Assessing the significance of conserved genomic aberrations using high resolution genomic microarrays

Genomic aberrations recurrent in a particular cancer type can be important prognostic markers for tumor progression. Typically in early tumorigenesis, cells incur a breakdown of the DNA replication machinery that results in an accumulation of genomic aberrations in the form of duplications, deletions, translocations, and other genomic alterations. Microarray methods allow for finer mapping of these aberrations than has previously been possible; however, data processing and analysis methods have not taken full advantage of this higher resolution. Attention has primarily been given to analysis on the single sample level, where multiple adjacent probes are necessarily used as replicates for the local region containing their target sequences. However, regions of concordant aberration can be short enough to be detected by only one, or very few, array elements. We describe a method called Multiple Sample Analysis for assessing the significance of concordant genomic aberrations across multiple experiments that does not require a-priori definition of aberration calls for each sample. If there are multiple samples, representing a class, then by exploiting the replication across samples our method can detect concordant aberrations at much higher resolution than can be derived from current single sample approaches. Additionally, this method provides a meaningful approach to addressing population-based questions such as determining important regions for a cancer subtype of interest or determining regions of copy number variation in a population. Multiple Sample Analysis also provides single sample aberration calls in the locations of significant concordance, producing high resolution calls per sample, in concordant regions. The approach is demonstrated on a dataset representing a challenging but important resource: breast tumors that have been formalin-fixed, paraffin-embedded, archived, and subsequently UV-laser capture microdissected and hybridized to two-channel BAC arrays using an amplification protocol. We demonstrate the accurate detection on simulated data, and on real datasets involving known regions of aberration within subtypes of breast cancer at a resolution consistent with that of the array. Similarly, we apply our method to previously published datasets, including a 250K SNP array, and verify known results as well as detect novel regions of concordant aberration. The algorithm has been fully implemented and tested and is freely available as a Java application at http://www.cbil.upenn.edu/MSA.     

Application of the salt stress to the protoplast cultures of the carrot (Daucus carota L.) and evaluation of the response of regenerants to soil salinity

This is the first study to generate carrot plants for enhanced salinity tolerance using a single-cell in vitro system. Protoplasts of three carrot accessions were exposed to treatment by seven different concentrations of NaCl (10–400 mM). Salt concentrations higher than 50 mM decreased plating efficiency and those of 200–400 mM of NaCl completely arrested mitotic divisions of cultured cells. The protoplast-derived plants from the control and 50–100 mM NaCl treatment were subjected to an 8-week salt stress in greenhouse conditions induced by salinized soil (EC 3 and 6 mS cm^?1). 50 mM NaCl stress applied in vitro induced polyploidy among regenerated plants. The regenerants obtained from the 50 and 100 mM NaCl-treated protoplast cultures grown in saline soil had a higher survival rate compared to the regenerants from the control cultures. The salt-stressed plants accumulated anthocyanins in petioles and produced denser hairs on leaves and petioles in comparison to the control plants. Salt stress influenced pollen viability and seed setting of obtained regenerants. The results suggest that salt stress applied in vitro in protoplast cultures creates variation which allows alleviating the negative effects of salt stress on the development and reproduction of the carrot.

     

Echiniscus testudo (Doyère, 1840) in New Zealand: anthropogenic dispersal or evidence for the ‘Everything is Everywhere’ hypothesis?

Many species of microscopic invertebrates, such as tardigrades or rotifers, have been traditionally regarded as cosmopolitan. This conviction was based on classical taxonomic observations of very similar phenotypes across the globe and hypothetical abilities of micrometazoans for large-scale dispersal with air and water currents. However, several recent studies showed that micrometazoan species may have restricted geographic ranges, suggesting that microscopic size is not the only determinant of dispersal. Here, we describe the first Australasian record of Echiniscus testudo, a heterotardigrade originally described from the Holarctic. Morphological and genetic analyses gave congruent conclusions and confirmed that European and New Zealand populations represent a single species. Importantly, however, without broader sampling in primeval localities, it is not possible to test whether the New Zealand record of E. testudo is a result of natural dispersal or whether the species was brought to Australasia by humans.     

Secreting oviduct epithelial cells of Coturnix coturnix japonica (QOEC) and changes to their proteome after nonviral transfection

The quail oviduct (Coturnix c. japonica) is a natural candidate avian bioreactor, while the secretive quail oviduct epithelial cells (QOECs) are potential in vitro producers of recombinant proteins and vaccines. In view of the need for highly performing and transformable cell lines, QOEC may potentially act as an alternative bioreactor platform to the existing ones, for example, to the Chinese hamster ovary. The aim of this work was to characterize QOECs and their response to nucleofection with a nonviral plasmid DNA carrying the human interferon-α 2a gene (hIFNλ2a), in vitro. Primary QOEC cultures from laying quails (10-15 weeks old) were characterized by their proliferation rate, doubling time, and multilineage differentiation. Electroporation to cell nuclei (nucleofection) was used to deliver nonviral plasmid DNA containing a reporter GFP and hIFN under the ovalbumin promoter. The posttransfection analysis included polymerase chain reaction, Western blot analysis, and liquid chromatography coupled to tandem mass spectrometry. QOEC showed a typical epithelial characteristic in a primary 2D monolayer culture system and retained secretive potential up to the first passage. QOEC showed differentiation into osteoblastic lineage after stimulation. The nucleofection mean efficiency was low (2.3%). Differences of up to 10% in the proteomic profiles between nontransfected and transfected QOEC were found, the most important of these were related to the absence of keratins and cell-adhesion proteins in the transfected QOEC. Concluding, with the practical information provided here, QOEC have the potential to serve as an avian secreting cellular platform. QOEC may be further transformed to cell lineage to meet the requirement for a stable, electrocompetent, and transfectable model. The first proteomic comparison of QOEC delivered in this study showed, in the majority, a stable proteome of the nontransfected vs transfected QOEC.     

Inadequate function of sterile tw5/tw32 spermatozoa overcome by intracytoplasmic sperm injection

Mice carrying two t complementary haplotypes (t^w5/t^w32) are totally sterile. Their spermatozoa have poor motility and fertilize neither zone-intact nor zona-free oocytes, even though they are structurally indistinguishable from control (wild-type) spermatozoa. However, when injected directly into oocytes, these infertile spermatozoa are able to participate in normal development. This suggests that infertility of t^w5/t^w32 male (spermatozoa) is more likely to be due to poor sperm-oocyte interaction than to genetic incompetence of sperm nuclei. © 1996 Wiley-Liss, Inc.     

Flavonoids as tyrosinase inhibitors in in silico and in vitro models: basic framework of SAR using a statistical modelling approach

Flavonoids are widely distributed in plants and constitute the most common polyphenolic phytoconstituents in the human diet. In this study, the in vitro inhibitory activity of 44 different flavonoids (1–44) against mushroom tyrosinase was studied, and an in silico study and type of inhibition for the most active compounds were evaluated too. Tyrosinase inhibitors block melanogenesis and take part in melanin production or distribution leading to pigmentation diseases. The in vitro study showed that quercetin was a competitive inhibitor (IC₅₀=44.38 ± 0.13 µM) and achieved higher antityrosinase activity than the control inhibitor kojic acid. The in silico results highlight the importance of the flavonoid core with a hydroxyl at C7 as a strong contributor of interference with tyrosinase activity. According to the developed statistical model, the activity of molecules depends on hydroxylation at C3 and methylation at C8, C7, and C3 in the benzo-γ-pyrane ring of the flavonoids.     

The race concept in six regions: variation without consensus

Race, once the central concept in physical anthropology worldwide, now varies in the degree of support it receives in different regions. We present the currently available information on the status of the concept in the United States, the Spanish language areas, Poland, Europe, Russia, and China. Rejection of race ranges from high to low with the highest rejection occurring among anthropologists in the United States (and Canada). Rejection of race is moderate in Europe, sizeable in Poland and Cuba, and lowest in Russia and China. A discussion on the scientific and contextual reasons influencing these variations is presented. The tension between scientific evidence and social influences varies from region to region. The methods used in the studies reported here included questionnaires and content analysis. Response rates to questionnaires were often around 50 percent (with exception of the Polish studies). We discuss reasons for the low rates. Although a uniform method of data gathering is desirable, it may not suit scientists working in different traditions of theory and research. We conclude that it is once again timely to discuss the race concept in international meetings where all scientific and political changes occurring throughout the world in recent past decades are taken into account.