The advantage of laser‐capture microdissection over whole tissue analysis in proteomic profiling studies

Laser‐capture microdissection (LCM) offers a reliable cell population enrichment tool and has been successfully coupled to MS analysis. Despite this, most proteomic studies employ whole tissue lysate (WTL) analysis in the discovery of disease biomarkers and in profiling analyses. Furthermore, the influence of tissue heterogeneity in WTL analysis, nor its impact in biomarker discovery studies have been completely elucidated. In order to address this, we compared previously obtained high resolution MS data from a cohort of 38 breast cancer tissues, of which both LCM enriched tumor epithelial cells and WTL samples were analyzed. Label‐free quantification (LFQ) analysis through MaxQuant software showed a significantly higher number of identified and quantified proteins in LCM enriched samples (3404) compared to WTLs (2837). Furthermore, WTL samples displayed a higher amount of missing data compared to LCM both at peptide and protein levels (p‐value < 0.001). 2D analysis on co‐expressed proteins revealed discrepant expression of immune system and lipid metabolisms related proteins between LCM and WTL samples. We hereby show that LCM better dissected the biology of breast tumor epithelial cells, possibly due to lower interference from surrounding tissues and highly abundant proteins. All data have been deposited in the ProteomeXchange with the dataset identifier PXD002381 ( http://proteomecentral.proteomexchange.org/dataset/PXD002381 ).     

UV-B treated algae exhibiting different responses as a food source for Daphnia magna

The response of green algae Scenedesmus quadricauda and Selenastrumcapricornutum to different levels of UV-B radiation was studied.Daphnia magna, the next step in the food web, was fed with UV-Btreated algae. UV-B radiation induced the synthesis of UV absorbingsubstances and photosynthetic pigments, enhanced the level ofrespiratory potential and suppressed growth of the treated algae.Biomass production in S. quadricauda was higher than in S. capricornutum,but the production of photosynthetic pigments and relative amountsof UV absorbing substances as well as respiratory potentialwere more pronounced in S. capricornutum. A short-term feedingexperiment with D. magna showed that in general, females ingestedhigher numbers of cells of algae treated with a high UV-B dose,but the biomass of ingested algae did not differ significantly.The exception was S. capricornutum treated with a high UV-Bdose; a lower biomass was ingested, probably reflecting thesmaller UV-treated cells.     

Thrombin inhibitors built on an azaphenylalanine scaffold

A series of azaphenylalanine derivatives were investigated as novel thrombin inhibitors based on the prodrug principle. By systematic structural modifications we have identified optimal groups for this series that led us to potent inhibitors of thrombin incorporating the benzamidine fragment at the P1 position, and their potentially orally active benzamidoxime prodrugs. The binding modes in the thrombin active site of two representative compounds were identified by X-ray crystallographic analysis.     

Role of platelet derived endothelial cell growth factor/thymidine phosphorylase in fluoropyrimidine sensitivity and potential role of deoxyribose-1-phosphate

Thymidine phosphorylase (TP) catalyzes the phosphorolytic cleavage of thymidine (TdR) to thymine and deoxyribose-1-phosphate (dR-1-P). TP, which is overexpressed in a wide variety of solid tumors, is involved in the activation and inactivation of fluoropyrimidines. We investigated the role of TP in 5'-deoxy-5-fluorouridine (5'DFUR), 5-fluorouracil (5FU) and trifluorothymidine (TFT) sensitivity. TP had no effect on TFT while it activated 5'DFUR and to a lesser extent 5FU. In order to provide an explanation for this difference in activation of 5'DFUR and 5FU, we studied the role of the 5FU co-substrate, dR-1-P, needed for its activation.     

Changes in particulate and dissolved organic matter in nutrient-enriched enclosures from an area influenced by mucilage: the northern Adriatic Sea

This study examined the partitioning of organic matter intoparticulate organic carbon (POC) and dissolved organic carbon(DOC) pools in nutrient-enriched enclosures containing naturalplankton from the Gulf of Trieste (northern Adriatic), an areaaffected by mucilage. The strategy of nutrient additions wasto introduce a pulse of new nutrients in concentrations thatmimic natural inputs and to survey community structure and organicmatter fluxes long enough so that plankton became nutrient-limited.Maximal bacterial biomass attained roughly double the initialvalue, while autotrophic carbon increased by nearly an orderof magnitude. The microflagellate-dominated community releasedmore DOC per unit biomass (5.5 ± 7.2 to 50.6 ±28.0 µg C µg Chl a^-1 day^-1 versus 3.4 ± 3.4to 10.8 ± 4.6 µg C µg Chl a^-1 day^-1 for diatom-dominatedphytoplankton), POC increase was modest (~300 µg C l^-1)and there was little change in DOC. Organic carbon partitioningduring two experiments in which diatoms prevailed was dominatedby POC (>800 µg C l^-1) in the exponential growth phasewith an increasing contribution of particulate carbohydratesthat paralleled gradual nutrient depletion. Transition to thestationary phase and the decay of autotrophic communities wereaccompanied by the net accumulation of a carbohydrate-rich DOC.     

Impact of urea on water structure: a clue to its properties as a denaturant?

A new investigation of the structure of urea-water solutions at a mole ratio of 1 urea to 4 water molecules is described. Neutron diffraction is used in conjunction with isotope labelling on the water and urea hydrogen atoms and on the nitrogen atom of urea. The diffraction data are analysed using the empirical potential structure refinement procedure to yield a set of site-site radial distribution functions and spatial density functions that are consistent with the diffraction data. The results are discussed in relation to recent and past X-ray and neutron diffraction experiments and theoretical studies of this system. It is found that urea incorporates readily into water, forming pronounced hydrogen bonds with water at both the amine and carbonyl headgroups. In addition the urea also hydrogen bonds to itself, forming chains or clusters consisting of up to approximately 60 urea molecules in a cluster. There, is however, little or no evidence of urea segregating itself from water, in marked contrast to a recent study of the methanol-water system. This behaviour is discussed in the context of the great propensity of urea to effect protein denaturation.     

IL-2 activation of NK cells: involvement of MKK1/2/ERK but not p38 kinase pathway

IL-2 stimulates extracellular signal-regulated protein kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) in various immune cell populations. The functional roles that these kinases play are still unclear. In this study, we examined whether MAPK kinase (MKK)/ERK and p38 MAPK pathways are necessary for IL-2 to activate NK cells. Using freshly isolated human NK cells, we established that an intact MKK/ERK pathway is necessary for IL-2 to activate NK cells to express at least four known biological responses: LAK generation, IFN-gamma secretion, and CD25 and CD69 expression. IL-2 induced ERK activation within 5 min. Treatment of NK cells with a specific inhibitor of MKK1/2, PD98059, during the IL-2 stimulation blocked in a dose-dependent manner each of four sequelae, with inhibition of lymphokine-activated killing induction being least sensitive to MKK/ERK pathway blockade. Activation of p38 MAPK by IL-2 was not detected in NK cells. In contrast to what was observed by others in T lymphocytes, SB203850, a specific inhibitor of p38 MAPK, did not inhibit IL-2-activated NK functions. This data indicate that p38 MAPK activation was not required for IL-2 to activate NK cells for the four functions examined. These results reveal selective signaling differences between NK cells and T lymphocytes; in NK cells, the MKK/ERK pathway and not p38 MAPK plays a critical positive regulatory role during activation by IL-2.     

Musical Minds: Attentional Blink Reveals Modality-Specific Restrictions

BackgroundFormal musical training is known to have positive effects on attentional and executive functioning, processing speed, and working memory. Consequently, one may expect to find differences in the dynamics of temporal attention between musicians and non-musicians. Here we address the question whether that is indeed the case, and whether any beneficial effects of musical training on temporal attention are modality specific or generalize across sensory modalities.Conclusion/SignificanceAB magnitude within one modality can generalize to another modality, but this turns out not to be the case for every individual. Formal musical training seems to have a domain-general, but modality-specific beneficial effect on selective attention. The results fit with the idea that a major source of attentional restriction as reflected in the AB lies in modality-specific, independent sensory systems rather than a central amodal system. The findings demonstrate that individual differences in AB magnitude can provide important information about the modular structure of human cognition.     

Vascular Pattern Analysis for the Prediction of Clinical Behaviour in Pheochromocytomas and Paragangliomas

Pheochromocytomas (PCCs) are neuroendocrine tumors arising from chromaffin cells of the adrenal medulla. Related tumors that arise from the paraganglia outside the adrenal medulla are called paragangliomas (PGLs). PCC/PGLs are usually benign, but approximately 17% of these tumors are malignant, as defined by the development of metastases. Currently, there are no generally accepted markers for identifying a primary PCC or PGL as malignant. In 2002, Favier et al. described the use of vascular architecture for the distinction between benign and malignant primary PCC/PGLs. The aim of this study was to validate the use of vascular pattern analysis as a test for malignancy in a large series of primary PCC/PGLs. Six independent observers scored a series of 184 genetically well-characterized PCCs and PGLs for the CD34 immunolabeled vascular pattern and these findings were correlated to the clinical outcome. Tumors were scored as malignant if an irregular vascular pattern was observed, including vascular arcs, parallels and networks, while tumors with a regular pattern of short straight capillaries were scored as benign. Mean sensitivity and specificity of vascular architecture, as a predictor of malignancy was 59.7% and 72.9%, respectively. There was significant agreement between the 6 observers (mean κ = 0.796). Mean sensitivity of vascular pattern analysis was higher in tumors >5 cm (63.2%) and in genotype cluster 2 tumors (100%). In conclusion, vascular pattern analysis cannot be used in a stand-alone manner as a prognostic tool for the distinction between benign and malignant PCC, but could be used as an indicator of malignancy and might be a useful tool in combination with other morphological characteristics.     

Leaf optical properties in amphibious plant species are affected by multiple leaf traits

The amphibious plant species Sagittaria sagittifolia and Ranunculus lingua here serve as model systems to study differences in leaf optical properties of different leaf types that develop in aquatic and terrestrial environments. We aimed to determine leaf traits that explain most of the variability in the reflectance and transmittance spectra in the range from 280 to 880 nm. Comparisons of leaves of the same form revealed marked differences in their structures and particularly in the content of total methanol-soluble UV-absorbing compounds. Submerged leaves transmit radiation over the whole range measured, but emerged leaves transmit only at wavelengths from 500 to 650 nm, and above 690 nm. Redundancy analysis shows that biochemical leaf traits, namely the UV-absorbing compounds chlorophyll a and b, together with the specific leaf area (SLA), significantly affect the reflectance spectra, explaining 60% of the spectra variability. Pigment levels negatively affect reflectance, while the effect of SLA is positive. Physical traits like thickness of the palisade mesophyll, SLA, and thickness of the lower and upper epidermis, along with anthocyanin content, explain 62% of the transmittance spectra variability. This study provides new insight into the understanding of data collected for aquatic and semi-aquatic plants based on spectral analyses.