Analysis of Cerebrospinal Fluid Extracellular Vesicles by Proximity Extension Assay: A Comparative Study of Four Isolation Kits

There is a lack of reliable biomarkers for disorders of the central nervous system (CNS), and diagnostics still heavily rely on symptoms that are both subjective and difficult to quantify. The cerebrospinal fluid (CSF) is a promising source of biomarkers due to its close connection to the CNS. Extracellular vesicles are actively secreted by cells, and proteomic analysis of CSF extracellular vesicles (EVs) and their molecular composition likely reflects changes in the CNS to a higher extent compared with total CSF, especially in the case of neuroinflammation, which could increase blood–brain barrier permeability and cause an influx of plasma proteins into the CSF. We used proximity extension assay for proteomic analysis due to its high sensitivity. We believe that this methodology could be useful for de novo biomarker discovery for several CNS diseases. We compared four commercially available kits for EV isolation: MagCapture and ExoIntact (based on magnetic beads), EVSecond L70 (size-exclusion chromatography), and exoEasy (membrane affinity). The isolated EVs were characterized by nanoparticle tracking analysis, ELISA (CD63, CD81 and albumin), and proximity extension assay (PEA) using two different panels, each consisting of 92 markers. The exoEasy samples did not pass the built-in quality controls and were excluded from downstream analysis. The number of detectable proteins in the ExoIntact samples was considerably higher (~150% for the cardiovascular III panel and ~320% for the cell regulation panel) compared with other groups. ExoIntact also showed the highest intersample correlation with an average Pearson’s correlation coefficient of 0.991 compared with 0.985 and 0.927 for MagCapture and EVSecond, respectively. The median coefficient of variation was 5%, 8%, and 22% for ExoIntact, MagCapture, and EVSecond, respectively. Comparing total CSF and ExoIntact samples revealed 70 differentially expressed proteins in the cardiovascular III panel and 17 in the cell regulation panel. To our knowledge, this is the first time that CSF EVs were analyzed by PEA. In conclusion, analysis of CSF EVs by PEA is feasible, and different isolation kits give distinct results, with ExoIntact showing the highest number of identified proteins with the lowest variability.     

Histone Methylation Marks on Circulating Nucleosomes as Novel Blood-Based Biomarker in Colorectal Cancer

Circulating nucleic acids (CNAs) are under investigation as a liquid biopsy in cancer as potential non-invasive biomarkers, as stable structure in circulation nucleosomes could be valuable sources for detection of cancer-specific alterations in histone modifications. Our interest is in histone methylation marks with a focus on colorectal cancer, one of the leading cancers respective the incidence and mortality. Our previous work included the analysis of trimethylations of lysine 9 on histone 3 (H3K9me3) and of lysine 20 on histone 4 (H4K20me3) by chromatin immuno- precipitation-related PCR in circulating nucleosomes. Here we asked whether global immunologic measurement of histone marks in circulation could be a suitable approach to show their potential as biomarkers. In addition to H3K9me3 and H4K20me3 we also measured H3K27me3 in plasma samples from CRC patients (n = 63) and cancer free individuals (n = 40) by ELISA-based methylation assays. Our results show that of three marks, the amounts of H3K27me3 (p = 0.04) and H4K20me3 (p < 0.001) were significantly lower in CRC patients than in healthy controls. For H3K9me3 similar amounts were measured in both groups. Areas under the curve (AUC) in receiver operating characteristic (ROC) curves indicating the power of CRC detection were 0.620 for H3K27me3, 0.715 for H4K20me3 and 0.769 for the combination of both markers. In conclusion, findings of this preliminary study reveal the potential of blood-based detection of CRC by quantification of histone methylation marks and the additive effect of the marker combination.     

Tracking influent inorganic suspended solids through wastewater treatment plants.

From an experimental and theoretical investigation of the continuity of influent inorganic suspended solids (ISS) along the links connecting the primary settling tank (PST), fully aerobic or N removal activated sludge (AS) and anaerobic and aerobic sludge digestion unit operations, it was found that the influent wastewater (fixed) ISS concentration is conserved through primary sludge anaerobic digestion, activated sludge and aerobic digestion unit operations. However, the measured ISS flux at different stages through a series of wastewater treatment plant (WWTP) unit operations is not equal to the influent ISS flux, because the ordinary heterotrophic organisms (OHO) biomass contributes to the ISS flux by differing amounts depending on the active fraction of the VSS solids at that stage.     

Prominent consequences of role stress: A meta-analytic review.

Role stress has received a lot of research attention in psychological, sociological, and organizational studies over the last several decades. Based on a literature review of about 300 journal articles, this article examines prominent consequences of role stress. Specific focus is on researching differences in relationships between facets of role stress (i.e., role ambiguity, role conflict, and role overload) and frequently cited consequences using techniques of meta-analysis. Findings indicate that each role stress facet has a different relationship with the eight consequences studied. Role stress research can benefit from looking at each facet individually in addition to role stress generally. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Co-digestion of manure and biowaste according to the EC animal by-products regulation and Finnish national regulations.

The objective of this study was to compare methane production and characteristics of digested material in anaerobic digestion concepts according to the Animal By-Products Regulation (ABP-Regulation) of the EC (hygienisation of biowaste for 1 hour at 70 degrees C, particle size < 12 mm) and Finnish national regulations (treatment temperature 55 degrees C, feeding interval 24 h, hydraulic retention time (HRT) 20 d, particle size < 40 mm) and with small variations in treatment methods for treating manure and biowaste. Moreover, the survival of three different salmonella bacteria in these processes was studied. Hygienisation of biowaste prior to digestion at 35 degrees C enhanced methane production by 14-18% compared to similar treatment without hygienisation. The differences in treatment temperature, HRT and hygienisation of biowaste prior to digestion did not significantly affect the characteristics of digested material. The concepts according to the ABP-Regulation and Finnish national regulations were effective in destroying salmonella bacteria to an undetectable level.     

Pilot Plant Study of Simultaneous Sewage Sludge Digestion and Metal Leaching

This research is related to a preindustrial pilot scale study of the performance of the simultaneous sewage sludge digestion and metal leaching (SSDML) process for decontamination and stabilization of sewage sludge. Ten batch tests were carried out in two 4?m3 bioreactors under various conditions of operation. Results indicated that the addition of 1.0 to 1.5?g?S0/L, which is the equivalent of approximately 30 to 50?kg?S0 per tons dry sludge, is sufficient to obtain conditions of acidity (pH<2.5) and oxido-reduction potential greater than 500 mV necessary for an effective solubilization of toxic metals. The final average of metal solubilization in the output sludge during the SSDML process varied in the following ranges: 25–78% Cd, 9–32% Cr, 48–100% Cu, 77–99% Mn, 15–53% Ni, 12–47% Pb, and 66–100% Zn. The N, P, and K contents were also preserved in the decontaminated sludge. Moreover, the use of low concentrations of elemental sulfur makes it possible to obtain decontaminated sludge with a low total sulfur content (1.4–1.5% S) compatible with agricultural use. The suspended solids removal calculated for the SSDML process was slightly lower, (2.5±0.4)% volatile suspended solids per day, than those reached using standard aerobic digestion. Finally, the SSDML process was found to be effective in removing bad odors and in the destruction (99–100%) of indicator bacteria.     

Sparse field level set method for non-convex Hamiltonians in 3D plasma etching profile simulations

Level set method [S. Osher, J. Sethian, J. Comput. Phys. 79 (1988) 12] is a highly robust and accurate computational technique for tracking moving interfaces in various application domains. It originates from the idea to view the moving front as a particular level set of a higher dimensional function, so the topological merging and breaking, sharp gradients and cusps can form naturally, and the effects of curvature can be easily incorporated. The resulting equations, describing interface surface evolution, are of Hamilton-Jacobi type and they are solved using techniques developed for hyperbolic equations. In this paper we describe an extension of the sparse field method for solving level set equations in the case of non-convex Hamiltonians, which are common in the simulations of the profile surface evolution during plasma etching and deposition processes. Sparse field method itself, developed by Whitaker [R. Whitaker, Internat. J. Comput. Vision 29 (3) (1998) 203] and broadly used in image processing community, is an alternative to the usual combination of narrow band and fast marching procedures for the computationally effective solving of level set equations. The developed procedure is applied to the simulations of 3D feature profile surface evolution during plasma etching process, that include the effects of ion enhanced chemical etching and physical sputtering, which are the primary causes of the Hamiltonian non-convexity.     

On the effect of spatial variances in historical rainfall time series to CSO performance evaluation.

Historical, high-resolution rain series are the backbone of modern combined sewer overflow (CSO) structure design. These rain series are the input to the computational estimation of the performance of the measures with respect to CSO pollution abatement. However, those historical precipitation measurements are available at only a few locations. Frequently rain series have to be used from gauging stations at a significant distance. In order to judge and to compensate for this influence an estimate between rain characteristics and combined sewer outflow (CSO) performance indicators would be useful. In this paper such correlations have been sought for a collection of 37 rain series covering large areas of Europe. It was found that the mean annual rain volume can explain most of the variances for the performance indicators Number of overflows and CSO volume. For explaining the spatial differences in the efficiency of the CSO structure another rain characteristic, i.e. the maximum event with a return period of one year, is to be used.     

A new polymer electrolyte poly(acrylonitrile)-dimethylsulphoxide-salt for electrochemical capacitors

The PAN-DMSO-Et₄NBF₄ and PAN-DMSO-Et₄NTf (Tf is triflate ion) electrolytes were prepared as white, turbid foils with a thickness in the range of 0.1-0.5 mm, using the casting technique. Room temperature conductance of the electrolytes, detected from ac impedance experiments, was at the level of 8 and 14 mS cm⁻¹ for Et₄NBF₄ and Et₄NTf salts, respectively. The electrochemical stability window of approximately 2.6-2.8 V was estimated using a glassy carbon electrode. Temperature dependence of the conductivity is of the Arrhenius-type for both electrolytes, with an activation energy of approximately 34 kJ mol⁻¹. The double-layer capacitors built with these electrolytes, serving both as separators and activated carbon powder (ACP) binders, showed a specific capacity of 50 F g⁻¹ of carbon material. Capacitors were assembled by sandwiching the PAN-DMSO-salt electrolyte between two PAN-salt-DMSO-ACP-AB electrodes and pressing across the system; the resulting devices had a coin-like shape with 8 mm diameter and thickness between 2.0 and 2.5 mm.