Development of an organotypic 3D in vitro model of normal human breast tissue: a tool for cancer initiation studies

The mechanisms involved in breast cancer initiation are not well understood. This may in part be due to a lack of an in vitro model that faithfully recapitulates the morphology, phenotype and in vivo architecture of the normal human mammary gland. Most in vitro models of normal breast have relied on the use of reconstituted basement membrane gels to induce luminal epithelial cell polarity and have neglected the role of myoepithelial cells and fibroblasts in this process. The aim of this thesis was to develop a three dimensional in vitro culture system of normal breast which included three of the major functional cell types of breast embedded in a more physiologically relevant collagen I matrix. It was then sought to use the system to investigate the mechanisms behind breast cancer initiation via genetic manipulation of well-known oncogenes and tumour suppressors involved in breast cancer progression. To achieve this, myoepithelial cells (Myo1089, originally isolated from breast reduction mammoplasty sample, gift of Dr Mike O’Hare) and fibroblasts (isolated and immortalised from breast reduction mammoplasty samples collected with ethical approval in house) were characterised by immunofluorescence to assess their suitability. Following characterisation, these were virally transfected with Turbo Green Fluorescent Protein (tGFP) and dsRed protein respectively to enable tracking. Three-dimensional tri-cultures were established in collagen I and included the non-tumorigenic luminal epithelial cell line HB2 with GFP Myo1089 cells and dsRed fibroblasts. Cells were cultured for three weeks in Transwell™ cell culture inserts. Following fixation these were analysed by haematoxylin and eosin staining, confocal microscopy and immunohistochemistry. Morphology and immunostaining profiles were compared to sections of a normal human in vivo breast tissue specimen. Immunohistochemical characterisation using the following antibodies: E-cadherin, epithelial membrane antigen, vimentin, laminin 5, collagen IV plus luminal and basal cytokeratins, demonstrated polarised epithelial structures with lumen formation and basement membrane production with a similar immunostaining profile to normal breast tissue. The importance of including myoepithelial cells and fibroblasts in maintaining these structures was demonstrated. We established this model was amenable to genetic engineering by overexpressing HER2 and HER3 in HB2 cells, and knocking out ERβ1 in Myo1089 cells and DOCK4 in fibroblast cell lines using siRNA/shRNA techniques respectively. These were included in separate models with morphological and phenotypic effects determined by haematoxylin and eosin staining, immunohistochemistry and quantification of HB2 structures formed. We further investigated the intracellular signalling cascades stimulated by heregulin in order to validate our findings upon overexpression of HER2 and HER3 and to investigate the cancer initiation potential of heregulin in the breast. In summary, an in vitro model of normal breast tissue that includes three of the major functional breast cell types cultured in a physiologically relevant three dimensional matrix has been developed. The morphology and protein expression profile of the model was validated against a human breast tissue specimen and confirmed that it is a suitable model of normal breast. The model proved to be reproducible, suitable for experimentation using genetic engineering and cell behaviour could be easily visualised using standard laboratory techniques. To conclude, this is a robust in vitro model of normal breast tissue offering an alternative cost-effective method of studying genes and processes involved in breast cancer initiation

The Mediational Effect of Self-Regulatory Capacity on the Relationship Between Temperament, Childhood Invalidation and Interpersonal Functioning: Testing a New Neuro-Regulatory Model.

Based on existing theories of personality and socio-emotional functioning (e.g. Clark, 2005; Lynch, Hempel & Clark, in press) a new model is proposed and tested. The model hypothesises that (i) temperament (reward and threat sensitivity) and childhood invalidation predict problems with interpersonal functioning, (ii) this effect is mediated by self-regulatory capacity; where self-regulatory capacity comprises self-control (ranging from emotional over-control to emotional under-control) and flexible control and (iii) self-regulatory capacity itself has a quadratic relationship with interpersonal functioning. A UK community sample (n= 512) completed a self-report survey, measuring each of the aforementioned latent variables. Structural equation modelling (SEM) was used to determine the goodness-of-fit of this and variations of this model. SEM identified that a non-mediation model provided the best fit (χ²=49.403, p< 0.001; CFI=0.98; RMSEA=0.056). Good-fit was obtained for a model including flexible control as a partial mediator (χ²=269.06, p< 0.001; CFI=0.956; RMSEA=0.081) and adequate-fit for a model including over-control as a partial mediator (χ²= 91.744, p < 0.001, CFI=0.932; RMSEA= 0.096). Correlation analyses suggested that over-control and under-control correlated positively with interpersonal problems. Results from SEM provided promising initial evidence for the mediating role of self-regulatory capacity, particularly for the flexible control component. Correlation analyses provided support for the non-linear relationship between self-regulatory capacity and interpersonal functioning, whereby extreme over-control or extreme under-control is associated with interpersonal problems. Findings have implications for identifying mechanisms of change for therapeutic approaches to emotion dysregulation and for understanding the over-controlled population, which has previously been overlooked

Tidally Driven Processes Leading to Near-Field Turbulence in a Channel at the Crest of the Mendocino Escarpment*

In situ observations of tidally driven turbulence were obtained in a small channel that transects the crest of the Mendocino Ridge, a site of mixed (diurnal and semidiurnal) tides. Diurnal tides are subinertial at this latitude, and once per day a trapped tide leads to large flows through the channel giving rise to tidal excursion lengths comparable to the width of the ridge crest. During these times, energetic turbulence is observed in the channel, with overturns spanning almost half of the full water depth. A high-resolution, nonhydrostatic, 2.5-dimensional simulation is used to interpret the observations in terms of the advection of a breaking tidal lee wave that extends from the ridge crest to the surface and the subsequent development of a hydraulic jump on the flanks of the ridge. Modeled dissipation rates show that turbulence is strongest on the flanks of the ridge and that local dissipation accounts for 28% of the energy converted from the barotropic tide into baroclinic motion.United States. Office of Naval Research (Grant ONR N00014- 12-1-0943)National Science Foundation (U.S.). (Grant NSF OCE 1129763

Winters are changing: snow effects on Arctic and alpine tundra ecosystems

Snow is an important driver of ecosystem processes in cold biomes. Snow accumulation determines ground temperature, light conditions, and moisture availability during winter. It also affects the growing season&rsquo;s start and end, and plant access to moisture and nutrients. Here, we review the current knowledge of the snow cover&rsquo;s role for vegetation, plant-animal interactions, permafrost conditions, microbial processes, and biogeochemical cycling. We also compare studies of natural snow gradients with snow experimental manipulation studies to assess time scale difference of these approaches. The number of tundra snow studies has increased considerably in recent years, yet we still lack a comprehensive overview of how altered snow conditions will affect these ecosystems. Specifically, we found a mismatch in the timing of snowmelt when comparing studies of natural snow gradients with snow manipulations. We found that snowmelt timing achieved by snow addition and snow removal manipulations (average 7.9 days advance and 5.5 days delay, respectively) were substantially lower than the temporal variation over natural spatial gradients within a given year (mean range 56 days) or among years (mean range 32 days). Differences between snow study approaches need to be accounted for when projecting snow dynamics and their impact on ecosystems in future climates. &nbsp;</p

School leaders reflections on their school's engagement in a program to foster health literacy development

Many health attitudes and behaviors formed during childhood are sustained through to adulthood, thereforechildhood is a critical time to develop health literacy. Primary schools provide an ideal environment to equipchildren with lifelong health skills, understanding and knowledge. Through semi-structured interviews, this studygathered primary school leaders’ reflections on the implementation of a program (HealthLit4Kids) designed tofoster health literacy development in their schools. The aim of this study was to determine how school leadersexperienced the HealthLit4Kids intervention. The results showed that leaders perceived the program had a positive effect on health literacy knowledge and understanding within the school community, as well as improvedhealth behaviors. School leaders’ statements indicated that key barriers such as parental engagement and anovercrowded curriculum would need to be navigated to ensure successful program sustainability

Independent Publishing: Making and Preserving Culture in a Global Literary Marketplace

First paragraph: This report results from a Programme of Enquiry funded and hosted by the Scottish Insight Universities Institute (Scottish Insight), on the theme of Independent Publishing: Making and Preserving Culture in a Global Literary Marketplace. A series of events was held from June-August 2011 in Scottish Insight's premises in Glasgow, with an additional event held at the Edinburgh International Book Festival in association with Publishing Scotland in August 2011. The events brought together publishers, authors, policy makers, government, librarians, academics from multidisciplinary backgrounds, publishing students, and others with an involvement in books and publishing from Scotland, the UK and beyond. The Programme was supplemented by a series of interviews with independent publishers

Building the capacity of rural allied health generalists through online postgraduate education: a qualitative evaluation

Introduction: Workforce development is a key strategy for building the capacity and capability of a workforce. Accordingly, rural and remote practising allied health professionals require relevant and accessible continuing professional development to enhance their knowledge and skills and improve consumer health outcomes. This study explored the impact of an online postgraduate allied health rural generalist education program, from the perspective of allied health professionals participating in the program and their supervisors and managers. Methods: A qualitative, exploratory descriptive study design was employed using semistructured interviews. This study formed the qualitative component of a larger convergent mixed-methods evaluation study aimed at evaluating the reach, quality and impact of an online rural generalist education program for allied health professionals in Australia. Allied health professionals from seven professions enrolled in an online postgraduate rural generalist education program, the rural generalist program (RGP). Their designated work-based supervisors and their managers who were responsible for the operational management of the study sites were invited to participate in the study. All participants were employed in rural and remote health services in 10 sites across four Australian states. Study participants’ experience and perceptions of the impact of the RGP on themselves, the healthcare service and the broader community were explored using semistructured interviews. Data were thematically analysed site by site, then across sites using Braun and Clarke’s (2012) systematic six-phase approach. Provisional codes were generated and iteratively compared, contrasted and collapsed into secondary, more advanced codes until final themes and subthemes were developed. Results: Semistructured interviews were conducted with 23 allied health professionals enrolled in the RGP and their 27 work-based supervisors and managers across the 10 study sites. Three final themes were identified that describe the impact of the RGP: building capability as rural generalist allied health professionals; recruiting and building a rural workforce; enhancing healthcare services and consumer outcomes. Conclusion: Allied health professionals working in rural and remote locations valued the RGP because it provided accessible postgraduate education that aligned with their professional and clinical needs. Integrated into a supportive, well-structured development pathway, the experience potentiated learning and facilitated safe clinical practice that met the needs of consumers and organisations. The findings demonstrate that effective work-integrated learning strategies can enhance the development of essential capabilities for rural practice and support early-career allied health professionals’ transition to rural and remote practice. These experiences can engage allied health professionals in a way that engenders a desire to remain working in rural and remote contexts

Exploring the origins of the power-law properties of energy landscapes: An egg-box model

Multidimensional potential energy landscapes (PELs) have a Gaussian distribution for the energies of the minima, but at the same time the distribution of the hyperareas for the basins of attraction surrounding the minima follows a power-law. To explore how both these features can simultaneously be true, we introduce an ``egg-box'' model. In these model landscapes, the Gaussian energy distribution is used as a starting point and we examine whether a power-law basin area distribution can arise as a natural consequence through the swallowing up of higher-energy minima by larger low-energy basins when the variance of this Gaussian is increased sufficiently. Although the basin area distribution is substantially broadened by this process,it is insufficient to generate power-laws, highlighting the role played by the inhomogeneous distribution of basins in configuration space for actual PELs.Comment: 7 pages, 8 figure