Antioxidant activity of blueberry fruit is impaired by association with milk

The antioxidant properties of dietary phenolics are believed to be reduced in vivo because of their affinity for proteins. In this study we assessed the bioavailability of phenolics and the in vivo plasma antioxidant capacity after the consumption of blueberries (Vaccinium corymbosum L.) with and without milk. In a crossover design, 11 healthy human volunteers consumed either (a) 200 g of blueberries plus 200 ml of water or (b) 200 g of blueberries plus 200 ml of whole milk. Venous samples were collected at baseline and at 1, 2, and 5 h postconsumption. Ingestion of blueberries increased plasma levels of reducing and chain-breaking potential (+ 6.1%, p < 0.001; + 11.1%, p < 0.05) and enhanced plasma concentrations of caffeic and ferulic acid. When blueberries and milk were ingested there was no increase in plasma antioxidant capacity. There was a reduction in the peak plasma concentrations of caffeic and ferulic acid (? 49.7%, p < 0.001, and ? 19.8%, p < 0.05, respectively) as well as the overall absorption (AUC) of caffeic acid (p < 0.001). The ingestion of blueberries in association with milk, thus, impairs the in vivo antioxidant properties of blueberries and reduces the absorption of caffeic acid.     

The P2Y4 receptor forms homo-oligomeric complexes in several CNS and PNS neuronal cells

It is well established that several cell surface receptors interact with each other to form dimers and oligomers, which are essential for their activation. Since little is known about the quaternary structure of P2Y receptors, in the present work, we investigated the expression of the G-protein-coupled P2Y₄ subunit as monomeric or higher-order complex protein. We examined both endogenously expressed P2Y₄ subtype with the aid of specific anti-P2Y₄ antiserum, and heterologously transfected P2Y₄-tagged receptors with the use of antitag antibodies. In both cases, we found the P2Y₄ receptor displaying molecular masses corresponding to monomeric, dimeric and oligomeric structures. Experiments performed in the absence of reducing agents demonstrated that there is a strict correlation among the multiple protein bands and that the multimeric forms are at least partially assembled by disulphide bonds. The direct demonstration of P2Y₄ homodimerisation comes instead from co–transfection and differential co–immunoprecipitation experiments, with the use of differently tagged P2Y₄ receptors and antitag antibodies. The structural propensity of the P2Y₄ protein to form homo-oligomers may open the possibility of a novel regulatory mechanism of physiopathological functions for this and additional P2Y receptors.     

Diversity of salt response among yeasts

Forty-two yeast strains from 27 species belonging to seven genera, selected for their ability to grow in 10% NaCl, have been analysed for their resistance to salt concentrations up to 5 M, by calculating the Minimum Inhibitory Concentrations (MIC). Using eight different NaCl concentrations from 0 to 5M, results show that halotolerance (MIC) ranges from 1.7 to 3.8 M NaCl, with an avera ge around 2.5 M and confirm that the most halotolerant strains belong to the speciesDebaryomyces hansenii. Since a real halophily could not be found in these isolates, and is generally questioned to be present among the yeast, the effects of NaCl has been measured as salt enhancement effect on growth (MSE), which is defined as the rate between the growth at a given NaCl concentration and theowth in the medium without addition of salt. The implications of these findings in food microbial ecology and technology are discussed.     

Bacterial Community Structure of Sediments of the Bizerte Lagoon (Tunisia), a Southern Mediterranean Coastal Anthropized Lagoon

In order to estimate how pollution affects the bacterial community structure and composition of sediments, chemical and molecular approaches were combined to investigate eight stations around the Bizerte lagoon. Terminal restriction fragment length polymorphism (T-RFLP) analysis of PCR-amplified 16S rRNA genes revealed that each station was characterized by a specific bacterial community structure. The combination of this data with those of chemical analysis showed a correlation between the bacterial fingerprint and the pollutant content, principally with hydrocarbon pollution. The composition of the bacterial community of two contrasted stations related to the pollution revealed sequences affiliated to α, β, γ, δ, ε subclass of the Proteobacteria, Actinobacteria, and Acidobacteria in both stations although in different extent. Gamma and delta subclass of the Proteobacteria were dominant and represent 70% of clones in the heavy-metal-contaminated station and 47% in the polyaromatic hydrocarbon (PAH)-contaminated. Nevertheless, most of the sequences found were unaffiliated to cultured bacteria. The adaptation of the bacterial community mainly to PAH compounds demonstrated here and the fact that these bacterial communities are mainly unknown suggest that the Bizerte lagoon is an interesting environment to understand the capacity of bacteria to cope with some pollutants.     

Placental thrombomodulin expression in recurrent miscarriage

Background  

Early pregnancy loss can be associated with trophoblast insufficiency and coagulation defects. Thrombomodulin is an endothelial-associated anticoagulant protein involved in the control of hemostasis and inflammation at the vascular beds and it's also a cofactor of the protein C anticoagulant pathway.     

Reduced adiposity and improved insulin sensitivity in obese mice with antisense suppression of 4E-BP2 expression

To investigate the possible role of eukaryotic initiation factor 4E-binding protein-2 (4E-BP2) in metabolism and energy homeostasis, high-fat diet-induced obese mice were treated with a 4E-BP2-specific antisense oligonucleotide (ASO) or a control 4E-BP2 ASO at a dose of 25 mg/kg body wt or with saline twice a week for 6 wk. 4E-BP2 ASO treatment reduced 4E-BP2 levels by >75% in liver and white (WAT) and brown adipose (BAT) tissues. Treatment did not change food intake but lowered body weight by approximately 7% and body fat content by approximately 18%. Treatment decreased liver triglyceride (TG) content by >50%, normalized plasma glucose and insulin levels, and reduced glucose excursion during glucose tolerance test. 4E-BP2 ASO-treated mice showed >8.5% increase in metabolic rate, >40% increase in UCP1 levels in BAT, >45% increase in beta(3)-adrenoceptor mRNA, and 40-55% decrease in mitochondrial dicarboxylate carrier, fatty acid synthase, and diacylglycerol acyltransferase 2 mRNA levels in WAT. 4E-BP2 ASO-transfected mouse hepatocytes showed an increased fatty acid oxidation rate and a decreased TG synthesis rate. In addition, 4E-BP2 ASO-treated mice demonstrated approximately 60 and 29% decreases in hepatic glucose-6-phosphatase and phosphoenolpyruvate carboxykinase mRNA, respectively, implying decreased hepatic glucose output. Furthermore, increased phosphorylation of Akt(Ser473) in both liver and fat of 4E-BP2 ASO-treated mice and increased GLUT4 levels in plasma membrane in WAT of the ASO-treated mice were observed, indicating enhanced insulin signaling and increased glucose uptake as a consequence of reduced 4E-BP2 expression. These data demonstrate for the first time that peripheral 4E-BP2 plays an important role in metabolism and energy homeostasis.     

Reduction of JNK1 expression with antisense oligonucleotide improves adiposity in obese mice

To investigate the role of JNK1 in metabolism, male ob/ob and diet-induced obese mice were treated with a JNK1-specific antisense oligonucleotide (ASO) or control ASO at 25 mg/kg or saline twice/wk for 6 and 7 wk, respectively. JNK1 ASO reduced JNK1 mRNA and activity by 65-95% in liver and fat tissues in both models. Compared with controls, treatment with JNK1 ASO did not change food intake but lowered body weight, fat pad weight, and whole body fat content. The treatment increased metabolic rate. In addition, the treatment markedly reduced plasma cholesterol levels and improved liver steatosis and insulin sensitivity. These positive observations were accompanied by the following changes: 1) increased mRNA levels of AR-beta(3) and UCP1 by >60% in BAT, 2) reduced mRNA levels of ACC1, ACC2, FAS, SCD1, DGAT1, DGAT2, and RBP4 by 30-60% in WAT, and 3) reduced mRNA levels of ACC1, FAS, G-6-Pase, and PKCepsilon by 40-70% and increased levels of UCP2 and PPARalpha by more than twofold in liver. JNK1 ASO-treated mice demonstrated reduced levels of pIRS-1 Ser(302) and pIRS-1 Ser(307) and increased levels of pAkt Ser(473) in liver and fat in response to insulin. JNK1 ASO-transfected mouse hepatocytes showed decreased rates of de novo sterol and fatty acid synthesis and an increased rate of fatty acid oxidation. These results indicate that inhibition of JNK1 expression in major peripheral tissues can improve adiposity via increasing fuel combustion and decreasing lipogenesis and could therefore provide clinical benefit for the treatment of obesity and related metabolic abnormalities.     

Discovery and analysis of antisense oligonucleotide activity in cell culture

In the past decade antisense oligonucleotides (ASOs) have proven to be a useful tool for dissection of gene function in molecular cell biology (Koller, E., Gaarde, W. A., and Monia, B. P. (2000) Trends Pharm. Sci., 21, 142-148), and validation of gene targets in animal models (Crooke, S. T. (1998) Biotechnol. Gen. Eng. Rev. 15, 121-157), as well as a means for therapeutic treatment of human diseases (Bennett, C. F. (1999) Exp. Opin. Invest. Drugs 8, 237-253). An important step toward usage of ASOs in the described applications is identification of an active ASO. This article describes the underlying basis and means for achieving this goal in cell culture.     

Antisense inhibition of membrane-bound human interleukin-5 receptor-alpha chain does not affect soluble receptor expression and induces apoptosis in TF-1 cells

Binding of human interleukin-5 (HuIL-5) to its membrane-anchored receptor (IL-5R) triggers multiple signaling pathways, cellular proliferation, and maturational responses, as well as protection from apoptosis. In contrast, soluble forms of the HuIL-5R have been shown to inhibit IL-5 signaling and, therefore, may represent naturally occurring negative regulators of IL-5 function. Because of the central role of IL-5 in promoting eosinophilia and airway hyperresponsiveness in animal models of asthma, antisense oligonucleotides specific either for the membrane form alone or for sequences shared between both the membrane and soluble forms of the HuIL-5Ralpha ligand binding chain were designed. The activities of these oligonucleotides were characterized in IL-5R-expressing erythroleukemic TF-1 cells. Herein we report that an antisense oligonucleotide targeted to a sequence unique to the alternatively spliced membrane-bound form of the HuIL-5Ralpha chain has been developed that selectively inhibits membrane, but not soluble, mRNA isoform expression. Both this membrane-specific oligonucleotide and an antisense oligonucleotide targeted to sequence common to both membrane and soluble isoforms were found to potently suppress cell surface IL-5Ralpha levels and IL-5-mediated cell survival by inducing apoptosis similar to IL-5 withdrawal. Thus, these oligonucleotides represent unique genetic agents with therapeutic potential for diseases with an eosinophilic component.