A comparison of self and proxy quality of life ratings for people with dementia and their carers: a European prospective cohort study.

Objectives: To identify correlates of self-rated and proxy-rated quality of life (QoL) in people with dementia on (i) a dementia-specific and (ii) a capability-wellbeing QoL measure at baseline and 12-month follow-up, and to consider such factors in the context of QoL intervention development.Method: Prospective clinical and demographic data were collected from 451 community-dwelling dyads (mild-moderate dementia) across eight European countries. QoL was measured using the QOL-AD and the ICECAP-O. Multivariate modelling identified correlates of self- and proxy-rated QoL at baseline and at 12-month follow-up.Results: Carer's proxy-ratings of QoL were significantly lower than self-ratings at all time-points for both measures. Proxy-ratings declined over time, but self-ratings remained stable. Baseline predictors of greater self-rated QoL were education, and greater functional ability and relationship quality. Greater proxy-rated QoL was associated with education and greater functional ability, relationship quality, carer social support and carer QoL, lower carer anxiety/depression and less severe neuropsychiatric symptoms in people with dementia. At follow-up, greater self-rated QoL was predicted by greater functional ability, relationship quality, carer social support and having a spousal carer. Greater proxy-rated QoL at follow-up was associated with the same factors as at baseline; however, the dyad living together was an additional predictive factor.Conclusion: Both proxy-ratings and self-ratings of QoL should be interpreted with caution and in the context of each individual caregiving relationship. Different functional, psychosocial, relational and contextual factors influence self- and proxy-ratings, and both sets of factors should be considered in the context of QoL intervention development for the dyad.

Access to community care for people with dementia and their informal carers : Case vignettes for a European comparison of structures and common pathways to formal care.

BACKGROUND: People with dementia and their informal carers often do not receive appropriate professional support or it is not received at the right time. OBJECTIVES: Description and comparison of common pathways to formal community dementia care in eight European countries as a part of the transnational Actifcare project. MATERIALS AND METHODS: The German team was responsible for creating an individual case scenario as a starting point. The research teams in Ireland, Italy, the Netherlands, Norway, Portugal, Sweden, and the United Kingdom were then asked to describe a common pathway to formal dementia care by writing their own vignette using the provided individual case scenario. RESULTS: A transnational qualitative content analysis was used to identify the following categories as being the most important: involved professionals, dementia-specific and team-based approaches, proactive roles, and financial aspects. General practitioners (GPs) are described as being the most important profession supporting the access to formal care in all the involved countries. In some countries other professionals take over responsibility for the access procedure. Dementia-specific approaches are rarely part of standard care; team-based approaches have differing significances in each of the countries. Informal carers are mainly proactive in seeking formal care. The Nordic countries demonstrate how financial support enhances access to the professional system. CONCLUSION: Enhanced cooperation between GPs and other professions might optimize access to formal dementia care. Team-based approaches focusing on dementia care should be developed further. Informal carers should be supported and relieved in their role. Financial barriers remain which should be further investigated and reduced.

[The costs and benefits of an integrated approach to dementia]. / De kosten en opbrengsten van een geïntegreerde aanpak bij dementie.

BACKGROUND: An integrated approach to dementia is generally recommended because no one discipline is adequately equipped it deal with the complex psychic, physical and social problems that are inherent in dementia. A multidisciplinary approach, however, leads inevitably to higher costs. It is not known what the cost/benefit ratio will be. AIM: To describe our research into the costs and benefits of an integrated approach to dementia involving the use of a diagnostic research centre for psycho-geriatrics and thereafter to compare our findings with the results of other studies of the costs and benefits of an integrated approach. METHOD: We performed a prospective and randomised efficiency study and we compared our findings with the results of other studies of the costs and benefits of an integrated approach. We reviewed recent literature. RESULTS: The DRC-PG was more effective than normal care as far as the patients' quality of life was concerned, but was not more expensive. It can therefore be regarded as a cost-effective facility for ambulatory patients with dementia. Three other studies provided additional empirical evidence of the success of a similar integrated approach in various sectors involved in the care of patients with dementia. CONCLUSION: An integrated approach with regard to the diagnosis, treatment and management of dementia produces favourable results. More research is needed into the efficacy and cost-effectiveness of integrated care programmes. This should result in improvements in the care and treatment of patients with dementia.

Cost-effectiveness of post-diagnosis treatment in dementia coordinated by Multidisciplinary Memory Clinics in comparison to treatment coordinated by general practitioners: an example of a pragmatic trial.

BACKGROUND: With the rising number of dementia patients with associated costs and the recognition that there is room for improvement in the provision of dementia care, the question arises on how to efficiently provide high quality dementia care. OBJECTIVE: To describe the design of a study to determine multidisciplinary memory clinics' (MMC) effectiveness and cost-effectiveness in post-diagnosis treatment and care-coordination of dementia patients and their caregivers compared to the post-diagnosis treatment and care-coordination by general practitioners (GP). Next, this article provides the theoretical background of pragmatic trials, often needed in complex interventions, with the AD- Euro study as an example of such a pragmatic approach in a clinical trial. METHOD: The study is a pragmatic multicentre, randomised clinical trial with an economic evaluation alongside, which aims to recruit 220 independently living patients with a new dementia diagnosis and their informal caregivers. After baseline measurements, patient and caregiver are allocated to the treatment arm MMC or GP and are visited for follow up measurements at 6 and 12 months. Primary outcome measures are Health Related Quality of Life of the patient as rated by the caregiver using the Quality of Life in Alzheimer's Disease instrument (Qol-AD) and self-perceived caregiving burden of the informal caregiver measured using the Sense of Competence Questionnaire (SCQ). To establish cost-effectiveness a cost-utility analysis using utilities generated by the EuroQol instrument (EQ-5D) will be conducted from a societal perspective. Analyses will be done in an intention-to-treat fashion. RESULTS: The inclusion period started in January 2008 and will commence until at least December 2008. After finalising follow up the results of the study are expected to be available halfway through 2010. DISCUSSION: The study will provide an answer to whether follow-up of dementia patients can best be done in specialised outpatient memory clinics or in primary care settings with regard to quality and costs. It will enable decision making on how to provide good and efficient health care services in dementia. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT00554047.

Localization and rearrangement modulation of the N-terminal arm of the membrane-bound major coat protein of bacteriophage M13.

During infection the major coat protein of the filamentous bacteriophage M13 is in the cytoplasmic membrane of the host Escherichia coli. This study focuses on the configurational properties of the N-terminal part of the coat protein in the membrane-bound state. For this purpose X-Cys substitutions are generated at coat protein positions 3, 7, 9, 10, 11, 12, 13, 14, 15, 17, 19, 21, 22, 23 and 24, covering the N-terminal protein part. All coat protein mutants used are successfully produced in mg quantities by overexpression in E. coli. Mutant coat proteins are labeled and reconstituted into mixed bilayers of phospholipids. Information about the polarity of the local environment around the labeled sites is deduced from the wavelength of maximum emission using AEDANS attached to the SH groups of the cysteines as a fluorescent probe. Additional information is obtained by determining the accessibility of the fluorescence quenchers acrylamide and 5-doxyl stearic acid. By employing uniform coat protein surroundings provided by TFE and SDS, local effects of the backbone of the coat proteins or polarity of the residues could be excluded. Our data suggest that at a lipid to protein ratio around 100, the N-terminal arm of the protein gradually enters the membrane from residue 3 towards residue 19. The hinge region (residues 17-24), connecting the helical parts of the coat protein, is found to be more embedded in the membrane. Substitution of one or more of the membrane-anchoring amino acid residues lysine 8, phenylalanine 11 and leucine 14, results in a rearrangement of the N-terminal protein part into a more extended conformation. The N-terminal arm can also be forced in this conformation by allowing less space per coat protein at the membrane surface by decreasing the lipid to protein ratio. The influence of the phospholipid headgroup composition on the rearrangement of the N-terminal part of the protein is found to be negligible within the range thought to be relevant in vivo. From our experiments we conclude that membrane-anchoring and space-limiting effects are key factors for the structural rearrangement of the N-terminal protein part of the coat protein in the membrane.

Conformational changes in oligo-nucleotides upon binding to a peptide representing the N-terminal region of a viral coat protein. An optical spectroscopy study.

Conformational changes of the oligonucleotides r(A)12, d(GC)5, and d(AT)5 upon binding to a pentacosapeptide representing the RNA-binding N-terminus of the coat protein of cowpea chlorotic mottle virus (CCMV) were studied by using absorption and circular dichroism spectroscopy. The peptide destabilizes the single-stranded structure of r(A)12 at pH 7.2, but disrupts the double-stranded structure of r(A)12 at pH 5.0. However, at pH 4.0, the peptide is not able to disrupt this double-stranded structure. The double-stranded structures of d(GC)5 and d(AT)5 with Watson-Crick base-pairing are stabilized upon binding to the peptide.

Deuterium nuclear magnetic resonance investigation of bacteriophage M13 coat protein in dimyristoylphosphatidylcholine liposomes using palmitic acid as a probe.

The effect of incorporation of various amounts of M13 bacteriophage coat protein on the bilayer order and acyl chain motion in dimyristoylphosphatidylcholine (DMPC) liposomes has been investigated using deuterium NMR of specifically deuterated palmitic acid as a bilayer probe, phosphorus NMR and additional spin-label electron spin resonance (ESR). The secondary structure of the M13 coat protein in these bilayers was determined from circular dichroism spectra. Phosphorus NMR spectra of the mixed liposomes are characteristic for DMPC organized in bilayers, also after incorporation of various levels of M13 protein. Circular dichroism spectra of the coat protein indicate that the protein conformation is predominantly a beta-structure (more than 75%). Various incorporation levels of M13 coat protein do not affect the order of the deuterium-labelled positions along the acyl chain at the carbon-2, 9 and 16 positions. In contrast, the spin-spin relaxation times decrease at higher protein levels, especially at the carbon-16 position. The spin-label ESR spectra of the same system using 14-doxylstearic acid as a label show a second, motionally restricted component, that is not observed by deuterium NMR. The NMR and ESR results are consistent with a model in which the fatty acid molecules are in a fast two-site exchange (at a rate of approx. 10(7) Hz) between the sites in the bulk of the lipid bilayer and the motionally restricted sites on the coat protein.

Conformation and orientation of the gene 9 minor coat protein of bacteriophage M13 in phospholipid bilayers.

The membrane-bound state of the gene 9 minor coat protein of bacteriophage M13 was studied in model membrane systems, which varied in lipid head group and lipid acyl chain composition. By using FTIR spectroscopy and subsequent band analysis a quantitative analysis of the secondary structure of the protein was obtained. The secondary structure of the gene 9 protein predominantly consists of alpha-helical (67%) and turn (33%) structures. The turn structure is likely to be located C-terminally where it has a function in recognizing the phage DNA during bacteriophage assembly. Attenuated total reflection FTIR spectroscopy was used to determine the orientation of gene 9 protein in the membrane, revealing that the alpha-helical domain is mainly transmembrane. The conformational and orientational measurements result in two models for the gene 9 protein in the membrane: a single transmembrane helix model and a two-helix model consisting of a 15 amino acid long transmembrane helix and a 10 amino acid long helix oriented parallel to the membrane plane. Potential structural consequences for both models are discussed.

Spontaneous insertion of gene 9 minor coat protein of bacteriophage M13 in model membranes.

Gene 9 minor coat protein from bacteriophage M13 is known to be located in the inner membrane after phage infection of Escherichia coli. The way of insertion of this small protein (32 amino acids) into membranes is still unknown. Here we show that the protein is able to insert in monolayers. The limiting surface pressure of 35 mN/m for 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphoglycerol lipid systems indicates that this spontaneous insertion can also occur in vivo. By carboxyfluorescein leakage experiments of vesicles it is demonstrated that protein monomers, or at least small aggregates, are more effective in releasing carboxyfluorescein than highly aggregated protein. The final orientation of the protein in the bilayer after insertion was addressed by proteinase K digestion, thereby making use of the unique C-terminal location of the antigenic binding site. After insertion the C-terminus is still available for the enzymatic digestion, while the N-terminus is not. This leads to the overall conclusion that the protein is able to insert spontaneously into membranes without the need of any machinery or transmembrane gradient, with the positively charged C-terminus remaining on the outside. The orientation after insertion of gene 9 protein is in agreement with the 'positive inside rule'.

Severe burn cases : a tentative analysis of the epidemiological, clinical and bacteriological data.

We have computerized the epidemiological, clinical and bacteriological data of 140 serious burn cases, hospitalized in our intensive care unit (I.C.U.). The most relevant conclusions are : Epidemiology : it is imperative to exert a prophylactic action against domestic burns through scalds and explosions. Clinic : the most frequent cause of death, during the phases of sept icemia, is cardiac failure. Lung burns significantly increase the mortality rate and they are impossible to codify in the classical systems of burn descriptions. Bacteriology : preventive antibiotic therapy determines, after three days, the strains which resist the usual antibiotics. An antibiotic loses some of its effect when used intensively in reanimation care. We express our concern with regard to the efficiency of antibiotic therapy in intensive care. We suggest to try and standardize data collection, so that multi-centre studies may help to increase the efficiency of their processing.

Aggregation-related conformational change of the membrane-associated coat protein of bacteriophage M13.

The state of the coat protein of bacteriophage M13, reconstituted into amphiphilic media, has been investigated. The in situ conformation of the coat protein has been determined by using circular dichroism. Minimum numbers for the protein aggregation in the system have been determined after disruption of the lipid-protein system and subsequent uptake of the protein in cholate micelles. The aggregational state and conformation of the protein were affected by (1) the method of coat protein isolation (phenol extraction vs cholate isolation), (2) the nature of amphiphiles used (variation in phospholipid headgroups and acyl chains), and (3) the ratio of amphiphiles and protein. Under all conditions, phenol-extracted coat protein was in a predominantly beta-structure and in a highly aggregated polymeric form. Cholate-isolated coat protein was initially oligomeric and contained a substantial amount of alpha-helix. Below an aggregation number of 20, this protein showed a reversible aggregation with no change in conformation. Upon further aggregation, a conformational change was observed, and aggregation was irreversible, resulting in predominantly beta-structured coat protein polymers. This effect was observed upon uptake in phospholipids at low lipid to protein molar ratios (L/P ratios) and with phosphatidylcholines (PC) and phosphatidic acids (PA) containing saturated acyl chains. After reconstitution in phospholipids with unsaturated acyl chains and with phosphatidylglycerols (PG) at high L/P ratios, the original alpha-helix-containing state of the coat protein was maintained. Cross-linking experiments demonstrated that the beta-polymers are able to form reversible superaggregates within the vesicle system. An aggregation-related conformational change mechanism for the coat protein in phospholipid systems is proposed.

Spin-label electron spin resonance study of bacteriophage M13 coat protein incorporation into mixed lipid bilayers.

The major coat protein of bacteriophage M13 was incorporated in mixed dimyristoylphosphatidylcholine/dimyristoylphosphatidylglycerol (80/20 w/w) vesicles probed with different spin-labeled phospholipids, labeled on the C-14 atom of the sn-2 chain. The specificity for a series of phospholipids was determined from a motionally restricted component seen in the electron spin resonance (ESR) spectra of vesicles with the coat protein incorporated. At 30 degrees C and pH 8, the fraction of motionally restricted phosphatidic acid spin-label is 0.36, 0.52, and 0.72 for lipid/protein ratios of 18, 14, and 9 mol/mol, respectively. The ESR spectra, analyzed by digital subtraction, resulted in a phospholipid preference following the pattern cardiolipin = phosphatidic acid greater than stearic acid = phosphatidylserine = phosphatidylglycerol greater than phosphatidylcholine = phosphatidylethanolamine. The specificities found are related to the composition of the target Escherichia coli cytoplasmic membrane.

Local dynamics of the M13 major coat protein in different membrane-mimicking systems.

The local environment of the transmembrane and C-terminal domain of M13 major coat protein was probed by site-directed ESR spin labeling when the protein was introduced into three membrane-mimicking systems, DOPC vesicles, sodium cholate micelles, and SDS micelles. For this purpose, we have inserted unique cysteine residues at specific positions in the transmembrane and C-terminal region, using site-directed mutagenesis. Seven viable mutants with reasonable yield were harvested: A25C, V31C, T36C, G38C, T46C, A49C, and S50C. The mutant coat proteins were indistinguishable from wild type M13 coat protein with respect to their conformational and aggregational properties. The ESR data suggest that the amino acid positions 25 and 46 of the coat protein in DOPC vesicles are located close to the membrane-water interface. In this way the lysines at positions 40, 43, and 44 and the phenylalanines at positions 42 and 45 act as hydrophilic and hydrophobic anchors, respectively. The ESR spectra of site specific maleimido spin-labeled mutant coat proteins reconstituted into DOPC vesicles and solubilized in sodium cholate or SDS indicate that the local dynamics of the major coat protein is significantly affected by its structural environment (micellar vs bilayer), location (aqueous vs hydrophobic), and lipid/protein ratio. The detergents SDS and sodium cholate sufficiently well solubilize the major coat protein and largely retain its secondary structure elements. However, the results indicate that they have a poorly defined protein-amphiphilic structure and lipid-water interface as compared to bilayers and thus are not a good substitute for lipid bilayers in biophysical studies.

Membrane assembly of the bacteriophage Pf3 major coat protein.

The Pf3 major coat protein of the Pf3 bacteriophage is stored in the inner membrane of the infected cell during the reproductive cycle. The protein consists of 44 amino acids, and contains an acidic amphipathic N-terminal domain, a hydrophobic domain, and a short basic C-terminal domain. The mainly alpha-helical membrane-bound protein traverses the membrane once, leaving the C-terminus in the cytoplasm and the N-terminus in the periplasm. A cysteine-scanning approach was followed to measure which part of the membrane-bound Pf3 protein is inside or outside the membrane. In this approach, the fluorescence probe N-[(iodoacetyl)amino]ethyl-1-sulfonaphthylamine (IAEDANS) was attached to single-cysteine mutants of the Pf3 coat protein. The labeled mutant coat proteins were reconstituted into the phospholipid DOPC/DOPG (80/20 molar ratio) and DOPE/DOPG (80/20 molar ratio) model membranes. We subsequently studied the fluorescence characteristics at the different positions in the protein. We measured the local polarity of the environment of the probe, as well as the accessibility of the probe to the fluorescence quencher acrylamide. The results of this study show a single membrane-spanning protein with both the C- and N-termini remaining close to the surface of the membrane. A nearly identical result was seen previously for the membrane-bound M13 coat protein. On the basis of a comparison between the results from both studies, we suggest an "L-shaped" membrane-bound model for the Pf3 coat protein. DOPE-containing model membranes revealed a higher polarity, and quenching efficiency at the membrane/water interface. Furthermore, from the outside to the inside of the membrane, a steeper polarity gradient was measured at the PE/PG interface as compared to the PC/PG interface. These results suggest a thinner interface for DOPE/DOPG than for DOPC/DOPG membranes.

In situ aggregational state of M13 bacteriophage major coat protein in sodium cholate and lipid bilayers.

The in situ aggregational behavior of the bacteriophage M13 major coat protein was determined for the protein isolated in sodium cholate and reconstituted into DOPC lipid bilayers. For this purpose, the cysteine mutants A49C and T36C of the major coat protein were labeled with either a maleimido spin-label or a fluorescence label (IAEDANS). The steric restrictions sensed by the spin-label were used to evaluate the local protein conformation and the extent of protein-protein interactions at the position of the labeled residue. In addition, fluorescent labels covalently attached to the protein were used to determine the polarity of the local environment. The labeled coat protein mutants were examined under different conditions of protein association (amphiphile environment, ionic strength, temperature, and pH). The aggregational state of the major coat protein solubilized from the phage particle in sodium cholate was not dependent on the ionic strength, but was strongly dependent on cholate concentration and pH during sample preparation. At pH 7.0 and high sodium cholate concentration, the protein was in a dimeric form. The unusually strong association properties of the protein dimer in sodium cholate at pH 7.0 were attributed to the inability of sodium cholate to disrupt the strong hydrophobic forces between neighboring protein subunits in the phage particle. Such a "structural protein dimer" was, however, completely and irreversibly disrupted at pH 10.0. Qualitatively the same aggregational tendency was found upon changing the pH for the coat protein reconstituted in DOPC lipid bilayers. This reveals that the dimer disruption process is primarily a protein property, because there are no titratable groups on DOPC in the experimental pH range. The results are interpreted in terms of a model relating the protein aggregational state in the assembled phage to the protein aggregational behavior in sodium cholate and lipid bilayers.

Characterization of the brome mosaic virus movement protein expressed in E. coli.

The biochemical and functional properties of the movement protein (MP) of brome mosaic virus (BMV) were investigated. Expression and purification of the BMV MP from Escherichia coli resulted in a pure and soluble protein preparation. Sucrose gradient centrifugation revealed that BMV MP forms oligomers consisting of two or more copies but no higher order multimers even when different ionic strengths and pHs were applied. Nitro-cellulose filter binding and gel retardation studies showed that in vitro the BMV MP preferentially bound to ss nucleic acids (RNA and DNA); the affinity to ssRNA was lower compared to BMV coat protein. The binding to ss nucleic acid was cooperative and not sequence specific and the hypothetical binding site was calculated to be around three to six nucleotides per MP monomer. The nucleic acid binding properties of the BMV MP are discussed in relation to the recent finding that this protein is also able to form tubular structures in infected protoplasts.

Conformational and aggregational properties of the gene 9 minor coat protein of bacteriophage M13 in membrane-mimicking systems.

The membrane-bound state of the gene 9 minor coat protein of bacteriophage M13 was studied in various membrane-mimicking systems, including organic solvents, detergent micelles, and phospholipid bilayers. For this purpose we determined the conformational and aggregational properties of the chemically synthesized protein by CD, FTIR, and HPSEC. The protein appears to be in a monomeric or small oligomeric alpha-helical state in TFE but adopts a mixture of alpha-helical and random structure after subsequent incorporation into SDS or DOPG. When solubilized by sodium cholate, however, the protein undergoes a transition in time into large aggregates, which contain mainly beta-sheet conformation. The rate of this beta-polymerization process was decreased at lower temperature and higher concentrations of sodium cholate. This aggregation was reversed only upon addition of high concentrations of the strong detergent SDS. By reconstitution of the cholate-solubilized protein into DOPG, it was found that the state of the protein, whether initially alpha-helical monomeric/oligomeric or beta-sheet aggregate, did not change. On the basis of our results, we propose that the principal conformational state of membrane-bound gene 9 protein in vivo is alpha-helical.