Apoptotic effects of over-expressed estrogen receptor-beta on LoVo colon cancer cell is mediated by p53 signalings in a ligand-dependent manner

Epidemiologic studies reported that the prevalence of hereditary non-polyposis colon cancer (HNPCC) in male is about 1.5-fold higher than that in female. Decreases in circulatory estradiol (E2) have been reported to downregulate the expression of E2 receptor (ER) and significantly increase the risk of colorectal cancer. Patients that received E2 replacement therapy were found to have a reduction in the incidence of colon adenoma and carcinoma. Furthermore, significant decreases in the expression of ER have been found in colorectal cancer specimens. These data strongly suggest the protective roles of E2 and ER against colorectal cancer. However, the mechanisms remain unexplored. LoVo cells were transient transfected to overexpress ER-beta, DNA fragmentation and caspase activity assay were performed to evaluate apoptotic effects. Western blotting was used to evaluate protein levels, and luciferase activity assay to measure the TNF-alpha promoter activity. Our data clearly demonstrated that E2 and ER-beta alone could upregulate p21 and p27 proteins, which further activate caspase-8 and caspase-9 to induce apoptosis in LoVo cell, and the ER-beta. effects were enhanced by E2. However, overexpressed ER-beta did not influence the expression and promoter activity of TNF-alpha. In addition, E2 and overexpressed ER-beta downregulated the beta-catenin proteins which cause the downregulation of its target genes, cyclin D1 and Rb, to inhibit the cell cycle and cell proliferation. The results indicate that overexpressed ER-beta may induce LoVo cell apoptosis and anti-proliferation by increasing p53 signaling in a ligand-dependent manner, and without hTNF-alpha involvement. Efforts aiming at enhancing ER-beta expression and/or activity may prove to be an attractive alternative therapy against colorectal cancer.     

Protein-protein interaction site predictions with three-dimensional probability distributions of interacting atoms on protein surfaces

Protein-protein interactions are key to many biological processes. Computational methodologies devised to predict protein-protein interaction (PPI) sites on protein surfaces are important tools in providing insights into the biological functions of proteins and in developing therapeutics targeting the protein-protein interaction sites. One of the general features of PPI sites is that the core regions from the two interacting protein surfaces are complementary to each other, similar to the interior of proteins in packing density and in the physicochemical nature of the amino acid composition. In this work, we simulated the physicochemical complementarities by constructing three-dimensional probability density maps of non-covalent interacting atoms on the protein surfaces. The interacting probabilities were derived from the interior of known structures. Machine learning algorithms were applied to learn the characteristic patterns of the probability density maps specific to the PPI sites. The trained predictors for PPI sites were cross-validated with the training cases (consisting of 432 proteins) and were tested on an independent dataset (consisting of 142 proteins). The residue-based Matthews correlation coefficient for the independent test set was 0.423; the accuracy, precision, sensitivity, specificity were 0.753, 0.519, 0.677, and 0.779 respectively. The benchmark results indicate that the optimized machine learning models are among the best predictors in identifying PPI sites on protein surfaces. In particular, the PPI site prediction accuracy increases with increasing size of the PPI site and with increasing hydrophobicity in amino acid composition of the PPI interface; the core interface regions are more likely to be recognized with high prediction confidence. The results indicate that the physicochemical complementarity patterns on protein surfaces are important determinants in PPIs, and a substantial portion of the PPI sites can be predicted correctly with the physicochemical complementarity features based on the non-covalent interaction data derived from protein interiors.     

Patterns of circulating hepatitis B surface antigen variants among vaccinated children born to hepatitis B surface antigen carrier and non-carrier mothers

Hepatitis B virus (HBV) variants that possessed missense mutation within the neutralization epitope of the major S antigen as defined by amino acid residues (aa#) 124–147, termed the a determinant variants, were identified through a population-based serosurvey of 2,305 children of the vaccinated birth cohorts born after 1986. Data on the 678 nucleotides encoding the S antigen of HBV were available for 75 HBV strains that were collected from 63 vaccinated children and 12 unvaccinated or incompletely vaccinated children, and 21 HBV strains from 25 unvaccinated adults. Among the diverse patterns of one to three amino acid substitutions within the a determinant, 145-Arg occurred most frequently (5/14); other variants were: 126-Ala, 127-Thr, 126-Ser/131-Asn/133-Thr, 129-His, 129-Arg, 123-Asn/131-Ile, 133-Leu, 141-Glu, and 141-Arg/144-Ala. Only one of these variants occurred in the 16 hepatitis B surface antigen (HBsAg)-carrier children born to HBsAg-negative mothers, whereas 12 of these variants occurred in the 20 (50%) children born to HBsAg-positive mothers. In addition, early administration of HBV vaccine within the noenatal period increased the likelihood of the emergence of these variants to 64.7% (11/17). Five of the 21 (23.8%) unvaccinated HBsAg-carrier adults harbored the a determinant variants possessing mutations within aa# 125–136, i.e. the putative first loop formed by the cysteine disulfide bonds. Vaccinated children were likely to harbor HBV variants possessing mutations involving altered charge of side chains and/or its hydrophobicity of amino acid residues within the putative second loop between aa#140 and 146. Our data suggest that emergence of these HBV S gene mutants in the phase of HBV vaccination program would be most common among populations in whom perinatal/vertical transmission of HBV is most common, i.e. southeast Asian and the Taiwanese.     

Resistin induces monocyte-endothelial cell adhesion by increasing ICAM-1 and VCAM-1 expression in endothelial cells via p38MAPK-dependent pathway

Resistin, firstly reported as an adipocyte-specific hormone, is suggested to be an important link between obesity and diabetes. Recent studies have suggested an association between resistin and atherogenic processes. The adhesion of circulating monocytes to endothelial cells is a critical step in the early stages of atherosclerosis. The purpose of the present study was to investigate the effect of resistin on the adhesion of THP-1 monocytes to human umbilical vein endothelial cells (HUVECs) and the underlying mechanism. Our results showed that resistin caused a significant increase in monocyte adhesion. In exploring the underlying mechanisms of resistin action, we found that resistin-induced monocyte adhesion was blocked by inhibition of p38MAPK activation using SB203580 and SB202190. Furthermore, resistin increased the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) by HUVECs and these effects were also p38MAPK-dependent. Resistin-induced monocyte adhesion was also blocked by monoclonal antibodies against ICAM-1 and VCAM-1. Taken together, these results show that resistin increases both the expression of ICAM-1 and VCAM-1 by endothelial cells and monocyte adhesion to HUVECs via p38MAPK-dependent pathways.     

ArrayFusion: a web application for multi-dimensional analysis of CGH, SNP and microarray data

ArrayFusion annotates conventional CGH results and various types of microarray data from a range of platforms (cDNA, expression, exon, SNP, array-CGH and ChIP-on-chip) and converts them into standard formats which can be visualized in genome browsers (Affymetrix Integrated Genome Browser and GBrowse in the HapMap Project). Converted files can then be imported simultaneously into a single genome browser to benefit a collective interpretation between different array results. ArrayFusion therefore provides a new type of tool facilitating the integration of CGH and array results to provide new experimental directions. AVAILABILITY: http://microarray.ym.edu.tw/tools/arrayfusion     

The Bacillus anthracis Protein MprF Is Required for Synthesis of Lysylphosphatidylglycerols and for Resistance to Cationic Antimicrobial Peptides

During inhalational anthrax, Bacillus anthracis survives and replicates in alveolar macrophages, followed by rapid invasion into the host's bloodstream, where it multiplies to cause heavy bacteremia. B. anthracis must therefore defend itself from host immune functions encountered during both the intracellular and the extracellular stages of anthrax infection. In both of these niches, cationic antimicrobial peptides are an essential component of the host's innate immune response that targets B. anthracis. However, the genetic determinants of B. anthracis contributing to resistance to these peptides are largely unknown. Here we generated Tn917 transposon mutants in the ΔANR strain (pXO1⁻ pXO2⁻) of B. anthracis and screened them for altered protamine susceptibility. A protamine-sensitive mutant identified carried the transposon inserted in the BA1486 gene encoding a putative membrane protein homologous to MprF known in several gram-positive pathogens. A mutant strain with the BAS1375 gene (the orthologue of BA1486) deleted in the Sterne 34F2 strain (pXO1⁺ pXO2⁻) of B. anthracis exhibited hypersusceptibility not only to protamine but also to α-helical cathelicidin LL-37 and β-sheet defensin human neutrophil peptide 1 compared to the wild-type Sterne strain. Analysis of membrane lipids using isotopic labeling demonstrated that the BAS1375 deletion mutant is unable to synthesize lysinylated phosphatidylglycerols, and this defect is rescued by genetic complementation. Further, we determined the structures of these lysylphosphatidylglycerols by using various mass spectrometric analyses. These results demonstrate that in B. anthracis a functional MprF is required for the biosynthesis of lysylphosphatidylglycerols, which is critical for resistance to cationic antimicrobial peptides.     

Characterization and structural analyses of nonspecific lipid transfer protein 1 from mung bean

Plant nonspecific lipid transfer proteins (nsLTPs) are thermal stable proteins that are capable of transferring lipid molecules between bilayers in vitro. This family of proteins, abundant in plants, is proposed to be involved in defense, pollination, and germination; the in vivo biological function remains, however, elusive. Here we report the purification and sequencing of an nsLTP1 from mung bean sprouts. We have also determined the solution structure of this nsLTP1, which represents the first 3D structure of the dicotyledonous nsLTP1 family. The global fold of mung bean nsLTP1 is similar to those of the monocotyledonous nsLTP1 structures and consists of four alpha-helices stabilized by four disulfide bonds. There are, however, some notable differences in the C-terminal tails and internal hydrophobic cavities. Circular dichroism and fluorescence spectroscopy were used to compare the thermodynamics and lipid transfer properties of mung bean nsLTP1 with those of rice nsLTP1. Docking of a lipid molecule into the solution structure of mung bean nsLTP1 reveals similar binding cavities and hydrophobic interactions as in rice nsLTP1, consistent with their comparable lipid transfer properties measured experimentally.     

Developmentally regulated sphingolipid synthesis in African trypanosomes

Sphingolipids are essential components of eukaryotic membranes, and many unicellular eukaryotes, including kinetoplastid protozoa, are thought to synthesize exclusively inositol phosphorylceramide (IPC). Here we characterize sphingolipids from Trypanosoma brucei, and a trypanosome sphingolipid synthase gene family (TbSLS1-4) that is orthologous to Leishmania IPC synthase. Procyclic trypanosomes contain IPC, but also sphingomyelin, while surprisingly bloodstream-stage parasites contain sphingomyelin and ethanolamine phosphorylceramide (EPC), but no detectable IPC. In vivo fluorescent ceramide labelling confirmed stage-specific biosynthesis of both sphingomyelin and IPC. Expression of TbSLS4 in Leishmania resulted in production of sphingomyelin and EPC suggesting that the TbSLS gene family has bi-functional synthase activity. RNAi silencing of TbSLS1-4 in bloodstream trypanosomes led to rapid growth arrest and eventual cell death. Ceramide levels were increased more than threefold by silencing suggesting a toxic downstream effect mediated by this potent intracellular messenger. Topology predictions support a revised six-transmembrane domain model for the kinetoplastid sphingolipid synthases consistent with the proposed mammalian sphingomyelin synthase structure. This work reveals novel diversity and regulation in sphingolipid metabolism in this important group of human parasites.     

N-Acetylcysteine ameliorates lipopolysaccharide-induced organ damage in conscious rats

Lipopolysaccharide is strongly associated with septic shock, leading to multiple organ failure. It can activate monocytes and macrophages to release proinflammatory mediators such as tumor necrosis factor- (TNF-), interleukin-1 (IL-1), and nitric oxide (NO). The present experiments were designed to induce endotoxin shock by an intravenous injection ofKlebsiella pneumoniae lipopolysaccharide (LPS, 10 mg/kg) in conscious rats. Arterial pressure and heart rate (HR) were continuously monitored for 48 h after LPS administration. N-Acetyl-cysteine was used to study its effects on organ damage. Biochemical substances were measured to reflect organ functions. Biochemical factors included blood urea nitrogen (BUN), creatinine (Cre), lactic dehydrogenase (LDH), creatine phosphokinase (CPK), aspartate transferase (GOT), alanine transferase (GPT), TNF-, IL-1, methyl guanidine (MG), and nitrites/nitrates. LPS caused significant increases in blood BUN, Cre, LDH, CPK, GOT, GPT, TNF-, IL-1, MG levels, and HR, as well as a decrease in mean arterial pressure and an elevation of nitrites/nitrates. N-Acetylcysteine suppressed the release of TNF-, IL-1, and MG, but enhanced NO production. These actions ameliorate LPS-induced organ damage in conscious rats. The beneficial effects may suggest a potential chemopreventive effect of this compound in sepsis prevention and treatment.     

Winter spatial distribution of fish larvae assemblages relative to the hydrography of the waters surrounding Taiwan

The aim of this study was to investigate the species composition and distribution of fish larvae in relation to hydrographic conditions in the waters surrounding Taiwan Island (TI) in February 2003. In total, 242 kinds of fish larvae belonging to 127 genera and 75 families were recognized. Among these, 109 taxa were identified to the family or genus level, others to the species level. The 12 predominant types, which constituted 71% of the total fish larvae, were Engraulis japonica, Scomber sp., Diaphus spp., Benthosema pterotum, Carangoides ferdau, Embolichthys mitsukurii, Maurolicus sp., unidentified Myctophidae, Gonostoma gracile, Trichiurus lepturus, unidentified Gobiidae, and Myctophum asperum. The distribution of fish larvae showed a clear association with water masses around TI, with higher abundances and lower species richness northwest of TI where the China Coastal Current prevails, and lower abundances and higher species diversity east of TI where the Kuroshio Current dominates. Cluster analysis distinguished three station groups and four species groups, and the distribution patterns of fish larvae also corresponded to hydrographic conditions. The total abundances of fish larvae and eight of the 12 predominant taxa showed significant and positive correlations with zooplankton abundance, which suggests that food source might be a key factor determining the abundance and distribution of fish larvae during the winter.