Upscaling e-mental health in Europe: a six-country qualitative analysis and policy recommendations from the eMEN project.

E-mental health (eMH) encompasses the use of digital technologies to deliver, support, or enhance mental health services. Despite the growing evidence for the effectiveness of eMH interventions, the process of implementation of eMH solutions in healthcare remains slow throughout Europe. To address this issue, the e-Mental Health Innovation and Transnational Implementation Platform North-West Europe (eMEN) project was initiated to increase the dissemination and quality of eMH services in Europe. In this project, status analyses regarding eMH in the six participating countries (i.e., Belgium, France, Germany, Ireland, The Netherlands, and the UK) were conducted and eight recommendations for eMH were developed. Expert teams from the six participating countries conducted status analyses regarding the uptake of eMH based on a narrative literature review and stakeholder interviews. Based on these status analyses, the eMEN consortium developed eight policy recommendations to further support the implementation of eMH in Europe. The status analyses showed that the participating countries are in different stages of implementing eMH into mental healthcare. Some barriers to implementing eMH were common among countries (e.g., a limited legal and regulatory framework), while others were country-specific (e.g., fragmented, federal policies). The policy recommendations included fostering awareness, creating strong political commitment, and setting reliable standards related to ethics and data security. The eMEN project has provided the initial recommendations to guide political and regulatory processes regarding eMH. Further research is needed to establish well-tailored implementation strategies and to assess the generalizability of the recommendations beyond the countries involved in the eMEN project.

Immuno-based detection of extracellular vesicles in urine as diagnostic marker for prostate cancer.

Extracellular vesicles (including the subclass exosomes) secreted by cells contain specific proteins and RNA that could be of interest in determining new markers. Isolation/characterization of PCa-derived exosomes from bodily fluids enables us to discover new markers for this disease. Unfortunately, isolation with current techniques (ultracentrifugation) is labor intensive and other techniques are still under development. The goal of our study was to develop a highly sensitive time-resolved fluorescence immunoassay (TR-FIA) for capture/detection of PCa-derived exosomes. In our assay, biotinylated capture antibodies against human CD9 or CD63 were incubated on streptavidin-coated wells. After application of exosomes, Europium-labeled detection antibodies (CD9 or CD63) were added. Cell medium from 37 cell lines was taken to validate this TR-FIA. Urine was collected (after digital rectal exam) from patients with PCa (n = 67), men without PCa (n = 76). As a control, urine was collected from men after radical prostatectomy (n = 13), women (n = 16) and patients with prostate cancer without digital rectal exam (n = 16). Signal intensities were corrected for urinary PSA and creatinine. This TR-FIA can measure purified exosomes with high sensitivity and minimal background signals. Exosomes can be measured in medium from 37 cell lines and in urine. DRE resulted in a pronounced increase in CD63 signals. After DRE and correction for urinary PSA, CD9 and CD63 were significantly higher in men with PCa. This TR-FIA enabled us to measure exosomes with high sensitivity directly from urine and cell medium. This TR-FIA forms the basis for testing different antibodies directed against exosome membrane markers to generate disease-specific detection assays.

A search for new metabolites of N,N',N''-triethylenethiophosphoramide.

An attempt was made to unravel the metabolic profile of the alkylating agent N,N',N''-triethylenethiophosphoramide (thioTEPA). thioTEPA and its metabolite N,N',N-triethylenephosphoramide (TEPA) were quantified in urine of treated patients by gas chromatography with selective nitrogen/phosphorous detection. Total alkylating activity was assessed by p-nitrobenzylpyridine reactivity. The total alkylating activity exceeded the amount of thioTEPA and TEPA, indicating the presence of other alkylating metabolites. Solid-phase extraction and liquid-liquid extractions followed by gas chromatography-mass spectrometry analysis revealed the conversion of an aziridinyl function of TEPA into a beta-chloroethyl moiety. This metabolite, N,N'-diethylene-N''-2-chloroethylphosphoramide, was quantified by gas chromatography with selective nitrogen/phosphorous detection and accounted for only 0.69% of the administered dose. Large volumes of urine were concentrated with solid-phase extraction and fractionated with high-performance liquid chromatography. Alkylating activity was determined for each 2-ml fraction and showed the presence of an alkylating compound eluting between 8 and 12 ml. The fractions with alkylating activity were collected, evaporated under a stream of nitrogen at room temperature to dryness, reconstituted in methanol, and subjected to fast atom bombardment-mass spectrometry and fast atom bombardment-tandem mass spectrometry. A new metabolite was found with a molecular mass of 352 Da, the same as that of thioTEPA-mercapturate. thioTEPA-mercapturate is likely the result of glutathione conjugation, after which the glutathione adduct loses two amino acid residues in separate stages. The fragmentation pattern and chromatographic properties of this new metabolite were identical to those of the reference, thioTEPA-mercapturate, which was obtained by incubation of thioTEPA with N-acetylcysteine at pH 11 and 95 degrees C for 30 min. Quantification of thioTEPA-mercapturate was carried out by liquid chromatography-mass spectrometry. The thioTEPA-mercapturate levels in urine accounted for 12.3% of the administered dose and exceeded the amount of TEPA, which was previously assumed to be the main metabolite of thioTEPA. The total excreted amount of thioTEPA and its metabolites accounts for 54-100% of the total alkylating activity, indicating the presence of still other alkylating metabolites.

Gamma-irradiation of liposomes composed of saturated phospholipids: effect of bilayer composition, size, concentration and absorbed dose on chemical degradation and physical destabilization of liposomes.

Liposomes composed of dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG), or mixtures of these two phospholipids were exposed to gamma-irradiation in an air environment. Disappearance of the mother compounds was monitored by HPLC analysis. Plotting of the logarithmic values of residual DPPC or DPPG concentration versus irradiation dose resulted in straight lines. The slopes of these lines (overall degradation constants) depended on the type of phospholipids, concentration of the liposomes and the size of the liposomes. Under the chosen conditions, addition of DPPG in DPPC-liposomes did not affect the degradation rate constant of DPPC and vice versa. The presence of phosphate buffer (pH 7.4), pH or presence of sodium chloride did not affect the irradiation damage either. Minor changes were found upon analysis of total fatty acids by GLC and upon measurement of water soluble phosphate compounds. These changes were less pronounced than the changes monitored by HPLC of phospholipids, because the HPLC analysis monitored the overall degradation of the liposomal phospholipids. Thin-layer chromatography/fast atom bombardment mass spectrometry (TLC/FAB-MS) analysis of irradiated and non-irradiated DPPC or DPPG provided information on the structure of several degradation products. Degradation routes which include these degradation products are proposed. Gamma-irradiation neither affected the size of the liposomes nor the bilayer rigidity as determined by dynamic light scattering and fluorescence anisotropy of the probe 1,6-diphenyl-1,3,5-hexatriene (DPH), respectively. However, upon gamma-irradiation, changes in the melting characteristics of the liposomes were found by differential scanning calorimetry (DSC) measurements. The pre-transition melting enthalpy of the liposomal bilayer decreased or disappeared and the main-transition broadened. The changes found in DSC scans correlated qualitatively well with the changes recorded after HPLC analysis of phospholipids.

Detection of intact megaDalton protein assemblies of vanillyl-alcohol oxidase by mass spectrometry.

Well-resolved ion signals of intact large protein assemblies, with molecular masses extending above one million Dalton, have been detected and mass analyzed using electrospray ionization mass spectrometry, with an uncertainty in mass of <0.2%. The mass spectral data seem to reflect known solution-phase behavior of the studied protein assembly and have therefore been directly used to probe the protein assembly topology and stability as a function of ionic strength and pH.

Measuring plasma exudation in nasal lavage fluid and in induced sputum as a tool for studying respiratory tract inflammation.

We performed nasal lavage (NAL) combined with induced sputum to determine exudative inflammation in the upper and lower airways in patients with chronic sinusitis and in controls. To monitor plasma exudation into the respiratory lumen and loss of size-selectivity of the mucosa, we determined the sample-to-serum ratio of albumin and alpha-2-macroglobulin, Qa1b and Qa2m, and the dilution independent Relative Coefficient of Excretion, RCE=Qa2m/Qa1b. To detect low protein levels in NAL and induced sputum we adapted an ELISA system for alpha-2-macroglobulin described by Out et al. [Clin. Chim. Acta, 165 (1987) 277-288], and modified this into a sensitive ELISA for albumin. Dithiothreitol, added to increase sputum solubility, did not interfere with the analysis, nor did N-ethylmaleimide, added to block dithiothreitol. In this study plasma exudation in induced sputum is significantly increased in patients with chronic sinusitis, compared to controls. Plasma exudation in NAL is also increased in patients, although not significant. The RCE in NAL and sputum is well-correlated in one of the three study visits. There is much variation in sample protein-levels partly due to differences in dilution and the heterogeneity of the studied population. Determination of plasma exudation together with RCE in NAL and induced sputum is a good, non-invasive way to quantify inflammation of airway mucosa.

Metastable ion formation and disparate charge separation in the gas-phase dissection of protein assemblies studied by orthogonal time-of-flight mass spectrometry.

The dissection of specific and nonspecific protein complexes in the gas phase is studied by collisionally activated decomposition. In particular, the gas phase dissection of multiple protonated homodimeric Human Galectin I, E. Coli Glyoxalase I, horse heart cytochrome c, and Hen egg Lysozyme have been investigated. Both the Human Galectin I and E. Coli Glyoxalase I enzymes are biologically active as a dimer, exhibiting molecular weights of approximately 30 kDa. Cytochrome c and Lysozyme are monomers, but may aggregate to some extent at high protein concentrations. The gas phase dissociation of these multiple protonated dimer assemblies does lead to the formation of monomers. The charge distribution over the two concomitant monomers following the dissociation of these multiple protonated dimers is found to be highly dissimilar. There is no evident correlation between the solution phase stability of the dimeric proteins and their gas-phase dissociation pattern. Additionally, in the collisionally activated decomposition spectra diffuse ion signals are observed, which are attributed to monomer ions formed via slow decay of the collisionally activated dimer ions inside the reflectron time-of-flight. Although, the formation of these diffuse metastable ions may complicate the interpretation of collisionally activated decomposition mass spectra, especially when studying noncovalent protein complexes, a simple mathematical equation may be used to reveal their origin and pathway of formation.

Plasma pharmacokinetics, tissue disposition, excretion and metabolism of vinleucinol in mice as determined by high-performance liquid chromatography.

We investigated the pharmacokinetics of the experimental semisynthetic vinca alkaloid vinleucinol (VileE; O4-deacetyl-3-de(methoxycarbonyl)-3-[[[1-ethoxycarbonyl-2- methylbutyl]amino]carbonyl]-vincaleukoblastine). The study was performed in male FVB mice receiving 10.5 mg/kg VileE i.v. or p.o. Plasma, urine, faeces and tissue samples were analysed by a selective method based on ion-exchange normal-phase high-performance liquid chromatography (HPLC) with fluorescence detection and liquid-liquid extraction for sample clean-up. Apart from the parent drug, two other metabolic compounds were detected. One of these metabolites is vinleucinol acid (VileA; O4-deacetyl-3-de(methoxycarbonyl)-3-[[[1-carboxyl-2- methylbutyl]amino]carbonyl]-vincaleukoblastine), which possesses no cytotoxic activity. The structure proposed for the second metabolite (VileX) was based on tandem mass spectrometry (MS-MS) and infrared (IR) spectroscopy data. Metabolization of VileE to VileX must occur in the amino acid moiety of the molecule, with a (beta- or gamma-) lactone ring being formed after oxidation of the (beta- or gamma) carbon of the amino acid. VileX is a major metabolite, which is excreted in faeces and urine after i.v. administration and accounting for up to 23% of the administered dose. The activity of VileX against cultured L1210 cells is four times that of the parent drug VileE and comparable with that of vinblastine (VBL). At 48 h after administration of VileE, the concentration of VileX exceeds that of the parent drug in many tissues. These findings indicate that the metabolite VileX may be at least largely responsible for the activity observed against xenografts in mice after administration of the parent drug, VileE.

A simple, whole blood method for assessment of platelet function: application to dietary intervention.

The propensity to thrombosis in an individual or population represents a significant risk factor in coronary heart disease, that ultimately may result in acute myocardial infarction or unstable angina. A variety of currently available tests assess the relative potential for platelets to be activated and then aggregate, including agonist-dependent platelet aggregation or flow cytometric analysis of platelet activation. However, all of these methods have certain limitations, ranging from being poorly quantifiable with limited sensitivity, to the necessity for specialized equipment. In the present study, we describe the development of a simple whole blood, radiolabel assay that measures the surface expression of the alpha-granule protein, P-selectin, by activated platelets. This assay is performed in the presence of GP IIb-IIIa blockade to allow quantitation without interference by platelet aggregate formation, and thus directly measures agonist dose-response without complications arising from secondary activation mediated by GP IIb-IIIa. The sensitivity of this assay method to dietary manipulation was investigated by administration of fish oil capsules at a dose known to decrease platelet aggregation.

Separation and identification of enzymatic sucrose hydrolysis products by high-performance anion-exchange chromatography with pulsed amperometric detection.

An accurate carbohydrate analysis method, namely high-performance anion-exchange chromatography with pulsed amperometric detection was successfully applied to the study of sucrose hydrolysis under enzymatic (baker's yeast invertase) conditions. The hydrolysis was monitored by determining sucrose degradation and the corresponding formation of D-glucose, D-fructose and five intermediate fructans using a CarboPac PA-100 (Dionex) analytical anion-exchange column. Highly reproducible results were obtained. The unknown fructans were collected from a semi-preparative CarboPac PA-100 (Dionex) column, neutralized and then desalted on a column containing mixed bed resin AG 501-X8 (D) before identification of the chemical structure. This procedure permitted us to obtain about 20 microg of pure product which is not enough for NMR analysis. Detailed GC-MS analytical data of the methylated compounds indicated that these oligosaccharides were beta-D-Fru-(2 --> 1)-beta-D-Fru-(2 --> 1)-alpha-D-glucopyranoside (1-kestose), beta-D-Fru-(2 --> 6)-alpha-D-glucopyranoside (6-beta fructofuranosylglucose), beta-D-Fru-(2 --> 1)-beta-D-fructofuranoside (inulobiose), beta-D-Fru-(2 --> 6)-beta-D-Fru-(2 --> 1)-alpha-D-glucopyranoside (6-kestose) and beta-D-Fru-(2 --> 6)-alpha-D-Glc-(1 --> 2)-beta-D-fructofuranoside (neokestose) coeluating with a disaccharide.

1-Phenyl-5-vinyl-2-imidazolidinethione, a proposed causative agent of Spanish toxic oil syndrome: synthesis, and identification in one of a group of case-associated oil samples.

A method is described for the synthesis and characterization of N-(2-hydroxy-3-butenyl)-N'-phenylthiourea, and its cyclization product, 1-phenyl-5-vinyl-2-imidazolidinethione (PVIZT). Fourteen coded oil samples associated with toxic oil syndrome cases in Spain were examined by gas chromatography-electron impact mass spectrometry for the presence of PVIZT. Although these samples were obtained from households where cases of toxic oil syndrome had been recorded, they differed extremely with regard to their anilide and sulphur contents. In one sample PVIZT was detected at an estimated concentration of 1 mg/kg.

Rolling Loop Scan: An Approach Featuring Ring-Closing Metathesis for Generating Libraries of Peptides with Molecular Shapes Mimicking Bioactive Conformations or Local Folding of Peptides and Proteins.

Libraries of loop-containing peptides (such as the one shown schematically) can be prepared from bis-N-alkylated peptides by ring-closing metathesis. In a general solid-phase procedure the peptides are accessible by site-specific N-alkylation. Since the amino acid side chains are not involved in cyclization, they remain available for interaction with, for example, a receptor.

Iron-regulated metabolites produced by Pseudomonas fluorescens WCS374r are not required for eliciting induced systemic resistance against Pseudomonas syringae pv. tomato in Arabidopsis.

The plant growth-promoting rhizobacterium Pseudomonas fluorescens WCS374r produces several iron-regulated metabolites, including the fluorescent siderophore pseudobactin (Psb374), salicylic acid (SA), and pseudomonine (Psm), a siderophore that contains a SA moiety. After purification of Psb374 from culture supernatant of WCS374r, its structure was determined following isoelectrofocusing and tandem mass spectrometry, and found to be identical to the fluorescent siderophore produced by P. fluorescens ATCC 13525. To study the role of SA and Psm production in colonization of Arabidopsis thaliana roots and in induced systemic resistance (ISR) against Pseudomonas syringae pv. tomato (Pst) by strain WCS374r, mutants disrupted in the production of these metabolites were obtained by homologous recombination. These mutants were further subjected to transposon Tn5 mutagenesis to generate mutants also deficient in Psb374 production. The mutants behaved similar to the wild type in both their Arabidopsis rhizosphere-colonizing capacity and their ability to elicit ISR against Pst. We conclude that Psb374, SA, and Psm production by P. fluorescens WCS374r are not required for eliciting ISR in Arabidopsis.

A low vitamin A status increases the susceptibility to cigarette smoke-induced lung emphysema in C57BL/6J mice.

Chronic obstructive pulmonary disease (COPD) is characterized by chronic airway inflammation. Cigarette smoke has been considered a major player in the pathogenesis of COPD. The inflamed airways of COPD patients contain several inflammatory cells. Vitamin A metabolites have been implicated in the repair of lung damage. Exposure to cigarette smoke has been shown to depress levels of retinol in lungs of rats. The purpose of this study was to investigate if a low, but not deficient, vitamin A status potentiated susceptibility to the development of cigarette smoke-induced lung emphysema in mice. Mice were bred that were the offspring's of 3 generations of mice that were fed a purified diet containing low levels of vitamin A and exposed to cigarette smoke for 3 months, every weekday. Then, levels of 9-cis, 13-cis, and all-trans retinoic acid, retinol and retinyl palmitate were measured in plasma, liver and right lung lobe. The left lung lobe was used to assess mean linear intercept (Lm), as a measure of smoke-induced lung damage. Average feed intakes were not different between treatment groups. We show that both retinol and retinyl palmitate levels were dramatically decreased in the storage organs of mice on the low vitamin A diet (retinol 2-fold in both lung and liver, and retinyl palmitate 5- fold in lung) which shows that the depletion was successful. However, this treatment did not result in the development of lung emphysema. However, smoke exposure led to a significant increase in Lm in mice with a low vitamin A status compared to the room air-breathing controls. Lung levels of acid retinoids were similar in all mice, irrespective of diet or smoke exposure. Concluding, a low vitamin A status increases the susceptibility to the development of cigarette smoke-induced lung emphysema, possibly because of decreased anti-oxidant capacity in the lungs due to locally reduced retinol and retinyl palmitate levels. These observations indicate that human populations with a low vitamin A status and a high prevalence of smoking may be at increased risk of developing lung emphysema.

Progression of cerebral white matter lesions is not associated with development of depressive symptoms in elderly subjects at risk of cardiovascular disease: The PROSPER Study.

BACKGROUND: Cerebral white matter hyperintensities on magnetic resonance imaging (MRI) scans have been associated with vascular disease and late-life depression, both in the general population and in psychiatric patients. Therefore, a cerebrovascular etiology for late-onset depression has been hypothesized. However, longitudinal studies on the causal role of white matter hyperintensities in the development of depressive symptoms in elderly adults are lacking. OBJECTIVE: To investigate the relation between white matter hyperintensities and depressive symptoms in elderly subjects at risk of cardiovascular disease. METHODS: In the Dutch sample of the PROSPER (PROspective Study of Pravastatine in the Elderly at Risk of cardiovascular disease) cohort, 527 non-demented elderly, all aged 70 years or older, received a cranial MRI scan and the 15-item Geriatric Depression Scale, at baseline and 33 months (SD 1.6) later. RESULTS: Presence of white matter hyperintensities at baseline was not related to baseline depressive symptoms nor to the development of depressive symptoms during follow-up. Moreover, no association was found between progression of white matter lesion volume and progression of depressive symptoms. CONCLUSION: This longitudinal study does not confirm the involvement of cerebrovascular disease expressed as MRI white matter hyperintensities in the development of depressive symptoms in elderly subjects.

Synthesis of cyclic peptides by ring-closing metathesis.

The synthesis of a series of "amide to amide" cyclized peptides by ring-closing metathesis (RCM) as well as a convenient synthesis for the linear precursors is described. In addition, the influence of the length of the alkene substituents and the influence of the peptide sequence is investigated, leading to a set of general rules to obtain "amide to amide" cyclized peptides by RCM.

Isolation and identification of 25-hydroxydihydrotachysterol2 1 alpha,25-dihydroxydihydrotachysterol2 and 1 beta,25-dihydroxydihydrotachysterol2.

Three metabolites of orally administered dihydrotachysterol2 have been isolated in impure form from serum of rats. These metabolites have been identified as 25-hydroxydihydrotachysterol2 and two epimers of formula 1-ambo,25-dihydroxydihydrotachysterol2 by means of gas chromatography-mass spectrometry and ultraviolet absorption spectrometry. For the first time this provides evidence for 9,10-seco steroid hydroxylation at pseudo C3. The stereochemistry of the 1-hydroxyl group of the two epimers could be established tentatively by quantitative comparison of the mass spectra of their respective trimethylsilyl derivatives. Since purity requirements were not achieved, biological activities could not be determined.

High-performance thin-layer chromatography/fast atom bombardment (tandem) mass spectrometry of Pseudomonas rhamnolipids.

Several rhamnolipid preparations from Pseudomonas strains were studied by thin-layer chromatography/fast atom bombardment (TLC/FAB) mass spectrometry and TLC/FAB tandem mass spectrometry (MS/MS). The preparations were separated with normal-phase (Silica 60) and reversed-phase (RP-8) chromatography. Silica 60 plates appeared to be very useful in the separation of rhamnolipids according to the number of monosaccharide residues present. Spectra which show characteristic fragment ions could be obtained from components of mixtures with a total sample size of less than 200 ng. Chromatography on RP-8 plates gave a good separation of the rhamnolipids based on the length of the fatty acid alkyl chain. MS/MS of the sodium cationized molecules gave information about the sequence of the building blocks. Particularly, heterogeneity in beta-hydroxy fatty acid composition was determined for the principal as well as minor components present in natural rhamnolipid mixtures.

5-Octadecenoic acid: evidence for a novel type of fatty acid modification in schistosomes.

The lipid metabolism of schistosomes is characterized by several intriguing adaptations to a parasitic way of living. The surface of the parasite consists of two closely apposed phospholipid bilayers, a structure unique to blood flukes. Schistosomes do not synthesize fatty acids de novo, but are able to modify fatty acids, which they obtain from the host, by chain elongation. Here we present evidence that schistosomes are capable of another type of fatty acid modification, resulting in the formation of 5-octadecenoic acid [C18:1(5)]. This highly unusual fatty acid, which is absent in the blood of the host, was shown to be almost exclusively located in the outer membrane complex of the schistosome. Within these membranes, it was almost exclusively present in one molecular phospholipid species, 1-palmitoyl-2,5-octadecenoyl phosphatidylcholine [C16:0-18:1(5)PtdCho]. Apart from dipalmitoyl phosphatidylcholine, this was the most abundant phosphatidylcholine species in the outer membrane complex. The specific synthesis by the schistosome of C18:1(5) and the highly specific localization of this fatty acid to the tegumental membranes suggest an important tegument-mediated role for this lipid.