Surgery (St Vincent's) - Theses
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ItemColorectal cancer in rural regional Australia( 2017)Colorectal cancer (CRC) is the second commonest cancer in Australia. The survival outcome of colorectal cancer patients within Australia is reported to vary with population density and between health services, with some literature showing poorer prognosis in rural regional and remote patients. Chapter one aims to outline some key issues in CRC such as epidemiology, diagnosis, treatment, surveillance, and outcome whilst chapter two aims to present a systematic review of how geographical disparity influences CRC survival. There are many potential factors that contribute to a poorer prognosis in rural regional CRC patients though the literature is limited, and at times, inconsistent. Thus, there is a need for regular audit, reporting and benchmarking of outcomes in CRC patients against agreed standards. I reviewed the long-term outcomes of CRC at Barwon Health, which serves the South West Victoria, a region with a population of some 500,000. My aim was to determine whether changes introduced to the management of CRC translated into improved survival after surgery. The literature to date has suggested that patients living in rural and regional Australia (grouped together) have worse colorectal cancer survival rates than those living in metropolitan Australia. This thesis was based on a prospectively maintained registry kept over a period of thirteen years from 2002 to 2014, that had accumulated 1079 patients who had undergone surgery at the University Hospital Geelong for CRC (744 colon cancers and 335 rectal cancers). The overall number of operations per year increased over time (p=0.037) but with similar proportions of elective and emergency surgery (p=0.75) and tumour stage (P=0.21). This lack of change in the proportion of elective cases was in spite of the Federal Government introducing a National Bowel Cancer Screening Program in 2006. The proportion of patients with severe comorbidities did increase (p=0.015) over the study period. The median survival after surgery by stage was 123 months, 141 months, 76 months and 17 months for stages I to IV CRC respectively. Overall, there were improvements observed in both peri-operative mortality (POMR) (p=0.028) and long- term survival (p=0.0025) of CRC patients in this major regional centre. I then reviewed the outcome of patients with metastatic disease. The Geelong database included 843 patients who had undergone resection and primary anastomosis for their primary tumour (661 colon cancers, 182 rectal cancers). Metastatic disease was present in 16% (135 patients) and was associated with an increased risk of anastomotic leakage (13% vs. 5%, p=0.003) and a higher peri- operative mortality rate (9.6% vs. 2.8%, p=0.0003). Patients with anastomotic leakage had a reduction in the overall survival (121 months vs. 66 months, p=0.02). The fifth chapter aimed to perform a regional study to identify patients with colorectal cancer at higher risk of developing metastatic disease. There were 503 patients (345 colon and 158 rectal) with non-metastatic (stage I-III) CRC who had resections and were followed up for at least five years. Metastatic progression was, as expected, significantly higher for patients with stage III disease (aHR 4.42 for colon cancer 95% CI 1.74 to 11.23, aHR 3.34 for rectal cancer 95% CI 1.36 to 8.22), and those with lymphovascular invasion (aHR 2.94 95% CI 1.70 to 5.06). Metastatic disease was also more likely to eventuate in those with severe comorbidities (aHR 2.18, 95% CI 0.26 to 0.86), and in colon cancer patients with the lowest socioeconomic status (aHR 2.03 95% CI 1.23 to 3.34). Gender, tumour location and geographical location (rural or regional) was not associated with metastatic progression. Before determining surveillance strategies targeting higher-risk patient groups in regional Victoria, these findings would require confirmation from similar studies in other regions of rural/regional Victoria, such as Bendigo, Albury / Wodonga, Latrobe Valley, or Shepparton.
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ItemRadiation-associated breast, thyroid and solid malignancies in patients attending the Peter MacCallum Cancer Centre Late Effects service( 2017)Background: Survivors of childhood, adolescent and young adulthood (CAYA) malignancies have an increased risk of subsequent primary malignancies, particularly after exposure to therapeutic radiation. The Peter MacCallum Cancer Centre Late Effects (PMCC LE) service provides individualize, multidisciplinary care and surveillance advice for survivors of malignancies, especially CAYA malignancies. Methods: A retrospective review was performed of patients exposed to therapeutic radiation attending the PMCC LE service from 1st January 2000 to 20th February 2013. All invasive malignancies, in-situ malignancies, benign tumours and deaths were evaluated. Separate time-to-event analyses was performed for radiation-associated breast, thyroid and solid malignancies in patients exposed to chest, thyroid and any therapeutic radiation respectively, measured from the date of first attendance to the PMCC LE service and stratified by the interval from completion of radiation to the first attendance. The incidence of breast and thyroid malignancies was compared to the Australian general population. Compliance with breast and thyroid surveillance recommendations was determined by assessing the number of screen events over the period of attendance to the service. Clinicopathological features and management of radiation-associated breast and thyroid and other solid malignancies was examined. Ultrasound and cytological workup of radiation exposed thyroid nodules was assessed. Results: After excluding 187 patients, 534 included patients developed 194 invasive malignancies; 147 were radiation-associated and 47 non-radiation associated. The most common malignancies were non-melanoma skin (37.1%), thyroid (17.0%) and breast (12.9%) malignancies. Patients whose first attendance was ≥15+ years after radiation exposure experienced the highest incidence of radiation-associated breast, thyroid and solid malignancies, with 23%, 8% and 27% affected after 10 years of subsequent follow-up respectively. The incidence of breast and thyroid malignancy was elevated 11.2 and 57.6 times respectively compared to the Australian general population (both p<0.001). Compliance with breast surveillance using mammography or any screening modality was observed in 18.4% and 28.6% of women at risk respectively. Twenty-eight radiation-associated breast malignancies occurred in 24 women (16.7% bilaterality). Breast malignancies diagnosed after the first attendance to the PMCC LE service were more likely screen-detected (p=0.002). Most were hormone receptor positive (87.5%), invasive ductal carcinomas (82.1%) managed with mastectomy (89.3%). Compliance with thyroid surveillance was observed in 76.9% of patients at risk. Ultrasound features of microcalcification and increased internal vascularity had a low sensitivity (62.5%) for predicting a malignant nodule, which improved when used in conjunction with a Bethesda IV-VI result (91.7%), although cytological assessment was not performed in 45.6% of operative cases. Thirty-three patients had a radiation-associated thyroid malignancy; 45.4% (n=15) were incidental. The majority were papillary thyroid cancers (88.9%); of which 12.5% were node positive and 34.4% were multifocal. Node positive thyroid cancers were more likely to present symptomatically (p=0.03). There were 36 deaths in the cohort (6.7%), most commonly attributable to radiation-associated malignancies (41.9%), especially brain, breast and sarcomatous malignancies. Conclusions: Patients attending the PMCC LE service have a high burden of subsequent malignancies that typically occur after a long latency. Ongoing long-term surveillance is essential and judicious management with adherence to guidelines is advocated in this unique population of patients.
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ItemPrediction and prognosis in anal cancer: developing models to improve patient outcome( 2017)Anal squamous cell carcinoma is a human papilloma virus–related disease for which definitive treatment comprises chemoradiotherapy that has not changed substantially for forty years. Few advances in treatment have been made since then, especially for those patients who develop disease relapse and for whom no surgical options exist. Predicting responses in patients for whom conventional treatment will fail remains elusive and is a significant clinical problem. As anal cancer is reasonably described as a rare cancer, innovative approaches are required to address this pressing clinical issue as large clinical trials are exceptionally challenging and are unlikely to be undertaken. This thesis describes a range of research strategies to identify potential avenues to predict and improve patient responses to existing and novel therapies. It comprises a combination of clinical and translational research. Using our institutional database I have assessed the utility of post-treatment imaging with FDG-PET as it may serve as a means of early detection of poor response to treatment. I found that a complete metabolic response on post-treatment PET scan was predictive of overall survival and disease-free survival. The database, which spans a thirty year period, was also interrogated to explore patterns of treatment failure, subsequent salvage treatment and outcomes. I found that multiple treatment modalities have been utilised to treat patients with persistent or recurrent disease, with satisfactory survival benefit in carefully selected patients. I also evaluated the literature that investigated the molecular biology of anal cancer finding that no clinically valuable biomarkers have emerged. Some suggestions have been reported that regulators of apoptosis, including survivin, and agents targeting the PI3K/AKT pathway, might offer opportunities for targeted therapy. Additionally, antibody therapy targeting epidermal growth factor receptor may prove efficacious although the safety profile in combination with standard chemoradiotherapy has proven to be suboptimal. In the laboratory, next generation RNA sequencing was utilised in eleven anal SCC patient samples. Through stratification of the tumours into clinically relevant groups and Bioinformatic analysis, eight genes with differential expression were chosen for further validation. One of these genes was identified as a novel target which could ultimately lead to expanding therapeutic options in anal cancer management. Due to a lack of pre-clinical models, including cell lines and mouse models for testing new therapies, I developed a new anal cancer model based upon patient-derived tumour xenografts. I used this model in a pilot experiment to assess the novel drug target identified by RNA-seq. The outcomes were promising with stand-alone efficacy of the novel drug observed with statistical significance, while also validating the feasibility of using xenografts for anal SCC. This thesis builds upon the clinical experience of decades of management of patients with anal cancer identifying both clinical and laboratory approaches to advance assessment and identify novel treatment possibilities for this group of patients.
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ItemDefining lymphatic microstructure and the genetic basis of lipolymphedema( 2017)The lymphatic system regulates tissue fluid homeostasis, intestinal fat absorption, and immune cell trafficing. Lymphedema is soft tissue swelling secondary to lymphatic dysfunction, which results in the accumulation of tissue fluid in the interstitial space. This might occur as a primary disorder of the developing lymphatic system, or alternatively lymphedema might be an acquired disorder secondary to lymphatic injury. For example, secondary lymphedema is a common problem following cancer and cancer treatments such as lymph node surgery and radiotherapy, resulting in significant morbidity. Radiotherapy is an established risk factor for lymphedema, and in addition to causing direct injury to the lymphatic vessel, it is possible that alternative mechanisms might also contribute to radiation‐induced lymphatic dysfunction, such as localized ischemia of the lymphatic wall. It is also likely that predisposing genetic risk factors are at play, as not all individuals exposed to the same risk factors will develop secondary lymphedema. Lipoedema is a different form of soft tissue swelling due to the abnormal accumulation of adipose tissue. Lipoedema and lymphatic dysfunction appear to be linked, as individuals frequently develop a degree of lymphedema, particularly as the condition progresses in severity, where it may be decribed as lipo‐lymphedema. The cause of lipoedema and the genetic basis of the condition are currently unknown. This thesis aims to discover and define alternative mechanisms for lymphtic dysfunction in the context of secondary lymphedema, particularly focussing on the supply of oxygenated blood to the lymphatic vessel wall. We also aim to describe inheritance patterns and the genetic factors involved in lipoedema and lipo‐lymphedema. Such knowledge might uncover therapeutic targets and facilitate the development of treatments for lymphedema and lipoedema, including gene therapy.
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ItemToward a functional and permanent interface between materials and skin( 2017)All medical devices that pass through skin are plagued by infection and other problems at the skin interface. Such devices include intravenous lines, catheters, robotics, osseointegrated rods, bone anchored hearing aids, and tracheostomy tubes. A robust and functional skin interface could permit long-term infection-free implantion of these types of devices. Failure of skin attachment to percutaneous devices results from skin avulsion, infection, and epidermal marsupialisation. Prevention of marsupialisation requires epidermal cell (keratinocyte) attachment and viable (living dynamic) subepidermal tissue integration into implant pores. To address these problems, a structure was designed, a “cap-scaffold,” to permit dynamic tissue integration aided by negative pressure wound therapy (NPWT). While the design and testing of a skin-material interface system for use with NPWT was the ultimate destination of this thesis, surface-optimisation to improve epidermal attachment was thought to be important. Hence, the majority of work reported herein preceded in vivo work and involved surface optimisation techniques and their measurement: a method of surface functionalisation with thiol groups and the conjugation of collagen and biotin to these groups with maleimide linkers; the novel use of conjugated collagen immunoassay as a method for assessing the efficacy of surface functionalisation; the novel finding that collagen 4 (C4) is more stable than collagen 1 (C1) in both adsorbed and covalently-bound forms; and the assessment of keratinocyte responses to collagen and laminin-332 (L332). Thiol groups can undergo a large variety of chemical reactions. They are used commonly in solution phase to conjugate bioactive molecules. Previous research on solid substrates with continuous phase glow discharge polymerisation of thiol-containing monomers might have been compromised by oxidation. Thiol surface functionalisation via glow discharge polymerisation has been reported as requiring pulsing. Herein, continuous phase glow discharge polymerisation of allyl mercaptan (2-propene-1-thiol) was used to generate significant densities of thiol groups on a mixed macrodiol polyurethane and tantalum. Three general classes of chemistry are used to conjugate proteins to thiol groups, with maleimide linkers being used most commonly. Herein the pH specificity of maleimide reactions was used effectively to conjugate surface-bound thiol groups to amine groups in collagen. XPS demonstrated surface-bound thiol groups without evidence of oxidation, along with the subsequent presence of maleimide and collagen. Glow discharge reactor parameters were optimised by testing the resistance of bound collagen to degradation by 8 M urea. The nature of the chemical bonding of collagen to surface thiol groups was effectively assessed by colourimetric assay (ELISA) of residual collagen after incubation in 8 M urea over eight days and after incubation with keratinocytes over fifteen days. The facile creation of useable solid-supported thiol groups via continuous phase glow discharge polymerisation of allyl mercaptan opens a route for attaching a vast array of bioactive molecules. Traditional methods of assessing surface functionalisation, including spectroscopy and chemical labelling, often involve significant error and conjecture about bonds. Proteins that improve cell attachment have specific pKa’s and optimum binding requirements that may differ from the conditions required for chemical labelling. The utility of collagen ELISA to optimise acetaldehyde glow discharge polymerisation (Aapp) reactor parameters was tested. Accurate stepwise increases in collagen conjugation strength were demonstrated by incubating specimens in 8 M urea for 5-8 days followed by ELISA to test for residual surface collagen. Surface modifications also were assessed by X-ray photoelectron spectroscopy (XPS). Results suggested that ELISA after bond-stressing with urea may suffice for optimising surface functionalisation and that traditional methods of analysis may be superfluous if protein conjugation is the aim of functionalisation. C1 is used commonly to improve biological responses to implant surfaces. The stability of C1 was compared with C4 on a Elast-Eon™, both adsorbed and covalently bound via Aapp and reductive amination. Substrate specimens were incubated in solutions of C1 and C4. The strength of conjugation was tested by incubation in 8 M urea followed by ELISA to measure residual C1 and C4. L332 was superimposed via adsorption on C4-treated specimens. Keratinocytes were grown on untreated, C1-treated, C4-treated, and C4 + L332 treated specimens, followed by measurement of cell area, proliferation, and focal adhesion density. Adsorbed C4 was shown to be significantly more stable than C1 and covalent conjugation conferred even greater stability, with no degradation of C4 over twenty days in 8 M urea. Cell growth was similar for C1 and C4, with no additional benefit conferred by superimposition of L332. The greater resistance of C4 to degradation may be consequent to cysteine residues and disulphide bonds in its non-collagenous domains. The use of C4 on implants, rather than C1, may improve their long-term stability in tissues. Following the completion of in vitro research and armed with the improved stability findings of covalently bound C4, thesis work progressed to pig studies. Six wounds were made on the backs of each of four 3-month old pigs. Four unmodified (no caps) scaffolds were implanted along with 20 cap-scaffolds. C4 was attached to 21 implants. NPWT then was applied. Structures were explanted and assessed histologically at day 7 and day 28. At day 28, there was close tissue apposition to scaffolds, without detectable reactions from defensive or interfering cells. Three cap-scaffolds explanted at day 28 showed likely attachment of epidermis to the cap or cap-scaffold junction, without deeper marsupialisation. NPWT appeared to facilitate dynamic integration with macroporous scaffolds, epidermis appeared to attach to caps in most full-thickness implanted cap-scaffolds, and there was no infection or inflammation at 28 days. This time span was insufficient to permit conclusions about long-term infection risk, but skin-interfaced devices implanted for 28 days could still be useful in clinical practice, e.g. a course of chemotherapy or dialysis. Implant numbers were insufficient to reveal effects of conjugated C4 on epidermal attachment in vivo. Given the improved epidermal attachment to implanted contact lenses optimised with C1 reported by others, the effects of conjugated C4 on skin-material interfacing warrants further investigation. While numbers were limited, qualitative results suggested that a robust skin-material interface system might be within reach using toric-shaped cap-scaffolds with NPWT.
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ItemUsing a murine bioengineering model to study cancer cell biology: the effects of mammographic density on breast cancer progression( 2017)Breast cancer (BC) remains a leading cause of cancer-related morbidity and mortality for women worldwide. Mammographic density (MD) has been well recognized as a strong risk factor for BC, independent of its masking effect for small tumours on a mammogram. Due to the common presentation of high MD (HMD) in the community, especially in pre-menopausal women, MD is arguably the most important risk factor for BC when taking into account of its high population-attributable risk. However, much remains to be learned about the biological mechanism underlying MD-associated BC risk. My team previously utilised a murine engineering biochamber model to show that human breast tissues of high and low MD not only remained viable, but also maintained their radiological and histological features in relation to their MD status for at least 6 weeks. Extending from the prior findings, this PhD study aimed to further explore the biological mechanisms behind MD-associated BC risk, and is divided into four main sections: (i) I first attempted to develop the murine biochamber model further by implanting collagenase digested and flow cytometry sorted single cells to pave the way for manipulations of specific cell types that might be responsible for HMD. I found that collagenase digested and flow cytometry sorted single cells of high or low MD breast tissue reconstituted glandular organoids in murine chambers, albeit in limited numbers; (ii) second, based on a collection of human high and low MD breast tissues from prophylactic mastectomy procedures over a period of 5 years, I evaluated the histological differences between within-individual high and low MD mammary specimens of all participants, and found that the HMD tissue microenvironment was significantly altered compared with that of LMD -- HMD was characterised by increased levels of collagen organisation and quantity, aromatase immunoreactivity and immune cell infiltration of various subtypes; (iii) The serendipitous finding of increased immune cell presence in HMD led to my subsequent examination of the potential differences in immune cell representation between patient-matched high and low MD tissues, the immune infiltrates of both innate and adaptive system, and cytokines such as IL-6 and IL-4, and I found that immune cells of various subtypes were significantly raised in HMD tissue compared with LMD; and (iv) parallel to the human breast tissue studies, I also tested whether high and low MD human tissue had any direct effect on cancer cell growth and dissemination; using our murine biochamber model, I showed that compared to co-inoculation with LMD tissue, HMD tissue stimulated the progression of MCF10DCIS.com cells that represented cells of ductal carcinoma in situ (DCIS), to lesions resembling invasive ductal carcinomas, as well as their metastases in the mice hosting the murine biochambers. Over the past twenty years, the body of evidence on various aspects of MD and its associated BC risk has been expanding, however, to the best of my knowledge, (i) my study contained the largest cohort of high-risk women in Australia to characterise immunohistochemical and immune cell differences between high and low MD, and (ii) the work presented in this thesis is the first to utilise the murine biochamber xenograft model to test human breast organoids formation from single cells and to evaluate the direct effect of MD on human breast cancer cell progression in vivo. Collectively my work has defined that HMD is characterised by an increase in stromal cells, extracellular matrix and inflammation. I have shown that HMD stimulated the progression of early stage BC cells, which highlights the importance of MD being considered for BC diagnoses, treatments and surveillances.
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ItemInvestigating epithelial-mesenchymal plasticity in breast cancer using circulating tumour cells and circulating tumour DNA( 2017)Breast cancer is the most frequent invasive cancer among women worldwide with mortality primarily caused by metastasis. One of the key proposed processes underlying metastasis, including the escape to the bloodstream, is epithelial- mesenchymal plasticity (EMP). This refers to the dynamic transition between epithelial and mesenchymal phenotypes of cells within the primary tumour mass. Within the bloodstream, tumour cells and tumour DNA, which are referred to as circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA), respectively, have showcased their potential use as liquid biopsy in cancer management. The presence of CTCs has been shown to associate with poor prognosis in metastatic cancers, which becomes worse with higher CTC numbers. By virtue of carrying genetic and epigenetic features of primary tumours, ctDNA has demonstrated its utility in detecting and monitoring cancer progression. Various studies conducted on the molecular characterisation of CTCs have generated data supporting the role of EMP in generating CTCs. However, the dynamic changes in expression, especially of genes associated with EMP, between primary tumours, CTCs and metastases remain far from conclusive. In keeping with this paradigm, as cancer cells in the primary tumours shift from the epithelial to the mesenchymal phenotype, any released ctDNA may have epithelial and/or mesenchymal features depending on its cellular origin. To date, research on the utility of EMP-associated methylation markers to detect ctDNA is lacking. In light of the suggested role of EMP in different key steps of cancer progression and metastasis, this thesis has aimed to study EMP reflected in CTCs and ctDNA to provide further insights into the CTC molecular characteristics and assist in the detection of ctDNA. This thesis is comprised of two principal areas of study: (1) the gene expression profiling of CTCs in two human breast cancer xenograft models, and (2) the locus- specific methylation profiling of breast cancer cell lines and breast tumours. Firstly, the expression profiles of EMP-associated genes were characterised in CTCs at the population level and the single cell level, and were compared with the expression profiles of primary tumours and (where possible) metastases, for two xenograft models, the MDA-MB-468 and ED-03. In pooled CTCs relative to primary tumours from both models, a significant increase in expression of mesenchymal markers (SNAI1, VIM, SERPINE1 and NOTCH1), and surprisingly, of a typical epithelial marker CDH1, were observed. A decrease/loss of EPCAM was reproducibly observed in CTCs of both models, while decreased CD24 and EGFR in CTCs were only seen in the MDA-MB-468 model. Genes associating with hypoxia (HIF1A, BNIP3 and APLN) and cellular metabolism (PPARGC1A) were also significantly elevated in CTCs of both models. In additional studies, a direct lysis method was successfully optimised to assist the gene expression study in single cells. The subsequent analysis of single CTCs revealed heterogeneity of CTCs, with co-expression of epithelial and mesenchymal markers, and high expression levels of epithelial markers in individual CTCs. The results reinforced the complex gene expression profiles and alterations seen in pooled CTCs. Secondly, a panel of DNA methylation markers, including those associated with EMP, was developed and tested in breast cancer cell lines, primary tumours and whole blood of normal controls to identify suitable markers for ctDNA detection. In a panel of breast cancer cell lines spanning the epithelial-mesenchymal spectrum, the majority of epithelial cell lines were methylated for cancer-associated markers (i.e., RASSF1A, RARß) and epithelial methylation-based markers (i.e. VIM, DKK3 and CRABP1). Mesenchymal cell lines were exclusively methylated for mesenchymal methylation-based markers (GRHL2, MIR200C and CDH1); however, the level of methylation was quite low. The methylation profiles of the studied genes classified primary tumours into intermediate phenotypes while few tumours were mesenchymal. In addition, MIR200C, RASSF1A, AKR1B1 and TWIST1 were methylated at high frequency in our cohort. Among these, RASSF1A and AKR1B1 showed no methylation in whole blood of normal controls, suggesting their potential use as markers for ctDNA detection from plasma of the breast cancer patients in our cohort. Preliminary experiments established ddPCR assays for these two markers, allowing further testing on patient cell-free DNA samples for the detection of ctDNA. The results of this thesis challenge the conventional model of EMT, where cells in epithelial tumours become mesenchymal, with associated migratory properties, and later re-epithelialise (MET) at a distant metastasis. Firstly, the complex and consistent alterations in the epithelial and mesenchymal markers in CTCs across the two models is suggestive of a ‘hybrid’ phenotype. The overall findings from the CTC work that CTCs were not as mesenchymal as expected also suggest that other processes than EMP directly influence the generation and survival of CTCs. Secondly, nearly all the examined breast tumours exhibited an intermediate rather than a strong epithelial phenotype based on the methylation profiles. This suggested that a plasticity is already present at the solid tumour state. These findings provide an alternative view of EMP in both primary breast cancer and the disseminated forms, and provide an important platform for further research in this field.
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ItemConstructing a neuromuscular-prosthetic interface and actuator system for limb reconstruction( 2017)A neuromuscular prosthesis provides an ideal solution to functional restoration of the limb after amputation. In such a system, the severed nerves at the stump are implanted into denervated muscles and the innervated muscle is coupled to a human-machine interface which detects the body's signals and transmits it to the actuator. This thesis will present studies that address key components of this bio-prosthetic actuator system, namely the neuromuscular junction, the muscle electrode junction and the actuator system. An in vitro nerve muscle co-culture system was established as a model platform for studying the neuromuscular junction. The effect of agrin and laminin on the innervation of muscle cells was studied with immunocytochemistry, real time PCR and liver cell imaging. Agrin and laminin were found to sensitize muscle cells to innervation by PC12 cells forming more functional neuromuscular junctions and promoting muscle maturation. An in vitro model of the neuromuscular prosthetic interface was created from PC12 neural and primary mice myoblasts grown on multi-electrode arrays. Electrodes of the array were further coated with a conducting polymer polypyrrole to create a low impedance interface between the muscle cells and the electrode. The effect of polypyrrole coating thickness on the quality of the cell recording was assessed. The thickness of polypyrrole coating had no impact on the strength of the cell recording. Finally biocompatibility studies were performed on trilayer polypyrrole based actuators. Trilayer polypyrrole based actuators are known for their superior work density compared to natural muscle and existing actuators. When implanted into rabbit muscle, actuators that had pores engineered into it to encourage tissue integration showed significant polypyrrole delamination from the actuator. The degradation was slowed by sealing the cut edges of the pores on the actuator with polypyrrole. The biocompatibility results provide valuable insight into required design upgrades to existing polypyrrole based trilayered actuators. The work presented in this thesis serve as a basis and platform for further studies in integrating the neuromuscular junction, muscle electrode junction and the actuator into one unit for future translation.
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ItemOptimising real-time surgical navigation interface design: enhancing spatial awareness while limiting distraction( 2017)Modern diagnostic imaging techniques allow surgeons to preoperatively scrutinise anatomy and pathology in an ever more precise fashion. Surgical planning may be honed to minimise access and perform targeted yet thorough procedures. Image guided surgery systems help integrate this valuable data into the operating theatre to aid spatial awareness during surgery. Advanced navigational displays show promise in providing surgeons with real-time image guidance presented in an intuitive manner during a procedure. In addition to standard 2-dimensional cross-sectional views, options for visual presentation include 3-dimensional virtual views and augmented reality. Live computer-assisted feedback may include a 3-dimensional visual representation of tool position and critical structure’s with proximity auditory alerts. Although orientation may be enhanced, additional stimuli inherently demand some attention from the operator. An ideal user interface would provide accurate and accessible navigational data with minimal distraction. This thesis explores, through pre-clinical navigation and dissection trials, ways to provide the potential benefits of surgical precision and efficiency while mitigating distracting cues. Cadaver dissection trials allowed structured testing of the new technology, which is directly compared to conventional systems. Real-time surgical navigation systems are shown to enhance spatial awareness while reducing task workload during complex endoscopic skull base surgery. These systems showed particular promise where there was high spatial demand, compromised visual landmarks and proximity to critical structures. World first surgical studies investigating the attentional cost of advanced navigational displays highlight the potential dangers of this technology. The concept of inattentional blindness is introduced into the surgical literature with experimental evidence of its existence. Augmented reality advanced navigational displays are shown to increase inattentional blindness with a reduced detection of unexpected but salient findings. The findings serve to caution injudicious adoption of this technology and reinforce the need for pre-clinical human factors testing.