Disease-dependent differently methylated regions (D-DMRs) of DNA are enriched on the X chromosome in uterine leiomyoma

Uterine leiomyoma is the most common benign tumor in women. Although responsible gene mutations have not been found in leiomyomas, they represent a progressive disease with irreversible symptoms. To characterize epigenetic features of uterine leiomyomas, the DNA methylation status of a paired sample of leiomyoma and normal myometrium was subjected to a microarray-based DNA methylation analysis with restriction tag-mediated amplification (D-REAM). In the leiomyoma, we identified an aberrant DNA methylation status for 463 hypomethylated and 318 hypermethylated genes. Although these changes occurred on all chromosomes, aberrantly hypomethylated genes were preferentially located on the X chromosome. Using paired samples of normal myometrium and leiomyoma from 6 hysterectomy patients, methylation-sensitive quantitative real-time PCR revealed 14 shared X chromosome genes with an abnormal DNA hypomethylation status (FAM9A, CPXCR1, CXORF45, TAF1, NXF5, VBP1, GABRE, DDX53, FHL1, BRCC3, DMD, GJB1, AP1S2 and PCDH11X) and one hypermethylated locus (HDAC8). Expression of XIST, which is involved in X chromosome inactivation, was equivalent in the normal myometrium and leiomyoma, indicating that the epigenetic abnormality on the X chromosome did not result from aberration of XIST gene expression. Based on these data, a unique epigenetic signature for uterine leiomyomas has emerged. The 14 hypomethylated and one hypermethylated loci provide valuable biomarkers for understanding the molecular pathogenesis of leiomyoma.     

Nitric oxide stimulates IP3 production via a cGMP/ PKG-dependent pathway in rat pancreatic acinar cells

In an attempt to explore the functioning of nitric oxide (NO) in pancreatic exocrine cells, we have recently obtained several lines of circumstantial evidence indicating that one of molecular targets of NO is phospholipase C (PLC), the activation of which leads to an increase in the cytosolic Ca2+ concentration ([Ca2+]i) via inositol 1, 4, 5-trisphosphate, IP3. However, whether IP3 is actually produced by NO has not yet been substantiated. The present study was therefore designed to directly measure the intracellular IP3, concentration ([IP3]i) for better understanding of the underlying mechanisms with the help of pharmacological tools. [IP3]i was measured using a fluorescence polarization technique (HitHunter). We obtained the following results: 1) varying concentrations of an NO donor, sodium nitroprusside (SNP), elevated [IP3]i, 2) this elevation was completely inhibited in the presence of the soluble guanylyl cyclase (sGC) inhibitor, 1H-[1, 2, 4] oxadiazolo [4, 3-a] quinoxalin-1-one (ODQ), 3) varying concentrations of the cGMP analogue, 8-Br-cGMP, also increased [IP3]i, 4) the cGMP analogue-induced IP3 production was abolished by pretreatment with either a PLC inhibitor, U73122, or a G-protein inhibitor, GP2A, and 5) KT5823, a potent and highly selective inhibitor of cGMP-dependent protein kinase G (PKG), also abolished the IP3 production induced by 8-Br-cGMP. These results suggest that the NO-induced [Ca2+]i increase is triggered by an increase in [IP3]i located downstream from intracellular cGMP elevation. In this intracellular pathway, each sGC, cGMP-dependent PKG, G-protein and PLC were suggested to be involved. The present work provides new insights into the intracellular signaling accelerated by NO. NO triggers a [Ca2+]I increase via cGMP and IP3 in pancreatic acinar cells.     

Validation of a direct time-resolved fluoroimmunoassay for progesterone in milk from dairy and beef cows

The aim of this study was to validate a direct time-resolved fluoroimmunoassay (TR-FIA) for quantifying progesterone concentrations in milk during the bovine oestrous cycle. Holstein-Friesian and suckled and non-suckled Japanese Black cows were used to demonstrate the relationship between milk and plasma progesterone concentrations and to monitor progesterone profiles in milk and plasma during the oestrous cycle. The minimum detection level of the assay was 1.53ng/mL. Progesterone concentrations in milk and plasma changed in a similar manner throughout the oestrous cycle in dairy and beef cows, and milk and plasma progesterone profiles were significantly correlated (P<0.001). The study confirmed that a direct TR-FIA can be used to monitor the oestrous cycle in cattle and to quantify progesterone concentrations in whole milk.     

Effect of supplementation of soy sauce oil and Ca salts of fatty acids on rumen fermentation,milk production and conjugated linoleic acid in milk of dairy cows

Eight cows were used to evaluate the effects of supplementation of soy sauce oil (SO) or Ca salts of fatty acids (FA) on rumen fermentation and milk production. The control diet (CO) consisted mainly of hay, corn silage and a concentrate. In the experimental diets, 400 g/day per cow of SO or FA (soybean oil and rapeseed oil) was supplemented to the CO diet. Experimental period for the three treatments was 14 days, and milk samples were taken during the last 2 days and rumen sample was taken on the last day. Dry matter intake was not affected by the treatments. The number of rumen protozoa at 0 h increased by SO and FA diets. Total volatile fatty acids at 2 h after feeding of SO diet was decreased compared to CO. The milk composition yield did not differ among treatments, although the percentages of fat and protein were decreased by SO and FA diets. The proportions of C8–C16 fatty acids in milk fat decreased, and those of C18 increased by SO and FA diets. The proportion of cis‐9, trans‐11 conjugated linoleic acid in milk fat by SO and FA diets increased by 120% and 135%, respectively. In spite of the slight suppression of rumen fermentation by SO diet, negative effects on feed intake and milk production were not detected.     

Complex patterning by vertical interchange atom manipulation using atomic force microscopy

The ability to incorporate individual atoms in a surface following predetermined arrangements may bring future atom-based technological enterprises closer to reality. Here, we report the assembling of complex atomic patterns at room temperature by the vertical interchange of atoms between the tip apex of an atomic force microscope and a semiconductor surface. At variance with previous methods, these manipulations were produced by exploring the repulsive part of the short-range chemical interaction between the closest tip-surface atoms. By using first-principles calculations, we clarified the basic mechanisms behind the vertical interchange of atoms, characterizing the key atomistic processes involved and estimating the magnitude of the energy barriers between the relevant atomic configurations that leads to these manipulations.     

Orbital motion in the radio galaxy 3C 66B: evidence for a supermassive black hole binary

Supermassive black hole binaries may exist in the centers of active galactic nuclei such as quasars and radio galaxies, and mergers between galaxies may result in the formation of supermassive binaries during the course of galactic evolution. Using the very-long-baseline interferometer, we imaged the radio galaxy 3C 66B at radio frequencies and found that the unresolved radio core of 3C 66B shows well-defined elliptical motions with a period of 1.05 +/- 0.03 years, which provides a direct detection of a supermassive black hole binary.     

Antagonistic effects of atipamezole,yohimbine, and prazosin on xylazine-induced diuresis in clinically normal cats

This study aimed to investigate and compare the antagonistic effects of atipamezole, yohimbine, and prazosin on xylazine-induced diuresis in clinically normal cats. Five cats were repeatedly used in each of the 9 groups. One group was not medicated. Cats in the other groups received 2 mg/kg BW xylazine intramuscularly, and saline (as the control); 160 μg/kg BW prazosin; or 40, 160, or 480 μg/kg BW atipamezole or yohimbine intravenously 0.5 h later. Urine and blood samples were collected 10 times over 8 h. Urine volume, pH, and specific gravity; plasma arginine vasopressin (AVP) concentration; and creatinine, osmolality, and electrolyte values in both urine and plasma were measured. Both atipamezole and yohimbine antagonized xylazine-induced diuresis, but prazosin did not. The antidiuretic effect of atipamezole was more potent than that of yohimbine but not dose-dependent, in contrast to the effect of yohimbine at the tested doses. Both atipamezole and yohimbine reversed xylazine-induced decreases in both urine specific gravity and osmolality, and the increase in free water clearance. Glomerular filtration rate, osmolar clearance, and plasma electrolyte concentrations were not significantly altered. Antidiuresis of either atipamezole or yohimbine was not related to the area under the curve for AVP concentration, although the highest dose of both atipamezole and yohimbine increased plasma AVP concentration initially and temporarily, suggesting that this may in part influence antidiuretic effects of both agents. The diuretic effect of xylazine in cats may be mediated by α₂-adrenoceptors but not α₁-adrenoceptors. Atipamezole and yohimbine can be used as antagonistic agents against xylazine-induced diuresis in clinically normal cats.     

Isoxaben analogs inhibit chitin synthesis in the cultured integument of the rice stem borer Chilo suppressalis

Benzoylphenylureas (BPUs) were discovered as novel type insecticides about a half century ago; many analogs have been launched as insecticides and acaricides. BPUs are known to inhibit chitin synthesis in insects and other arthropods, but they have no effect against microorganisms such as fungi. We designed new chitin synthesis inhibitors based on the hypothesis that biomolecules that play important roles in cellulose and chitin biosynthesis are similar. In the full automatic modeling system (FAMS), the cellulose synthase was selected as a template three-dimensional structure. Thus, we focused on the structure of cellulose synthase inhibitor, isoxaben, to develop new chemistry. The 1,1-diethylethyl [-C(CH₃)(CH₂CH₃)₂] group of isoxaben was changed to a 4-substituted phenyl group bearing Cl, Et, or Ph. These compounds significantly inhibited chitin synthesis in the cultured integument of the rice stem borer Chilo suppressalis. The activity of the 4-ethylphenyl analog was enhanced 30-fold by adding piperonyl butoxide to the culture medium.     

Nonsteroidal ecdysone receptor agonists use a water channel for binding to the ecdysone receptor complex EcR/USP

The ecdysone receptor (EcR) possesses the remarkable capacity to adapt structurally to different types of ligands. EcR binds ecdysteroids, including 20-hydroxyecdysone (20E), as well as nonsteroidal synthetic agonists such as insecticidal dibenzoylhydrazines (DBHs). Here, we report the crystal structures of the ligand-binding domains of Heliothis virescens EcR/USP bound to the DBH agonist BYI09181 and to the imidazole-type compound BYI08346. The region delineated by helices H7 and H10 opens up to tightly fit a phenyl ring of the ligands to an extent that depends on the bulkiness of ring substituent. In the structure of 20E-bound EcR, this part of the ligand-binding pocket (LBP) contains a channel filled by water molecules that form an intricate hydrogen bond network between 20E and LBP. The water channel present in the nuclear receptor bound to its natural hormone acts as a critical molecular adaptation spring used to accommodate synthetic agonists inside its binding cavity.