Sex hormone modulation of cell growth and apoptosis of the human monocytic/macrophage cell line

Sex hormones seem to modulate the immune/inflammatory responses by different mechanisms in female and male rheumatoid arthritis patients. The effects of 17β-oestradiol and of testosterone were tested on the cultured human monocytic/macrophage cell line (THP-1) activated with IFN-γ in order to investigate their role in cell proliferation and apoptosis. Activated human THP-1 cells were cultured in the presence of 17β-oestradiol and testosterone (final concentration, 10 nM). The evaluation of markers of cell proliferation included the NF-κB DNA-binding assay, the NF-κB inhibition complex, the proliferating cell nuclear antigen expression and the methyl-tetrazolium salt test. Apoptosis was detected by the annexin V-propidium assay and by the cleaved poly-ADP ribose polymerase expression. Specific methods included flow analysis cytometry scatter analysis, immunocytochemistry and western blot analysis. Cell growth inhibition and increased apoptosis were observed in testosterone-treated THP-1 cells. Increased poly-ADP ribose polymerase-cleaved expression and decreased proliferating cell nuclear antigen expression, as well as an increase of IκB-α and a decrease of the IκB-α phosphorylated form (ser 32), were found in testosterone-treated THP-1 cells. However, the NF-κB DNA binding was found increased in 17β-oestradiol-treated THP-1 cells. The treatment with staurosporine (enhancer of apoptosis) induced decreased NF-κB DNA binding in all conditions, but particularly in testosterone-treated THP-1 cells. Treatment of THP-1 by sex hormones was found to influence cell proliferation and apoptosis. Androgens were found to increase the apoptosis, and oestrogens showed a protective trend on cell death – both acting as modulators of the NF-κB complex.     

Characterization of a mutation that results in independence of oxidosqualene cyclase (Erg7) activity from the downstream 3-ketoreductase (Erg27) in the yeast ergosterol biosynthetic pathway

In yeast, deletion of ERG27, which encodes the sterol biosynthetic enzyme, 3-keto-reductase, results in a concomitant loss of the upstream enzyme, Erg7p, an oxidosqualene cyclase (OSC). However, this phenomenon occurs only in fungi, as mammalian Erg27p orthologues are unable to rescue yeast Erg7p activity. In this study, an erg27 mutant containing the mouse ERG27 orthologue was isolated that was capable of growing without sterol supplementation (FGerg27). GC/MS analysis of this strain showed an accumulation of squalene epoxides, 3-ketosterones, and ergosterol. This strain which was crossed to a wildtype and daughter segregants showed an accumulation of squalene epoxides as well as ergosterol indicating that the mutation entailed a leaky block at ERG7. Upon sequencing the yeast ERG7 gene an A598S alteration was found in a conserved alpha helical region. We theorize that this mutation stabilizes Erg7p in a conformation that mimics Erg27p binding. This mutation, while decreasing OSC activity still retains sufficient residual OSC activity such that the strain in the presence of the mammalian 3-keto reductase enzyme functions and no longer requires the yeast Erg27p. Because sterol biosynthesis occurs in the ER, a fusion protein was synthesized combining Erg7p and Erg28p, a resident ER protein and scaffold of the C-4 demethyation complex. Both FGerg27 and erg27 strains containing this fusion plasmid and the mouse ERG27 orthologue showed restoration of ergosterol biosynthesis with minimal accumulation of squalene epoxides. These results indicate retention of Erg7p in the ER increases its activity and suggest a novel method of regulation of ergosterol biosynthesis.     

Chemical characterization (GC/MS and NMR Fingerprinting) and bioactivities of South-African Pelargonium capitatum (L.) L' Her. (Geraniaceae) essential oil

Chemical fingerprinting of commercial Pelargonium capitatum (Geraniaceae) essential oil samples of south African origin was performed by GC, GC/MS, and (13) C- and (1) H-NMR. Thirty-seven compounds were identified, among which citronellol (32.71%) and geraniol (19.58%) were the most abundant. NMR Spectra of characteristic chemicals were provided. Broad-spectrum bioactivity properties of the oil were evaluated and compared with those of commercial Thymus vulgaris essential oil with the aim to obtain a functional profile in terms of efficacy and safety. P. capitatum essential oil provides a good performance as antimicrobial, with particular efficacy against Candida albicans strains. Antifungal activity performed against dermatophyte and phytopathogen strains revealed the latter as more sensitive, while antibacterial activity was not remarkable against both Gram-positive and Gram-negative bacteria. P. capitatum oil provided a lower antioxidant activity (IC(50) ) than that expressed by thyme essential oil, both in the 1,1-diphenyl-2-picrylhydrazyl (DPPH) and β-carotene bleaching tests. Results in photochemiluminescence (PCL) assay were negligible. To test the safety aspects of P. capitatum essential oil, mutagenic and toxicity properties were assayed by Ames test, with and without metabolic activation. Possible efficacy of P. capitatum essential oil as mutagenic protective agent against NaN(3) , 2-nitrofluorene, and 2-aminoanthracene was also assayed, providing interesting and significant antigenotoxic properties.     

Toxic HypF-N Oligomers Selectively Bind the Plasma Membrane to Impair Cell Adhesion Capability

The deposition of fibrillar protein aggregates in human organs is the hallmark of several pathological states, including highly debilitating neurodegenerative disorders and systemic amyloidoses. It is widely accepted that small oligomers arising as intermediates in the aggregation process, released by fibrils, or growing in secondary nucleation steps are the cytotoxic entities in protein-misfolding diseases, notably neurodegenerative conditions. Increasing evidence indicates that cytotoxicity is triggered by the interaction between nanosized protein aggregates and cell membranes, even though little information on the molecular details of such interaction is presently available. In this work, we propose what is, to our knowledge, a new approach, based on the use of single-cell force spectroscopy applied to multifunctional substrates, to study the interaction between protein oligomers, cell membranes, and/or the extracellular matrix. We compared the interaction of single Chinese hamster ovary cells with two types of oligomers (toxic and nontoxic) grown from the N-terminal domain of the Escherichia coli protein HypF. We were able to quantify the affinity between both oligomer type and the cell membrane by measuring the mechanical work needed to detach the cells from the aggregates, and we could discriminate the contributions of the membrane lipid and protein fractions to such affinity. The fundamental role of the ganglioside GM1 in the membrane-oligomers interaction was also highlighted. Finally, we observed that the binding of toxic oligomers to the cell membrane significantly affects the functionality of adhesion molecules such as Arg-Gly-Asp binding integrins, and that this effect requires the presence of the negatively charged sialic acid moiety of GM1.     

Landscape simplification increases vineyard pest outbreaks and insecticide use

Diversifying agricultural landscapes may mitigate biodiversity declines and improve pest management. Yet landscapes are rarely managed to suppress pests, in part because researchers seldom measure key variables related to pest outbreaks and insecticides that drive management decisions. We used a 13‐year government database to analyse landscape effects on European grapevine moth (Lobesia botrana) outbreaks and insecticides across c. 400 Spanish vineyards. At harvest, we found pest outbreaks increased four‐fold in simplified, vineyard‐dominated landscapes compared to complex landscapes in which vineyards are surrounded by semi‐natural habitats. Similarly, insecticide applications doubled in vineyard‐dominated landscapes but declined in vineyards surrounded by shrubland. Importantly, pest population stochasticity would have masked these large effects if numbers of study sites and years were reduced to typical levels in landscape pest‐control studies. Our results suggest increasing landscape complexity may mitigate pest populations and insecticide applications. Habitat conservation represents an economically and environmentally sound approach for achieving sustainable grape production.     

Genome Size: A Novel Genomic Signature in Support of Afrotheria

Molecular phylogenetic analyses suggest an emerging phylogeny for the extant Placentalia (eutherian) that radically departs from morphologically based constructions of the past. Placental mammals are partitioned into four supraordinal clades: Afrotheria, Xenarthra, Laurasiatheria, and Euarchontoglires. Afrotheria form an endemic African clade that includes elephant shrews, golden moles, tenrecs, aardvarks, hyraxes, elephants, dugongs, and manatees. Datamining databases of genome size (GS) shows that till today just one afrotherian GS has been evaluated, that of the aardvark Orycteropus afer. We show that the GSs of six selected representatives across the Afrotheria supraordinal group are among the highest for the extant Placentalia, providing a novel genomic signature of this enigmatic group. The mean GS value of Afrotheria, 5.3 ± 0.7 pg, is the highest reported for the extant Placentalia. This should assist in planning new genome sequencing initiatives. [Reviewing Editor: Dmitri Petrov]     

Microplastic ingestion ubiquitous in marine turtles

Despite concerns regarding the environmental impacts of microplastics, knowledge of the incidence and levels of synthetic particles in large marine vertebrates is lacking. Here, we utilize an optimized enzymatic digestion methodology, previously developed for zooplankton, to explore whether synthetic particles could be isolated from marine turtle ingesta. We report the presence of synthetic particles in every turtle subjected to investigation (n = 102) which included individuals from all seven species of marine turtle, sampled from three ocean basins (Atlantic [ATL]: n = 30, four species; Mediterranean (MED): n = 56, two species; Pacific (PAC): n = 16, five species). Most particles (n = 811) were fibres (ATL: 77.1% MED: 85.3% PAC: 64.8%) with blue and black being the dominant colours. In lesser quantities were fragments (ATL: 22.9%: MED: 14.7% PAC: 20.2%) and microbeads (4.8%; PAC only; to our knowledge the first isolation of microbeads from marine megavertebrates). Fourier transform infrared spectroscopy (FT‐IR) of a subsample of particles (n = 169) showed a range of synthetic materials such as elastomers (MED: 61.2%; PAC: 3.4%), thermoplastics (ATL: 36.8%: MED: 20.7% PAC: 27.7%) and synthetic regenerated cellulosic fibres (SRCF; ATL: 63.2%: MED: 5.8% PAC: 68.9%). Synthetic particles being isolated from species occupying different trophic levels suggest the possibility of multiple ingestion pathways. These include exposure from polluted seawater and sediments and/or additional trophic transfer from contaminated prey/forage items. We assess the likelihood that microplastic ingestion presents a significant conservation problem at current levels compared to other anthropogenic threats.