What is important, what needs treating? How GPs perceive older patients' multiple health problems: a mixed method research study

ABSTRACT: BACKGROUND: GPs increasingly deal with multiple health problems of their older patients. They have to apply a hierarchical management approach that considers priorities to balance competing needs for treatment. Yet, the practice of setting individual priorities in older patients is largely unexplored. This paper analyses the GPs' perceptions on important and unimportant health problems and how these affect their treatment. METHODS: GPs appraised the importance of health problems for a purposive sample of their older patients in semi-structured interviews. Prior to the interviews, the GPs had received a list of their patients' health problems resulting from a geriatric assessment and were asked to rate the importance of each identified problem. In the interviews the GPs subsequently explained why they considered certain health problems important or not and how this affected treatment. Data was analysed using qualitative content analysis and quantitative methods. RESULTS: The problems GPs perceive as important are those that are medical and require active treatment or monitoring, or that induce empathy or awareness but cannot be assisted further. Unimportant problems are those that are well managed problems and need no further attention as well as age-related conditions or functional disabilities that provoke fatalism, or those considered outside the GPs' responsibility. Statements of professional actions are closely linked to explanations of important problems and relate to physical problems rather than functional and social patient issues. CONCLUSIONS: GPs tend to prioritise treatable clinical conditions. Treatment approaches are, however, vague or missing for complex chronic illnesses and disabilities. Here, patient empowerment strategies are of value and need to be developed and implemented. The professional concepts of ageing and disability should not impede but rather foster treatment and care. To this end, GPs need to be able to delegate care to a functioning primary care team. Trial Registration German Trial Register (DRKS): 00000792.     

Combining ultracentrifugation and peptide termini group-specific immunoprecipitation for multiplex plasma protein analysis

Blood plasma is a valuable source of potential biomarkers. However, its complexity and the huge dynamic concentration range of its constituents complicate its analysis. To tackle this problem, an immunoprecipitation strategy was employed using antibodies directed against short terminal epitope tags (triple X proteomics antibodies), which allow the enrichment of groups of signature peptides derived from trypsin-digested plasma. Isolated signature peptides are subsequently detected using MALDI-TOF/TOF mass spectrometry. Sensitivity of the immunoaffinity approach was, however, compromised by the presence of contaminant peaks derived from the peptides of nontargeted high abundant proteins. A closer analysis of the enrichment strategy revealed nonspecific peptide binding to the solid phase affinity matrix as the major source of the contaminating peptides. We therefore implemented a sucrose density gradient ultracentrifugation separation step into the procedure. This yielded a 99% depletion of contaminating peptides from a sucrose fraction containing 70% of the peptide-antibody complexes and enabled the detection of the previously undetected low abundance protein filamin-A. Assessment of this novel approach using 15 different triple X proteomics antibodies demonstrated a more consistent detection of a greater number of targeted peptides and a significant reduction in the intensity of nonspecific peptides. Ultracentrifugation coupled with immunoaffinity MS approaches presents a powerful tool for multiplexed plasma protein analysis without the requirement for demanding liquid chromatography separation techniques.     

Find Pairs: The Module for Protein Quantification of the PeakQuant Software Suite

Abstract Accurate quantification of proteins is one of the major tasks in current proteomics research. To address this issue, a wide range of stable isotope labeling techniques have been developed, allowing one to quantitatively study thousands of proteins by means of mass spectrometry. In this article, the FindPairs module of the PeakQuant software suite is detailed. It facilitates the automatic determination of protein abundance ratios based on the automated analysis of stable isotope-coded mass spectrometric data. Furthermore, it implements statistical methods to determine outliers due to biological as well as technical variance of proteome data obtained in replicate experiments. This provides an important means to evaluate the significance in obtained protein expression data. For demonstrating the high applicability of FindPairs, we focused on the quantitative analysis of proteome data acquired in (14)N/(15)N labeling experiments. We further provide a comprehensive overview of the features of the FindPairs software, and compare these with existing quantification packages. The software presented here supports a wide range of proteomics applications, allowing one to quantitatively assess data derived from different stable isotope labeling approaches, such as (14)N/(15)N labeling, SILAC, and iTRAQ. The software is publicly available at http://www.medizinisches-proteom-center.de/software and free for academic use.     

A molecular modeling analysis of diethylstilbestrol conformations and their similarity to estradiol-17β

Crystallographic and computer modeling studies throughout the last 25 years have shown the structure of diethylstilbestrol (DES) to exist in two symmetrical or one asymmetrical conformation. As a result of specific comparisons to estradiol-17β (E₂), the asymmetrical DES conformer has been suggested as the geometry possessing estrogenic activity. In the present study, a more complete set of DES conformations has been elucidated through the use of computer modeling. All previously defined DES geometries were found within this new set of ten structural forms. Differences between the molecular mechanics heat of formation energies of the ten conformers, as well as the transition energies separating them from each other, were found to be less than I kcal/mol. Additionally, a computer-based molecular alignment method was employed to quantitatively compare the steric and electrostatic molecular features of each DES conformer relative to E₂. All ten DES structures were found to have shape relationships similar to E₂. Thus, a model for the estrogen action of DES is presented whereby this stilbene can favorably interact with the estrogen receptor regardless of the conformation or orientation of the initial ligand-receptor association.     

Myosin-10 and actin filaments are essential for mitotic spindle function

Mitotic spindles are microtubule-based structures responsible for chromosome partitioning during cell division. Although the roles of microtubules and microtubule-based motors in mitotic spindles are well established, whether or not actin filaments (F-actin) and F-actin-based motors (myosins) are required components of mitotic spindles has long been controversial. Based on the demonstration that myosin-10 (Myo10) is important for assembly of meiotic spindles, we assessed the role of this unconventional myosin, as well as F-actin, in mitotic spindles. We find that Myo10 localizes to mitotic spindle poles and is essential for proper spindle anchoring, normal spindle length, spindle pole integrity, and progression through metaphase. Furthermore, we show that F-actin localizes to mitotic spindles in dynamic cables that surround the spindle and extend between the spindle and the cortex. Remarkably, although proper anchoring depends on both F-actin and Myo10, the requirement for Myo10 in spindle pole integrity is F-actin independent, whereas F-actin and Myo10 actually play antagonistic roles in maintenance of spindle length.     

Pilosulin 5, a novel histamine-releasing peptide of the Australian ant, Myrmecia pilosula (Jack Jumper Ant)

Venom of the Australian ant species Myrmecia pilosula contains a number of allergenic peptides including pilosulins. To obtain novel cDNA clones that encode the pilosulin-related bioactive peptides, mRNA of M. pilosula species complex was subjected to RT-PCR in which the forward primer corresponds to a nucleotide sequence in the leader sequences of pilosulins. We isolated a cDNA clone encoding the novel bioactive peptide pilosulin 5. Tandem mass analysis was entirely consistent with the cDNA derived sequence, and indicated that pilosulin 5 is connected by a single disulfide bridge to create a dimmer peptide of 8546 Da. Synthetic pilosulin 5 peptide caused a significant histamine release in a dose-dependent manner, and the mastoparan homologous region of pilosulin 5 was responsible for the activity.