Phosphodiesterase 7A1 is expressed in human CD4+ naïve T cells at higher levels than in CD4+ memory cells and is not required during their CD3/CD28-dependent activation

PDE7A1 is a cAMP-hydrolyzing phosphodiesterase expressed in lymphoid tissue, where its possible role during T cell activation remains unclear. We have characterized the functional relevance of PDE7A1 in the na?ve (CD4+CD45RA+) and memory (CD4+CD45RO+) subsets of human peripheral CD4+ T cells during CD3/CD28-dependent stimulation. Our results indicate that PDE7A1 is expressed in resting na?ve CD4+ T cells at higher levels than in the corresponding memory cells and that levels of PDE7A1 mRNA are not upregulated upon CD3/CD28 mediated stimulation of these T cell subsets. Treatment with a selective inhibitor of PDE7A1 does not impair CD3/CD28 induced activation of na?ve or memory CD4+ T cells, nor does it increase intracellular cAMP in CD4+ T cells. We conclude that PDE7A1 is not required during CD3/CD28-dependent activation of na?ve and memory CD4+ T cells, but cannot rule out other regulatory roles of PDE7A1 during maturation of CD4+ T cells.     

Modulation of paraoxonase 1 and 3 expression after moderate exercise training in the rat

Paraoxonases (PONs) are a small family of antioxidant enzymes whose antiatherogenic activity is well known. The aim of the present study was the evaluation of the effects of moderate aerobic training on their expression using a rat model. In order to discriminate between PON1 and PON3 enzymatic activity, we took advantage of some differences in their substrate preferences. PON1 and PON3 enzymatic activities and their protein levels were analyzed in plasma and in liver microsomes, and their mRNA levels in the liver. Exercise training did not affect PON1 expression or enzymatic activity but increased PON3 mRNA, protein levels, and enzymatic activity. Training also induced variations in plasma membrane composition, including an increase in polyunsaturated and a decrease in mono- and di-unsaturated fatty acids. On the other hand, acute exercise inhibited PON activities while increasing PON3 protein content in liver microsomes and reversing the relative composition in mono-, di-, and poly-unsaturated fatty acids, suggesting that physical stress, by altering membrane composition, may impair PON release from liver membranes. In conclusion, we documented, for the first time, the presence of PON3 in rat serum and, notably, found that the upregulation of PON3, rather than PON1, appears to be associated with physical training.     

Biologic effects of prolonged exposure to ELF electromagnetic fields in rats. I. 50 Hz electric fields

A three-year investigation was conducted on the biological effects of high-intensity electric field exposures of rats for up to 18% of their life span. Two hundred and forty adult male rats, divided into groups of 20 animals each, were exposed at ground potential for 8 h/ day at 25-kV/m and 100-kV/m 50-Hz electric fields or were sham exposed for 280, 440, and 1240 h. The corresponding ages at sacrifice were 140, 164, and 315 days. An additional group of 40 rats was investigated under similar experimental conditions after 440 h of exposure at floating potential. Independent of exposure duration, mode of grounding, and field strength, no statistical differences in body weight, morphology, and histology of the liver, heart, mesenteric lymph nodes, and blood variables (hematology and serum chemistry) were found in comparison with sham-exposed animals. Plasma levels of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (TS)at sacrifice varied widely among experimental animals in the same group but did not differ in exposed compared with sham-exposed rats. A nonsignificant tendency toward a decrease in the testes/body weight ratio was found after 1240 h of exposure. Microscopic examination of a large number of specimens showed no quantitative or qualitative statistical differences in testes alterations either among exposed animals or between exposed and their corresponding sham-exposed groups. We conclude that 50-Hz electric field exposure, even of long duration at very high field strengths, does not induce harmful effects on tissues with high cellular turnover rates and does not impair the reproductive function of rats. Moreover, after exposure, all variables investigated were well within the normal physiological range. © 1993 Wiley-Liss. Inc.     

Speciation of ionic alkyllead compounds in human urine by gas chromatography–mass spectrometry after butylation through a Grignard reaction

One analytical procedure for the determination of ionic alkyllead in human urine has been studied. The system consists of the extraction of Me₃Pb⁺, Et₃Pb⁺ and Pb²⁺ at pH 9.0 with diethyldithiocarbamate to an organic phase. Then, the ionic compounds are butylated with BuMgCl and the final organic solution is analyzed by GC–MS–SIM. The elimination of both foam and gels in the extraction step and the general procedure for the urine are discussed. The recovery of compounds ranges from 105.1% for Me₃Pb⁺ to 97.2% for Et₃Pb⁺ using hexane as extracting agent and detection limits are 18.4 pg/ml of Me₃Pb⁺ and 19.2 pg/ml of Et₃Pb⁺ in urine. The speciation of ionic alkylleads in the urine of a petrol station worker showed a value of 27.9 pg/ml of Me₃Pb⁺ in urine and Et₃Pb⁺ was below the detection limit.     

Regulation of the low-CO2-inducible polypeptides in Chlamydomonas reinhardtii

Polypeptides of 21, 36 and 37 kDa are induced in the unicellular green alga Chlamydomonas reinhardtii Dang. when cells are transferred from high (2%) to low (0.03%) CO₂ concentrations. The synthesis of these polypeptides is correlated with the induction of the CO₂-concentrating mechanism. In this work we studied the effect of the growth conditions on the synthesis of these polypeptides with the aim of clarifying whether the induction of all three of these low-CO₂-inducible polypeptides requires the same environmental factor. Our results showed that induction of the 21- and 36-kDa polypeptides under low-CO₂ conditions occurred only in the light, while the 37-kDa periplasmic carbonic anhydrase (EC 4.2.1.1) was induced in light, darkness, and in both synchronous and asynchronous cultures. In addition, induction of these polypeptides appeared to be determined more by the O₂/CO₂ ratio than by the CO₂ concentrations. None of these polypeptides could be induced in either of two different mutants of C. reinhardtii, one lacking ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) and the other with inactive enzyme. Our results indicate that the 21- and 36-kDa polypeptides are regulated by a mechanism different from that controlling the 37-kDa polypeptide.Abbreviations pCA (periplasmic) carbonic anhydrase - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - TAP Trisacetate phosphate medium The authors thank Prof. M. Spalding (Iowa State University, USA) for providing antisera to LIP-21 and LIP-36. We thank Prof. S. Bartlett and Dr. J. Moroney (Louisiana State University, USA) for providing antibodies to C. reinhardtii, Rubisco and 37-kDa pCA, respectively. This work was supported by the Instituto Tecnologico de Canarias.     

Effect of Chronic Ethanol Treatment on the γ-Aminobutyric Acid-Mediated Enhancement of [3H]Flunitrazepam Binding in Rat Cortex and Hippocampus

The mechanism by which ethanol affects the gamma-aminobutyric acid (GABA)/benzodiazepine complex is not clear. It is known that ethanol enhances the Cl- influx mediated by the GABAA receptor complex, and although chronic ethanol administration does not change the KD or Bmax for [3H]flunitrazepam binding, some reports have suggested that it could modify the modulation of benzodiazepine binding produced by GABA. In the present work, we studied the effect of chronic ethanol treatment on the modulation by GABA of [3H]flunitrazepam binding, using light microscopic autoradiography. This technique allows the measurement of densities of benzodiazepine receptors in different brain areas, the visual cortex and hippocampus, which appear to constitute the anatomical support for the behavioral and physiological responses affected by ethanol. We found enhancement of benzodiazepine binding by GABA at concentrations of greater than 10(-6) M for the various cortical and hippocampal areas studied from both control and ethanol-treated animals; this enhancement peaked at 10(-4) M GABA but decreased at 10(-3) M GABA. We found a clear effect of ethanol treatment on the modulatory properties of GABAA receptor, in both cortex and hippocampus, although only in cortex were the differences statistically significant between control and ethanol-treated animals.     

Evidence for a protein transported through the secretory pathway en route to the higher plant chloroplast

In contrast to animal and fungal cells, green plant cells contain one or multiple chloroplasts, the organelle(s) in which photosynthetic reactions take place. Chloroplasts are believed to have originated from an endosymbiotic event and contain DNA that codes for some of their proteins. Most chloroplast proteins are encoded by the nuclear genome and imported with the help of sorting signals that are intrinsic parts of the polypeptides. Here, we show that a chloroplast-located protein in higher plants takes an alternative route through the secretory pathway, and becomes N-glycosylated before entering the chloroplast.     

Effect of Ethanol Treatment on Rate and Equilibrium Constants for [3H]Muscimol Binding to Rat Brain Membranes: Alteration of Two Affinity States of the GABAA Receptor

Abstract: Equilibrium binding curves were biphasic in control and ethanol-treated rats. [³H]Muscimol binds to sites of high ( K _DA of ∼10 n M ) and low ( K _DB of ∼0.3–0.4 µ M ) affinity. Chronic ethanol treatment produced a decrease in B _maxA value, and the hyperbolic binding profiles were progressively affected by the chronic and in vitro ethanol treatments, with most of this effect corresponding to the high-affinity site. IC₅₀ and K _i values were calculated for several competing ligands, using membranes from both control and ethanol-treated animals. The association and dissociation curves were also biphasic, using a radioligand concentration precluding a significant occupancy of the low-affinity sites, which suggests the existence of two forms or affinity states of the monoliganded receptor. Chronic ethanol treatment did not produce changes in the values of the dissociation rate constants (fast and slow phases). By contrast, we report for the first time a decrease in the values of the association rate constants, with this decrease being higher for the slow phase. Consequently, the dissociation equilibrium constants are two times higher in chronically ethanol-treated animals for both phases.     

Characterization of BCAM0224, a Multifunctional Trimeric Autotransporter from the Human Pathogen Burkholderia cenocepacia

Members of the trimeric autotransporter adhesin (TAA) family play a crucial role in adhesion of Gram-negative pathogens to host cells. Moreover, these proteins are multifunctional virulence factors involved in several other biological traits, including invasion into host cells and evasion of the host immune system. In cystic fibrosis epidemic Burkholderia cenocepacia strain J2315, we identified a unique TAA (BCAM0224)-encoding gene, previously described as being implicated in virulence. Here, we characterized this multifunctional protein, trying to establish its role in B. cenocepacia pathogenicity. We show that BCAM0224 occurs on the bacterial surface and adopts a trimeric conformation. Furthermore, we demonstrated that BCAM0224 is needed for earlier stages of biofilm formation and is required for swarming motility. In addition, BCAM0224 plays an important role in evasion of the human innate immune system, providing resistance against the bactericidal activity of serum via the complement classical pathway. Finally, BCAM0224 mediates bacterial adhesion to and invasion of cultured human bronchial epithelial cells. Together, these data reveal the high versatility of the BCAM0224 protein as a virulence factor in the pathogenic bacterium B. cenocepacia.