The role of PAR2 in the myeloid compartment and the regulation of osteoclastogenesis

Protease activated receptor 2 (PAR2) is one member of a family of G-protein coupled receptors. A defining feature of this family of receptors is their protease mediated activation. An emerging role for PAR2 in both the immune system and also in bone biology suggests a potential function for this receptor in osteoimmunology and its associated pathologies. Inflammatory arthritic conditions such as rheumatoid arthritis (RA), are pathologies that include both inflammation and bone destruction, making them a prime model for osteoimmunological studies. Previous murine studies identified that loss of PAR2 results in protection from inflammatory adjuvant-induced arthritis. These animals experienced an attenuated form of arthritis, with significantly reduced joint inflammation and damage. In addition, PAR2 is known to be upregulated in both macrophages (from tissue biopsies) and peripheral blood monocytes in patients with RA. However, the functional impact of protease signalling via PAR2 in monocytes in RA and how this receptor influences joint destruction via monocytes and their tissue differentials is still unknown. The central aim of this doctoral study was to understand the function of PAR2 in monocytes and how protease signalling via PAR2 would influence the differentiation potential of these cells. Specific focus was placed on osteoclasts (OCs), and how protease signalling may further contribute to bone erosion through the action of PAR2 on these bone resorbing cells. Initial work confirmed the expression of the PAR2 on the plasma membrane of monocytes, with the highest expression consistently found on classical monocytes that have the highest osteoclastogenic potential. In order to study the impact of PAR2 signalling on OC formation and activity, in vitro OC differentiation assays were set up using both WT and par2-/- cells. Both standard homeostatic and inflammatory TNF enhanced OC assays were utilised. These studies revealed that during homeostatic OC formation PAR2 contributed to the regulation of OC formation and prevented excessive fusion of precursors (OCPs) into giant cells. The PAR2 mediated regulation of OC formation was found to be important both via OCPs directly, and via the stromal compartment. An absence of PAR2 in osteoblast (OB) -like cells resulted in a more osteoclastogenic stroma and contributed to enhanced OC formation. However, during inflammatory3 driven OC formation, the role of PAR2 in osteoclastogenesis was reversed, and protease signals via PAR2 enhanced the formation of OCs. The observations made in murine in vitro systems were replicated in human cell cultures when monocytes were exposed to PAR2 inhibitors during the process of OC differentiation. Combined, this work indicates that PAR2 has an impact on the process of osteoclastogenesis. Whether PAR2 signalling regulates or enhances OC formation is dependent upon the environment and the combination of signals received by precursor cells

Exploring the microbiome: diversity of the microbial community of three foam nesting frogs, Genus: Polypedates, across a developmental gradient

Characterization of microbial biodiversity, including that of the amphibian skin-associated microbiome, is a frontier of research recently made accessible through advances in sequencing technology. Microbial interaction with a host has been determined to have profound influences on host health across a wide range of macroscopic organisms. For amphibians, the influence of the skin-associated microbiome has been found to have particular importance, as amphibians are currently one of the fastest disappearing vertebrate groups on the planet, largely in part to skin-associated diseases caused by pathogenic microbes. Therefore, it is important to characterize the amphibian skin-associated microbiome, particularly for species with no existing microbiome data, and to delineate relationships that may influence host health. In determining the microbial community of amphibian skin, it is important to outline baseline native microbial presence and gain insight into how these microbes become established. This study focused on being the first to characterize the cutaneous microbial diversity of three Southeast Asian tree frogs in the family Rhacophoridae (genus: Polypedates) that reproduce via the specialized breeding strategy of building a foam nest and comparing the amphibian microbiome across initial development to that of the environment. Microbes associated with reproducing adults, foam nests, tadpoles before and after environmental interaction, and the surrounding environment were characterized using 16S amplicon sequencing. The phylum Proteobacteria comprised the majority of communities across amphibian and environmental samples at 57% relative abundance with Firmicutes (16%) and Bacteroidetes (13%) as the next most dominant phyla. In comparing amphibian and environmental samples, no amphibian microbial communities mirrored that of their immediate environment. Interestingly, tadpole skin-associated microbes differed in relative abundance and microbial taxa between nest-extracted tadpoles and those that were sampled after interaction with a pond environment. This demonstrates the necessity of further research into microbial community establishment, host selection processes, and microbial transmission. Gaining baseline knowledge of the skin-associated microbiome contributes to our knowledge of the natural world and preliminary delineation of ecological relationships between host, microbe, and environment provides an example of the need for continued research in this area which has the potential to broadly inform conservation efforts for amphibians worldwide

Medical school clinical placements – the optimal method for assessing the clinical educational environment from a graduate entry perspective

Background: Educational environment is a strong determinant of student satisfaction and achievement. The learning environments of medical students on clinical placements are busy workplaces, composed of many variables. There is no universally accepted method of evaluating the clinical learning environment, nor is there consensus on what concepts or aspects should be measured. The aims of this study were to compare the Dundee ready educational environment measure (DREEM - the current de facto standard) and the more recently developed Manchester clinical placement index (MCPI) for the assessment of the clinical learning environment in a graduate entry medical student cohort by correlating the scores of each and analysing free text comments. This study also explored student perceptionof how the clinical educational environment is assessed. Methods: An online, anonymous survey comprising of both the DREEM and MCPI instruments was delivered to students on clinical placement in a graduate entry medical school. Additional questions explored students’ perceptions of instruments for giving feedback. Numeric variables (DREEM score, MCPI score, ratings) were tested for normality and summarised. Pearson’s correlation coefficient was used to measure the strength of the association between total DREEM score and total MCPI scores. Thematic analysis was used to analyse the free text comments. Results: The overall response rate to the questionnaire was 67% (n = 180), with a completed response rate for the MCPI of 60% (n = 161) and for the DREEM of 58% (n = 154). There was a strong, positive correlation between total DREEM and MCPI scores (r = 0.71, p < 0.001). On a scale of 0 to 7, the mean rating for how worthwhile students found completing the DREEM was 3.27 (SD 1.41) and for the MCPI was 3.49 (SD 1.57). ‘Finding balance’ and ‘learning at work’ were among the themes to emerge from analysis of free text comments. Conclusions: The present study confirms that DREEM and MCPI total scores are strongly correlated. Graduate entry students tended to favour this method of evaluation over the DREEM with the MCPI prompting rich description of the clinical learning environment. Further study is warranted to determine if this finding is transferable to all clinical medical student cohorts. Keywords: Clinical education, DREEM, Manchester clinical placement inde

Causation in metaphysics and moral theory

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Linguistics and Philosophy, 2002.Includes bibliographical references.Chapter 1: The causal relata. Ordinary talk suggests that entities from different ontological categories can cause and be caused: Kathy's throw, the fact that Kathy threw, and Kathy herself can all cause the window to break. But according to the majority view, causation exclusively relates events. This chapter defends the contrary view that the causal relata are as miscellaneous as ordinary talk suggests. A question remains: is there an ontological kind K such that causal relations on entities of that kind are somehow more fundamental than causal relations on the non-Ks? I argue that there is such a kind: facts. I defend this claim against objections. Chapter 2. Causation by omission. Ordinary talk also suggests that omissions can be causes. For example, if Barry promised to water Alice's plant, didn't water it, and the plant then dried up and died, then Barry's not watering the plant-his omitting to water the plant-is a cause of its death. But there are reasons to think that either there is no causation by omission, or there is far more of it than common sense allows. I argue that neither disjunct is acceptable, and propose that we avoid the dilemma by embracing the view that causation has a normative component. The proposal faces the objection that causation is a paradigmatic example of a natural, and so entirelynon-normative, relation.(cont.) I argue that the objection can be defused once we are clear about the kind of normativity that plays a role in causation by omission. Chapter 3. Causation and the Making/Allowing Distinction. Common sense morality suggests that it can matter morally whether an agent makes an outcome occur or merely allows it to occur. For example, it is far worse to pinch your little brother than to allow him to be pinched. I argue against the assumption that the making/allowing distinction is exclusive: in fact, the categories of making and allowing overlap. I go on to offer a positive account of makings, and a positive account of allowings.by Sarah McGrath.Ph.D

Multimessenger Approaches to Supermassive Black Hole Binary Detection and Parameter Estimation II: Optimal Strategies for a Pulsar Timing Array

Pulsar timing arrays (PTAs) are Galactic-scale gravitational wave (GW) detectors consisting of precisely-timed pulsars distributed across the sky. Within the decade, PTAs are expected to detect the nanohertz GWs emitted by close-separation supermassive black hole binaries (SMBHBs), thereby opening up the low frequency end of the GW spectrum for science. Individual SMBHBs which power active galactic nuclei are also promising multi-messenger sources; they may be identified via theoretically predicted electromagnetic (EM) signatures and be followed up by PTAs for GW observations. In this work, we study the detection and parameter estimation prospects of a PTA which targets EM-selected SMBHBs. Adopting a simulated Galactic millisecond pulsar population, we envisage three different pulsar timing campaigns which observe three mock sources at different sky locations. We find that an all-sky PTA which times the best pulsars is an optimal and feasible approach to observe EM-selected SMBHBs and measure their source parameters to high precision (i.e., comparable to or better than conventional EM measurements). We discuss the implications of our findings in the context of the future PTA experiment with the planned Deep Synoptic Array-2000 and the multi-messenger studies of SMBHBs such as the well-known binary candidate OJ 287.Comment: 14 pages, 6 figures, 3 tables; ApJ accepted; data will be available with the ApJ publicatio