Evaluation of fibroin-based scaffolds for ocular tissue reconstruction

The epithelium of the corneolimbus contains stem cells for regenerating the corneal epithelium. Diseases and injuries affecting the limbus can lead to a condition known as limbal stem cell deficiency (LSCD), which results in loss of the corneal epithelium, and subsequent chronic inflammation and scarring of the ocular surface. Advances in the treatment of LSCD have been achieved through use of cultured human limbal epithelial (HLE) grafts to restore epithelial stem cells of the ocular surface. These epithelial grafts are usually produced by the ex vivo expansion of HLE cells on human donor amniotic membrane (AM), but this is not without limitations. Although AM is the most widely accepted substratum for HLE transplantation, donor variation, risk of disease transfer, and rising costs have led to the search for alternative biomaterials to improve the surgical outcome of LSCD. Recent studies have demonstrated that Bombyx mori silk fibroin (hereafter referred to as fibroin) membranes support the growth of primary HLE cells, and thus this thesis aims to explore the possibility of using fibroin as a biomaterial for ocular surface reconstruction. Optimistically, the grafted sheets of cultured epithelium would provide a replenishing source of epithelial progenitor cells for maintaining the corneal epithelium, however, the HLE cells lose their progenitor cell characteristics once removed from their niche. More severe ocular surface injuries, which result in stromal scarring, damage the epithelial stem cell niche, which subsequently leads to poor corneal re-epithelialisation post-grafting. An ideal solution to repairing the corneal limbus would therefore be to grow and transplant HLE cells on a biomaterial that also provides a means for replacing underlying stromal cells required to better simulate the normal stem cell niche. The recent discovery of limbal mesenchymal stromal cells (L-MSC) provides a possibility for stromal repair and regeneration, and therefore, this thesis presents the use of fibroin as a possible biomaterial to support a three dimensional tissue engineered corneolimbus with both an HLE and underlying L-MSC layer. Investigation into optimal scaffold design is necessary, including adequate separation of epithelial and stromal layers, as well as direct cell-cell contact. Firstly, the attachment, morphology and phenotype of HLE cells grown on fibroin were directly compared to that observed on donor AM, the current clinical standard substrate for HLE transplantation. The production, transparency, and permeability of fibroin membranes were also evaluated in this part of the study. Results revealed that fibroin membranes could be routinely produced using a custom-made film casting table and were found to be transparent and permeable. Attachment of HLE cells to fibroin after 4 hours in serum-free medium was similar to that supported by tissue culture plastic but approximately 6-fold less than that observed on AM. While HLE cultured on AM displayed superior stratification, epithelia constructed from HLE on fibroin maintained evidence of corneal phenotype (cytokeratin pair 3/12 expression; CK3/12) and displayed a comparable number and distribution of ÄNp63+ progenitor cells to that seen in cultures grown on AM. These results confirm the suitability of membranes constructed from silk fibroin as a possible substrate for HLE cultivation. One of the most important aspects in corneolimbal tissue engineering is to consider the reconstruction of the limbal stem cell niche to help form the natural limbus in situ. MSC with similar properties to bone marrow derived-MSC (BM-MSC) have recently been grown from the limbus of the human cornea. This thesis evaluated methods for culturing L-MSC and limbal keratocytes using various serum-free media. The phenotype of resulting cultures was examined using photography, flow cytometry for CD34 (keratocyte marker), CD45 (bone marrow-derived cell marker), CD73, CD90, CD105 (collectively MSC markers), CD141 (epithelial/vascular endothelial marker), and CD271 (neuronal marker), immunocytochemistry (alpha-smooth muscle actin; á-sma), differentiation assays (osteogenesis, adipogenesis and chrondrogenesis), and co-culture experiments with HLE cells. While all techniques supported to varying degrees establishment of keratocyte and L-MSC cultures, sustained growth and serial propagation was only achieved in serum-supplemented medium or the MesenCult-XF„¥ culture system (Stem Cell Technologies). Cultures established in MesenCult-XF„¥ grew faster than those grown in serum-supplemented medium and retained a more optimal MSC phenotype. L-MSC cultivated in MesenCult-XFR were also positive for CD141, rarely expressed £\-sma, and displayed multi-potency. L-MSC supported growth of HLE cells, with the largest epithelial islands being observed in the presence of L-MSC established in MesenCult-XF„¥ medium. All HLE cultures supported by L-MSC widely expressed the progenitor cell marker £GNp63, along with the corneal differentiation marker CK3/12. Our findings conclude that MesenCult-XFR is a superior culture system for L-MSC, but further studies are required to explore the significance of CD141 expression in these cells. Following on from the findings of the previous two parts, silk fibroin was tested as a novel dual-layer construct containing both an epithelium and underlying stroma for corneolimbal reconstruction. In this section, the growth and phenotype of HLE cells on non-porous versus porous fibroin membranes was compared. Furthermore, the growth of L-MSC in either serum-supplemented medium or the MesenCult-XFR culture system within fibroin fibrous mats was investigated. Lastly, the co-culture of HLE and L-MSC in serum-supplemented medium on and within fibroin dual-layer constructs was also examined. HLE on porous membranes displayed a flattened and squamous monolayer; in contrast, HLE on non-porous fibroin appeared cuboidal and stratified closer in appearance to a normal corneal epithelium. Both constructs maintained CK3/12 expression and distribution of £GNp63+ progenitor cells. Dual-layer fibroin scaffolds consisting of HLE cells and L-MSC maintained a similar phenotype as on the single layers alone. Overall, the present study proposed to create a three dimensional limbal tissue substitute of HLE cells and L-MSC together, ultimately for safe and beneficial transplantation back into the human eye. The results show that HLE and L-MSC can be cultivated separately and together whilst maintaining a clinically feasible phenotype containing a majority of progenitor cells. In addition, L-MSC were able to be cultivated routinely in the MesenCult-XF® culture system while maintaining a high purity for the MSC characteristic phenotype. However, as a serum-free culture medium was not found to sustain growth of both HLE and L-MSC, the combination scaffold was created in serum-supplemented medium, indicating that further refinement of this cultured limbal scaffold is required. This thesis has also demonstrated a potential novel marker for L-MSC, and has generated knowledge which may impact on the understanding of stromal-epithelial interactions. These results support the feasibility of a dual-layer tissue engineered corneolimbus constructed from silk fibroin, and warrant further studies into the potential benefits it offers to corneolimbal tissue regeneration. Further refinement of this technology should explore the potential benefits of using epithelial-stromal co-cultures with MesenCult-XF® derived L-MSC. Subsequent investigations into the effects of long-term culture on the phenotype and behaviour of the cells in the dual-layer scaffolds are also required. While this project demonstrated the feasibility in vitro for the production of a dual-layer tissue engineered corneolimbus, further studies are required to test the efficacy of the limbal scaffold in vivo. Future in vivo studies are essential to fully understand the integration and degradation of silk fibroin biomaterials in the cornea over time. Subsequent experiments should also investigate the use of both AM and silk fibroin with epithelial and stromal cell co-cultures in an animal model of LSCD. The outcomes of this project have provided a foundation for research into corneolimbal reconstruction using biomaterials and offer a stepping stone for future studies into corneolimbal tissue engineering.

Critical Literacy in the Early Years : Emergence and Sustenance in an Age of Accountability

The work of early childhood educators in facilitating young children’s literacy acquisition has never received more attention than in the new millennium. Media hype about literacy crises, falling standards, teacher quality and government promises of minimum standards for all children have simultaneously increased the ‘visibility’ of literacy and the stakes for school performance. Indeed the last two decades could be seen as an age of pronouncements with respect to literacy, with politicians internationally promising to cure supposed low literacy with standardized tests and mandated programmes. As the rhetoric around literacy intensifies many late-capitalist economies are experiencing shifts that have increased the gaps between rich and poor, changed the very nature of work, and fundamentally altered the cultural mix of their populations. More and more children attending schools where English is the language of instruction speak it as a second or third language. Many children have experienced the effects of war, terrorism, migration and poverty. Many live in fractured, fragmented and changing families. Teacher populations are changing too. In some places aging teacher workforces mean that there is already a shortage of qualified teachers. Literacy is also changing as the impact of digital technologies on global and local communication, economies and knowledges begins to bite in everyday and working lives. It is challenging to think about how spaces for the emergence and sustenance of critical literacy in early childhood education might be created.

Post-disaster reconstruction: a statistical overview of the barriers for community participation in Afghanistan

When a community already torn by an event such as a prolonged war, is then hit by a natural disaster, the negative impact of this subsequent disaster in the longer term can be extremely devastating. Natural disasters further damage already destabilised and demoralised communities, making it much harder for them to be resilient and recover. Communities often face enormous challenges during the immediate recovery and the subsequent long term reconstruction periods, mainly due to the lack of a viable community involvement process. In post-war settings, affected communities, including those internally displaced, are often conceived as being completely disabled and are hardly ever consulted when reconstruction projects are being instigated. This lack of community involvement often leads to poor project planning, decreased community support, and an unsustainable completed project. The impact of war, coupled with the tensions created by the uninhabitable and poor housing provision, often hinders the affected residents from integrating permanently into their home communities. This paper outlines a number of fundamental factors that act as barriers to community participation related to natural disasters in post-war settings. The paper is based on a statistical analysis of, and findings from, a questionnaire survey administered in early 2012 in Afghanistan.

It's just an old apron : aprons, artefacts and art - representations of personal and private 'her-stories', as public histories

This paper examines art and artefact in the representation and recollection of deeply personal WWII women’s experiences as POW’s under the Japanese. This kind of treatment of internees in the Tjideng Women and Children’s internment camp (and others) in Batavia under the Japanese in WWII, stands in stark and brutal contrast to the idyllic life lived by many families up to that time in what was then known as the Dutch East Indies (Indonesia). The deprivation and brutality of the Japanese incarceration of these women and children evoked responses - not military, but certainly militant, if muted. Representations of those responses – as both art and artefact - may be found in the most unlikely places and unexpected forms - and are still being unearthed to this day. However close we might personally be to these artists and artisans, can we, as observers from a distance, ever truly comprehend through spoken or written words alone, the day-today realities of those extraordinary times?

Posttraumatic stress disorder and traumatic stress : from bench to bedside, from war to disaster

War is a tragic event and its mental health consequences can be profound. Recent studies indicate substantial rates of posttraumatic stress disorder and other behavioral alterations because of war exposure. Understanding the psychological, behavioral, and neurobiological mechanism of mental health and behavioral changes related to war exposure is critical to helping those in need of care. Substantial work to encourage bench to bedside to community knowledge and communication is a core component of addressing this world health need.

Design of a traumatic injury simulator for assessing lower limb response to high loading rates

Current military conflicts are characterized by the use of the improvised explosive device. Improvements in personal protection, medical care, and evacuation logistics have resulted in increasing numbers of casualties surviving with complex musculoskeletal injuries, often leading to lifelong disability. Thus, there exists an urgent requirement to investigate the mechanism of extremity injury caused by these devices in order to develop mitigation strategies. In addition, the wounds of war are no longer restricted to the battlefield; similar injuries can be witnessed in civilian centers following a terrorist attack. Key to understanding such mechanisms of injury is the ability to deconstruct the complexities of an explosive event into a controlled, laboratory-based environment. In this article, a traumatic injury simulator, designed to recreate in the laboratory the impulse that is transferred to the lower extremity from an anti-vehicle explosion, is presented and characterized experimentally and numerically. Tests with instrumented cadaveric limbs were then conducted to assess the simulator’s ability to interact with the human in two mounting conditions, simulating typical seated and standing vehicle passengers. This experimental device will now allow us to (a) gain comprehensive understanding of the load-transfer mechanisms through the lower limb, (b) characterize the dissipating capacity of mitigation technologies, and (c) assess the bio-fidelity of surrogates.

Global Politics of Health

International responses to the outbreak of SARS, the spread of HIV/AIDS, and the promotion of health as a human right all demonstrate how global politics have a profound effect on the way we think about and respond to major health challenges. Despite a growing interest in the relationship between health and international relations there has yet to be a systematic study of the links between them. Global Health Issues aims to fill this gap – ultimately showing how world politics can be good, or bad, for your health. This book calls for a more nuanced understanding of the nature of the current global health crisis and the political dilemmas faced by those responsible for the development and implementation of responses to it. By charting these debates and showing how they shape the way actors think about key issues relating to health, such as people movement; infectious disease; the business of health; and the consequences of war; this volume provides an innovative and comprehensive introduction to health and international relations for students of global politics, health studies and related disciplines.

Heat strain evaluation of overt and covert body armour in a hot and humid environment

The aim of this study was to elucidate the thermophysiological effects of wearing lightweight non-military overt and covert personal body armour (PBA) in a hot and humid environment. Eight healthy males walked on a treadmill for 120 min at 22% of their heart rate reserve in a climate chamber simulating 31 °C (60%RH) wearing either no armour (control), overt or covert PBA in addition to a security guard uniform, in a randomised controlled crossover design. No significant difference between conditions at the end of each trial was observed in core temperature, heart rate or skin temperature (P > 0.05). Covert PBA produced a significantly greater amount of body mass change (−1.81 ± 0.44%) compared to control (−1.07 ± 0.38%, P = 0.009) and overt conditions (−1.27 ± 0.44%, P = 0.025). Although a greater change in body mass was observed after the covert PBA trial; based on the physiological outcome measures recorded, the heat strain encountered while wearing lightweight, non-military overt or covert PBA was negligible compared to no PBA. Practitioner summary The wearing of bullet proof vests or body armour is a requirement of personnel engaged in a wide range of occupations including police, security, customs and even journalists in theatres of war. This randomised controlled crossover study is the first to examine the thermophysiological effects of wearing lightweight non-military overt and covert personal body armour (PBA) in a hot and humid environment. We conclude that the heat strain encountered while wearing both overt and covert lightweight, non-military PBA was negligible compared to no PBA.

The Soldier's Wife (LP)

Audio recording of 10 tracks funded by Legacy, The Australia Council, and Arts Queensland. The recordings explore the untold stories of soldiers' wives through song. It features the vocal and songwriting talents of Jackie Marshall, Bertie Page, Sahara Beck, Emma Bosworth, Roz Pappalardo, and Kristy Apps. Recorded, Mixed, Mastered, and Co-Produced by Phil Graham.

Alliances in the shadow of conflict

Victorious alliances often fight about the spoils of war. This article presents an experiment on the determinants of whether alliances break up and fight internally after having defeated a joint enemy. First, if peaceful sharing yields an asymmetric rent distribution, this increases the likelihood of fighting. In turn, anticipation of the higher likelihood of internal fight reduces the alliance’s ability to succeed against the outside enemy. Second, the option to make nonbinding nonaggression declarations between alliance members does not make peaceful settlement within the alliance more likely. Third, higher differences in the alliance players’ contributions to alliance effort lead to more internal conflict and more intense fighting.

Insomnia : A Self Help Handbook

This book is a practical and useful tool for getting your sleep back on track. Even if you have suffered from insomnia for many years, this book contains simple, easy to learn strategies to manage your sleep loss through evidence-based techniques such as cognitive therapy and stimulus control. Dr. Sacre will guide you through these approaches and explain how they work and why they are recommended above other approaches. There is a chapter on special populations that tells you what to do if you are a shift worker, long distance traveller, parent, older adult, woman (including pregnancy and menopause) or an elite athlete. If you want to enjoy natural, healthy and satisfying sleep again, this handbook gives you all the tools you need to achieve it. You only need to have the motivation and discipline to apply the strategies and stick to them over time. This handbook first explains what normal sleep is all about and challenges some myths about sleep and insomnia. Then you will be guided through a thorough sleep assessment. Insomnia is then described in detail including different types of insomnia and the kinds of factors that contribute to sleep loss. Through the following chapters, you will be shown step-by-step what to do to bring about change in your sleeping patterns and habits, through addressing the factors that perpetuate poor sleep. These factors mainly revolve around unhelpful thinking, compensatory behaviors, poor sleep hygiene and environmental influences. These are all things that are within your control and Dr. Sacre will show you how. Dr. Sacre has worked in the fields of sleep health, mental health and addictive disorders for 25 years and over that time, she has encountered hundreds of people who have struggled with insomnia and sleep loss due to other causes. She currently heads the Therapy Programs department at Belmont Private Hospital in Brisbane, Australia, where there is an emphasis on Cognitive Behavioral Therapy, including a Cognitive Behavioral Therapy for Insomnia (CBT-i) program. A psychologist and nurse, Dr. Sacre is a long-time member of the Australasian Sleep Association and the Australian Psychological Society. She has conducted research into the function of dreaming, online sleep surveys and the usefulness of sleep self-help guides for students, older adults and carers of people with dementia. She has also published on diverse topics, including the management of nightmares in war veterans. She is an Adjunct Associate Professor at Queensland University of Technology, Brisbane and lectures professionals, including psychologists, school counselors and psychiatrists, on sleep disorders and their management as well as Cognitive Behavioral Therapy.

Soluble mediators in platelet concentrates modulate dendritic cell inflammatory responses in an experimental model of transfusion

The transfusion of platelet concentrates (PCs) is widely used to treat thrombocytopenia and severe trauma. Ex vivo storage of PCs is associated with a storage lesion characterized by partial platelet activation and the release of soluble mediators, such as soluble CD40 ligand (sCD40L), RANTES, and interleukin (IL)-8. An in vitro whole blood culture transfusion model was employed to assess whether mediators present in PC supernatants (PC-SNs) modulated dendritic cell (DC)-specific inflammatory responses (intracellular staining) and the overall inflammatory response (cytometric bead array). Lipopolysaccharide (LPS) was included in parallel cultures to model the impact of PC-SNs on cell responses following toll-like receptor-mediated pathogen recognition. The impact of both the PC dose (10%, 25%) and ex vivo storage period was investigated [day 2 (D2), day 5 (D5), day 7 (D7)]. PC-SNs alone had minimal impact on DC-specific inflammatory responses and the overall inflammatory response. However, in the presence of LPS, exposure to PC-SNs resulted in a significant dose associated suppression of the production of DC IL-12, IL-6, IL-1a, tumor necrosis factor-a (TNF-a), and macrophage inflammatory protein (MIP)-1b and storage-associated suppression of the production of DC IL-10, TNF-a, and IL-8. For the overall inflammatory response, IL-6, TNF-a, MIP-1a, MIP-1b, and inflammatory protein (IP)-10 were significantly suppressed and IL-8, IL-10, and IL-1b significantly increased following exposure to PC-SNs in the presence of LPS. These data suggest that soluble mediators present in PCs significantly suppress DC function and modulate the overall inflammatory response, particularly in the presence of an infectious stimulus. Given the central role of DCs in the initiation and regulation of the immune response, these results suggest that modulation of the DC inflammatory profile is a probable mechanism contributing to transfusion-related complications.

Expulsion of Afghan refugees from Pakistan: A violation of principle of non-refoulement

This article addresses the need of an implementation mechanism for the protection of refugees’ rights. However, it is contended that the principle forms part of Customary International Law, under which it is binding on all states irrespective of whether or not they are parties to the Convention Relating to the Status of Refugees 1951 or its Protocol 1967. Since last decade, U.S and its allies have been fighting to curve terrorism which has raised many issues such as human rights violation, deportation, expulsion, extradition, rendition and many more. Pakistan has played a very critical role in War against Terrorism, particularly in reference of war in Afghanistan. Particular concern of this article is the violation of refugees’ rights in Pakistan in 2008 and 2010. This article would highlight the legislation regarding non-expulsion of Afghan refugees from Pakistan to a territory where they have well founded fear of persecution. Article is divided into three parts, the first one deals with “Principle of Non-Refoulement”, the second one deals with “exceptions to the principle” whereas the last one discusses the violation of the very principle in Pakistan with reference to Afghan refugees.

A subculture of mental toughness in an Australian Football League club

Objective The current study aimed to provide a subcultural analysis of mental toughness in a high-performance context in sport. Design Using Schein's (1990) framework of organisational culture, an exploratory qualitative analysis, employing focus group and individual interviews, was used to investigate mental toughness in an Australian Football League club. Method Nine senior coaches and players participated in focus group and individual interviews. Photo elicitation was used as a method to capture mental toughness through the identification of prominent club artefacts. Participants were considered to have significant subcultural knowledge of their football club and were willing to describe personal experiences and perceptions of mental toughness through this cultural lens. Deductive and inductive analyses were conducted to capture the core themes of mental toughness across the disparate levels of Schein's organisational framework. Results Mental toughness was found to be a socially derived term marked by unrelenting standards and sacrificial displays. These acts were underpinned by subcultural values emphasising a desire for constant improvement, a team first ethos, relentless effort, and the maintenance of an infallible image. At its core, mental toughness was assumed to be an internal concept, epitomised an idealised form of masculinity, elitist values, and was rhetorically depicted through metaphors of war. Conclusions It may be difficult to understand mental toughness without giving attention to the contextual norms related to the term. Appreciating how people promote, instil, and internalise prized ideals coveted as mental toughness could be intriguing for future research in sport psychology.

Phytogeny of the Quambalariaceae fam. nov., including important Eucalyptus pathogens in South Africa and Australia

The genus Quambalaria consists of plant-pathogenic fungi causing disease on leaves and shoots of species of Eucalyptus and its close relative, Corymbia. The phylogenetic relationship of Quambalaria spp., previously classified in genera such as Sporothrix and Ramularia, has never been addressed. It has, however, been suggested that they belong to the basidiomycete orders Exobasidiales or Ustilaginales. The aim of this study was thus to consider the ordinal relationships of Q. eucalypti and Q. pitereka using ribosomal LSU sequences. Sequence data from the ITS nrDNA were used to determine the phylogenetic relationship of the two Quambalaria species together with Fugomyces (= Cerinosterus) cyanescens. In addition to sequence data, the ultrastructure of the septal pores of the species in question was compared. From the LSU sequence data it was concluded that Quambalaria spp. and F. cyanescens form a monophyletic clade in the Microstromatales, an order of the Ustilaginomycetes. Sequences from the ITS region confirmed that Q. pitereka and Q. eucalypti are distinct species. The ex-type isolate of F. cyanescens, together with another isolate from Eucalyptus in Australia, constitute a third species of Quambalaria, Q. cyanescens (de Hoog & G.A. de Vries) Z.W. de Beer, Begerow & R. Bauer comb. nov. Transmission electron-microscopic studies of the septal pores confirm that all three Quambalaria spp. have dolipores with swollen lips, which differ from other members of the Microstromatales (i.e. the Microstromataceae and Volvocisporiaceae) that have simple pores with more or less rounded pore lips. Based on their unique ultrastructural features and the monophyly of the three Quambalaria spp. in the Microstromatales, a new family, Quambalariaceae Z.W. de Beer, Begerow & R. Bauer fam. nov., is described.