De novo DNA methylation independent establishment of maternal imprint on X chromosome in mouse oocytes

In female mouse embryos, the paternal X chromosome (Xp) is preferentially inactivated during preimplantation development and trophoblast differentiation. This imprinted X-chromosome inactivation (XCI) is partly due to an activating imprint on the maternal X chromosome (Xm), which is set during oocyte growth. However, the nature of this imprint is unknown. DNA methylation is one candidate, and therefore we examined whether disruptions of the two de novo DNA methyltransferases in growing oocytes affect imprinted XCI. We found that accumulation of histone H3 lysine-27 trimethylation, a hallmark of XCI, occurs normally on the Xp, and not on the Xm, in female blastocysts developed from the mutant oocytes. Furthermore, the allelic expression patterns of X-linked genes including Xist and Tsix were unchanged in preimplantation embryos and also in the trophoblast. These results show that a maternal disruption of the DNA methyltransferases has no effect on imprinted XCI and argue that de novo DNA methylation is dispensable for Xm imprinting. This underscores the difference between imprinted XCI and autosomal imprinting.     

Determination of minimum biofilm eradication concentrations of orbifloxacin for canine bacterial uropathogens over different treatment periods

Biofilm formation can cause refractory urinary tract infections (UTIs) in dogs; however, minimum biofilm eradication concentrations (MBECs) of veterinary drugs against canine uropathogens remain to be investigated. In this study, the MBECs of orbifloxacin (OBFX), trimethoprim‐sulfamethoxazole (TMS) and amoxicillin/clavulanate (ACV) over different time periods for treatment of canine uropathogenic Escherichia coli (n = 10) were determined. The MBECs of OBFX for other bacterial uropathogens, including Staphylococcus pseudintermedius (n = 5), Pseudomonas aeruginosa (n = 5), Klebsiella pneumoniae (n = 5) and Proteus mirabilis (n = 5) were also determined. Minimum inhibitory concentrations (MICs) were identified for all strains by broth microdilution, and MBECs were determined at 24, 72, and 168 hr using the Calgary biofilm method. The 24 hr MBECs of OBFX, TMS and ACV for the E. coli strains were significantly higher than the MICs (P < 0.05), and the 72 and 168 hr MBECs were significantly lower than those at 24 hr (P < 0.05). In addition, the 24 hr OBFX MBECs for the four other uropathogens were significantly higher than the corresponding MICs (P < 0.05). The 72 and/or 168 hr OBFX MBECs for S. pseudintermedius, K. pneumoniae and P. mirabilis were significantly lower than the 24 hr concentrations (P < 0.05), whereas for P. aeruginosa, no significant difference was found between any of the MBECs (P > 0.05). These data indicate that the administration period and uropathogenic bacterial species are important factors affecting the efficacy of OBFX treatment of biofilm‐related UTIs in dogs.     

Free fatty acids alter the electrophoretic mobility of serum lipoproteins

When normal human serum was analysed after addition of free fatty acids (FFA: oleic acids), the lipoproteins showed abnormal mobility on agarose-gel electrophoresis. β- and α-lipoproteins from the serum with a FFA concentration of 2.04 μEq/ml or more migrated with a mobility showing a more negative charge than native ones. Although the ultracentrifugal profile of the lipoprotein cholesterol was not altered with the excess-FFA, FFA were detected in the lipoprotein fractions as well as in free proteins. The lipoproteins isolated from the excess-FFA aerum also showed abnormal mobility on agarosegel electrophoresis.These results suggest that, in the serum containing excess-FFA, FFA attach themselves to the lipoproteins and alter their charge.     

Okadaic acid and dinophysistoxin-1, non-TPA-type tumor promoters, stimulate prostaglandin E2 production in rat peritoneal macrophages

Okadaic acid and dinophysistoxin-1 isolated from a black sponge, Halichondria okadai are non-12-O-tetrade-canoylphorbol 13-acetate (non-TPA)-type tumor promoters of mouse skin. Okadaic acid at concentrations of 10-100 ng/ml stimulated prostaglandin E2 production in rat peritoneal macrophages. Dinophysistoxin-1 (35-methylokadaic acid) stimulated prostaglandin E2 production as strong as okadaic acid, but okadaic acid tetramethyl ether, an inactive compound as a tumor promoter, did not. Okadaic acid at 10 ng/ml (12.4 nM) stimulated prostaglandin E2 production as strongly as TPA at 10 ng/ml (16.2 nM) 20 h after incubation. Unlike TPA-type tumor promoters, okadaic acid required a lag phase before stimulation. The duration of this lag phase was dependent on the concentration of okadaic acid. Indomethacin inhibited okadaic acid-induced preostaglandin E2 production in a dose-dependent manner, and its inhibition was more strongly observed in okadaic acid-induced prostaglandin E2 production. Cycloheximide inhibited okadaic acid-induced release of radioactivity from [3H]arachidonic acid-labeled macrophages and prostaglandin E2 production dose dependently, suggesting that protein synthesis is a prerequisite for the stimulation of arachidonic acid metabolism. These results support our idea that tumor promoters, at very low concentrations, are able to stimulate arachidonic acid metabolism in rat peritoneal macrophages.     

Spontaneous development of dermatitis in DS-Nh mice under specific pathogen-free conditions.

Spontaneous development of dermatitis in DS-Nh mice under specific pathogen-free conditions was examined to verify the hypothesis [Exp. Anim. 46: 225-229, 1997] that Stapylococcus aureus (S. aureus) infection is causally associated with the dermatitis. Observation of the mice up to 28 weeks of age indicated that obvious dermatitis does occur under S. aureus-free conditions, though the incidence was low (six of 42 females and two of 90 males). Skin lesions in the absence of this bacterium showed histological changes very similar to those that can be observed under conventional conditions. In addition, hyperproduction of serum IgE was demonstrated in the dermatitis-positive mouse. These findings suggested that the dermatitis is triggered by IgE-mediated allergic reactions.     

Purification and properties of a polyamine oxidase from Zea mays

Polyamine oxidase, purified 260-fold from maize shoots, was light yellow in colour. Maximum light-absorption was at 450 nm and was decreased by the addition of either sodium dithionite or spermidine, but not by putrescine. Under aerobic conditions, the enzyme could use p-benzoquinone as an electron acceptor. Cu²⁺ inhibited the enzyme activity, while SO₃⁻ was stimulatory. Several metal-binding agents and thiol reagents were without effect.     

A nucleotide-phosphohydrolyzing activity in non-histone proteins of pea cotyledon inhibits in vitro RNA synthesis

Concerning in vitro RNA synthesis with E. coli RNA polymerase,an inhibitor which exists in non-histone proteins of germinatedpea cotyledon was investigated. The inhibition was attributedto the phosphohydrolysis of UTP as a substrate in the assaysystem. UTP was degraded to UMP and 2 Pi but ATP was hardlyaffected. (Received February 13, 1978; )