In vivo dose measurement using TLDs and MOSFET dosimeters for cardiac radiosurgery.

In vivo measurements were made of the dose delivered to animal models in an effort to develop a method for treating cardiac arrhythmia using radiation. This treatment would replace RF energy (currently used to create cardiac scar) with ionizing radiation. In the current study, the pulmonary vein ostia of animal models were irradiated with 6 MV X-rays in order to produce a scar that would block aberrant signals characteristic of atrial fibrillation. The CyberKnife radiosurgery system was used to deliver planned treatments of 20-35 Gy in a single fraction to four animals. The Synchrony system was used to track respiratory motion of the heart, while the contractile motion of the heart was untracked. The dose was measured on the epicardial surface near the right pulmonary vein and on the esophagus using surgically implanted TLD dosimeters, or in the coronary sinus using a MOSFET dosimeter placed using a catheter. The doses measured on the epicardium with TLDs averaged 5% less than predicted for those locations, while doses measured in the coronary sinus with the MOSFET sensor nearest the target averaged 6% less than the predicted dose. The measurements on the esophagus averaged 25% less than predicted. These results provide an indication of the accuracy with which the treatment planning methods accounted for the motion of the target, with its respiratory and cardiac components. This is the first report on the accuracy of CyberKnife dose delivery to cardiac targets

BMP9 is a potent inducer of chondrogenesis, volumetric expansion and collagen type II accumulation in bovine auricular cartilage chondroprogenitors

Reconstruction of the outer ear currently requires harvesting of cartilage from the posterior of the auricle or ribs leading to pain and donor site morbidity. An alternative source for auricular reconstruction is in vitro tissue engineered cartilage using stem/progenitor cells. Several candidate cell-types have been studied with tissue-specific auricular cartilage progenitor cells (AuCPC) of particular interest. Whilst chondrogenic differentiation of competent stem cells using growth factor TGFβ1 produces cartilage this tissue is frequently fibrocartilaginous and lacks the morphological features of hyaline cartilage. Recent work has shown that growth factor BMP9 is a potent chondrogenic and morphogenetic factor for articular cartilage progenitor cells, and we hypothesised that this property extends to cartilage-derived progenitors from other tissues. In this study we show monoclonal populations of AuCPCs from immature and mature bovine cartilage cultured with BMP9 produced cartilage pellets have 3-5-fold greater surface area in sections than those grown with TGFβ1. Increased volumetric growth using BMP9 was due to greater sGAG deposition in immature pellets and significantly greater collagen accumulation in both immature and mature progenitor pellets. Polarised light microscopy and immunohistochemical analyses revealed that the organisation of collagen fibrils within pellets is an important factor in the growth of pellets. Additionally, chondrocytes in BMP9 stimulated cell pellets had larger lacunae and were more evenly dispersed throughout the extracellular matrix. Interestingly, BMP9 tended to normalise the response of immature AuCPC monoclonal cell lines to differentiation cues whereas cells exhibited more variation under TGFβ1. In conclusion, BMP9 appears to be a potent inducer of chondrogenesis and volumetric growth for AuCPCs a property that can be exploited for tissue engineering strategies for reconstructive surgery though with the caveat of negligible elastin production following 21-day treatment with either growth factor

Electric Field and Thermal Analysis of 132 kV Ceramic Oil-filled Cable Sealing Ends

This paper reports on a method combining the use of finite element simulations and external measurements to provide preliminary condition diagnosis of oil-filled cable sealing ends (CSEs) without requiring downtime. Good agreement has been obtained between the predictions from the electric field and thermal simulations and the measurements on a 132 kV oil-filled CSE. The electrostatic computation combined with electric field measurements can provide information regarding the electric field distribution inside the CSE and help in identifying potential issues with the CSE design, the materials or the cable termination process. The thermal computation combined with thermal imaging can reveal potential problems, such as high resistance connection to the busbar, and provide information regarding the cooling efficiency of the liquid dielectric. The method presented can provide the starting point for prioritizing maintenance operations on CSEs

The regulation of human mesenchymal stem cell chondrogenesis through multiaxial load

The repair of damaged articular cartilage remains a clinical problem despite the development of numerous surgical approaches for cartilage regeneration. As result new options for therapeutic approaches are being sought. One of the candidate cell types for cartilage repair are mesenchymal stem cells (MSCs). These cells can be isolated from a number of different tissues and have the ability to differentiate down several different mesenchymal lineages. This thesis focused on the use of MSCs for repairing damaged articular cartilage. Specifically I investigated the effect of producing regenerative medicine type constructs containing different populations of MSCs on the induction of chondrogenesis in response to mechanical load, compared the induction of chondrogenesis in MSCs through the application of exogenous TGF-β1 and multiaxial mechanical load and identified potentially novel markers of MSC chondrogenesis. The results presented in this thesis show that the induction of chondrogenesis in MSCs can be manipulated by producing constructs that contain separate populations of MSCs. The work demonstrated that seeding a layer of MSCs on the loaded surface of a fibrin-poly(ester-urethane) scaffold could increase the deposition of histologically detectable matrix. However, it was not possible to determine the mechanism responsible for this. Comparison of the secretomes of MSCs stimulated with TGF-β1 and mechanical load showed that these two forms of chondrogenic stimulation are not analogous and that a number of markers, including GRO and MMP13 may be useful for monitoring the progression of MSCs through chondrogenesis and hypertrophy. These data provide further insights into the effect of joint-like load on MSCs within tissue engineering/regenerative medicine style constructs, and the chondrogenic response of MSCs to this stimulation, which may prove to be useful for the development of constructs for cartilage repair

Short-sighted virus evolution and a germline hypothesis for chronic viral infections

This work was funded by The Wellcome Trust and The Royal Society grant numbers wtvm055984 (KAL) and 107653/Z/15/Z (JG), The Natural Environment Research Council grant number NE/K009524/1 (AG), and The European Research Council under the European Union’s Seventh Framework Programme (FP7/2007-2013)/ERC grant number 614725-PATHPHYLODYN (OGP).With extremely short generation times and high mutability, many viruses can rapidly evolve and adapt to changing environments. This ability is generally beneficial to viruses as it allows them to evade host immune responses, evolve new behaviours, and exploit ecological niches. However, natural selection typically generates adaptation in response to the immediate selection pressures that a virus experiences in its current host. Consequently, we argue that some viruses, particularly those characterised by long durations of infection and ongoing replication, may be susceptible to short-sighted evolution, whereby a virus’ adaptation to its current host will be detrimental to its onward transmission within the host population. Here we outline the concept of short-sighted viral evolution and provide examples of how it may negatively impact viral transmission among hosts. We also propose that viruses that are vulnerable to short-sighted evolution may exhibit strategies that minimise its effects. We speculate on the various mechanisms by which this may be achieved, including viral life history strategies that result in low rates of within-host evolution, or the establishment of a ‘germline’ lineage of viruses that avoids short-sighted evolution. These concepts provide a new perspective on the way in which some viruses have been able to establish and maintain global pandemics.Publisher PDFPeer reviewe

Phosphodiesterase 4D activity in acrodysostosis-associated neural pathology: too much or too little?

Members of the phosphodiesterase 4 (PDE4) enzyme family regulate the availability of the secondary messenger cyclic adenosine monophosphate (cAMP) and, by doing so, control cellular processes in health and disease. In particular, PDE4D has been associated with Alzheimer's disease and the intellectual disability seen in fragile X syndrome. Furthermore, single point mutations in critical PDE4D regions cause acrodysostosis type 2(ACRDYS2, also referred to as inactivating PTH/PTHrP signalling disorder 5 or iPPSD5), where intellectual disability is seen in ∼90% of patients alongside the skeletal dysmorphologies that are characteristic of acrodysostosis type 1 (ACRDYS1/iPPSD4) and ACRDYS2. Two contrasting mechanisms have been proposed to explain how mutations in PDE4D cause iPPSD5. The first mechanism, the 'over-activation hypothesis', suggests that cAMP/PKA (cyclic adenosine monophosphate/protein kinase A) signalling is reduced by the overactivity of mutant PDE4D, whilst the second, the 'over-compensation hypothesis' suggests that mutations reduce PDE4D activity. That reduction in activity is proposed to cause an increase in cellular cAMP, triggering the overexpression of other PDE isoforms. The resulting over-compensation then reduces cellular cAMP and the levels of cAMP/PKA signalling. However, neither of these proposed mechanisms accounts for the fine control of PDE activation and localization, which are likely to play a role in the development of iPPSD5. This review will draw together our understanding of the role of PDE4D in iPPSD5 and present a novel perspective on possible mechanisms of disease

Bone Morphogenetic Protein-9 Is a Potent Chondrogenic and Morphogenic Factor for Articular Cartilage Chondroprogenitors

Articular cartilage contains a subpopulation of tissue-specific progenitors that are an ideal cell type for cell therapies and generating neo-cartilage for tissue engineering applications. However, it is unclear whether the standard chondrogenic medium employing transforming growth factor-β (TGFβ) isoforms is optimal to differentiate these cells. We therefore used pellet culture to screen progenitors from immature bovine articular cartilage with a number of chondrogenic factors and discovered that bone morphogenetic factor-9 (BMP9) precociously induces their differentiation. This difference was apparent with toluidine blue staining and confirmed by biochemical and transcriptional analyses with BMP9 treated progenitors exhibiting 11-fold and 5-fold greater aggrecan and collagen type II gene expression than TGFβ1 treated progenitors. Quantitative gene expression analysis over 14 days highlighted the rapid and phased nature of BMP9 induced chondrogenesis with sequential activation of aggrecan then collagen type II, and negligible collagen type X gene expression. The extracellular matrix of TGFβ1treated progenitors analysed using atomic force microscopy was fibrillar and stiff whist BMP9-induced matrix of cells more compliant and correspondingly less fibrillar. Polarised light microscopy revealed an annular pattern of collagen fibril deposition typified by TGFβ1 treated pellets, whereas BMP9 treated pellets displayed a birefringence pattern that was more anisotropic. Remarkably, differentiated immature chondrocytes incubated as high-density cultures in vitro with BMP9 generated a pronounced anisotropic organisation of collagen fibrils indistinguishable from mature adult articular cartilage, with cells in deeper zones arranged in columnar fashion. This contrasted with cells grown with TGFβ1 where a concentric pattern of collagen fibrils was visualised within tissue pellets. In summary, BMP9 is a potent chondrogenic factor for articular cartilage progenitors and is also capable of inducing morphogenesis of adult-like cartilage, a highly desirable attribute for in vitro tissue-engineered cartilage

TAMMiCol: Tool for analysis of the morphology of microbial colonies.

Many microbes are studied by examining colony morphology via two-dimensional top-down images. The quantification of such images typically requires each pixel to be labelled as belonging to either the colony or background, producing a binary image. While this may be achieved manually for a single colony, this process is infeasible for large datasets containing thousands of images. The software Tool for Analysis of the Morphology of Microbial Colonies (TAMMiCol) has been developed to efficiently and automatically convert colony images to binary. TAMMiCol exploits the structure of the images to choose a thresholding tolerance and produce a binary image of the colony. The images produced are shown to compare favourably with images processed manually, while TAMMiCol is shown to outperform standard segmentation methods. Multiple images may be imported together for batch processing, while the binary data may be exported as a CSV or MATLAB MAT file for quantification, or analysed using statistics built into the software. Using the in-built statistics, it is found that images produced by TAMMiCol yield values close to those computed from binary images processed manually. Analysis of a new large dataset using TAMMiCol shows that colonies of Saccharomyces cerevisiae reach a maximum level of filamentous growth once the concentration of ammonium sulfate is reduced to 200 μM. TAMMiCol is accessed through a graphical user interface, making it easy to use for those without specialist knowledge of image processing, statistical methods or coding

A place to grow older… alone? Living and ageing as a single older lifestyle migrant in the Azores

International retirement migration has been typically regarded as a couple’s project. Much less is known about single lifestyle migrants who settle and age abroad and those who, after the loss of their spouse, decide to stay put. Informed by a larger pool of 36 life-narrative interviews with later-life lifestyle migrants in the Azores, and structured around four selected portrayals, this paper seeks to explore meanings of place in later life and to unpack the complex set of negotiations that the ageing self undergoes in a context of migration. Drawing on relational and unbound understandings of place, I question traditional assumptions that see single older migrants abroad as vulnerable and dependent, emphasising a more complex set of needs, desires and expectations in (and for) later-life. For these migrants, emplaced living and ageing experiences are ambiguously mixed. The islands are described as places of freedom and self-actualisation, and also “safe ports” in later-life. Yet, this does not prevent intermittent feelings of isolation and loneliness, including the desire to restore some sense of companionship and romantic life