Comparative molecular field analysis of colchicine inhibition and tubulin polymerization for combretastatins binding to the colchicine binding site on beta-tubulin

A molecular modeling study using Comparative Molecular Field Analysis (CoMFA) was undertaken to develop a predictive model for combretastatin binding to the colchicine binding site of tubulin. Furthermore, we examined the potential contribution of lipophilicity (log P) and molecular dipole moment and were unable to correlate these properties to the observed biological data. In this study we first confirmed that tubulin polymerization inhibition (IC50) correlated (R2 = 0.92) with [3H]colchicine displacement. Although these data correlated quite well, we developed two independent models for each set of data to quantify structural features that may contribute to each biological property independently. To develop our predictive model we first examined a series of molecular alignments for the training set and ultimately found that overlaying the respective trimethoxyphenyl rings (A ring) of the analogues generated the best correlated model. The CoMFA yielded a cross-validated R2 = 0.41 (optimum number of components equal to 5) for the tubulin polymerization model and an R2 = 0.38 (optimum number of components equal to 5) for [3H]colchicine inhibition. Final non-cross-validation generated models for tubulin polymerization (R2 of 0.93) and colchicine inhibition (R2 of 0.91). These models were validated by predicting both biological properties for compounds not used in the training set. These models accurately predicted the IC50 for tubulin polymerization with an R2 of 0.88 (n = 6) and those of [3H]colchicine displacement with an R2 of 0.80 (n = 7). This study represents the first predictive model for the colchicine binding site over a wide range of combretastatin analogues.     

Spatial distribution and differentiation potential of stem cells in hatchlings and adults in the marine platyhelminth macrostomum sp.: a bromodeoxyuridine analysis

Stem cells (neoblasts) in Platyhelminthes are pluripotent, and likely totipotent, undifferentiated cells which retain throughout adult life the capacity to proliferate and from which all somatic cells as well as the germ cells derive. However, basic data on the pool and heterogeneity of neoblasts, their rates of differentiation into sets and subsets of differentiated cells, and their migration to different body regions are still lacking. To fill this gap, S-phase cells in the macrostomid Macrostomum sp. were labeled with the thymidine analog 5-bromo-2'-deoxyuridine (BrdU). S-phase cells were found to be neoblasts and to be distributed in two bands along the lateral sides of the body leaving unlabeled the median axis of the body and the region anterior to the eyes. This distribution is parallel to that of mitotic cells demonstrated using an antibody to phosphorylated histone H3. At different chase times, clusters of BrdU-labeled cells appear, labeled cells migrate to formerly unlabeled areas, and they differentiate into several somatic cell types and into germ cells. Finally, continuous exposure to BrdU shows an extensive renewal of the epithelial cells. Altogether, these results strengthen the idea of platyhelminth neoblasts as an unparalleled stem-cell system within the Animal Kingdom calling for further investigation.     

Microvideography of gas bladder inflation in larval walleye

High-resolution microvideograph observations supported the hypothesis that first filling of the gas bladder of larval walleye Stizostedion vitreum is accomplished by the fish penetrating the air-water interface to gulp air, then transmitting a swallowed air bubble through the gut and the pneumatic duct to the gas bladder. The snout of the larva penetrated the water surface with extension of most of the mouth into the air for only 1.5 s. An air bubble ( c. 100 μm) in the foregut was broken up into progressively smaller bubbles (10–15 μm), presumably by the combined effect of surfactant derived from the gall bladder and mechanical (peristaltic) action of the gut. These smaller bubbles seemed to be aligned in the pneumatic duct before being forced to the gas bladder by pressures generated from peristalsis.     

Phylogenetic utility of the nuclear gene arginine decarboxylase: an example from Brassicaceae

Arginine decarboxylase (ADC) is an important enzyme in the production of putrescine and polyamines in plants. It is encoded by a single or low-copy nuclear gene that lacks introns in sequences studied to date. The rate of Adc amino acid sequence evolution is similar to that of ndhF for the angiosperm family studied. Highly conserved regions provide several target sites for PCR priming and sequencing and aid in nucleotide and amino acid sequence alignment across a range of taxonomic levels, while a variable region provides an increased number of potentially informative characters relative to ndhF for the taxa surveyed. The utility of the Adc gene in plant molecular systematic studies is demonstrated by analysis of its partial nucleotide sequences obtained from 13 representatives of Brassicaceae and 3 outgroup taxa, 2 from the mustard oil clade (order Capparales) and 1 from the related order Malvales. Two copies of the Adc gene, Adc1 and Adc2, are found in all members of the Brassicaceae studied to data except the basal genus Aethionema. The resulting Adc gene tree provides robust phylogenetic data regarding relationships within the complex mustard family, as well as independent support for proposed tribal realignments based on other molecular data sets such as those from chloroplast DNA.      

Epidemic Spread of the Rust Fungus Puccinia lagenophorae and its Impact on the Competitive Ability of Senecio vulgaris in Celeriac During Early Development

The 'system management approach' of biological weed control was applied in a small-scale field experiment with celeriac (root celery), intersown with an inbred line of the annual weed Senecio vulgaris L. The naturalized rust fungus Puccinia lagenophorae Cooke (Basidiomycetes: Uredinales), a common and widespread pathogen of S. vulgaris in Europe, was introduced into parts of the plots, and its impact on the competitive balance between the crop and weed in the presence and absence of an additional herbicide treatment was studied. Competition by S. vulgaris (at a realized density of only 50 plants/m2) during the first 10 weeks of growth was substantial, reducing the fresh weight of the celeriac bulbs by 28%. The epidemic spread of the rust fungus was relatively fast, and the time to infection was similar to that in full-area applications. Inoculation with the rust fungus strongly reduced crop losses due to competition with S. vulgaris . The fresh weight of the celeriac bulbs in plots with both S. vulgaris and the fungus was not statistically diVerent from the celeriac yield in plots without S. vulgaris . This eVect was mainly the result of the reduced biomass of S. vulgaris , and not reduced survival. Infected plants may, therefore, still contribute to soil cover and may help to suppress later germinating weed species. Older plant stages were found to be infected earlier than younger stages. No significant interactions were observed between the eVects of the fungal infection and a low-dose application of the herbicide chlorbromuron on weed performance. Basic studies necessary to develop the system management approach further are discussed.     

On the Subjective Acceptance during Cardiovascular Magnetic Resonance Imaging at 7.0 Tesla

Purpose

This study examines the subjective acceptance during UHF-CMR in a cohort of healthy volunteers who underwent a cardiac MR examination at 7.0T.

Methods

Within a period of two-and-a-half years (January 2012 to June 2014) a total of 165 healthy volunteers (41 female, 124 male) without any known history of cardiac disease underwent UHF-CMR. For the assessment of the subjective acceptance a questionnaire was used to examine the participants experience prior, during and after the UHF-CMR examination. For this purpose, subjects were asked to respond to the questionnaire in an exit interview held immediately after the completion of the UHF-CMR examination under supervision of a study nurse to ensure accurate understanding of the questions. All questions were answered with “yes” or “no” including space for additional comments.

Results

Transient muscular contraction was documented in 12.7% of the questionnaires. Muscular contraction was reported to occur only during periods of scanning with the magnetic field gradients being rapidly switched. Dizziness during the study was reported by 12.7% of the subjects. Taste of metal was reported by 10.1% of the study population. Light flashes were reported by 3.6% of the entire cohort. 13% of the subjects reported side effects/observations which were not explicitly listed in the questionnaire but covered by the question about other side effects. No severe side effects as vomiting or syncope after scanning occurred. No increase in heart rate was observed during the UHF-CMR exam versus the baseline clinical examination.

Conclusions

This study adds to the literature by detailing the subjective acceptance of cardiovascular magnetic resonance imaging examinations at a magnetic field strength of 7.0T. Cardiac MR examinations at 7.0T are well tolerated by healthy subjects. Broader observational and multi-center studies including patient cohorts with cardiac diseases are required to gain further insights into the subjective acceptance of UHF-CMR examinations.     

Association of CMV-Specific T Cell-Mediated Immunity with CMV DNAemia and Development of CMV Disease in HIV-1–Infected Individuals

Background

Among HIV-1–infected individuals, cytomegalovirus (CMV) reactivation and disease occur in the setting of advanced immunosuppression. The value of a standardized assessment of CMV-specific T-cell mediated immunity by the CMV QuantiFERON assay (CMV-QFT) has not yet been thoroughly investigated in HIV-1–infected subjects.

Methods

Prospective, longitudinal study in 153 HIV-1–infected subjects with a CD4⁺ T cell count < 350/μL who simultaneously underwent CMV-QFT, CMV serology testing and CMV-DNA quantification. Factors associated with CMV-QFT were evaluated. Clinical screening for CMV manifestations was then performed every 3 months.

Results

Among the 141 CMV IgG-seropositive individuals the CMV-QFT assay yielded reactive results in 84% (118/141), negative results in 15% (21/141) and indeterminate (negative mitogen IFN-gamma response) results in 1% (2/141) of subjects. The mean actual CD4⁺ T cell count was significantly higher in CMV-QFT reactive subjects, when compared to CMV-QFT non-reactive individuals (183 ± 102 vs. 126 ± 104 cells/μL, P = 0.015). A significantly lower proportion of CMV-QFT reactive vs. non-reactive patients displayed CMV DNAemia > 100 copies/mL (23% (27/118) vs. 48% (11/23), P = 0.02). Furthermore, a statistically significant inverse association between mitogen IFN-gamma response and CMV-DNAemia > 1000 copies/mL was observed (P < 0.001). During the observational period, 5 CMV end-organ manifestations were observed. In three of the CMV cases the CMV-QFT yielded indeterminate results.

Conclusions

While CMV-QFT reactivity indicates CMV-specific immunity, indeterminate results due to negative mitogen IFN-gamma response might reflect HIV-1-induced immunodeficiency. Thus, dependency upon CD4⁺ T cell count should be considered when interpreting CMV-QFT results.     

International External Quality Assessment Study for Molecular Detection of Lassa Virus

Lassa virus (LASV) is a causative agent of hemorrhagic fever in West Africa. In recent years, it has been imported several times to Europe and North America. The method of choice for early detection of LASV in blood is RT-PCR. Therefore, the European Network for Diagnostics of ‘Imported’ Viral Diseases (ENIVD) performed an external quality assessment (EQA) study for molecular detection of LASV. A proficiency panel of 13 samples containing various concentrations of inactivated LASV strains Josiah, Lib-1580/121, CSF, or AV was prepared. Samples containing the LASV-related lymphocytic choriomeningitis virus (LCMV) and negative sera were included as specificity controls. Twenty-four laboratories from 17 countries (13 European, one African, one Asian, two American countries) participated in the study. Thirteen laboratories (54%) reported correct results, 4 (17%) laboratories reported 1 to 2 false-negative results, and 7 (29%) laboratories reported 3 to 5 false-negative results. This EQA study indicates that most participating laboratories have a good or acceptable performance in molecular detection of LASV. However, several laboratories need to review and improve their diagnostic procedures.