The in vitro unscheduled DNA synthesis (UDS) assay in rat primary hepatocytes: evaluation of 2-furoic acid and 7 drug candidates

The in vitro unscheduled DNA synthesis assay (UDS) is part of the routine genetic toxicology screening at The Upjohn Company. The purpose of this paper is to report results for 8 compounds which were tested in the in-house genetic toxicology program. These compounds represent diverse chemical structure and most of them entered the screening program because they are biologically active in efficacy screens. All tests were carried out under Good Laboratory Practices Regulations of the U.S. Food and Drug Administration. None of the materials reported here produced an increase in UDS and therefore the UDS results with these compounds do not suggest potential for genotoxicity.     

Effect of cimetidine on the heart conduction system]

An effect of cimetidine on ECG records has been investigated in a group of 40 patients with gastric or duodenal ulcers and coexisting circulatory disorders. For this purpose ECG has been recorded with Holter's technique (Medilog 2000) together with ECG-recording using high amplifying technique. An intravenous injection of 200 mg of cimetidine produced in some of patients inhibition of the sinus rhythm atrio-ventricular conduction disturbances as well as changes in the end phase of ECG ventricular image. The authors suggest, that intravenous administration of cimetidine to patients with cardiac diseases should be monitored with ECG recording.     

Extracting protein regulatory networks with graphical models

During the last decade the development of high-throughput biotechnologies has resulted in the production of exponentially expanding quantities of biological data, such as genomic and proteomic expression data. One fundamental problem in systems biology is to learn the architecture of biochemical pathways and regulatory networks in an inferential way from such postgenomic data. Along with the increasing amount of available data, a lot of novel statistical methods have been developed and proposed in the literature. This article gives a non-mathematical overview of three widely used reverse engineering methods, namely relevance networks, graphical Gaussian models, and Bayesian networks, whereby the focus is on their relative merits and shortcomings. In addition the reverse engineering results of these graphical methods on cytometric protein data from the RAF-signalling network are cross-compared via AUROC scatter plots.     

Statistics for proteomics: a review of tools for analyzing experimental data

Most proteomics experiments make use of 'high throughput' technologies such as 2-DE, MS or protein arrays to measure simultaneously the expression levels of thousands of proteins. Such experiments yield large, high-dimensional data sets which usually reflect not only the biological but also technical and experimental factors. Statistical tools are essential for evaluating these data and preventing false conclusions. Here, an overview is given of some typical statistical tools for proteomics experiments. In particular, we present methods for data preprocessing (e.g. calibration, missing values estimation and outlier detection), comparison of protein expression in different groups (e.g. detection of differentially expressed proteins or classification of new observations) as well as the detection of dependencies between proteins (e.g. protein clusters or networks). We also discuss questions of sample size planning for some of these methods.     

Comparative evaluation of reverse engineering gene regulatory networks with relevance networks, graphical gaussian models and bayesian networks

MOTIVATION: An important problem in systems biology is the inference of biochemical pathways and regulatory networks from postgenomic data. Various reverse engineering methods have been proposed in the literature, and it is important to understand their relative merits and shortcomings. In the present paper, we compare the accuracy of reconstructing gene regulatory networks with three different modelling and inference paradigms: (1) Relevance networks (RNs): pairwise association scores independent of the remaining network; (2) graphical Gaussian models (GGMs): undirected graphical models with constraint-based inference, and (3) Bayesian networks (BNs): directed graphical models with score-based inference. The evaluation is carried out on the Raf pathway, a cellular signalling network describing the interaction of 11 phosphorylated proteins and phospholipids in human immune system cells. We use both laboratory data from cytometry experiments as well as data simulated from the gold-standard network. We also compare passive observations with active interventions. RESULTS: On Gaussian observational data, BNs and GGMs were found to outperform RNs. The difference in performance was not significant for the non-linear simulated data and the cytoflow data, though. Also, we did not observe a significant difference between BNs and GGMs on observational data in general. However, for interventional data, BNs outperform GGMs and RNs, especially when taking the edge directions rather than just the skeletons of the graphs into account. This suggests that the higher computational costs of inference with BNs over GGMs and RNs are not justified when using only passive observations, but that active interventions in the form of gene knockouts and over-expressions are required to exploit the full potential of BNs. AVAILABILITY: Data, software and supplementary material are available from http://www.bioss.sari.ac.uk/staff/adriano/research.html     

Adaptation of the phosphotungstate method to determine reduced and oxidized vitamin C in blood plasma

The phosphotungstate reagent (PTR) was used for quantitative spectrophotometric determination of physiological forms of vitamin C in blood plasma. An immediate action of PTR on the first half of the tested samples allowed to determine reduced vitamin C concentrations (I) at 700 nm. 10 mM dithiothreitol added to the second half of the samples reduced oxidized vitamin C in it--hence the total amount of this vitamin was reduced with a concentration (II) determined as above (remains of dithiothreitol were removed with N-ethylmaleimide). The difference of results (II) and (I) gave the concentration of oxidized vitamin C. The method is characterised by fault-less analytical parameters: correlation coefficients of analytical curves > 0.99, recovery factor 100.5%, variation coefficients intra- and inter-serial < 3% and < 5%, respectively, detection limit 0.05 microM. The simplicity of the method enables an easy control of the ratio of oxidized and reduced vitamin C concentrations in blood plasma--the biomarker of the level of oxidative damage to cells.     

Mineral Content of the Pulp and Peel of Various Citrus Fruit Cultivars

Biological Trace Element Research - The aim of the study was to compare the mineral content between the peel and the pulp of citrus fruits and to determine which citrus fruit, among orange (Citrus...     

Synthesis and Antibacterial Evaluation of Mannich Bases Derived from 1,2,4‐Triazole

The series of novel Mannich bases were synthesized and evaluated for their in vitro antibacterial activity against Gram‐positive and Gram‐negative bacterial strains. The results showed that all compounds were less active than the drugs used as reference, but some of them had moderate potency against Staphylococcus epidermidis ATCC 12228 and Bacillus subtilis ATCC 6633. The presence of a phenyl ring in the position 4 of piperazine seems to be necessary for antibacterial activity in this class of compounds.     

Strain differences in in vitro rat hepatocyte unscheduled DNA synthesis (UDS): effect of UV is independent of strain while increased sensitivity is apparent using Fischer-344 instead of Sprague-Dawley rats

The unscheduled DNA synthesis (UDS) assay measures DNA repair following in vitro treatment of rat primary hepatocytes. This report compares the UDS response of primary hepatocytes from 2 widely used rat strains, the Fischer-344 (F344) and Sprague-Dawley (SD) strains. Ultraviolet (UV) light and 5 known genotoxic chemicals were evaluated in each strain in parallel experiments. The chemicals tested were 2-acetylaminofluorene (2-AAF), 4-aminobiphenyl (4-AB), benzidine, dimethylnitrosamine (DMN) and N-propyl-N'-nitro-N-nitrosoguanidine (PNNG). Four of these compounds (2-AAF, 4-AB, benzidine and DMN) require metabolic activation. Benzidine and PNNG were both negative using SD rat hepatocytes, but were weakly positive using F344 rat hepatocytes. In the first of 2 experiments, 4-AB was inconclusive in SD hepatocytes, but strongly positive in F344 cells. In the second experiment, 4-AB was positive in hepatocytes from both strains. 2-AAF was more strongly positive in F344 cells than in SD cells. DMN and UV light induced positive dose responses with little or no differences between strains. It is concluded that hepatocytes from F344 rats may be more sensitive, qualitatively and quantitatively, than hepatocytes from SD rats as indicators of UDS. This difference is not due to intrinsic differences in DNA repair mechanisms but is probably due to differences in drug-metabolizing enzymes between these strains. Thus, for routine screening, F344 rats are preferable for measurement of the in vitro UDS-inducing potential of compounds.