Quantitative appreciation of steroidogenic gene expression in mouse tissues: new roles for type 2 5alpha-reductase, 20alpha-hydroxysteroid dehydrogenase and estrogen sulfotransferase

We have recently developed an improved method for the RealTime PCR quantification of reversed transcribed mRNA (Q_RTPCR) that allows to obtain absolute mRNA expression levels with high sensitivity and accuracy. Using this Q_RTPCR method to assess the mRNA expression levels of genes encoding steroidogenic enzymes in male and female mouse tissues allows us to gain quantitative appreciation of the function of these genes. We could thus identify the existence of two types of steroidogenic tissues: those of classical endocrine glands such as the testis, ovary and adrenals which deliver steroids into the circulation, and in which millions of copies/mug total RNA are detected, and those of peripheral intracrine tissues where steroids are synthesized locally and exert their action at the site where they are produced (prostate, uterus, etc.), and in which the expression level of steroidogenic enzymes is much lower. We also observed an abnormally high expression levels of type 2 5alpha-reductase and 20alpha-HSD in the male and female adrenals, respectively, thus indirectly suggesting new roles for these sex-specific enzymes. On the other hand estrogen sulfotransferase, the enzyme that inactivates estrogen, has been found selectively expressed in male tissues, thus suggesting a role for this enzyme to protect male-specific tissues against estrogenic activity.     

Synthesis and structure-activity relationships of analogs of EM-652 (acolbifene), a pure selective estrogen receptor modulator. Study of nitrogen substitution

EM-652 (acolbifene) analogs have been synthesized as selective estrogen receptor modulators. Substitution on the nitrogen atom of these 2H-1-benzopyran derivatives has been studied for its influence on antiestrogenic activity. Binding to the rat estrogen receptor, inhibition of estradiol-stimulated proliferation of T-47D breast cancer cells, as well as antiuterotrophic and uterotrophic activities in ovariectomized mice have been evaluated. 2H-1-Benzopyran 1b (EM-343, racemic form of EM-652), which contains a piperidine ring, shows the best pharmacological profile; RBA = 380, IC50 value = 0.110 nM (in T-47D cells), as well as 63% and 84% antiuterotrophic inhibitions at the 7.5 and 75 nmol doses, respectively.     

Interaction between epithelial basement membrane and migrating mesoblast cells in the avian blastoderm

We investigated the remodeling of glucosamine-containing basement-membrane components in chimaeric avian embryos during gastrulation. Epiblast grafts metabolically labelled with tritiated glucosamine were excised from gastrulating quail embryos and implanted orthotopically into chicken embryos at the same developmental stage. The chimaerae were allowed to develop in culture for 5-7 h before autoradiographic processing. The resulting autoradiographs not only showed the presence of silver grains in the grafted quail tissue and at the level of its basement membrane, but also revealed labelling in the basement-membrane region of the chicken tissue lateral to the graft, i.e. between the mesoblast and epiblast. This last labelling extended as far as at the edge of the area pellucida, i.e. in a region of chicken tissue situated more laterally than the initial position of the graft. No labelling was observed medial, anterior, or posterior to the graft. This observation argues against the interpretation that our results were due to diffusion of labelled compounds within the basement membrane. We also provide evidence to exclude the possibility that quail epiblast cells migrated on their own underlying basement membrane, leaving behind a carpet of labelled material. Taking into account, firstly, the morphogenetic movements that occur during gastrulation, i.e. the movement of epiblast cells towards the primitive streak where they ingress, and the migration of mesoblast cells along the basement membrane towards the periphery of the area pellucida, and secondly, the medial movement of the basement membrane, it is suggested that mesoblast cells picked up labelled compounds in the basement membrane of the graft and left these behind during their lateral migration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Spatial extent and historical context of North Sea oxygen depletion in August 2010

Prompted by recent observations of seasonal low dissolved oxygen from two moorings in the North Sea, a hydrographic survey in August 2010 mapped the spatial extent of summer oxygen depletion. Typical near-bed dissolved oxygen saturations in the stratified regions of the North Sea were 75–80 % while the well-mixed regions of the southern North Sea reached 90 %. Two regions of strong thermal stratification, the area between the Dooley and Central North Sea Currents and the area known as the Oyster Grounds, had oxygen saturations as low as 65 and 70 % (200 and 180 μmol dm^?3) respectively. Low dissolved oxygen was apparent in regions characterised by low advection, high stratification, elevated organic matter production from the spring bloom and a deep chlorophyll maximum. Historical data over the last century from the International Council for the Exploration of the Sea oceanographic database highlight an increase in seasonal oxygen depletion and a warming over the past 20 years. The 2010 survey is consistent with, and reinforces, the signal of recent depleted oxygen at key locations seen in the (albeit sparse) historical data.     

Global warming will affect the genetic diversity and uniqueness of Lycaena helle populations

The climate warming of the postglacial has strongly reduced the distribution of cold‐adapted species over most of Central Europe. Such taxa have therefore become extinct over most of the lowlands and shifted to higher altitudes where they have survived to the present day. The lycaenid butterfly Lycaena helle follows this pattern of former widespread distribution and later restriction to mountain areas such as the European middle mountains. We sampled 203 individuals from 10 populations representing six mountain ranges (Pyrenees, Jura, Massif Central, Morvan, Vosges and Ardennes) over the species' western distribution. Allozyme and microsatellite polymorphisms were analysed to study the genetic status of these highly fragmented populations. Both molecular marker systems revealed a strong genetic differentiation among the analysed populations, coinciding with the orographic structure and highly restricted gene flow among them. The large‐scale genetic differentiation is more pronounced in allozymes (F_CT: 0.326) than in microsatellites (R_CT: 0.113), but microsatellites show a higher resolution on the regional scale (R_SC: 0.082) compared with allozymes (F_SC: n.s.). For both analytical tools, we found private alleles occurring exclusively in a single mountain area. The highly fragmented and isolated occurrence of populations is supported by the distribution pattern of potentially suitable climate suggested by species distribution models. Model projections under two climate warming scenarios predict a decline of climatically suitable areas, which will result in the extinction of most of the populations showing unique genetic characteristics.     

Experimental climate effect on seasonal variability of polyphenol/phenoloxidase interplay along a narrow fen–bog ecological gradient in Sphagnum fallax

Extracellular phenoloxidase enzymes play an important role in the stability of soil carbon storage by contributing to the cycling of complex recalcitrant phenolic compounds. Climate warming could affect peatland functioning through an alteration of polyphenol/phenoloxidase interplay, which could lead them to becoming weaker sinks of carbon. Here, we assessed the seasonal variability of total phenolics and phenoloxidases subjected to 2–3 °C increase in air temperature using open‐top chambers. The measurements were performed along a narrow fen–bog ecological gradient over one growing season. Climate warming had a weak effect on phenoloxidases, but reduced phenolics in both fen and bog areas. Multivariate analyses revealed a split between the areas and also showed that climate warming exacerbated the seasonal variability of polyphenols, culminating in a destabilization of the carbon cycle. A negative relationship between polyphenols and phenoloxidases was recorded in controls and climate treatments suggesting an inhibitory effect of phenolics on phenoloxidases. Any significant decrease of phenolics through repeatedly elevated temperature would greatly impact the ecosystem functioning and carbon cycle through an alteration of the interaction of polyphenols with microbial communities and the production of extracellular enzymes. Our climate treatments did not have the same impact along the fen–bog gradient and suggested that not all the peatland habitats would respond similarly to climate forcing.     

Rhein inhibits angiogenesis and the viability of hormone-dependent and -independent cancer cells under normoxic or hypoxic conditions in vitro

Hypoxia is a hallmark of solid tumors, including breast cancer, and the extent of tumor hypoxia is associated with treatment resistance and poor prognosis. Considering the limited treatment of hypoxic tumor cells and hence a poor prognosis of breast cancer, the investigation of natural products as potential chemopreventive anti-angiogenic agents is of paramount interest. Rhein (4,5-dihydroxyanthraquinone-2-carboxylic acid), the primary anthraquinone in the roots of Cassia alata L., is a naturally occurring quinone which exhibits a variety of biologic activities including anti-cancer activity. However, the effect of rhein on endothelial or cancer cells under hypoxic conditions has never been delineated. Therefore, the aim of this study was to investigate whether rhein inhibits angiogenesis and the viability of hormone-dependent (MCF-7) or -independent (MDA-MB-435s) breast cancer cells in vitro under normoxic or hypoxic conditions. Rhein inhibited vascular endothelial growth factor (VEGF₁₆₅)-stimulated human umbilical vein endothelial cell (HUVEC) tube formation, proliferation and migration under normoxic and hypoxic conditions. In addition, rhein inhibited in vitro angiogenesis by suppressing the activation of phosphatidylinositol 3-kinase (PI3K), phosphorylated-AKT (p-AKT) and phosphorylated extracellular signal-regulated kinase (p-ERK) but showed no inhibitory effects on total AKT or ERK. Rhein dose-dependently inhibited the viability of MCF-7 and MDA-MB-435s breast cancer cells under normoxic or hypoxic conditions, and inhibited cell cycle in both cell lines. Furthermore, Western blotting demonstrated that rhein inhibited heat shock protein 90alpha (Hsp90α) activity to induce degradation of Hsp90 client proteins including nuclear factor-kappa B (NF-κB), COX-2, and HER-2. Rhein also inhibited the expression of hypoxia-inducible factor-1 alpha (HIF-1α), vascular endothelial growth factor (VEGF₁₆₅), epidermal growth factor (EGF), and the phosphorylation of inhibitor of NF-κB (I-κB) under normoxic or hypoxic conditions. Taken together, these data indicate that rhein is a promising anti-angiogenic compound for breast cancer cell viability and growth. Therefore, further studies including in vivo and pre-clinical need to be performed.