Caspase-8 binding to cardiolipin in giant unilamellar vesicles provides a functional docking platform for bid

Caspase-8 is involved in death receptor-mediated apoptosis in type II cells, the proapoptotic programme of which is triggered by truncated Bid. Indeed, caspase-8 and Bid are the known intermediates of this signalling pathway. Cardiolipin has been shown to provide an anchor and an essential activating platform for caspase-8 at the mitochondrial membrane surface. Destabilisation of this platform alters receptor-mediated apoptosis in diseases such as Barth Syndrome, which is characterised by the presence of immature cardiolipin which does not allow caspase-8 binding. We used a simplified in vitro system that mimics contact sites and/or cardiolipin-enriched microdomains at the outer mitochondrial surface in which the platform consisting of caspase-8, Bid and cardiolipin was reconstituted in giant unilamellar vesicles. We analysed these vesicles by flow cytometry and confirm previous results that demonstrate the requirement for intact mature cardiolipin for caspase-8 activation and Bid binding and cleavage. We also used confocal microscopy to visualise the rupture of the vesicles and their revesiculation at smaller sizes due to alteration of the curvature following caspase-8 and Bid binding. Biophysical approaches, including Laurdan fluorescence and rupture/tension measurements, were used to determine the ability of these three components (cardiolipin, caspase-8 and Bid) to fulfil the minimal requirements for the formation and function of the platform at the mitochondrial membrane. Our results shed light on the active functional role of cardiolipin, bridging the gap between death receptors and mitochondria

Fostering implementation of health services research findings into practice: a consolidated framework for advancing implementation science

Abstract Background Many interventions found to be effective in health services research studies fail to translate into meaningful patient care outcomes across multiple contexts. Health services researchers recognize the need to evaluate not only summative outcomes but also formative outcomes to assess the extent to which implementation is effective in a specific setting, prolongs sustainability, and promotes dissemination into other settings. Many implementation theories have been published to help promote effective implementation. However, they overlap considerably in the constructs included in individual theories, and a comparison of theories reveals that each is missing important constructs included in other theories. In addition, terminology and definitions are not consistent across theories. We describe the Consolidated Framework For Implementation Research (CFIR) that offers an overarching typology to promote implementation theory development and verification about what works where and why across multiple contexts. Methods We used a snowball sampling approach to identify published theories that were evaluated to identify constructs based on strength of conceptual or empirical support for influence on implementation, consistency in definitions, alignment with our own findings, and potential for measurement. We combined constructs across published theories that had different labels but were redundant or overlapping in definition, and we parsed apart constructs that conflated underlying concepts. Results The CFIR is composed of five major domains: intervention characteristics, outer setting, inner setting, characteristics of the individuals involved, and the process of implementation. Eight constructs were identified related to the intervention (e.g., evidence strength and quality), four constructs were identified related to outer setting (e.g., patient needs and resources), 12 constructs were identified related to inner setting (e.g., culture, leadership engagement), five constructs were identified related to individual characteristics, and eight constructs were identified related to process (e.g., plan, evaluate, and reflect). We present explicit definitions for each construct. Conclusion The CFIR provides a pragmatic structure for approaching complex, interacting, multi-level, and transient states of constructs in the real world by embracing, consolidating, and unifying key constructs from published implementation theories. It can be used to guide formative evaluations and build the implementation knowledge base across multiple studies and settings.http://deepblue.lib.umich.edu/bitstream/2027.42/78272/1/1748-5908-4-50.xmlhttp://deepblue.lib.umich.edu/bitstream/2027.42/78272/2/1748-5908-4-50-S1.PDFhttp://deepblue.lib.umich.edu/bitstream/2027.42/78272/3/1748-5908-4-50-S3.PDFhttp://deepblue.lib.umich.edu/bitstream/2027.42/78272/4/1748-5908-4-50-S4.PDFhttp://deepblue.lib.umich.edu/bitstream/2027.42/78272/5/1748-5908-4-50.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/78272/6/1748-5908-4-50-S2.PDFPeer Reviewe

On the effects of mechanical stress of biological membranes in modeling of swelling dynamics of biological systems

We highlight mechanical stretching and bending of membranes and the importance of membrane deformations in the analysis of swelling dynamics of biological systems, including cells and subcellular organelles. Membrane deformation upon swelling generates tensile stress and internal pressure, contributing to volume changes in biological systems. Therefore, in addition to physical (internal/external) and chemical factors, mechanical properties of the membranes should be considered in modeling analysis of cellular swelling. Here we describe an approach that considers mechanical properties of the membranes in the analysis of swelling dynamics of biological systems. This approach includes membrane bending and stretching deformations into the model, producing a more realistic description of swelling. We also discuss the effects of membrane stretching on swelling dynamics. We report that additional pressure generated by membrane bending is negligible, compared to pressures generated by membrane stretching, when both membrane surface area and volume are variable parameters. Note that bending deformations are reversible, while stretching deformation may be irreversible, leading to membrane disruption when they exceed a certain threshold level. Therefore, bending deformations need only be considered in reversible physiological swelling, whereas stretching deformations should also be considered in pathological irreversible swelling. Thus, the currently proposed approach may be used to develop a detailed biophysical model describing the transition from physiological to pathological swelling mode.National Aeronautics & Space Administration (NASA):80NSSC19M0049; PR Space Grant (NASA):NNX15AI11Hinfo:eu-repo/semantics/publishedVersio

Identification of serum angiopoietin-2 as a biomarker for clinical outcome of colorectal cancer patients treated with bevacizumab-containing therapy

BACKGROUND: The combination of chemotherapy with the vascular endothelial growth factor (VEGF) antibody bevacizumab is a standard of care in advanced colorectal cancer (CRC). However, biomarkers predicting outcome of bevacizumab-containing treatment are lacking. As angiopoietin-2 (Ang-2) is a key regulator of vascular remodelling in concert with VEGF, we investigated its role as a biomarker in metastatic CRC. METHODS: Serum Ang-2 levels were measured in 33 healthy volunteers and 90 patients with CRC. Of these, 34 had metastatic disease and received bevacizumab-containing therapy. To determine the tissue of origin of Ang-2, quantitative real-time PCR was performed on microdissected cryosections of human CRC and in a murine xenograft model of CRC using species-specific amplification. RESULTS: Ang-2 originated from the stromal compartment of CRC tissues. Serum Ang-2 levels were significantly elevated in patients with metastatic CRC compared with healthy controls. Amongst patients receiving bevacizumab-containing treatment, low pre-therapeutic serum Ang-2 levels were associated with a significant better response rate (82 vs 31%; P<0.01), a prolonged median progression-free survival (14.1 vs 8.5 months; P<0.01) and a reduction of 91% in the hazard of death (P<0.05). CONCLUSION: Serum Ang-2 is a candidate biomarker for outcome of patients with metastatic CRC treated with bevacizumab-containing therapy, and it should be further validated to customise combined chemotherapeutic and anti-angiogenic treatment. British Journal of Cancer (2010) 103, 1407-1414. doi: 10.1038/sj.bjc.6605925 www.bjcancer.com Published online 5 October 2010 (C) 2010 Cancer Research U

Perturbing Dynamin Reveals Potent Effects on the Drosophila Circadian Clock

BACKGROUND: Transcriptional feedback loops are central to circadian clock function. However, the role of neural activity and membrane events in molecular rhythms in the fruit fly Drosophila is unclear. To address this question, we expressed a temperature-sensitive, dominant negative allele of the fly homolog of dynamin called shibire(ts1) (shi(ts1)), an active component in membrane vesicle scission. PRINCIPAL FINDINGS: Broad expression in clock cells resulted in unexpectedly long, robust periods (>28 hours) comparable to perturbation of core clock components, suggesting an unappreciated role of membrane dynamics in setting period. Expression in the pacemaker lateral ventral neurons (LNv) was necessary and sufficient for this effect. Manipulation of other endocytic components exacerbated shi(ts1)'s behavioral effects, suggesting its mechanism is specific to endocytic regulation. PKA overexpression rescued period effects suggesting shi(ts1) may downregulate PKA pathways. Levels of the clock component PERIOD were reduced in the shi(ts1)-expressing pacemaker small LNv of flies held at a fully restrictive temperature (29 degrees C). Less restrictive conditions (25 degrees C) delayed cycling proportional to observed behavioral changes. Levels of the neuropeptide PIGMENT-DISPERSING FACTOR (PDF), the only known LNv neurotransmitter, were also reduced, but PERIOD cycling was still delayed in flies lacking PDF, implicating a PDF-independent process. Further, shi(ts1) expression in the eye also results in reduced PER protein and per and vri transcript levels, suggesting that shibire-dependent signaling extends to peripheral clocks. The level of nuclear CLK, transcriptional activator of many core clock genes, is also reduced in shi(ts1) flies, and Clk overexpression suppresses the period-altering effects of shi(ts1). CONCLUSIONS: We propose that membrane protein turnover through endocytic regulation of PKA pathways modulates the core clock by altering CLK levels and/or activity. These results suggest an important role for membrane scission in setting circadian period

Models of care for musculoskeletal health: A cross-sectional qualitative study of Australian stakeholders' perspectives on relevance and standardised evaluation

Background: The prevalence and impact of musculoskeletal conditions are predicted to rapidly escalate in the coming decades. Effective strategies are required to minimise 'evidence-practice', 'burden-policy' and 'burden-service' gaps and optimise health system responsiveness for sustainable, best-practice healthcare. One mechanism by which evidence can be translated into practice and policy is through Models of Care (MoCs), which provide a blueprint for health services planning and delivery. While evidence supports the effectiveness of musculoskeletal MoCs for improving health outcomes and system efficiencies, no standardised national approach to evaluation in terms of their 'readiness' for implementation and 'success' after implementation, is yet available. Further, the value assigned to MoCs by end users is uncertain. This qualitative study aimed to explore end users' views on the relevance of musculoskeletal MoCs to their work and value of a standardised evaluation approach. Methods: A cross-sectional qualitative study was undertaken. Subject matter experts (SMEs) with health, policy and administration and consumer backgrounds were drawn from three Australian states. A semi-structured interview schedule was developed and piloted to explore perceptions about musculoskeletal MoCs including: i) aspects important to their work (or life, for consumers) ii) usefulness of standardised evaluation frameworks to judge 'readiness' and 'success' and iii) challenges associated with standardised evaluation. Verbatim transcripts were analysed by two researchers using a grounded theory approach to derive key themes. Results: Twenty-seven SMEs (n = 19; 70.4 % female) including five (18.5 %) consumers participated in the study. MoCs were perceived as critical for influencing and initiating changes to best-practice healthcare planning and delivery and providing practical guidance on how to implement and evaluate services. A 'readiness' evaluation framework assessing whether critical components across the health system had been considered prior to implementation was strongly supported, while 'success' was perceived as an already familiar evaluation concept. Perceived challenges associated with standardised evaluation included identifying, defining and measuring key 'readiness' and 'success' indicators; impacts of systems and context changes; cost; meaningful stakeholder consultation and developing a widely applicable framework. Conclusions: A standardised evaluation framework that includes a strong focus on 'readiness' is important to ensure successful and sustainable implementation of musculoskeletal MoCs

The Effect of Performance-Based Financial Incentives on Improving Patient Care Experiences: A Statewide Evaluation

Patient experience measures are central to many pay-for-performance (P4P) programs nationally, but the effect of performance-based financial incentives on improving patient care experiences has not been assessed. The study uses Clinician &amp; Group CAHPS data from commercially insured adult patients (n = 124,021) who had visits with 1,444 primary care physicians from 25 California medical groups between 2003 and 2006. Medical directors were interviewed to assess the magnitude and nature of financial incentives directed at individual physicians and the patient experience improvement activities adopted by groups. Multilevel regression models were used to assess the relationship between performance change on patient care experience measures and medical group characteristics, financial incentives, and performance improvement activities. Over the course of the study period, physicians improved performance on the physician-patient communication (0.62 point annual increase, p &lt; 0.001), care coordination (0.48 point annual increase, p &lt; 0.001), and office staff interaction (0.22 point annual increase, p = 0.02) measures. Physicians with lower baseline performance on patient experience measures experienced larger improvements (p &lt; 0.001). Greater emphasis on clinical quality and patient experience criteria in individual physician incentive formulas was associated with larger improvements on the care coordination (p &lt; 0.01) and office staff interaction (p &lt; 0.01) measures. By contrast, greater emphasis on productivity and efficiency criteria was associated with declines in performance on the physician communication (p &lt; 0.01) and office staff interaction (p &lt; 0.001) composites. In the context of statewide measurement, reporting, and performance-based financial incentives, patient care experiences significantly improved. In order to promote patient-centered care in pay for performance and public reporting programs, the mechanisms by which program features influence performance improvement should be clarified

The Serotonin 5-HT7Dro Receptor Is Expressed in the Brain of Drosophila, and Is Essential for Normal Courtship and Mating

The 5-HT7 receptor remains one of the less well characterized serotonin receptors. Although it has been demonstrated to be involved in the regulation of mood, sleep, and circadian rhythms, as well as relaxation of vascular smooth muscles in mammals, the precise mechanisms underlying these functions remain largely unknown. The fruit fly, Drosophila melanogaster, is an attractive model organism to study neuropharmacological, molecular, and behavioral processes that are largely conserved with mammals. Drosophila express a homolog of the mammalian 5-HT7 receptor, as well as homologs for the mammalian 5-HT1A, and 5-HT2, receptors. Each fly receptor couples to the same effector pathway as their mammalian counterpart and have been demonstrated to mediate similar behavioral responses. Here, we report on the expression and function of the 5-HT7Dro receptor in Drosophila. In the larval central nervous system, expression is detected postsynaptically in discreet cells and neuronal circuits. In the adult brain there is strong expression in all large-field R neurons that innervate the ellipsoid body, as well as in a small group of cells that cluster with the PDF-positive LNvs neurons that mediate circadian activity. Following both pharmacological and genetic approaches, we have found that 5-HT7Dro activity is essential for normal courtship and mating behaviors in the fly, where it appears to mediate levels of interest in both males and females. This is the first reported evidence of direct involvement of a particular serotonin receptor subtype in courtship and mating in the fly

Quantitative Image Analysis Reveals Distinct Structural Transitions during Aging in Caenorhabditis elegans Tissues

Aging is associated with functional and structural declines in many body systems, even in the absence of underlying disease. In particular, skeletal muscles experience severe declines during aging, a phenomenon termed sarcopenia. Despite the high incidence and severity of sarcopenia, little is known about contributing factors and development. Many studies focus on functional aspects of aging-related tissue decline, while structural details remain understudied. Traditional approaches for quantifying structural changes have assessed individual markers at discrete intervals. Such approaches are inadequate for the complex changes associated with aging. An alternative is to consider changes in overall morphology rather than in specific markers. We have used this approach to quantitatively track tissue architecture during adulthood and aging in the C. elegans pharynx, the neuromuscular feeding organ. Using pattern recognition to analyze aged-grouped pharynx images, we identified discrete step-wise transitions between distinct morphologies. The morphology state transitions were maintained in mutants with pharynx neurotransmission defects, although the pace of the transitions was altered. Longitudinal measurements of pharynx function identified a predictive relationship between mid-life pharynx morphology and function at later ages. These studies demonstrate for the first time that adult tissues undergo distinct structural transitions reflecting postdevelopmental events. The processes that underlie these architectural changes may contribute to increased disease risk during aging, and may be targets for factors that alter the aging rate. This work further demonstrates that pattern analysis of an image series offers a novel and generally accessible approach for quantifying morphological changes and identifying structural biomarkers

Evidence for biological roots in the transgenerational transmission of intimate partner violence

Intimate partner violence is a ubiquitous and devastating phenomenon for which effective interventions and a clear etiological understanding are still lacking. A major risk factor for violence perpetration is childhood exposure to violence, prompting the proposal that social learning is a major contributor to the transgenerational transmission of violence. Using an animal model devoid of human cultural factors, we showed that male rats became highly aggressive against their female partners as adults after exposure to non-social stressful experiences in their youth. Their offspring also showed increased aggression toward females in the absence of postnatal father–offspring interaction or any other exposure to violence. Both the females that cohabited with the stressed males and those that cohabited with their male offspring showed behavioral (including anxiety- and depression-like behaviors), physiological (decreased body weight and basal corticosterone levels) and neurobiological symptoms (increased activity in dorsal raphe serotonergic neurons in response to an unfamiliar male) resembling the alterations described in abused and depressed women. With the caution required when translating animal work to humans, our findings extend current psychosocial explanations of the transgenerational transmission of intimate partner violence by strongly suggesting an important role for biological factors