A Top-Down LC-FTICR MS-Based Strategy for Characterizing Oxidized Calmodulin in Activated Macrophages

A liquid chromatography-mass spectrometry (LC-MS)-based approach for characterizing the degree of nitration and oxidation of intact calmodulin (CaM) has been used to resolve ∼250 CaM oxiforms using only 500 ng of protein. The analysis was based on high-resolution data of the intact CaM isoforms obtained by Fourier-transform ion cyclotron resonance mass spectrometry (FTICR MS) coupled with an on-line reversed-phase LC separation. Tentative identifications of post-translational modifications (PTMs), such as oxidation or nitration, have been assigned by matching observed protein mass to a database containing all theoretically predicted oxidation products of CaM and verified through a combination of tryptic peptide information (generated from bottom-up analyses) and on-line collisionally induced dissociation (CID) tandem mass spectrometry (MS/MS) at the intact protein level. The reduction in abundance and diversity of oxidatively modified CaM (i.e., nitrated tyrosines and oxidized methionines) induced by macrophage activation has been explored and semiquantified for different oxidation degrees (i.e., no oxidation, moderate, and high oxidation). This work demonstrates the power of the top-down approach to identify and quantify hundreds of combinations of PTMs for single protein target such as CaM and implicate competing repair and peptidase activities to modulate cellular metabolism in response to oxidative stress

The plasma membrane calcium ATPase (PMCA) of neurones is electroneutral and exchanges 2 H+ for each Ca2+ or Ba2+ ion extruded

The coupling between Ca2+ extrusion and H+ uptake by the ubiquitous plasma membrane calcium ATPase (PMCA) has not been measured in any neurone. I have investigated this with Ca2+- and pH-sensitive microelectrodes in large voltage-clamped snail neurones, which have no Na+–Ca2+ exchangers. The recovery of [Ca2+]i and surface pH after a brief depolarization or Ca2+ injection was not slowed by hyperpolarization to −90 mV from a holding potential of −50 mV, consistent with a 1 Ca2+ : 2 H+ coupling ratio. Since Ca2+ injections proved difficult to quantify, and Ca2+ currents through channels were obscured by K+ currents, Ba2+ was used as a substitute. When the cell was bathed in Ca2+-free Ba2+ Ringer solution, the K+ currents were blocked and large inward currents were revealed on depolarization. The Ca2+-sensitive microelectrodes were sensitive to intracellular Ba2+ as well as Ca2+. With equal depolarizations Ba2+ entry appeared larger than Ca2+ entry and generated similar but slower pH changes. Ba2+ extrusion was insensitive to hyperpolarization, blocked by eosin or high pH, and about 5 times slower than Ca2+ extrusion. The ratio of the pH change caused by the extrusion of unit charge of Ba2+ influx to that caused by unit charge of H+ injection was 0.85 ± 0.08 (s.e.m., n= 8), corresponding to a Ba2+ : H+ ratio of 1 : 1.7. Both this ratio and the electroneutrality of the PMCA suggest that the Ca2+ : H+ ratio is 1 : 2, ensuring that after a Ca2+ influx [Ca2+]i recovery is not influenced by the membrane potential and maximizes the conversion of Ca2+ influxes into possible pH signals

The psychological correlates of distinct neural states occurring during wakeful rest

When unoccupied by an explicit external task, humans engage in a wide range of different types of self-generated thinking. These are often unrelated to the immediate environment and have unique psychological features. Although contemporary perspectives on ongoing thought recognise the heterogeneity of these self-generated states, we lack both a clear understanding of how to classify the specific states, and how they can be mapped empirically. In the current study, we capitalise on advances in machine learning that allow continuous neural data to be divided into a set of distinct temporally re-occurring patterns, or states. We applied this technique to a large set of resting state data in which we also acquired retrospective descriptions of the participants' experiences during the scan. We found that two of the identified states were predictive of patterns of thinking at rest. One state highlighted a pattern of neural activity commonly seen during demanding tasks, and the time individuals spent in this state was associated with descriptions of experience focused on problem solving in the future. A second state was associated with patterns of activity that are commonly seen under less demanding conditions, and the time spent in it was linked to reports of intrusive thoughts about the past. Finally, we found that these two neural states tended to fall at either end of a neural hierarchy that is thought to reflect the brain's response to cognitive demands. Together, these results demonstrate that approaches which take advantage of time-varying changes in neural function can play an important role in understanding the repertoire of self-generated states. Moreover, they establish that important features of self-generated ongoing experience are related to variation along a similar vein to those seen when the brain responds to cognitive task demands

Academic doctors' views of complementary and alternative medicine (CAM) and its role within the NHS: an exploratory qualitative study

<p>Abstract</p> <p>Background</p> <p>There has been a marked increase in the use of complementary and alternative medicine (CAM) in the UK population in recent years. Surveys of doctors' perspectives on CAM have identified a variety of views and potential information needs. While these are useful for describing the proportions of doctors who hold particular attitudes towards CAM, they are less helpful for understanding why. In addition, while the views of non-academic doctors have begun to be studied, the perspective and rationales of academic doctors remains under-researched. It seems important to investigate the views of those with a research-orientation, given the emphasis on the need for more scientific evidence in recent debates on CAM.</p> <p>Methods</p> <p>This exploratory study used qualitative methods to explore academic doctors' views of CAM and the rationales they provided for their views. A purposeful sampling strategy was used to identify doctors with a dual clinical and academic role in the Bristol area, with an anticipated variety of views on CAM. Semi-structured interviews were conducted with nine doctors. The data were analysed thematically, drawing on the Framework Approach.</p> <p>Results</p> <p>The doctors expressed a spectrum of views on CAM, falling into three broad groups: the 'enthusiasts', the 'sceptics' and the 'undecided'. Scepticism or uncertainty about the value of CAM was prominent, except among those practising a form of CAM. A variety of rationales underpinned their perspectives on CAM, a key recurring rationale being their perspective on the scientific evidence base. The main themes arising included: the role of doctors' professional experiences of conventional medicine and CAM in shaping their attitudes towards CAM, doctor-patient communication about CAM and patient disclosure, whether there is a need for training and education in CAM for doctors, a hierarchy of acceptability of CAM and the nature of evidence; and the role of CAM within the NHS.</p> <p>Conclusion</p> <p>Despite the caution or scepticism towards CAM expressed by doctors in this study, more open doctor-patient communication about CAM may enable doctors' potential concerns about CAM to be addressed, or at least enhance their knowledge of what treatments or therapies their patients are using. Offering CAM to patients may serve to enhance patients' treatment choices and even increase doctors' fulfilment in their practice. However, given the recurring concerns about lack of scientific evidence expressed by the doctors in this study, perceptions of the evidence base may remain a significant barrier to greater integration of CAM within the NHS.</p

Stress Propagation through Frictionless Granular Material

We examine the network of forces to be expected in a static assembly of hard, frictionless spherical beads of random sizes, such as a colloidal glass. Such an assembly is minimally connected: the ratio of constraint equations to contact forces approaches unity for a large assembly. However, the bead positions in a finite subregion of the assembly are underdetermined. Thus to maintain equilibrium, half of the exterior contact forces are determined by the other half. We argue that the transmission of force may be regarded as unidirectional, in contrast to the transmission of force in an elastic material. Specializing to sequentially deposited beads, we show that forces on a given buried bead can be uniquely specified in terms of forces involving more recently added beads. We derive equations for the transmission of stress averaged over scales much larger than a single bead. This derivation requires the Ansatz that statistical fluctuations of the forces are independent of fluctuations of the contact geometry. Under this Ansatz, the $d(d+1)/2$-component stress field can be expressed in terms of a d-component vector field. The procedure may be generalized to non-sequential packings. In two dimensions, the stress propagates according to a wave equation, as postulated in recent work elsewhere. We demonstrate similar wave-like propagation in higher dimensions, assuming that the packing geometry has uniaxial symmetry. In macroscopic granular materials we argue that our approach may be useful even though grains have friction and are not packed sequentially.=17Comment: 15 pages, 4 figures, revised vertion for Phys. Rev.

The oceanography and marine ecology of Ningaloo, a world heritage area

The Ningaloo coast of north-western Australia (eastern Indian Ocean) hosts one of the world’s longest and most extensive fringing coral reef systems, along with globally significant abundances of large marine fauna such as whale sharks. These characteristics – which have contributed to its inscription on the World Heritage list – exist because of the unique climatic, geomorphologic and oceanographic conditions. The region is hot and arid, so runoff of water from land is low, facilitating clear water that allows corals to grow close to the shore. The poleward-flowing Leeuwin Current is an important influence, bringing warm water and generally suppressing coastal upwelling. During the austral summer, strong southerly winds generate the equatorward-flowing Ningaloo Current on the inner shelf – this current facilitates sporadic upwelling events that enhance concentrations of nutrients, which in turn enhance pelagic primary productivity that supports the reef’s biota. The coast has experienced several marine heatwaves since 2011 that have caused mortality of corals and probably seagrass, albeit relatively less than elsewhere along the coast. Wind-generated surface waves break over the fringing reef crest, causing cooling currents that tend to dampen warming – although this mechanism seems not to have prevented some areas from experiencing damaging heat, and corals in places that do not receive the wave-generated currents have experienced substantial mortality. Herbivores, from fish to green turtles, are abundant, and in the lagoon, extensive stands of large brown algae provide an important habitat for newly recruited fish. There has been a decline in abundance of some fish. Predictions of future pressures include a weaker but more variable Leeuwin Current and increased human use. The ability of Ningaloo’s ecosystems to withstand growing pressures will depend partly on the rate and magnitude of global warming but also on actions that manage local pressures from increasing human use. These actions will rely on continued science to provide the evidence needed to identify the pressures, the changes they create and the ways that we can mitigate them

The impact of NHS based primary care complementary therapy services on health outcomes and NHS costs: a review of service audits and evaluations

<p>Abstract</p> <p>Background</p> <p>The aim of this study was to review evaluations and audits of primary care complementary therapy services to determine the impact of these services on improving health outcomes and reducing NHS costs. Our intention is to help service users, service providers, clinicians and NHS commissioners make informed decisions about the potential of NHS based complementary therapy services.</p> <p>Methods</p> <p>We searched for published and unpublished studies of NHS based primary care complementary therapy services located in England and Wales from November 2003 to April 2008. We identified the type of information included in each document and extracted comparable data on health outcomes and NHS costs (e.g. prescriptions and GP consultations).</p> <p>Results</p> <p>Twenty-one documents for 14 services met our inclusion criteria. Overall, the quality of the studies was poor, so few conclusions can be made. One controlled and eleven uncontrolled studies using SF36 or MYMOP indicated that primary care complementary therapy services had moderate to strong impact on health status scores. Data on the impact of primary care complementary therapy services on NHS costs were scarcer and inconclusive. One controlled study of a medical osteopathy service found that service users did not decrease their use of NHS resources.</p> <p>Conclusion</p> <p>To improve the quality of evaluations, we urge those evaluating complementary therapy services to use standardised health outcome tools, calculate confidence intervals and collect NHS cost data from GP medical records. Further discussion is needed on ways to standardise the collection and reporting of NHS cost data in primary care complementary therapy services evaluations.</p

Proteomic Analysis of Salmonella enterica Serovar Typhimurium Isolated from RAW 264.7 Macrophages

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Chapter 4 The Oceanography and Marine Ecology of Ningaloo, A World Heritage Area

The Ningaloo coast of north-western Australia (eastern Indian Ocean) hosts one of the world’s longest and most extensive fringing coral reef systems, along with globally-significant abundances of large marine fauna such as whale sharks. These characteristics — which have contributed to its inscription on the World Heritage list — exist because of the unique climatic, geomorphologic and oceanographic conditions. The region is hot and arid, so runoff of water from land is low, facilitating clear water that allows corals to grow close to the shore. The poleward-flowing Leeuwin Current is an important influence, bringing warm water and generally suppressing coastal upwelling. During the austral summer, strong southerly winds generate the equatorward-flowing Ningaloo Current on the inner shelf — this current facilitates sporadic upwelling events that enhance concentrations of nutrients, which in turn enhances pelagic primary productivity that supports the reef’s biota. The coast has experienced several marine heatwaves since 2011 that have caused mortality of corals, and probably seagrass, albeit relatively less than elsewhere along the coast. Wind-generated surface waves break over the fringing reef crest, causing cooling currents that tend to dampen warming — although this mechanism seems not to have prevented some areas from experiencing damaging heat, and corals in places that do not experience the wave-generated currents have experienced substantial mortality. Herbivores, from fish to green turtles, are abundant, and in the lagoon extensive stands of large brown algae provide an important habitat for newly-recruited fish. There has been a decline in abundance of some fish. Predictions of future pressures include a weaker but more variable Leeuwin Current, and increased human use. The ability of Ningaloo’s ecosystems to withstand growing pressures will depend partly on the rate and magnitude of global warming, but also on actions that manage local pressures from increasing human use. These actions will rely on continued science to provide the evidence needed to identify the pressures, the changes they create and the ways that we can mitigate them