In vivo oestrogenicity and binding characteristics of alpha-zearalanol (P-1496) to different classes of oestrogen binding proteins in rat liver

It is now well established that the mycotoxin zearalenone and some of its derivatives possess oestrogenic activity. In the present study, the binding characteristics of [3H]zearalanol (P-1496) to different classes of sites including [1] the oestrogen receptor, [2] the higher capacity lower affinity (HCLA) sites, [3] the antioestrogen sites and [4] a new class of binding sites apparently specific for P-1496 were examined in rat liver. Analysis of the binding by sucrose density gradient centrifugation confirmed that P-1496 binds to the oestrogen receptor but not to the higher capacity lower affinity sites for oestradiol-17 beta. Furthermore, saturation experiments using partially-purified fractions showed that P-1496 binds to the oestrogen receptor with an affinity very similar to that of oestradiol-17 beta (apparent dissociation constants ranged from 0.1-0.3 nM). Competition studies using partially purified cytosolic oestrogen receptor suggested that P-1496 binds to a second high affinity site distinct from the oestrogen receptor. This binding site was further characterized as selective for P-1496 by saturation analysis following the complete occupancy of oestrogen receptor by oestradiol-17 beta. The in vitro binding characteristics of P-1496 were then compared with in vivo effects on concentrations of serum triglycerides. Treatment of ovariectomized female rats daily with 1.5 or 2 mg P-1496/kg body weight resulted in marked increases in the concentrations of serum triglycerides associated with the very low density lipoprotein (VLDL) fraction. Dose-response studies indicated that there was no sex difference with respect to the dose necessary to produce significant increases in serum triglycerides. The present study shows striking similarities between the binding of P-1496 and oestradiol-17 beta to liver oestrogen receptor in vitro. However, differences are observed with respect to their binding to other cytoplasmic components of liver. In addition, although P-1496 is capable of eliciting in vivo oestrogenic effects in liver, it is much less potent than oestradiol-17 beta.     

Robustness and Epistasis in the C. elegans Vulval Signaling Network Revealed by Pathway Dosage Modulation

1. Download : Download high-res image (275KB)
2. Download : Download full-size image

Highlights? A phase map of cell fate patterns is built as a function of signaling pathway dose ? The vulva system can buffer a 4-fold variation in mean lin-3/egf mRNA number ? The major role of LIN-12/Notch in the vulva is to promote the 2° fate ? Inhibition of 1° fate by LIN-12 is important when lin-3 dose is mildly increased     

MicroRNAs Act as Cofactors in Bicoid-Mediated Translational Repression

1. Download : Download high-res image (146KB)
2. Download : Download full-size image

     

An alternative strategy for the membrane proteome analysis of the green sulfur bacterium Chlorobium tepidum using blue native PAGE and 2-D PAGE on purified membranes

To avoid the specific problems concerning intrinsic membrane proteins in proteome analysis, an alternative strategy is described that is complementary to previous investigations using 2-D polyacrylamide gel electrophoresis (PAGE) techniques. The strategy involves (a) obtaining purified preparations of the membranes from Chlorobium tepidum by washing with 2 M NaBr, which removed membrane-associated soluble proteins and membrane-associated organelles; (b) separation of membrane protein complexes using 1-D Blue-native polyacrylamide gel electrophoresis (BN-PAGE) after solubilization with n-dodecyl-beta-d-maltoside (DDM); (c) combination of the BN with Tricine-SDS-PAGE; (d) high-throughput mass spectrometric analysis after gel band excision, in-gel digestion, and MALDI target spotting; and (e) protein identification from mixtures of tryptic peptides by peptide mass fingerprinting. Using this approach, we identified 143 different proteins, 70 of which have not been previously reported using 2-D PAGE techniques. Membrane proteins with up to 14 transmembrane helices were found, and this procedure proved to be efficient with proteins within a wide pI range (4.4-11.6). About 54% of the identified membrane proteins belong to various functional categories like energy metabolism, transport, signal transduction, and protein translocation, while for the others, a function is not yet known, indicating the potential of the method for the elucidation of the membrane proteomes in general.     

Application of discriminant analysis and quantitative cytologic examination to gastric lesions

OBJECTIVE: To investigate of the potential value of morphometry and discriminant analysis for the classification of benign and malignant gastric cells and lesions. STUDY DESIGN: The data set consisted of 13,300 cells from 120 cases composed of 30 cases of cancer, 26 cases of gastritis and 64 cases of ulcer according to the final histologic diagnosis. The cytologic diagnosis was divided into 5 categories (gastritis, ulcer, inflammatory dysplasia, cancer and true dysplasia). Classification was attempted at 2 levels: the cell level to classify individual cells and the case level to classify individual cases. For the cellular classification the measured cells from 50% of available cases were selected as a training set to construct a model. The cells from the remaining cases were used as a test set to validate the model. Similarly for case classification, the same 50% of cases that were used for cell classification were used as a training set and the remaining cases as a test set. Images of routinely processed gastric smears stained by the Papanicolaou technique were analyzed by a customized image analysis system. RESULTS: Application of discriminant analysis on the test set gave correct classification of 98.4% of benign cells and 67.1% of malignant cells. On case classification, 100% accuracy was achieved for benign and malignant cases, both for the training and test sets. CONCLUSION: The application of discriminant analysis described in this paper could produce significant classification results at the cellular and individual case level.     

Protein and lipid composition of a vitellin isolated from eggs of Sparus aurata

The protein and lipid composition of a vitellin isolated from eggs of Sparus aurata were characterized by SDS PAGE, N-terminal sequence analysis and lipid analysis by thin layer chromatography and gas chromatography. The lipoprotein complex contains proteins with apparent molecular weights of 69, 59, 23, 21 and 12 kDa and were characterized as vitellinogenin fragments by N-terminal sequencing. Lipid extraction and analysis indicate an association of cholesterol and phospholipids with the protein subunits. The phospholipids contain fatty acids with 14, 16 and 18 carbon atoms as determined by GC/MS.