Plasmid-coded degradation of ethylbenzene and 1-phenylethanol in Pseudomonas fluorescens

Abstract Pseudomonas fluorescens EB carries genes for the catabolism of ethylbenzene and 1-phenylethanol on a plasmid. The size of the plasmid as measured by analysis of agarose electrophoresis gels after restriction endonuclease hydrolysis, was 253–267 kb. By treatment with Mitomycin C, mutants of EB strain were obtained bearing a plasmid which had undergone an extensive deletion of about 80 kb. These mutants have lost the ability to grow on ethylbenzene and 1-phenylethanol as well as to synthesize meta-cleavage enzymes.     

Ethanol Administration in the Rat Decreases Prostacyclin Production by Isolated Brain Microvessels

The effects of short- and long-term ethanol administration to rats on basal levels and formation of prostacyclin (PGI2) measured as 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), and on lipid class content and fatty acid composition of isolated brain microvessels (BMV) were studied. After acute treatment (2 h, at the peak of plasma ethanol concentration) basal 6-keto-PGF1 alpha levels in BMV and release on incubation were reduced to 50% of control values. After chronic administration (15 days), PGI2 release was reduced to about 40% of control values, without changes in basal levels. Total lipid, phospholipid, and cholesterol levels in BMV, measured after prolonged administration of alcohol, were not modified. Also, only minor changes in the fatty acid composition of individual phospholipid classes were detected. The observed reduction of PGI2 synthesis in BMV thus could not be related to changes of the fatty acid precursor pool in the preparation. Precursor release and/or the biosynthetic pathways may be affected by ethanol administration.     

Ultrastructural cytochemical and autoradiographic demonstration of nonspecific esterase(s) in guinea pig basophils

We used ultrastructural autoradiographic and cytochemical methods to localize esterase activities in unstimulated guinea pig basophils and in basophils undergoing degranulation or recovery from degranulation. We used tritium-labeled diisopropylfluorophosphate (DFP) as a probe for serine enzymes and localized this probe by ultrastructural autoradiography to cytoplasmic granules of immature or mature unstimulated basophils, as well as to granules released by degranulating basophils. Ultrastructural cytochemistry using alpha naphthyl acetate (ANA) as substrate localized nonspecific esterase activity to extruded granules, either within the interiors of degranulation sacs or within granules completely separated from degranulating basophils. Extruded granules retained their esterase activity for as long as 24 hr after antigen-induced degranulation. The plasma membranes of unstimulated or degranulating basophils, as well as of basophils recovering from degranulation, displayed prominent cell surface ANA esterase ectoenzyme activity. Lipid bodies, organelles present in the cytoplasm of both control and recovering basophils, were also alpha naphthyl acetate esterase (ANAE)-positive. Thus, cytochemical and autoradiographic techniques localized esterase and/or [3H]-DFP-binding activities to cytoplasmic granules, lipid bodies, and cell surface of basophils, and these enzyme activities persisted during both degranulation and recovery from degranulation.     

Stimulation of arachidonic acid metabolism by CCl4 in isolated rat hepatocytes

Arachidonic acid metabolism was evaluated in isolated rat hepatocytes after CCl4 exposure. CCl4 induced dose-dependently the synthesis and release of prostacyclin (PGI2) and thromboxane (TXB2). Treatment with prostaglandin E2 (PGE2) 30 min after exposure to CCl4, significantly reduced the cell damage as well as the release of TXB2 from the cells.     

Dietary n-9, n-6, and n-3 fatty acids modify linoleic acid more than arachidonic acid levels in plasma and platelet lipids and minimally affect platelet thromboxane formation in the rabbit

We have studied the effects of semisynthetic diets containing 5% by weight (12% of the energy) of either olive oil (70% oleic acid, OA) or corn oil (58% linoleic acid), or fish oil (Max EPA, containing about 30% eicosapentaenoic, EPA C 20:5 n-3, plus docosahexaenoic, DHA C 22:6 n-3, acids, and less than 2% linoleic acid), fed to male rabbits for a period of five weeks, on plasma and platelet fatty acids and platelet thromboxane formation. Aim of the study was to quantitate the absolute changes of n-6 and n-3 fatty acid levels in plasma and platelet lipid pools after dietary manipulations and to correlate the effects on eicosanoid-precursor fatty acids with those on platelet thromboxane formation. The major differences were found when comparing the group fed fish oil and depleted linoleic acid vs the other groups. The accumulation of n-3 fatty acids in various lipid classes was associated with modifications in the distribution of linoleic acid and arachidonic acid in different lipid pools. In platelets maximal incorporation of n-3 fatty acids occurred in phosphatidyl ethanolamine, which also participated in most of the total arachidonic acid reduction occurring in platelets, and linoleic acid, more than archidonic acid, was replaced by n-3 fatty acids in various phospholipids. The archidonic acid content of phosphatidyl choline was unaffected and that of phosphatidyl inositol only marginally reduced. Thromboxane formation by thrombin stimulated platelets did not differ among the three groups, and this may be related to the minimal changes of arachidonic acid in phosphatidyl choline and phosphatidyl inositol.     

Chromosome abnormalities in tuberous sclerosis

Summary In fibroblasts cultured from biopsies of the skin lesions of six patients with tuberous sclerosis (TS) there was a variable but consistent degree of karyotypic variation. Premature centromere disjunction (PCD) of all or part of the chromosomes, micronuclei, an increased incidence of breaks, dicentric chromosomes and the presence of polyploid metaphases were found in all cultures. The PCD was of the type encountered in Roberts syndrome and its frequency varied from 8% to 30%. In metaphases with PCD of one and of two chromosomes, the chromosome involved were identified, and chromosome 3 was involved 21 times among 59 chromosomes with PCD. Chromosome 3 tends to be preferentially involved in dicentric formation. In lymphocyte cultures from the same patients there were no metaphases with PCD, but there was a slight increase of breaks and the presence of dicentric chromosomes, also involving chromosome 3. Polyploid metaphases were increased in some of the cases. Karyotypic variation can be considered a cellular phenotypic characteristic of TS in fibroblasts cultured from the skin lesions, and its type indicates disturbances in the mechanics of centromere division and of chromosome distribution at cell division.     

Unilateral reduction mammaplasty: sculpturing the breast from the undersurface

Various traditional mammaplasty techniques have been suggested for unilateral breast reduction, and an inverted-T incision is still the most popular approach. However, unilaterally performed traditional techniques can rarely provide long-lasting symmetry because the operated and the unoperated breasts react differently to aging, weight changes, and pregnancy. Considerable residual scarring, interference with clinical and mammographic evaluation, and limited versatility are all major drawbacks of traditional procedures. We have performed unilateral mammaplasties on 47 patients with various types of congenital and acquired asymmetries, reducing and sculpturing the breast from the undersurface by means of minimal incisions, always avoiding horizontal scarring in the inframammary crease. Through a vertical infra-areolar incision, the breast is completely detached from the underlying pectoralis fascia and hooked up, thus completely exposing the undersurface of the mammary cone. The breast can thereafter be reshaped according to the size and shape of the contralateral breast by means of a discoid resection and/or selective sectoral removal of excessive subcutaneous tissues; modifications of the basic discoid resection can increase anterior projection of the new breast mound and can change the inclination of the anteroposterior breast axis on the anterior chest wall both on the horizontal and vertical planes. The results show that if criteria for patient selection are carefully respected, the procedure can provide long-lasting symmetry with minimal residual scarring and fully preserve the breast anatomy, function, and vascularization.     

The AT-rich tract of the SV40 ori core: negative synergism and specific recognition by single stranded and duplex DNA binding proteins.

The SV40 origin of replication comprises a run of thymine and adenine residues. Integrity of this AT-rich sequence is known to be essential for replication. We set out to study whether or not these elements can work synergistically to sustain replication. Quite surprisingly, additional copies of the AT stretch linked to a functional SV40 ori core dramatically reduce its replication in Cosl cells, probably by creating some physical block. Interestingly, the same inhibiting effect can be observed with the addition in cis of the yeast ARS consensus, which is homologous to the SV40 AT stretch. This modulation is possibly due to the action of cellular factors that recognize either of the two sequences. In fact, we demonstrate the existence of factor(s) in Cosl crude nuclear extracts that in vitro can specifically bind to either of them. Moreover, we show that these sequence-specific factor(s) (MW about 50 kDa), named SOAP, recognize both single (T-rich strand) and double stranded forms of the AT tracts. Binding to single stranded AT stretches can be specifically inhibited by the corresponding duplex form, but not vice versa.     

Frozen bovine semen quality and bovine cervical mucus penetration test

Research has been carried out to test bovine cervical mucus penetration (penetration) as a means for evaluating frozen-thawed bovine semen. A commercially available cervical mucus penetration test kit (the kit) was used. A total of 158 previously frozen semen samples collected from 61 bulls were thawed in a 37 C water-bath for 2 minutes. Four ways to estimate penetration were compared using the distance traveled during 90 minutes 1) at 21 C, or 2) at 37 C, by 3) the first solitary mobile spermatozoon, or by 4) the front of the mass of the mobile spermatozoa. Penetration was measured using phase contrast microscopy and a millimeter grid. Spermatozoal quality parameters (concentration, total motility, progressive motility, acrosome integrity, total sperm integrity and cytoplasmic droplets) were measured and the correlation to penetration was calculated. The best way to assay penetration with the kit was by measuring the penetration of the first solitary mobile spermatozoon at 37 C. Semen quality variability was significant (P < 0.05) relative to penetration. Linear correlations between penetration and acrosome integrity r=0.42 as well as between penetration and total sperm integrity r=0.53 were highly significant (P < 0.001). There was significant linear multiple regression between penetration and acrosome integrity (expressed as percentage and number) and total sperm integrity (expressed as percentage and number) (r=0.62; F=23.5147; P<0.0001). There was a significant difference between the average progressive motility of samples with penetration > 20 mm and samples with penetration 20%), but it is not useful to define the fertility level of semen samples.