10 research outputs found
Fiber design for high-power low-cost Yb:Al-doped fiber laser operating at 980nm
No full textWe investigated an Yb:Al-doped depressed-clad hollow optical fiber (DCHOF) for cladding-pumped 980nm laser operation. With a careful design, the nonzero fundamental-mode cutoff characteristics of a DCHOF allows the competing 1030-1060nm emission to be filtered out, despite being quite close to 980nm. The laser yielded over 3W of output power in a diffraction limited beam (M -parameter degrades to 2.7, as a result of increased cladding-mode lasing, as the cladding thickness is reduced
Advanced fibre Bragg gratings: application to dispersion compensation
No full textThis thesis presents the design and fabrication issues related to advanced fibre Bragg gratings with particular emphasis on realising novel devices with application to dispersion compensation. Developments of fabrication techniques presented herein resulted in the first demonstration of continuously-written metre-long chirped gratings. Subsequent development led to the realisation of gratings with deliberate spectral shaping (without affecting the group delay response) and the first demonstration of gratings designed to compensate both 2nd and 3rd order dispersion of dispersion-shifted fibre. Transfer of this fabrication technology to a commercial fabrication facility resulted in several successful system trials. Results are presented of the first demonstration of chirped FBGs with the application of dispersion-managed soliton transmission. Error-free transmission was obtained over 1000 km of non-dispersion shifted fibre. Application of a new reverse-engineering grating design concept has allowed the fabrication of gratings designed to meet the needs of DWDM dispersion compensation on a 50 GHz grid. The first experimental results of devices designed by this technique are presented, and have sufficient bandwidth utilisation for application in long-haul transmission and optically-transparent networks
The agency of service user and carer engagement in health and social care education
No full textService user and carer involvement in health and social care education in the UK has gained momentum over the last two decades, largely driven by consumerist and democratic ideologies. This is reinforced by the health and social care regulatory bodies such as the Nursing and Midwifery Council (NMC) and the Health and Care Professions Council (HCPC).
This thesis presents a series of eight peer reviewed papers that have focussed on the agency of service user and carer involvement in health and social care education. The accompanying commentary draws the papers together and locates them within an overarching theoretical framework, ‘The Ladder of involvement’. This portfolio of evidence demonstrates a coherent approach that draws on underlying philosophies and theoretical underpinnings and displays contribution to knowledge in five distinct sections: Contribution to the literature with new findings, location of the findings within the current literature, location of the findings within the theoretical framework, contribution to the refinement and development of theory and contribution to dialogue and debate.
The key message from the studies undertaken as part of this portfolio of evidence is that service user and care involvement in health and social care education enhances student learning and influences their future practice. However, there must be a well-developed infrastructure within higher education institutions that recognises the complexities of user involvement for the key stakeholders. There is a pressing need for additional research to further substantiate the benefit of user involvement for all parties concerned, in order for user involvement to take its place as a core component of health and social care education
Transcription Factor AP-2 Regulatory Signatures in Breast Cancer
No full textPhDAP-2 transcription factors are highly conserved basic helix-span-helix proteins whose
members ((x, ß, y, S and c) are crucial regulators of bryonic development. They also
play an important role in human neoplasia. uohis ochemical studies have detected
high levels of AP-2y expression in primary tumo of breast cancer patients. This high
expression has been correlated with reduced survival in all patients and reduced survival
in an ERa positive subset treated with hormone therapy. In breast cancer cell lines, AP-
2 factors have been implicated in the regulation of the ERBB2 proto-oncogene and ERa.
In an effort to further understand the role of AP-2y in breast carcinoma, this study has
sought to identify additional AP-2 activated cellular pathways and ultimately novel
transcriptional targets for AP-2 through the use of gene expression profiling.
RNAi using three independent AP-2y targeting sequences, has been used to deplete AP-
2y levels in the ERa positive MCF-7 breast carcinoma cell line, chosen as it exclusively
expresses the AP-2y family member. Microarrays were then utilised to create an AP-2y
dependent transcription profile. Statistical comparisons between non-silencing control
siRNA and AP-2y targeting siRNA groups identified a total of 162 gene expression
changes (p<0.01). These changes implicate AP-2y in the control of cell cycle
progression and developmental signalling. Indeed a role for AP-2y in the control of cell
cycle, in particular at the GUS transition, has been verified using flow cytometry.
Several of these gene expression changes, including IGFBP3, Transgelin and
KIAA1324, have been confirmed using qPCR and immunoblotting.
Finally, elevated levels of p21 mRNA and protein have been observed following AP-2y
silencing in MCF-7 cells. Additionally, the activity of a p21 promoter reporter is
repressed following transfection with an AP-2y expression construct in HepG2 cells.
These results coupled with ChIP experiments showing AP-2y occupancy at the proximal
promoter region of p21 in cycling MCF-7 cells, implicate AP-2y in the repression of
p21 transcription and suggest a role for AP2y in- the, control of cell cycle in breast
carcinoma in part through the transcriptional repression of p21
Gene expression in Rhipicephalus (Boophilus) microplus: Identification of tick antigens and investigation of the tick RNA interference pathway
No full textThe cattle tick Rhipicephalus microplus is a hematophagous ectoparasite of major veterinary importance both globally and in Australia. Australian beef cattle producers in the tropical and subtropical regions of the continent suffer estimated annual losses of AU$175m. The development of a novel tick vaccine has therefore been identified as a high priority research goal by all stakeholders of the Australian beef industry (Playford 2005). A novel bioinformatics methodology for the identification of potential vaccine targets in the R. microplus expressed sequence tags sequences was developed and applied resulting in the identification of 68 potential tick vaccine candidates. Furthermore, the applicability of RNA interference (RNAi) for tick gene functional studies was assessed. This study comprehensively characterized the major components of the RNAi pathway in both R. microplus and Ixodes scapularis, which resulted in the identification of an I. scapularis homologue of Dicer protein, as well as the identification of Argonaute-1 and -2 homologues in both I. scapularis and R. microplus. Most notably, homologues of a C. elegans RNA-dependent RNA polymerase were found in both R. microplus and I. scapularis, a protein involved in the amplification of the RNAi signal and previously not identified in any other arthropod species. A comparative RNAi study targeting 10 highly conserved R. microplus homologues of Drosophila genes with known RNAi phenotypes was conducted. The R. microplus homologue of D. melanogaster Ubiquitin-63E showed the strongest phenotypic changes in both cell culture and in adult female ticks injected with dsRNA. This Ubiquitin-63Esilencing effect in fully engorged females was subsequently investigated by microarray analysis. The contribution of potential off-target effects to phenotypic changes was assessed by developing an in silico analysis and combining the prediction results with the microarray data. A total of 224 unique transcripts were detected as differentially expressed. The off-target analysis revealed that 11 off-targets predicted by the in silico analysis were present in the list of differentially expressed genes. The research presented in this thesis has contributed to the development of novel approaches for reverse vaccinology in ectoparasites and it has furthermore improved the understanding of the mechanisms of RNAi and the delivery of double-stranded RNA in ticks
Exploring the Role of Topoisomerase II Beta in Macrophage Maturation and Pro-inflammatory Cytokine Production
No full textAlthough it is known that DNA topo IIβis required for the regulation of transcription during neural development and differentiation, it is not clear whether the enzyme is required during differentiation of human monocytes into macrophages and/or the subsequent transcription of
cytokine genes. To test this, a robust model of differentiation of monocyte-like cells into macrophage-like cells using U937 and HL-60 cells treated with phorbol 12-myristate 13-acetate (PMA) and Lipopolysaccharide (LPS) was validated. Differentiation was determined by morphological and growth characteristics and CD11b surface antigen expression as determined by flow cytometry. qRT-PCR was also used to measure mRNA transcript levels of key genes known to be up-regulated during monocyte differentiation and the secretion of pro-inflammatory cytokines produced by differentiated cells were measured using ELISA.
siRNA topo IIβknockdown did not hinder monocyte-like cells from undergoing differentiation, however experiments revealed a correlation between topo IIβknockdown and secreted TNFα, with the latter decreasing when topo IIβwas reduced. This pattern was also noted when measuring IL-1βsecretion. Similar results were seen using a Murine transgenic fibroblast cell line lacking topo IIβ, which when stimulated with LPS secreted significantly lower levels of IL-6 compared to the wild type cells. Thus topo IIβexpression is necessary for
secretion of normal levels of the cytokines, TNFα, IL-1βand IL-6 in response to LPS at certain time points. In addition in the macrophage-like state of the two cell lines, the relative levels of the βisoform (mRNA and protein) were shown to be significantly increased
compared to α, further outlining the importance of topo IIβin the differentiated state. Chromatin immuno-precipitation followed by qPCR showed however that topo IIβwas not associated at three defined proximal promoter regions of either the TNFαand IL-1βgenes, although further studies are required to rule out a direct association of topo IIβwith these and other regions of the genes.
Down regulation of topo IIβprotein using the inhibitor ICRF-193 did not hinder monocyte-like cells from undergoing differentiation either. However, contrary to the knockdown results, a 6 h pre-treatment with 1 nM ICRF-193 increased TNFαlevels in these cells, both at the
mRNA and the protein level, along with a slight increase in secreted TNFα. NF-κB, EGR2, TLR4 and TLR2 transcript levels were also increased under these conditions. Thus further studies are required to determine if these increases are due to additional cellular effects of the
drug or whether topo IIβmay play an inhibitory effect on transcription.
Thus it is clear that topo IIβplays an important role in expression of cytokines and understanding the exact nature of this requires further research that may yield potential new avenues for treatment of disease
Investigation of the role of t(4;6) and 6q15 deletion in prostate cancer development and progression
No full textPhDThe recent identification of TMPRSS2:ETS gene fusions highlighted the importance of chromosomal rearrangement and fusion gene development in prostate tumourigenesis. We previously reported a recurrent translocation, t(4;6), in prostate cancer (PCa) with the breakpoints identified at 4q22 and 6q15 in the LNCaP cell line. A small deletion adjacent to the 6q15 breakpoint was found, which is consistent with the frequently lost chromosome region previously reported and detected by our micro-array analysis. Here I accessed the prevalence of the t(4;6)(q22;q15) and the 6q15 deletion and also looked for relevant candidate tumour suppressor genes (TSGs) in PCa. Using fluorescence in situ hybridisation (FISH) analysis, t(4;6)(q22;q15) was detected in 78 of 667 clinical, localised PCa samples. Statistical analysis showed it was not independently associated with patient outcome but occurred more frequently in high clinical T stage, high tumour volume specimens and in those with high baseline PSA (prostate-specific antigen) (P=0.001, 0.001 and 0.01 respectively). We hypothesised that both t(4;6)(q22;q15) and the 6q15 deletion contribute to the inactivation of TSGs at 6q15, which are associated with PCa development and progression. Therefore I investigated the TSGs located in this region. Using FISH analysis, this deletion was confirmed in 46% (13 in 28) of the PCa samples. Using Exon array to systematically detect inactivated genes in this region, four candidate genes, CNR1, PNRC1, GJA10 and BACH2 were identified with common down-regulation. The real-time PCR analysis validated these exon array results. However, with additional controls and clinical cancer samples, only two genes (CNR1 and BACH2) still showed significant down-regulation. CNR1 protein expression was absent in 76.8% (43 in 56) of PCa cases whereas its
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expression was shown in 83.9% (26 in 31) of benign prostatic hyperplasia (BPH) samples as demonstrated by immunohistochemistry (IHC) analysis (p<0.001). In contrast, cytoplasmic expression of BACH2 was slightly less in the examined PCa cases (14.0%, 12 in 86) compared with BPH samples (6.9%, 2 in 29). However, nuclear expression of BACH2 was significantly higher in prostate cancer cases (45.3%) than it was in BPH samples (10.3%) (p=0.001). Five of the 43 PCa samples without CNR1 expression were selected for sequencing and one of the five samples had insertion in the CNR1 genomic DNA. The cellular function study of PCa cell lines indicated that the CNR1 might function as a tumour suppressor involved in cell proliferation and invasion. In conclusion, both t(4;6)(4q22;6q15) and the 6q15 deletion are frequent events and the gene CNR1 is a candidate TSG at this deletion region in PCa
Genetic Susceptibility Factors for Eczema
No full textEczema (atopic dermatitis) is one of the most common chronic inflammatory skin diseases in infants and children with prevalence rates of up to 20%. The disease frequently co-occurs with other atopic disorders such as asthma and rhinitis and is often accompanied by elevated levels of immunoglobulin E (IgE) antibodies and aberrant IgE-mediated responses to otherwise harmless environmental agents. Eczema is considered a polygenic disease, caused by a complex interplay of various predisposing genes, which additionally interact with environmental, non-genetic components. This work focuses on the identification of genetic factors contributing to the aetiology of eczema. To this end several candidate gene association studies were performed and the first and to date only genome-wide association studies (GWAS) on eczema as well as on total serum IgE levels were conducted. This thesis provides the first independent replication study on the filaggrin gene (FLG). FLG is located in the epidermal differentiation complex (EDC) on chromosome 1q21 and encodes a structural protein with key functions in the formation of the epidermal barrier. In a family-based approach it was clearly shown that the two loss-of-function mutations R501X and 2282del4 in this gene strongly predispose for eczema (Odds Ratio (OR) 2.73; P-value 5.1x10-8). Along with a consistent and prominent association between these mutations and eczema subsequent investigations delineated their impact on distinct eczema subtypes as well as on eczema-related traits like asthma and hay fever. Strong associations with allergic sensitization (P 2.3x10-7), increased total IgE (P 9.8x10-8) and the atopic (OR 3.66; P 4.6x10-5), but not the non-atopic form of eczema were observed. Both disease alleles predispose to the early-onset and severe form of eczema (OR 5.21 and 2.65; P 2.8x10-6 and 0.0043, respectively). It was demonstrated that mutant FLG alleles increase the risk for allergic rhinitis (OR 2.64; P 2.5x 10-6), and patients with FLG-related eczema were shown to be at higher risk to develop additionally allergic asthma (OR 3.49; P 1.0x10-5). Moreover, it was possible to evaluate the relevance of FLG risk alleles on the population level in a German cross-sectional study. On the basis of an observed carrier frequency of 7.4% for the four most prevalent mutations the Population Attributable Risk (PAR) was estimated as high as 13.5% and the penetrance reached 38.5%. The remarkably strong effect of the FLG gene on eczema and asthma risk was finally confirmed with the help of a comprehensive meta-analysis based on 24 studies published until 2008. This analysis emphasizes the important role of FLG as the first validated and strong genetic risk factor for eczema, and provides a general measure of its impressive effect size. FLG mutant alleles were shown to cause a more than threefold increased eczema risk and clearly convey predisposition to the particular phenotype of asthma occurring in the context of eczema, but apparently not to asthma independent of eczema. These observations point towards a genetically disturbed epidermal barrier as key event in the pathogenesis of eczema and as risk factor for allergic sensitization and concomitant respiratory disease. The first GWA study on eczema published in 2009 identified a novel susceptibility locus on chromosome 11q (OR 1.22; P 7.64x10-10), as well as putative additional risk variants (OR 1.20; P 3.52x10-5) apart from FLG within the EDC. The associated variant on chromosome 11 is located in an intergenic region between the two plausible candidates C11orf30 (chromosome 11 open reading frame 30) and LRRC32 (leucine rich repeat containing 32). Hence the gene affected by the identified risk allele on chromosome 11 is still elusive and subject of current investigations. With the help of an independent GWAS on total IgE levels it could be shown that variants within the gene encoding the alpha chain of the high affinity receptor for IgE (FCER1A) strongly influence serum IgE levels (P 7.08x10-19). Presence of the homozygous state of the rs2427837 minor allele was estimated to decrease serum IgE levels by 35%, and was strongly associated with decreased FCER1A cell surface expression on basophils. Concurrently, the study identified the cytokine cluster on chromosome 5q31 as a second susceptibility locus for altered IgE concentrations (P 6.28x10-7 - 4.46x10-8). Taken together, the discovery and comprehensive description of novel risk variants and potential susceptibility genes for eczema and related phenotypes change and complete our understanding of the complex aetiology of eczema and atopic diseases. Although to date immunological alterations have been the focus of pathophysiological considerations, new concepts recognize the impairment of the skin barrier as well as an abnormal reactivity of the adaptive and innate immune system as important factors. Eczema causes significant problems in everyday life and has been shown to lead to decreased quality of life, vitality, and mental health of patients, concurrently constituting a high economic burden with direct and indirect costs similar to those of asthma. Hence, the development of new prognostic and therapeutic tools is of essential relevance. Knowledge about the genetic background of eczema translates into a clearer understanding of mechanisms of the disease and ideally into the future development and refinement of diagnostic and therapeutic strategies. The challenge of future studies will be the further dissection of the genetic architecture of eczema and the confirmation of observed relations between variations in the human genome and eczema by functional approaches in order to illuminate the pathophysiological nature of this complex disease
