Impact of Mistletoe Triterpene Acids on the Uptake of Mistletoe Lectin by Cultured Tumor Cells

Complementary treatment possibilities for the therapy of cancer are increasing in demand due to the severe side effects of the standard cytostatics used in the first-line therapy. A common approach as a complementary treatment is the use of aqueous extracts of Viscum album L. (Santalaceace). The therapeutic activity of these extracts is attributed to Mistletoe lectins which are Ribosome-inactivating proteins type II. Besides these main constituents the extract of Viscum album L. comprises also a mixture of lipophilic ingredients like triterpene acids of the oleanane, lupane and ursane type. However, these constituents are not contained in commercially available aqueous extracts due to their high lipophilicity and insolubility in aqueous extraction media. To understand the impact of the extract ingredients in cancer therapy, the intracellular uptake of the mistletoe lectin I (ML) by cultured tumor cells was investigated in relation to the mistletoe triterpene acids, mainly oleanolic acid. Firstly, these hydrophobic triterpene acids were solubilized using cyclodextrins ("TT" extract). Afterwards, the uptake of either single compounds (isolated ML and the aqueous "viscum" extract) or in combination with the TT extract (ML+TT, viscumTT), was analyzed. The uptake of ML was studied inTHP-1-, HL-60-, 143B- and Ewing TC-71-cells and determined after 30, 60 and 120 minutes by an enzyme linked immunosorbent assay which quantifies the A-chain of the hololectin. It could be shown that the intracellular uptake after 120 minutes amounted to 20 % in all cell lines after incubation with viscumTT. The studies further revealed that the uptake in THP-1-, HL-60- and Ewing TC-71-cells was independent of the addition of TT extract. Interestingly, the uptake of ML by 143B-cells could only be measured after addition of triterpenes pointing to resistance to mistletoe lectin

Bridging from syntactic to statistical methods: Classification with automatically segmented features from sequences

To Integrate The Benefits Of Statistical Methods Into Syntactic Pattern Recognition, A Bridging Approach Is Proposed: (I) Acquisition Of A Grammar Per Recognition Class (Ii) Comparison Of The Obtained Grammars In Order To Find Substructures Of Interest Represented As Sequences Of Terminal And/Or Non-Terminal Symbols And Filling The Feature Vector With Their Counts (Iii) Hierarchical Feature Selection And Hierarchical Classification, Deducing And Accounting For The Domain Taxonomy. The Bridging Approach Has The Benefits Of Syntactic Methods: Preserves Structural Relations And Gives Insights Into The Problem. Yet, It Does Not Imply Distance Calculations And, Thus, Saves A Non-Trivial Task-Dependent Design Step. Instead It Relies On Statistical Classification From Many Features. Our Experiments Concern A Difficult Problem Of Chemical Toxicity Prediction. The Code And The Data Set Are Open-Source. (C) 2015 Elsevier Ltd. All Rights Reserved

Estimation of the in vivo recombination rate for a plant RNA virus

[EN] Phylogenomic evidence suggested that recombination is an important evolutionary force for potyviruses, one of the larger families of plant RNA viruses. However, mixed-genotype potyvirus infections are marked by low levels of cellular coinfection, precluding template switching and recombination events between virus genotypes during genomic RNA replication. To reconcile these conflicting observations, we evaluated the in vivo recombination rate (r(g)) of Tobacco etch virus (TEV; genus Potyvirus, family Potyviridae) by coinfecting plants with pairs of genotypes marked with engineered restriction sites as neutral markers. The recombination rate was then estimated using two different approaches: (i) a classical approach that assumed recombination between marked genotypes can occur in the whole virus population, rendering an estimate of r(g)=7.762x10(-8) recombination events per nucleotide site per generation, and (ii) an alternative method that assumed recombination between marked genotypes can occur only in coinfected cells, rendering a much higher estimate of r(g)=3.427x10(-8) recombination events per nucleotide site per generation. This last estimate is similar to the TEV mutation rate, suggesting that recombination should be at least as important as point mutation in creating variability. Finally, we compared our mutation and recombination rate estimates to those reported for animal RNA viruses. Our analysis suggested that high recombination rates may be an unavoidable consequence of selection for fast replication at the cost of low fidelity.We thank Francisca de la Iglesia and Angels Prosper for excellent technical assistance, Jose A. Dare's for methodological advice, Jose M. Cuevas for critical reading of the manuscript, and other lab members for helpful discussions. This work was supported by the Spanish Secretaria de Estado de Investigacion, Desarrollo e Innovacion (grants BFU2009-06993 and BFU2012-30805). N. T. was supported,by a pre-doctoral fellowship from the former Spanish Ministerio de Ciencia e Innovacion.Tromas, N.; Zwart, MP.; Poulain, M.; Elena Fito, SF. (2014). Estimation of the in vivo recombination rate for a plant RNA virus. Journal of General Virology. 95:724-732. https://doi.org/10.1099/vir.0.060822-0S7247329

Langmuir monolayers composed of single and double tail sulfobetaine lipids

AbstractHypothesisOwing to structural similarities between sulfobetaine lipids and phospholipids it should be possible to form stable Langmuir monolayers from long tail sulfobetaines. By modification of the density of lipid tail group (number of carbon chains) it should also be possible to modulate the two-dimensional phase behaviour of these lipids and thereby compare with that of equivalent phospholipids. Potentially this could enable the use of such lipids for the wide array of applications that currently use phospholipids. The benefit of using sulfobetaine lipids is that they can be synthesised by a one-step reaction from cheap and readily available starting materials and will degrade via different pathways than natural lipids. The molecular architecture of the lipid can be easily modified allowing the design of lipids for specific purposes. In addition the reversal of the charge within the sulfobetaine head group relative to the charge orientation in phospholipids may modify behaviour and thereby allow for novel uses of these surfactants.ExperimentsStable Langmuir monolayers were formed composed of single and double tailed sulfobetaine lipids. Surface pressure-area isotherm, Brewster Angle Microscopy and X-ray and neutron reflectometry measurements were conducted to measure the two-dimensional phase behaviour and out-of-plane structure of the monolayers as a function of molecular area.FindingsSulfobetaine lipids are able to form stable Langmuir monolayers with two dimensional phase behaviour analogous to that seen for the well-studied phospholipids. Changing the number of carbon tail groups on the lipid from one to two promotes the existence of a liquid condensed phase due to increased Van der Waals interactions between the tail groups. Thus the structure of the monolayers appears to be defined by the relative sizes of the head and tail groups in a predictable way. However, the presence of sub-phase ions has little effect on the monolayer structure, behaviour that is surprisingly different to that seen for phospholipids

A universal BMV-based RNA recombination system—how to search for general rules in RNA recombination

At present, there is no doubt that RNA recombination is one of the major factors responsible for the generation of new RNA viruses and retroviruses. Numerous experimental systems have been created to investigate this complex phenomenon. Consequently, specific RNA structural motifs mediating recombination have been identified in several viruses. Unfortunately, up till now a unified model of genetic RNA recombination has not been formulated, mainly due to difficulties with the direct comparison of data obtained for different RNA-based viruses. To solve this problem, we have attempted to construct a universal system in which the recombination activity of various RNA sequences could be tested. To this end, we have used brome mosaic virus, a model (+)RNA virus of plants, for which the structural requirements of RNA recombination are well defined. The effectiveness of the new homomolecular system has been proven in an experiment involving two RNA sequences derived from the hepatitis C virus genome. In addition, comparison of the data obtained with the homomolecular system with those generated earlier using the heteromolecular one has provided new evidence that the mechanisms of homologous and non-homologous recombination are different and depend on the virus' mode of replication

Diversity of Plant Virus Populations: A Valuable Tool for Epidemiological Studies

Plant viruses, as any other living organisms, differ genetically from each other as a result of processes (such as mutation, recombination and other forms of genetic exchange) that generate genetic variation in each generation during their reproduction and processes (such as selection, migration and genetic drift) that modulate this variation, determine the distribution of the genetic variants within a population (i.e., the genetic structure of the population) and how it changes with time, in a dynamical phenomenon called evolution. For plant viruses, evolutionary forces that generate and modulate the genetic diversity of their populations are often associated to different phases in their biology and ecology, such as virus-host interactions and host to host transmission. Forces that shape the evolution of plant viruses are at the same time key factors affecting their pathogenic properties, including their ability to cause diseases (an aspect that is studied in the field of epidemiology). The present chapter aims to illustrate how measurement and analysis of genetic diversity and structure of plant virus populations are essential to the current knowledge on the evolutionary biology of plant viruses and how evolutionary factors have a relevant role in the dynamics of virus populations and therefore, in the epidemiology of plant virus diseases