the mRNA and protein levels. Furthermore, the expression levels of DNMT31 and DNMT3B, as well as of p-STAT3 were downregulated by BRB anthocyanins in these animals. Taken together, these results suggested that BRB anthocyanins could modulate the composition of gut commensal microbiota, and changes in inflammation and the methylation status of the SFRP2 gene may play a central role in the chemoprevention of CRC.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Olsen, Todd Andrew; Brandt, Craig C.; Brooks, Scott C.

Mercury (Hg) methylation and methylmercury (MMHg) demethylation activity of periphyton biofilms from East Fork Poplar Creek, Tennessee, USA (EFPC) were measured during 2014-2015 using stable Hg isotopic rate assays. 201Hg II and MM 202Hg were added to intact periphyton samples and the formation of MM 201Hg and loss of MM 202Hg were monitored over time and used to calculate first-order rate constants for methylation and demethylation, respectively. The influence of location, temperature/season, light exposure and biofilm structure on methylation and demethylation were examined. Between-site differences in net methylation for samples collected from an upstream versus downstream location were driven bymore » differences in the demethylation rate constant (k d). In contrast, the within-site seasonal difference in net methylation was driven by changes in the methylation rate constant (k m). Samples incubated in the dark had lower net methylation due to km values that were 60% less than those incubated in the light. Disrupting the biofilm structure decreased km by 50% and resulted in net demethylating conditions. Overall, the measured rates resulted in a net excess of MMHg generated which could account for 27-85% of the MMHg flux in EFPC and suggests intact, actively photosynthesizing periphyton biofilms harbor zones of MMHg production.« less

Periphyton biofilms influence net methylmercury production in an industrially contaminated system

DOE PAGES

Olsen, Todd Andrew; Brandt, Craig C.; Brooks, Scott C.

2016-09-12

Mercury (Hg) methylation and methylmercury (MMHg) demethylation activity of periphyton biofilms from East Fork Poplar Creek, Tennessee, USA (EFPC) were measured during 2014-2015 using stable Hg isotopic rate assays. 201Hg II and MM 202Hg were added to intact periphyton samples and the formation of MM 201Hg and loss of MM 202Hg were monitored over time and used to calculate first-order rate constants for methylation and demethylation, respectively. The influence of location, temperature/season, light exposure and biofilm structure on methylation and demethylation were examined. Between-site differences in net methylation for samples collected from an upstream versus downstream location were driven bymore » differences in the demethylation rate constant (k d). In contrast, the within-site seasonal difference in net methylation was driven by changes in the methylation rate constant (k m). Samples incubated in the dark had lower net methylation due to km values that were 60% less than those incubated in the light. Disrupting the biofilm structure decreased km by 50% and resulted in net demethylating conditions. Overall, the measured rates resulted in a net excess of MMHg generated which could account for 27-85% of the MMHg flux in EFPC and suggests intact, actively photosynthesizing periphyton biofilms harbor zones of MMHg production.« less

Intramolecular Oxidative O-Demethylation of an Oxoferryl Porphyrin Complexed with a Per-O-methylated β-Cyclodextrin Dimer.

PubMed

Kitagishi, Hiroaki; Kurosawa, Shun; Kano, Koji

2016-11-22

The intramolecular oxidation of ROCH 3 to ROCH 2 OH, where the latter compound spontaneously decomposed to ROH and HCHO, was observed during the reaction of the supramolecular complex (met-hemoCD3) with cumene hydroperoxide in aqueous solution. Met-hemoCD3 is composed of meso-tetrakis(4-sulfonatophenyl)porphinatoiron(III) (Fe III TPPS) and a per-O-methylated β-cyclodextrin dimer having an -OCH 2 PyCH 2 O- linker (Py=pyridine-3,5-diyl). The O=Fe IV TPPS complex was formed by the reaction of met-hemoCD3 with cumene hydroperoxide, and isolated by gel-filtration chromatography. Although the isolated O=Fe IV TPPS complex in the cyclodextrin cage was stable in aqueous solution at 25 °C, it was gradually converted to Fe II TPPS (t 1/2 =7.6 h). This conversion was accompanied by oxidative O-demethylation of an OCH 3 group in the cyclodextrin dimer. The results indicated that hydrogen abstraction by O=Fe IV TPPS from ROCH 3 yields HO-Fe III TPPS and ROCH 2 . . This was followed by radical coupling to afford Fe II TPPS and ROCH 2 OH. The hemiacetal (ROCH 2 OH) immediately decomposed to ROH and HCHO. This study revealed the ability of oxoferryl porphyrin to induce two-electron oxidation. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biogeochemical factors influencing net mercury methylation in contaminated freshwater sediments from the St. Lawrence River in Cornwall, Ontario, Canada.

PubMed

Avramescu, Mary-Luyza; Yumvihoze, Emmanuel; Hintelmann, Holger; Ridal, Jeff; Fortin, Danielle; Lean, David R S

2011-02-01

The activity of various anaerobic microbes, including sulfate reducers (SRB), iron reducers (FeRP) and methanogens (MPA) has been linked to mercury methylation in aquatic systems, although the relative importance of each microbial group in the overall process is poorly understood in natural sediments. The present study focused on the biogeochemical factors (i.e. the relative importance of various groups of anaerobic microbes (FeRP, SRB, and MPA) that affect net monomethylmercury (MMHg) formation in contaminated sediments of the St. Lawrence River (SRL) near Cornwall (Zone 1), Ontario, Canada. Methylation and demethylation potentials were measured separately by using isotope-enriched mercury species ((200)Hg(2+) and MM(199)Hg(+)) in sediment microcosms treated with specific microbial inhibitors. Sediments were sampled and incubated in the dark at room temperature in an anaerobic chamber for 96h. The potential methylation rate constants (K(m)) and demethylation rates (K(d)) were found to differ significantly between microcosms. The MPA-inhibited microcosm had the highest potential methylation rate constant (0.016d(-1)), whereas the two SRB-inhibited microcosms had comparable potential methylation rate constants (0.003d(-1) and 0.002d(-1), respectively). The inhibition of methanogens stimulated net methylation by inhibiting demethylationand by stimulating methylation along with SRB activity. The inhibition of both methanogens and SRB was found to enhance the iron reduction rates but did not completely stop MMHg production. The strong positive correlation between K(m) and Sulfate Reduction Rates (SRR) and between K(d) and Methane Production Rates (MPR) supports the involvement of SRB in Hg methylation and MPA in MMHg demethylation in the sediments. In contrast, the strong negative correlation between K(d) and Iron Reduction Rates (FeRR) shows that the increase in FeRR corresponds to a decrease in demethylation, indicating that iron reduction may influence net

Novel activation process for Mg-implanted GaN

NASA Astrophysics Data System (ADS)

Hashimoto, Shin; Nakamura, Takao; Honda, Yoshio; Amano, Hiroshi

2014-02-01

A novel activation process for Mg-implanted GaN was demonstrated. As opposed to the conventional thermal annealing process, an H2/NH3 alternating supply annealing process achieved better optical activation, stronger near-ultraviolet luminescence and weaker yellow luminescence in the photoluminescence spectroscopy. After this process, small hexagonal hillocks were observed on the surface, which indicated that crystal regrowth was induced by this process, consisting of decomposition of GaN by H2 supplies and re-crystallization by NH3 supplies. It was revealed that the implanted Mg could easily be located at the activation site by means of crystal regrowth by this process.

Molecular Level Understanding of the Factors Affecting the Stability of Dimethoxy Benzene Catholyte Candidates from First-Principles Investigations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Assary, Rajeev S.; Zhang, Lu; Huang, Jinhua

First-principles simulations are performed to gain molecular level insights into the factors affecting the stability of seven 1,4-dimethoxybenzene (DMB) derivatives. These molecules are potential catholyte candidates for nonaqueous redox flow battery systems. Computations are performed to predict oxidation potentials in various dielectric mediums, intrinsic-reorganization energies, and structural changes of these representative catholyte molecules during the redox process. In order to understand the stability of the DMB-based radical cations, the thermodynamic feasibility of the following reactions is computed using density functional theory: (a) deprotonation, (b) dimerization, (c) hydrolysis, and (d) demethylation. The computations indicate that radical cations of the 2,3-dimethyl andmore » 2,5-dimethyl derivatives are the most stable among the DMB derivatives considered in this study. In the presence of solvents with high-proton solvating ability (water, DMSO, acetonitrile), degradation of cation radical occurring via deprotonation is the most likely mechanism. In the presence of solvents such as propylene carbonate (PC), demethylation was found to be the most likely reaction that causes degradation of radical cations. From the computed enthalpy of activation (Delta H-double dagger) for a demethylation reaction in PC, the 2,5-dimethyl DMB cation radical would exhibit better kinetic stability in comparison to the other candidates. Finally, this investigation suggests that computational studies of structural properties such as redox potentials, reorganization energies, and the computed reaction energetics (deprotonation and demethylation) of charged species can be used to predict the relative stability of a large set of molecules required for the discovery of novel redox active materials for flow battery applications« less

Molecular Level Understanding of the Factors Affecting the Stability of Dimethoxy Benzene Catholyte Candidates from First-Principles Investigations

DOE PAGES

Assary, Rajeev S.; Zhang, Lu; Huang, Jinhua; ...

2016-06-14

First-principles simulations are performed to gain molecular level insights into the factors affecting the stability of seven 1,4-dimethoxybenzene (DMB) derivatives. These molecules are potential catholyte candidates for nonaqueous redox flow battery systems. Computations are performed to predict oxidation potentials in various dielectric mediums, intrinsic-reorganization energies, and structural changes of these representative catholyte molecules during the redox process. In order to understand the stability of the DMB-based radical cations, the thermodynamic feasibility of the following reactions is computed using density functional theory: (a) deprotonation, (b) dimerization, (c) hydrolysis, and (d) demethylation. The computations indicate that radical cations of the 2,3-dimethyl andmore » 2,5-dimethyl derivatives are the most stable among the DMB derivatives considered in this study. In the presence of solvents with high-proton solvating ability (water, DMSO, acetonitrile), degradation of cation radical occurring via deprotonation is the most likely mechanism. In the presence of solvents such as propylene carbonate (PC), demethylation was found to be the most likely reaction that causes degradation of radical cations. From the computed enthalpy of activation (Delta H-double dagger) for a demethylation reaction in PC, the 2,5-dimethyl DMB cation radical would exhibit better kinetic stability in comparison to the other candidates. Finally, this investigation suggests that computational studies of structural properties such as redox potentials, reorganization energies, and the computed reaction energetics (deprotonation and demethylation) of charged species can be used to predict the relative stability of a large set of molecules required for the discovery of novel redox active materials for flow battery applications« less

Interactive effects of polymethoxy flavones from Citrus on cell growth inhibition in human neuroblastoma SH-SY5Y cells.

PubMed

Akao, Yukihiro; Itoh, Tomohiro; Ohguchi, Kenji; Iinuma, Munekazu; Nozawa, Yoshinori

2008-03-15

Much evidence indicates that typical phytochemicals such as resveratrol, epigallocatechin gallate, and curcumin have a growth inhibitory effect against cancer cells when each is tested separately. However, when fruits and vegetables including a mixture of phytochemicals are consumed, it is unclear whether this anti-proliferative activity is elicited in the body. Initially, we found that nobiletin, a typical polymethoxy flavone from Citrus, had a preventive effect on H(2)O(2)-induced apoptosis at 20-30 microM in human neuroblastoma SH-SY5Y cells. Nobiletin acted as a signal modulator to attenuate the activation of the intrinsic pathway of the apoptosis induced by H(2)O(2) exposure. On the other hand, tangeretin and 5-demethyl nobiletin, which are also polymethoxy flavones from Citrus, were shown to have a growth inhibitory effect by us and others. These results led us to investigate the interactive effects of these polymethoxy flavones on cell growth. In the present study, we found that tangeretin, nobiletin, and 5-demethyl nobiletin exhibited a cancelling, synergistic, or additive effect when combinations of two of these three compounds were tested. As to the structure-activity relationship, the methyl group at C-5 in nobiletin was shown to contribute to the anti-proliferative effect. By the combined treatment with tangeretin and 5-demethyl nobiletin, the apoptotic cell population and the activity of caspase-3 were synergistically elevated. The finding that tangeretin and 5-demethyl nobiletin induced apoptosis by reducing the mitochondrial membrane potential suggested that an intrinsic pathway of apoptosis was synergistically activated by the combination treatment with tangeretin and 5-demethyl nobiletin. On the other hand, in the combined treatment including nobiletin, the growth inhibitory activity of tangeretin was reduced. These results indicate the relevance of the combination of phytochemicals for the enhancement of the anticancer effect.

Base Excision Repair of Tandem Modifications in a Methylated CpG Dinucleotide*

PubMed Central

Sassa, Akira; Çağlayan, Melike; Dyrkheeva, Nadezhda S.; Beard, William A.; Wilson, Samuel H.

2014-01-01

Cytosine methylation and demethylation in tracks of CpG dinucleotides is an epigenetic mechanism for control of gene expression. The initial step in the demethylation process can be deamination of 5-methylcytosine producing the TpG alteration and T:G mispair, and this step is followed by thymine DNA glycosylase (TDG) initiated base excision repair (BER). A further consideration is that guanine in the CpG dinucleotide may become oxidized to 7,8-dihydro-8-oxoguanine (8-oxoG), and this could affect the demethylation process involving TDG-initiated BER. However, little is known about the enzymology of BER of altered in-tandem CpG dinucleotides; e.g. Tp8-oxoG. Here, we investigated interactions between this altered dinucleotide and purified BER enzymes, the DNA glycosylases TDG and 8-oxoG DNA glycosylase 1 (OGG1), apurinic/apyrimidinic (AP) endonuclease 1, DNA polymerase β, and DNA ligases. The overall TDG-initiated BER of the Tp8-oxoG dinucleotide is significantly reduced. Specifically, TDG and DNA ligase activities are reduced by a 3′-flanking 8-oxoG. In contrast, the OGG1-initiated BER pathway is blocked due to the 5′-flanking T:G mispair; this reduces OGG1, AP endonuclease 1, and DNA polymerase β activities. Furthermore, in TDG-initiated BER, TDG remains bound to its product AP site blocking OGG1 access to the adjacent 8-oxoG. These results reveal BER enzyme specificities enabling suppression of OGG1-initiated BER and coordination of TDG-initiated BER at this tandem alteration in the CpG dinucleotide. PMID:24695738

Simultaneous determination of the novel tyrosine kinase inhibitor meditinib and its active metabolite demethylation meditinib in monkey plasma by liquid chromatography-tandem mass spectrometry and its application to pharmacokinetic studies.

PubMed

Liang, Feng; Kong, Qi; Guo, Yongqi; Wang, Yu; Sun, Dejie; Liu, Shi; Cai, Jinling; Guan, Yongbiao; Ding, Rigao

2015-08-01

Meditinib (ME) is a novel tyrosine kinase inhibitor used as an antichronic myeloid leukemia drug. A simple, sensitive and specific LC/MS/MS method was developed and validated for the analysis of ME and its metabolite demethylation meditinib (PI) in monkey plasma using naltrexone as the internal standard. Sample preparation involved protein precipitation with methanol. The analysis was carried out on an Agilent C8 column (3.5 µm, 2.1 × 50 mm). Elution was achieved with a mobile phase gradient varying the proportion of a water solution containing 0.1% formic acid (solvent A) and a 0.1% formic acid in methanol solution (solvent B) at a flow rate of 300 μL/min. The method had a linear calibration curve over the concentration range of 2-1000 ng/mL for ME and 2-1000 ng/mL for PI. The lower limits of quantification of ME and PI were 2 and 2 ng/mL, respectively. The intra- and inter-day precision values were <15% and accuracy values were within ±10.0%. The mean recoveries of ME and PI from plasma were >85%. The assay has been successfully used for pharmacokinetic evaluation of ME and PI using the monkey as an animal model, and those data are reported for the first time. Copyright © 2015 John Wiley & Sons, Ltd.

PI3K/AKT/mTOR Signaling Mediates Valproic Acid-Induced Neuronal Differentiation of Neural Stem Cells through Epigenetic Modifications.

PubMed

Zhang, Xi; He, Xiaosong; Li, Qingqing; Kong, Xuejian; Ou, Zhenri; Zhang, Le; Gong, Zhuo; Long, Dahong; Li, Jianhua; Zhang, Meng; Ji, Weidong; Zhang, Wenjuan; Xu, Liping; Xuan, Aiguo

2017-05-09

Although valproic acid (VPA), has been shown to induce neuronal differentiation of neural stem cells (NSCs), the underlying mechanisms remain poorly understood. Here we investigated if and how mammalian target of rapamycin (mTOR) signaling is involved in the neuronal differentiation of VPA-induced NSCs. Our data demonstrated that mTOR activation not only promoted but also was necessary for the neuronal differentiation of NSCs induced by VPA. We further found that inhibition of mTOR signaling blocked demethylation of neuron-specific gene neurogenin 1 (Ngn1) regulatory element in induced cells. These are correlated with the significant alterations of passive DNA demethylation and the active DNA demethylation pathway in the Ngn1 promoter, but not the suppression of lysine-specific histone methylation and acetylation in the promoter region of Ngn1. These findings highlight a potentially important role for mTOR signaling, by working together with DNA demethylation, to influence the fate of NSCs via regulating the expression of Ngn1 in VPA-induced neuronal differentiation of NSCs. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Epigenetic hierarchy governing Nestin expression.

PubMed

Han, Dong Wook; Do, Jeong Tae; Araúzo-Bravo, Marcos J; Lee, Sung Ho; Meissner, Alexander; Lee, Hoon Taek; Jaenisch, Rudolf; Schöler, Hans R

2009-05-01

Nestin is an intermediate filament protein expressed specifically in neural stem cells and progenitor cells of the central nervous system. DNA demethylation and histone modifications are two types of epigenetic modifications working in a coordinate or synergistic manner to regulate the expression of various genes. This study investigated and elucidated the epigenetic regulation of Nestin gene expression during embryonic differentiation along the neural cell lineage. Nestin exhibits differential DNA methylation and histone acetylation patterns in Nestin-expressing and nonexpressing cells. In P19 embryonic carcinoma cells, activation of Nestin expression is mediated by both trichostatin A and 5-aza-2'-deoxycytidine treatment, concomitant with histone acetylation, but not with DNA demethylation. Nestin transcription is also mediated by treatment with retinoic acid, again in the absence of DNA demethylation. Thus, histone acetylation is sufficient to mediate the activation of Nestin transcription. This study proposed that the regulation of Nestin gene expression can be used as a model to study the epigenetic regulation of gene expression mediated by histone acetylation, but not by DNA demethylation.

Control of DEMETER DNA demethylase gene transcription in male and female gamete companion cells in Arabidopsis thaliana

PubMed Central

Park, Jin-Sup; Frost, Jennifer M.; Park, Kyunghyuk; Ohr, Hyonhwa; Park, Guen Tae; Kim, Seohyun; Eom, Hyunjoo; Lee, Ilha; Brooks, Janie S.; Fischer, Robert L.; Choi, Yeonhee

2017-01-01

The DEMETER (DME) DNA glycosylase initiates active DNA demethylation via the base-excision repair pathway and is vital for reproduction in Arabidopsis thaliana. DME-mediated DNA demethylation is preferentially targeted to small, AT-rich, and nucleosome-depleted euchromatic transposable elements, influencing expression of adjacent genes and leading to imprinting in the endosperm. In the female gametophyte, DME expression and subsequent genome-wide DNA demethylation are confined to the companion cell of the egg, the central cell. Here, we show that, in the male gametophyte, DME expression is limited to the companion cell of sperm, the vegetative cell, and to a narrow window of time: immediately after separation of the companion cell lineage from the germline. We define transcriptional regulatory elements of DME using reporter genes, showing that a small region, which surprisingly lies within the DME gene, controls its expression in male and female companion cells. DME expression from this minimal promoter is sufficient to rescue seed abortion and the aberrant DNA methylome associated with the null dme-2 mutation. Within this minimal promoter, we found short, conserved enhancer sequences necessary for the transcriptional activities of DME and combined predicted binding motifs with published transcription factor binding coordinates to produce a list of candidate upstream pathway members in the genetic circuitry controlling DNA demethylation in gamete companion cells. These data show how DNA demethylation is regulated to facilitate endosperm gene imprinting and potential transgenerational epigenetic regulation, without subjecting the germline to potentially deleterious transposable element demethylation. PMID:28130550

30 CFR 785.22 - In situ processing activities.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 30 Mineral Resources 3 2011-07-01 2011-07-01 false In situ processing activities. 785.22 Section... REGULATORY PROGRAMS REQUIREMENTS FOR PERMITS FOR SPECIAL CATEGORIES OF MINING § 785.22 In situ processing... reclamation operations utilizing in situ processing activities. (b) Any application for a permit for...

30 CFR 785.22 - In situ processing activities.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 30 Mineral Resources 3 2013-07-01 2013-07-01 false In situ processing activities. 785.22 Section... REGULATORY PROGRAMS REQUIREMENTS FOR PERMITS FOR SPECIAL CATEGORIES OF MINING § 785.22 In situ processing... reclamation operations utilizing in situ processing activities. (b) Any application for a permit for...

30 CFR 785.22 - In situ processing activities.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 30 Mineral Resources 3 2012-07-01 2012-07-01 false In situ processing activities. 785.22 Section... REGULATORY PROGRAMS REQUIREMENTS FOR PERMITS FOR SPECIAL CATEGORIES OF MINING § 785.22 In situ processing... reclamation operations utilizing in situ processing activities. (b) Any application for a permit for...

30 CFR 785.22 - In situ processing activities.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 30 Mineral Resources 3 2010-07-01 2010-07-01 false In situ processing activities. 785.22 Section... REGULATORY PROGRAMS REQUIREMENTS FOR PERMITS FOR SPECIAL CATEGORIES OF MINING § 785.22 In situ processing... reclamation operations utilizing in situ processing activities. (b) Any application for a permit for...

30 CFR 785.22 - In situ processing activities.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 30 Mineral Resources 3 2014-07-01 2014-07-01 false In situ processing activities. 785.22 Section... REGULATORY PROGRAMS REQUIREMENTS FOR PERMITS FOR SPECIAL CATEGORIES OF MINING § 785.22 In situ processing... reclamation operations utilizing in situ processing activities. (b) Any application for a permit for...

12 CFR 211.604 - Data processing activities.

Code of Federal Regulations, 2011 CFR

2011-01-01

... processing activities under Regulation K (12 CFR part 211). This question has arisen as a result of the fact... (12 CFR part 225) at that time, as the Regulation K authority permitted limited non-financial data... 12 Banks and Banking 2 2011-01-01 2011-01-01 false Data processing activities. 211.604 Section 211...

12 CFR 211.604 - Data processing activities.

Code of Federal Regulations, 2012 CFR

2012-01-01

... processing activities under Regulation K (12 CFR part 211). This question has arisen as a result of the fact... (12 CFR part 225) at that time, as the Regulation K authority permitted limited non-financial data... 12 Banks and Banking 2 2012-01-01 2012-01-01 false Data processing activities. 211.604 Section 211...

12 CFR 211.604 - Data processing activities.

Code of Federal Regulations, 2013 CFR

2013-01-01

... processing activities under Regulation K (12 CFR part 211). This question has arisen as a result of the fact... (12 CFR part 225) at that time, as the Regulation K authority permitted limited non-financial data... 12 Banks and Banking 2 2013-01-01 2013-01-01 false Data processing activities. 211.604 Section 211...

12 CFR 211.604 - Data processing activities.

Code of Federal Regulations, 2014 CFR

2014-01-01

... processing activities under Regulation K (12 CFR part 211). This question has arisen as a result of the fact... (12 CFR part 225) at that time, as the Regulation K authority permitted limited non-financial data... 12 Banks and Banking 2 2014-01-01 2014-01-01 false Data processing activities. 211.604 Section 211...

12 CFR 211.604 - Data processing activities.

Code of Federal Regulations, 2010 CFR

2010-01-01

... processing activities under Regulation K (12 CFR part 211). This question has arisen as a result of the fact... (12 CFR part 225) at that time, as the Regulation K authority permitted limited non-financial data... 12 Banks and Banking 2 2010-01-01 2010-01-01 false Data processing activities. 211.604 Section 211...

Distributions of Sensitivities to Three Sterol Demethylation Inhibitor Fungicides Among Populations of Uncinula necator Sensitive and Resistant to Triadimefon.

PubMed

Erickson, E O; Wilcox, W F

1997-08-01

ABSTRACT Single-conidial isolates of Uncinula necator from (i) a population representing two vineyards with no previous exposure to sterol demethylation inhibitor (DMI) fungicides ("unexposed," n = 77) and (ii) a population representing two vineyards in which powdery mildew was poorly controlled by triadimefon after prolonged DMI use ("selected," n = 82) were assayed to determine distributions of sensitivities to the DMI fungicides triadimenol (the active form of triadimefon), myclobutanil, and fenarimol. Median 50% effective dose (ED(50)) values (micrograms per milliliter) in the selected versus unexposed populations were 0.06 versus 1.9 for triadimenol, 0.03 versus 0.23 for myclobutanil, and 0.03 versus 0.07 for fenarimol, respectively. Isolates were grouped into sensitivity classes according to their ED(50) values, and those from the selected population were categorized as resistant if the frequency of their sensitivity class had increased significantly relative to levels found in the unexposed population (ED(50) values exceeding 0.56, 0.18, and 0.18 mug/ml for triadimenol, myclobutanil, and fenarimol, respectively). Of the 76 isolates defined as resistant to triadimenol, 64% were classified as cross-resistant to myclobutanil, 18% were classified as cross-resistant to fenarimol, and 17% were classified as resistant to all three fungicides; 25% of the isolates classified as resistant to myclobutanil also were classified as resistant to fenarimol. Similar cross-resistance relationships were revealed when all isolates were examined by regressing log ED(50) values for each fungicide against those for the remaining two fungicides to determine the correlation coefficients (e.g., r = 0.85 for triadimenol versus myclobutanil and 0.56 for triadimenol versus fenarimol). The restricted levels of cross-resistance indicated by these data, particularly between fenarimol and the other two fungicides, is in sharp contrast to the high levels of cross-resistance among DMIs

Methyl mercury toxicity in plant cultures: modification of resistance and demethylation by light and/or 2,4-dichlorophenoxyacetic acid

DOE Office of Scientific and Technical Information (OSTI.GOV)

Czuba, M.

1987-04-01

Cultures of Daucus carota, Ca-68-10, and Lactuca sativa, Le-67, were grown at increasing methyl mercury (MeHg) concentrations ranging from initial doses of 0.05 to 5.0 micrograms/ml per day for 4 days with or without 0.15 microgram/ml 2,4-dichlorophenoxyacetic acid (2,4-D) in the presence or absence of light. The presence of 2,4-D interacted with light synergistically in the expression of MeHg toxicity within the whole range of concentrations. Demethylation patterns increased or decreased depending on the species, the 2,4-D concentration in the medium, and methyl mercury concentration used in the treatment. Lettuce was more sensitive to this interaction than carrot. In lettuce,more » the presence of 2,4-D in the light lowered the concentration of total Hg (or MeHg) required to reduce growth by 50%, about 13 times relative to that in the dark (i.e., it sensitized the cells). In the absence of 2,4-D the pattern was reversed. In carrot the pattern was similar but less pronounced. This suggests that, in these cell populations, MeHg toxicity is partly a hormone-mediated and light-sensitive event.« less

Overexpression of Tet3 in donor cells enhances goat somatic cell nuclear transfer efficiency.

PubMed

Han, Chengquan; Deng, Ruizhi; Mao, Tingchao; Luo, Yan; Wei, Biao; Meng, Peng; Zhao, Lu; Zhang, Qing; Quan, Fusheng; Liu, Jun; Zhang, Yong

2018-05-23

Ten-eleven translocation 3 (TET3) mediates active DNA demethylation of paternal genomes during mouse embryonic development. However, the mechanism of DNA demethylation in goat embryos remains unknown. In addition, aberrant DNA methylation reprogramming prevalently occurs in embryos cloned by somatic cell nuclear transfer (SCNT). In this study, we reported that TET3 is a key factor in DNA demethylation in goat pre-implantation embryos. Knockdown of Tet3 hindered DNA demethylation at the two- to four-cell stage in goat embryos and decreased Nanog expression in blastocysts. Overexpression of Tet3 in somatic cells can initiate DNA demethylation, reduce 5-methylcytosine level, increase 5-hydroxymethylcytosine level and promote the expression of key pluripotency genes. After SCNT, overexpression of Tet3 in donor cells corrected abnormal DNA hypermethylation of cloned embryos and significantly enhanced in vitro and in vivo developmental rate (P < 0.05). We conclude that overexpression of Tet3 in donor cells significantly improves goat SCNT efficiency. © 2018 Federation of European Biochemical Societies.

Discovery of gemfibrozil analogues that activate PPARα and enhance the expression of gene CPT1A involved in fatty acids catabolism.

PubMed

De Filippis, Barbara; Giancristofaro, Antonella; Ammazzalorso, Alessandra; D'Angelo, Alessandra; Fantacuzzi, Marialuigia; Giampietro, Letizia; Maccallini, Cristina; Petruzzelli, Michele; Amoroso, Rosa

2011-10-01

A new series of gemfibrozil analogues conjugated with α-asarone, trans-stilbene, chalcone, and their bioisosteric modifications were synthesized and evaluated to develop PPARα agonists. In this attempt, we have removed the methyls on the phenyl ring of gemfibrozil and introduced the above scaffolds in para position synthesizing two series of derivatives, keeping the dimethylpentanoic skeleton of gemfibrozil unaltered or demethylated. Four compounds exhibited good activation of the PPARα receptor and were also screened for their activity on PPARα-regulated gene CPT1A. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Spectrum of the Reductive Dehalogenation Activity of Desulfitobacterium frappieri PCP-1

PubMed Central

Dennie, D.; Gladu, I.; Lépine, F.; Villemur, R.; Bisaillon, J.-G.; Beaudet, R.

1998-01-01

Desulfitobacterium frappieri PCP-1 was induced for ortho- and para-dechlorinating activities by different chlorophenols. Dehalogenation rates ranging from 25 to 1,158 nmol/min/mg of cell protein were observed according to the chlorophenol tested and the position of the chlorine removed. D. frappieri shows a broad substrate specificity; in addition to tetrachloroethylene and pentachloropyridine, strain PCP-1 can dehalogenate at ortho, meta, and para positions a large variety of aromatic molecules with substituted hydroxyl or amino groups. Reactions of O demethylation and reduction of nitro to amino substituents on aromatic molecules were also observed. PMID:9797330

Expression of ZmMET1, a gene encoding a DNA methyltransferase from maize, is associated not only with DNA replication in actively proliferating cells, but also with altered DNA methylation status in cold-stressed quiescent cells.

PubMed

Steward, N; Kusano, T; Sano, H

2000-09-01

A cDNA fragment encoding part of a DNA methyltransferase was isolated from maize. The putative amino acid sequence identically matched that deduced from a genomic sequence in the database (accession no. AF063403), and the corresponding gene was designated as ZmMET1. Bacterially expressed ZmMET1 actively methylated DNA in vitro. Transcripts of ZmMET1 could be shown to exclusively accumulate in actively proliferating cells of the meristems of mesocotyls and root apices, suggesting ZmMET1 expression to be associated with DNA replication. This was confirmed by simultaneous decrease of transcripts of ZmMET1 and histone H3, a marker for DNA replication, in seedlings exposed to wounding, desiccation and salinity, all of which suppress cell division. Cold stress also depressed both transcripts in root tissues. In contrast, however, accumulation of ZmMET1 transcripts in shoot mesocotyls was not affected by cold stress, whereas those for H3 sharply decreased. Such a differential accumulation of ZmMET1 transcripts was consistent with ZmMET1 protein levels as revealed by western blotting. Expression of ZmMET1 is thus coexistent, but not completely dependent on DNA replication. Southern hybridization analysis with a methylation-sensitive restriction enzyme revealed that cold treatment induced demethylation of DNA in the Ac/Ds transposon region, but not in other genes, and that such demethylation primarily occurred in roots. These results suggested that the methylation level was decreased selectively by cold treatment, and that ZmMET1 may, at least partly, prevent such demethylation.

Effects of thermal processing and various chemical substances on formaldehyde and dimethylamine formation in squid Dosidicus gigas.

PubMed

Zhu, Junli; Li, Jianrong; Jia, Jia

2012-09-01

Trimethylamine oxide (TMAO) in squid is demethylated to dimethylamine (DMA) and formaldehyde (FA) during storage and processing. This study examined the effects of thermal processing and various chemical substances on FA and DMA formation in squid. The thermal conversion of TMAO was assessed by analysing four squid and four gadoid fish species, which revealed that FA, DMA and trimethylamine (TMA) were gradually produced in squid, whereas TMA increased and FA decreased in gadoid fish. A significant increase in both FA and DMA levels was observed in the supernatant of jumbo squid with increased heating temperature and extended heating time at pH 6-7. Ferrous chloride combined with cysteine and/or ascorbate had a significantly positive effect on FA formation in the heated supernatant of jumbo squid. No significant difference was observed in the levels of Cu and Fe in squid and gadoid fish. The capability of Fe(2+) to promote the formation of FA and DMA was not completely attributable to its reducing power in squid. Non-enzymatic decomposition of TMAO was a key pathway during the thermal processing of jumbo squid, and Fe(2+) was a crucial activator in the formation of FA and DMA. Copyright © 2012 Society of Chemical Industry.

CD137+CD154- Expression As a Regulatory T Cell (Treg)-Specific Activation Signature for Identification and Sorting of Stable Human Tregs from In Vitro Expansion Cultures.

PubMed

Nowak, Anna; Lock, Dominik; Bacher, Petra; Hohnstein, Thordis; Vogt, Katrin; Gottfreund, Judith; Giehr, Pascal; Polansky, Julia K; Sawitzki, Birgit; Kaiser, Andrew; Walter, Jörn; Scheffold, Alexander

2018-01-01

Regulatory T cells (Tregs) are an attractive therapeutic tool for several different immune pathologies. Therapeutic Treg application often requires prolonged in vitro culture to generate sufficient Treg numbers or to optimize their functionality, e.g., via genetic engineering of their antigen receptors. However, purity of clinical Treg expansion cultures is highly variable, and currently, it is impossible to identify and separate stable Tregs from contaminating effector T cells, either ex vivo or after prior expansion. This represents a major obstacle for quality assurance of expanded Tregs and raises significant safety concerns. Here, we describe a Treg activation signature that allows identification and sorting of epigenetically imprinted Tregs even after prolonged in vitro culture. We show that short-term reactivation resulted in expression of CD137 but not CD154 on stable FoxP3+ Tregs that displayed a demethylated Treg-specific demethylated region, high suppressive potential, and lack of inflammatory cytokine expression. We also applied this Treg activation signature for rapid testing of chimeric antigen receptor functionality in human Tregs and identified major differences in the signaling requirements regarding CD137 versus CD28 costimulation. Taken together, CD137+CD154- expression emerges as a universal Treg activation signature ex vivo and upon in vitro expansion allowing the identification and isolation of epigenetically stable antigen-activated Tregs and providing a means for their rapid functional testing in vitro .

Potential Active Processes in Porter Crater

NASA Image and Video Library

2015-07-15

The extended-mission status of the Mars Reconnaissance Orbiter and the HiRISE camera has greatly increased our understanding of numerous active processes occurring on Mars. By taking carefully planned repeat images of surface, we now have an important record of how the surface evolves for a maximum of 5 Mars years. This image shows the central peak in Porter Crater. Although there are no repeat images here we can infer several active geologic processes, based on morphologic evidence and lessons learned from past well-monitored sites. Shallow gullies are located on the south and east facing slopes of the central peak. These features might have been carved by volatiles, such as carbon dioxide frost, sometime in the recent geologic past. Meanwhile on the northern slopes are several smaller slope features that have a morphology hinting at recurring slope lineae (RSL). Alternatively, these features could be the remnants of past active gullies. Several more HiRISE images would be needed to characterize their behavior and confirm their status as RSL (see "Recurring Slope Lineae in Equatorial Regions of Mars"). Southward on the slopes below the peak is a large dune field. Dunes show sharp crests with prominent ripples, both signs of actively migrating dunes. Also, we can see dust devil tracks crossing the nearby dusty surfaces and clear evidence for ongoing modification by swirling winds that persistently remove surface dust. Ongoing operations by HiRISE are dedicated to studying all of the active surface and atmospheric processes operating on Mars. http://photojournal.jpl.nasa.gov/catalog/PIA19853

CD137+CD154− Expression As a Regulatory T Cell (Treg)-Specific Activation Signature for Identification and Sorting of Stable Human Tregs from In Vitro Expansion Cultures

PubMed Central

Nowak, Anna; Lock, Dominik; Bacher, Petra; Hohnstein, Thordis; Vogt, Katrin; Gottfreund, Judith; Giehr, Pascal; Polansky, Julia K.; Sawitzki, Birgit; Kaiser, Andrew; Walter, Jörn; Scheffold, Alexander

2018-01-01

Regulatory T cells (Tregs) are an attractive therapeutic tool for several different immune pathologies. Therapeutic Treg application often requires prolonged in vitro culture to generate sufficient Treg numbers or to optimize their functionality, e.g., via genetic engineering of their antigen receptors. However, purity of clinical Treg expansion cultures is highly variable, and currently, it is impossible to identify and separate stable Tregs from contaminating effector T cells, either ex vivo or after prior expansion. This represents a major obstacle for quality assurance of expanded Tregs and raises significant safety concerns. Here, we describe a Treg activation signature that allows identification and sorting of epigenetically imprinted Tregs even after prolonged in vitro culture. We show that short-term reactivation resulted in expression of CD137 but not CD154 on stable FoxP3+ Tregs that displayed a demethylated Treg-specific demethylated region, high suppressive potential, and lack of inflammatory cytokine expression. We also applied this Treg activation signature for rapid testing of chimeric antigen receptor functionality in human Tregs and identified major differences in the signaling requirements regarding CD137 versus CD28 costimulation. Taken together, CD137+CD154− expression emerges as a universal Treg activation signature ex vivo and upon in vitro expansion allowing the identification and isolation of epigenetically stable antigen-activated Tregs and providing a means for their rapid functional testing in vitro. PMID:29467769

Are 8-oxoguanine (8-oxoGua) and 5-hydroxymethyluracil (5-hmUra) oxidatively damaged DNA bases or transcription (epigenetic) marks?

PubMed

Zarakowska, Ewelina; Gackowski, Daniel; Foksinski, Marek; Olinski, Ryszard

2014-04-01

The oxidatively modified DNA base 8-oxo-7,8-dihydroguanine (8-oxoGua) is nontoxic and weakly mutagenic. Here we report on new data suggesting a potential for 8-oxoGua to affect the expression of several genes via epigenetic changes resulting in chromatin relaxation. Using pig thymus extract, we analyzed the distribution of 8-oxoGua among different nuclei fractions representative of transcriptionally active and silenced regions. The levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) found in transcriptionally active euchromatin (4.37/10(6) nucleotides) and in the matrix fraction (4.16/10(6) nucleotides) were about 5 times higher than in transcriptionally silenced heterochromatin (0.91/10(6) nucleotides). Other experimental data are presented which suggest that 8-oxoGua present in specific DNA sequences may be widely used for transcription regulation. Like 8-oxoGua, 5-hydroxymethyluracil (5-hmUra) is another oxidatively modified DNA base (the derivative is formed by thymine oxidation). Recent experimental evidence supports the notion that 5-hmUra plays an important role in active DNA demethylation. This involves overexpression of activation-induced cytidine deaminase (AID) and ten-eleven translocation 1 (TET1) protein (the key proteins involved in active demethylation), which leads to global accumulation of 5-hmUra. Our preliminary data demonstrate a significant increase of the 5-hmUra levels in pig brain extract when compared with liver extract. The lack of 5-hmUra in Escherichia coli DNA also speaks for a role of this modification in the active demethylation process. It is concluded that 8-oxodG and 5-hmUra in DNA may be considered as epigenetic marks. Copyright © 2013 Elsevier B.V. All rights reserved.

Zinc deficiency enhanced inflammatory response by increasing immune cell activation and inducing IL6 promoter demethylation

PubMed Central

Wong, Carmen P.; Rinaldi, Nicole A.; Ho, Emily

2015-01-01

Scope Zinc deficiency results in immune dysfunction and promotes systemic inflammation. The objective of this study was to examine the effects of zinc deficiency on cellular immune activation and epigenetic mechanisms that promote inflammation. This work is potentially relevant to the aging population given that age-related immune defects, including chronic inflammation, coincide with declining zinc status. Methods and results An in vitro cell culture system and the aged mouse model were used to characterize immune activation and DNA methylation profiles that may contribute to the enhanced proinflammatory response mediated by zinc deficiency. Zinc deficiency up-regulated cell activation markers ICAM1, MHC class II, and CD86 in THP1 cells, that coincided with increased IL1β and IL6 responses following LPS stimulation. A decreased zinc status in aged mice was similarly associated with increased ICAM1 and IL6 gene expression. Reduced IL6 promoter methylation was observed in zinc deficient THP1 cells, as well as in aged mice and human lymphoblastoid cell lines derived from aged individuals. Conclusion Zinc deficiency induced inflammatory response in part by eliciting aberrant immune cell activation and altered promoter methylation. Our results suggested potential interactions between zinc status, epigenetics, and immune function, and how their dysregulation could contribute to chronic inflammation. PMID:25656040

Process of activation of a palladium catalyst system

DOEpatents

Sobolevskiy, Anatoly [Orlando, FL; Rossin, Joseph A [Columbus, OH; Knapke, Michael J [Columbus, OH

2011-08-02

Improved processes for activating a catalyst system used for the reduction of nitrogen oxides are provided. In one embodiment, the catalyst system is activated by passing an activation gas stream having an amount of each of oxygen, water vapor, nitrogen oxides, and hydrogen over the catalyst system and increasing a temperature of the catalyst system to a temperature of at least 180.degree. C. at a heating rate of from 1-20.degree./min. Use of activation processes described herein leads to a catalyst system with superior NOx reduction capabilities.

Importance of sulfate reducing bacteria in mercury methylation and demethylation in periphyton from Bolivian Amazon region.

PubMed

Achá, Darío; Hintelmann, Holger; Yee, Janet

2011-02-01

Sulfate reducing bacteria (SRB) are important mercury methylators in sediments, but information on mercury methylators in other compartments is ambiguous. To investigate SRB involvement in methylation in Amazonian periphyton, the relationship between Hg methylation potential and SRB (Desulfobacteraceae, Desulfobulbaceae and Desulfovibrionaceae) abundance in Eichhornia crassipes and Polygonum densiflorum root associated periphyton was examined. Periphyton subsamples of each macrophyte were amended with electron donors (lactate, acetate and propionate) or inhibitors (molybdate) of sulfate reduction to create differences in SRB subgroup abundance, which was measured by quantitative real-time PCR with primers specific for the 16S rRNA gene. Mercury methylation and demethylation potentials were determined by a stable isotope tracer technique using 200HgCl and CH3(202)HgCl, respectively. Relative abundance of Desulfobacteraceae (<0.01-12.5%) and Desulfovibrionaceae (0.01-6.8%) were both highly variable among samples and subsamples, but a significant linear relationship (p<0.05) was found between Desulfobacteraceae abundance and net methylmercury formation among treatments of the same macrophyte periphyton and among all P. densiflorum samples, suggesting that Desulfobacteraceae bacteria are the most important mercury methylators among SRB families. Yet, molybdate only partially inhibited mercury methylation potentials, suggesting the involvement of other microorganisms as well. The response of net methylmercury production to the different electron donors and molybdate was highly variable (3-1104 pg g(-1) in 12 h) among samples, as was the net formation in control samples (17-164 pg g(-1) in 12 h). This demonstrates the importance of community variability and complexity of microbial interactions for the overall methylmercury production in periphyton and their response to external stimulus. Copyright © 2010 Elsevier Ltd. All rights reserved.

Measurement of action spectra of light-activated processes

NASA Astrophysics Data System (ADS)

Ross, Justin; Zvyagin, Andrei V.; Heckenberg, Norman R.; Upcroft, Jacqui; Upcroft, Peter; Rubinsztein-Dunlop, Halina H.

2006-01-01

We report on a new experimental technique suitable for measurement of light-activated processes, such as fluorophore transport. The usefulness of this technique is derived from its capacity to decouple the imaging and activation processes, allowing fluorescent imaging of fluorophore transport at a convenient activation wavelength. We demonstrate the efficiency of this new technique in determination of the action spectrum of the light mediated transport of rhodamine 123 into the parasitic protozoan Giardia duodenalis.

Transcriptional activation of transposable elements in mouse zygotes is independent of Tet3-mediated 5-methylcytosine oxidation.

PubMed

Inoue, Azusa; Matoba, Shogo; Zhang, Yi

2012-12-01

The methylation state of the paternal genome is rapidly reprogrammed shortly after fertilization. Recent studies have revealed that loss of 5-methylcytosine (5mC) in zygotes correlates with appearance of 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). This process is mediated by Tet3 and the 5mC oxidation products generated in zygotes are gradually lost during preimplantation development through a replication-dependent dilution process. Despite these findings, the biological significance of Tet3-mediated oxidation of 5mC to 5hmC/5fC/5caC in zygotes is unknown. DNA methylation plays an important role in silencing gene expression including the repression of transposable elements (TEs). Given that the activation of TEs during preimplantation development correlates with loss of DNA methylation, it is believed that paternal DNA demethylation may have an important role in TE activation. Here we examined this hypothesis and found that Tet3-mediated 5mC oxidation does not have a significant contribution to TE activation. We show that the expression of LINE-1 (long interspersed nucleotide element 1) and ERVL (endogenous retroviruses class III) are activated from both paternal and maternal genomes in zygotes. Inhibition of 5mC oxidation by siRNA-mediated depletion of Tet3 affected neither TE activation, nor global transcription in zygotes. Thus, our study provides the first evidence demonstrating that activation of both TEs and global transcription in zygotes are independent of Tet3-mediated 5mC oxidation.

Stress enhances the gene expression and enzyme activity of phenylalanine ammonia-lyase and the endogenous content of salicylic acid to induce flowering in pharbitis.

PubMed

Wada, Kaede C; Mizuuchi, Kaori; Koshio, Aya; Kaneko, Kentaro; Mitsui, Toshiaki; Takeno, Kiyotoshi

2014-07-01

The involvement of salicylic acid (SA) in the regulation of stress-induced flowering in the short-day plant pharbitis (also called Japanese morning glory) Ipomoea nil (formerly Pharbitis nil) was studied. Pharbitis cv. Violet was induced to flower when grown in 1/100-strength mineral nutrient solution under non-inductive long-day conditions. All fully expanded true leaves were removed from seedlings, leaving only the cotyledons, and flowering was induced under poor-nutrition stress conditions. This indicates that cotyledons can play a role in the regulation of poor-nutrition stress-induced flowering. The expression of the pharbitis homolog of PHENYLALANINE AMMONIA-LYASE, the enzyme activity of phenylalanine ammonia-lyase (PAL; E.C. 4.3.1.5) and the content of SA in the cotyledons were all up-regulated by the stress treatment. The Violet was also induced to flower by low-temperature stress, DNA demethylation and short-day treatment. Low-temperature stress enhanced PAL activity, whereas non-stress factors such as DNA demethylation and short-day treatment decreased the activity. The PAL enzyme activity was also examined in another cultivar, Tendan, obtaining similar results to Violet. The exogenously applied SA did not induce flowering under non-stress conditions but did promote flowering under weak stress conditions in both cultivars. These results suggest that stress-induced flowering in pharbitis is induced, at least partly, by SA, and the synthesis of SA is promoted by PAL. Copyright © 2014 Elsevier GmbH. All rights reserved.

Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

PubMed

Tampe, Björn; Steinle, Ulrike; Tampe, Désirée; Carstens, Julienne L; Korsten, Peter; Zeisberg, Elisabeth M; Müller, Gerhard A; Kalluri, Raghu; Zeisberg, Michael

2017-01-01

Acute kidney injury (AKI) and progressive chronic kidney disease (CKD) are intrinsically tied syndromes. In this regard, the acutely injured kidney often does not achieve its full regenerative capacity and AKI directly transitions into progressive CKD associated with tubulointerstitial fibrosis. Underlying mechanisms of such AKI-to-CKD progression are still incompletely understood and specific therapeutic interventions are still elusive. Because epigenetic modifications play a role in maintaining tissue fibrosis, we used a murine model of ischemia-reperfusion injury to determine whether aberrant promoter methylation of RASAL1 contributes causally to the switch between physiological regeneration and tubulointerstitial fibrogenesis, a hallmark of AKI-to-CKD progression. It is known that the antihypertensive drug hydralazine has demethylating activity, and that its optimum demethylating activity occurs at concentrations below blood pressure-lowering doses. Administration of low-dose hydralazine effectively induced expression of hydroxylase TET3, which catalyzed RASAL1 hydroxymethylation and subsequent RASAL1 promoter demethylation. Hydralazine-induced CpG promoter demethylation subsequently attenuated renal fibrosis and preserved excretory renal function independent of its blood pressure-lowering effects. In comparison, RASAL1 demethylation and inhibition of tubulointerstitial fibrosis was not detected upon administration of the angiotensin-converting enzyme inhibitor Ramipril in this model. Thus, RASAL1 promoter methylation and subsequent transcriptional RASAL1 suppression plays a causal role in AKI-to-CKD progression. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

15 CFR 400.33 - Restrictions on manufacturing and processing activity.

Code of Federal Regulations, 2010 CFR

2010-01-01

...-TRADE ZONES BOARD Manufacturing and Processing Activity-Reviews § 400.33 Restrictions on manufacturing and processing activity. (a) In general. In approving manufacturing or processing activity for a zone... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Restrictions on manufacturing and...

Cold acclimation alters DNA methylation patterns and confers tolerance to heat and increases growth rate in Brassica rapa

PubMed Central

Liu, Tongkun; Li, Ying; Duan, Weike; Huang, Feiyi

2017-01-01

Abstract Epigenetic modifications are implicated in plant adaptations to abiotic stresses. Exposure of plants to one stress can induce resistance to other stresses, a process termed cross-adaptation, which is not well understood. In this study, we aimed to unravel the epigenetic basis of elevated heat-tolerance in cold-acclimated Brassica rapa by conducting a genome-wide DNA methylation analysis of leaves from control (CK) and cold-acclimated (CA) plants. We found that both methylation and demethylation occurred during cold acclimation. Two significantly altered pathways, malate dehydrogenase activity and carbon fixation, and 1562 differentially methylated genes, including BramMDH1, BraKAT2, BraSHM4, and Bra4CL2, were identified in CA plants. Genetic validation and treatment of B. rapa with 5-aza-2-deoxycytidine (Aza) suggested that promoter demethylation of four candidate genes increased their transcriptional activities. Physiological analysis suggested that elevated heat-tolerance and high growth rate were closely related to increases in organic acids and photosynthesis, respectively. Functional analyses demonstrated that the candidate gene BramMDH1 (mMDH: mitochondrial malate dehydrogenase) directly enhances organic acids and photosynthesis to increase heat-tolerance and growth rate in Arabidopsis. However, Aza-treated B. rapa, which also has elevated BramMDH1 levels, did not exhibit enhanced heat-tolerance. We therefore suggest that DNA demethylation alone is not sufficient to increase heat-tolerance. This study demonstrates that altered DNA methylation contributes to cross-adaptation. PMID:28158841

Methylation of an intragenic alternative promoter regulates transcription of GARP.

PubMed

Haupt, Sonja; Söntgerath, Viktoria Sophie Apollonia; Leipe, Jan; Schulze-Koops, Hendrik; Skapenko, Alla

2016-02-01

Alternative promoter usage has been proposed as a mechanism regulating transcriptional and translational diversity in highly elaborated systems like the immune system in humans. Here, we report that transcription of human glycoprotein A repetitions predominant (GARP) in regulatory CD4 T cells (Tregs) is tightly regulated by two alternative promoters. An intragenic promoter contains several CpGs and acts as a weak promoter that is demethylated and initiates transcription Treg-specifically. The strong up-stream promoter containing a CpG-island is, in contrast, fully demethylated throughout tissues. Transcriptional activity of the strong promoter was surprisingly down-regulated upon demethylation of the weak promoter. This demethylation-induced transcriptional attenuation regulated the magnitude of GARP expression and correlated with disease activity in rheumatoid arthritis. Treg-specific GARP transcription was initiated by synergistic interaction of forkhead box protein 3 (Foxp3) with nuclear factor of activated T cells (NFAT) and was underpinned by permissive chromatin remodeling caused by release of the H3K4 demethylase, PLU-1. Our findings describe a novel function of alternative promoters in regulating the extent of transcription. Moreover, since GARP functions as a transporter of transforming growth factor β (TGFβ), a cytokine with broad pleiotropic traits, GARP transcriptional attenuation by alternative promoters might provide a mechanism regulating peripheral TGFβ to avoid unwanted harmful effects. Copyright © 2015 Elsevier B.V. All rights reserved.

Ethanol-induced changes in Poly (ADP ribose) Polymerase and neuronal developmental gene expression

PubMed Central

Gavin, David P.; Kusumo, Handojo; Sharma, Rajiv P.; Guizzetti, Marina

2016-01-01

Prenatal alcohol exposure has profound effects on neuronal growth and development. Poly-ADP Ribose Polymerase (PARP) enzymes are perhaps unique in the field of epigenetics in that they directly participate in histone modifications, transcription factor modifications, DNA methylation/demethylation and are highly inducible by ethanol. It was our hypothesis that ethanol would induce PARP enzymatic activity leading to alterations in neurodevelopmental gene expression. Mouse E18 cortical neurons were treated with ethanol, PARP inhibitors, and nuclear hormone receptor transcription factor PPARγ agonists and antagonists. Subsequently, we measured PARP activity and changes in Bdnf, OKSM (Oct4, Klf4, Sox2, c-Myc), DNA methylating/demethylating factors, and Pparγ mRNA expression, promoter 5-methylcytosine (5MC) and 5-hydroxymethylcytosine (5HMC), and PPARγ promoter binding. We found that ethanol reduced Bdnf4, 9a, and Klf4 mRNA expression, and increased c-Myc expression. These changes were reversed with a PARP inhibitor. In agreement with its role in DNA demethylation PARP inhibition increased 5MC levels at the c-Myc promoter. In addition, we found that elevated PARP enzymatic activity reduced PPARγ promoter binding, and this corresponded to decreased Bdnf and Klf4 mRNA expression. Our results suggest that PARP participates in DNA demethylation and reduces PPARγ promoter binding. The current study underscores the importance of PARP in ethanol-induced changes to neurodevelopmental gene expression. PMID:27497606

The Histone Demethylase Jhdm1a Regulates Hepatic Gluconeogenesis

PubMed Central

Zou, Tie; Yao, Annie Y.; Cooper, Marcus P.; Boyartchuk, Victor; Wang, Yong-Xu

2012-01-01

Hepatic gluconeogenesis is required for maintaining blood glucose homeostasis; yet, in diabetes mellitus, this process is unrestrained and is a major contributor to fasting hyperglycemia. To date, the impacts of chromatin modifying enzymes and chromatin landscape on gluconeogenesis are poorly understood. Through catalyzing the removal of methyl groups from specific lysine residues in the histone tail, histone demethylases modulate chromatin structure and, hence, gene expression. Here we perform an RNA interference screen against the known histone demethylases and identify a histone H3 lysine 36 (H3K36) demethylase, Jhdm1a, as a key negative regulator of gluconeogenic gene expression. In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level. Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity. Mechanistically, we find that Jhdm1a regulates the expression of a major gluconeogenic regulator, C/EBPα. This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus. Our work provides compelling evidence that links histone demethylation to transcriptional regulation of gluconeogenesis and has important implications for the treatment of diabetes. PMID:22719268

Method for recovering and using lignin in adhesive resins

DOEpatents

Schroeder, Herbert A.

1993-01-01

Lignin, or a lignin derived material, which has been significantly demethylated (e.g., the demethylated lignin found in the raffinate produced as a by-product of dimethyl sulfide production which can be carried out using the spent liquor from wood pulping operations) can be isolated by a process wherein an organic solvent is added to a lignin-containing aqueous solution. The organic solvent is typically a polar, and at least a partially water-immiscible substance such as, for example, ethyl acetate. The resulting lignin-containing aqueous solution/organic solvent mixture is acidified to produce a water layer which is discarded and an organic solvent layer which contains the demethylated lignin. Upon its recovery, the demethylated lignin is dissolved in an alkaline solution to which an aldehyde source is added to produce a resol-type resin. The aldehyde source may be formaldehyde in solution, paraformaldehyde, hexamethylenetetramine, or other aldehydes including acetaldehyde, furfural, and their derivatives.

Degradation of crystal violet by an FeGAC/H2O2 process.

PubMed

Chen, Chiing-Chang; Chen, Wen-Ching; Chiou, Mei-Rung; Chen, Sheng-Wei; Chen, Yao Yin; Fan, Huan-Jung

2011-11-30

Because of the growing concern over highly contaminated crystal violet (CV) wastewater, an FeGAC/H(2)O(2) process was employed in this research to treat CV-contaminated wastewater. The experimental results indicated that the presence of iron oxide-coated granular activated carbon (FeGAC) greatly improved the oxidative ability of H(2)O(2) for the removal of CV. For instance, the removal efficiencies of H(2)O(2), GAC, FeGAC, GAC/H(2)O(2) and FeGAC/H(2)O(2) processes were 10%, 44%, 40%, 43% and 71%, respectively, at test conditions of pH 3 and 7.4mM H(2)O(2). FeGAC/H(2)O(2) combined both the advantages of FeGAC and H(2)O(2). FeGAC had a good CV adsorption ability and could effectively catalyze the hydrogen peroxide oxidation reaction. Factors (including pH, FeGAC dosage and H(2)O(2) dosage) affecting the removal of CV by FeGAC/H(2)O(2) were investigated in this research as well. In addition, the reaction intermediates were separated and identified using HPLC-ESI-MS. The N-demethylation step might be the main reaction pathway for the removal of CV. The reaction mechanisms for the process proposed in this research might be useful for future application of this technology to the removal of triphenylmethane (TPM) dyes. Copyright © 2011 Elsevier B.V. All rights reserved.

Effect of diethyldithiocarbamate (DDC) and ticlopidine on CYP1A2 activity and caffeine metabolism: an in vitro comparative study with human cDNA-expressed CYP1A2 and liver microsomes.

PubMed

Kot, Marta; Daniel, Władysława A

2009-01-01

The aim of the present study was to test the effect of diethyldithiocarbamate (DDC), which is regarded as a cytochrome P450 (CYP) CYP2A6 and CYP2E1 inhibitor, and ticlopidine, an efficient CYP2B6, CYP2C19 and CYP2D6 inhibitor, on the activity of human CYP1A2 and the metabolism of caffeine (1-N-, 3-N- and 7-N-demethylation, and C-8-hydroxylation). The experiment was carried out in vitro using human cDNA-expressed CYP1A2 (Supersomes) and human pooled liver microsomes. The effects of DDC and ticlopidine were compared to those of furafylline (a strong CYP1A2 inhibitor). A comparative in vitro study provides clear evidence that ticlopidine and DDC, applied at concentrations that inhibit the above-mentioned CYP isoforms, potently (as compared to furafylline) inhibit human CYP1A2 and caffeine metabolism, in particular 1-N- and 3-N-demethylation.

Active non-volatile memory post-processing

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kannan, Sudarsun; Milojicic, Dejan S.; Talwar, Vanish

A computing node includes an active Non-Volatile Random Access Memory (NVRAM) component which includes memory and a sub-processor component. The memory is to store data chunks received from a processor core, the data chunks comprising metadata indicating a type of post-processing to be performed on data within the data chunks. The sub-processor component is to perform post-processing of said data chunks based on said metadata.

An epigenetic view of developmental diseases: new targets, new therapies.

PubMed

Xie, Pei; Zang, Li-Qun; Li, Xue-Kun; Shu, Qiang

2016-08-01

Function of epigenetic modifications is one of the most competitive fields in life science. Over the past several decades, it has been revealed that epigenetic modifications play essential roles in development and diseases including developmental diseases. In the present review, we summarize the recent progress about the function of epigenetic regulation, especially DNA and RNA modifications in developmental diseases. Original research articles and literature reviews published in PubMed-indexed journals. DNA modifications including methylation and demethylation can regulate gene expression, and are involved in development and multiple diseases including Rett syndrome, Autism spectrum disorders, congenital heart disease and cancer, etc. RNA methylation and demethylation play important roles in RNA processing, reprogramming, circadian, and neuronal activity, and then modulate development. DNA and RNA modifications play important roles in development and diseases through regulating gene expression. Epigenetic components could serve as novel targets for the treatment of developmental diseases.

Effects of Conazole Fungicides on Spontaneous Activity in Neural Networks

EPA Science Inventory

Hexaconazole (HEX), Tetraconazole (TET), Fluconazole (FLU), and Triadimefon (TRI) are conazole fungicides, used to control powdery mildews on crops, and as veterinary and clinical treatments. TRI, a demethylation inhibitor, is neurotoxic in vivo, and previous in vitro experiments...

Speech perception as an active cognitive process

PubMed Central

Heald, Shannon L. M.; Nusbaum, Howard C.

2014-01-01

One view of speech perception is that acoustic signals are transformed into representations for pattern matching to determine linguistic structure. This process can be taken as a statistical pattern-matching problem, assuming realtively stable linguistic categories are characterized by neural representations related to auditory properties of speech that can be compared to speech input. This kind of pattern matching can be termed a passive process which implies rigidity of processing with few demands on cognitive processing. An alternative view is that speech recognition, even in early stages, is an active process in which speech analysis is attentionally guided. Note that this does not mean consciously guided but that information-contingent changes in early auditory encoding can occur as a function of context and experience. Active processing assumes that attention, plasticity, and listening goals are important in considering how listeners cope with adverse circumstances that impair hearing by masking noise in the environment or hearing loss. Although theories of speech perception have begun to incorporate some active processing, they seldom treat early speech encoding as plastic and attentionally guided. Recent research has suggested that speech perception is the product of both feedforward and feedback interactions between a number of brain regions that include descending projections perhaps as far downstream as the cochlea. It is important to understand how the ambiguity of the speech signal and constraints of context dynamically determine cognitive resources recruited during perception including focused attention, learning, and working memory. Theories of speech perception need to go beyond the current corticocentric approach in order to account for the intrinsic dynamics of the auditory encoding of speech. In doing so, this may provide new insights into ways in which hearing disorders and loss may be treated either through augementation or therapy. PMID

Enantiomeric separation of some demethylated analogues of clofibric acid by capillary zone electrophoresis and nano-liquid chromatography.

PubMed

Fantacuzzi, Marialuigia; Bettoni, Giancarlo; D'Orazio, Giovanni; Fanali, Salvatore

2006-03-01

The enantiomeric separation of some demethylated analogues of clofibric acid, namely 2-(6-chloro-benzothiazol-2-ylsulfanyl)-, 2-(6-methoxy-benzothiazol-2-ylsulfanyl)-, 2-(quinolin-2-yloxy)-, 2-(6-chloro-quinolin-2-yloxy)-, 2-(7-chloro-quinolin-4-yloxy)-propionic acid (compounds A-E, respectively), has been studied by CZE and nano-LC using for the first technique two beta-CD derivatives and vancomycin added to the BGE and vancomycin-modified silica particles for the second one, with the aim to find the optimum experimental conditions for the baseline resolution. The type and the concentration of the chiral selector added to the BGE, the buffer pH, the type of organic modifier and its concentration, the capillary temperature and the applied voltage played a very important role in the enantioresolution of the analysed compounds. The use of 6-monodeoxy-6-monoamino-beta-CD allowed to achieve baseline resolution of four of five clofibric acid derivatives in less than 10 min while heptakis-(2,3,6-tri-O-methyl)-beta-CD partially resolved the same compounds in their enantiomers. Employing vancomycin as the chiral selector in CZE, the counter-current partial filling method was chosen achieving baseline resolution of four analytes. All the studied compounds were enantioresolved employing a capillary column packed with vancomycin stationary phase by nano-LC, and the resolution was strongly influenced by the concentration of the organic modifier and by the pH of the mobile phase. The best results were achieved at pH 4.5 in presence of 60% of methanol (MeOH). However, longer analysis times were observed in the experiments carried out by nano-LC.

[Experimental rationale for the parameters of a rapid method for oxidase activity determination].

PubMed

Butorina, N N

2010-01-01

Experimental rationale is provided for the parameters of a rapid (1-2-min) test to concurrently determine the oxidase activity of all bacteria grown on the membrane filter after water filtration. Oxidase reagents that are the aqueous solutions of tetramethyl-p-phenylenediamine dihydrochloride and demethyl-p-phenylenediamine dihydrochloride have been first ascertained to exert no effect on the viability and enzymatic activity of bacteria after one-hour contact. An algorithm has been improved for the rapid oxidase activity test: the allowable time for bacteria to contact oxidase reagents and procedures for minimizing the effect on bacterial biochemical activity following the contact. An accelerated method based on lactose medium with tergitol 7 and Endo agar has been devised to determine coliform bacteria, by applying the rapid oxidase test: the time of a final response is 18-24 hours. The method has been included into GOST 52426-2005.

Behavioral activation system modulation on brain activation during appetitive and aversive stimulus processing.

PubMed

Barrós-Loscertales, Alfonso; Ventura-Campos, Noelia; Sanjuán-Tomás, Ana; Belloch, Vicente; Parcet, Maria-Antònia; Avila, César

2010-03-01

The reinforcement sensitivity theory (RST) proposed the behavioral activation system (BAS) as a neurobehavioral system that is dependent on dopamine-irrigated structures and that mediates the individual differences in sensitivity and reactivity to appetitive stimuli associated with BAS-related personality traits. Theoretical developments propose that high BAS sensitivity is associated with both enhanced appetitive stimuli processing and the diminished processing of aversive stimuli. The objective of this study was to analyze how individual differences in BAS functioning were associated with brain activation during erotic and aversive picture processing while subjects were involved in a simple goal-directed task. Forty-five male participants took part in this study. The task activation results confirm the activation of the reward and punishment brain-related structures while viewing erotic and aversive pictures, respectively. The SR scores show a positive correlation with activation of the left lateral prefrontal cortex, the mesial prefrontal cortex and the right occipital cortex while viewing erotic pictures, and a negative correlation with the right lateral prefrontal cortex and the left occipital cortex while viewing aversive pictures. In summary, the SR scores modulate the activity of the cortical areas in the prefrontal and the occipital cortices that are proposed to modulate the BAS and the BIS-FFFS.

Quantification of citalopram or escitalopram and their demethylated metabolites in neonatal hair samples by liquid chromatography-tandem mass spectrometry.

PubMed

Frison, Giampietro; Favretto, Donata; Vogliardi, Susanna; Terranova, Claudio; Ferrara, Santo Davide

2008-08-01

Citalopram and escitalopram are highly selective serotonin reuptake inhibitors widely used in the treatment of depression. They exhibit adverse drug reactions and side effects, however, and the development of specific methods for their determination is of great interest in clinical and forensic toxicology. A liquid chromatography-tandem mass spectrometry method has been developed and validated for the assay of citalopram, escitalopram, and their demethylated metabolites in 10-mg hair samples. The analytes were extracted by incubation in methanol and liquid/liquid extraction with diethyl ether/dichloromethane. Gradient elution on a narrow bore C18 column was realized using clomipramine-d3 as an internal standard. Positive ion electrospray ionization and tandem mass spectrometry determination by collision-induced dissociation were performed in an ion trap mass spectrometer. The method exhibited a linear range of 25 to 2000 pg/mg, a quantification limit of 25 pg/mg for all analytes, relative standard deviations in the range of 12.10 to 9.80 (intraassay), and 13.80 to 11.78 (interassay), and accuracies (as percent recovery of the spiked standards) in the range of 90% to 110%; it was applied to the determination of citalopram and escitalopram and their metabolites in hair samples of two newborns to document their in utero exposure to the drugs. The method proved suitable for neonatal hair analysis of citalopram or escitalopram and was applied to two real cases of gestational exposure.

Persistence of Cytosine Methylation of DNA following Fertilisation in the Mouse

PubMed Central

Li, Yan; O'Neill, Chris

2012-01-01

Normal development of the mammalian embryo requires epigenetic reprogramming of the genome. The level of cytosine methylation of CpG-rich (5meC) regions of the genome is a major epigenetic regulator and active global demethylation of 5meC throughout the genome is reported to occur within the first cell-cycle following fertilization. An enzyme or mechanism capable of catalysing such rapid global demethylation has not been identified. The mouse is a widely used model for studying developmental epigenetics. We have reassessed the evidence for this phenomenon of genome-wide demethylation following fertilisation in the mouse. We found when using conventional methods of immunolocalization that 5meC showed a progressive acid-resistant antigenic masking during zygotic maturation which gave the appearance of demethylation. Changing the unmasking strategy by also performing tryptic digestion revealed a persistence of a methylated state. Analysis of methyl binding domain 1 protein (MBD1) binding confirmed that the genome remained methylated following fertilisation. The maintenance of this methylated state over the first several cell-cycles required the actions of DNA methyltransferase activity. The study shows that any 5meC remodelling that occurs during early development is not explained by a global active loss of 5meC staining during the cleavage stage of development and global loss of methylation following fertilization is not a major component of epigenetic reprogramming in the mouse zygote. PMID:22292019

Ethanol-induced changes in poly (ADP ribose) polymerase and neuronal developmental gene expression.

PubMed

Gavin, David P; Kusumo, Handojo; Sharma, Rajiv P; Guizzetti, Marina

2016-11-01

Prenatal alcohol exposure has profound effects on neuronal growth and development. Poly-ADP Ribose Polymerase (PARP) enzymes are perhaps unique in the field of epigenetics in that they directly participate in histone modifications, transcription factor modifications, DNA methylation/demethylation and are highly inducible by ethanol. It was our hypothesis that ethanol would induce PARP enzymatic activity leading to alterations in neurodevelopmental gene expression. Mouse E18 cortical neurons were treated with ethanol, PARP inhibitors, and nuclear hormone receptor transcription factor PPARγ agonists and antagonists. Subsequently, we measured PARP activity and changes in Bdnf, OKSM (Oct4, Klf4, Sox2, c-Myc), DNA methylating/demethylating factors, and Pparγ mRNA expression, promoter 5-methylcytosine (5MC) and 5-hydroxymethylcytosine (5HMC), and PPARγ promoter binding. We found that ethanol reduced Bdnf4, 9a, and Klf4 mRNA expression, and increased c-Myc expression. These changes were reversed with a PARP inhibitor. In agreement with its role in DNA demethylation PARP inhibition increased 5MC levels at the c-Myc promoter. In addition, we found that inhibition of PARP enzymatic activity increased PPARγ promoter binding, and this corresponded to increased Bdnf and Klf4 mRNA expression. Our results suggest that PARP participates in DNA demethylation and reduces PPARγ promoter binding. The current study underscores the importance of PARP in ethanol-induced changes to neurodevelopmental gene expression. Published by Elsevier Ltd.

Metabolism of Kaempferia parviflora polymethoxyflavones by human intestinal bacterium Bautia sp. MRG-PMF1.

PubMed

Kim, Mihyang; Kim, Nayoung; Han, Jaehong

2014-12-24

Poylmethoxyflavones (PMFs) are major bioactive flavonoids, which exhibit various biological activities, such as anticancer effects. The biotransformation of PMFs and characterization of a PMF-metabolizing human intestinal bacterium were studied herein for the first time. Hydrolysis of aryl methyl ether functional groups by human fecal samples was observed from the bioconversion of various PMFs. Activity-guided screening for PMF-metabolizing intestinal bacteria under anaerobic conditions resulted in the isolation of a strict anaerobic bacterium, which was identified as Blautia sp. MRG-PMF1. The isolated MRG-PMF1 was able to metabolize various PMFs to the corresponding demethylated flavones. The microbial conversion of bioactive 5,7-dimethoxyflavone (5,7-DMF) and 5,7,4'-trimethoxyflavone (5,7,4'-TMF) was studied in detail. 5,7-DMF and 5,7,4'-TMF were completely metabolized to 5,7-dihydroxyflavone (chrysin) and 5,7,4'-trihydroxyflavone (apigenin), respectively. From a kinetics study, the methoxy group on the flavone C-7 position was found to be preferentially hydrolyzed. 5-Methoxychrysin, the intermediate of 5,7-DMF metabolism by Blautia sp. MRG-PMF1, was isolated and characterized by nuclear magnetic resonance spectroscopy. Apigenin was produced from the sequential demethylation of 5,7,4'-TMF, via 5,4'-dimethoxy-7-hydroxyflavone and 7,4'-dihydroxy-5-methoxyflavone (thevetiaflavone). Not only demethylation activity but also deglycosylation activity was exhibited by Blautia sp. MRG-PMF1, and various flavonoids, including isoflavones, flavones, and flavanones, were found to be metabolized to the corresponding aglycones. The unprecedented PMF demethylation activity of Blautia sp. MRG-PMF1 will expand our understanding of flavonoid metabolism in the human intestine and lead to novel bioactive compounds.

Kinetics of Methylmercury Production Revisited

DOE PAGES

Olsen, Todd A.; Muller, Katherine A.; Painter, Scott L.; ...

2018-01-27

Laboratory measurements of the biologically mediated methylation of mercury (Hg) to the neurotoxin monomethylmercury (MMHg) often exhibit kinetics that are inconsistent with first-order kinetic models. Using time-resolved measurements of filter passing Hg and MMHg during methylation/demethylation assays, a multisite kinetic sorption model, and reanalyses of previous assays, we show in this paper that competing kinetic sorption reactions can lead to time-varying availability and apparent non-first-order kinetics in Hg methylation and MMHg demethylation. The new model employing a multisite kinetic sorption model for Hg and MMHg can describe the range of behaviors for time-resolved methylation/demethylation data reported in the literature includingmore » those that exhibit non-first-order kinetics. Additionally, we show that neglecting competing sorption processes can confound analyses of methylation/demethylation assays, resulting in rate constant estimates that are systematically biased low. Finally, simulations of MMHg production and transport in a hypothetical periphyton biofilm bed illustrate the implications of our new model and demonstrate that methylmercury production may be significantly different than projected by single-rate first-order models.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

Palsamy, Periyasamy; Ayaki, Masahiko; Elanchezhian, Rajan

Highlights: Black-Right-Pointing-Pointer We found significant Keap1 promoter demethylation in diabetic cataractous lenses. Black-Right-Pointing-Pointer Demethylation of Keap1 gene upregulated the expression of Keap1 mRNA and protein. Black-Right-Pointing-Pointer Elevated levels of Keap1 are known to decrease the levels of Nrf2. Black-Right-Pointing-Pointer Thereby, the levels of antioxidant enzymes are suppressed by decreased Nrf2 level. -- Abstract: Age-related cataracts (ARCs) are the major cause of visual impairments worldwide, and diabetic adults tend to have an earlier onset of ARCs. Although age is the strongest risk factor for cataracts, little is known how age plays a role in the development of ARCs. It is knownmore » that oxidative stress in the lens increases with age and more so in the lenses of diabetics. One of the central adaptive responses against the oxidative stresses is the activation of the nuclear transcriptional factor, NF-E2-related factor 2 (Nrf2), which then activates more than 20 different antioxidative enzymes. Kelch-like ECH associated protein 1 (Keap1) targets and binds to Nrf2 for proteosomal degradation. We hypothesized that hyperglycemia will lead to a dysfunction of the Nrf2-dependent antioxidative protection in the lens of diabetics. We studied the methylation status of the CpG islands in 15 clear and 21 diabetic cataractous lenses. Our results showed significant levels of demethylated DNA in the Keap1 promoter in the cataractous lenses from diabetic patients. In contrast, highly methylated DNA was found in the clear lens and tumorized human lens epithelial cell (HLEC) lines (SRA01/04). HLECs treated with a demethylation agent, 5-aza-2 Prime deoxycytidine (5-Aza), had a 10-fold higher levels of Keap1 mRNA, 3-fold increased levels of Keap1 protein, produced higher levels of ROS, and increased cell death. Our results indicated that demethylation of the CpG islands in the Keap1 promoter will activate the expression of Keap1 protein

Process for preparing active oxide powders

DOEpatents

Berard, Michael F.; Hunter, Jr., Orville; Shiers, Loren E.; Dole, Stephen L.; Scheidecker, Ralph W.

1979-02-20

An improved process for preparing active oxide powders in which cation hydroxide gels, prepared in the conventional manner are chemically dried by alternately washing the gels with a liquid organic compound having polar characteristics and a liquid organic compound having nonpolar characteristics until the mechanical water is removed from the gel. The water-free cation hydroxide is then contacted with a final liquid organic wash to remove the previous organic wash and speed drying. The dried hydroxide treated in the conventional manner will form a highly sinterable active oxide powder.

Complete solids retention activated sludge process.

PubMed

Amanatidou, E; Samiotis, G; Trikoilidou, E; Pekridis, G; Tsikritzis, L

2016-01-01

In a slaughterhouse's full-scale extended aeration activated sludge wastewater treatment plant (WWTP), operating under complete solids retention time, the evolution of mixed liquor suspended solids (MLSS) and mixed liquor volatile suspended solids (MLVSS) concentration, food to micro-organisms ratio (F/M) and substrate utilization rate (SUR) were studied for over a year. Biomass growth phases in correlation to sludge biological and morphological characteristics were studied. Three distinguished growth phases were observed during the 425 days of monitoring. The imposed operational conditions led the process to extended biomass starvation conditions, minimum F/M, minimum SUR and predator species growth. MLSS and MLVSS reached a stabilization phase (plateau phase) where almost zero sludge accumulation was observed. The concept of degradation of the considered non-biodegradable particulate compounds in influent and in biomass (cell debris) was also studied. Comparison of evolution of observed sludge yields (Yobs) in the WWTP with Yobs predictions by activated sludge models verified the degradation concept for the considered non-biodegradable compounds. Control of the sedimentation process was achieved, by predicting the solids loading rate critical point using state point analysis and stirred/unstirred settling velocity tests and by applying a high return activated sludge rate. The nitrogen gas related sedimentation problems were taken into consideration.

A predictive computational model of the kinetic mechanism of stimulus-induced transducer methylation and feedback regulation through CheY in archaeal phototaxis and chemotaxis.

PubMed

Streif, Stefan; Oesterhelt, Dieter; Marwan, Wolfgang

2010-03-18

Photo- and chemotaxis of the archaeon Halobacterium salinarum is based on the control of flagellar motor switching through stimulus-specific methyl-accepting transducer proteins that relay the sensory input signal to a two-component system. Certain members of the transducer family function as receptor proteins by directly sensing specific chemical or physical stimuli. Others interact with specific receptor proteins like the phototaxis photoreceptors sensory rhodopsin I and II, or require specific binding proteins as for example some chemotaxis transducers. Receptor activation by light or a change in receptor occupancy by chemical stimuli results in reversible methylation of glutamate residues of the transducer proteins. Both, methylation and demethylation reactions are involved in sensory adaptation and are modulated by the response regulator CheY. By mathematical modeling we infer the kinetic mechanisms of stimulus-induced transducer methylation and adaptation. The model (deterministic and in the form of ordinary differential equations) correctly predicts experimentally observed transducer demethylation (as detected by released methanol) in response to attractant and repellent stimuli of wildtype cells, a cheY deletion mutant, and a mutant in which the stimulated transducer species is methylation-deficient. We provide a kinetic model for signal processing in photo- and chemotaxis in the archaeon H. salinarum suggesting an essential role of receptor cooperativity, antagonistic reversible methylation, and a CheY-dependent feedback on transducer demethylation.

Changes in cytochrome P4501A activity during development in common tern chicks fed polychlorinated biphenyls, as measured by the caffeine breath test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feyk, L.A.; Giesy, J.P.; Bosveld, A.T.C.

2000-03-01

Cytochrome P4501A (CYPIA) activity is often used as a biomarker of exposure of wildlife to polyhalogenated diaromatic hydrocarbons and is usually measured ex vivo in liver tissue. A caffeine breath test (CBT) with radiolabeled substrate ({sup 14}C-caffeine) was used to measure in vivo CYP1A activity twice during development in 14 common tern (Sterna hirundo) chicks treated with polyhalogenated diaromatic hydrocarbons. Tern hatchlings were fed fish spiked with 3,3{prime}, 4,4{prime},5-pentachlorobiphenyl (PCB 126) and 2,2{prime},4,4{prime},5,5{prime}-hexachlorobiphenyl (PCB 153) such that the diet contained an average of 23, 99, or 561 pg of 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents per gram of fish for 21 d. Sixteen additionalmore » common tern chicks were similarly dosed with polyhalogenated diaromatic hydrocarbons but were not subjected to the CBT procedure. In weeks 1 and 2, caffeine N-demethylation and ethoxyresorufin-O-deethylation activity on day 21 were elevated in birds that received the greatest PCB dose. There was less constitutive and greater induction of ethoxyresorufin-O-deethylation activity than caffeine N-demethylation. The {sup 14}C-CBT was less invasive than the ethoxyresorufin-O-deethylase assay. Only one morphological parameter differed significantly between CBT subjects and no-CBT subjects fed the same level of PCBs. Bursa weight was significantly less in control CBT subjects than in control no-CBT subjects, but bursa weights did not differ among CBT and no-CBT birds from the two PCB treatment groups. No alterations of survival or growth occurred in CBT subjects compared with no-CBT subjects.« less

New macrocyclic lactones with acaricidal and nematocidal activities from a genetically engineered strain Streptomyces bingchenggensis BCJ60.

PubMed

Li, Jian-Song; Zhang, Hui; Zhang, Shao-Yong; Wang, Hai-Yan; Zhang, Ji; Chen, An-Liang; Wang, Ji-Dong; Xiang, Wen-Sheng

2017-04-01

Two new macrocyclic lactones, 4,25-diethyl-4,25-demethyl-milbemycin β 3 (1) and 27-formaldehyde-milbemycin β 14 (2), were isolated from a genetically engineered strain Streptomyces bingchenggensis BCJ60. Their structures were determined on the basis of spectroscopic analysis, including 1D and 2D NMR techniques as well as ESI-MS and comparison with data from the literature. The acaricidal and nematocidal capacities of compounds 1 and 2 were evaluated against Tetranychus cinnabarinus and Bursaphelenchus xylophilus, respectively. The results showed that the two new macrocyclic lactones 1 and 2 possessed potent acaricidal and nematocidal activities.

Gadd45a Is an RNA Binding Protein and Is Localized in Nuclear Speckles

PubMed Central

Sytnikova, Yuliya A.; Kubarenko, Andriy V.; Schäfer, Andrea; Weber, Alexander N. R.; Niehrs, Christof

2011-01-01

Background The Gadd45 proteins play important roles in growth control, maintenance of genomic stability, DNA repair, and apoptosis. Recently, Gadd45 proteins have also been implicated in epigenetic gene regulation by promoting active DNA demethylation. Gadd45 proteins have sequence homology with the L7Ae/L30e/S12e RNA binding superfamily of ribosomal proteins, which raises the question if they may interact directly with nucleic acids. Principal Findings Here we show that Gadd45a binds RNA but not single- or double stranded DNA or methylated DNA in vitro. Sucrose density gradient centrifugation experiments demonstrate that Gadd45a is present in high molecular weight particles, which are RNase sensitive. Gadd45a displays RNase-sensitive colocalization in nuclear speckles with the RNA helicase p68 and the RNA binding protein SC35. A K45A point mutation defective in RNA binding was still active in DNA demethylation. This suggests that RNA binding is not absolutely essential for demethylation of an artificial substrate. A point mutation at G39 impared RNA binding, nuclear speckle localization and DNA demethylation, emphasizing its relevance for Gadd45a function. Significance The results implicate RNA in Gadd45a function and suggest that Gadd45a is associated with a ribonucleoprotein particle. PMID:21249130

Periphyton Biofilms Influence Net Methylmercury Production in an Industrially Contaminated System.

PubMed

Olsen, Todd A; Brandt, Craig C; Brooks, Scott C

2016-10-18

Mercury (Hg) methylation and methylmercury (MMHg) demethylation activity of periphyton biofilms from the industrially contaminated East Fork Poplar Creek, Tennessee (EFPC) were measured during 2014-2016 using stable Hg isotopic rate assays. 201 Hg II and MM 202 Hg were added to intact periphyton samples in ambient streamwater and the formation of MM 201 Hg and loss of MM 202 Hg were monitored over time and used to calculate first-order rate potentials for methylation and demethylation. The influences of location, temperature/season, light exposure and biofilm structure on methylation and demethylation potentials were examined. Between-site differences in net methylation for samples collected from an upstream versus downstream location were driven by differences in the demethylation rate potential (k d ). In contrast, the within-site temperature-dependent difference in net methylation was driven by changes in the methylation rate potential (k m ). Samples incubated in the dark had lower net methylation due to lower k m values than those incubated in the light. Disrupting the biofilm structure decreased k m and resulted in lower net methylation. Overall, the measured rates resulted in a net excess of MMHg generated which could account for 3.71-7.88 mg d -1 MMHg flux in EFPC and suggests intact, actively photosynthesizing periphyton biofilms harbor zones of MMHg production.

Active Learning for Automatic Audio Processing of Unwritten Languages (ALAPUL)

DTIC Science & Technology

2016-07-01

AFRL-RH-WP-TR-2016-0074 ACTIVE LEARNING FOR AUTOMATIC AUDIO PROCESSING OF UNWRITTEN LANGUAGES (ALAPUL) Dimitra Vergyri Andreas Kathol Wen Wang...June 2015-July 2016 4. TITLE AND SUBTITLE Active Learning for Automatic Audio Processing of Unwritten Languages (ALAPUL) 5a. CONTRACT NUMBER...5430, 27 October 2016 1. SUMMARY The goal of the project was to investigate development of an automatic spoken language processing (ASLP) system

The co-existence of transcriptional activator and transcriptional repressor MEF2 complexes influences tumor aggressiveness

PubMed Central

Di Giorgio, Eros; Franforte, Elisa; Cefalù, Sebastiano; Rossi, Sabrina; Dei Tos, Angelo Paolo; Polano, Maurizio; Maestro, Roberta; Paluvai, Harikrishnareddy

2017-01-01

The contribution of MEF2 TFs to the tumorigenic process is still mysterious. Here we clarify that MEF2 can support both pro-oncogenic or tumor suppressive activities depending on the interaction with co-activators or co-repressors partners. Through these interactions MEF2 supervise histone modifications associated with gene activation/repression, such as H3K4 methylation and H3K27 acetylation. Critical switches for the generation of a MEF2 repressive environment are class IIa HDACs. In leiomyosarcomas (LMS), this two-faced trait of MEF2 is relevant for tumor aggressiveness. Class IIa HDACs are overexpressed in 22% of LMS, where high levels of MEF2, HDAC4 and HDAC9 inversely correlate with overall survival. The knock out of HDAC9 suppresses the transformed phenotype of LMS cells, by restoring the transcriptional proficiency of some MEF2-target loci. HDAC9 coordinates also the demethylation of H3K4me3 at the promoters of MEF2-target genes. Moreover, we show that class IIa HDACs do not bind all the regulative elements bound by MEF2. Hence, in a cell MEF2-target genes actively transcribed and strongly repressed can coexist. However, these repressed MEF2-targets are poised in terms of chromatin signature. Overall our results candidate class IIa HDACs and HDAC9 in particular, as druggable targets for a therapeutic intervention in LMS. PMID:28419090

Krebs cycle dysfunction shapes epigenetic landscape of chromatin: novel insights into mitochondrial regulation of aging process.

PubMed

Salminen, Antero; Kaarniranta, Kai; Hiltunen, Mikko; Kauppinen, Anu

2014-07-01

Although there is a substantial literature that mitochondria have a crucial role in the aging process, the mechanism has remained elusive. The role of reactive oxygen species, mitochondrial DNA injuries, and a decline in mitochondrial quality control has been proposed. Emerging studies have demonstrated that Krebs cycle intermediates, 2-oxoglutarate (also known as α-ketoglutarate), succinate and fumarate, can regulate the level of DNA and histone methylation. Moreover, citrate, also a Krebs cycle metabolite, can enhance histone acetylation. Genome-wide screening studies have revealed that the aging process is linked to significant epigenetic changes in the chromatin landscape, e.g. global demethylation of DNA and histones and increase in histone acetylation. Interestingly, recent studies have revealed that the demethylases of DNA (TET1-3) and histone lysines (KDM2-7) are members of 2-oxoglutarate-dependent dioxygenases (2-OGDO). The 2-OGDO enzymes are activated by oxygen, iron and the major Krebs cycle intermediate, 2-oxoglutarate, whereas they are inhibited by succinate and fumarate. Considering the endosymbiont origin of mitochondria, it is not surprising that Krebs cycle metabolites can control the gene expression of host cell by modifying the epigenetic landscape of chromatin. It seems that age-related disturbances in mitochondrial metabolism can induce epigenetic reprogramming, which promotes the appearance of senescent phenotype and degenerative diseases. Copyright © 2014 Elsevier Inc. All rights reserved.

Meditation-induced neuroplastic changes in amygdala activity during negative affective processing.

PubMed

Leung, Mei-Kei; Lau, Way K W; Chan, Chetwyn C H; Wong, Samuel S Y; Fung, Annis L C; Lee, Tatia M C

2018-06-01

Recent evidence suggests that the effects of meditation practice on affective processing and resilience have the potential to induce neuroplastic changes within the amygdala. Notably, literature speculates that meditation training may reduce amygdala activity during negative affective processing. Nonetheless, studies have thus far not verified this speculation. In this longitudinal study, participants (N = 21, 9 men) were trained in awareness-based compassion meditation (ABCM) or matched relaxation training. The effects of meditation training on amygdala activity were examined during passive viewing of affective and neutral stimuli in a non-meditative state. We found that the ABCM group exhibited significantly reduced anxiety and right amygdala activity during negative emotion processing than the relaxation group. Furthermore, ABCM participants who performed more compassion practice had stronger right amygdala activity reduction during negative emotion processing. The lower right amygdala activity after ABCM training may be associated with a general reduction in reactivity and distress. As all participants performed the emotion processing task in a non-meditative state, it appears likely that the changes in right amygdala activity are carried over from the meditation practice into the non-meditative state. These findings suggest that the distress-reducing effects of meditation practice on affective processing may transfer to ordinary states, which have important implications on stress management.

Active processes in one dimension

NASA Astrophysics Data System (ADS)

Demaerel, Thibaut; Maes, Christian

2018-03-01

We consider the thermal and athermal overdamped motion of particles in one-dimensional geometries where discrete internal degrees of freedom (spin) are coupled with the translational motion. Adding a driving velocity that depends on the time-dependent spin constitutes the simplest model of active particles (run-and-tumble processes) where the violation of the equipartition principle and of the Sutherland-Einstein relation can be studied in detail even when there is generalized reversibility. We give an example (with four spin values) where the irreversibility of the translational motion manifests itself only in higher-order (than two) time correlations. We derive a generalized telegraph equation as the Smoluchowski equation for the spatial density for an arbitrary number of spin values. We also investigate the Arrhenius exponential law for run-and-tumble particles; due to their activity the slope of the potential becomes important in contrast to the passive diffusion case and activity enhances the escape from a potential well (if that slope is high enough). Finally, in the absence of a driving velocity, the presence of internal currents such as in the chemistry of molecular motors may be transmitted to the translational motion and the internal activity is crucial for the direction of the emerging spatial current.

Students' Learning Activities While Studying Biological Process Diagrams

ERIC Educational Resources Information Center

Kragten, Marco; Admiraal, Wilfried; Rijlaarsdam, Gert

2015-01-01

Process diagrams describe how a system functions (e.g. photosynthesis) and are an important type of representation in Biology education. In the present study, we examined students' learning activities while studying process diagrams, related to their resulting comprehension of these diagrams. Each student completed three learning tasks. Verbal…

Nonoxidative removal of organics in the activated sludge process

PubMed Central

Modin, Oskar; Persson, Frank; Wilén, Britt-Marie; Hermansson, Malte

2016-01-01

ABSTRACT The activated sludge process is commonly used to treat wastewater by aerobic oxidation of organic pollutants into carbon dioxide and water. However, several nonoxidative mechanisms can also contribute to removal of organics. Sorption onto activated sludge can remove a large fraction of the colloidal and particulate wastewater organics. Intracellular storage of, e.g., polyhydroxyalkanoates (PHA), triacylglycerides (TAG), or wax esters can convert wastewater organics into precursors for high-value products. Recently, several environmental, economic, and technological drivers have stimulated research on nonoxidative removal of organics for wastewater treatment. In this paper, we review these nonoxidative removal mechanisms as well as the existing and emerging process configurations that make use of them for wastewater treatment. Better utilization of nonoxidative processes in activated sludge could reduce the wasteful aerobic oxidation of organic compounds and lead to more resource-efficient wastewater treatment plants. PMID:27453679

Aging process alters hippocampal and cortical secretase activities of Wistar rats.

PubMed

Bertoldi, Karine; Cechinel, Laura Reck; Schallenberger, Bruna; Meireles, Louisiana; Basso, Carla; Lovatel, Gisele Agustini; Bernardi, Lisiane; Lamers, Marcelo Lazzaron; Siqueira, Ionara Rodrigues

2017-01-15

A growing body of evidence has demonstrated amyloid plaques in aged brain; however, little attention has been given to amyloid precursor protein (APP) processing machinery during the healthy aging process. The amyloidogenic and non-amyloidogenic pathways, represented respectively by β- and α-secretases (BACE and TACE), are responsible for APP cleavage. Our working hypothesis is that the normal aging process could imbalance amyloidogenic and non-amyloidogenic pathways specifically BACE and TACE activities. Besides, although it has been showed that exercise can modulate secretase activities in Alzheimer Disease models the relationship between exercise effects and APP processing during healthy aging process is rarely studied. Our aim was to investigate the aging process and the exercise effects on cortical and hippocampal BACE and TACE activities and aversive memory performance. Young adult and aged Wistar rats were subjected to an exercise protocol (20min/day for 2 weeks) and to inhibitory avoidance task. Biochemical parameters were evaluated 1h and 18h after the last exercise session in order to verify transitory and delayed exercise effects. Aged rats exhibited impaired aversive memory and diminished cortical TACE activity. Moreover, an imbalance between TACE and BACE activities in favor of BACE activity was observed in aged brain. Moderate treadmill exercise was unable to alter secretase activities in any brain areas or time points evaluated. Our results suggest that aging-related aversive memory decline is partly linked to decreased cortical TACE activity. Additionally, an imbalance between secretase activities can be related to the higher vulnerability to neurodegenerative diseases induced by aging. Copyright © 2016 Elsevier B.V. All rights reserved.

Basic Science Process Skills. An Inservice Workshop Kit: Outlines and Activities.

ERIC Educational Resources Information Center

Rowland, Paul; And Others

A science process skill project was developed to help elementary teachers meet competency standards in New Mexico for teaching the process approach in their science classes. An outline of the process skills along with recommended activities are presented in this document. Performance objectives are identified and a sample activity form is…

Elucidation of the mechanism of N-demethylation catalyzed by cytochrome P450 monooxygenase is facilitated by exploiting nitrogen-15 heavy isotope effects.

PubMed

Kwiecień, Renata A; Molinié, Roland; Paneth, Piotr; Silvestre, Virginie; Lebreton, Jacques; Robins, Richard J

2011-06-01

(15)N heavy isotope effects are especially useful when detail is sought pertaining to the reaction mechanism for the cleavage of a C-N bond. Their potential in assisting to describe the mechanism of N-demethylation of tertiary amines by the action of cytochrome P450 monooxygenase has been investigated. As a working model for the first step, oxidation of the N-methyl group to N-methoxyl, tropine and a cytochrome P450 monooxygenase reaction centre composed of a truncated heme with sulfhydryl as the axial ligand were used. It is apparent that this first step of the reaction proceeds via a hydrogen atom transfer mechanism. Transition states for this step are described for both the high spin ((4)TS(H)) and low spin ((2)TS(H)) pathways in both gas and solvation states. Hence, overall normal secondary (15)N KIE could be calculated for the reaction path modeled in the low spin state, and inverse for the reaction modeled in the high spin state. This partial reaction has been identified as the probable rate limiting step. The model for the second step, fission of the C-N bond, consisted of N-methoxylnortropine and two molecules of water. A transition state described for this step, TS(CN), gives a strongly inverse overall theoretical (15)N KIE. Copyright © 2011 Elsevier Inc. All rights reserved.

Enantiomeric metabolic interactions and stereoselective human methadone metabolism.

PubMed

Totah, Rheem A; Allen, Kyle E; Sheffels, Pamela; Whittington, Dale; Kharasch, Evan D

2007-04-01

Methadone is administered as a racemate, although opioid activity resides in the R-enantiomer. Methadone disposition is stereoselective, with considerable unexplained variability in clearance and plasma R/S ratios. N-Demethylation of methadone in vitro is predominantly mediated by cytochrome P450 CYP3A4 and CYP2B6 and somewhat by CYP2C19. This investigation evaluated stereoselectivity, models, and kinetic parameters for methadone N-demethylation by recombinant CYP2B6, CYP3A4, and CYP2C19, and the potential for interactions between enantiomers during racemate metabolism. CYP2B6 metabolism was stereoselective. CYP2C19 was less active, and stereoselectivity was opposite that for CYP2B6. CYP3A4 was not stereoselective. With all three isoforms, enantiomer N-dealkylation rates in the racemate were lower than those of (R)-(6-dimethyamino-4,4-diphenyl-heptan-3-one) hydrochloride (R-methadone) or (S)-(6-dimethyamino-4,4-diphenyl-heptan-3-one) hydrochloride (S-methadone) alone, suggesting an enantiomeric interaction and mutual metabolic inhibition. For CYP2B6, the interaction between enantiomers was stereoselective, with S-methadone as a more potent inhibitor of R-methadone N-demethylation than R-of S-methadone. In contrast, enantiomer interactions were not stereoselective with CYP2C19 or CYP3A4. For all three cytochromes P450, methadone N-demethylation was best described by two-site enzyme models with competitive inhibition. There were minor model differences between cytochromes P450 to account for stereoselectivity of metabolism and enantiomeric interactions. Changes in plasma R/S methadone ratios observed after rifampin or troleandomycin pretreatment in humans in vivo were successfully predicted by CYP2B6- but not CYP3A4-catalyzed methadone N-demethylation. CYP2B6 is a predominant catalyst of stereoselective methadone metabolism in vitro. In vivo, CYP2B6 may be a major determinant of methadone metabolism and disposition, and CYP2B6 activity and stereoselective metabolic

Trimethoprim degradation by Fenton and Fe(II)-activated persulfate processes.

PubMed

Wang, Shizong; Wang, Jianlong

2018-01-01

Trimethoprim is a pollutant ubiquitous in the environment due to its extensive application, and it cannot be effectively removed by conventional wastewater treatment processes. In this study, the Fenton and the Fe(II)-activated persulfate processes were employed to degrade trimethoprim in an aqueous solution. The results showed that the concentration of persulfate, H 2 O 2 and Fe(II) a had significant influence on the degradation of trimethoprim in both processes. De-ionized water spiked with trimethoprim resulted in the complete degradation of trimethoprim (0.05 mM) by the mineralization of 54.9% of Fenton's reagent when the concentrations of H 2 O 2 and Fe(II) were 1 mM and 0.05 mM, respectively. In contrast, 73.4% of trimethoprim was degraded by the mineralization of 40.5% of the Fe(II)-activated persulfate process when the concentration of persulfate and Fe(II) were each 4 mM. Intermediate compounds with different m/z were detected for the Fenton and the Fe(II)-activated persulfate processes, indicating alternative degradation pathways. In the actual wastewater spiked with trimethoprim, the removal efficiency of trimethoprim decreased to 35.8% and 43.6%, respectively, for the Fenton and the Fe(II)-activated persulfate processes. In addition, the decomposition efficiencies for hydrogen peroxide and persulfate were 43.8% and 92.5%, respectively, which was lower than those in the de-ionized water system. These results demonstrated that wastewater components had a negative influence on trimethoprim degradation and the decomposition of the oxidants (persulfate and H 2 O 2 ). In summary, the Fe(II)-activated persulfate process could be used as an alternative technology for treating trimethoprim-containing wastewater. Copyright © 2017 Elsevier Ltd. All rights reserved.

Differential Processing for Actively Ignored Pictures and Words.

PubMed

Walker, Maegen; Ciraolo, Margeaux; Dewald, Andrew; Sinnett, Scott

2017-01-01

Previous work suggests that, when attended, pictures may be processed more readily than words. The current study extends this research to assess potential differences in processing between these stimulus types when they are actively ignored. In a dual-task paradigm, facilitated recognition for previously ignored words was found provided that they appeared frequently with an attended target. When adapting the same paradigm here, previously unattended pictures were recognized at high rates regardless of how they were paired with items during the primary task, whereas unattended words were later recognized at higher rates only if they had previously been aligned with primary task targets. Implicit learning effects obtained by aligning unattended items with attended task-targets may apply only to conceptually abstract stimulus types, such as words. Pictures, on the other hand, may maintain direct access to semantic information, and are therefore processed more readily than words, even when being actively ignored.

Differential Processing for Actively Ignored Pictures and Words

PubMed Central

Ciraolo, Margeaux

2017-01-01

Previous work suggests that, when attended, pictures may be processed more readily than words. The current study extends this research to assess potential differences in processing between these stimulus types when they are actively ignored. In a dual-task paradigm, facilitated recognition for previously ignored words was found provided that they appeared frequently with an attended target. When adapting the same paradigm here, previously unattended pictures were recognized at high rates regardless of how they were paired with items during the primary task, whereas unattended words were later recognized at higher rates only if they had previously been aligned with primary task targets. Implicit learning effects obtained by aligning unattended items with attended task-targets may apply only to conceptually abstract stimulus types, such as words. Pictures, on the other hand, may maintain direct access to semantic information, and are therefore processed more readily than words, even when being actively ignored. PMID:28122022

Selective attention modulates high-frequency activity in the face-processing network.

PubMed

Müsch, Kathrin; Hamamé, Carlos M; Perrone-Bertolotti, Marcela; Minotti, Lorella; Kahane, Philippe; Engel, Andreas K; Lachaux, Jean-Philippe; Schneider, Till R

2014-11-01

Face processing depends on the orchestrated activity of a large-scale neuronal network. Its activity can be modulated by attention as a function of task demands. However, it remains largely unknown whether voluntary, endogenous attention and reflexive, exogenous attention to facial expressions equally affect all regions of the face-processing network, and whether such effects primarily modify the strength of the neuronal response, the latency, the duration, or the spectral characteristics. We exploited the good temporal and spatial resolution of intracranial electroencephalography (iEEG) and recorded from depth electrodes to uncover the fast dynamics of emotional face processing. We investigated frequency-specific responses and event-related potentials (ERP) in the ventral occipito-temporal cortex (VOTC), ventral temporal cortex (VTC), anterior insula, orbitofrontal cortex (OFC), and amygdala when facial expressions were task-relevant or task-irrelevant. All investigated regions of interest (ROI) were clearly modulated by task demands and exhibited stronger changes in stimulus-induced gamma band activity (50-150 Hz) when facial expressions were task-relevant. Observed latencies demonstrate that the activation is temporally coordinated across the network, rather than serially proceeding along a processing hierarchy. Early and sustained responses to task-relevant faces in VOTC and VTC corroborate their role for the core system of face processing, but they also occurred in the anterior insula. Strong attentional modulation in the OFC and amygdala (300 msec) suggests that the extended system of the face-processing network is only recruited if the task demands active face processing. Contrary to our expectation, we rarely observed differences between fearful and neutral faces. Our results demonstrate that activity in the face-processing network is susceptible to the deployment of selective attention. Moreover, we show that endogenous attention operates along the whole

Study on extraction process and activity of plant polysaccharides

NASA Astrophysics Data System (ADS)

Ma, Xiaogen; Wang, Xiaojing; Fan, Shuangli; Chen, Jiezhong

2017-10-01

Recent studies have shown that plant polysaccharides have many pharmacological activities, such as hypoglycemic, anti-inflammatory and tumor inhibition. The pharmacological activities of plant polysaccharides were summarized. The extraction methods of plant polysaccharides were discussed. Finally, the extraction process of Herba Taraxaci polysaccharides was optimized by ultrasonic assisted extraction. Through single factor experiments and orthogonal experiment to optimize the optimum extraction process from dandelion polysaccharide, optimum conditions of dandelion root polysaccharide by ultrasonic assisted extraction method for ultrasonic power 320W, temperature 80°C, extraction time 40min, can get higher dandelion polysaccharide extract.

Computational implications of activity-dependent neuronal processes

NASA Astrophysics Data System (ADS)

Goldman, Mark Steven

Synapses, the connections between neurons, often fail to transmit a large percentage of the action potentials that they receive. I describe several models of synaptic transmission at a single stochastic synapse with an activity-dependent probability of transmission and demonstrate how synaptic transmission failures may increase the efficiency with which a synapse transmits information. Spike trains in the visual cortex of freely viewing monkeys have positive auto correlations that are indicative of a redundant representation of the information they contain. I show how a synapse with activity-dependent transmission failures modeled after those occurring in visual cortical synapses can remove this redundancy by transmitting a decorrelated subset of the spike trains it receives. I suggest that redundancy reduction at individual synapses saves synaptic resources while increasing the sensitivity of the postsynaptic neuron to information arriving along many inputs. For a neuron receiving input from many decorrelating synapses, my analysis leads to a prediction of the number of visual inputs to a neuron and the cross-correlations between these inputs and suggests that the time scale of synaptic dynamics observed in sensory areas corresponds to a fundamental time scale for processing sensory information. Systems with activity-dependent changes in their parameters, or plasticity, often display a wide variability in their individual components that belies the stability of their function, Motivated by experiments demonstrating that identified neurons with stereotyped function can have a large variability in the densities of their ion channels, or ionic conductances, I build a conductance-based model of a single neuron. The neuron's firing activity is relatively insensitive to changes in certain combinations of conductances, but markedly sensitive to changes in other combinations. Using a combined modeling and experimental approach, I show that neuromodulators and regulatory

Controlling Contagion Processes in Activity Driven Networks

NASA Astrophysics Data System (ADS)

Liu, Suyu; Perra, Nicola; Karsai, Márton; Vespignani, Alessandro

2014-03-01

The vast majority of strategies aimed at controlling contagion processes on networks consider the connectivity pattern of the system either quenched or annealed. However, in the real world, many networks are highly dynamical and evolve, in time, concurrently with the contagion process. Here, we derive an analytical framework for the study of control strategies specifically devised for a class of time-varying networks, namely activity-driven networks. We develop a block variable mean-field approach that allows the derivation of the equations describing the coevolution of the contagion process and the network dynamic. We derive the critical immunization threshold and assess the effectiveness of three different control strategies. Finally, we validate the theoretical picture by simulating numerically the spreading process and control strategies in both synthetic networks and a large-scale, real-world, mobile telephone call data set.

Conformational Transition Pathway in the Activation Process of Allosteric Glucokinase

PubMed Central

Shi, Ting; Zhao, Yaxue; Chen, Yingyi; Li, Xiaobai; Liu, Xinyi; Huang, Zhimin; Zhang, Jian

2013-01-01

Glucokinase (GK) is a glycolytic enzyme that plays an important role in regulating blood glucose level, thus acting as a potentially attractive target for drug discovery in the treatment of diabetes of the young type 2 and persistent hyperinsulinemic hypoglycemia of infancy. To characterize the activation mechanism of GK from the super-open state (inactive state) to the closed state (active state), a series of conventional molecular dynamics (MD) and targeted MD (TMD) simulations were performed on this enzyme. Conventional MD simulation showed a specific conformational ensemble of GK when the enzyme is inactive. Seven TMD simulations depicted a reliably conformational transition pathway of GK from the inactive state to the active state, and the components important to the conformational change of GK were identified by analyzing the detailed structures of the TMD trajectories. In combination with the inactivation process, our findings showed that the whole conformational pathway for the activation-inactivation-activation of GK is a one-direction circulation, and the active state is less stable than the inactive state in the circulation. Additionally, glucose was demonstrated to gradually modulate its binding pose with the help of residues in the large domain and connecting region of GK during the activation process. Furthermore, the obtained energy barriers were used to explain the preexisting equilibrium and the slow binding kinetic process of the substrate by GK. The simulated results are in accordance with the recent findings from the mutagenesis experiments and kinetic analyses. Our observations reveal a complicated conformational process in the allosteric protein, resulting in new knowledge about the delicate mechanisms for allosteric biological macromolecules that will be useful in drug design for targeting allosteric proteins. PMID:23409066

Process-Based Governance in Public Administrations Using Activity-Based Costing

NASA Astrophysics Data System (ADS)

Becker, Jörg; Bergener, Philipp; Räckers, Michael

Decision- and policy-makers in public administrations currently lack on missing relevant information for sufficient governance. In Germany the introduction of New Public Management and double-entry accounting enable public administrations to get the opportunity to use cost-centered accounting mechanisms to establish new governance mechanisms. Process modelling in this case can be a useful instrument to help the public administrations decision- and policy-makers to structure their activities and capture relevant information. In combination with approaches like Activity-Based Costing, higher management level can be supported with a reasonable data base for fruitful and reasonable governance approaches. Therefore, the aim of this article is combining the public sector domain specific process modelling method PICTURE and concept of activity-based costing for supporting Public Administrations in process-based Governance.

DNA methylation dynamics in mouse preimplantation embryos revealed by mass spectrometry.

PubMed

Okamoto, Yoshinori; Yoshida, Naoko; Suzuki, Toru; Shimozawa, Nobuhiro; Asami, Maki; Matsuda, Tomonari; Kojima, Nakao; Perry, Anthony C F; Takada, Tatsuyuki

2016-01-11

Following fertilization in mammals, paternal genomic 5-methyl-2'-deoxycytidine (5 mC) content is thought to decrease via oxidation to 5-hydroxymethyl-2'-deoxycytidine (5 hmC). This reciprocal model of demethylation and hydroxymethylation is inferred from indirect, non-quantitative methods. We here report direct quantification of genomic 5 mC and 5 hmC in mouse embryos by small scale liquid chromatographic tandem mass spectrometry (SMM). Profiles of absolute 5 mC levels in embryos produced by in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) were almost identical. By 10 h after fertilization, 5 mC levels had declined by ~40%, consistent with active genomic DNA demethylation. Levels of 5 mC in androgenotes (containing only a paternal genome) and parthenogenotes (containing only a maternal genome) underwent active 5 mC loss in the first 6 h, showing that both parental genomes can undergo demethylation independently. We found no evidence for net loss of 5 mC 10-48 h after fertilization, implying that any passive 'demethylation' following DNA replication was balanced by active 5 mC maintenance methylation. However, levels of 5 mC declined during development after 48 h, to 1% (measured as a fraction of G-residues) in blastocysts (~96 h). 5 hmC levels were consistently low (<0.2% of G-residues) throughout development in normal diploid embryos. This work directly quantifies the dynamics of global genomic DNA modification in mouse preimplantation embryos, suggesting that SMM will be applicable to other biomedical situations with limiting sample sizes.

Alteration of the Alkaloid Profile in Genetically Modified Tobacco Reveals a Role of Methylenetetrahydrofolate Reductase in Nicotine N-Demethylation1[C][W][OA

PubMed Central

Hung, Chiu-Yueh; Fan, Longjiang; Kittur, Farooqahmed S.; Sun, Kehan; Qiu, Jie; Tang, She; Holliday, Bronwyn M.; Xiao, Bingguang; Burkey, Kent O.; Bush, Lowell P.; Conkling, Mark A.; Roje, Sanja; Xie, Jiahua

2013-01-01

Methylenetetrahydrofolate reductase (MTHFR) is a key enzyme of the tetrahydrofolate (THF)-mediated one-carbon (C1) metabolic network. This enzyme catalyzes the reduction of 5,10-methylene-THF to 5-methyl-THF. The latter donates its methyl group to homocysteine, forming methionine, which is then used for the synthesis of S-adenosyl-methionine, a universal methyl donor for numerous methylation reactions, to produce primary and secondary metabolites. Here, we demonstrate that manipulating tobacco (Nicotiana tabacum) MTHFR gene (NtMTHFR1) expression dramatically alters the alkaloid profile in transgenic tobacco plants by negatively regulating the expression of a secondary metabolic pathway nicotine N-demethylase gene, CYP82E4. Quantitative real-time polymerase chain reaction and alkaloid analyses revealed that reducing NtMTHFR expression by RNA interference dramatically induced CYP82E4 expression, resulting in higher nicotine-to-nornicotine conversion rates. Conversely, overexpressing NtMTHFR1 suppressed CYP82E4 expression, leading to lower nicotine-to-nornicotine conversion rates. However, the reduced expression of NtMTHFR did not affect the methionine and S-adenosyl-methionine levels in the knockdown lines. Our finding reveals a new regulatory role of NtMTHFR1 in nicotine N-demethylation and suggests that the negative regulation of CYP82E4 expression may serve to recruit methyl groups from nicotine into the C1 pool under C1-deficient conditions. PMID:23221678

Digital image processing and analysis for activated sludge wastewater treatment.

PubMed

Khan, Muhammad Burhan; Lee, Xue Yong; Nisar, Humaira; Ng, Choon Aun; Yeap, Kim Ho; Malik, Aamir Saeed

2015-01-01

Activated sludge system is generally used in wastewater treatment plants for processing domestic influent. Conventionally the activated sludge wastewater treatment is monitored by measuring physico-chemical parameters like total suspended solids (TSSol), sludge volume index (SVI) and chemical oxygen demand (COD) etc. For the measurement, tests are conducted in the laboratory, which take many hours to give the final measurement. Digital image processing and analysis offers a better alternative not only to monitor and characterize the current state of activated sludge but also to predict the future state. The characterization by image processing and analysis is done by correlating the time evolution of parameters extracted by image analysis of floc and filaments with the physico-chemical parameters. This chapter briefly reviews the activated sludge wastewater treatment; and, procedures of image acquisition, preprocessing, segmentation and analysis in the specific context of activated sludge wastewater treatment. In the latter part additional procedures like z-stacking, image stitching are introduced for wastewater image preprocessing, which are not previously used in the context of activated sludge. Different preprocessing and segmentation techniques are proposed, along with the survey of imaging procedures reported in the literature. Finally the image analysis based morphological parameters and correlation of the parameters with regard to monitoring and prediction of activated sludge are discussed. Hence it is observed that image analysis can play a very useful role in the monitoring of activated sludge wastewater treatment plants.

Processing of Color Words Activates Color Representations

ERIC Educational Resources Information Center

Richter, Tobias; Zwaan, Rolf A.

2009-01-01

Two experiments were conducted to investigate whether color representations are routinely activated when color words are processed. Congruency effects of colors and color words were observed in both directions. Lexical decisions on color words were faster when preceding colors matched the color named by the word. Color-discrimination responses…

Turboexpanders with dry gas seals and active magnetic bearings in hydrocarbon processing

DOE Office of Scientific and Technical Information (OSTI.GOV)

Agahi, R.R.

1999-07-01

Since its first application in hydrocarbon processing in the early 1960s, turboexpander design has changed, evolved and improved tremendously. Today, hydrocarbon process designers use turboexpanders for almost all hydrocarbon liquid rejection and hydrocarbon dew point control for onshore and offshore installations. There are presently more than 3,000 turboexpanders operating in hydrocarbon gas processing plants worldwide. Due to the wide application of turboexpanders in hydrocarbon processing, the API-617 committee has assigned a task force to prepare an appendix to API-617 to cover design and manufacturing standards for turboexpanders. Dry gas seals (DGS) were cautiously introduced in the early 1980s for compressorsmore » used in hydrocarbon processing. It took almost a decade before dry gas seals found their application in turboexpanders. Dry gas seals were originally utilized to protect cryogenic hydrocarbon process gas from contamination by lubricating oil. Later on, dry gas seals were used to minimized hydrocarbon process gas leakage and also to provide an inert-gas-purged environment for both oil bearings and active magnetic bearings. The former eliminates the lubricating oil dilution problem and the latter made certification of active magnetic bearings by international certifying agencies possible. Active magnetic bearings (AMB), similar to dry gas seals, were originally introduced into hydrocarbon process gas compressors in the mid 1980s. The hydrocarbon processing industry waited half a decade to adopt this innovative technology for turboexpanders in the hydrocarbon process. The first turboexpander with active magnetic bearings was installed on an offshore platform in 1991. High reliability, low capital investment, low capital investment, low operating costs and more compact design have accelerated demand in recent years for turboexpanders with active magnetic bearings. In this paper, the author describes the technology of turboexpanders with dry gas

Process for producing an activated carbon adsorbent with integral heat transfer apparatus

NASA Technical Reports Server (NTRS)

Jones, Jack A. (Inventor); Yavrouian, Andre H. (Inventor)

1996-01-01

A process for producing an integral adsorbent-heat exchanger apparatus useful in ammonia refrigerant heat pump systems. In one embodiment, the process wets an activated carbon particles-solvent mixture with a binder-solvent mixture, presses the binder wetted activated carbon mixture on a metal tube surface and thereafter pyrolyzes the mixture to form a bonded activated carbon matrix adjoined to the tube surface. The integral apparatus can be easily and inexpensively produced by the process in large quantities.

Engineering a self-sufficient Mycobacterium tuberculosis CYP130 by gene fusion with the reductase-domain of CYP102A1 from Bacillus megaterium.

PubMed

Ortega Ugalde, Sandra; Luirink, Rosa A; Geerke, Daan P; Vermeulen, Nico P E; Bitter, Wilbert; Commandeur, Jan N M

2018-03-01

CYP130 belongs to the subset of cytochrome P450s from Mycobacterium tuberculosis (Mtb) that have been structurally characterized. Despite several efforts for its functional characterization, CYP130 is still considered an orphan enzyme for which no endogenous or exogenous substrate has been identified. In addition, functional redox-partners for CYP130 have not been clearly established yet, hampering the elucidation of its physiological role. In the present study, a catalytically active fusion protein involving CYP130 and the NADPH reductase-domain of CYP102A1 from Bacillus megaterium was created. By screening a panel of known substrates of human P450s, dextromethorphan N-demethylation was identified as a reaction catalyzed by CYP130. The fusion enzyme showed higher catalytic activity, when compared to CYP130 reconstituted with a selection of non-native redox-partners. Molecular dynamics simulation studies based on the crystal structure of CYP130 revealed two primary docking poses of dextromethorphan within the active site consistent with the experimentally observed N-demethylation reaction during the entire molecular dynamics simulation. The dextromethorphan N-demethylation reaction was strongly inhibited by azole-drugs and maybe applied to identify mechanism-based inhibitors of CYP130. Furthermore, the present active CYP130-fusion protein may facilitate the identification of endogenous substrates from Mtb. Copyright © 2017 Elsevier Inc. All rights reserved.

Active chatter suppression with displacement-only measurement in turning process

NASA Astrophysics Data System (ADS)

Ma, Haifeng; Wu, Jianhua; Yang, Liuqing; Xiong, Zhenhua

2017-08-01

Regenerative chatter is a major hindrance for achieving high quality and high production rate in machining processes. Various active controllers have been proposed to mitigate chatter. However, most of existing controllers were developed on the basis of multi-states feedback of the system and state observers were usually needed. Moreover, model parameters of the machining process (mass, damping and stiffness) were required in existing active controllers. In this study, an active sliding mode controller, which employs a dynamic output feedback sliding surface for the unmatched condition and an adaptive law for disturbance estimation, is designed, analyzed, and validated for chatter suppression in turning process. Only displacement measurement is required by this approach. Other sensors and state observers are not needed. Moreover, it facilitates a rapid implementation since the designed controller is established without using model parameters of the turning process. Theoretical analysis, numerical simulations and experiments on a computer numerical control (CNC) lathe are presented. It shows that the chatter can be substantially attenuated and the chatter-free region can be significantly expanded with the presented method.

Phytochemical profiles and antioxidant activity of processed brown rice products.

PubMed

Gong, Er Sheng; Luo, Shunjing; Li, Tong; Liu, Chengmei; Zhang, Guowen; Chen, Jun; Zeng, Zicong; Liu, Rui Hai

2017-10-01

The phytochemical profiles and antioxidant activity of free, soluble-conjugated, and bound fractions of brown rice and its processed products (textured rice, cooked rice and rice noodle) were studied. Nineteen phenolic acids were identified. Trans-ferulic acid was the most abundant monomeric phenolic acid with trans-trans-8-O-4' diferulic acid being most abundant diferulic acid. Processing increased the content of free phenolic acids, but decreased the content of soluble-conjugated phenolic acids. The content of bound phenolic acids was increased by improved extrusion cooking technology and cooking, but not affected by rice noodle extrusion. The total phenolic contents and antioxidant activities of free and soluble-conjugated fractions were decreased after processing, whereas those of bound fraction were increased by improved extrusion cooking technology and cooking, but not affected by rice noodle extrusion. Results indicated that whole foods designed for reducing chronic disease risk need to consider the effects of processing on phytochemical profiles and antioxidant activity of whole grains. Copyright © 2017 Elsevier Ltd. All rights reserved.

5-Azacytidine treatment induces demethylation of DAPK1 and MGMT genes and inhibits growth in canine mammary gland tumor cells.

PubMed

Ren, Xiaoli; Li, Huatao; Song, Xianyi; Wu, Yuhong; Liu, Yun

2018-01-01

Canine mammary gland tumors (CMGTs) are the most common, spontaneous types of neoplasias in female dogs. Aberrant DAPK1 and MGMT methylation associated with tumor formation and development in various cancers. 5-Azacytidine is a known specific demethylation drug that covalently binds to DNA methyltransferase. However, the methylation of the DAPK1 and MGMT is unknown with respect to CMGTs. Therefore, we sought to demonstrate the effects of 5-azacytidine on the proliferation of CMGTs cell, and elucidate the potential molecular mechanisms of action in these cancerous cells. The effects of 5-azacytidine on CHMm and CHMp cell proliferation were evaluated by MTT assay. The DAPK1 and MGMT gene methylation patterns in CHMm and CHMp cells and CMGTs blood/tissue samples were analyzed by MSP assay. Effect of 5-azacytidine on the methylation of DAPK1 and MGMT gene, and DAPK1 and MGMT mRNA expression in CHMm and CHMp cells were analyzed by MSP assay and qRT-PCR assay, respectively. 5-Azacytidine may suppress the proliferation of CHMm and CHMp cells. Furthermore, the DAPK1 and MGMT genes were hypermethylated in CHMm/CHMp cells and clinical malignant tumor samples, but not in normal female dogs' blood and tissue. However, the DAPK1 and MGMT genes were re-inducible in CHMm and CHMp cells treated with 5 μM 5-azacytidine. Meanwhile, 5-azacytidine increased the expression of DAPK1 and MGMT mRNA. These results suggest that DAPK1 and MGMT methylation can serve as sensitive diagnostic biomarkers and therapeutic targets for CMGTs. 5-Azacytidine also could be a potential therapeutic candidate for CMGTs.

The position of imidazopyridine and metabolic activation are pivotal factors in the antimutagenic activity of novel imidazo[1,2-a]pyridine derivatives.

PubMed

El-Sayed, Wael M; Hussin, Warda A; Al-Faiyz, Yasair S; Ismail, Mohamed A

2013-09-05

The antimutagenic activity of eight novel imidazo[1,2-a]pyridine derivatives (I-VIII) against sodium azide (NaN3) and benzo[a]pyrene (B[a]P) was evaluated using the Salmonella reverse mutation assay. At non-toxic concentrations (12.5-50 µM), imidazopyridines I, II, III, and V with a terminal imidazopyridine group were mutagenic, while derivatives VII and VIII with a central imidazopyridine group were not mutagenic. Compounds IV, VII, and VIII exerted a moderate antimutagenic activity against NaN3 under pre-exposure conditions, and a strong activity (>40%) against B[a]P in the presence of S9 under both pre- and co-exposure conditions and mostly independent on the dose. Imidazopyridines possibly inhibited the microsomal-dependent activation of B[a]P. The demethylated derivative VII was the most active antimutagen. All imidazopyridines had a low to moderate antioxidant activity. The antibacterial activity of imidazopyridines was sporadic and moderate probably due to the failure of bacteria to convert imidazopyridines into active metabolites. The position of imidazopyridine was a pivotal factor in the mutagenic/antimutagenic activity. The strong antimutagenic compounds were dicationic planar compounds with a centered imidazo[1,2-a]pyridine spacer. With LD50 of 60 mg/kg in mice for both derivatives VII and VIII, it is safe to investigate the anticancer activity of these derivatives in animal models. © 2013 Elsevier B.V. All rights reserved.

Arabidopsis ERG28 Tethers the Sterol C4-Demethylation Complex to Prevent Accumulation of a Biosynthetic Intermediate That Interferes with Polar Auxin Transport[C][W

PubMed Central

Mialoundama, Alexis Samba; Jadid, Nurul; Brunel, Julien; Di Pascoli, Thomas; Heintz, Dimitri; Erhardt, Mathieu; Mutterer, Jérôme; Bergdoll, Marc; Ayoub, Daniel; Van Dorsselaer, Alain; Rahier, Alain; Nkeng, Paul; Geoffroy, Philippe; Miesch, Michel; Camara, Bilal; Bouvier, Florence

2013-01-01

Sterols are vital for cellular functions and eukaryotic development because of their essential role as membrane constituents. Sterol biosynthetic intermediates (SBIs) represent a potential reservoir of signaling molecules in mammals and fungi, but little is known about their functions in plants. SBIs are derived from the sterol C4-demethylation enzyme complex that is tethered to the membrane by Ergosterol biosynthetic protein28 (ERG28). Here, using nonlethal loss-of-function strategies focused on Arabidopsis thaliana ERG28, we found that the previously undetected SBI 4-carboxy-4-methyl-24-methylenecycloartanol (CMMC) inhibits polar auxin transport (PAT), a key mechanism by which the phytohormone auxin regulates several aspects of plant growth, including development and responses to environmental factors. The induced accumulation of CMMC in Arabidopsis erg28 plants was associated with diagnostic hallmarks of altered PAT, including the differentiation of pin-like inflorescence, loss of apical dominance, leaf fusion, and reduced root growth. PAT inhibition by CMMC occurs in a brassinosteroid-independent manner. The data presented show that ERG28 is required for PAT in plants. Furthermore, it is accumulation of an atypical SBI that may act to negatively regulate PAT in plants. Hence, the sterol pathway offers further prospects for mining new target molecules that could regulate plant development. PMID:24326590

Dynamic DNA methylation reconfiguration during seed development and germination.

PubMed

Kawakatsu, Taiji; Nery, Joseph R; Castanon, Rosa; Ecker, Joseph R

2017-09-15

Unlike animals, plants can pause their life cycle as dormant seeds. In both plants and animals, DNA methylation is involved in the regulation of gene expression and genome integrity. In animals, reprogramming erases and re-establishes DNA methylation during development. However, knowledge of reprogramming or reconfiguration in plants has been limited to pollen and the central cell. To better understand epigenetic reconfiguration in the embryo, which forms the plant body, we compared time-series methylomes of dry and germinating seeds to publicly available seed development methylomes. Time-series whole genome bisulfite sequencing reveals extensive gain of CHH methylation during seed development and drastic loss of CHH methylation during germination. These dynamic changes in methylation mainly occur within transposable elements. Active DNA methylation during seed development depends on both RNA-directed DNA methylation and heterochromatin formation pathways, whereas global demethylation during germination occurs in a passive manner. However, an active DNA demethylation pathway is initiated during late seed development. This study provides new insights into dynamic DNA methylation reprogramming events during seed development and germination and suggests possible mechanisms of regulation. The observed sequential methylation/demethylation cycle suggests an important role of DNA methylation in seed dormancy.

Running Memory for Clinical Handoffs: A Look at Active and Passive Processing.

PubMed

Anderson-Montoya, Brittany L; Scerbo, Mark W; Ramirez, Dana E; Hubbard, Thomas W

2017-05-01

The goal of the present study was to examine the effects of domain-relevant expertise on running memory and the ability to process handoffs of information. In addition, the role of active or passive processing was examined. Currently, there is little research that addresses how individuals with different levels of expertise process information in running memory when the information is needed to perform a real-world task. Three groups of participants differing in their level of clinical expertise (novice, intermediate, and expert) performed an abstract running memory span task and two tasks resembling real-world activities, a clinical handoff task and an air traffic control (ATC) handoff task. For all tasks, list length and the amount of information to be recalled were manipulated. Regarding processing strategy, all participants used passive processing for the running memory span and ATC tasks. The novices also used passive processing for the clinical task. The experts, however, appeared to use more active processing, and the intermediates fell in between. Overall, the results indicated that individuals with clinical expertise and a developed mental model rely more on active processing of incoming information for the clinical task while individuals with little or no knowledge rely on passive processing. The results have implications about how training should be developed to aid less experienced personnel identify what information should be included in a handoff and what should not.

Active in-database processing to support ambient assisted living systems.

PubMed

de Morais, Wagner O; Lundström, Jens; Wickström, Nicholas

2014-08-12

As an alternative to the existing software architectures that underpin the development of smart homes and ambient assisted living (AAL) systems, this work presents a database-centric architecture that takes advantage of active databases and in-database processing. Current platforms supporting AAL systems use database management systems (DBMSs) exclusively for data storage. Active databases employ database triggers to detect and react to events taking place inside or outside of the database. DBMSs can be extended with stored procedures and functions that enable in-database processing. This means that the data processing is integrated and performed within the DBMS. The feasibility and flexibility of the proposed approach were demonstrated with the implementation of three distinct AAL services. The active database was used to detect bed-exits and to discover common room transitions and deviations during the night. In-database machine learning methods were used to model early night behaviors. Consequently, active in-database processing avoids transferring sensitive data outside the database, and this improves performance, security and privacy. Furthermore, centralizing the computation into the DBMS facilitates code reuse, adaptation and maintenance. These are important system properties that take into account the evolving heterogeneity of users, their needs and the devices that are characteristic of smart homes and AAL systems. Therefore, DBMSs can provide capabilities to address requirements for scalability, security, privacy, dependability and personalization in applications of smart environments in healthcare.

The regulation of Jmjd3 upon the expression of NF-κB downstream inflammatory genes in LPS activated vascular endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Shaoqing; Graduate School of Medicine, Nanchang University, Nanchang; Chen, Xia

Inflammatory mediators and adhesion molecules have been implicated in a variety of diseases including atherosclerosis. As both the mediator-releasing and targeted cells, vascular endothelial cells play key role in pathological processes. NF-κB signaling regulates a cluster of inflammatory factors in LPS-activated vascular endothelial cells but the underlying mechanisms remain largely unknown. Here, we investigated the epigenetic regulation of LPS upon the expression of inflammatory mediators and adhesion molecules. We found that LPS treatment promoted jmjd3 expression, enhanced Jmjd3 nuclear accumulation in human vascular endothelial cells. In addition, LPS enhanced the demethylation of H3K27me3, a specific substrate of Jmjd3. LPS treatmentmore » recruited Jmjd3 and NF-κB to the promoter region of target genes, suggesting Jmjd3 synergizes with NF-κB to activate the expression of target genes. We further found that Jmjd3 attenuated the methylation status in promoter region of target genes, culminating in target gene expression. Our findings unveil epigenetic regulations of LPS upon NF-κB pathway and identify Jmjd3 as a critical modulator of NF-κB pathway and potential therapeutic target for NF-κB related diseases including atherosclerosis.« less

Using cell membrane chromatography and HPLC-TOF/MS method for in vivo study of active components from roots of Aconitum carmichaeli

PubMed Central

Cao, Yan; Chen, Xiao-Fei; Lü, Di-Ya; Dong, Xin; Zhang, Guo-Qing; Chai, Yi-Feng

2012-01-01

An offline two-dimensional system combining a rat cardiac muscle cell membrane chromatography time-of-flight mass spectrometry (CMC-TOF/MS) with a high Performance liquid chromatography time-of-flight mass spectrometry (HPLC-TOF/MS) was established for investigating the parent components and metabolites in rat urine samples after administration of the roots of Aconitum carmichaeli. On the basis ofthe analysis of the first dimension, retention components of the urine sample were collected into 30 fractions (one fraction per minute). Then offline analysis of the second dimension was carried out. 34 compounds including 24 parent alkaloids and 10 potential metabolites were identified from the dosed rat urine, and then binding affinities of different compounds on cell membranes were compared and influences of some functional groups on activity were estimated with the semi-quantification and curve fitting method. As a result, binding affinities decreased along with the process of deacylation, debenzoylation and demethylation, which may be related to the alleviation of toxicity in the procedure of herb processing or metabolism. Moreover, some minor components in rat urine (Songorine, 14-benzoylneoline, Deoxyaconitine, etc.) exerted relatively strong affinity on cell membranes are worth exploring. The results delivered by the System suggest that the CMC can be applied to in vivo study. PMID:29403691

Identification of epoxybergamottin as a CYP3A4 inhibitor in grapefruit peel.

PubMed

Wangensteen, H; Molden, E; Christensen, H; Malterud, K E

2003-02-01

The oral availability of many drugs metabolised by the enzyme cytochrome P(450) 3A4 (CYP3A4) is increased if co-administered with grapefruit juice. Extracts from grapefruit peel have also demonstrated inhibitory activity and, during commercial manufacturing of grapefruit juice, inhibitory components might be squeezed into the juice from the peel. Thus, the aim of this in vitro study was to identify CYP3A4 inhibitors in grapefruit peel. Grapefruit peel was extracted with diethyl ether, and the extract was further fractionated by normal-phase chromatography. Fractions demonstrating significant CYP3A4 inhibitory activity, as measured by the relative reduction in N-demethylation of diltiazem in transfected human liver epithelial cells, were subsequently separated by preparative thin-layer chromatography. Constituents of the fractions and isolated compounds were identified by nuclear magnetic resonance spectroscopy. Analysis of diltiazem and N-demethyl-diltiazem was performed using high-performance liquid chromatography. Of the identified components in grapefruit peel, only epoxybergamottin demonstrated a concentration-dependent inhibition of the CYP3A4-mediated N-demethylation of diltiazem. The IC(50) value was calculated to be 4.2+/-1.1 micro M. Coumarins without the furan ring and flavonoids isolated from grapefruit peel did not interfere with the metabolism of diltiazem. The results indicated the presence of other CYP3A4 inhibitors in grapefruit peel, but these agents were lost during the purification process excluding their identification. The furanocoumarin epoxybergamottin, present in grapefruit peel, is an inhibitor of CYP3A4. In commercial manufacturing of grapefruit juice, epoxybergamottin is possibly distributed into the juice. During manufacturing, however, epoxybergamottin may be hydrolysed to 6',7'-dihydroxybergamottin, which has been suggested as an important CYP3A4 inhibitor in grapefruit juice.

Identification of three new phase II metabolites of a designer drug methylone formed in rats by N-demethylation followed by conjugation with dicarboxylic acids.

PubMed

Židková, Monika; Linhart, Igor; Balíková, Marie; Himl, Michal; Dvořáčková, Veronika; Lhotková, Eva; Páleníček, Tomáš

2018-06-01

1. Methylone (3,4-methylenedioxy-N-methylcathinone, MDMC), which appeared on the illicit drug market in 2004, is a frequently abused synthetic cathinone derivative. Known metabolic pathways of MDMC include N-demethylation to normethylone (3,4-methylenedioxycathinone, MDC), aliphatic chain hydroxylation and oxidative demethylenation followed by monomethylation and conjugation with glucuronic acid and/or sulphate. 2. Three new phase II metabolites, amidic conjugates of MDC with succinic, glutaric and adipic acid, were identified in the urine of rats dosed subcutaneously with MDMC.HCl (20 mg/kg body weight) by LC-ESI-HRMS using synthetic reference standards to support identification. 3. The main portion of administered MDMC was excreted unchanged. Normethylone, was a major urinary metabolite, of which a minor part was conjugated with dicarboxylic acids. 4. Previously identified ring-opened metabolites 4-hydroxy-3-methoxymethcathinone (4-OH-3-MeO-MC), 3-hydroxy-4-methoxymeth-cathinone (3-OH-4-MeO-MC) and 3,4-dihydroxymethcathinone (3,4-di-OH-MC) mostly in conjugated form with glucuronic and/or sulphuric acids were also detected. 5. Also, ring-opened metabolites derived from MDC, namely, 4-hydroxy-3-methoxycathinone (4-OH-3-MeO-C), 3-hydroxy-4-methoxycathinone (3-OH-4-MeO-C) and 3,4-dihydroxycathinone (3,4-di-OH-C) were identified for the first time in vivo.

Available processing resources influence encoding-related brain activity before an event

PubMed Central

Galli, Giulia; Gebert, A. Dorothea; Otten, Leun J.

2013-01-01

Effective cognitive functioning not only relies on brain activity elicited by an event, but also on activity that precedes it. This has been demonstrated in a number of cognitive domains, including memory. Here, we show that brain activity that precedes the effective encoding of a word into long-term memory depends on the availability of sufficient processing resources. We recorded electrical brain activity from the scalps of healthy adult men and women while they memorized intermixed visual and auditory words for later recall. Each word was preceded by a cue that indicated the modality of the upcoming word. The degree to which processing resources were available before word onset was manipulated by asking participants to make an easy or difficult perceptual discrimination on the cue. Brain activity before word onset predicted later recall of the word, but only in the easy discrimination condition. These findings indicate that anticipatory influences on long-term memory are limited in capacity and sensitive to the degree to which attention is divided between tasks. Prestimulus activity that affects later encoding can only be engaged when the necessary cognitive resources can be allocated to the encoding process. PMID:23219383

Clinical process analysis and activity-based costing at a heart center.

PubMed

Ridderstolpe, Lisa; Johansson, Andreas; Skau, Tommy; Rutberg, Hans; Ahlfeldt, Hans

2002-08-01

Cost studies, productivity, efficiency, and quality of care measures, the links between resources and patient outcomes, are fundamental issues for hospital management today. This paper describes the implementation of a model for process analysis and activity-based costing (ABC)/management at a Heart Center in Sweden as a tool for administrative cost information, strategic decision-making, quality improvement, and cost reduction. A commercial software package (QPR) containing two interrelated parts, "ProcessGuide and CostControl," was used. All processes at the Heart Center were mapped and graphically outlined. Processes and activities such as health care procedures, research, and education were identified together with their causal relationship to costs and products/services. The construction of the ABC model in CostControl was time-consuming. However, after the ABC/management system was created, it opened the way for new possibilities including process and activity analysis, simulation, and price calculations. Cost analysis showed large variations in the cost obtained for individual patients undergoing coronary artery bypass grafting (CABG) surgery. We conclude that a process-based costing system is applicable and has the potential to be useful in hospital management.

Meltlets(®) of soy isoflavones: process optimization and the effect of extrusion spheronization process parameters on antioxidant activity.

PubMed

Deshmukh, Ketkee; Amin, Purnima

2013-07-01

In the current research work an attempt was made to develop "Melt in mouth pellets" (Meltlets(®)) containing 40% herbal extract of soy isoflavones that served to provide antioxidants activity in menopausal women. The process of extrusion-spheronization was optimized for extruder speed, extruder screen size, spheronization speed, and time. While doing so the herbal extract incorporated in the pellet matrix was subjected to various processing conditions such as the effect of the presence of other excipients, mixing or kneading to prepare wet mass, heat generated during the process of extrusion, spheronization, and drying. Thus, the work further investigates the effect of these processing parameters on the antioxidant activity of the soy isoflavone herbal extract incorporated in the formula. Thereby, the antioxidant activity of the soya bean herbal extract, Meltlets(®) and of the placebo pellets was evaluated using DPPH free radical scavenging assay and total reduction capacity.

Left ventral occipitotemporal activation during orthographic and semantic processing of auditory words.

PubMed

Ludersdorfer, Philipp; Wimmer, Heinz; Richlan, Fabio; Schurz, Matthias; Hutzler, Florian; Kronbichler, Martin

2016-01-01

The present fMRI study investigated the hypothesis that activation of the left ventral occipitotemporal cortex (vOT) in response to auditory words can be attributed to lexical orthographic rather than lexico-semantic processing. To this end, we presented auditory words in both an orthographic ("three or four letter word?") and a semantic ("living or nonliving?") task. In addition, a auditory control condition presented tones in a pitch evaluation task. The results showed that the left vOT exhibited higher activation for orthographic relative to semantic processing of auditory words with a peak in the posterior part of vOT. Comparisons to the auditory control condition revealed that orthographic processing of auditory words elicited activation in a large vOT cluster. In contrast, activation for semantic processing was only weak and restricted to the middle part vOT. We interpret our findings as speaking for orthographic processing in left vOT. In particular, we suggest that activation in left middle vOT can be attributed to accessing orthographic whole-word representations. While activation of such representations was experimentally ascertained in the orthographic task, it might have also occurred automatically in the semantic task. Activation in the more posterior vOT region, on the other hand, may reflect the generation of explicit images of word-specific letter sequences required by the orthographic but not the semantic task. In addition, based on cross-modal suppression, the finding of marked deactivations in response to the auditory tones is taken to reflect the visual nature of representations and processes in left vOT. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

The Land Processes Distributed Active Archive Center (LP DAAC)

USGS Publications Warehouse

Golon, Danielle K.

2016-10-03

The Land Processes Distributed Active Archive Center (LP DAAC) operates as a partnership with the U.S. Geological Survey and is 1 of 12 DAACs within the National Aeronautics and Space Administration (NASA) Earth Observing System Data and Information System (EOSDIS). The LP DAAC ingests, archives, processes, and distributes NASA Earth science remote sensing data. These data are provided to the public at no charge. Data distributed by the LP DAAC provide information about Earth’s surface from daily to yearly intervals and at 15 to 5,600 meter spatial resolution. Data provided by the LP DAAC can be used to study changes in agriculture, vegetation, ecosystems, elevation, and much more. The LP DAAC provides several ways to access, process, and interact with these data. In addition, the LP DAAC is actively archiving new datasets to provide users with a variety of data to study the Earth.

A cluster expansion model for predicting activation barrier of atomic processes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rehman, Tafizur; Jaipal, M.; Chatterjee, Abhijit, E-mail: achatter@iitk.ac.in

2013-06-15

We introduce a procedure based on cluster expansion models for predicting the activation barrier of atomic processes encountered while studying the dynamics of a material system using the kinetic Monte Carlo (KMC) method. Starting with an interatomic potential description, a mathematical derivation is presented to show that the local environment dependence of the activation barrier can be captured using cluster interaction models. Next, we develop a systematic procedure for training the cluster interaction model on-the-fly, which involves: (i) obtaining activation barriers for handful local environments using nudged elastic band (NEB) calculations, (ii) identifying the local environment by analyzing the NEBmore » results, and (iii) estimating the cluster interaction model parameters from the activation barrier data. Once a cluster expansion model has been trained, it is used to predict activation barriers without requiring any additional NEB calculations. Numerical studies are performed to validate the cluster expansion model by studying hop processes in Ag/Ag(100). We show that the use of cluster expansion model with KMC enables efficient generation of an accurate process rate catalog.« less

Effect of alcohol consumption on CpG methylation in the differentially methylated regions of H19 and IG-DMR in male gametes: implications for fetal alcohol spectrum disorders.

PubMed

Ouko, Lillian A; Shantikumar, Katpaham; Knezovich, Jaysen; Haycock, Philip; Schnugh, Desmond J; Ramsay, Michèle

2009-09-01

Exposure to alcohol in utero is the main attributable cause of fetal alcohol spectrum disorders (FASD) which in its most severe form is characterized by irreversible behavioral and cognitive disability. Paternal preconception drinking is not considered to be a significant risk factor, even though animal studies have demonstrated that chronic paternal alcohol consumption has a detrimental effect on the physical and mental development of offspring even in the absence of in utero alcohol exposure. It has been documented that alcohol can reduce the levels and activity of DNA methyltransferases resulting in DNA hypomethylation and that reduced methyltransferase activity can cause activation of normally silenced genes. The aim of this study was to establish a link between alcohol use in men and hypomethylation of paternally imprinted loci in sperm DNA in genomic regions critical for embryonic development, thus providing a mechanism for paternal effects in the aetiology of FASD. Sperm DNA from male volunteers was bisulfite treated and the methylation patterns of 2 differentially methylated regions (DMRs), H19 and IG-DMR, analyzed following sequencing of individual clones. The methylation patterns were correlated with the alcohol consumption levels of the volunteer males. There was a pattern of increased demethylation with alcohol consumption at the 2 imprinted loci with a significant difference observed at the IG-DMR between the nondrinking and heavy alcohol consuming groups. Greater inter-individual variation in average methylation was observed at the H19 DMR and individual clones were more extensively demethylated than those of the IG-DMR. CpG site #4 in the IG-DMR was preferentially demethylated among all individuals and along with the H19 DMR CpG site #7 located within the CTCF binding site 6 showed significant demethylation in the alcohol consuming groups compared with the control group. This study demonstrates a correlation between chronic alcohol use and

Valuation of OSA process and folic acid addition as excess sludge minimization alternatives applied in the activated sludge process.

PubMed

Martins, C L; Velho, V F; Ramos, S R A; Pires, A S C D; Duarte, E C N F A; Costa, R H R

2016-01-01

The aim of this study was to investigate the ability of the oxic-settling-anaerobic (OSA)-process and the folic acid addition applied in the activated sludge process to reduce the excess sludge production. The study was monitored during two distinct periods: activated sludge system with OSA-process, and activated sludge system with folic acid addition. The observed sludge yields (Yobs) were 0.30 and 0.08 kgTSS kg(-1) chemical oxygen demand (COD), control phase and OSA-process (period 1); 0.33 and 0.18 kgTSS kg(-1) COD, control phase and folic acid addition (period 2). The Yobs decreased by 73 and 45% in phases with the OSA-process and folic acid addition, respectively, compared with the control phases. The sludge minimization alternatives result in a decrease in excess sludge production, without negatively affecting the performance of the effluent treatment.

Early neural activation during facial affect processing in adolescents with Autism Spectrum Disorder.

PubMed

Leung, Rachel C; Pang, Elizabeth W; Cassel, Daniel; Brian, Jessica A; Smith, Mary Lou; Taylor, Margot J

2015-01-01

Impaired social interaction is one of the hallmarks of Autism Spectrum Disorder (ASD). Emotional faces are arguably the most critical visual social stimuli and the ability to perceive, recognize, and interpret emotions is central to social interaction and communication, and subsequently healthy social development. However, our understanding of the neural and cognitive mechanisms underlying emotional face processing in adolescents with ASD is limited. We recruited 48 adolescents, 24 with high functioning ASD and 24 typically developing controls. Participants completed an implicit emotional face processing task in the MEG. We examined spatiotemporal differences in neural activation between the groups during implicit angry and happy face processing. While there were no differences in response latencies between groups across emotions, adolescents with ASD had lower accuracy on the implicit emotional face processing task when the trials included angry faces. MEG data showed atypical neural activity in adolescents with ASD during angry and happy face processing, which included atypical activity in the insula, anterior and posterior cingulate and temporal and orbitofrontal regions. Our findings demonstrate differences in neural activity during happy and angry face processing between adolescents with and without ASD. These differences in activation in social cognitive regions may index the difficulties in face processing and in comprehension of social reward and punishment in the ASD group. Thus, our results suggest that atypical neural activation contributes to impaired affect processing, and thus social cognition, in adolescents with ASD.

Identification of methylated genes in salivary gland adenoid cystic carcinoma xenografts using global demethylation and methylation microarray screening

PubMed Central

LING, SHIZHANG; RETTIG, ELENI M.; TAN, MARIETTA; CHANG, XIAOFEI; WANG, ZHIMING; BRAIT, MARIANA; BISHOP, JUSTIN A.; FERTIG, ELANA J.; CONSIDINE, MICHAEL; WICK, MICHAEL J.; HA, PATRICK K.

2016-01-01

Salivary gland adenoid cystic carcinoma (ACC) is a rare head and neck malignancy without molecular biomarkers that can be used to predict the chemotherapeutic response or prognosis of ACC. The regulation of gene expression of oncogenes and tumor suppressor genes (TSGs) through DNA promoter methylation may play a role in the carcinogenesis of ACC. To identify differentially methylated genes in ACC, a global demethylating agent, 5-aza-2′-deoxycytidine (5-AZA) was utilized to unmask putative TSG silencing in ACC xenograft models in mice. Fresh xenografts were passaged, implanted in triplicate in mice that were treated with 5-AZA daily for 28 days. These xenografts were then evaluated for genome-wide DNA methylation patterns using the Illumina Infinium HumanMethylation27 BeadChip array. Validation of the 32 candidate genes was performed by bisulfite sequencing (BS-seq) in a separate cohort of 6 ACC primary tumors and 6 normal control salivary gland tissues. Hypermethylation was identified in the HCN2 gene promoter in all 6 control tissues, but hypomethylation was found in all 6 ACC tumor tissues. Quantitative validation of HCN2 promoter methylation level in the region detected by BS-seq was performed in a larger cohort of primary tumors (n=32) confirming significant HCN2 hypomethylation in ACCs compared with normal samples (n=10; P=0.04). HCN2 immunohistochemical staining was performed on an ACC tissue microarray. HCN2 staining intensity and H-score, but not percentage of the positively stained cells, were significantly stronger in normal tissues than those of ACC tissues. With our novel screening and sequencing methods, we identified several gene candidates that were methylated. The most significant of these genes, HCN2, was actually hypomethylated in tumors. However, promoter methylation status does not appear to be a major determinant of HCN2 expression in normal and ACC tissues. HCN2 hypomethylation is a biomarker of ACC and may play an important role in the

Regio- and Stereo-Selective Oxidation of a Cardiovascular Drug, Metoprolol, Mediated by Cytochrome P450 2D and 3A Enzymes in Marmoset Livers.

PubMed

Uehara, Shotaro; Ishii, Sakura; Uno, Yasuhiro; Inoue, Takashi; Sasaki, Erika; Yamazaki, Hiroshi

2017-08-01

A β -blocker, metoprolol, is one of the in vivo probes for human cytochrome P450 (P450) 2D6. Investigation of nonhuman primate P450 enzymes helps to improve the accuracy of the extrapolation of pharmacokinetic data from animals into humans. Common marmosets ( Callithrix jacchus ) are a potential primate model for preclinical research, but the detailed roles of marmoset P450 enzymes in metoprolol oxidation remain unknown. In this study, regio- and stereo-selectivity of metoprolol oxidations by a variety of P450 enzymes in marmoset and human livers were investigated in vitro. Although liver microsomes from cynomolgus monkeys and rats preferentially mediated S -metoprolol O -demethylation and R -metoprolol α -hydroxylation, respectively, those from humans, marmosets, minipigs, and dogs preferentially mediated R -metoprolol O -demethylation, in contrast to the slow rates of R - and S -metoprolol oxidation in mouse liver microsomes. R - and S -metoprolol O -demethylation activities in marmoset livers were strongly inhibited by quinidine and ketoconazole, and were significantly correlated with bufuralol 1'-hydroxylation and midazolam 1'-hydroxylation activities and also with P450 2D and 3A4 contents, which is different from the case in human livers that did not have any correlations with P450 3A-mediated midazolam 1'-hydroxylation. Recombinant human P450 2D6 enzyme and marmoset P450 2D6/3A4 enzymes effectively catalyzed R -metoprolol O -demethylation, comparable to the activities of human and marmoset liver microsomes, respectively. These results indicated that the major roles of P450 2D enzymes for the regio- and stereo-selectivity of metoprolol oxidation were similar between human and marmoset livers, but the minor roles of P450 3A enzymes were unique to marmosets. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

Production of activated carbon by using pyrolysis process in an ammonia atmosphere

NASA Astrophysics Data System (ADS)

Indayaningsih, N.; Destyorini, F.; Purawiardi, R. I.; Insiyanda, D. R.; Widodo, H.

2017-04-01

Activated carbon is materials that have wide applications, including supercapacitor materials, absorbent in chemical industry, and absorbent material in the chemical industry. This study has carried out for the manufacturing of activated carbon from inexpensive materials through efficient processes. Carbon material was made from coconut fibers through pyrolysis process at temperature of 650, 700, 750 and 800°C. Aim of this study was to obtain carbon material that has a large surface area. Pyrolysis process is carried out in an inert atmosphere (N2 gas) at a temperature of 450°C for 30 minutes, followed by pyrolysis process in an ammonia atmosphere at 800°C for 2 hours. The pyrolysis results showed that the etching process in ammonia is occurred; as it obtained some greater surface area when compared with the pyrolisis process in an atmosphere by inert gas only. The resulted activated carbon also showed to have good properties in surface area and total pore volume.

Active In-Database Processing to Support Ambient Assisted Living Systems

PubMed Central

de Morais, Wagner O.; Lundström, Jens; Wickström, Nicholas

2014-01-01

As an alternative to the existing software architectures that underpin the development of smart homes and ambient assisted living (AAL) systems, this work presents a database-centric architecture that takes advantage of active databases and in-database processing. Current platforms supporting AAL systems use database management systems (DBMSs) exclusively for data storage. Active databases employ database triggers to detect and react to events taking place inside or outside of the database. DBMSs can be extended with stored procedures and functions that enable in-database processing. This means that the data processing is integrated and performed within the DBMS. The feasibility and flexibility of the proposed approach were demonstrated with the implementation of three distinct AAL services. The active database was used to detect bed-exits and to discover common room transitions and deviations during the night. In-database machine learning methods were used to model early night behaviors. Consequently, active in-database processing avoids transferring sensitive data outside the database, and this improves performance, security and privacy. Furthermore, centralizing the computation into the DBMS facilitates code reuse, adaptation and maintenance. These are important system properties that take into account the evolving heterogeneity of users, their needs and the devices that are characteristic of smart homes and AAL systems. Therefore, DBMSs can provide capabilities to address requirements for scalability, security, privacy, dependability and personalization in applications of smart environments in healthcare. PMID:25120164

Hypoxia, Epithelial-Mesenchymal Transition, and TET-Mediated Epigenetic Changes

PubMed Central

Kao, Shih-Han; Wu, Kou-Juey; Lee, Wen-Hwa

2016-01-01

Tumor hypoxia is a pathophysiologic outcome of disrupted microcirculation with inadequate supply of oxygen, leading to enhanced proliferation, epithelial-mesenchymal transition (EMT), metastasis, and chemo-resistance. Epigenetic changes induced by hypoxia are well documented, and they lead to tumor progression. Recent advances show that DNA demethylation mediated by the Ten-eleven translocation (TET) proteins induces major epigenetic changes and controls key steps of cancer development. TET enzymes serve as 5mC (5-methylcytosine)-specific dioxygenases and cause DNA demethylation. Hypoxia activates the expression of TET1, which also serves as a co-activator of HIF-1α transcriptional regulation to modulate HIF-1α downstream target genes and promote epithelial-mesenchymal transition. As HIF is a negative prognostic factor for tumor progression, hypoxia-activated prodrugs (HAPs) may provide a favorable therapeutic approach to lessen hypoxia-induced malignancy. PMID:26861406

[Research on the treatment of wastewater containing PVA by ozonation-activated sludge process].

PubMed

Xing, Xiao-Qiong; Huang, Cheng-Lan; Liu, Min; Chen, Ying

2012-11-01

The wastewater containing polyvinyl alcohol (PVA) was characterized with poor biodegradability, and was difficult to remove. In order to find an economically reasonable and practical technology, the research on the removal efficiency of different concentration wastewater containing PVA by ozonation-activated sludge process was studied, and the result was compared with the traditional activated sludge process. The results showed that the ozonation-activated sludge process was not suitable for treating influent with COD below 500 mg x L(-1) and the wastewater PVA concentration was 10-30 mg x L(-1). When the influent COD was between 500-800 mg x L(-1) and the PVA concentration was 15-60 mg x L(-1), the system had advantages on dealing with this kind of wastewater, and the average removal efficiency of COD and PVA were 92.8% and 57.4%, which were better than the traditional activated sludge process 4.1% and 15.2% respectively. In addition, the effluent concentrations of COD could keep between 30-60 mg x L(-1). When the influent COD was 1 000-1 200 mg x L(-1) and the PVA concentration was 20-70 mg x L(-1), the average removal efficiencies of COD and PVA were 90.9% and 45.3%, which were better than the traditional activated sludge process 12.8% and 12.1% respectively, but the effluent should to be further treated. Compared with the traditional activated sludge process, ozonation-activated sludge process had high treatment efficiency, stable running effect, and effectively in dealing with industrial wastewater containing PVA.

Overview of processing activities aimed at higher efficiencies and economical production

NASA Technical Reports Server (NTRS)

Bickler, D. B.

1985-01-01

An overview of processing activities aimed at higher efficiencies and economical production were presented. Present focus is on low-cost process technology for higher-efficiency cells of up to 18% or higher. Process development concerns center on the use of less than optimum silicon sheet, the control of production yields, and making uniformly efficient large-area cells. High-efficiency cell factors that require process development are bulk material perfection, very shallow junction formation, front-surface passivation, and finely detailed metallization. Better bulk properties of the silicon sheet and the keeping of those qualities throughout large areas during cell processing are required so that minority carrier lifetimes are maintained and cell performance is not degraded by high doping levels. When very shallow junctions are formed, the process must be sensitive to metallizatin punch-through, series resisitance in the cell, and control of dopant leaching during surface passivation. There is a need to determine the sensitivity to processing by mathematical modeling and experimental activities.

Processing of Building Binder Materials to Increase their Activation

NASA Astrophysics Data System (ADS)

Fediuk, R. S.; Garmashov, I. S.; Kuzmin, D. E.; Stoyushko, N. Yu; Gladkova, N. A.

2018-01-01

The paper deals modern physical methods of activation of building powder materials. During mechanical activation a composite binder active molecules cement minerals occur in the destruction of the molecular defects in the areas of packaging and breaking metastable phase decompensation intermolecular forces. The process is accompanied by a change in the kinetics of hardening of Portland cement. Activated concrete has a number of features that are used as design characteristics of structures and are due to the structure of the activated binder and its contacts with concrete aggregates. These features also have a significant impact on the nature of the destruction of concrete under load, changing the boundaries of its microcracks and durability.

The role of CYP2D6 in primary and secondary oxidative metabolism of dextromethorphan: in vitro studies using human liver microsomes.

PubMed Central

Kerry, N L; Somogyi, A A; Bochner, F; Mikus, G

1994-01-01

1. The enzyme kinetics of dextromethorphan O-demethylation in liver microsomes from three extensive metabolisers (EM) with respect to CYP2D6 indicated high (Km1 2.2-9.4 microM) and low (Km2 55.5-307.3 microM) affinity sites whereas microsomes from two poor metabolisers (PM) indicated a single site (Km 560 and 157 microM). Similar differences were shown for 3-methoxymorphinan O-demethylation to 3-hydroxymorphinan (Km 6.9-9.6 microM in EM subjects; Km 307 and 213 microM in PM subjects). 2. Dextromethorphan O-demethylation was inhibited competitively by quinidine (Ki 0.1 microM), rac-perhexiline (Ki 0.4 microM), dextropropoxyphene (Ki 6 microM), rac-methadone (Ki 8 microM), and 3-methoxymorphinan (Ki 15 microM). These compounds were also potent inhibitors of 3-methoxymorphinan O-demethylation with IC50 values ranging from 0.02-12 microM. Anti-LKM1 serum inhibited both dextromethorphan and 3-methoxymorphinan O-demethylations in a titre-dependent manner. 3. The Michaelis-Menten constant for dextromethorphan N-demethylation to 3-methoxymorphinan (Km 632-977 microM) and dextrorphan N-demethylation to 3-hydroxymorphinan (Km 1571-4286 microM) did not differ between EM and PM microsomes. These N-demethylation reactions were not inhibited by quinidine and rac-methadone or LKM1 antibodies. 4. Dextromethorphan and 3-methoxymorphinan are metabolised by the same P450 isoform, CYP2D6, whereas the N-demethylation reactions are not carried out by CYP2D6. PMID:7826826

Blogs and Social Network Sites as Activity Systems: Exploring Adult Informal Learning Process through Activity Theory Framework

ERIC Educational Resources Information Center

Heo, Gyeong Mi; Lee, Romee

2013-01-01

This paper uses an Activity Theory framework to explore adult user activities and informal learning processes as reflected in their blogs and social network sites (SNS). Using the assumption that a web-based space is an activity system in which learning occurs, typical features of the components were investigated and each activity system then…

Determination of oxidation products of 5-methylcytosine in plants by chemical derivatization coupled with liquid chromatography/tandem mass spectrometry analysis.

PubMed

Tang, Yang; Xiong, Jun; Jiang, Han-Peng; Zheng, Shu-Jian; Feng, Yu-Qi; Yuan, Bi-Feng

2014-08-05

Cytosine methylation (5-methylcytosine, 5-mC) in DNA is an important epigenetic mark that has regulatory roles in various biological processes. In plants, active DNA demethylation can be achieved through direct cleavage by DNA glycosylases, followed by replacement of 5-mC with cytosine by base excision repair (BER) machinery. Recent studies in mammals have demonstrated 5-mC can be sequentially oxidized to 5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-foC), and 5-carboxylcytosine (5-caC) by Ten-eleven translocation (TET) proteins. The consecutive oxidations of 5-mC constitute the active DNA demethylation pathway in mammals, which raised the possible presence of oxidation products of 5-mC (5-hmC, 5-foC, and 5-caC) in plant genomes. However, there is no definitive evidence supporting the presence of these modified bases in plant genomic DNA, especially for 5-foC and 5-caC. Here we developed a chemical derivatization strategy combined with liquid chromatography-electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) method to determine 5-formyl-2'-deoxycytidine (5-fodC) and 5-carboxyl-2'-deoxycytidine (5-cadC). Derivatization of 5-fodC and 5-cadC by Girard's reagents (GirD, GirT, and GirP) significantly increased the detection sensitivities of 5-fodC and 5-cadC by 52-260-fold. Using this method, we demonstrated the widespread existence of 5-fodC and 5-cadC in genomic DNA of various plant tissues, indicating that active DNA demethylation in plants may go through an alternative pathway similar to mammals besides the pathway of direct DNA glycosylases cleavage combined with BER. Moreover, we found that environmental stresses of drought and salinity can change the contents of 5-fodC and 5-cadC in plant genomes, suggesting the functional roles of 5-fodC and 5-cadC in response to environmental stresses.

Human cortical activity evoked by contextual processing in attentional orienting.

PubMed

Zhao, Shuo; Li, Chunlin; Uono, Shota; Yoshimura, Sayaka; Toichi, Motomi

2017-06-07

The ability to assess another person's direction of attention is paramount in social communication, many studies have reported a similar pattern between gaze and arrow cues in attention orienting. Neuroimaging research has also demonstrated no qualitative differences in attention to gaze and arrow cues. However, these studies were implemented under simple experiment conditions. Researchers have highlighted the importance of contextual processing (i.e., the semantic congruence between cue and target) in attentional orienting, showing that attentional orienting by social gaze or arrow cues could be modulated through contextual processing. Here, we examine the neural activity of attentional orienting by gaze and arrow cues in response to contextual processing using functional magnetic resonance imaging. The results demonstrated that the influence of neural activity through contextual processing to attentional orienting occurred under invalid conditions (when the cue and target were incongruent versus congruent) in the ventral frontoparietal network, although we did not identify any differences in the neural substrates of attentional orienting in contextual processing between gaze and arrow cues. These results support behavioural data of attentional orienting modulated by contextual processing based on the neurocognitive architecture.

Modeling DNA methylation by analyzing the individual configurations of single molecules

PubMed Central

Affinito, Ornella; Scala, Giovanni; Palumbo, Domenico; Florio, Ermanno; Monticelli, Antonella; Miele, Gennaro; Avvedimento, Vittorio Enrico; Usiello, Alessandro; Chiariotti, Lorenzo; Cocozza, Sergio

2016-01-01

ABSTRACT DNA methylation is often analyzed by reporting the average methylation degree of each cytosine. In this study, we used a single molecule methylation analysis in order to look at the methylation conformation of individual molecules. Using D-aspartate oxidase as a model gene, we performed an in-depth methylation analysis through the developmental stages of 3 different mouse tissues (brain, lung, and gut), where this gene undergoes opposite methylation destiny. This approach allowed us to track both methylation and demethylation processes at high resolution. The complexity of these dynamics was markedly simplified by introducing the concept of methylation classes (MCs), defined as the number of methylated cytosines per molecule, irrespective of their position. The MC concept smooths the stochasticity of the system, allowing a more deterministic description. In this framework, we also propose a mathematical model based on the Markov chain. This model aims to identify the transition probability of a molecule from one MC to another during methylation and demethylation processes. The results of our model suggest that: 1) both processes are ruled by a dominant class of phenomena, namely, the gain or loss of one methyl group at a time; and 2) the probability of a single CpG site becoming methylated or demethylated depends on the methylation status of the whole molecule at that time. PMID:27748645

Advanced oxidation processes on doxycycline degradation: monitoring of antimicrobial activity and toxicity.

PubMed

Spina-Cruz, Mylena; Maniero, Milena Guedes; Guimarães, José Roberto

2018-05-08

Advanced oxidation processes (AOPs) have been highly efficient in degrading contaminants of emerging concern (CEC). This study investigated the efficiency of photolysis, peroxidation, photoperoxidation, and ozonation at different pH values to degrade doxycycline (DC) in three aqueous matrices: fountain, tap, and ultrapure water. More than 99.6% of DC degradation resulted from the UV/H 2 O 2 and ozonation processes. Also, to evaluate the toxicity of the original solution and throughout the degradation time, antimicrobial activity tests were conducted using Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacteria, and acute toxicity test using the bioluminescent marine bacterium (Vibrio fischeri). Antimicrobial activity reduced as the drug degradation increased in UV/H 2 O 2 and ozonation processes, wherein the first process only 6 min was required to reduce 100% of both bacteria activity. In ozonation, 27.7 mg L -1 of ozone was responsible for reducing 100% of the antimicrobial activity. When applied the photoperoxidation process, an increase in the toxicity occurred as the high levels of degradation were achieved; it means that toxic intermediates were formed. The ozonated solutions did not present toxicity.

The Role of Informal and Formal Leisure Activities in the Disablement Process

ERIC Educational Resources Information Center

Janke, Megan C.; Payne, Laura L.; Van Puymbroeck, Marieke

2008-01-01

The disablement process model has been used as a framework to investigate factors that accelerate or decelerate disablement among older adults. Although very little is known about the direct and moderating effects of involvement in leisure activities on the disablement process, research has suggested that participation in leisure activities may…

Antioxidant Activity, Acetylcholinesterase, and Carbonic Anhydrase Inhibitory Properties of Novel Ureas Derived from Phenethylamines.

PubMed

Aksu, Kadir; Özgeriş, Bünyamin; Taslimi, Parham; Naderi, Ali; Gülçin, İlhami; Göksu, Süleyman

2016-12-01

A series of ureas derived from phenethylamines were synthesized and evaluated for human carbonic anhydrase (hCA) I and II, acetylcholinesterase (AChE), and butyrylcholinesterase (BChE) enzyme inhibitory activities and antioxidant properties. The ureas were synthesized from the reactions of substituted phenethylamines with N,N-dimethylcarbamoyl chloride; then, the synthesized compounds were converted to their corresponding phenolic derivatives via O-demethylation. hCA I and II were effectively inhibited by the newly synthesized compounds, with K i values in the range of 0.307-0.432 nM for hCA I and 0.149-0.278 nM for hCA II. On the other hand, the K i parameters of these compounds for AChE and BChE were determined in the range of 0.129-0.434 and 0.095-0.207 nM, respectively. Phenolic ureas also showed good antioxidant activities. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Instructional Transaction Theory: Knowledge Relationships among Processes, Entities, and Activities.

ERIC Educational Resources Information Center

Merrill, M. David; And Others

1993-01-01

Discussion of instructional transaction theory focuses on knowledge representation in an automated instructional design expert system. A knowledge structure called PEA-Net (processes, entities, and activities) is explained; the refrigeration process is used as an example; text resources and graphic resources are described; and simulations are…

Degradation of the UV-filter benzophenone-3 in aqueous solution using persulfate activated by heat, metal ions and light.

PubMed

Pan, Xiaoxue; Yan, Liqing; Qu, Ruijuan; Wang, Zunyao

2018-04-01

The goals of this study were to bring forward new data and insights into the effect of activation methods, operational variables and reaction pathways during sulfate radicals-based oxidation of benzophenone-3 (BP-3) in aqueous solution. Heat, transition metal ions (Fe 2+ , Cu 2+ , Co 2+ ), UV and visible light irradiation were used to activate persulfate (PS) to degrade BP-3. The results showed that these three activation methods can remarkably enhance BP-3 removal efficiency. Under the conditions of [BP-3] 0 : [PS] 0  = 1: 500, pH = 7.0, and 40 °C, complete removal of BP-3 (1.31 μM) was observed in 3 h. In the pH range of 3.0-9.0, the degradation of BP-3 decreased with increasing pH. Increasing the PS dosage accelerated the reaction, while the presence of humic acid (HA) significantly inhibited the efficiency of BP-3 removal. Based on electron paramagnetic resonance (EPR) and radical quenching studies, sulfate and hydroxyl radicals contributed to the oxidation process. According to the evolution of BP-3 and its 7 by-products, as well as frontier electron densities (FED) calculation, two routes were proposed involving hydroxylation, demethylation and direct oxidation. On the whole, this work is a unique contribution to the systematic elucidation of BP-3 removal by PS. Copyright © 2017 Elsevier Ltd. All rights reserved.

Correlation of neural activity with behavioral kinematics reveals distinct sensory encoding and evidence accumulation processes during active tactile sensing.

PubMed

Delis, Ioannis; Dmochowski, Jacek P; Sajda, Paul; Wang, Qi

2018-07-15

Many real-world decisions rely on active sensing, a dynamic process for directing our sensors (e.g. eyes or fingers) across a stimulus to maximize information gain. Though ecologically pervasive, limited work has focused on identifying neural correlates of the active sensing process. In tactile perception, we often make decisions about an object/surface by actively exploring its shape/texture. Here we investigate the neural correlates of active tactile decision-making by simultaneously measuring electroencephalography (EEG) and finger kinematics while subjects interrogated a haptic surface to make perceptual judgments. Since sensorimotor behavior underlies decision formation in active sensing tasks, we hypothesized that the neural correlates of decision-related processes would be detectable by relating active sensing to neural activity. Novel brain-behavior correlation analysis revealed that three distinct EEG components, localizing to right-lateralized occipital cortex (LOC), middle frontal gyrus (MFG), and supplementary motor area (SMA), respectively, were coupled with active sensing as their activity significantly correlated with finger kinematics. To probe the functional role of these components, we fit their single-trial-couplings to decision-making performance using a hierarchical-drift-diffusion-model (HDDM), revealing that the LOC modulated the encoding of the tactile stimulus whereas the MFG predicted the rate of information integration towards a choice. Interestingly, the MFG disappeared from components uncovered from control subjects performing active sensing but not required to make perceptual decisions. By uncovering the neural correlates of distinct stimulus encoding and evidence accumulation processes, this study delineated, for the first time, the functional role of cortical areas in active tactile decision-making. Copyright © 2018 Elsevier Inc. All rights reserved.

Granular activated carbon promoted ozonation of a food-processing secondary effluent.

PubMed

Alvarez, Pedro M; Pocostales, J Pablo; Beltrán, Fernando J

2011-01-30

This paper reports on the application of a simultaneous combination of ozone and a granular activated carbon (O(3)/GAC) as a tertiary treatment of a wastewater generated from the activity of various food-processing industries. Prior to the O(3)/GAC treatment, the wastewater was subjected to conventional primary and secondary treatments in a full-scale wastewater treatment plant (WWTP). The effluent from the WWTP presented high organic load (COD>500 mg/l and TOC>150 mg/l), which could be much reduced by the O(3)/GAC treatment. Results from the O(3)/GAC experiments were compared with those obtained in single ozonation, single adsorption onto GAC and sequential O(3)-GAC adsorption experiments. While single processes and the sequential one showed limited capacity to remove organic matter for the food-processing effluent (COD removal <40%), the simultaneous O(3)/GAC process led to decreases of COD up to 82% at the conditions here applied. The combined process also improved the ozone consumption, which decreased from about 19 g O(3)/g TOC (single ozonation process) to 8.2-10.7 g O(3)/g TOC (O(3)/GAC process). The reusability of the GAC throughout a series of consecutive O(3)/GAC experiments was studied with no apparent loss of activity for a neutral GAC (PZC = 6.7) but for a basic GAC (PZC = 9.1). Copyright © 2010 Elsevier B.V. All rights reserved.

Evaluation of Control Parameters for the Activated Sludge Process

ERIC Educational Resources Information Center

Stall, T. Ray; Sherrard, Josephy H.

1978-01-01

An evaluation of the use of the parameters currently being used to design and operate the activated sludge process is presented. The advantages and disadvantages for the use of each parameter are discussed. (MR)

Activated zeolite--suitable carriers for microorganisms in anaerobic digestion processes?

PubMed

Weiß, S; Lebuhn, M; Andrade, D; Zankel, A; Cardinale, M; Birner-Gruenberger, R; Somitsch, W; Ueberbacher, B J; Guebitz, G M

2013-04-01

Plant cell wall structures represent a barrier in the biodegradation process to produce biogas for combustion and energy production. Consequently, approaches concerning a more efficient de-polymerisation of cellulose and hemicellulose to monomeric sugars are required. Here, we show that natural activated zeolites (i.e. trace metal activated zeolites) represent eminently suitable mineral microhabitats and potential carriers for immobilisation of microorganisms responsible for anaerobic hydrolysis of biopolymers stabilising related bacterial and methanogenic communities. A strategy for comprehensive analysis of immobilised anaerobic populations was developed that includes the visualisation of biofilm formation via scanning electron microscopy and confocal laser scanning microscopy, community and fingerprint analysis as well as enzyme activity and identification analyses. Using SDS polyacrylamide gel electrophoresis, hydrolytical active protein bands were traced by congo red staining. Liquid chromatography/mass spectroscopy revealed cellulolytical endo- and exoglucanase (exocellobiohydrolase) as well as hemicellulolytical xylanase/mannase after proteolytic digestion. Relations to hydrolytic/fermentative zeolite colonisers were obtained by using single-strand conformation polymorphism analysis (SSCP) based on amplification of bacterial and archaeal 16S rRNA fragments. Thereby, dominant colonisers were affiliated to the genera Clostridium, Pseudomonas and Methanoculleus. The specific immobilisation on natural zeolites with functional microbes already colonising naturally during the fermentation offers a strategy to systematically supply the biogas formation process responsive to population dynamics and process requirements.

Decreased medial prefrontal cortex activation during self-referential processing in bipolar mania.

PubMed

Herold, Dorrit; Usnich, Tatiana; Spengler, Stephanie; Sajonz, Bastian; Bauer, Michael; Bermpohl, Felix

2017-09-01

Patients with bipolar disorder in mania exhibit symptoms pointing towards altered self-referential processing, such as decreased self-focus, flight of ideas and high distractibility. In depression, the opposite pattern of symptoms has been connected to increased activation of medial prefrontal cortex (mPFC) during self-referential processing. In this study, we hypothesized that (1) patients with mania will exhibit decreased activation in the mPFC during self-referential processing and (2) will be more alexithymic and that levels of alexithymia will correlate negatively with mPFC activation. The neural response to standardized pictures was compared in 14 patients with bipolar I disorder in mania to 14 healthy controls using blood oxygen level dependent contrast magnetic resonance imaging. Participants were asked to indicate with button press during the scanning session for each picture whether the pictures personally related to them or not. Toronto alexithymia scale (TAS) scores were recorded from all participants. In the group analysis, patients with mania exhibited decreased activation in a predefined region of interest in the mPFC during self-referential processing compared to healthy controls. Patients with mania showed significantly higher levels of alexithymia, attributable to difficulties in identifying and describing emotions. Activation in the mPFC correlated negatively with levels of alexithymia. Results presented here should be replicated in a larger group, potentially including unmedicated patients. The finding of decreased mPFC activation during self-referential processing in mania may reflect decreased self-focus and high distractibility. Support for this view comes from the negative correlation between higher alexithymia scores and decreased mPFC activation. These findings represent an opposite clinical and neuroimaging pattern to findings in depression. Copyright © 2017. Published by Elsevier B.V.

Adsorption behavior of sulfamethazine in an activated sludge process treating swine wastewater.

PubMed

Ben, Weiwei; Qiang, Zhimin; Yin, Xiaowei; Qu, Jiuhui; Pan, Xun

2014-08-01

Swine wastewater is an important pollution source of antibiotics entering the aquatic environment. In this work, the adsorption behavior of sulfamethazine (SMN), a commonly-used sulfonamide antibiotic, on activated sludge from a sequencing batch reactor treating swine wastewater was investigated. The results show that the adsorption of SMN on activated sludge was an initially rapid process and reached equilibrium after 6hr. The removal efficiency of SMN from the water phase increased with an increasing concentration of mixed liquor suspended solids, while the adsorbed concentration of SMN decreased. Solution pH influenced both the speciation of SMN and the surface properties of activated sludge, thus significantly impacting the adsorption process. A linear partition model could give a good fit for the equilibrium concentrations of SMN at the test temperatures (i.e., 10, 20 and 30°C). The partition coefficient (Kd) was determined to be 100.5L/kg at 20°C, indicating a quite high adsorption capacity for SMN. Thermodynamic analysis revealed that SMN adsorption on activated sludge was an exothermic process. This study could help to clarify the fate and behavior of sulfonamide antibiotics in the activated sludge process and assess consequent environmental risks arising from sludge disposal as well. Copyright © 2014. Published by Elsevier B.V.

Salicylic acid and nitric oxide alleviate high temperature induced oxidative damage in Lablab purpureus L plants by regulating bio-physical processes and DNA methylation.

PubMed

Rai, Krishna Kumar; Rai, Nagendra; Rai, Shashi Pandey

2018-07-01

Salicylic acid (SA) and sodium nitroprusside (SNP, NO donor) modulates plant growth and development processes and recent findings have also revealed their involvement in the regulation of epigenetic factors under stress condition. In the present study, some of these factors were comparatively studied in hyacinth bean plants subjected to high temperature (HT) environment (40-42 °C) with and without exogenous application of SA and SNP under field condition. Exogenous application of SA and SNP substantially modulated the growth and biophysical process of hyacinth bean plants under HT environment. Exogenous application of SA and SNP also remarkably regulated the activities of antioxidant enzymes, modulated mRNA level of certain enzymes, improves plant water relation, enhance photosynthesis and thereby increasing plant defence under HT. Coupled restriction enzyme digestion-random amplification (CRED-RA) technique revealed that many methylation changes were "dose dependent" and HT significantly increased DNA damages as evidenced by both increase and decrease in bands profiles, methylation and de-methylation pattern. Thus, the result of the present study clearly shows that exogenous SA and SNP regulates DNA methylation pattern, modulates stress-responsive genes and can impart transient HT tolerance by synchronizing growth and physiological acclimatization of plants, thus narrowing the gaps between physio-biochemical and molecular events in addressing HT tolerance. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

MTHFR methylation moderates the impact of smoking on DNA methylation at AHRR for African American young adults.

PubMed

Beach, Steven R H; Lei, Man Kit; Ong, Mei Ling; Brody, Gene H; Dogan, Meeshanthini V; Philibert, Robert A

2017-09-01

Smoking has been shown to have a large, reliable, and rapid effect on demethylation of AHRR, particularly at cg05575921, suggesting that methylation may be used as an index of cigarette consumption. Because the availability of methyl donors may also influence the degree of demethylation in response to smoking, factors that affect the activity of methylene tetrahydrofolate reductase (MTHFR), a key regulator of methyl group availability, may be of interest. In the current investigation, we examined the extent to which individual differences in methylation of MTHFR moderated the association between smoking and demethylation at cg05575921 as well as at other loci on AHRR associated with a main effect of smoking. Using a discovery sample (AIM, N = 293), and a confirmatory sample (SHAPE, N = 368) of young adult African Americans, degree of methylation of loci in the first exon of MTHFR was associated with amplification of the association between smoking and AHRR demethylation at cg05575921. However, genetic variation at a commonly studied MTHFR variant, C677T, did not influence cg05575921 methylation. The significant interaction between MTHFR methylation and the smoking-induced response at cg05575921 suggests a role for individual differences in methyl cycle regulation in understanding the effects of cigarette consumption on genome wide DNA methylation. © 2017 Wiley Periodicals, Inc.

Altered chromatin condensation of heat-stressed spermatozoa perturbs the dynamics of DNA methylation reprogramming in the paternal genome after in vitro fertilisation in cattle.

PubMed

Rahman, Mohammad Bozlur; Kamal, Md Mostofa; Rijsselaere, Tom; Vandaele, Leen; Shamsuddin, Mohammed; Van Soom, Ann

2014-10-01

Shortly after penetration of the oocyte, sperm DNA is actively demethylated, which is required for totipotent zygotic development. Aberrant DNA methylation is thought to be associated with altered chromatin condensation of spermatozoa. The objectives of this study were to investigate the dynamics of DNA methylation reprogramming in the paternal pronucleus and subsequent fertilisation potential of heat-stressed bull spermatozoa having altered chromatin condensation. Hence, bovine zygotes (n=1239) were collected at three different time points (12, 18 and 24h post insemination, hpi), and stained with an antibody against 5-methylcytosine. Fluorescence intensities of paternal and maternal pronuclei were measured by ImageJ. DNA methylation patterns in paternal pronuclei derived from heat-stressed spermatozoa did not differ between time points (P>0.05), whereas control zygotes clearly showed demethylation and de novo methylation at 18 and 24hpi, respectively. Moreover, heat-stressed spermatozoa showed a highly reduced (P<0.01) fertilisation rate compared with non-heat-stressed or normal control spermatozoa (53.7% vs 70.2% or 81.5%, respectively). Our data show that the normal pattern of active DNA demethylation followed by de novo methylation in the paternal pronucleus is perturbed when oocytes are fertilised with heat-stressed spermatozoa, which may be responsible for decreased fertilisation potential.

Proteolytic processing and activation of Clostridium perfringens epsilon toxin by caprine small intestinal contents.

PubMed

Freedman, John C; Li, Jihong; Uzal, Francisco A; McClane, Bruce A

2014-10-21

Epsilon toxin (ETX), a pore-forming toxin produced by type B and D strains of Clostridium perfringens, mediates severe enterotoxemia in livestock and possibly plays a role in human disease. During enterotoxemia, the nearly inactive ETX prototoxin is produced in the intestines but then must be activated by proteolytic processing. The current study sought to examine ETX prototoxin processing and activation ex vivo using the intestinal contents of a goat, a natural host species for ETX-mediated disease. First, this study showed that the prototoxin has a KEIS N-terminal sequence with a molecular mass of 33,054 Da. When the activation of ETX prototoxin ex vivo by goat small intestinal contents was assessed by SDS-PAGE, the prototoxin was processed in a stepwise fashion into an ~27-kDa band or higher-molecular-mass material that could be toxin oligomers. Purified ETX corresponding to the ~27-kDa band was cytotoxic. When it was biochemically characterized by mass spectrometry, the copresence of three ETX species, each with different C-terminal residues, was identified in the purified ~27-kDa ETX preparation. Cytotoxicity of each of the three ETX species was then demonstrated using recombinant DNA approaches. Serine protease inhibitors blocked the initial proteotoxin processing, while carboxypeptidase inhibitors blocked further processing events. Taken together, this study provides important new insights indicating that, in the intestinal lumen, serine protease (including trypsin and possibly chymotrypsin) initiates the processing of the prototoxin but other proteases, including carboxypeptidases, then process the prototoxin into multiple active and stable species. Importance: Processing and activation by intestinal proteases is a prerequisite for ETX-induced toxicity. Previous studies had characterized the activation of ETX using only arbitrarily chosen amounts of purified trypsin and/or chymotrypsin. Therefore, the current study examined ETX activation ex vivo by natural

Dynamic Stimuli And Active Processing In Human Visual Perception

NASA Astrophysics Data System (ADS)

Haber, Ralph N.

1990-03-01

Theories of visual perception traditionally have considered a static retinal image to be the starting point for processing; and has considered processing both to be passive and a literal translation of that frozen, two dimensional, pictorial image. This paper considers five problem areas in the analysis of human visually guided locomotion, in which the traditional approach is contrasted to newer ones that utilize dynamic definitions of stimulation, and an active perceiver: (1) differentiation between object motion and self motion, and among the various kinds of self motion (e.g., eyes only, head only, whole body, and their combinations); (2) the sources and contents of visual information that guide movement; (3) the acquisition and performance of perceptual motor skills; (4) the nature of spatial representations, percepts, and the perceived layout of space; and (5) and why the retinal image is a poor starting point for perceptual processing. These newer approaches argue that stimuli must be considered as dynamic: humans process the systematic changes in patterned light when objects move and when they themselves move. Furthermore, the processing of visual stimuli must be active and interactive, so that perceivers can construct panoramic and stable percepts from an interaction of stimulus information and expectancies of what is contained in the visual environment. These developments all suggest a very different approach to the computational analyses of object location and identification, and of the visual guidance of locomotion.

Design, Synthesis, and Structure--Activity Relationship of New 2-Aryl-3,4-dihydro-β-carbolin-2-ium Salts as Antifungal Agents.

PubMed

Hou, Zhe; Zhu, Li-Fei; Yu, Xin-chi; Sun, Ma-Qiang; Miao, Fang; Zhou, Le

2016-04-13

Twenty-two 2-aryl-9-methyl-3,4-dihydro-β-carbolin-2-ium bromides along with four 9-demethylated derivatives were synthesized and characterized by spectroscopic analysis. By using the mycelium growth rate method, the compounds were evaluated for antifungal activities in vitro against six plant pathogenic fungi, and structure-activity relationships (SAR) were derived. Almost all of the compounds showed obvious inhibition activity on each of the fungi at 150 μM. For all of the fungi, 10 of the compounds showed average inhibition rates of >80% at 150 μM, and most of their EC50 values were in the range of 2.0-30.0 μM. SAR analysis showed that the substitution pattern of the N-aryl ring significantly influences the activity; N9-alkylation improves the activity, whereas aromatization of ring-C reduces the activity. It was concluded that the present research provided a series of new 2-aryl-9-alkyl-3,4-dihydro-β-carbolin-2-iums with excellent antifungal potency and structure optimization design for the development of new carboline antifungal agents.

Dormancy activation mechanism of oral cavity cancer stem cells.

PubMed

Chen, Xiang; Li, Xin; Zhao, Baohong; Shang, Dehao; Zhong, Ming; Deng, Chunfu; Jia, Xinshan

2015-07-01

Radiotherapy and chemotherapy are targeted primarily at rapidly proliferating cancer cells and are unable to eliminate cancer stem cells in the G0 phase. Thus, these treatments cannot prevent the recurrence and metastasis of cancer. Understanding the mechanisms by which cancer stem cells are maintained in the dormant G0 phase, and how they become active is key to developing new cancer therapies. The current study found that the anti-cancer drug 5-fluorouracil, acting on the oral squamous cell carcinoma KB cell line, selectively killed proliferating cells while sparing cells in the G0 phase. Bisulfite sequencing PCR showed that demethylation of the Sox2 promoter led to the expression of Sox2. This then resulted in the transformation of cancer stem cells from the G0 phase to the division stage and suggested that the transformation of cancer stem cells from the G0 phase to the division stage is closely related to an epigenetic modification of the cell.

Optimizing the antioxidant activity of Kelakai (Stenochlaena palustris) through multiplestage extraction process

NASA Astrophysics Data System (ADS)

Wijaya, Elza; Widiputri, Diah Indriani; Rahmawati, Della

2017-11-01

Kelakai is known as traditional remedy for treating several diseases, such as fever, anemia, and stimulate the production of breast milk for breastfeeding mother. Instead of those benefits, kelakai also proved has several kinds of antioxidant properties. Therefore, extracting antioxidant properties from kelakai is one way to discover the amount of antioxidant activity contained in kelakai. In this research, the multiple-stage extraction process was done in order to optimize the antioxidant activity. Moreover, based on data obtained from single stage extraction process, the most suitable condition was discovered. It turns out that the use of milled sample in water solvent for 12 hours at 44°C produce the highest antioxidant activity, which is 919.95 mg to inhibit 50% of DPPH. Referred to the experiment, the antioxidant activity of the extract which gained from multiple-stage was higher than from single stage. Multiple-stage process has proven the increasing of antioxidant activity up to 72.43%, which is need 404 mg to inhibit 50% of DPPH.

Carbonate-activated hydrogen peroxide oxidation process for azo dye decolorization: Process, kinetics, and mechanisms.

PubMed

Li, Yang; Li, Lei; Chen, Zi-Xi; Zhang, Jie; Gong, Li; Wang, Yi-Xuan; Zhao, Han-Qing; Mu, Yang

2018-02-01

Advanced oxidation processes offer effective solutions in treating wastewater from various industries. This study is the first time to investigate the potential of carbonate-activated hydrogen peroxide (CAP) oxidation process for the removal of organic pollutant from highly alkaline wastewaters. Azo dye acid orange 7 (AO7) was selected as a model pollutant. The influences of various parameters on AO7 decolorization by the CAP oxidation were evaluated. Furthermore, the active species involved in AO7 degradation were explored using scavenger experiments and electron spin resonance analysis. Additionally, AO7 degradation products by the CAP oxidation were identified to elucidate possible transformation pathways. Results showed that the CAP oxidation had better AO7 decolorization performance compared to bicarbonate-activated hydrogen peroxide method. The AO7 decolorization efficiency augmented from 3.70 ± 0.76% to 54.27 ± 2.65% when carbonate concentration was increased from 0 to 50 mM at pH 13.0, and then changed slightly with further increasing carbonate concentration to 70 mM. It increased almost linearly from 5.95 ± 0.32% to 94.03 ± 0.39% as H 2 O 2 concentration was increased from 5 to 50 mM. Moreover, trace amount of Co(II) could facilitate AO7 decolorization by the CAP reaction. Superoxide and carbonate radicals might be the main reactive oxygen species involved in the CAP process. Finally, a possible degradation pathway of AO7 by the CAP oxidation was proposed based on the identified products. Copyright © 2017 Elsevier Ltd. All rights reserved.

Active control of complex, multicomponent self-assembly processes

NASA Astrophysics Data System (ADS)

Schulman, Rebecca

The kinetics of many complex biological self-assembly processes such as cytoskeletal assembly are precisely controlled by cells. Spatiotemporal control over rates of filament nucleation, growth and disassembly determine how self-assembly occurs and how the assembled form changes over time. These reaction rates can be manipulated by changing the concentrations of the components needed for assembly by activating or deactivating them. I will describe how we can use these principles to design driven self-assembly processes in which we assemble and disassemble multiple types of components to create micron-scale networks of semiflexible filaments assembled from DNA. The same set of primitive components can be assembled into many different, structures depending on the concentrations of different components and how designed, DNA-based chemical reaction networks manipulate these concentrations over time. These chemical reaction networks can in turn interpret environmental stimuli to direct complex, multistage response. Such a system is a laboratory for understanding complex active material behaviors, such as metamorphosis, self-healing or adaptation to the environment that are ubiquitous in biological systems but difficult to quantitatively characterize or engineer.

Disentangling brain activity related to the processing of emotional visual information and emotional arousal.

PubMed

Kuniecki, Michał; Wołoszyn, Kinga; Domagalik, Aleksandra; Pilarczyk, Joanna

2018-05-01

Processing of emotional visual information engages cognitive functions and induces arousal. We aimed to examine the modulatory role of emotional valence on brain activations linked to the processing of visual information and those linked to arousal. Participants were scanned and their pupil size was measured while viewing negative and neutral images. The visual noise was added to the images in various proportions to parametrically manipulate the amount of visual information. Pupil size was used as an index of physiological arousal. We show that arousal induced by the negative images, as compared to the neutral ones, is primarily related to greater amygdala activity while increasing visibility of negative content to enhanced activity in the lateral occipital complex (LOC). We argue that more intense visual processing of negative scenes can occur irrespective of the level of arousal. It may suggest that higher areas of the visual stream are fine-tuned to process emotionally relevant objects. Both arousal and processing of emotional visual information modulated activity within the ventromedial prefrontal cortex (vmPFC). Overlapping activations within the vmPFC may reflect the integration of these aspects of emotional processing. Additionally, we show that emotionally-evoked pupil dilations are related to activations in the amygdala, vmPFC, and LOC.

Structure-Activity Relationship of Benzophenanthridine Alkaloids from Zanthoxylum rhoifolium Having Antimicrobial Activity

PubMed Central

Tavares, Luciana de C.; Zanon, Graciane; Weber, Andréia D.; Neto, Alexandre T.; Mostardeiro, Clarice P.; Da Cruz, Ivana B. M.; Oliveira, Raul M.; Ilha, Vinicius; Dalcol, Ionara I.; Morel, Ademir F.

2014-01-01

Zanthoxylum rhoifolium (Rutaceae) is a plant alkaloid that grows in South America and has been used in Brazilian traditional medicine for the treatment of different health problems. The present study was designed to evaluate the antimicrobial activity of the steam bark crude methanol extract, fractions, and pure alkaloids of Z. rhoifolium. Its stem bark extracts exhibited a broad spectrum of antimicrobial activity, ranging from 12.5 to 100 µg/mL using bioautography method, and from 125 to 500 µg/mL in the microdilution bioassay. From the dichloromethane basic fraction, three furoquinoline alkaloids (1–3), and nine benzophenanthridine alkaloids (4–12) were isolated and the antimicrobial activity of the benzophenanthridine alkaloids is discussed in terms of structure-activity relationships. The alkaloid with the widest spectrum of activity was chelerythrine (10), followed by avicine (12) and dihydrochelerythrine (4). The minimal inhibitory concentrations of chelerythrine, of 1.50 µg/mL for all bacteria tested, and between 3.12 and 6.25 µg/mL for the yeast tested, show this compound to be a more powerful antimicrobial agent when compared with the other active alkaloids isolated from Z. rhoifolium. To verify the potential importance of the methylenedioxy group (ring A) of these alkaloids, chelerythrine was selected to represent the remainder of the benzophenanthridine alkaloids isolated in this work and was subjected to a demethylation reaction giving derivative 14. Compared to chelerythrine, the derivative (14) was less active against the tested bacteria and fungi. Kinetic measurements of the bacteriolytic activities of chelerythrine against the bacteria Bacillus subtilis (Gram-positive) and Escherichia coli (Gram-negative) were determined by optical density based on real time assay, suggesting that its mechanism of action is not bacteriolytic. The present study did not detect hemolytic effects of chelerythrine on erythrocytes and found a protective effect

Probing for the Activities of Arsenic and Selenium Metabolizing Microbes

NASA Astrophysics Data System (ADS)

Stolz, J. F.

2007-12-01

Microbial activities can directly impact the mobility and toxicity of arsenic and selenium in the environment. Arsenic is cycled through oxidation/reduction and methylation/demethylation reactions as part of resistance and respiratory processes. The requirement for selenium is primarily for incorporation into selenocysteine and its function in selenoenzymes. Selenium oxyanions can also serve as an electron acceptor in anaerobic respiration. Both culture and culture-independent methods have been developed to detect the presence and activity of organisms capable of arsenic and selenium transformations. Enrichment media have been successful at cultivating arsenate respiring bacteria from a variety of environments, however, both electron donor and the concentration of arsenic can exert strong selective pressure. Thus, the organisms in the enrichment culture may not be the dominant organisms in the environment. Culture-independent methods, including immunological approaches (e.g., polyclonal antibodies to ArrA) and PCR-based technologies, have also had mixed success. PCR-primers designed to amplify portions of genes involved in resistance (e.g., arsC, acr3), respiration (e.g., arrA), and oxidation (e.g., aoxB) have been useful in several environments. Applications include T-RFLP, rt-PCR, and DGGE analyses. Nevertheless, these primers do not work with certain organisms suggesting the existence of additional enzymes and pathways. Although the biosynthetic pathway (and the proteins involved) for selenocysteine has been described in detail, much less is known about selenium methylation, assimilation and respiration. Only one respiratory selenate reductase has been characterized and its close sequence identity with chlorate and perchlorate reductases has complicated efforts to design a functional probe. Thus many aspects of the biogeochemical cycle of selenium remains to be explored.

Drilling to investigate processes in active tectonics and magmatism

NASA Astrophysics Data System (ADS)

Shervais, J.; Evans, J.; Toy, V.; Kirkpatrick, J.; Clarke, A.; Eichelberger, J.

2014-12-01

Coordinated drilling efforts are an important method to investigate active tectonics and magmatic processes related to faults and volcanoes. The US National Science Foundation (NSF) recently sponsored a series of workshops to define the nature of future continental drilling efforts. As part of this series, we convened a workshop to explore how continental scientific drilling can be used to better understand active tectonic and magmatic processes. The workshop, held in Park City, Utah, in May 2013, was attended by 41 investigators from seven countries. Participants were asked to define compelling scientific justifications for examining problems that can be addressed by coordinated programs of continental scientific drilling and related site investigations. They were also asked to evaluate a wide range of proposed drilling projects, based on white papers submitted prior to the workshop. Participants working on faults and fault zone processes highlighted two overarching topics with exciting potential for future scientific drilling research: (1) the seismic cycle and (2) the mechanics and architecture of fault zones. Recommended projects target fundamental mechanical processes and controls on faulting, and range from induced earthquakes and earthquake initiation to investigations of detachment fault mechanics and fluid flow in fault zones. Participants working on active volcanism identified five themes: the volcano eruption cycle; eruption sustainability, near-field stresses, and system recovery; eruption hazards; verification of geophysical models; and interactions with other Earth systems. Recommended projects address problems that are transferrable to other volcanic systems, such as improved methods for identifying eruption history and constraining the rheological structure of shallow caldera regions. Participants working on chemical geodynamics identified four major themes: large igneous provinces (LIPs), ocean islands, continental hotspot tracks and rifts, and

Indicators of activity-friendly communities: an evidence-based consensus process.

PubMed

Brennan Ramirez, Laura K; Hoehner, Christine M; Brownson, Ross C; Cook, Rebeka; Orleans, C Tracy; Hollander, Marla; Barker, Dianne C; Bors, Philip; Ewing, Reid; Killingsworth, Richard; Petersmarck, Karen; Schmid, Thomas; Wilkinson, William

2006-12-01

Regular physical activity, even at modest intensities, is associated with many health benefits. Most Americans, however, do not engage in the recommended levels. As practitioners seek ways to increase population rates of physical activity, interventions and advocacy efforts are being targeted to the community level. Yet, advocates, community leaders, and researchers lack the tools needed to assess local barriers to and opportunities for more active, healthy lifestyles. Investigators used a systematic review process to identify key indicators of activity-friendly communities that can assess and improve opportunities for regular physical activity. Investigators conducted a comprehensive literature review of both peer-reviewed literature and fugitive information (e.g., reports and websites) to generate an initial list of indicators for review (n=230). The review included a three-tiered, modified Delphi consensus-development process that incorporated input of international, national, state, and local researchers and practitioners from academic institutions, federal and state government agencies, nonprofit organizations, and funding agencies in public health, transportation, urban planning, parks and recreation, and public policy. Ten promising indicators of activity-friendly communities were identified: land use environment, access to exercise facilities, transportation environment, aesthetics, travel patterns, social environment, land use economics, transportation economics, institutional and organizational policies, and promotion. Collaborative, multidisciplinary approaches are underway to test, refine, and expand this initial list of indicators and to develop measures that communities, community leaders, and policymakers can use to design more activity-friendly community environments.

[Treatment of carbonization effluent by the ultrasonic radiation and activated sludge process].

PubMed

Ning, Ping; Xu, Jinqiu; Huang, Dongbin; Ma, Xiaoli; Xu, Xiaojun; Li, Ziyan

2003-05-01

The paper deals with the degradation of organic pollutants by the ultrasonic irradiation-activated sludge process. The treatment of the real coking wastewater of Kunming coke making-gas plant was studied with the water quality model. Using the ultrasonic irradiation-activated sludge process the organic pollutants in the real coking wastewater can be degraded effectively. The influence factors of the ultrasonic degradation effect such as initial concentration, aerated gas and ultrasonic density were investigated and mechanism was explored. The result shows that the ultrasonic degradation effect was high with the decrease of initial concentration of the CODCr, the presence of aerated gas and the increase of ultrasonic density. At the initial CODCr concentration of 807 mg/L, when air acted as aerated gas and only air itself (no ultrasound) was exerted on the wastewater, the degradation rate of the CODCr will be 4.5%. However, when the ultrasound of the intensity of 119.4 kW/m2 was exerted on the wastewater, the degradation rate of the CODCr will be 65%. Compared with the activated sludge process alone, the combination of the ultrasonic irradiation and activated sludge process can increase the degradation rate of the CODCr from 45% to 81%. The oxygen consumption rate of the carbonization effluent obviously decreased in the presence of the activated sludge. This shows the carbonization effluent is not biotoxic behind the ultrasonic irradiation.

Materials and Process Activities for NASA's Composite Crew Module

NASA Technical Reports Server (NTRS)

Polis, Daniel L.

2012-01-01

In January 2007, the NASA Administrator and Associate Administrator for the Exploration Systems Mission Directorate chartered the NASA Engineering and Safety Center (NESC) to design, build, and test a full-scale Composite Crew Module (CCM). The overall goal of the CCM project was to develop a team from the NASA family with hands-on experience in composite design, manufacturing, and testing in anticipation of future space exploration systems being made of composite materials. The CCM project was planned to run concurrently with the Orion project s baseline metallic design within the Constellation Program so that features could be compared and discussed without inducing risk to the overall Program. The materials and process activities were prioritized based on a rapid prototype approach. This approach focused developmental activities on design details with greater risk and uncertainty, such as out-of-autoclave joining, over some of the more traditional lamina and laminate building block levels. While process development and associated building block testing were performed, several anomalies were still observed at the full-scale level due to interactions between process robustness and manufacturing scale-up. This paper describes the process anomalies that were encountered during the CCM development and the subsequent root cause investigations that led to the final design solutions. These investigations highlight the importance of full-scale developmental work early in the schedule of a complex composite design/build project.

Overview of materials processing in space activity at Marshall Space Flight Center

NASA Technical Reports Server (NTRS)

Williams, J. R.; Chassay, R. P.; Moore, W. W.; Ruff, R. C.; Yates, I. C.

1984-01-01

An overview of activities involving the Space Transportation System (STS), now in the operational phase, and results of some of the current space experiments, as well as future research opportunities in microgravity environment, are presented. The experiments of the Materials Processing in Space Program flown on the STS, such as bioseparation processes, isoelectric focusing, solidification and crystal growth processes, containerless processes, and the Materials Experiment Assembly experiments are discussed. Special consideration is given to the experiments to be flown aboard the Spacelab 3 module, the Fluids Experiments System, and the Vapor Crystal Growth System. Ground-based test facilities and planned space research facilities, as well as the nature of the commercialization activities, are briefly explained.

Color vision predicts processing modes of goal activation during action cascading.

PubMed

Jongkees, Bryant J; Steenbergen, Laura; Colzato, Lorenza S

2017-09-01

One of the most important functions of cognitive control is action cascading: the ability to cope with multiple response options when confronted with various task goals. A recent study implicates a key role for dopamine (DA) in this process, suggesting higher D1 efficiency shifts the action cascading strategy toward a more serial processing mode, whereas higher D2 efficiency promotes a shift in the opposite direction by inducing a more parallel processing mode (Stock, Arning, Epplen, & Beste, 2014). Given that DA is found in high concentration in the retina and modulation of retinal DA release displays characteristics of D2-receptors (Peters, Schweibold, Przuntek, & Müller, 2000), color vision discrimination might serve as an index of D2 efficiency. We used color discrimination, assessed with the Lanthony Desaturated Panel D-15 test, to predict individual differences (N = 85) in a stop-change paradigm that provides a well-established measure of action cascading. In this task it is possible to calculate an individual slope value for each participant that estimates the degree of overlap in task goal activation. When the stopping process of a previous task goal has not finished at the time the change process toward a new task goal is initiated (parallel processing), the slope value becomes steeper. In case of less overlap (more serial processing), the slope value becomes flatter. As expected, participants showing better color vision were more prone to activate goals in a parallel manner as indicated by a steeper slope. Our findings suggest that color vision might represent a predictor of D2 efficiency and the predisposed processing mode of goal activation during action cascading. Copyright © 2017 Elsevier Ltd. All rights reserved.

Biodegradation of Basic Violet 3 by Candida krusei isolated from textile wastewater.

PubMed

Deivasigamani, Charumathi; Das, Nilanjana

2011-11-01

Basic Violet 3 (BV) belongs to the most important group of synthetic colorants and is used extensively in textile industries. It is considered as xenobiotic compound which is recalcitrant to biodegradation. As Candida krusei could not use BV as sole carbon source, experiments were conducted to study the effect of cosubstrates on decolorization of BV in semi synthetic medium using glucose, sucrose, lactose, maltose, yeast extract, peptone, urea and ammonium sulphate. Maximum decolorization (74%) was observed in media supplemented with sucrose. Use of sugarcane bagasse extract as sole nutrient source showed 100% decolorization of BV within 24 h under optimized condition. UV-visible, FTIR spectral analysis and HPLC analysis confirmed the biodegradation of BV. Six degradation products were isolated and identified. We propose the biodegradation pathway for BV which occurs via stepwise reduction and demethylation process to yield mono-, di-, tri-, tetra-, penta- and hexa-demethylated BV species which was degraded completely. The study of the enzymes responsible for decolorization showed the activities of lignin peroxidase, lacasse, tyrosinase, NADH-DCIP reductase, MG reductase and azoreductase in cells before and after decolorization. A significant increase in activities of NADH-DCIP reductase and laccase was observed in the cells after decolorization. The yeast C. krusei could show the ability to decolorize the textile dye BV using inexpensive source like sugarcane bagasse extract for decolorization.

Zscan4 Inhibits Maintenance DNA Methylation to Facilitate Telomere Elongation in Mouse Embryonic Stem Cells.

PubMed

Dan, Jiameng; Rousseau, Philippe; Hardikar, Swanand; Veland, Nicolas; Wong, Jiemin; Autexier, Chantal; Chen, Taiping

2017-08-22

Proper telomere length is essential for embryonic stem cell (ESC) self-renewal and pluripotency. Mouse ESCs (mESCs) sporadically convert to a transient totipotent state similar to that of two-cell (2C) embryos to recover shortened telomeres. Zscan4, which exhibits a burst of expression in 2C-like mESCs, is required for telomere extension in these cells. However, the mechanism by which Zscan4 extends telomeres remains elusive. Here, we show that Zscan4 facilitates telomere elongation by inducing global DNA demethylation through downregulation of Uhrf1 and Dnmt1, major components of the maintenance DNA methylation machinery. Mechanistically, Zscan4 recruits Uhrf1 and Dnmt1 and promotes their degradation, which depends on the E3 ubiquitin ligase activity of Uhrf1. Blocking DNA demethylation prevents telomere elongation associated with Zscan4 expression, suggesting that DNA demethylation mediates the effect of Zscan4. Our results define a molecular pathway that contributes to the maintenance of telomere length homeostasis in mESCs. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Comparison of laboratory-scale thermophilic biofilm and activated sludge processes integrated with a mesophilic activated sludge process.

PubMed

Suvilampi, J; Lehtomäki, A; Rintala, J

2003-07-01

A combined thermophilic-mesophilic wastewater treatment was studied using a laboratory-scale thermophilic activated sludge process (ASP) followed by mesophilic ASP or a thermophilic suspended carrier biofilm process (SCBP) followed by mesophilic ASP, both systems treating diluted molasses (dilution factor 1:500 corresponding GF/A-filtered COD (COD(filt)) of 1900+/-190 mgl(-1)). With hydraulic retention times (HRTs) of 12-18 h the thermophilic ASP and thermophilic SCBP removed 60+/-13% and 62+/-7% of COD(filt), respectively, with HRT of 8 h the removals were 48+/-1% and 69+/-4%. The sludge volume index (SVI) was notably lower in the thermophilic SCBP (measured from suspended sludge) than in the thermophilic ASP. Under the lowest HRT the mesophilic ASP gave better performance (as SVI, COD(filt), and COD(tot) removals) after the thermophilic SCBP than after the thermophilic ASP. Measured sludge yields were low (less than 0.1 kg suspended solids (SS) kg COD(filt removed)(-1)) in all processes. Both thermophilic treatments removed 80-85% of soluble COD (COD(sol)) whereas suspended COD (COD(susp)) and colloidal COD (COD(col)) were increased. Both mesophilic post-treatments removed all COD(col) and most of the COD(susp) from the thermophilic effluents. In conclusion, combined thermophilic-mesophilic treatment appeared to be easily operable and produced high effluent quality.

Executive Functions in Learning Processes: Do They Benefit from Physical Activity?

ERIC Educational Resources Information Center

Barenberg, Jonathan; Berse, Timo; Dutke, Stephan

2011-01-01

As executive functions play an essential role in learning processes, approaches capable of enhancing executive functioning are of particular interest to educational psychology. Recently, the hypothesis has been advanced that executive functioning may benefit from changes in neurobiological processes induced by physical activity. The present…

Structural and Functional Coordination of DNA and Histone Methylation

PubMed Central

Cheng, Xiaodong

2014-01-01

One of the most fundamental questions in the control of gene expression in mammals is how epigenetic methylation patterns of DNA and histones are established, erased, and recognized. This central process in controlling gene expression includes coordinated covalent modifications of DNA and its associated histones. This article focuses on structural aspects of enzymatic activities of histone (arginine and lysine) methylation and demethylation and functional links between the methylation status of the DNA and histones. An interconnected network of methyltransferases, demethylases, and accessory proteins is responsible for changing or maintaining the modification status of specific regions of chromatin. PMID:25085914

Arginine methylation of HSP70 regulates retinoid acid-mediated RARβ2 gene activation

PubMed Central

Gao, Wei-wei; Xiao, Rong-quan; Peng, Bing-ling; Xu, Huan-teng; Shen, Hai-feng; Huang, Ming-feng; Shi, Tao-tao; Yi, Jia; Zhang, Wen-juan; Wu, Xiao-nan; Gao, Xiang; Lin, Xiang-zhi; Dorrestein, Pieter C.; Rosenfeld, Michael G.; Liu, Wen

2015-01-01

Although “histone” methyltransferases and demethylases are well established to regulate transcriptional programs and to use nonhistone proteins as substrates, their possible roles in regulation of heat-shock proteins in the nucleus have not been investigated. Here, we report that a highly conserved arginine residue, R469, in HSP70 (heat-shock protein of 70 kDa) proteins, an evolutionarily conserved protein family of ATP-dependent molecular chaperone, was monomethylated (me1), at least partially, by coactivator-associated arginine methyltransferase 1/protein arginine methyltransferase 4 (CARM1/PRMT4) and demethylated by jumonji-domain–containing 6 (JMJD6), both in vitro and in cultured cells. Functional studies revealed that HSP70 could directly regulate retinoid acid (RA)-induced retinoid acid receptor β2 (RARβ2) gene transcription through its binding to chromatin, with R469me1 being essential in this process. HSP70’s function in gene transcriptional regulation appears to be distinct from its protein chaperon activity. R469me1 was shown to mediate the interaction between HSP70 and TFIIH, which involves in RNA polymerase II phosphorylation and thus transcriptional initiation. Our findings expand the repertoire of nonhistone substrates targeted by PRMT4 and JMJD6, and reveal a new function of HSP70 proteins in gene transcription at the chromatin level aside from its classic role in protein folding and quality control. PMID:26080448

Dopamine modulates reward system activity during subconscious processing of sexual stimuli.

PubMed

Oei, Nicole Y L; Rombouts, Serge Arb; Soeter, Roelof P; van Gerven, Joop M; Both, Stephanie

2012-06-01

Dopaminergic medication influences conscious processing of rewarding stimuli, and is associated with impulsive-compulsive behaviors, such as hypersexuality. Previous studies have shown that subconscious subliminal presentation of sexual stimuli activates brain areas known to be part of the 'reward system'. In this study, it was hypothesized that dopamine modulates activation in key areas of the reward system, such as the nucleus accumbens, during subconscious processing of sexual stimuli. Young healthy males (n=53) were randomly assigned to two experimental groups or a control group, and were administered a dopamine antagonist (haloperidol), a dopamine agonist (levodopa), or placebo. Brain activation was assessed during a backward-masking task with subliminally presented sexual stimuli. Results showed that levodopa significantly enhanced the activation in the nucleus accumbens and dorsal anterior cingulate when subliminal sexual stimuli were shown, whereas haloperidol decreased activations in those areas. Dopamine thus enhances activations in regions thought to regulate 'wanting' in response to potentially rewarding sexual stimuli that are not consciously perceived. This running start of the reward system might explain the pull of rewards in individuals with compulsive reward-seeking behaviors such as hypersexuality and patients who receive dopaminergic medication.

The Role of Small-Scale Processes in Solar Active Region Decay

NASA Astrophysics Data System (ADS)

Meyer, Karen; Mackay, Duncan

2017-08-01

Active regions are locations of intense magnetic activity on the Sun, whose evolution can result in highly energetic eruptive phenomena such as solar flares and coronal mass ejections (CMEs). Therefore, fast and accurate simulation of their evolution and decay is essential in the prediction of Space Weather events. In this talk we present initial results from our new model for the photospheric evolution of active region magnetic fields. Observations show that small-scale processes appear to play a role in the dispersal and decay of solar active regions, for example through cancellation at the boundary of sunspot outflows and erosion of flux by surrounding convective cells. Our active region model is coupled to our existing model for the evolution of small-scale photospheric magnetic features. Focusing first on the active region decay phase, we consider the evolution of its magnetic field due to both large-scale (e.g. differential rotation) and small-scale processes, such as its interaction with surrounding small-scale magnetic features and convective flows.This project is funded by The Carnegie Trust for the Universities of Scotland, through their Research Incentives Grant scheme.

Integration of Metabolomics and In vitro Metabolism Assays for Investigating the Stereoselective Transformation of Triadimefon in Rainbow Trout

EPA Science Inventory

Triadimefon is a systemic agricultural fungicide of the triazole class whose major metabolite, triadimenol, also a commercial fungicide, provides the majority of the actual fungicidal activity; i.e., inhibition of steroid demethylation. Both chemicals are chiral: triadimefon has...

The influence of spontaneous activity on stimulus processing in primary visual cortex.

PubMed

Schölvinck, M L; Friston, K J; Rees, G

2012-02-01

Spontaneous activity in the resting human brain has been studied extensively; however, how such activity affects the local processing of a sensory stimulus is relatively unknown. Here, we examined the impact of spontaneous activity in primary visual cortex on neuronal and behavioural responses to a simple visual stimulus, using functional MRI. Stimulus-evoked responses remained essentially unchanged by spontaneous fluctuations, combining with them in a largely linear fashion (i.e., with little evidence for an interaction). However, interactions between spontaneous fluctuations and stimulus-evoked responses were evident behaviourally; high levels of spontaneous activity tended to be associated with increased stimulus detection at perceptual threshold. Our results extend those found in studies of spontaneous fluctuations in motor cortex and higher order visual areas, and suggest a fundamental role for spontaneous activity in stimulus processing. Copyright © 2011. Published by Elsevier Inc.

Validity of an Integrative Method for Processing Physical Activity Data.

PubMed

Ellingson, Laura D; Schwabacher, Isaac J; Kim, Youngwon; Welk, Gregory J; Cook, Dane B

2016-08-01

Accurate assessments of both physical activity and sedentary behaviors are crucial to understand the health consequences of movement patterns and to track changes over time and in response to interventions. The study evaluates the validity of an integrative, machine learning method for processing activity monitor data in relation to a portable metabolic analyzer (Oxycon mobile [OM]) and direct observation (DO). Forty-nine adults (age 18-40 yr) each completed 5-min bouts of 15 activities ranging from sedentary to vigorous intensity in a laboratory setting while wearing ActiGraph (AG) on the hip, activPAL on the thigh, and OM. Estimates of energy expenditure (EE) and categorization of activity intensity were obtained from the AG processed with Lyden's sojourn (SOJ) method and from our new sojourns including posture (SIP) method, which integrates output from the AG and activPAL. Classification accuracy and estimates of EE were then compared with criterion measures (OM and DO) using confusion matrices and comparisons of the mean absolute error of log-transformed data (MAE ln Q). The SIP method had a higher overall classification agreement (79%, 95% CI = 75%-82%) than the SOJ (56%, 95% CI = 52%-59%) based on DO. Compared with OM, estimates of EE from SIP had lower mean absolute error of log-transformed data than SOJ for light-intensity (0.21 vs 0.27), moderate-intensity (0.33 vs 0.42), and vigorous-intensity (0.16 vs 0.35) activities. The SIP method was superior to SOJ for distinguishing between sedentary and light activities as well as estimating EE at higher intensities. Thus, SIP is recommended for research in which accuracy of measurement across the full range of activity intensities is of interest.

LSD Increases Primary Process Thinking via Serotonin 2A Receptor Activation

PubMed Central

Kraehenmann, Rainer; Pokorny, Dan; Aicher, Helena; Preller, Katrin H.; Pokorny, Thomas; Bosch, Oliver G.; Seifritz, Erich; Vollenweider, Franz X.

2017-01-01

Rationale: Stimulation of serotonin 2A (5-HT2A) receptors by lysergic acid diethylamide (LSD) and related compounds such as psilocybin has previously been shown to increase primary process thinking – an ontologically and evolutionary early, implicit, associative, and automatic mode of thinking which is typically occurring during altered states of consciousness such as dreaming. However, it is still largely unknown whether LSD induces primary process thinking under placebo-controlled, standardized experimental conditions and whether these effects are related to subjective experience and 5-HT2A receptor activation. Therefore, this study aimed to test the hypotheses that LSD increases primary process thinking and that primary process thinking depends on 5-HT2A receptor activation and is related to subjective drug effects. Methods: Twenty-five healthy subjects performed an audio-recorded mental imagery task 7 h after drug administration during three drug conditions: placebo, LSD (100 mcg orally) and LSD together with the 5-HT2A receptor antagonist ketanserin (40 mg orally). The main outcome variable in this study was primary index (PI), a formal measure of primary process thinking in the imagery reports. State of consciousness was evaluated using the Altered State of Consciousness (5D-ASC) rating scale. Results: LSD, compared with placebo, significantly increased primary index (p < 0.001, Bonferroni-corrected). The LSD-induced increase in primary index was positively correlated with LSD-induced disembodiment (p < 0.05, Bonferroni-corrected), and blissful state (p < 0.05, Bonferroni-corrected) on the 5D-ASC. Both LSD-induced increases in primary index and changes in state of consciousness were fully blocked by ketanserin. Conclusion: LSD induces primary process thinking via activation of 5-HT2A receptors and in relation to disembodiment and blissful state. Primary process thinking appears to crucially organize inner experiences during both dreams and psychedelic

LSD Increases Primary Process Thinking via Serotonin 2A Receptor Activation.

PubMed

Kraehenmann, Rainer; Pokorny, Dan; Aicher, Helena; Preller, Katrin H; Pokorny, Thomas; Bosch, Oliver G; Seifritz, Erich; Vollenweider, Franz X

2017-01-01

Rationale: Stimulation of serotonin 2A (5-HT2A) receptors by lysergic acid diethylamide (LSD) and related compounds such as psilocybin has previously been shown to increase primary process thinking - an ontologically and evolutionary early, implicit, associative, and automatic mode of thinking which is typically occurring during altered states of consciousness such as dreaming. However, it is still largely unknown whether LSD induces primary process thinking under placebo-controlled, standardized experimental conditions and whether these effects are related to subjective experience and 5-HT2A receptor activation. Therefore, this study aimed to test the hypotheses that LSD increases primary process thinking and that primary process thinking depends on 5-HT2A receptor activation and is related to subjective drug effects. Methods: Twenty-five healthy subjects performed an audio-recorded mental imagery task 7 h after drug administration during three drug conditions: placebo, LSD (100 mcg orally) and LSD together with the 5-HT2A receptor antagonist ketanserin (40 mg orally). The main outcome variable in this study was primary index (PI), a formal measure of primary process thinking in the imagery reports. State of consciousness was evaluated using the Altered State of Consciousness (5D-ASC) rating scale. Results: LSD, compared with placebo, significantly increased primary index ( p < 0.001, Bonferroni-corrected). The LSD-induced increase in primary index was positively correlated with LSD-induced disembodiment ( p < 0.05, Bonferroni-corrected), and blissful state ( p < 0.05, Bonferroni-corrected) on the 5D-ASC. Both LSD-induced increases in primary index and changes in state of consciousness were fully blocked by ketanserin. Conclusion: LSD induces primary process thinking via activation of 5-HT2A receptors and in relation to disembodiment and blissful state. Primary process thinking appears to crucially organize inner experiences during both dreams and psychedelic

The Metabolism Distribution and Effect of Thiamethoxam After Oral Exposure in Mongolian racerunner (Eremias argus).

PubMed

Wang, Yinghuan; Zhang, Yang; Xu, Peng; Guo, Baoyuan; Li, Wei

2018-06-20

Systematically evaluation of the metabolism, distribution and effect of thiamethoxam in mongolian racerunner (Eremias argus) were carried out after oral exposure. The HPLC equipped with Q Exactive focus was used for identification and concentration analysis of thiamethoxam and its metabolites. Percutaneous and urine excretions were the primary ways for the elimination of thiamethoxam and its metabolites, and the limiting factor was urine output. Demethylation thiamethoxam and clothianidin were the main metabolites of thiamethoxam in lizard. The CYP3A4, CYP3A7 and CYP2C9 played a crucial role in the metabolism process. Aldehyde oxidase only dominated the nitro-reduction process of demethylation thiamethoxam and clothianidin. Glutathione S-transferase might be related to the clearance process of thiamethoxam and its metabolites. The findings indicated that thiamethoxam might pose potential carcinogenic and hepatic injury risk to lizards. The results enrich and supplement the knowledge of the environmental fate of thiamethoxam in reptiles.

Hanford's Simulated Low Activity Waste Cast Stone Processing

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Young

2013-08-20

Cast Stone is undergoing evaluation as the supplemental treatment technology for Hanford’s (Washington) high activity waste (HAW) and low activity waste (LAW). This report will only cover the LAW Cast Stone. The programs used for this simulated Cast Stone were gradient density change, compressive strength, and salt waste form phase identification. Gradient density changes show a favorable outcome by showing uniformity even though it was hypothesized differently. Compressive strength exceeded the minimum strength required by Hanford and greater compressive strength increase seen between the uses of different salt solution The salt waste form phase is still an ongoing process asmore » this time and could not be concluded.« less

Operational Control Procedures for the Activated Sludge Process: Appendix.

ERIC Educational Resources Information Center

West, Alfred W.

This document is the appendix for a series of documents developed by the National Training and Operational Technology Center describing operational control procedures for the activated sludge process used in wastewater treatment. Categories discussed include: control test data, trend charts, moving averages, semi-logarithmic plots, probability…

Between strain and tissue differences exist in global hydroxymethylation after acute ozone exposure.

EPA Science Inventory

Epigenetics have been increasingly recognized as a mechanism linking environment and gene expression with both normal physiologic function as well as disease states. Demethylation of cysteine residues, generally leading to gene activation, is an oxygen-dependent reaction and crea...

Fluconazole Binding and Sterol Demethylation in Three CYP51 Isoforms Indicate Differences in Active Site Topology

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bellamine, A.; Lepesheva, Galina I.; Waterman, Mike

2010-11-16

14{alpha}-Demethylase (CYP51) is a key enzyme in all sterol biosynthetic pathways (animals, fungi, plants, protists, and some bacteria), catalyzing the removal of the C-14 methyl group following cyclization of squalene. Based on mutations found in CYP51 genes from Candida albicans azole-resistant isolates obtained after fluconazole treatment of fungal infections, and using site-directed mutagenesis, we have found that fluconazole binding and substrate metabolism vary among three different CYP51 isoforms: human, fungal, and mycobacterial. In C. albicans, the Y132H mutant from isolates shows no effect on fluconazole binding, whereas the F145L mutant results in a 5-fold increase in its IC{sub 50} formore » fluconazole, suggesting that F145 (conserved only in fungal 14{alpha}-demethylases) interacts with this azole. In C. albicans, F145L accounts, in part, for the difference in fluconazole sensitivity reported between mammals and fungi, providing a basis for treatment of fungal infections. The C. albicans Y132H and human Y145H CYP51 mutants show essentially no effect on substrate metabolism, but the Mycobacterium tuberculosis F89H CYP51 mutant loses both its substrate binding and metabolism. Because these three residues align in the three isoforms, the results indicate that their active sites contain important structural differences, and further emphasize that fluconazole and substrate binding are uncoupled properties.« less

Novel, Highly Specific N-Demethylases Enable Bacteria To Live on Caffeine and Related Purine Alkaloids

PubMed Central

Summers, Ryan M.; Louie, Tai Man; Yu, Chi-Li; Gakhar, Lokesh; Louie, Kailin C.

2012-01-01

The molecular basis for the ability of bacteria to live on caffeine as a sole carbon and nitrogen source is unknown. Pseudomonas putida CBB5, which grows on several purine alkaloids, metabolizes caffeine and related methylxanthines via sequential N-demethylation to xanthine. Metabolism of caffeine by CBB5 was previously attributed to one broad-specificity methylxanthine N-demethylase composed of two subunits, NdmA and NdmB. Here, we report that NdmA and NdmB are actually two independent Rieske nonheme iron monooxygenases with N1- and N3-specific N-demethylation activity, respectively. Activity for both enzymes is dependent on electron transfer from NADH via a redox-center-dense Rieske reductase, NdmD. NdmD itself is a novel protein with one Rieske [2Fe-2S] cluster, one plant-type [2Fe-2S] cluster, and one flavin mononucleotide (FMN) per enzyme. All ndm genes are located in a 13.2-kb genomic DNA fragment which also contained a formaldehyde dehydrogenase. ndmA, ndmB, and ndmD were cloned as His6 fusion genes, expressed in Escherichia coli, and purified using a Ni-NTA column. NdmA-His6 plus His6-NdmD catalyzed N1-demethylation of caffeine, theophylline, paraxanthine, and 1-methylxanthine to theobromine, 3-methylxanthine, 7-methylxanthine, and xanthine, respectively. NdmB-His6 plus His6-NdmD catalyzed N3-demethylation of theobromine, 3-methylxanthine, caffeine, and theophylline to 7-methylxanthine, xanthine, paraxanthine, and 1-methylxanthine, respectively. One formaldehyde was produced from each methyl group removed. Activity of an N7-specific N-demethylase, NdmC, has been confirmed biochemically. This is the first report of bacterial N-demethylase genes that enable bacteria to live on caffeine. These genes represent a new class of Rieske oxygenases and have the potential to produce biofuels, animal feed, and pharmaceuticals from coffee and tea waste. PMID:22328667

Orchard factors associated with resistance and cross resistance to sterol demethylation inhibitor fungicides in populations of Venturia inaequalis from Pennsylvania.

PubMed

Pfeufer, Emily E; Ngugi, Henry K

2012-03-01

Orchard management practices, such as destroying of overwintered inoculum and limiting the number of fungicide applications, are often recommended as tactics for slowing the development of resistance to sterol demethylation-inhibitor (DMI) fungicides in populations of Venturia inaequalis. However, there is little quantitative evidence relating the use of such practices to levels of resistance in orchards. The aim of this study was to evaluate the sensitivity of V. inaequalis isolates from Pennsylvania to DMI fungicides, and to identify orchard management factors related to the incidence of resistant isolates. In total, 644 single-spore V. inaequalis cultures obtained from 20 apple orchards in 2008 or 2009 were tested for sensitivity to myclobutanil, fenbuconazole, or difenoconazole. Growers provided management history of the sampled plots. Widespread shifts toward resistance to the three fungicides were noted, with mean effective concentration for 50% inhibition (EC(50)) values of 2.136, 0.786, and 0.187 μg/ml for myclobutanil, fenbuconazole, and difenoconazole, respectively. Cross resistance to the three fungicides was documented in high correlation (Spearman's r > 0.6) between mean EC(50) values for 14 orchards. Based on a 0.5-μg/ml threshold, 66 and 26% of isolates were resistant to myclobutanil and fenbuconazole, respectively, and 22% were cross resistant to the two fungicides. A significant between-year shift toward increased resistance was noted in two of three orchards surveyed in both years. Failure to use dormant copper sprays, older trees, larger orchards, orchards with ≤10 cultivars, and application of >4 DMI sprays were positively correlated (0.0001 < P < 0.05) with the incidence of resistant isolates. Isolates from orchards with >4 DMI sprays were four times as likely to be resistant to fenbuconazole (odds ratio = 4.57; P = 0.015). Isolates from orchards without dormant copper sprays were twice as likely to be cross-shifted toward resistance to all

The Aorta-Gonad-Mesonephros Organ Culture Recapitulates 5hmC Reorganization and Replication-Dependent and Independent Loss of DNA Methylation in the Germline.

PubMed

Calvopina, Joseph Hargan; Cook, Helene; Vincent, John J; Nee, Kevin; Clark, Amander T

2015-07-01

Removal of cytosine methylation from the genome is critical for reprogramming and transdifferentiation and plays a central role in our understanding of the fundamental principles of embryo lineage development. One of the major models for studying cytosine demethylation is the mammalian germ line during the primordial germ cell (PGC) stage of embryo development. It is now understood that oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) is required to remove cytosine methylation in a locus-specific manner in PGCs; however, the mechanisms downstream of 5hmC are controversial and hypothesized to involve either active demethylation or replication-coupled loss. In the current study, we used the aorta-gonad-mesonephros (AGM) organ culture model to show that this model recapitulates germ line reprogramming, including 5hmC reorganization and loss of cytosine methylation from Snrpn and H19 imprinting control centers (ICCs). To directly address the hypothesis that cell proliferation is required for cytosine demethylation, we blocked PI3-kinase-dependent PGC proliferation and show that this leads to a G1 and G2/M cell cycle arrest in PGCs, together with retained levels of cytosine methylation at the Snrpn ICC, but not at the H19 ICC. Taken together, the AGM organ culture model is an important tool to evaluate mechanisms of locus-specific demethylation and the role of PI3-kinase-dependent PGC proliferation in the locus-specific removal of cytosine methylation from the genome.

Enzyme induction in neonates after fetal exposure to antiepileptic drugs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rating, D.; Jaeger-Roman, E.; Nau, H.

1983-01-01

The /sup 13/C-AP breath test is shown to be a convenient, noninvasive method to monitor velocity and capacity of P450-dependent AP N-demethylation in infancy and childhood. According to /sup 13/C-AP breath tests, neonates have a very low capacity to eliminate /sup 13/CO/sub 2/, which is only 15 to 21% of the activity in adults. During the first year of life AP N-demethylation increases to reach its maximum at about 2 years; afterwards a slight decrease occurs. In 25 neonates exposed prenatally to different antiepileptic drugs /sup 13/C-AP breath test was efficiently used to prove that cytochrome AP N-demethylation was considerablymore » stimulated. After primidone/phenobarbitone, especially in combination with phenytoin, /sup 13/C elimination reaches and even surpasses the range for older children. Valproate exposure during fetal life is not consistently followed by a significant increase in AP N-demethylation. The enzyme induction demonstrated by /sup 13/C-AP breath test was often accompanied by accelerated metabolic clearance and shortened half-life times of transplacentally acquired antiepileptic drugs. There was good agreement between /sup 13/C-AP breath tests and pharmacokinetic data for primidone/phenobarbitone but not for phenytoin. In contrast, in the case of phenytoin exposure during pregnancy the pharmacokinetic parameters and the /sup 13/C breath test data will transport very different informations about enzyme induction in these neonates.« less

Development of a heat-processing method for koji to enhance its antioxidant activity.

PubMed

Okutsu, Kayu; Yoshizaki, Yumiko; Takamine, Kazunori; Tamaki, Hisanori; Ito, Kiyoshi; Sameshima, Yoshihiro

2012-03-01

We developed a heat-processing method to enhance the antioxidant activity of koji. The superoxide anion scavenging activity (SOSA) and oxygen radical absorbance capacity (ORAC) of heat-processed koji (HP-koji) at 55 °C for 7 days were 4.9 times and 4.2 times, respectively, those of unheated koji. These results showed that heat processing effectively enhances the antioxidant activity of koji. Analysis of the antioxidant activities of koji subjected to a range of temperatures (45-75 °C) revealed that the SOSA is enhanced by heating at higher temperatures, which might be catalyzed by Maillard reaction, whereas the ORAC was enhanced by heating at lower temperatures, which might be catalyzed by an enzymatic reaction. Assuming these enhancements in antioxidant activities are contributed by both Maillard and enzyme reactions, we hypothesized that the antioxidant activity of HP-koji could be more effectively amplified by heating at a higher temperature after the progression of the enzymatic reaction at a moderate temperature. Therefore, we evaluated the effect of heating of koji in a stepwise manner, first at 55 °C for 2 days and then at 75 °C for 5days. The antioxidant activities of stepwise-heated HP-koji were higher than those of koji heated at either 55 °C or 75 °C. The SOSA and ORAC of stepwise-heated HP-koji were 94 times and 6 times, respectively, those of unheated koji. This result suggests that enzymatic reaction followed by Maillard reaction can effectively enhance the antioxidant activity of HP-koji. Thus, we developed a novel heat-processing method to enhance the antioxidant activity of koji. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

USING CHIRALITY TO INFORM THE METABOLISM OF TRIADIMEFON TO TRIADIMENOL: A CROSS-SPECIES EXAMINATION

EPA Science Inventory

Triadimefon is a systemic agricultural fungicide of the conazole class whose metabolite, triadimenol, provides the majority of the actual fungicidal activity; i.e. inhibition of steroid demethylation. Triadimenol is also registered and used as a fungicide. Both chemicals are ch...

Land processes distributed active archive center product lifecycle plan

USGS Publications Warehouse

Daucsavage, John C.; Bennett, Stacie D.

2014-01-01

The U.S. Geological Survey (USGS) Earth Resources Observation and Science (EROS) Center and the National Aeronautics and Space Administration (NASA) Earth Science Data System Program worked together to establish, develop, and operate the Land Processes (LP) Distributed Active Archive Center (DAAC) to provide stewardship for NASA’s land processes science data. These data are critical science assets that serve the land processes science community with potential value beyond any immediate research use, and therefore need to be accounted for and properly managed throughout their lifecycle. A fundamental LP DAAC objective is to enable permanent preservation of these data and information products. The LP DAAC accomplishes this by bridging data producers and permanent archival resources while providing intermediate archive services for data and information products.

Measuring temperature-dependent activation energy in thermally activated processes: a 2D Arrhenius plot method.

PubMed

Li, Jian V; Johnston, Steven W; Yan, Yanfa; Levi, Dean H

2010-03-01

Thermally activated processes are characterized by two key quantities, activation energy (E(a)) and pre-exponential factor (nu(0)), which may be temperature dependent. The accurate measurement of E(a), nu(0), and their temperature dependence is critical for understanding the thermal activation mechanisms of non-Arrhenius processes. However, the classic 1D Arrhenius plot-based methods cannot unambiguously measure E(a), nu(0), and their temperature dependence due to the mathematical impossibility of resolving two unknown 1D arrays from one 1D experimental data array. Here, we propose a 2D Arrhenius plot method to solve this fundamental problem. Our approach measures E(a) at any temperature from matching the first and second moments of the data calculated with respect to temperature and rate in the 2D temperature-rate plane, and therefore is able to unambiguously solve E(a), nu(0), and their temperature dependence. The case study of deep level emission in a Cu(In,Ga)Se(2) solar cell using the 2D Arrhenius plot method reveals clear temperature dependent behavior of E(a) and nu(0), which has not been observable by its 1D predecessors.

Right Occipital Cortex Activation Correlates with Superior Odor Processing Performance in the Early Blind

PubMed Central

Grandin, Cécile B.; Dricot, Laurence; Plaza, Paula; Lerens, Elodie; Rombaux, Philippe; De Volder, Anne G.

2013-01-01

Using functional magnetic resonance imaging (fMRI) in ten early blind humans, we found robust occipital activation during two odor-processing tasks (discrimination or categorization of fruit and flower odors), as well as during control auditory-verbal conditions (discrimination or categorization of fruit and flower names). We also found evidence for reorganization and specialization of the ventral part of the occipital cortex, with dissociation according to stimulus modality: the right fusiform gyrus was most activated during olfactory conditions while part of the left ventral lateral occipital complex showed a preference for auditory-verbal processing. Only little occipital activation was found in sighted subjects, but the same right-olfactory/left-auditory-verbal hemispheric lateralization was found overall in their brain. This difference between the groups was mirrored by superior performance of the blind in various odor-processing tasks. Moreover, the level of right fusiform gyrus activation during the olfactory conditions was highly correlated with individual scores in a variety of odor recognition tests, indicating that the additional occipital activation may play a functional role in odor processing. PMID:23967263

Overexpression of ShCYP51B and ShatrD in Sclerotinia homoeocarpa Isolates Exhibiting Practical Field Resistance to a Demethylation Inhibitor Fungicide

PubMed Central

Hulvey, Jon; Popko, James T.; Sang, Hyunkyu; Berg, Andrew

2012-01-01

We investigated genetic factors that govern the reduced propiconazole sensitivity of Sclerotinia homoeocarpa field isolates collected during a 2-year field efficacy study on dollar spot disease of turf in five New England sites. These isolates displayed a >50-fold range of in vitro sensitivity to a sterol demethylation inhibitor (DMI) fungicide, propiconazole, making them ideal for investigations of genetic mechanisms of reduced DMI sensitivity. The CYP51 gene homolog in S. homoeocarpa (ShCYP51B), encoding the enzyme target of DMIs, is likely a minor genetic factor for reduced propiconazole sensitivity, since there were no differences in constitutive relative expression (RE) values and only 2-fold-higher induced RE values for insensitive than for sensitive isolate groups. Next, we mined RNA-Seq transcriptome data for additional genetic factors and found evidence for the overexpression of a homolog of Botrytis cinerea atrD (BcatrD), ShatrD, a known efflux transporter of DMI fungicides. The ShatrD gene showed much higher constitutive and induced RE values for insensitive isolates. Several polymorphisms were found upstream of ShatrD but were not definitively linked to overexpression. The screening of constitutive RE values of ShCYP51B and ShatrD in isolates from two golf courses that exhibited practical field resistance to propiconazole uncovered evidence for significant population-specific overexpression of both genes. However, linear regression demonstrated that the RE of ShatrD displays a more significant relationship with propiconazole sensitivity than that of ShCYP51B. In summary, our results suggest that efflux is a major determinant of the reduced DMI sensitivity of S. homoeocarpa genotypes in New England, which may have implications for the emergence of practical field resistance in this important turfgrass pathogen. PMID:22798361

Methods for Measuring Specific Rates of Mercury Methylation and Degradation and Their Use in Determining Factors Controlling Net Rates of Mercury Methylation

PubMed Central

Ramlal, Patricia S.; Rudd, John W. M.; Hecky, Robert E.

1986-01-01

A method was developed to estimate specific rates of demethylation of methyl mercury in aquatic samples by measuring the volatile 14C end products of 14CH3HgI demethylation. This method was used in conjunction with a 203Hg2+ radiochemical method which determines specific rates of mercury methylation. Together, these methods enabled us to examine some factors controlling the net rate of mercury methylation. The methodologies were field tested, using lake sediment samples from a recently flooded reservoir in the Southern Indian Lake system which had developed a mercury contamination problem in fish. Ratios of the specific rates of methylation/demethylation were calculated. The highest ratios of methylation/demethylation were calculated. The highest ratios of methylation/demethylation occurred in the flooded shorelines of Southern Indian Lake. These results provide an explanation for the observed increases in the methyl mercury concentrations in fish after flooding. PMID:16346959

Plan before You Play: An Activity for Teaching the Managerial Process

ERIC Educational Resources Information Center

Althouse, Norm R.; Hedges, Peggy L.

2015-01-01

This article describes a 60-minute classroom activity using LEGO® bricks that demonstrates and reinforces the importance of the managerial process. The activity, Plan Before You Play (PBP), is targeted to introductory business classes, and differs from others in that it requires little investment or up-front planning, is easily scalable, and, with…

Mercury Methylation in Alaskan Peatlands Spanning a Large Range of Trophic Structure

NASA Astrophysics Data System (ADS)

Krabbenhoft, D. P.; Zhang, L.; Hines, M. E.; Barkay, T.; Schaefer, J.; Aiken, G.

2015-12-01

The process of mercury (Hg) methylation has long been recognized as a key area of research in order to understand spatial and temporal variability of toxic methylmercury (MeHg) on the landscape. Numerous factors affect MeHg production, the most important generally falling into those that affect inorganic Hg(II) bioavailability (e.g., Hg(II) concentration and ligand composition), and those that affect microbial community composition and activity. The principal goal of this project is to decipher the details of MeHg production in Alaskan peatlands exhibiting a range of trophic status, including those lacking in electron acceptors that support the traditional respiratory pathway of MeHg production (e.g., sulfate reduction). MeHg production is carried out by a diverse group of microorganisms that possess the gene cluster (hgcAB), including the well-studied sulfate and iron- reducing bacteria (SRB and FeRB). However, less well known bacteria also possess the hgcAB genes, including: syntrophs, methanogens, acetogens, and fermenters. Methylation and demethylation activities were determined by injecting trace levels of the stable isotope (198Hg and Me204Hg) into intact peat cores. In addition, the short-lived radioisotope 197Hg was used in laboratory incubations. Laboratory studies also included assays for changes in diagnostic gas concentrations (CH4, CO2, H2) and LMW organic acids (formate, acetate, propionate, butyrate) to infer specific microbial processes, and the use of genomics to confirm microbial assemblages and the presence/absence of hgcAB genes. Overall, we observed Hg methylation rates were greatest at minerotrophic sites with active syntrophy and methanogenesis. Methylation and demethylation rates corresponded significantly across sites. There was no evidence of SO4- reduction in these samples, and addition of SO4- did not stimulate methylation suggesting that methylation was conducted by SRB that were metabolizing syntrophically and/or by fermentation.

Effect of Thermal Processing and Maceration on the Antioxidant Activity of White Beans

PubMed Central

Huber, Karina; Brigide, Priscila; Bretas, Eloá Bolis; Canniatti-Brazaca, Solange Guidolin

2014-01-01

Phenolic compounds, which naturally occur in beans, are known to have antioxidant activity, which may be partially lost during the processing of this legume. This study evaluated the effect of thermal processing and maceration on the phenolic acid and flavonoids profile and content and on the antioxidant activity of white beans. According to the results obtained from the 2,2-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) method, there were no significant differences among treatment groups analysed. When was using 1,1-diphenyl-2-pycrylhydrazyl method (DPPH), beans cooked without maceration present the higher antioxidant activity, and raw beans the lower. The phenolic acids found in greater amounts were gallic acid and chlorogenic acid. Kaempferol was only detected in the soaked and cooked samples; catechin and kaempferol-3-rutinoside were found in the highest concentrations. Quercetin and kaempferol-3-glucoside were not affected by the cooking process, either with or without maceration. In general, the heat treatment increased the antioxidant activity. PMID:24991931

Hold My Calls: An Activity for Introducing the Statistical Process

ERIC Educational Resources Information Center

Abel, Todd; Poling, Lisa

2015-01-01

Working with practicing teachers, this article demonstrates, through the facilitation of a statistical activity, how to introduce and investigate the unique qualities of the statistical process including: formulate a question, collect data, analyze data, and interpret data.

Fabrication of Ag-decorated BiOBr-mBiVO4 dual heterojunction composite with enhanced visible light photocatalytic performance for degradation of malachite green

NASA Astrophysics Data System (ADS)

Regmi, Chhabilal; Dhakal, Dipesh; Kim, Tae-Ho; Yamaguchi, Takutaro; Wohn Lee, Soo

2018-04-01

A visible light active Ag-decorated BiVO4-BiOBr dual heterojunction photocatalyst was prepared using a facile hydrothermal method, followed by the photodeposition of Ag. The photocatalytic activity of the synthesized samples was investigated by monitoring the change in malachite green (MG) concentration upon visible light irradiation. The synthesized sample was highly effective for the degradation of non-biodegradable MG. The enhanced activity observed was ascribed to the efficient separation and transfer of charge carriers across the dual heterojunction structure as verified by photoluminescence measurements. The removal of MG was primarily initiated by hydroxyl radicals and holes based on scavenger’s effect. To gain insight into the degradation mechanism, both high performance liquid chromatography and high resolution-quantitative time of flight, electrospray ionization mass spectrometry measurements during the degradation process were carried out. The degradation primarily followed the hydroxylation and N-demethylation process. A possible reaction pathway is proposed on the basis of all the information obtained under various experimental conditions.

η-Secretase processing of APP inhibits neuronal activity in the hippocampus.

PubMed

Willem, Michael; Tahirovic, Sabina; Busche, Marc Aurel; Ovsepian, Saak V; Chafai, Magda; Kootar, Scherazad; Hornburg, Daniel; Evans, Lewis D B; Moore, Steven; Daria, Anna; Hampel, Heike; Müller, Veronika; Giudici, Camilla; Nuscher, Brigitte; Wenninger-Weinzierl, Andrea; Kremmer, Elisabeth; Heneka, Michael T; Thal, Dietmar R; Giedraitis, Vilmantas; Lannfelt, Lars; Müller, Ulrike; Livesey, Frederick J; Meissner, Felix; Herms, Jochen; Konnerth, Arthur; Marie, Hélène; Haass, Christian

2015-10-15

Alzheimer disease (AD) is characterized by the accumulation of amyloid plaques, which are predominantly composed of amyloid-β peptide. Two principal physiological pathways either prevent or promote amyloid-β generation from its precursor, β-amyloid precursor protein (APP), in a competitive manner. Although APP processing has been studied in great detail, unknown proteolytic events seem to hinder stoichiometric analyses of APP metabolism in vivo. Here we describe a new physiological APP processing pathway, which generates proteolytic fragments capable of inhibiting neuronal activity within the hippocampus. We identify higher molecular mass carboxy-terminal fragments (CTFs) of APP, termed CTF-η, in addition to the long-known CTF-α and CTF-β fragments generated by the α- and β-secretases ADAM10 (a disintegrin and metalloproteinase 10) and BACE1 (β-site APP cleaving enzyme 1), respectively. CTF-η generation is mediated in part by membrane-bound matrix metalloproteinases such as MT5-MMP, referred to as η-secretase activity. η-Secretase cleavage occurs primarily at amino acids 504-505 of APP695, releasing a truncated ectodomain. After shedding of this ectodomain, CTF-η is further processed by ADAM10 and BACE1 to release long and short Aη peptides (termed Aη-α and Aη-β). CTFs produced by η-secretase are enriched in dystrophic neurites in an AD mouse model and in human AD brains. Genetic and pharmacological inhibition of BACE1 activity results in robust accumulation of CTF-η and Aη-α. In mice treated with a potent BACE1 inhibitor, hippocampal long-term potentiation was reduced. Notably, when recombinant or synthetic Aη-α was applied on hippocampal slices ex vivo, long-term potentiation was lowered. Furthermore, in vivo single-cell two-photon calcium imaging showed that hippocampal neuronal activity was attenuated by Aη-α. These findings not only demonstrate a major functionally relevant APP processing pathway, but may also indicate potential

Activation of bean (Phaseolus vulgaris) [alpha]-amylase inhibitor requires proteolytic processing of the proprotein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pueyo, J.J.; Hunt, D.C.; Chrispeels, M.J.

Seeds of the common bean (Phaseolus vulgaris) contain a plant defense protein that inhibits the [alpha]-amylases of mammals and insects. This [alpha]-amylase inhibitor ([alpha]Al) is synthesized as a proprotein on the endoplasmic reticulum and is proteolytically processed after arrival in the protein storage vacuoles to polypeptides of relative molecular weight (M[sub r]) 15,000 to 18,000. The authors report two types of evidence that proteolytic processing is linked to activation of the inhibitory activity. First, by surveying seed extracts of wild accessions of P. vulgaris and other species in the genus Phaseolus, they found that antibodies to [alpha]Al recognize large (M[submore » r] 30,000-35,000) polypeptides as well as typical [alpha]Al processing products (M[sub r] 15,000-18,000). [alpha]Al activity was found in all extracts that had the typical [alpha]Al processed polypeptides, but was absent from seed extracts that lacked such polypeptides. Second, they made a mutant [alpha]Al in which asparagine-77 is changed to aspartic acid-77. This mutation slows down the proteolytic processing of pro-[alpha]Al when the gene is expressed in tobacco. When pro-[alpha]Al was separated from mature [alpha]Al by gel filtration, pro-[alpha]Al was found not to have [alpha]-amylase inhibitory activity. The authors interpret these results to mean that formation of the active inhibitor is causally related to proteolytic processing of the proprotein. They suggest that the polypeptide cleavage removes a conformation constraint on the precursor to produce the biochemically active molecule. 43 refs., 5 figs., 1 tab.« less

A neural network ActiveX based integrated image processing environment.

PubMed

Ciuca, I; Jitaru, E; Alaicescu, M; Moisil, I

2000-01-01

The paper outlines an integrated image processing environment that uses neural networks ActiveX technology for object recognition and classification. The image processing environment which is Windows based, encapsulates a Multiple-Document Interface (MDI) and is menu driven. Object (shape) parameter extraction is focused on features that are invariant in terms of translation, rotation and scale transformations. The neural network models that can be incorporated as ActiveX components into the environment allow both clustering and classification of objects from the analysed image. Mapping neural networks perform an input sensitivity analysis on the extracted feature measurements and thus facilitate the removal of irrelevant features and improvements in the degree of generalisation. The program has been used to evaluate the dimensions of the hydrocephalus in a study for calculating the Evans index and the angle of the frontal horns of the ventricular system modifications.

Brain activity patterns induced by interrupting the cognitive processes with online advertising.

PubMed

Rejer, Izabela; Jankowski, Jarosław

2017-11-01

As a result of the increasing role of online advertising and strong competition among advertisers, intrusive techniques are commonly used to attract web users' attention. Moreover, since marketing content is usually delivered to the target audience when they are performing typical online tasks, like searching for information or reading online content, its delivery interrupts the web user's current cognitive process. The question posed by many researchers in the field of online advertising is: how should we measure the influence of interruption of cognitive processes on human behavior and emotional state? Much research has been conducted in this field; however, most of this research has focused on monitoring activity in the simulated environment, or processing declarative responses given by users in prepared questionnaires. In this paper, a more direct real-time approach is taken, and the effect of the interruption on a web user is analyzed directly by studying the activity of his brain. This paper presents the results of an experiment that was conducted to find the brain activity patterns associated with interruptions of the cognitive process by showing internet advertisements during a text-reading task. Three specific aspects were addressed in the experiment: individual patterns, the consistency of these patterns across trials, and the intra-subject correlation of the individual patterns. Two main effects were observed for most subjects: a drop in activity in the frontal and prefrontal cortical areas across all frequency bands, and significant changes in the frontal/prefrontal asymmetry index.

Activities of the Institute of Chemical Processing of Coal at Zabrze

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dreszer, K.

1995-12-31

The Institute of Chemical Processing of Coal at Zabrze was established in 1955. The works on carbochemical technologies have been, therefore, carried out at the Institute for 40 years. The targets of the Institute`s activities are research, scientific and developing works regarding a sensible utilization of fuels via their processing into more refined forms, safe environment, highly efficient use of energy carriers and technological products of special quality. The Institute of Chemical Processing of Coal has been dealing with the following: optimized use of home hard coals; improvement of classic coal coking technologies, processing and utilization of volatile coking products;more » production technologies of low emission rate fuels for communal management; analyses of coal processing technologies; new technologies aimed at increasing the efficiency of coal utilization for energy-generating purposes, especially in industry and studies on the ecological aspects of these processes; production technologies of sorbents and carbon activating agents and technologies of the utilization; rationalization of water and wastes management in the metallurgical and chemical industries in connection with removal of pollution especially dangerous to the environment from wastes; utilization technologies of refined materials (electrode cokes, binders, impregnating agents) for making electrodes, refractories and new generation construction carbon materials; production technologies of high quality bituminous and bituminous and resin coating, anti-corrosive and insulation materials; environmentally friendly utilization technologies for power station, mine and other wastes, and dedusting processes in industrial gas streams.« less

Stabilization of waste-activated sludge through the anoxic-aerobic digestion process

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hashimoto, S.; Fujita, M.; Terai, K.

1982-08-01

During the aerobic digestion process, the nitrogen which had been embedded in the activated sludge is solubilized to form ammoniacal and nitric nitrogen which are in turn transferred to the liquor and cause the increase of nitrogen loading in the sewage treatment plant. In this study, the anoxic-aerobic sludge digestion system which is a modified form of the conventional aerobic sludge digestion is made up of aerobic and anoxic tanks and are designed to remove both the volatile suspended solids and the total nitrogen (TN) simultaneously. The removal efficiencies of both VSS and TN were investigated by feeding waste-activated sludgemore » continuously and semicontinuously. The maximum percent reduction of both VSS and TN was achieved at a Q /SUB r/ /Q /SUB s/ ratio of 2 in the continuous process. The semicontinuous process was used to improve the nitrogen removal efficiency further. In the semicontinuous process, the VSS reduction efficiency as well as the nitrogen removal efficiency increased remarkably under a constant Q /SUB r/ /Q /SUB s/ ratio of 2. This process also achieved a VSS reduction efficiency higher than the aerobic digestion process (control). It was suggested that the additional anoxic tank enhanced the sludge digestion. Furthermore, the anoxic-aerobic digestion system can be applied to other treatment media like the primary sludge, industrial sludge, animal manure, etc.« less

Stabilization of waste-activated sludge through the anoxic-aerobic digestion process.

PubMed

Hashimoto, S; Fujita, M; Terai, K

1982-08-01

During the aerobic digestion process, the nitrogen which had been embedded in the activated sludge is solubilized to form ammoniacal and nitric nitrogen which are in turn transferred to the liquor and cause the increase of nitrogen loading in the sewage treatment plant. In this study, the anoxic-aerobic sludge digestion system which is a modified form of the conventional aerobic sludge digestion is made up of aerobic and anoxic tanks and are designed to remove both the volatile suspended solids and the total nitrogen (TN) simultaneously. The removal efficiencies of both VSS and TN were investigated by feeding waste-activated sludge continuously and semicontinuously. The maximum percent reduction of both VSS and TN was achieved at a Q(r)/Q(s) ratio of 2 in the continuous process. The semicontinuous process was used to improve the nitrogen removal efficiency further. In the semicontinuous process, the VSS reduction efficiency as well as the nitrogen removal efficiency increased remarkably under a constant Q(r)/Q(s) ratio of 2. This process also achieved a VSS reduction efficiency higher than the aerobic digestion process (control). It was suggested that the additional anoxic tank enhanced the sludge digestion. Furthermore, the anoxic-aerobic digestion system can be applied to other treatment media like the primary sludge, industrial sludge, animal manure, etc.

Higher-order cognitive training effects on processing speed-related neural activity: a randomized trial.

PubMed

Motes, Michael A; Yezhuvath, Uma S; Aslan, Sina; Spence, Jeffrey S; Rypma, Bart; Chapman, Sandra B

2018-02-01

Higher-order cognitive training has shown to enhance performance in older adults, but the neural mechanisms underlying performance enhancement have yet to be fully disambiguated. This randomized trial examined changes in processing speed and processing speed-related neural activity in older participants (57-71 years of age) who underwent cognitive training (CT, N = 12) compared with wait-listed (WLC, N = 15) or exercise-training active (AC, N = 14) controls. The cognitive training taught cognitive control functions of strategic attention, integrative reasoning, and innovation over 12 weeks. All 3 groups worked through a functional magnetic resonance imaging processing speed task during 3 sessions (baseline, mid-training, and post-training). Although all groups showed faster reaction times (RTs) across sessions, the CT group showed a significant increase, and the WLC and AC groups showed significant decreases across sessions in the association between RT and BOLD signal change within the left prefrontal cortex (PFC). Thus, cognitive training led to a change in processing speed-related neural activity where faster processing speed was associated with reduced PFC activation, fitting previously identified neural efficiency profiles. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Diminished frontostriatal activity during processing of monetary rewards and losses in pathological gambling.

PubMed

Balodis, Iris M; Kober, Hedy; Worhunsky, Patrick D; Stevens, Michael C; Pearlson, Godfrey D; Potenza, Marc N

2012-04-15

Mesocorticolimbic neurocircuitry and impulsivity have both been implicated in pathological gambling (PG) and in reward processing. However, the neural underpinnings of specific phases of reward and loss processing in PG and their relationships to impulsivity remain only partially understood. The present functional magnetic resonance imaging study examined brain activity associated with different phases of reward and loss processing in PG. Given an inverse relationship between ventral striatal recruitment during anticipation of monetary rewards and impulsivity in alcohol dependence, the current study explored whether a similar association might also be present in PG. Fourteen adults with PG and 14 control comparison participants performed the Monetary Incentive Delay Task to identify brain activation changes associated with reward/loss prospect, reward/loss anticipation, and reward/loss notification. Impulsivity was assessed separately using the Barratt Impulsiveness Scale. Relative to the control comparison group, the PG group exhibited significantly reduced activity in the ventromedial prefrontal cortex, insula, and ventral striatum during several phases, including the prospect and anticipation phases of both gains and losses. Activity in the ventral striatum correlated inversely with levels of impulsivity in PG participants, consistent with prior findings in alcohol dependence. Relatively decreased activity in corticostriatal neurocircuitry during multiple phases of reward processing suggests consistent alterations in neurocircuitry underlying incentive valuation and loss prediction. Together with findings in alcohol dependence, these results suggest that impulsive tendencies in addictions may be reflected in diminished ventral striatal activations to reward anticipation and may represent targets for treatment development in addictions. Copyright © 2012 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Neural Networks Involved in Adolescent Reward Processing: An Activation Likelihood Estimation Meta-Analysis of Functional Neuroimaging Studies

PubMed Central

Silverman, Merav H.; Jedd, Kelly; Luciana, Monica

2015-01-01

Behavioral responses to, and the neural processing of, rewards change dramatically during adolescence and may contribute to observed increases in risk-taking during this developmental period. Functional MRI (fMRI) studies suggest differences between adolescents and adults in neural activation during reward processing, but findings are contradictory, and effects have been found in non-predicted directions. The current study uses an activation likelihood estimation (ALE) approach for quantitative meta-analysis of functional neuroimaging studies to: 1) confirm the network of brain regions involved in adolescents’ reward processing, 2) identify regions involved in specific stages (anticipation, outcome) and valence (positive, negative) of reward processing, and 3) identify differences in activation likelihood between adolescent and adult reward-related brain activation. Results reveal a subcortical network of brain regions involved in adolescent reward processing similar to that found in adults with major hubs including the ventral and dorsal striatum, insula, and posterior cingulate cortex (PCC). Contrast analyses find that adolescents exhibit greater likelihood of activation in the insula while processing anticipation relative to outcome and greater likelihood of activation in the putamen and amygdala during outcome relative to anticipation. While processing positive compared to negative valence, adolescents show increased likelihood for activation in the posterior cingulate cortex (PCC) and ventral striatum. Contrasting adolescent reward processing with the existing ALE of adult reward processing (Liu et al., 2011) reveals increased likelihood for activation in limbic, frontolimbic, and striatal regions in adolescents compared with adults. Unlike adolescents, adults also activate executive control regions of the frontal and parietal lobes. These findings support hypothesized elevations in motivated activity during adolescence. PMID:26254587

ACTIVATED CARBON PROCESS FOR TREATMENT OF WASTEWATERS CONTAINING HEXAVALENT CHROMIUM

EPA Science Inventory

The removal of hexavalent chromium, Cr(VI), from dilute aqueous solution by an activated carbon process has been investigated. Two removal mechanisms were observed; hexavalent chromium species were removed by adsorption onto the interior carbon surface and/or through reduction to...

Recombination activity of grain boundaries in high-performance multicrystalline Si during solar cell processing

NASA Astrophysics Data System (ADS)

Adamczyk, Krzysztof; Søndenâ, Rune; Stokkan, Gaute; Looney, Erin; Jensen, Mallory; Lai, Barry; Rinio, Markus; Di Sabatino, Marisa

2018-02-01

In this work, we applied internal quantum efficiency mapping to study the recombination activity of grain boundaries in High Performance Multicrystalline Silicon under different processing conditions. Wafers were divided into groups and underwent different thermal processing, consisting of phosphorus diffusion gettering and surface passivation with hydrogen rich layers. After these thermal treatments, wafers were processed into heterojunction with intrinsic thin layer solar cells. Light Beam Induced Current and Electron Backscatter Diffraction were applied to analyse the influence of thermal treatment during standard solar cell processing on different types of grain boundaries. The results show that after cell processing, most random-angle grain boundaries in the material are well passivated, but small-angle grain boundaries are not well passivated. Special cases of coincidence site lattice grain boundaries with high recombination activity are also found. Based on micro-X-ray fluorescence measurements, a change in the contamination level is suggested as the reason behind their increased activity.

Design, synthesis, antibacterial activity and docking study of some new trimethoprim derivatives.

PubMed

Rashid, Umer; Ahmad, Waqas; Hassan, Syed Fahad; Qureshi, Naveeda Akhtar; Niaz, Basit; Muhammad, Bakhtiar; Imdad, Sameera; Sajid, Muhammad

2016-12-01

In present study, nineteen novel trimethoprim (TMP) derivatives were designed, synthesized and evaluated for their antibacterial potential. Hydroxy trimethoprim 2 (HTMP) was synthesized by following the demethylation of 4-methoxy group at trimethoxy benzyl ring of TMP. Structure-activity relationship (SAR) studies were explored on HTMP by incorporating various substituents leading to the identification of some new compounds with improved antibacterial activities. The results revealed that the introduction of benzyloxy (4a-e) and phenyl ethanone (5a-e) group at 4-position of dimethoxy benzyl ring leads to overall increase in the antibacterial activity. The most potent antibacterial compound discovered is benzyloxy derivative 4b with MIC value of 5.0μM against Staphylococcus aureus and 4.0μM against Escherichia coli strains higher than the standard TMP (22.7μM against S. aureus and 55.1μM against E. coli). Substitution at 4-NH 2 group was not tolerated and the resulting Schiff base derivatives 3a-h demonstrated very little or no antibacterial activity in the tested concentration domain. We further performed exploratory docking studies on dihydrofolate reductase (DHFR) to rationalize the in vitro biological data and to demonstrate the mechanism of antibacterial activity. For the ability to cross lipophilic outer membrane, logP was computed. It was found that the compounds possessing high hydrophobicity have high activity against E. coli. Copyright © 2016 Elsevier Ltd. All rights reserved.

Distributive Processing by the Iron(II)/α-Ketoglutarate-Dependent Catalytic Domains of the TET Enzymes Is Consistent with Epigenetic Roles for Oxidized 5-Methylcytosine Bases.

PubMed

Tamanaha, Esta; Guan, Shengxi; Marks, Katherine; Saleh, Lana

2016-08-03

The ten-eleven translocation (TET) proteins catalyze oxidation of 5-methylcytosine ((5m)C) residues in nucleic acids to 5-hydroxymethylcytosine ((5hm)C), 5-formylcytosine ((5f)C), and 5-carboxycytosine ((5ca)C). These nucleotide bases have been implicated as intermediates on the path to active demethylation, but recent reports have suggested that they might have specific regulatory roles in their own right. In this study, we present kinetic evidence showing that the catalytic domains (CDs) of TET2 and TET1 from mouse and their homologue from Naegleria gruberi, the full-length protein NgTET1, are distributive in both chemical and physical senses, as they carry out successive oxidations of a single (5m)C and multiple (5m)C residues along a polymethylated DNA substrate. We present data showing that the enzyme neither retains (5hm)C/(5f)C intermediates of preceding oxidations nor slides along a DNA substrate (without releasing it) to process an adjacent (5m)C residue. These findings contradict a recent report by Crawford et al. ( J. Am. Chem. Soc. 2016 , 138 , 730 ) claiming that oxidation of (5m)C by CD of mouse TET2 is chemically processive (iterative). We further elaborate that this distributive mechanism is maintained for TETs in two evolutionarily distant homologues and posit that this mode of function allows the introduction of (5m)C forms as epigenetic markers along the DNA.

Enhancement of activated sludge disintegration and dewaterability by Fenton process

NASA Astrophysics Data System (ADS)

Heng, G. C.; Isa, M. H.

2016-06-01

Municipal and industrial wastewater treatment plants produce large amounts of sludge. This excess sludge is an inevitable drawback inherent to the activated sludge process. In this study, the waste activated sludge was obtained from the campus wastewater treatment plant at Universiti Teknologi PETRONAS (UTP), Malaysia. Fenton pretreatment was optimized by using the response surface methodology (RSM) to study the effects of three operating conditions including the dosage of H2O2 (g H2O2/kg TS), the molar ratio of H2O2/Fe2+ and reaction time. The optimum operating variables to achieve MLVSS removal 65%, CST reduction 28%, sCOD 11000 mg/L and EPS 500 mg/L were: 1000 g H2O2/kg TS, H2O2/Fe2+ molar ratio 70 and reaction time 45 min. Fenton process was proved to be able to enhance the sludge disintegration and dewaterability.

Zebra Mussel Antifouling Activity of the Marine Natural Product Aaptamine and Analogs

PubMed Central

Diers, Jeffrey A.; Bowling, John J.; Duke, Stephen O.; Wahyuono, Subagus; Kelly, Michelle; Hamann, Mark T.

2016-01-01

Several aaptamine derivatives were selected as potential zebra mussel (Dreissena polymorpha) antifoulants because of the noteworthy absence of fouling observed on Aaptos sponges. Sponges of the genus Aaptos collected in Manado, Indonesia consistently produce aaptamine-type alkaloids. To date, aaptamine and its derivatives have not been carefully evaluated for their antifoulant properties. Structure–activity relationship studies were conducted using several aaptamine derivatives in a zebra mussel antifouling assay. From these data, three analogs have shown significant antifouling activity against zebra mussel attachment. Aaptamine, isoaaptamine, and the demethylated aaptamine compounds used in the zebra mussel assay produced EC50 values of 24.2, 11.6, and 18.6 μM, respectively. In addition, neither aaptamine nor isoaaptamine produced a phytotoxic response (as high as 300 μM) toward a nontarget organism, Lemna pausicostata, in a 7-day exposure. The use of these aaptamine derivatives from Aaptos sp. as potential environmentally benign antifouling alternatives to metal-based paints and preservatives is significant, not only as a possible control of fouling organisms, but also to highlight the ecological importance of these and similar biochemical defenses. PMID:16718618

Enhanced phenol removal in an innovative lignite activated coke-assisted biological process.

PubMed

Zhang, Chen; Li, Jianfeng; Cheng, Fangqin; Liu, Yu

2018-07-01

In this study, a lignite activated coke (LAC)-assisted activated sludge (AS) process was developed for enhancing biodegradation of phenol, while the effects of LAC on sludge properties and microbial community structure were investigated. It was found that more than 90% of phenol was removed within 1 h in the LAC/AS, which was 3 times higher than the conventional AS process. Moreover, the floc size and settleability were also significantly improved in the LAC/AS. These results suggested that LAC could serve as the nucleating agent to promote the formation of compact floc, which was beneficial for toxicity mitigation and system stability. The microbial community analysis by 16S high-throughput pyrosequencing technology further revealed a more abundant bacterial richness and diversity in the LAC/AS process loaded with phenol, while some phenol degraders, such as Propionibacteriaceae were enriched. Engineering implications further suggests the LAC-assisted AS process is technically sound and economically viable. Copyright © 2018 Elsevier Ltd. All rights reserved.

76 FR 30696 - Reimbursement for Costs of Remedial Action at Active Uranium and Thorium Processing Sites

Federal Register 2010, 2011, 2012, 2013, 2014

2011-05-26

... DEPARTMENT OF ENERGY Reimbursement for Costs of Remedial Action at Active Uranium and Thorium...) acceptance of claims in FY 2011 from eligible active uranium and thorium processing site licensees for... incurred by licensees at active uranium and thorium processing sites to remediate byproduct material...

Role of invertase activity in processing quality of potatoes: Effect of storage temperature and duration.

PubMed

Bandana; Sharma, Vineet; Singh, Brajesh; Raigond, Pinky; Kaushik, S K

2016-03-01

Invertase activity and processing attributes of three potato cultivars were studied to find the reason for deterioration of processing quality during their prolonged storage in commercial cold stores (4°C) as compared to elevated temperature storage (12 ± 0.5°C), with CIPC {Isopropyl-N-(3-Cholorophenyl) carbamate}. Lower storage temperature (4°C) tended to be more effective in increasing invertase activity of potato tubers than elevated temperature. Non-processing cultivar viz., Kufri Pukhraj resulted in accumulation of more invertase activity than relatively two processing cultivars. Kufri Chipsona-1 and Kufri Chipsona-3 at 12 ± 0.5°C possessed basal invertase activity ranging from 39.3 to 79.8 and 54.1 to 93.8 (pmoles hexose h⁻¹ g⁻¹ f.wt.) respectively, during two years. Total invertase activity at 4°C increased abruptly and remained high from 30 to 60 days of storage. The activity progressively reached 90.6 to 106.6 and 81.4 to 101.3 during both the years respectively, after 60 days of storage to that observed initially. Reducing sugar content increased from 23.3 to 105.7 and 389.0 to 1138.2 (mg 100g⁻¹ f.wt.) after 90 days of storage at 12 ± 0.5°C and 4°C, respectively. Studies concluded that basal and total invertase, were responsible for cold-induced sweetening and resulted in deterioration of processing quality of potatoes during storage at 4°C. Since this activity is low at 12 ± 0.5°C, the processing traits remained acceptable to industry and consumers.

GoActive: a protocol for the mixed methods process evaluation of a school-based physical activity promotion programme for 13-14year old adolescents.

PubMed

Jong, Stephanie T; Brown, Helen Elizabeth; Croxson, Caroline H D; Wilkinson, Paul; Corder, Kirsten L; van Sluijs, Esther M F

2018-05-21

Process evaluations are critical for interpreting and understanding outcome trial results. By understanding how interventions function across different settings, process evaluations have the capacity to inform future dissemination of interventions. The complexity of Get others Active (GoActive), a 12-week, school-based physical activity intervention implemented in eight schools, highlights the need to investigate how implementation is achieved across a variety of school settings. This paper describes the mixed methods GoActive process evaluation protocol that is embedded within the outcome evaluation. In this detailed process evaluation protocol, we describe the flexible and pragmatic methods that will be used for capturing the process evaluation data. A mixed methods design will be used for the process evaluation, including quantitative data collected in both the control and intervention arms of the GoActive trial, and qualitative data collected in the intervention arm. Data collection methods will include purposively sampled, semi-structured interviews and focus group interviews, direct observation, and participant questionnaires (completed by students, teachers, older adolescent mentors, and local authority-funded facilitators). Data will be analysed thematically within and across datasets. Overall synthesis of findings will address the process of GoActive implementation, and through which this process affects outcomes, with careful attention to the context of the school environment. This process evaluation will explore the experience of participating in GoActive from the perspectives of key groups, providing a greater understanding of the acceptability and process of implementation of the intervention across the eight intervention schools. This will allow for appraisal of the intervention's conceptual base, inform potential dissemination, and help optimise post-trial sustainability. The process evaluation will also assist in contextualising the trial

Reversing DNA Methylation: Mechanisms, Genomics, and Biological Functions

PubMed Central

Wu, Hao; Zhang, Yi

2014-01-01

Methylation of cytosines in the mammalian genome represents a key epigenetic modification and is dynamically regulated during development. Compelling evidence now suggests that dynamic regulation of DNA methylation is mainly achieved through a cyclic enzymatic cascade comprised of cytosine methylation, iterative oxidation of methyl group by TET dioxygenases, and restoration of unmodified cytosines by either replication-dependent dilution or DNA glycosylase-initiated base excision repair. In this review, we discuss the mechanism and function of DNA demethylation in mammalian genomes, focusing particularly on how developmental modulation of the cytosine-modifying pathway is coupled to active reversal of DNA methylation in diverse biological processes. PMID:24439369

The Novel Bacterial N-Demethylase PdmAB Is Responsible for the Initial Step of N,N-Dimethyl-Substituted Phenylurea Herbicide Degradation

PubMed Central

Gu, Tao; Zhou, Chaoyang; Sørensen, Sebastian R.; Zhang, Ji; He, Jian; Yu, Peiwen; Li, Shunpeng

2013-01-01

The environmental fate of phenylurea herbicides has received considerable attention in recent decades. The microbial metabolism of N,N-dimethyl-substituted phenylurea herbicides can generally be initiated by mono-N-demethylation. In this study, the molecular basis for this process was revealed. The pdmAB genes in Sphingobium sp. strain YBL2 were shown to be responsible for the initial mono-N-demethylation of commonly used N,N-dimethyl-substituted phenylurea herbicides. PdmAB is the oxygenase component of a bacterial Rieske non-heme iron oxygenase (RO) system. The genes pdmAB, encoding the α subunit PdmA and the β subunit PdmB, are organized in a transposable element flanked by two direct repeats of an insertion element resembling ISRh1. Furthermore, this transposable element is highly conserved among phenylurea herbicide-degrading sphingomonads originating from different areas of the world. However, there was no evidence of a gene for an electron carrier (a ferredoxin or a reductase) located in the immediate vicinity of pdmAB. Without its cognate electron transport components, expression of PdmAB in Escherichia coli, Pseudomonas putida, and other sphingomonads resulted in a functional enzyme. Moreover, coexpression of a putative [3Fe-4S]-type ferredoxin from Sphingomonas sp. strain RW1 greatly enhanced the catalytic activity of PdmAB in E. coli. These data suggested that PdmAB has a low specificity for electron transport components and that its optimal ferredoxin may be the [3Fe-4S] type. PdmA exhibited low homology to the α subunits of previously characterized ROs (less than 37% identity) and did not cluster with the RO group involved in O- or N-demethylation reactions, indicating that PdmAB is a distinct bacterial RO N-demethylase. PMID:24123738

Cortex and hippocampus DNA epigenetic response to a long-term arsenic exposure via drinking water.

PubMed

Du, Xiaoyan; Tian, Meiping; Wang, Xiaoxue; Zhang, Jie; Huang, Qingyu; Liu, Liangpo; Shen, Heqing

2018-03-01

The neurotoxicity of arsenic is a serious health problem, especially for children. DNA epigenetic change may be an important pathogenic mechanism, but the molecular pathway remains obscure. In this study, the weaned male Sprague-Dawly (SD) rats were treated with arsenic trioxide via drinking water for 6 months, simulating real developmental exposure situation of children. Arsenic exposure impaired the cognitive abilities, and altered the expression of neuronal activity-regulated genes. Total arsenic concentrations of cortex and hippocampus tissues were significantly increased in a dose-dependent manner. The reduction in 5-methylcytosine (5 mC) and 5-hydroxymethylcytosine (5hmC) levels as well as the down-regulation of DNA methyltransferases (DNMTs) and ten-eleven translocations (TETs) expression suggested that DNA methylation/demethylation processes were significantly suppressed in brain tissues. S-adenosylmethionine (SAM) level wasn't changed, but the expression of the important indicators of oxidative/anti-oxidative balance and tricarboxylic acid (TCA) cycle was significantly deregulated. Overall, arsenic can disrupt oxidative/anti-oxidative balance, further inhibit TETs expression through TCA cycle and alpha-ketoglutarate (α-KG) pathway, and consequently cause DNA methylation/demethylation disruption. The present study implies oxidative stress but not SAM depletion may lead to DNA epigenetic alteration and arsenic neurotoxicity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Deep processing activates the medial temporal lobe in young but not in old adults.

PubMed

Daselaar, Sander M; Veltman, Dick J; Rombouts, Serge A R B; Raaijmakers, Jeroen G W; Jonker, Cees

2003-11-01

Age-related impairments in episodic memory have been related to a deficiency in semantic processing, based on the finding that elderly adults typically benefit less than young adults from deep, semantic as opposed to shallow, nonsemantic processing of study items. In the present study, we tested the hypothesis that elderly adults are not able to perform certain cognitive operations under deep processing conditions. We further hypothesised that this inability does not involve regions commonly associated with lexical/semantic retrieval processes, but rather involves a dysfunction of the medial temporal lobe (MTL) memory system. To this end, we used functional MRI on rather extensive groups of young and elderly adults to compare brain activity patterns obtained during a deep (living/nonliving) and a shallow (uppercase/lowercase) classification task. Common activity in relation to semantic classification was observed in regions that have been previously related to semantic retrieval, including mainly left-lateralised activity in the inferior prefrontal, middle temporal, and middle frontal/anterior cingulate gyrus. Although the young adults showed more activity in some of these areas, the finding of mainly overlapping activation patterns during semantic classification supports the idea that lexical/semantic retrieval processes are still intact in elderly adults. This received further support by the finding that both groups showed similar behavioural performances as well on the deep and shallow classification tasks. Importantly, though, the young revealed significantly more activity than the elderly adults in the left anterior hippocampus during deep relative to shallow classification. This finding is in line with the idea that age-related impairments in episodic encoding are, at least partly, due to an under-recruitment of the medial temporal lobe memory system.

Krebs cycle intermediates regulate DNA and histone methylation: epigenetic impact on the aging process.

PubMed

Salminen, Antero; Kauppinen, Anu; Hiltunen, Mikko; Kaarniranta, Kai

2014-07-01

Many aging theories have proposed that mitochondria and energy metabolism have a major role in the aging process. There are recent studies indicating that Krebs cycle intermediates can shape the epigenetic landscape of chromatin by regulating DNA and histone methylation. A growing evidence indicates that epigenetics plays an important role in the regulation of healthspan but also is involved in the aging process. 2-Oxoglutarate (α-ketoglutarate) is a key metabolite in the Krebs cycle but it is also an obligatory substrate for 2-oxoglutarate-dependent dioxygenases (2-OGDO). The 2-OGDO enzyme family includes the major enzymes of DNA and histone demethylation, i.e. Ten-Eleven Translocation (TETs) and Jumonji C domain containing (JmjC) demethylases. In addition, 2-OGDO members can regulate collagen synthesis and hypoxic responses in a non-epigenetical manner. Interestingly, succinate and fumarate, also Krebs cycle intermediates, are potent inhibitors of 2-OGDO enzymes, i.e. the balance of Krebs cycle reactions can affect the level of DNA and histone methylation and thus control gene expression. We will review the epigenetic mechanisms through which Krebs cycle intermediates control the DNA and histone methylation. We propose that age-related disturbances in the Krebs cycle function induce stochastic epigenetic changes in chromatin structures which in turn promote the aging process. Copyright © 2014 Elsevier B.V. All rights reserved.

Urinary metabolites of isorhynchophylline in rats and their neuroprotective activities in the HT22 cell assay

PubMed Central

Chen, Fangfang; Qi, Wen; Sun, Jiahong; Simpkins, James W.; Yuan, Dan

2015-01-01

Isorhynchophylline is one of the major alkaloids from the Uncaria hook possessing the effects of lowered blood pressure, vasodilatation and protection against ischemia-induced neuronal damage. However, the metabolic pathway of isorhynchophylline has not been fully reported yet. In this paper, the metabolism of isorhynchophylline was investigated in rats. Five metabolites were isolated by using solvent extraction and repeated chromatographic methods, and identified by spectroscopic methods including UV, MS, NMR and CD experiments. Three new compounds were identified as 5-oxoisorhynchophyllic acid-22-O-β-D-glucuronide (M1), 17-O-demethyl-16,17-dihydro isorhynchophylline (M2) and 5-oxoisorhynchophyllic acid (M4) together with two known compounds isorhynchophylline (M0) and rhynchophylline (M3). Possible metabolic pathways of isorhynchophylline are proposed. Furthermore, the activity assay for all the metabolites showed that isorhynchophylline (M0) exhibited potent neuroprotective effects against glutamate-induced HT22 cell death. However, little or weak neuroprotective activities were observed for M1–M4. Our present study is important to further understand its metabolic fate and disposition in humans. PMID:24910000

Relative Contributions of Specific Activity Histories and Spontaneous Processes to Size Remodeling of Glutamatergic Synapses

PubMed Central

Dvorkin, Roman; Ziv, Noam E.

2016-01-01

The idea that synaptic properties are defined by specific pre- and postsynaptic activity histories is one of the oldest and most influential tenets of contemporary neuroscience. Recent studies also indicate, however, that synaptic properties often change spontaneously, even in the absence of specific activity patterns or any activity whatsoever. What, then, are the relative contributions of activity history-dependent and activity history-independent processes to changes synapses undergo? To compare the relative contributions of these processes, we imaged, in spontaneously active networks of cortical neurons, glutamatergic synapses formed between the same axons and neurons or dendrites under the assumption that their similar activity histories should result in similar size changes over timescales of days. The size covariance of such commonly innervated (CI) synapses was then compared to that of synapses formed by different axons (non-CI synapses) that differed in their activity histories. We found that the size covariance of CI synapses was greater than that of non-CI synapses; yet overall size covariance of CI synapses was rather modest. Moreover, momentary and time-averaged sizes of CI synapses correlated rather poorly, in perfect agreement with published electron microscopy-based measurements of mouse cortex synapses. A conservative estimate suggested that ~40% of the observed size remodeling was attributable to specific activity histories, whereas ~10% and ~50% were attributable to cell-wide and spontaneous, synapse-autonomous processes, respectively. These findings demonstrate that histories of naturally occurring activity patterns can direct glutamatergic synapse remodeling but also suggest that the contributions of spontaneous, possibly stochastic, processes are at least as great. PMID:27776122

Write! Write! Write! Ready-to-Use Writing Process Activities for Grades 4-8.

ERIC Educational Resources Information Center

Behrman, Carol H.

This handbook contains over 265 reproducible writing process activities that help make writing fun for students in grades 4-8. The handbook provides stimulating activities to give students the directed practice they need to learn to write clearly and competently. Designed for minimal teacher input, activities are complete with directions geared to…

Neural Activations of Guided Imagery and Music in Negative Emotional Processing: A Functional MRI Study.

PubMed

Lee, Sang Eun; Han, Yeji; Park, HyunWook

2016-01-01

The Bonny Method of Guided Imagery and Music uses music and imagery to access and explore personal emotions associated with episodic memories. Understanding the neural mechanism of guided imagery and music (GIM) as combined stimuli for emotional processing informs clinical application. We performed functional magnetic resonance imaging (fMRI) to demonstrate neural mechanisms of GIM for negative emotional processing when personal episodic memory is recalled and re-experienced through GIM processes. Twenty-four healthy volunteers participated in the study, which used classical music and verbal instruction stimuli to evoke negative emotions. To analyze the neural mechanism, activated regions associated with negative emotional and episodic memory processing were extracted by conducting volume analyses for the contrast between GIM and guided imagery (GI) or music (M). The GIM stimuli showed increased activation over the M-only stimuli in five neural regions associated with negative emotional and episodic memory processing, including the left amygdala, left anterior cingulate gyrus, left insula, bilateral culmen, and left angular gyrus (AG). Compared with GI alone, GIM showed increased activation in three regions associated with episodic memory processing in the emotional context, including the right posterior cingulate gyrus, bilateral parahippocampal gyrus, and AG. No neural regions related to negative emotional and episodic memory processing showed more activation for M and GI than for GIM. As a combined multimodal stimulus, GIM may increase neural activations related to negative emotions and episodic memory processing. Findings suggest a neural basis for GIM with personal episodic memories affecting cortical and subcortical structures and functions. © the American Music Therapy Association 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Patterns of frontoparietal activation as a marker for unsuccessful visuospatial processing in healthy aging.

PubMed

Drag, Lauren L; Light, Sharee N; Langenecker, Scott A; Hazlett, Kathleen E; Wilde, Elisabeth A; Welsh, Robert; Steinberg, Brett A; Bieliauskas, Linas A

2016-09-01

Visuospatial abilities are sensitive to age-related decline, although the neural basis for this decline (and its everyday behavioral correlates) is as yet poorly understood. fMRI was employed to examine age-related differences in patterns of functional activation that underlie changes in visuospatial processing. All participants completed a brief neuropsychological battery and also a figure ground task (FGT) assessing visuospatial processing while fMRI was recorded. Participants included 16 healthy older adults (OA; aged 69-82 years) and 16 healthy younger adults (YA; aged 20-35 years). We examined age-related differences in behavioral performance on the FGT in relation to patterns of fMRI activation. OA demonstrated reduced performance on the FGT task and showed increased activation of supramarginal parietal cortex as well as increased activation of frontal and temporal regions compared to their younger counterparts. Performance on the FGT related to increased supramarginal gyrus activity and increased medial prefrontal activity in OAs, but not YAs. Our results are consistent with an anterior-posterior compensation model. Successful FGT performance requires the perception and integration of multiple stimuli and thus it is plausible that healthy aging may be accompanied by changes in visuospatial processing that mimic a subtle form of dorsal simultanagnosia. Overall, decreased visuospatial processing in OA relates to an altered frontoparietal neurobiological signature that may contribute to the general phenomenon of increasingly fragmented execution of behavior associated with normal aging.

Complete activation of autophagic process attenuates liver injury and improves survival in septic mice.

PubMed

Lin, Chih-Wen; Lo, Steven; Perng, Daw-Shyong; Wu, David Bin-Chia; Lee, Po-Huang; Chang, Ya-Fang; Kuo, Po-Lin; Yu, Ming-Lung; Yuan, Shyng-Shiou F; Hsieh, Ya-Ching

2014-03-01

The accumulation of autophagosomes in the terminal step of the autophagic process has recently emerged as a potentially maladaptive process in the septic heart and lung. However, the role of autophagy in the septic liver has not been ascertained. This study was investigated by first examining the entire sequence of the autophagic process in the liver of septic mice. Second, a novel pharmacotherapeutic approach was utilized to treat sepsis with autophagy enhancer/inhibitor. Sepsis was induced by cecal ligation and puncture (CLP). C57BL/6 mice received autophagy enhancer carbamazepine (CBZ), autophagy inhibitor 3-methyladenine (inhibition of autophagosomal formation), or chloroquine (impairment of autophagosomal clearance). We found that the whole autophagic process was activated at 4 h after CLP; however, it did not proceed to completion during the 4- to 24-h time period, as indicated by accumulated autophagosomes and decreased autophagic flux. Carbamazepine, which induced complete activation of the autophagic process, improved CLP survival. This protective effect was also associated with decreased cell death, inflammatory responses, and hepatic injury. However, disruption of autophagosomal clearance with chloroquine abolished the above protective effects in CBZ-treated CLP mice. 3-Methyladenine, which resulted in inhibition of the autophagosomal formation, did not show any above beneficial effects in CLP mice. Impaired autophagosome-lysome fusion resulting in incomplete activation of autophagy may contribute to sepsis-induced liver injury. Treatment with CBZ may serve a protective role in the septic liver, possibly through the effect of complete activation of autophagic process.

Differentiating maturational and training influences on fMRI activation during music processing.

PubMed

Ellis, Robert J; Norton, Andrea C; Overy, Katie; Winner, Ellen; Alsop, David C; Schlaug, Gottfried

2012-04-15

Two major influences on how the brain processes music are maturational development and active musical training. Previous functional neuroimaging studies investigating music processing have typically focused on either categorical differences between "musicians versus nonmusicians" or "children versus adults." In the present study, we explored a cross-sectional data set (n=84) using multiple linear regression to isolate the performance-independent effects of age (5 to 33 years) and cumulative duration of musical training (0 to 21,000 practice hours) on fMRI activation similarities and differences between melodic discrimination (MD) and rhythmic discrimination (RD). Age-related effects common to MD and RD were present in three left hemisphere regions: temporofrontal junction, ventral premotor cortex, and the inferior part of the intraparietal sulcus, regions involved in active attending to auditory rhythms, sensorimotor integration, and working memory transformations of pitch and rhythmic patterns. By contrast, training-related effects common to MD and RD were localized to the posterior portion of the left superior temporal gyrus/planum temporale, an area implicated in spectrotemporal pattern matching and auditory-motor coordinate transformations. A single cluster in right superior temporal gyrus showed significantly greater activation during MD than RD. This is the first fMRI which has distinguished maturational from training effects during music processing. Copyright © 2012 Elsevier Inc. All rights reserved.

Emergent cell and tissue dynamics from subcellular modeling of active biomechanical processes

NASA Astrophysics Data System (ADS)

Sandersius, S. A.; Weijer, C. J.; Newman, T. J.

2011-08-01

Cells and the tissues they form are not passive material bodies. Cells change their behavior in response to external biochemical and biomechanical cues. Behavioral changes, such as morphological deformation, proliferation and migration, are striking in many multicellular processes such as morphogenesis, wound healing and cancer progression. Cell-based modeling of these phenomena requires algorithms that can capture active cell behavior and their emergent tissue-level phenotypes. In this paper, we report on extensions of the subcellular element model to model active biomechanical subcellular processes. These processes lead to emergent cell and tissue level phenotypes at larger scales, including (i) adaptive shape deformations in cells responding to slow stretching, (ii) viscous flow of embryonic tissues, and (iii) streaming patterns of chemotactic cells in epithelial-like sheets. In each case, we connect our simulation results to recent experiments.

Emotion modulates activity in the 'what' but not 'where' auditory processing pathway.

PubMed

Kryklywy, James H; Macpherson, Ewan A; Greening, Steven G; Mitchell, Derek G V

2013-11-15

Auditory cortices can be separated into dissociable processing pathways similar to those observed in the visual domain. Emotional stimuli elicit enhanced neural activation within sensory cortices when compared to neutral stimuli. This effect is particularly notable in the ventral visual stream. Little is known, however, about how emotion interacts with dorsal processing streams, and essentially nothing is known about the impact of emotion on auditory stimulus localization. In the current study, we used fMRI in concert with individualized auditory virtual environments to investigate the effect of emotion during an auditory stimulus localization task. Surprisingly, participants were significantly slower to localize emotional relative to neutral sounds. A separate localizer scan was performed to isolate neural regions sensitive to stimulus location independent of emotion. When applied to the main experimental task, a significant main effect of location, but not emotion, was found in this ROI. A whole-brain analysis of the data revealed that posterior-medial regions of auditory cortex were modulated by sound location; however, additional anterior-lateral areas of auditory cortex demonstrated enhanced neural activity to emotional compared to neutral stimuli. The latter region resembled areas described in dual pathway models of auditory processing as the 'what' processing stream, prompting a follow-up task to generate an identity-sensitive ROI (the 'what' pathway) independent of location and emotion. Within this region, significant main effects of location and emotion were identified, as well as a significant interaction. These results suggest that emotion modulates activity in the 'what,' but not the 'where,' auditory processing pathway. Copyright © 2013 Elsevier Inc. All rights reserved.

Instruction-specific brain activations during episodic encoding. a generalized level of processing effect.

PubMed

Petersson, Karl Magnus; Sandblom, Johan; Elfgren, Christina; Ingvar, Martin

2003-11-01

In a within-subject design we investigated the levels-of-processing (LOP) effect using visual material in a behavioral and a corresponding PET study. In the behavioral study we characterize a generalized LOP effect, using pleasantness and graphical quality judgments in the encoding situation, with two types of visual material, figurative and nonfigurative line drawings. In the PET study we investigate the related pattern of brain activations along these two dimensions. The behavioral results indicate that instruction and material contribute independently to the level of recognition performance. Therefore the LOP effect appears to stem both from the relative relevance of the stimuli (encoding opportunity) and an altered processing of stimuli brought about by the explicit instruction (encoding mode). In the PET study, encoding of visual material under the pleasantness (deep) instruction yielded left lateralized frontoparietal and anterior temporal activations while surface-based perceptually oriented processing (shallow instruction) yielded right lateralized frontoparietal, posterior temporal, and occipitotemporal activations. The result that deep encoding was related to the left prefrontal cortex while shallow encoding was related to the right prefrontal cortex, holding the material constant, is not consistent with the HERA model. In addition, we suggest that the anterior medial superior frontal region is related to aspects of self-referential semantic processing and that the inferior parts of the anterior cingulate as well as the medial orbitofrontal cortex is related to affective processing, in this case pleasantness evaluation of the stimuli regardless of explicit semantic content. Finally, the left medial temporal lobe appears more actively engaged by elaborate meaning-based processing and the complex response pattern observed in different subregions of the MTL lends support to the suggestion that this region is functionally segregated.

Activity of LPO Processes in Women with Polycystic Ovarian Syndrome and Infertility.

PubMed

Kolesnikova, L I; Kolesnikov, S I; Darenskaya, M A; Grebenkina, L A; Nikitina, O A; Lazareva, L M; Suturina, L V; Danusevich, I N; Druzhinina, E B; Semendyaev, A A

2017-01-01

Specific features of LPO processes and antioxidant defense were studied in patients with polycystic ovarian syndrome (PCOS) and infertility. Changes in LPO processes in patients with PCOS were compensatory, which manifested in increased α-tocopherol and retinol concentrations and moderate decrease in superoxide dismutase activity. Intensification of prooxidant processes was found in the group of patients with infertility without PCOS. The observed changes necessitate differentiated approach to the treatment of these patients.

Encoding-related brain activity during deep processing of verbal materials: a PET study.

PubMed

Fujii, Toshikatsu; Okuda, Jiro; Tsukiura, Takashi; Ohtake, Hiroya; Suzuki, Maki; Kawashima, Ryuta; Itoh, Masatoshi; Fukuda, Hiroshi; Yamadori, Atsushi

2002-12-01

The recent advent of neuroimaging techniques provides an opportunity to examine brain regions related to a specific memory process such as episodic memory encoding. There is, however, a possibility that areas active during an assumed episodic memory encoding task, compared with a control task, involve not only areas directly relevant to episodic memory encoding processes but also areas associated with other cognitive processes for on-line information. We used positron emission tomography (PET) to differentiate these two kinds of regions. Normal volunteers were engaged in deep (semantic) or shallow (phonological) processing of new or repeated words during PET. Results showed that deep processing, compared with shallow processing, resulted in significantly better recognition performance and that this effect was associated with activation of various brain areas. Further analyses revealed that there were regions directly relevant to episodic memory encoding in the anterior part of the parahippocampal gyrus, inferior frontal gyrus, supramarginal gyrus, anterior cingulate gyrus, and medial frontal lobe in the left hemisphere. Our results demonstrated that several regions, including the medial temporal lobe, play a role in episodic memory encoding.

Efficient mineralization of the antibiotic trimethoprim by solar assisted photoelectro-Fenton process driven by a photovoltaic cell.

PubMed

Zhang, Yanyu; Wang, Aimin; Tian, Xiujun; Wen, Zhenjun; Lv, Hanjiao; Li, Desheng; Li, Jiuyi

2016-11-15

In this study, a novel self-sustainable solar assisted photoelectro-Fenton (SPEF) system driven by a solar photovoltaic cell was developed for the efficient mineralization of antibiotic trimethoprim (TMP) in water. A comparative degradation of 200mgL(-1) TMP by RuO2/Ti anodic oxidation (AO), anodic oxidation with H2O2 electrogeneration (AO-H2O2), electro-Fenton (EF) and SPEF was investigated. SPEF was proved to exhibit the highest oxidation power, i.e., more than 80% TOC was removed after 360min SPEF treatment of 200mgL(-1) of TMP under optimal conditions at pH 3.0, 1.0mM Fe(2+) and 18mAcm(-2). Influences of current density, pH, initial Fe(2+) and initial TMP concentration on SPEF process were also studied. Ten aromatic intermediates generated from hydroxylation, carbonylation and demethylation reactions were identified using UPLC-QTOF-MS/MS system during the SPEF treatment, together with three carboxylic acids (oxamic, oxalic and formic acids) and two inorganic ions (NH4(+) and NO3(-)) measured. Therefore, a reasonable pathway of TMP degradation in SPEF process was proposed. Copyright © 2016 Elsevier B.V. All rights reserved.

CYP3A5 Contributes significantly to CYP3A-mediated drug oxidations in liver microsomes from Japanese subjects.

PubMed

Yamaori, Satoshi; Yamazaki, Hiroshi; Iwano, Shunsuke; Kiyotani, Kazuma; Matsumura, Keiko; Honda, Goro; Nakagawa, Kazuko; Ishizaki, Takashi; Kamataki, Tetsuya

2004-04-01

The purpose of this study was to evaluate a contribution of polymorphic cytochrome P450 (CYP) 3A5 to the oxidation of diltiazem, midazolam and testosterone by liver microsomes from Japanese subjects. Twenty-seven liver samples were classified into three groups according to the CYP3A5 genotypes; CYP3A5(*)1/(*)1 (n=3), (*)1/(*)3 (n=12) and (*)3/(*)3 (n=12). The results of genotyping and immunochemical quantitation of CYP3A5 protein showed a good accordance between the CYP3A5 genotype and CYP3A5 content but not CYP3A4 content in liver microsomes. The expression levels of hepatic CYP3A5 protein ranged from 20 to 60% of the sum of CYP3A4 and CYP3A5 contents in subjects with at least one wild type allele ((*)1). The CYP3A5 contents correlated well with liver microsomal activities of diltiazem N-demethylation, midazolam 1'- and 4-hydroxylations and testosterone 6beta-hydroxylation among subjects carrying at least one (*)1 allele. In addition, the correlation coefficients of CYP3A5 contents with the rates of diltiazem N-demethylation, midazolam 1'-hydroxylation and testosterone 6beta- hydroxylation were higher than those of CYP3A4, although the value of CYP3A5 with the midazolam 4-hydroxylation rate was similar to that of CYP3A4. Kinetic analyses revealed a biphasic diltiazem N-demethylation in liver microsomes from subjects carrying the (*)1 allele. The apparent V(max)/K(m) values for recombinant CYP3A5 indicated the greater contributions to diltiazem N-demethylation and midazolam 1'-hydroxylation as compared with CYP3A4. These results suggest that polymorphic CYP3A5 contributes markedly to the drug oxidations, particularly diltiazem N-demethylation, midazolam 1'- hydroxylation and testosterone 6beta-hydroxylation by liver microsomes from Japanese subjects.

Solid-state 13C NMR studies of a large fossil gymnosperm from the Yallourn Open Cut, Latrobe Valley, Australia

USGS Publications Warehouse

Bates, A.L.; Hatcher, P.G.

1989-01-01

A series of samples taken from the cross section of a 3-m-diameter fossilized gymnospermous log (Araucariaceae) in the Yallourn Seam of the Australian brown coals was examined by solid state 13C nuclear magnetic resonance to delineate chemical changes related to the combined processes of peatification and coalification. The results show that cellulosic materials were degraded and lost on the periphery of the log, however, the degree of such degradation in the central core is substantially less. The lignin is uniformly altered by coalification reactions to a macromolecular substance displaying decreased aryl ether linkages but significantly greater amounts of carbon linkages compared to modern lignin. Changes in the methoxyl carbon contents of lignin in cross section reveal demethylation reactions, but these do not appear to be related to degree of carbon linking. Both the degredation of cellulosic materials and demethylation of lignin appear to be early diagenetic processes occurring during peatification independently of the coalification reactions. ?? 1989.

The Roles of Structured Input Activities in Processing Instruction and the Kinds of Knowledge They Promote

ERIC Educational Resources Information Center

Marsden, Emma; Chen, Hsin-Ying

2011-01-01

This study aimed to isolate the effects of the two input activities in Processing Instruction: referential activities, which force learners to focus on a form and its meaning, and affective activities, which contain exemplars of the target form and require learners to process sentence meaning. One hundred and twenty 12-year-old Taiwanese learners…

Correlated activity supports efficient cortical processing

PubMed Central

Hung, Chou P.; Cui, Ding; Chen, Yueh-peng; Lin, Chia-pei; Levine, Matthew R.

2015-01-01

Visual recognition is a computational challenge that is thought to occur via efficient coding. An important concept is sparseness, a measure of coding efficiency. The prevailing view is that sparseness supports efficiency by minimizing redundancy and correlations in spiking populations. Yet, we recently reported that “choristers”, neurons that behave more similarly (have correlated stimulus preferences and spontaneous coincident spiking), carry more generalizable object information than uncorrelated neurons (“soloists”) in macaque inferior temporal (IT) cortex. The rarity of choristers (as low as 6% of IT neurons) indicates that they were likely missed in previous studies. Here, we report that correlation strength is distinct from sparseness (choristers are not simply broadly tuned neurons), that choristers are located in non-granular output layers, and that correlated activity predicts human visual search efficiency. These counterintuitive results suggest that a redundant correlational structure supports efficient processing and behavior. PMID:25610392

Effects of Different Heat Processing on Fucoxanthin, Antioxidant Activity and Colour of Indonesian Brown Seaweeds

NASA Astrophysics Data System (ADS)

Susanto, Eko; Suhaeli Fahmi, A.; Winarni Agustini, Tri; Rosyadi, Septian; Dita Wardani, Ayunda

2017-02-01

Fucoxanthin (Fx) is major carotenoids in brown algae. It showed many health beneficial effects for oxidative stress. Fucoxanthin is lower stability which may cause problem in the application for functional food. The objective of this study was to evaluate the effects of various heat processing on Fx, antioxidant activity (IC50), total phenolic content, and colour stability of Sargassum ilicifolium. The various heat processing methods showed were not significantly affected to fucoxanthin and antioxidant activities however all treatments lower affected to brown seaweeds colour. Moreover, this study showed a useful proved in the design of brown seaweeds processing which minimize Fx, antioxidant activity and colour changes.

Spätzle-Processing Enzyme-independent Activation of the Toll Pathway in Drosophila Innate Immunity.

PubMed

Yamamoto-Hino, Miki; Goto, Satoshi

2016-05-07

The Toll pathway regulates innate immunity in insects and vertebrates. The Drosophila Toll receptor is activated by a processed form of a ligand, Spätzle. Spätzle-processing enzyme (SPE) is the only enzyme identified to date that functions in converting Spätzle to an active form during the immune response. In the present study, Toll activation induced by immune challenge was almost suppressed in spätzle mutant larvae and adults, whereas it was present in SPE mutant larvae challenged with Micrococcus luteus and adults challenged with Bacillus subtilis. Our data suggest that an unidentified protease besides SPE processes Spätzle under conditions of microbial challenge.

Sex differences in functional activation patterns revealed by increased emotion processing demands.

PubMed

Hall, Geoffrey B C; Witelson, Sandra F; Szechtman, Henry; Nahmias, Claude

2004-02-09

Two [O(15)] PET studies assessed sex differences regional brain activation in the recognition of emotional stimuli. Study I revealed that the recognition of emotion in visual faces resulted in bilateral frontal activation in women, and unilateral right-sided activation in men. In study II, the complexity of the emotional face task was increased through tje addition of associated auditory emotional stimuli. Men again showed unilateral frontal activation, in this case to the left; whereas women did not show bilateral frontal activation, but showed greater limbic activity. These results suggest that when processing broader cross-modal emotional stimuli, men engage more in associative cognitive strategies while women draw more on primary emotional references.

In vitro Anti-Thrombotic Activity of Extracts from Blacklip Abalone (Haliotis rubra) Processing Waste.

PubMed

Suleria, Hafiz Ansar Rasul; Hines, Barney M; Addepalli, Rama; Chen, Wei; Masci, Paul; Gobe, Glenda; Osborne, Simone A

2016-12-31

Waste generated from the processing of marine organisms for food represents an underutilized resource that has the potential to provide bioactive molecules with pharmaceutical applications. Some of these molecules have known anti-thrombotic and anti-coagulant activities and are being investigated as alternatives to common anti-thrombotic drugs, like heparin and warfarin that have serious side effects. In the current study, extracts prepared from blacklip abalone ( Haliotis rubra ) processing waste, using food grade enzymes papain and bromelain, were found to contain sulphated polysaccharide with anti-thrombotic activity. Extracts were found to be enriched with sulphated polysaccharides and assessed for anti-thrombotic activity in vitro through heparin cofactor-II (HCII)-mediated inhibition of thrombin. More than 60% thrombin inhibition was observed in response to 100 μg/mL sulphated polysaccharides. Anti-thrombotic potential was further assessed as anti-coagulant activity in plasma and blood, using prothrombin time (PT), activated partial thromboplastin time (aPTT), and thromboelastography (TEG). All abalone extracts had significant activity compared with saline control. Anion exchange chromatography was used to separate extracts into fractions with enhanced anti-thrombotic activity, improving HCII-mediated thrombin inhibition, PT and aPTT almost 2-fold. Overall this study identifies an alternative source of anti-thrombotic molecules that can be easily processed offering alternatives to current anti-thrombotic agents like heparin.

In vitro Anti-Thrombotic Activity of Extracts from Blacklip Abalone (Haliotis rubra) Processing Waste

PubMed Central

Suleria, Hafiz Ansar Rasul; Hines, Barney M.; Addepalli, Rama; Chen, Wei; Masci, Paul; Gobe, Glenda; Osborne, Simone A.

2016-01-01

Waste generated from the processing of marine organisms for food represents an underutilized resource that has the potential to provide bioactive molecules with pharmaceutical applications. Some of these molecules have known anti-thrombotic and anti-coagulant activities and are being investigated as alternatives to common anti-thrombotic drugs, like heparin and warfarin that have serious side effects. In the current study, extracts prepared from blacklip abalone (Haliotis rubra) processing waste, using food grade enzymes papain and bromelain, were found to contain sulphated polysaccharide with anti-thrombotic activity. Extracts were found to be enriched with sulphated polysaccharides and assessed for anti-thrombotic activity in vitro through heparin cofactor-II (HCII)-mediated inhibition of thrombin. More than 60% thrombin inhibition was observed in response to 100 μg/mL sulphated polysaccharides. Anti-thrombotic potential was further assessed as anti-coagulant activity in plasma and blood, using prothrombin time (PT), activated partial thromboplastin time (aPTT), and thromboelastography (TEG). All abalone extracts had significant activity compared with saline control. Anion exchange chromatography was used to separate extracts into fractions with enhanced anti-thrombotic activity, improving HCII-mediated thrombin inhibition, PT and aPTT almost 2-fold. Overall this study identifies an alternative source of anti-thrombotic molecules that can be easily processed offering alternatives to current anti-thrombotic agents like heparin. PMID:28042854

Independent active and thermodynamic processes govern the nucleolus assembly in vivo

PubMed Central

Falahati, Hanieh; Wieschaus, Eric

2017-01-01

Membraneless organelles play a central role in the organization of protoplasm by concentrating macromolecules, which allows efficient cellular processes. Recent studies have shown that, in vitro, certain components in such organelles can assemble through phase separation. Inside the cell, however, such organelles are multicomponent, with numerous intermolecular interactions that can potentially affect the demixing properties of individual components. In addition, the organelles themselves are inherently active, and it is not clear how the active, energy-consuming processes that occur constantly within such organelles affect the phase separation behavior of the constituent macromolecules. Here, we examine the phase separation model for the formation of membraneless organelles in vivo by assessing the two features that collectively distinguish it from active assembly, namely temperature dependence and reversibility. We use a microfluidic device that allows accurate and rapid manipulation of temperature and examine the quantitative dynamics by which six different nucleolar proteins assemble into the nucleoli of Drosophila melanogaster embryos. Our results indicate that, although phase separation is the main mode of recruitment for four of the studied proteins, the assembly of the other two is irreversible and enhanced at higher temperatures, behaviors indicative of active recruitment to the nucleolus. These two subsets of components differ in their requirements for ribosomal DNA; the two actively assembling components fail to assemble in the absence of ribosomal DNA, whereas the thermodynamically driven components assemble but lose temporal and spatial precision. PMID:28115706

Self-efficacy mediates the relationship between behavioral processes of change and physical activity in older breast cancer survivors.

PubMed

Loprinzi, Paul D; Cardinal, Bradley J

2013-01-01

The degree to which breast cancer survivors use behavioral processes of change has not been investigated. Additionally, the relationship between behavioral processes and other theory-based mediators of adult physical activity behavior has not been extensively studied among breast cancer survivors. The objectives of this study were to: (1) determine the extent to which breast cancer survivors use behavioral processes associated with physical activity behavior change, and (2) examine the inter-relationships between behavioral processes, self-efficacy, and physical activity behavior among breast cancer survivors. Sixty-nine breast cancer survivors completed surveys examining behavioral processes and exercise-specific self-efficacy. Six months later they completed a self-report physical activity questionnaire. Findings showed the majority of breast cancer survivors did not use approximately half of the behavioral processes on a regular basis, and self-efficacy completely mediated the relationship between behavioral processes and physical activity. Health care professionals may help enhance self-efficacy and ultimately increase physical activity behavior in breast cancer survivors by teaching behavior skills such as enlisting social support.

Diminished fronto-striatal activity during processing of monetary rewards and losses in pathological gambling

PubMed Central

Balodis, Iris M.; Kober, Hedy; Worhunsky, Patrick D.; Stevens, Michael C.; Pearlson, Godfrey D.; Potenza, Marc N.

2012-01-01

Background Mesocorticolimbic neurocircuitry and impulsivity have both been implicated in pathological gambling (PG) and in reward processing. However, the neural underpinnings of specific phases of reward and loss processing in PG and their relationships to impulsivity remain only partially understood. The present functional magnetic resonance imaging study examined brain activity associated with different phases of reward and loss processing in PG. Given an inverse relationship between ventral striatal recruitment during anticipation of monetary rewards and impulsivity in alcohol dependence, the current study explored whether a similar association might also be present in PG. Methods Fourteen adults with PG and 14 control comparison (CC) participants performed the Monetary Incentive Delay Task (MIDT) to identify brain activation changes associated with reward/loss prospect, reward/loss anticipation and reward/loss notification. Impulsivity was assessed separately using the Barratt Impulsiveness Scale. Results Relative to the CC group, the PG group exhibited significantly reduced activity in the ventromedial prefrontal cortex, insula and ventral striatum during several phases, including the prospect and anticipation phases of both gain and losses. Activity in the ventral striatum correlated inversely with levels of impulsivity in PG participants, consistent with prior findings in alcohol dependence. Conclusions Relatively decreased activity in cortico-striatal neurocircuitry during multiple phases of reward processing suggests consistent alterations in neurocircuitry underlying incentive valuation and loss prediction. Together with findings in alcohol dependence, these results suggest that impulsive tendencies in addictions may be reflected in diminished ventral striatal activations to reward anticipation and may represent targets for treatment development in addictions. PMID:22336565

Does physics instruction foster university students' cognitive processes?: A descriptive study of teacher activities

NASA Astrophysics Data System (ADS)

Ferguson-Hessler, Monica G. M.; de Jong, Ton

This study aims at giving a systematic description of the cognitive activities involved in teaching physics. Such a description of instruction in physics requires a basis in two models, that is, the cognitive activities involved in learning physics and the knowledge base that is the foundation of expertise in that subject. These models have been provided by earlier research. The model of instruction distinguishes three main categories of instruction process: presenting new information, integrating (i.e., bringing structure into) new knowledge, and connecting elements of new knowledge to prior knowledge. Each of the main categories has been divided into a number of specific instruction processes. Hereby any limited and specific cognitive teacher activity can be described along the two dimensions of process and type of knowledge. The model was validated by application to lectures and problem-solving classes of first year university courses. These were recorded and analyzed as to instruction process and type of knowledge. Results indicate that teachers are indeed involved in the various types of instruction processes defined. The importance of this study lies in the creation of a terminology that makes it possible to discuss instruction in an explicit and specific way.

Targeting CXCR4 reverts the suppressive activity of T-regulatory cells in renal cancer.

PubMed

Santagata, Sara; Napolitano, Maria; D'Alterio, Crescenzo; Desicato, Sonia; Maro, Salvatore Di; Marinelli, Luciana; Fragale, Alessandra; Buoncervello, Maria; Persico, Francesco; Gabriele, Lucia; Novellino, Ettore; Longo, Nicola; Pignata, Sandro; Perdonà, Sisto; Scala, Stefania

2017-09-29

With the intent to identify biomarkers in renal cell carcinoma (RCC) the functional status of T-regulatory cells (Tregs) was investigated in primary RCC. Tregs were isolated from tumoral-(TT), peritumoral tissue-(PT) and peripheral blood-(PB) of 42 primary RCC patients and function evaluated through effector T cells (Teff) proliferation, cytokines release and demethylation of Treg Specific Region (TSDR). The highest value of Tregs was detected in TT with the uppermost amount of effector-Tregs-(CD4 + CD25 hi FOXP3 hi CD45RA - ). PB-RCC Tregs efficiently suppress Teff proliferation compared to healthy donor (HD)-Tregs and, at the intrapatient evaluation, TT-derived Tregs were the most suppressive. Higher demethylation TSDR was detected in TT- and PB-RCC Tregs vs HD-Tregs ( P <0,001). CXCR4 is highly expressed on Tregs, thus we wished to modulate Tregs function through CXCR4 inhibition. CXCR4 antagonism, elicited by a new peptidic antagonist, Peptide-R29, efficiently reversed Tregs suppression of Teff proliferation. Thus Tregs functional evaluation precisely reflects Tregs status and may be a reliable biomarker of tumoral immune response. In addition, treatment with CXCR4 antagonist, impairing Tregs function, could improve the anticancer immune response, in combination with conventional therapy and/or immunotherapy such as checkpoints inhibitors.

Targeting CXCR4 reverts the suppressive activity of T-regulatory cells in renal cancer

PubMed Central

Santagata, Sara; Napolitano, Maria; D'Alterio, Crescenzo; Desicato, Sonia; Maro, Salvatore Di; Marinelli, Luciana; Fragale, Alessandra; Buoncervello, Maria; Persico, Francesco; Gabriele, Lucia; Novellino, Ettore; Longo, Nicola; Pignata, Sandro; Perdonà, Sisto; Scala, Stefania

2017-01-01

With the intent to identify biomarkers in renal cell carcinoma (RCC) the functional status of T-regulatory cells (Tregs) was investigated in primary RCC. Tregs were isolated from tumoral-(TT), peritumoral tissue-(PT) and peripheral blood-(PB) of 42 primary RCC patients and function evaluated through effector T cells (Teff) proliferation, cytokines release and demethylation of Treg Specific Region (TSDR). The highest value of Tregs was detected in TT with the uppermost amount of effector-Tregs-(CD4+CD25hiFOXP3hiCD45RA-). PB-RCC Tregs efficiently suppress Teff proliferation compared to healthy donor (HD)-Tregs and, at the intrapatient evaluation, TT-derived Tregs were the most suppressive. Higher demethylation TSDR was detected in TT- and PB-RCC Tregs vs HD-Tregs (P <0,001). CXCR4 is highly expressed on Tregs, thus we wished to modulate Tregs function through CXCR4 inhibition. CXCR4 antagonism, elicited by a new peptidic antagonist, Peptide-R29, efficiently reversed Tregs suppression of Teff proliferation. Thus Tregs functional evaluation precisely reflects Tregs status and may be a reliable biomarker of tumoral immune response. In addition, treatment with CXCR4 antagonist, impairing Tregs function, could improve the anticancer immune response, in combination with conventional therapy and/or immunotherapy such as checkpoints inhibitors. PMID:29100374

Activity of two chlorinated lincomycin analogues against chloroquine-resistant falciparum malaria in owl monkeys.

PubMed

Powers, K G; Jacobs, R L

1972-01-01

The chloroquine-resistant Oak Knoll strain of Plasmodium falciparum, recently adapted to the owl monkey (Aotus trivirgatus), was insusceptible to chloroquine therapy. Two chlorinated lincomycin analogues tested in this host-parasite system cured blood-induced infections. Acute infections were treated orally for 7 consecutive days with either 15 or 75 mg of clindamycin hydrochloride (U-21) per kg per day, 10 or 50 mg of N-demethyl-4'-pentyl clindamycin hydrochloride (U-24) per kg per day, or 20 mg of chloroquine base per kg per day. These lincomycin analogues cleared trophozoites from the peripheral blood by the end of the 7-day treatment period. The speed of clearance of parasites was not dose-related, but curative activity appeared dependent upon the amount of drug given as well as the number of daily treatments. The efficacy of U-21 and U-24 is of particular interest since they represent major structural departures from compounds commonly used in the treatment of malaria.

Characterization of emission factors related to source activity for trichloroethylene degreasing and chrome plating processes.

PubMed

Wadden, R A; Hawkins, J L; Scheff, P A; Franke, J E

1991-09-01

A study at an automotive parts fabrication plant evaluated four metal surface treatment processes during production conditions. The evaluation provides examples of how to estimate process emission factors from activity and air concentration data. The processes were open tank and enclosed tank degreasing with trichloroethylene (TCE), chromium conversion coating, and chromium electroplating. Area concentrations of TCE and chromium (Cr) were monitored for 1-hr periods at three distances from each process. Source activities at each process were recorded during each sampling interval. Emission rates were determined by applying appropriate mass balance models to the concentration patterns around each source. The emission factors obtained from regression analysis of the emission rate and activity data were 16.9 g TCE/basket of parts for the open-top degreaser; 1.0 g TCE/1000 parts for the enclosed degreaser; 1.48-1.64 mg Cr/1000 parts processed in the hot CrO3/HNO3 tank for the chrome conversion coating; and 5.35-9.17 mg Cr/rack of parts for chrome electroplating. The factors were also used to determine the efficiency of collection for the local exhaust systems serving each process. Although the number of observations were limited, these factors may be useful for providing initial estimates of emissions from similar processes in other settings.

Human ecstasy (MDMA) polydrug users have altered brain activation during semantic processing

PubMed Central

Watkins, Tristan J.; Raj, Vidya; Lee, Junghee; Dietrich, Mary S.; Cao, Aize; Blackford, Jennifer U.; Salomon, Ronald M.; Park, Sohee; Benningfield, Margaret M.; Di Iorio, Christina R.; Cowan, Ronald L.

2012-01-01

Rationale Ecstasy (MDMA) polydrug users have verbal memory performance that is statistically significantly lower than comparison control subjects. Studies have correlated long-term MDMA use with altered brain activation in regions that play a role in verbal memory. Objectives The aim of our study was to examine the association of lifetime ecstasy use with semantic memory performance and brain activation in ecstasy polydrug users. Methods 23 abstinent ecstasy polydrug users (age=24.57) and 11 controls (age=22.36) performed a two-part fMRI semantic encoding and recognition task. To isolate brain regions activated during each semantic task, we created statistical activation maps in which brain activation was greater for word stimuli than for non-word stimuli (corrected p<0.05). Results During the encoding phase, ecstasy polydrug users had greater activation during semantic encoding bilaterally in language processing regions, including Brodmann Areas 7, 39, and 40. Of this bilateral activation, signal intensity with a peak T in the right superior parietal lobe was correlated with lifetime ecstasy use (rs=0.43, p=0.042). Behavioral performance did not differ between groups. Conclusions These findings demonstrate that ecstasy polydrug users have increased brain activation during semantic processing. This increase in brain activation in the absence of behavioral deficits suggests that ecstasy polydrug users have reduced cortical efficiency during semantic encoding, possibly secondary to MDMA-induced 5-HT neurotoxicity. Although pre-existing differences cannot be ruled out, this suggests the possibility of a compensatory mechanism allowing ecstasy polydrug users to perform equivalently to controls, providing additional support for an association of altered cerebral neurophysiology with MDMA exposure. PMID:23241648

Severe tremor after cotrimoxazole-induced elevation of venlafaxine serum concentrations in a patient with major depressive disorder.

PubMed

Geber, Christian; Ostad Haji, Elnaz; Schlicht, Konrad; Hiemke, Christoph; Tadić, André

2013-06-01

: We describe a female patient who was an extensive metabolizer of cytochrome P450 isoenzyme (CYP) 2D6 and an intermediate metabolizer of CYP2C19 (genotype: CYP2C19 *1/*2). She exhibited high serum concentrations of venlafaxine and O-desmethylvenlafaxine and developed severe tremor after comedication with cotrimoxazole (sulfamethazole/trimethoprim). Venlafaxine is mainly metabolized by O- and N-demethylation. O-demethylation is catalyzed by the highly polymorphic CYP2D6 and N-demethylation by several enzymes, CYP2C19, CYP2C9, and CYP3A4. The observed overall pharmacokinetic effect was most probably the result of decreased N-demethylation of venlafaxine by (1) reduced expression of CYP2C19 due to a genetic deficit and (2) inhibition of CYP2C9 by cotrimoxazole.

PROCESS FOR THE PRODUCTION OF AN ACTIVATED FORM OF UO$sub 2$

DOEpatents

Polissar, M.J.

1957-09-24

A process for producing a highly active form of UO/sub 2/ characterized both by rapid oxidation in air and by rapid chlorination with CCl/sub 4/ vapor at an elevated temperature is reported. In accordance with the process, commercial UO/sub 2/, is subjected to a series of oxidation-reduction operations to produce a form of UC/sub 2/ of enhanced reactivity. By treatimg commercial UO/sub 2/ at a temperature between 335 and 485 deg C with methane, then briefly with an oxygen containing gas and followimg this by a second treatment with a methane containing gas, the original relatively stable charge of UO/sub 2/ will be transformed into an active form of UO/sub 2/.

Enhanced Passive and Active Processing of Syllables in Musician Children

ERIC Educational Resources Information Center

Chobert, Julie; Marie, Celine; Francois, Clement; Schon, Daniele; Besson, Mireille

2011-01-01

The aim of this study was to examine the influence of musical expertise in 9-year-old children on passive (as reflected by MMN) and active (as reflected by discrimination accuracy) processing of speech sounds. Musician and nonmusician children were presented with a sequence of syllables that included standards and deviants in vowel frequency,…

Alterations in protein kinase C activity and processing during zinc-deficiency-induced cell death.

PubMed

Chou, Susan S; Clegg, Michael S; Momma, Tony Y; Niles, Brad J; Duffy, Jodie Y; Daston, George P; Keen, Carl L

2004-10-01

Protein kinases C (PKCs) are a family of serine/threonine kinases that are critical for signal transduction pathways involved in growth, differentiation and cell death. All PKC isoforms have four conserved domains, C1-C4. The C1 domain contains cysteine-rich finger-like motifs, which bind two zinc atoms. The zinc-finger motifs modulate diacylglycerol binding; thus, intracellular zinc concentrations could influence the activity and localization of PKC family members. 3T3 cells were cultured in zinc-deficient or zinc-supplemented medium for up to 32 h. Cells cultured in zinc-deficient medium had decreased zinc content, lowered cytosolic classical PKC activity, increased caspase-3 processing and activity, and reduced cell number. Zinc-deficient cytosols had decreased activity and expression levels of PKC-alpha, whereas PKC-alpha phosphorylation was not altered. Inhibition of PKC-alpha with Gö6976 had no effect on cell number in the zinc-deficient group. Proteolysis of the novel PKC family member, PKC-delta, to its 40-kDa catalytic fragment occurred in cells cultured in the zinc-deficient medium. Occurrence of the PKC-delta fragment in mitochondria was co-incident with caspase-3 activation. Addition of the PKC-delta inhibitor, rottlerin, or zinc to deficient medium reduced or eliminated proteolysis of PKC-delta, activated caspase-3 and restored cell number. Inhibition of caspase-3 processing by Z-DQMD-FMK (Z-Asp-Gln-Met-Asp-fluoromethylketone) did not restore cell number in the zinc-deficient group, but resulted in processing of full-length PKC-delta to a 56-kDa fragment. These results support the concept that intracellular zinc concentrations influence PKC activity and processing, and that zinc-deficiency-induced apoptosis occurs in part through PKC-dependent pathways.

Loss of the retinoblastoma binding protein 2 (RBP2) histone demethylase suppresses tumorigenesis in mice lacking Rb1 or Men1

PubMed Central

Lin, Wenchu; Cao, Jian; Liu, Jiayun; Beshiri, Michael L.; Fujiwara, Yuko; Francis, Joshua; Cherniack, Andrew D.; Geisen, Christoph; Blair, Lauren P.; Zou, Mike R.; Shen, Xiaohua; Kawamori, Dan; Liu, Zongzhi; Grisanzio, Chiara; Watanabe, Hideo; Minamishima, Yoji Andrew; Zhang, Qing; Kulkarni, Rohit N.; Signoretti, Sabina; Rodig, Scott J.; Bronson, Roderick T.; Orkin, Stuart H.; Tuck, David P.; Benevolenskaya, Elizaveta V.; Meyerson, Matthew; Kaelin, William G.; Yan, Qin

2011-01-01

Aberrations in epigenetic processes, such as histone methylation, can cause cancer. Retinoblastoma binding protein 2 (RBP2; also called JARID1A or KDM5A) can demethylate tri- and dimethylated lysine 4 in histone H3, which are epigenetic marks for transcriptionally active chromatin, whereas the multiple endocrine neoplasia type 1 (MEN1) tumor suppressor promotes H3K4 methylation. Previous studies suggested that inhibition of RBP2 contributed to tumor suppression by the retinoblastoma protein (pRB). Here, we show that genetic ablation of Rbp2 decreases tumor formation and prolongs survival in Rb1+/− mice and Men1-defective mice. These studies link RBP2 histone demethylase activity to tumorigenesis and nominate RBP2 as a potential target for cancer therapy. PMID:21788502

Seasonal mercury transformation and surficial sediment detoxification by bacteria of Marano and Grado lagoons

NASA Astrophysics Data System (ADS)

Baldi, Franco; Gallo, Michele; Marchetto, Davide; Fani, Renato; Maida, Isabel; Horvat, Milena; Fajon, Vesna; Zizek, Suzana; Hines, Mark

2012-11-01

Marano and Grado lagoons are polluted by mercury from the Isonzo River and a chlor-alkali plant, yet despite this contamination, clam cultivation is one of the main activities in the region. Four stations (MA, MB, MC and GD) were chosen for clam seeding and surficial sediments were monitored in autumn, winter and summer to determine the Hg detoxifying role of bacteria. Biotransformation of Hg species in surficial sediments of Marano and Grado lagoons was investigated while taking into consideration the speciation of organic matter in the biochemical classes of PRT (proteins), CHO (carbohydrates) and LIP (lipids), water-washed cations and anions, bacterial biomass, Hg-resistant bacteria, some specific microbial activities such as sulfate reduction rates, Hg methylation rates, Hg-demethylation rates, and enzymatic ionic Hg reduction. MeHg in sediments was well correlated with PRT content, whereas total Hg in sediments correlated with numbers of Hg-resistant bacteria. Correlations of the latter with Hg-demethylation rates in autumn and winter suggested a direct role Hg-resistant bacteria in Hg detoxification by producing elemental Hg (Hg0) from ionic Hg and probably also from MeHg. MeHg-demethylation rates were ˜10 times higher than Hg methylation rates, were highest in summer and correlated with high sulfate reduction rates indicating that MeHg was probably degraded in summer by sulfate-reducing bacteria via an oxidative pathway. During the summer period, aerobic heterotrophic Hg-resistant bacteria decreased to <2% compared to 53% in winter. Four Hg-resistant bacterial strains were isolated, two Gram-positive (Staphylococcus and Bacillus) and two Gram-negative (Stenotrophomonas and Pseudomonas). Two were able to produce Hg0, but just one contained a merA gene; while other two strains did not produce Hg0 even though they were able to grow at 5 μg ml of HgCl2. Lagoon sediments support a strong sulfur cycle in summer that controls Hg methylation and demethylation