Biochemical Characterization of Branched Chain Amino Acids Uptake in Trypanosoma cruzi

Trypanosoma cruzi is the etiological agent of Chagas disease. During its life cycle, it alternates among vertebrate and invertebrate hosts. Metabolic flexibility is a main biochemical characteristic of this parasite, which is able to obtain energy by oxidizing a variety of nutrients that can be transported from the extracellular medium. Moreover, several of these metabolites, more specifically amino acids, have a variety of functions beyond being sources of energy. Branched chain amino acids (BCAA), beyond their role in ATP production, are involved in sterol biosynthesis; for example, leucine is involved as a negative regulator of the parasite differentiation process occurring in the insect midgut. BCAA are essential metabolites in most nonphotosynthetic eukaryotes, including trypanosomes. In view of this, the metabolism of BCAA in T. cruzi depends mainly on their transport into the cell. In this work, we kinetically characterized the BCAA transport in T. cruzi epimastigotes. Our data point to BCAA as being transported by a single saturable transport system able to recognize leucine, isoleucine and valine. In view of this, we used leucine to further characterize this system. The transport increased linearly with temperature from 10 to 45 °C, allowing the calculation of an activation energy of 51.30 kJ/mol. Leucine uptake was an active process depending on ATP production and a H⁺ gradient, but not on a Na⁺ or K⁺ gradient at the cytoplasmic membrane level.     

Multimodal optical analysis of meningioma and comparison with histopathology

Meningioma is the most frequent primary central nervous system tumor. The risk of recurrence and the prognosis are correlated with the extent of the resection that ideally encompasses the infiltrated dura mater and, if required, the infiltrated bone. No device can deliver real‐time intraoperative histopathological information on the tumor environment to help the neurosurgeon to achieve a gross total removal. This study assessed the abilities of nonlinear microscopy to provide relevant and real‐time data to help resection of meningiomas. Nine human meningioma samples (four World Health Organization Grade I, five Grade II) were analyzed using different optical modalities: spectral analysis and imaging, lifetime measurements, fluorescence lifetime imaging microscopy, fluorescence emitted under one‐ and two‐photon excitation and the second‐harmonic generation signal imaging using a multimodal setup. Nonlinear microscopy produced images close to histopathology as a gold standard. The second‐harmonic generation signal delineated the collagen background and two‐photon fluorescence underlined cell cytoplasm. The matching between fluorescence images and Hematoxylin and Eosin staining was possible in all cases. Grade I meningioma emitted less autofluorescence than Grade II meningioma and Grade II meningioma exhibited a distinct lifetime value. Autofluorescence was correlated with the proliferation rates and seemed to explain the observed differences between Grade I and II meningiomas. This preliminary multimodal study focused on human meningioma samples confirms the potential of tissue autofluorescence analysis and nonlinear microscopy in helping intraoperatively neurosurgeons to reach the actual boundaries of the tumor infiltration.

Correspondence between H&E staining (top pictures) and the two‐photon fluorescence imaging (bottom pictures)     

Infrastructure for the life sciences: design and implementation of the UniProt website

Background  

The UniProt consortium was formed in 2002 by groups from the Swiss Institute of Bioinformatics (SIB), the European Bioinformatics Institute (EBI) and the Protein Information Resource (PIR) at Georgetown University, and soon afterwards the website was set up as a central entry point to UniProt resources. Requests to this address were redirected to one of the three organisations' websites. While these sites shared a set of static pages with general information about UniProt, their pages for searching and viewing data were different. To provide users with a consistent view and to cut the cost of maintaining three separate sites, the consortium decided to develop a common website for UniProt. Following several years of intense development and a year of public beta testing, the domain was switched to the newly developed site described in this paper in July 2008.     

HZE ⁵⁶Fe-ion irradiation induces endothelial dysfunction in rat aorta: role of xanthine oxidase

Ionizing radiation has been implicated in the development of significant cardiovascular complications. Since radiation exposure is associated with space exploration, astronauts are potentially at increased risk of accelerated cardiovascular disease. This study investigated the effect of high atomic number, high-energy (HZE) iron-ion radiation on vascular and endothelial function as a model of space radiation. Rats were exposed to a single whole-body dose of iron-ion radiation at doses of 0, 0.5 or 1 Gy. In vivo aortic stiffness and ex vivo aortic tension responses were measured 6 and 8 months after exposure as indicators of chronic vascular injury. Rats exposed to 1 Gy iron ions demonstrated significantly increased aortic stiffness, as measured by pulse wave velocity. Aortic rings from irradiated rats exhibited impaired endothelial-dependent relaxation consistent with endothelial dysfunction. Acute xanthine oxidase (XO) inhibition or reactive oxygen species (ROS) scavenging restored endothelial-dependent responses to normal. In addition, XO activity was significantly elevated in rat aorta 4 months after whole-body irradiation. Furthermore, XO inhibition, initiated immediately after radiation exposure and continued until euthanasia, completely inhibited radiation-dependent XO activation. ROS production was elevated after 1 Gy irradiation while production of nitric oxide (NO) was significantly impaired. XO inhibition restored NO and ROS production. Finally, dietary XO inhibition preserved normal endothelial function and vascular stiffness after radiation exposure. These results demonstrate that radiation induced XO-dependent ROS production and nitroso-redox imbalance, leading to chronic vascular dysfunction. As a result, XO is a potential target for radioprotection. Enhancing the understanding of vascular radiation injury could lead to the development of effective methods to ameliorate radiation-induced vascular damage.     

Biomass changes and trophic amplification of plankton in a warmer ocean

Ocean warming can modify the ecophysiology and distribution of marine organisms, and relationships between species, with nonlinear interactions between ecosystem components potentially resulting in trophic amplification. Trophic amplification (or attenuation) describe the propagation of a hydroclimatic signal up the food web, causing magnification (or depression) of biomass values along one or more trophic pathways. We have employed 3‐D coupled physical‐biogeochemical models to explore ecosystem responses to climate change with a focus on trophic amplification. The response of phytoplankton and zooplankton to global climate‐change projections, carried out with the IPSL Earth System Model by the end of the century, is analysed at global and regional basis, including European seas (NE Atlantic, Barents Sea, Baltic Sea, Black Sea, Bay of Biscay, Adriatic Sea, Aegean Sea) and the Eastern Boundary Upwelling System (Benguela). Results indicate that globally and in Atlantic Margin and North Sea, increased ocean stratification causes primary production and zooplankton biomass to decrease in response to a warming climate, whilst in the Barents, Baltic and Black Seas, primary production and zooplankton biomass increase. Projected warming characterized by an increase in sea surface temperature of 2.29 ± 0.05 °C leads to a reduction in zooplankton and phytoplankton biomasses of 11% and 6%, respectively. This suggests negative amplification of climate driven modifications of trophic level biomass through bottom‐up control, leading to a reduced capacity of oceans to regulate climate through the biological carbon pump. Simulations suggest negative amplification is the dominant response across 47% of the ocean surface and prevails in the tropical oceans; whilst positive trophic amplification prevails in the Arctic and Antarctic oceans. Trophic attenuation is projected in temperate seas. Uncertainties in ocean plankton projections, associated to the use of single global and regional models, imply the need for caution when extending these considerations into higher trophic levels.     

A novel DAD type and folic acid conjugated fluorescent monomer as a targeting probe for imaging of folate receptor overexpressed cells

We describe a modification and post‐functionalization technique for a donor–acceptor–donor type monomer; 6‐(4,7‐bis(2,3‐dihydrothieno[3,4‐b][1,4]dioxin‐5‐yl)‐2H‐benzo[d][1,2, 3]triazol‐2‐yl)hexan‐1‐amine. Folic acid was attached to the fluorescent structure. The conjugation was confirmed via NMR and Fourier transform infrared analyses. Cytotoxicity was investigated and the comparison of association of targeted monomeric structures in tumor cells was monitored via fluorescence microscopy. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:952–959, 2014     

Thyrotrophin-releasing hormone inactivation by human postmortem brain

The mechanisms of inactivation of thyrotrophin-releasing hormone (TRH) by peptidases in several areas of normal human postmortem brain have been investigated by radioimmunoassay and high-performance liquid chromatography. Of the several brain regions studied, the cerebral cortex (Brodman's area, BA10) had the highest TRH-degrading activity in both subcellular fractions. Deamidated-TRH (TRH-OH) was the only product formed by the soluble fraction whereas the histidyl-proline diketopiperazine, cyclo(His-Pro), and a small amount of TRH-OH were formed by the particulate fraction. Several centrally acting TRH analogues showed varying degrees of resistance to degradation by the peptidases in the two fractions, the most stable analogue being RX77368 (pGlu-His-3,3'-dimethyl(ProNH2]. Areas of human postmortem brain appear to contain two of the enzymes capable of degrading TRH, a proline endopeptidase forming TRH-OH and a pyroglutamyl aminopeptidase forming cyclo(His-Pro). The use of the assay procedures in further studies on the inactivation of TRH by peptidases from brain areas of patients with neurological disorders may provide complementary information on the dynamics of TRH in these disorders. The stability of the centrally acting TRH analogues may prove useful in examining their therapeutic potential.     

In vitro and in vivo mutagenicity studies with airborne particulate extracts

The contribution of nitro compounds to airborne particulate mutagenicity was studied with Salmonella typhimurium strains TA98, TA98NR, TA98/1,8DNP6. The results obtained indicate that nitropyrenes play a minor role in air particulate mutagenicity. Seasonal variations indicate a relatively greater contribution of nitro compounds to the mutagenicity of spring and summer samples. Fractionation of extracts into acidic, neutral and basic components shows that neutral compounds account for about two-thirds of the total mutagenic activity. Attempts to extract mutagens adsorbed onto particulate matter with aqueous media were almost completely negative. No significant mutagenicity was detected in urine and faecal extracts and in plasma samples of Sprague-Dawley rats treated with air particulate extracts at 80 mg/kg either per os or by i.p. injection. Negative results were obtained in the micronucleus test with Swiss mice treated at 200 and 400 mg/kg (twice by i.p. injection). A significant decrease in liver aminopyrine-N-demethylase was observed in Swiss mice injected with air particulate extracts or its basic and neutral fractions. In vitro experiments suggest a direct interaction of test materials with microsomal cytochrome P-450.