Effects of pretreatment with a xanthine oxidase inhibitor on free radical levels during carotid endarterectomy

Objective: Free radicals contribute to the tissue damage caused by ischaemia-reperfusion. The aim of the present study was to investigate whether preoperative antioxidant therapy (allopurinol) affects free radical levels in cerebral venous blood in connection with surgery for carotid artery stenosis.

Materials and methods: Twenty-five patients were randomised into the study. Thirteen were controls and 12 were pretreated with allopurinol the day before surgery. Before, during and after surgery, blood samples were drawn from the ipsilateral jugular vein. Radical levels were measured using the spin trap technique ex vivo using OXANOH as the spin trap. Multivariate statistics were used with Principal Component Analysis and Partial Least Square regression analysis.

Results: Radical levels increased with diabetes, high leukocyte count, high creatinine and a high degree of contralateral stenosis. Radical levels decreased with high age, blood pressure, collateral circulation as well as operation for left-side carotid artery stenosis. After pretreatment with allopurinol, several of the relationships noted in the control group were eliminated, i.e. leukocyte count, side of operation, Betapred pretreatment and collateral circulation.

Conclusions: Radical levels can be determined in connection with surgery for carotid artery stenosis using an ex vivo spin trap method. With preoperative antioxidant therapy the relationships between enhanced radical levels and clinical data, as seen in control subjects, disappeared. This might indicate a beneficial effect of preoperative pretreatment with antioxidants.     

Regulation of cell motility by tyrosine phosphorylated villin

Temporal and spatial regulation of the actin cytoskeleton is vital for cell migration. Here, we show that an epithelial cell actin-binding protein, villin, plays a crucial role in this process. Overexpression of villin in doxycyline-regulated HeLa cells enhanced cell migration. Villin-induced cell migration was modestly augmented by growth factors. In contrast, tyrosine phosphorylation of villin and villin-induced cell migration was significantly inhibited by the src kinase inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) as well as by overexpression of a dominant negative mutant of c-src. These data suggest that phosphorylation of villin by c-src is involved in the actin cytoskeleton remodeling necessary for cell migration. We have previously shown that villin is tyrosine phosphorylated at four major sites. To further investigate the role of tyrosine phosphorylated villin in cell migration, we used phosphorylation site mutants (tyrosine to phenylalanine or tyrosine to glutamic acid) in HeLa cells. We determined that tyrosine phosphorylation at residues 60, 81, and 256 of human villin played an essential role in cell migration as well as in the reorganization of the actin cytoskeleton. Collectively, these studies define how biophysical events such as cell migration are actuated by biochemical signaling pathways involving tyrosine phosphorylation of actin binding proteins, in this case villin.     

GObar: A Gene Ontology based analysis and visualization tool for gene sets

Background  

Microarray experiments, as well as other genomic analyses, often result in large gene sets containing up to several hundred genes. The biological significance of such sets of genes is, usually, not readily apparent.     

DNA environment of the aerobactin iron uptake system genes in prototypic ColV plasmids.

The aerobactin iron uptake system genes in the prototypic plasmid pColV-K30 are flanked by inverted copies of insertion sequence IS1 and by two distinct replication regions. To address the question of how these flanking regions may facilitate the maintenance and spread of the aerobactin system among the plasmids and chromosomes of enteric species, we investigated the DNA environment of 12 ColV plasmids. We found that the aerobactin system-specific genes are conserved in every plasmid phenotypically positive for the aerobactin system. The upstream IS1 and its overlapping replication region (REPI) are also conserved. This replication region was cloned from several ColV plasmids and found to be functional by transforming these cloned derivatives into a polA bacterial host. In contrast, the downstream flanking region is variable. This includes the downstream copy of IS1 and the downstream replication region (REPII). We infer from these results that sequences in addition to the two flanking copies of IS1, in particular the upstream region including REPI, have been instrumental in the preservation and possible spread of aerobactin genes among ColV plasmids and other members of the FI incompatibility group.     

EDTA chelation therapy does not selectively increase chromium losses

Chelation therapy and supplemental Cr have both been shown to lead to improved blood glucose, lipids, and insulin activity. Chelation therapy leads to the removal of toxic as well as essential metals. To determine if chelation therapy leads to increased urinary Cr losses and altered Cr homeostasis, 2 groups of subjects (1 group that had undergone only 1 or no chelation therapy and 1 group in which all subjects had undergone at least 19 chelation sessions) were evaluated for differences in possible Cr homeostasis based on urinary Cr losses. There were no significant differences in urinary Cr losses between the two groups of subjects and there were no significant increases in urinary Cr losses resulting from chelation therapy. Increases in urinary Cr losses were strongly influenced by supplementation but not chelation therapy.     

Inherited X-chromosome inverted tandem duplication in a male traced to a grandparental mitotic error.

A male infant was referred for cytogenetic evaluation because of dysmorphic features and developmental delay. In both lymphocytes and skin fibroblasts, a modal number of 46 chromosomes was obtained with an obvious elongation of the long arm of the X chromosome (Xq+). Studies of seven members in 3 generations of this family showed that the proband's mother, sister, and maternal grandmother were phenotypically normal carriers of this abnormal X chromosome. High resolution GTG- and RBG-banding defined the extra chromatin material as an inverted duplication of Xq21----Xq24. This was supported by an approximate twofold increase in alpha-galactosidase A activity, localized to Xq21----q24, observed in the proband's lymphocytes and fibroblasts. BrdU-incorporation studies of the mother's lymphocytes showed the abnormal X to be late replicating in all 100 cells studied and normal alpha-galactosidase A levels. Cytogenetic analysis of the maternal grandmother revealed cytogenetic mosaicism with one cell line containing the abnormal X (37%), and the other, a normal female karyotype (63%). This family is instructive since: (1) it represents only the second case of a dysmorphic male demonstrating a confirmed interstitial partial Xq duplication, and (2) the origin of this familial structural rearrangement has been traced to a grandparental mitotic error.     

Spontaneous and photosensitiser-induced DNA single-strand breaks and formamidopyrimidine-DNA glycosylase sensitive sites at nucleotide resolutionin the nuclear and mitochondrial DNA of Saccharomyces cerevisiae.

A system is described for mapping oxidative DNA damage (sites sensitive to formamidopyrimidine-DNA glycosylase and single-strand breaks) at nucleotide resolution in the nuclear and mitochondrial DNA of Saccharomyces cerevisiae. Our 3' end labelling method is sensitive and was first developed using the well-studied inducer of oxidative DNA damage, methylene blue (MB) plus light. We treated yeast DNA in vitro with this so as to maximise levels of damage for assay development. Unfortunately, MB does not remain in yeast cells and yeast DNA repair mutants sensitive to active oxygen species are not sensitive to this agent, thus for in vivo experiments we turned to a polycyclic aromatic, RO 19-8022 (RO). This resulted in oxidative DNA damage when light was applied to yeast cells in its presence. The spectra of enzyme-sensitive sites and single-strand breaks induced by MB in vitro or by RO plus light in vivo or in vitro were examined in two yeast reporter genes: the nuclear MFA2 and the mitochondrial OLI1. The experiments revealed that most of the enzyme-sensitive sites and single-strand breaks induced by MB or RO plus light are at the same positions in these sequences, and that these are guanines.     

Effect of Nitrofurans and Chlortetracycline on Microorganisms Associated with Shrimp

Nitrofuran AF-2 displayed greater inhibitory effect than did nitrofuran Z when a mixed bacterial culture, including several proteolytic bacteria, isolated from shrimp was subjected to these compounds in vitro. Nitrofuran Z exhibited greater bactericidal properties than did chlortetracycline in all cultures used. Only 10 mug of nitrofuran AF-2 per ml was sufficient to inhibit the growth of mixed bacteria in nutrient broth, whereas 50 mug of nitrofuran Z per ml was necessary to accomplish the same inhibition. A 50-mug amount of chlortetracycline per ml displayed about the same inhibitory effect as either 10 mug of AF-2 per ml or 20 mug of Z per ml. The isolated proteolytic bacteria showed greater suppression of growth when subjected to AF-2 than when subjected to Z; however, both nitrofurans were effective in preventing growth. The addition of either 1 mug of AF-2 per ml or 5 mug of Z per ml to nutrient broth inhibited the growth of Achromobacter aquarmarinus, whereas chlortetracycline was less effective, requiring about 20 mug to suppress growth to the same degree.     

Strategies for the use of computational SAR methods in assessing genotoxicity

The relationship between computational SAR studies and relevant data gathering and generation activities is complex. First, the chemical class to be studied is selected on the basis of information requirements for hazard identification and assessment. Membership in the class is determined by consideration of chemical structure and reactivity. Compilation of the existing bioassay data for this chemical class follows immediately from the specification of the class. Bioassay data, qualitative knowledge of general chemical reactivities in this class, and knowledge concerning potential interactions with biomolecular targets all contribute to the derivation of possible mechanisms for biological activity. Computational studies based on modeling the proposed mechanism of action and/or the existing data base can provide a quantitative basis for the differentiation between chemicals. There is the opportunity for continuing feedback between the quantitative computational studies and the development of a relevant bioassay data base for this chemical class. The qualitative and quantitative information on the potential biological responses obtained will provide a rational basis for extrapolation from the extant data base to the chemicals of interest, and to biological responses significant to the assessment for which complete data are unavailable. Knowledge concerning possible mechanisms of action and preexisting data determine the type of computational study that will be most useful.