Na(+)-dependent, active nucleoside transport in mouse spleen lymphocytes, leukemia cells, fibroblasts and macrophages, but not in equivalent human or pig cells; dipyridamole enhances nucleoside salvage by cells with both active and facilitated transport

Formycin B influx studies have shown that P388 and L1210 mouse leukemia cells, mouse L929 cells, mouse RAW 309 Cr.1 cells, LK35.2 mouse B-cell hybridoma cells and cultured mouse peritoneal macrophages express both Na(+)-dependent, active and nonconcentrative, facilitated nucleoside transport systems. In the mouse cell lines, active transport represented only a minor nucleoside transport component and was detected only by measuring formycin B uptake in the presence of dipyridamole or nitrobenzylthioinosine, strong inhibitors of facilitated, but not of active, nucleoside transport. Inhibition of facilitated transport resulted in the concentrative accumulation of formycin B in cells expressing active nucleoside transport. Concentrative formycin B accumulation was abolished by treatment of the cells with gramicidin or absence of Na+ in the extracellular medium and strongly inhibited by ATP depletion or ouabain treatment. Mouse macrophages accumulated formycin B to 70-times the extracellular concentration in the absence of dipyridamole during 90 min of incubation at 37 degrees C. Thus active transport represents a major nucleoside transport system of these cells, similarly as previously reported for mouse spleen lymphocytes. In contrast to the various types of mouse cells, active formycin B transport was not detected in human HeLa cells, human H9, Jurkat and CEM T lymphoidal cells and pig spleen lymphocytes. These cells expressed only facilitated nucleoside transport with kinetic properties similar to those of the facilitated transporters of other mammalian cells.     

Species-specific determinants in the IgG CH3 domain enable Fab-arm exchange by affecting the noncovalent CH3-CH3 interaction strength

A distinctive feature of human IgG4 is its ability to recombine half molecules (H chain and attached L chain) through a dynamic process termed Fab-arm exchange, which results in bispecific Abs. It is becoming evident that the process of Fab-arm exchange is conserved in several mammalian species, and thereby represents a mechanism that impacts humoral immunity more generally than previously thought. In humans, Fab-arm exchange has been attributed to the IgG4 core-hinge sequence (226-CPSCP-230) in combination with unknown determinants in the third constant H chain domain (CH3). In this study, we investigated the role of the CH3 domain in the mechanism of Fab-arm exchange, and thus identified amino acid position 409 as the critical CH3 determinant in human IgG, with R409 resulting in exchange and K409 resulting in stable IgG. Interestingly, studies with IgG from various species showed that Fab-arm exchange could not be assigned to a common CH3 domain amino acid motif. Accordingly, in rhesus monkeys (Macaca mulatta), aa 405 was identified as the CH3 determinant responsible (in combination with 226-CPACP-230). Using native mass spectrometry, we demonstrated that the ability to exchange Fab-arms correlated with the CH3-CH3 dissociation constant. Species-specific adaptations in the CH3 domain thus enable Fab-arm exchange by affecting the inter-CH3 domain interaction strength. The redistribution of Ag-binding domains between molecules may constitute a general immunological and evolutionary advantage. The current insights impact our view of humoral immunity and should furthermore be considered in the design and evaluation of Ab-based studies and therapeutics.     

Global Distribution of Bartonella Infections in Domestic Bovine and Characterization of Bartonella bovis Strains Using Multi-Locus Sequence Typing

Bartonella bovis is commonly detected in cattle. One B. bovis strain was recently isolated from a cow with endocarditis in the USA, suggesting its role as an animal pathogen. In the present study, we investigated bartonella infections in 893 cattle from five countries (Kenya, Thailand, Japan, Georgia, and Guatemala) and 103 water buffaloes from Thailand to compare the prevalence of the infection among different regions and different bovid hosts. We developed a multi-locus sequence typing (MLST) scheme based on nine loci (16S rRNA, gltA, ftsZ, groEL, nuoG, ribC, rpoB, ssrA, and ITS) to compare genetic divergence of B. bovis strains, including 26 representatives from the present study and two previously described reference strains (one from French cows and another from a cow with endocarditis in the USA). Bartonella bacteria were cultured in 6.8% (7/103) of water buffaloes from Thailand; all were B. bovis. The prevalence of bartonella infections in cattle varied tremendously across the investigated regions. In Japan, Kenya, and the Mestia district of Georgia, cattle were free from the infection; in Thailand, Guatemala, and the Dusheti and Marneuli districts of Georgia, cattle were infected with prevalences of 10–90%. The Bartonella isolates from cattle belonged to three species: B. bovis (n=165), B. chomelii (n=9), and B. schoenbuchensis (n=1), with the latter two species found in Georgia only. MLST analysis suggested genetic variations among the 28 analyzed B. bovis strains, which fall into 3 lineages (I, II, and III). Lineages I and II were found in cattle while lineage III was restricted to water buffaloes. The majority of strains (17/28), together with the strain causing endocarditis in a cow in the USA, belonged to lineage I. Further investigations are needed to determine whether B. bovis causes disease in bovids.     

Automated mode-of-action detection by metabolic profiling

Rapid classification and identification of the mode-of-action of bioactive compounds applied to plants can be achieved by a robust and easy-to-use metabolic-profiling method. This method uses artificial neural network analysis of one-dimensional proton NMR spectra of aqueous plant extracts to rapidly classify changes in the total metabolic profile caused by application of crop protection chemicals.     

Derivation of human embryonic stem cells at the Center of Regenerative Medicine in Barcelona

We report here the legislative issues related to embryo research and human embryonic stem cell (hESC) research in Spain and the derivation of nine hESC lines at the Center of Regenerative Medicine in Barcelona. You can find the information for obtaining our lines for research purposes at blc@cmrb.eu.     

Enhancement of total lipid yield by nitrogen,carbon, and iron supplementation in isolated microalgae

The biochemical contents and biodiesel production ability of three microalgal strains grown under different sodium nitrate, sodium carbonate, and ferric ammonium citrate (iron) levels were investigated. The highest biomass and lipid contents were found in Scenedesmus sp., Chlorella sp., and Chlamydomonas sp. when grown in normal BG‐11 containing sodium carbonate concentration at 0.03 g · L^?1, and in normal BG‐11 containing iron concentration (IC) at 0.009 or 0.012 g · L^?1. Increasing the sodium nitrate level increased the biomass content, but decreased the lipid content in all three microalgae. Among the three microalgae, Scenedesmus sp. showed the highest total lipid yield of 0.69 g · L^?1 under the IC of 0.012 g · L^?1. Palmitic and oleic acids were the major fatty acids of Scenedesmus sp. and Chlamydomonas sp. lipids. On the other hand, Chlorella sp. lipids were rich in palmitic, oleic, and linolenic acids, and henceforth contributing to poor biodiesel properties below the standard limits. The three isolated strains had a potential for biodiesel production. Nevertheless, Scenedesmus sp. from stone quarry pond water was the most suitable source for biodiesel production with tolerance toward the high concentration of sodium carbonate without the loss of its biodiesel properties.     

Association of adenosine deaminase with erythrocyte and platelet plasma membrane: an immunological study using light and electron microscopy

Adenosine deaminase has been localized in the plasma membrane of erythrocytes and platelets by means of immunological techniques using light and electron microscopy with cells in suspension. In erythrocytes, adenosine deaminase is associated with the external side of the plasma membrane. In platelets, the enzyme is associated with the external side of the plasma membrane, which is known to extend through the canalicular system of these cells. These results confirm our previous findings, based on biochemical studies, concerning the attachment of the enzyme to cell membranes.     

Increase in local protein concentration by field-inversion gel electrophoresis

Background  

Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and non-specific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein separation efficiency. Enhancement of protein separation efficiency was investigated by implementing pulsed field-inversion gel electrophoresis (FIGE).     

Initial treatment choices to achieve sustained response in major depression: a systematic review and network meta‐analysis

Major depression is often a relapsing disorder. It is therefore important to start its treatment with therapies that maximize the chance of not only getting the patients well but also keeping them well. We examined the associations between initial treatments and sustained response by conducting a network meta‐analysis of randomized controlled trials (RCTs) in which adult patients with major depression were randomized to acute treatment with a psychotherapy (PSY), a protocolized antidepressant pharmacotherapy (PHA), their combination (COM), standard treatment in primary or secondary care (STD), or pill placebo, and were then followed up through a maintenance phase. By design, acute phase treatment could be continued into the maintenance phase, switched to another treatment or followed by discretionary treatment. We included 81 RCTs, with 13,722 participants. Sustained response was defined as responding to the acute treatment and subsequently having no depressive relapse through the maintenance phase (mean duration: 42.2±16.2 weeks, range 24‐104 weeks). We extracted the data reported at the time point closest to 12 months. COM resulted in more sustained response than PHA, both when these treatments were continued into the maintenance phase (OR=2.52, 95% CI: 1.66‐3.85) and when they were followed by discretionary treatment (OR=1.80, 95% CI: 1.21‐2.67). The same applied to COM in comparison with STD (OR=2.90, 95% CI: 1.68‐5.01 when COM was continued into the maintenance phase; OR=1.97, 95% CI: 1.51‐2.58 when COM was followed by discretionary treatment). PSY also kept the patients well more often than PHA, both when these treatments were continued into the maintenance phase (OR=1.53, 95% CI: 1.00‐2.35) and when they were followed by discretionary treatment (OR=1.66, 95% CI: 1.13‐2.44). The same applied to PSY compared with STD (OR=1.76, 95% CI: 0.97‐3.21 when PSY was continued into the maintenance phase; OR=1.83, 95% CI: 1.20‐2.78 when PSY was followed by discretionary treatment). Given the average sustained response rate of 29% on STD, the advantages of PSY or COM over PHA or STD translated into risk differences ranging from 12 to 16 percentage points. We conclude that PSY and COM have more enduring effects than PHA. Clinical guidelines on the initial treatment choice for depression may need to be updated accordingly.