Correlation between loss of pro-chiral hydrogen and E,Z geometry in isoprenoid biosynthesis

Biosynthesis of several mono- and sesqui-terpenes that possess E or Z double bonds, or which are generally considered to be derived from precursors possessing such geometries, involved loss of the pro-4S hydrogen of mevalonate in the construction of the double bond. These results confirm and extend previous observations. A recent claim to have newly discovered such a stereochemical correlation is rejected.     

Dielectric Properties of the Plasma Membrane of Cultured Murine Fibroblasts Treated with a Nonterpenoid Extract of Azadirachta indica Seeds

Neem oil is a natural product obtained from the seeds of the tree Azadirachta indica. In this report, we investigate the alterations of the biophysical properties of the plasma membrane caused by treatment with the nonterpenoid fraction of neem oil that we defined as methanolic extract (MEX). The dose-response effect was evaluated and a MEX-dependent cytoxicity evidenced. The effect of MEX on the plasma membrane was studied by a well-established dielectric spectroscopy technique: electrorotation, which allows single-cell analysis. Our results show a structural/functional alteration of the plasma membrane with an evident increase of specific capacitance and conductance. The biological implications of this effect are discussed.     

Interactions of chlorpromazine with phospholipid monolayers: effects of the ionization state of the drug

Molecular dynamics simulations have been performed to investigate the interactions between chlorpromazine (CPZ) and Langmuir monolayers of the zwitterionic dipalmitoylphosphatidylcholine (DPPC) and the anionic dipalmitoylphosphatidylglycerol (DPPG). Simulations for a fixed surface density and different charge states - neutral and protonated CPZ - were able to capture important features of the CPZ-phospholipid monolayer interaction. Neutral CPZ is predominantly found in the hydrophobic tail region, whereas protonated CPZ is located at the lipid-water interface. Specific interactions (hydrogen bonds) between protonated CPZ and the lipid head groups were found for both zwitterionic and anionic monolayers. We computed lipid tail order parameters and investigated the effects of the drug upon tail ordering. We also computed electrostatic surface potentials and found qualitative good agreement with experimental results.     

Insecticidal effects of Flourensia oolepis Blake (Asteraceae) essential oil

Flourensia oolepis Blake (Asteraceae) essential oil had a complex chemical composition with τ-muurolene (6.14%), santolinetriene (6.22%), 2-methylene-4,8,8-trimethyl-4-vinyl-bicyclo[5.2.0]nonane (10.15%), δ-cadinene (10.27%) and γ-gurjunene (20.69%) comprising more than 50% of the oil. This oil had repellent and toxic effects on Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) adults, acting as a contact toxin. Myzus persicae (Sulzer) (Homoptera: Aphididae) and Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae) adults showed behavioral sensibility to this oil.     

Mapping the protein interaction network in methicillin-resistant Staphylococcus aureus

Mortality attributable to infection with methicillin-resistant Staphylococcus aureus (MRSA) has now overtaken the death rate for AIDS in the United States, and advances in research are urgently needed to address this challenge. We report the results of the systematic identification of protein-protein interactions for the hospital-acquired strain MRSA-252. Using a high-throughput pull-down strategy combined with quantitative proteomics to distinguish specific from nonspecific interactors, we identified 13,219 interactions involving 608 MRSA proteins. Consecutive analyses revealed that this protein interaction network (PIN) exhibits scale-free organization with the characteristic presence of highly connected hub proteins. When clinical and experimental antimicrobial targets were queried in the network, they were generally found to occupy peripheral positions in the PIN with relatively few interacting partners. In contrast, the hub proteins identified in this MRSA PIN that are essential for network integrity and stability have largely been overlooked as drug targets. Thus, this empirical MRSA-252 PIN provides a rich source for identifying critical proteins essential for network stability, many of which can be considered as prospective antimicrobial drug targets.     

Preclinical models for neuroblastoma: establishing a baseline for treatment

Background

Preclinical models of pediatric cancers are essential for testing new chemotherapeutic combinations for clinical trials. The most widely used genetic model for preclinical testing of neuroblastoma is the TH-MYCN mouse. This neuroblastoma-prone mouse recapitulates many of the features of human neuroblastoma. Limitations of this model include the low frequency of bone marrow metastasis, the lack of information on whether the gene expression patterns in this system parallels human neuroblastomas, the relatively slow rate of tumor formation and variability in tumor penetrance on different genetic backgrounds. As an alternative, preclinical studies are frequently performed using human cell lines xenografted into immunocompromised mice, either as flank implant or orthtotopically. Drawbacks of this system include the use of cell lines that have been in culture for years, the inappropriate microenvironment of the flank or difficult, time consuming surgery for orthotopic transplants and the absence of an intact immune system.

Principal Findings

Here we characterize and optimize both systems to increase their utility for preclinical studies. We show that TH-MYCN mice develop tumors in the paraspinal ganglia, but not in the adrenal, with cellular and gene expression patterns similar to human NB. In addition, we present a new ultrasound guided, minimally invasive orthotopic xenograft method. This injection technique is rapid, provides accurate targeting of the injected cells and leads to efficient engraftment. We also demonstrate that tumors can be detected, monitored and quantified prior to visualization using ultrasound, MRI and bioluminescence. Finally we develop and test a “standard of care” chemotherapy regimen. This protocol, which is based on current treatments for neuroblastoma, provides a baseline for comparison of new therapeutic agents.

Significance

The studies suggest that use of both the TH-NMYC model of neuroblastoma and the orthotopic xenograft model provide the optimal combination for testing new chemotherapies for this devastating childhood cancer.     

Nestin-GFP transgene reveals neural precursor cells in adult skeletal muscle

Background

Therapy for neural lesions or degenerative diseases relies mainly on finding transplantable active precursor cells. Identifying them in peripheral tissues accessible for biopsy, outside the central nervous system, would circumvent the serious immunological and ethical concerns impeding cell therapy.

Methodology/Principal Findings

In this study, we isolated neural progenitor cells in cultured adult skeletal muscle from transgenic mice in which nestin regulatory elements control GFP expression. These cells also expressed the early neural marker Tuj1 and light and heavy neurofilament but not S100β, indicating that they express typical neural but not Schwann cell markers. GFP+/Tuj1+ cells were also negative for the endothelial and pericyte markers CD31 and α-smooth muscle actin, respectively. We established their a) functional response to glutamate in patch-clamp recordings; b) interstitial mesenchymal origin; c) replicative capacity; and d) the environment necessary for their survival after fluorescence-activated cell sorting.

Conclusions/Significance

We propose that the decline in nestin-GFP expression in muscle progenitor cells and its persistence in neural precursor cells in muscle cultures provide an invaluable tool for isolating a population of predifferentiated neural cells with therapeutic potential.     

Phylodynamics of hepatitis C virus subtype 2c in the province of Córdoba, Argentina

The Hepatitis C Virus Genotype 2 subtype 2c (HCV-2c) is detected as a low prevalence subtype in many countries, except in Southern Europe and Western Africa. The current epidemiology of HCV in Argentina, a low-prevalence country, shows the expected low prevalence for this subtype. However, this subtype is the most prevalent in the central province of Córdoba. Cruz del Eje (CdE), a small rural city of this province, shows a prevalence for HCV infections of 5%, being 90% of the samples classified as HCV-2c. In other locations of Córdoba Province (OLC) with lower prevalence for HCV, HCV-2c was recorded in about 50% of the samples. The phylogenetic analysis of samples from Córdoba Province consistently conformed a monophyletic group with HCV-2c sequences from all the countries where HCV-2c has been sequenced. The phylogeographic analysis showed an overall association between geographical traits and phylogeny, being these associations significant (α = 0.05) for Italy, France, Argentina (places other than Córdoba), Martinique, CdE and OLC. The coalescence analysis for samples from CdE, OLC and France yielded a Time for the Most Common Recent Ancestor of about 140 years, whereas its demographic reconstruction showed a “lag” phase in the viral population until 1880 and then an exponential growth until 1940. These results were also obtained when each geographical area was analyzed separately, suggesting that HCV-2c came into Córdoba province during the migration process, mainly from Europe, which is compatible with the history of Argentina of the early 20th century. This also suggests that the spread of HCV-2c occurred in Europe and South America almost simultaneously, possibly as a result of the advances in medicine technology of the first half of the 20th century.