Engineering

Publication Search Results

Now showing 1 - 10 of 25
  • (2021) Fu, Lu
    Thesis
    Angiogenesis is a vital step of cancer growth and metastasis and has become a promising target for cancer therapy. Due to the abnormal vascularisation and poor blood supply, tumours are usually hypoxic and lack nutrients, which triggers tumour aggressiveness and hampers efficient drug delivery. The success of traditional anti-angiogenic strategies, which focuses on decreasing the vascular supply to tumours, is limited by insufficient drug delivery or tumour resistance. Thus, vascular promotion therapy based on promoting angiogenesis is emerging as a complementary cancer therapy. Heparin is a linear negatively charged polysaccharide that, in addition to its anticoagulant action, can promote angiogenesis by potentiating angiogenic growth factors. Thus, heparin is a promising bioactive for cancer drug delivery applications. Cerium oxide nanoparticles (CNPs), a cytocompatible redox biomaterial, has gained attention as it actively scavenges reactive oxygen species (ROS) in biological systems which are related to cancer aggressiveness and tumour hypoxia. In this thesis, CNPs were functionalised with heparin with different chain lengths and density, aiming to investigate their mechanisms of interaction with the vasculature as blood vessel-targeting cancer therapeutics. CNPs were synthesised via precipitation and then functionalised with different amounts of unfractionated heparin (UFH) and low molecular weight heparin (LMWH) via an organosilane linker. CNPs and heparin-CNPs were approximately 12 nm in crystal size, exhibited face-centred cubic phase structure in morphology and as determined by high-resolution transmission electron microscopy (HRTEM) and X-ray diffraction (XRD), respectively. The successful functionalisation of CNPs with heparin was qualitatively verified by attenuated total reflectance-Fourier transform infra-red spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy (XPS). Additionally, thermogravimetric analysis (TGA) demonstrated the conjugation of heparin onto the surface of CNPs with approximately 1.5 μmol UFH, and either 1.8 μmol or 5.7 μmol LMWH conjugated per gram of CNPs. In addition, CNPs and heparin-CNPs showed a dose-dependent cytotoxicity in human umbilical vein endothelial cells (HUVECs), indicating that concentrations of heparin-CNPs above 5 μg/mL can act as anti-angiogenic agents. Heparin-CNPs at a dose of 1.5 μg/mL had limited redox activity which correlated with their limited effect on vascular endothelial growth factor receptor 2 (VEGFR2) expression, a key receptor in angiogenesis regulated via redox. However, the heparin-CNPs signalled fibroblast growth factor 2 (FGF-2) in vitro and in vivo, a key growth factor in angiogenesis, indicating their potential to support angiogenesis. In vivo analyses of the heparin-CNPs indicated that they dose-dependently promoted angiogenesis in the CAM assay at doses up to 1 mg/mL. Moreover, compared to CNPs, heparin-CNPs reduced intracellular ROS level in melanoma cells (MM200), reducing the hypoxic level of tumour cells, which indicated their potential as an anti-cancer agent. A human tumour model in the CAM was developed in this thesis to investigate the effects of nanoparticles on tumour angiogenesis and their interactions with the tumour vasculature. It was found that heparin-CNPs, compared to CNPs, dose-dependently promoted angiogenesis in the human tumour model in the CAM and exhibited a higher level of penetration into the tumour mass via light sheet microscopy and histological analysis. Overall, heparin-CNPs dose-dependently promoted angiogenesis by modulating key processes in angiogenesis and address tumour aggressiveness, showing their potential application in cancer therapeutics.

  • (2021) Li, Bitong
    Thesis
    In the past decade, the discovery of the CRISPR/Cas system launched a new era of genome editing and has rapidly become a universal research tool. The specific recognition between guide RNA and target nucleic acids enables CRISPR/Cas systems to exhibit high specificity compared to gene editing systems such as zinc finger or transcription activator-like effector nucleases. In this thesis, the CRISPR/Cas12a and CRISPR/Cas13a systems were designed to modify DNA or RNA associated with the HSPG2 gene which encodes perlecan, the major extracellular heparan sulphate (HS) proteoglycan in basement membranes. Perlecan has essential roles in organ development and contributes angiogenesis in pathological processes including cancer. The Cas12a and Cas13a nucleases were expressed in transformed E. coli and purified via His-tag affinity followed by size exclusion chromatography. The purified Cas12a nuclease exhibited high activity and specificity in a collateral activity assay in which the reporter sequence was only cleaved in the presence of the Cas12a protein and target ssDNA. Similarly, Cas13a nuclease exhibited high activity and specificity in a collateral activity assay with target ssRNA. A CRISPR/Cas12a gRNA was designed to target exon 2 of HSPG2 and was able to cleave amplified genomic DNA extracted from human melanoma cell line, MM200. Additionally, the collateral activity assay revealed that Cas12a nuclease dose-dependently cleaved the reporter ssDNA when used with target HSPG2 DNA. Similarly, a CRISPR/Cas13a gRNA was designed to target exon 2 of HSPG2 RNA and was able to cleave the target RNA extracted from MM200 cells and was active in the collateral activity assay when used with target HSPG2 RNA. Modification of HSPG2 nucleic acids in both MM200 cells and human umbilical vein endothelial cells (HUVECs) was also established. The CRISPR/Cas12a system resulted in up to 39 and 24 % reduction in HSPG2 gene expression in MM200 and HUVECs, respectively. Moreover, the CRISPR/Cas13a system achieved up to 69 and 99% reduction in HSPG2 RNA in MM200 and HUVECs, respectively. The HSPG2 mRNA modification in both MM200 and HUVECs resulted in decreased expression of FGF2 and VEGF-A, genes involved in the perlecan signalling pathway networks and associated with angiogenesis. The established CRISPR/Cas12a and CRISPR/Cas13a systems provide novel and efficient nucleic acid editing tools to further study the functions of perlecan in vitro and potentially in vivo. In addition, the LbCas12a or LwCas13a-based collateral cleavage assay enabled efficient and specific detection of the HSPG2 genome or transcripts, suggestive of its potential in perlecan-related disease diagnoses, such as cancer or genetic disorders.

  • (2022) Li, Yi
    Thesis
    CRISPR/Cas9-based gene editing is no doubt among the most intensively studied topics in bio-related fields in the recent decade, and certain new programmable Cas nucleases have been exploited recently for the development of a variety of biological tools far beyond gene editing, particularly for biosensor development. Although CRISPR/Cas-based biosensing has brought about a revolution in the area of nucleic acid diagnostic with their superior performances, its advantages were challenged when attempting to expand towards a broader range of non-nucleic acid targets. The currently reported methods for non-nucleic acid targets has successfully demonstrated the versality of CRISPR/Cas components in combining with other biosensing elements, however, combining these elements without proper optimization or controllable bioreaction environments could also bring additional variable factors into the system, hence potentially leading to compromised sensitivity or overall increase of system complexity. In this project, two novel CRISPR/Cas12a-based biosensing systems have been developed to realise simple, sensitive and universal non-nucleic acid detections. Both of these systems are established on a standard 96-well plate format, similar to the widely used ELISA diagnostic approach. The first system utilised an aptamer as its recognition molecule for rapid detection of two small proteins with fM-level sensitivity within 1.5 hours. In the second system, antibody was used as a recognition molecule, allowing to draw on a huge pool of commercially available antibodies to support its universality. This system exhibits ultra-high sensitivity down to 1 fg/mL (aM-level) for the detection of two protein targets. With these two successfully developed CRISPR/Cas12a-based non-nucleic acid biosensing systems, the universality and feasibility to deal with two practical bio-detection scenarios was investigated. The antibody-based system with minor modifications was directly used as a ready-to-use sensitivity enhancer onto a commercial IFN-γ ELISA kit, which resulted in a 2-log increase in sensitivity without changing its original protocol. Then, a similar modification strategy was used to re-direct the antibody-based system to detect whole pathogenic microorganism, Cryptosporidium parvum oocyst. Without the need for specialised instruments, the results show a successful detection of this pathogen with single oocyst sensitivity and capable of applying in challenging environmental samples. All these results serve as successful demonstration of the great potential in CRISPR/Cas-based biosensing technology to achieve affordable, translatable, and deployable solutions for various clinical, industrial and research diagnostic needs.

  • (2020) Armitage, Lucy
    Thesis
    Many people with transtibial amputation rate the fit between their residual limb and prosthetic socket as the most important factor in the use of their prosthesis. Current research suggests that poor fit between the residual limb and the prosthetic socket can cause pain, skin breakdown or alteration of gait. This thesis focused on measurement methods to assess residual limb – prosthetic socket fit in people with transtibial amputation. Three aspects of fit were examined and new measurement methods for each aspect were explored. Fit affects the amount of relative movement that occurs between the residual limb and prosthetic socket. A method to measure this movement was proposed using motion capture analysis. The development and attempted validation of the method were outlined. Results were unpredictable and subject to confounding factors such as marker movement artefact. Therefore, the method was deemed not to be appropriate as a fit measurement tool in its current form. Based on these findings it was deemed necessary to investigate other measures of fit Fluctuation in residual limb volume can also affect fit. The reliability and validity of current residual limb volume measurement techniques were examined in a systematic review. Results demonstrated that circumferential measurements are the most reliable and valid tool currently available. The results from this review informed a study investigating the use of a low-cost optical scanner as a clinical volume measurement tool. Results found that the scanner was reliable when used by the same assessor but was not valid. The loading state at the prosthetic socket-residual limb interface is also an important factor in assessment of fit. The effect of pressure feedback to the prosthetist on the design of prosthetic sockets was assessed. It found that feedback resulted in reduction in pressure over anatomical regions of concern, and improved consistency between prosthetists. A new sensor to measure pressure and shear at the socket-limb interface was developed and benchtop testing was performed. These methods have the potential to aid in an improved understanding of the mechanisms that affect fit at the socket-limb interface of people with transtibial amputation. It is hoped that this can lead to improved fit and comfort for prosthetic users.

  • (2020) Leong, Chin Neng
    Thesis
    Myocardial infarction (MI) is one of the diseases with the highest mortality rate. Following MI, myocardium experiences abrupt changes in its loading condition due to the presence of infarct. In response to such changes, myocytes undergo adaptations to maintain homeostasis. However, maladaptation can happen and lead to remodelling, in which the left ventricle (LV) gradually loses its function and eventually turns into heart failure. Nevertheless, the mechanisms underlying LV remodelling are still poorly understood. In this study, a generic LV model was developed, incorporating realistic fibre orientation and excitable contracting myocardium. It was demonstrated that the developed model is capable of reproducing physiological LV functions, including action potential propagation, LV pressure and cavity volume, LV twisting and wall thickening. Subsequently, the generic LV model was utilised to investigate the effects of the infarct state on LV regional mechanics, including the interaction between non-contractile infarct and contractile myocardium. It was found that infarct transmural extent (TME) is more important than infarct size in determining LV regional mechanics impairments. Neighbouring contractile myocardium and non-contractile infarct induce a mechanical tethering effect, which elevates with infarct TME, at the border zone (BZ). Such mechanical tethering causes the BZ to have high systolic fibre stress, elevated energy expenditure and reduced myocardial energy efficiency, which are believed to give rise to infarct extension. The generic LV model was then modified into a patient-specific model, incorporating patient-specific infarcted LV geometry and optimised regional material properties, to study the correlation between infarct extension and myocardial mechanics impairments, including the underlying mechanisms responsible for the impairments. Among the observed myocardial mechanics impairments, only the depressed myocardial energy efficiency was found to be correlated with infarct extension. The depressed myocardial energy efficiency was due to inadequate generation of contraction force, which, at least in part, owing to inadequate stretching of myocardium at end-diastole (the Frank-Starling law). Although a stiff infarct can prevent infarct expansion, results of this study showed that it can also cause the neighbouring myocardium to be under-stretched at end-diastole, thereby depressing the generated contraction force and energy efficiency during systole, which were found to be correlated with infarct extension of the neighbouring myocardium.

  • (2022) Tang, Junma
    Thesis
    Converting natural resources or greenhouse gases into value-added species with low carbon footprint, is essential for the development and sustainability of modern society. However, the goal for sustainable and cost-effective conversion by using many current technologies, including photo-, electro- and thermal-based catalytic reaction systems, has been largely underachieved. Hence, it is a necessity to explore and develop new approaches to fulfill this objective. In this thesis, three hybrid catalytic systems, containing liquid gallium (Ga) and solid materials as co-catalysts, are demonstrated, which realize the gaseous and liquid feedstocks conversion through nano-tribo-electrochemical reaction pathways. In the first stage of this PhD thesis, the author reports a green carbon capture and conversion technology for mitigating CO2 emissions. The technology uses suspensions of Ga liquid metal to reduce CO2 into solid carbonaceous products and O2 at near room temperature. The solid co-contributor of silver-Ga rods ensures a cyclic sustainable process. The overall process relies on mechanical energy as the input, which drives nano dimensional triboelectrochemical reactions. In the next stage, for the gaseous feedstock conversion, the author demonstrates an approach based on Ga liquid metal droplets and Ni(OH)2 co-catalysts for CH4 conversion into H2 and carbon. Mainly driven by the triboelectric voltage, originating from the joint contributions of the co-catalysts during agitation, CH4 is converted at the Ga and Ni(OH)2 interfaces. The efficiency of the system is enhanced when the reaction is performed at an increased pressure. The dehydrogenation of other non-gaseous hydrocarbons using this approach is also demonstrated. In the final stage, the author explores and realizes the liquid biofuels conversion, including canola oil and other liquid hydrocarbons, with H2 and C2H4 as the main products by employing Ga and nickel particles as the co-catalysts and mechanical energy as the stimulus. Altogether, the work of this PhD research offers novel pathways for low energy and green conversion of gaseous and liquid feedstocks that can be implemented in large scale conversion systems of the future.

  • (2022) Zhang, Chengchen
    Thesis
    Liquid metals (LMs) are a class of metals and their alloys which have low melting points near or below room temperature, and they are mainly composed of post-transition elements. The low melting points of LMs make them easily stay in a liquid state and readily be broken into tens or hundreds of nanometers, which are called LM nanoparticles (LMNPs). In this thesis, the author investigates LMNPs for three exciting applications of creating conductive polymer-LMNPs compositions and explores the potential utilization of LMNPs in biological applications. In the first phase of this research, the author develops nanocomposites of Ga-based LMNPs (EGaIn NPs) with conductive polymer polyaniline (PANI). This work reports a method of growing PANI nanofibers on the EGaIn NPs by firstly providing initial functionalization sites at the interfaces for the formation of PANI nanofibrous network. The nanocomposites provide synergistic effects of PANI nanofibers and EGaIn NPs for the applications of environmental sensing and molecular separation. In the second phase of the research, the author focused on the exploration of LMNPs for their anti-inflammatory applications. Considering that Ga ions (Ga3+), have been historically utilized as anti-inflammatory agents by interfering with the Fe homeostasis of immune cells. The study presents the anti-inflammatory effects of Ga by delivering Ga nanoparticles (Ga NPs) into lipopolysaccharide-induced macrophages. The Ga NPs show a selective anti-inflammatory effect by modulating nitric oxide production without disturbing other pro-inflammatory mediators. This work reveals the different anti-inflammatory effects between Ga NPs and Ga3+ come from their different endocytic pathways: transferrin receptor independent and dependent endocytosis for Ga NPs and Ga3+, respectively. In the final phase, the author studies the interactions between LMNPs and macrophages at a light microscopic level. The mechanistic responses of macrophages to LMNPs with different densities were observed, in comparison to some other commonly studied nanoparticles. This work discovers the mobility of macrophages is very much density-dependent. This thesis comprehensively studies the interactions between LMNPs and polymeric and biological systems, at both molecular and microscopic levels, which provides a basis and road map for utilizing LMNPs in various fields such as electronics and biomedical engineering.

  • (2022) Cao, Jun
    Thesis
    This thesis focuses on the development and applications of magnetic resonance electrical properties tomography (MREPT), which is an emerging imaging modality to noninvasively obtain the electrical properties of tissues, such as conductivity and permittivity. Chapter 2 describes the general information about human research ethics, MRI scanner, MR sequence and the method of phase-based MREPT implemented in this thesis. Chapter 3 examines the repeatability of phase-based MREPT in the brain conductivity measurement using balanced fast field echo (bFFE) and turbo spin echo (TSE) sequences, and investigate the effects of compressed SENSE, whole-head B_1 shimming and video watching during scan on the measurement precision. Chapter 4 investigates the conductivity signal in response to short-duration visual stimulus, compares the signal and functional activation pathway with that of BOLD, and tests the consistency of functional conductivity imaging (funCI) with visual stimulation across participants. Chapter 5 extends the use of functional conductivity imaging to somatosensory stimulation and trigeminal nerve stimulation to evaluate the consistency of functional conductivity activation across different types of stimuli. In addition, visual adaptation experiment is performed to test if the repetition suppression effect can be observed using funCI. Chapter 6 explores if resting state conductivity networks can be reliably constructed using resting state funCI, evaluates the consistency of persistent homology architectures, and compares the links between nodes in the whole brain. Chapter 7 investigates the feasibility of prostate conductivity imaging using MREPT, and distinctive features in the conductivity distribution between healthy participants and participants with suspected abnormalities.

  • (2022) Kaur, Jagjit
    Thesis
    Secreted by pancreatic β-cells, insulin is the major anabolic hormone, regulating the metabolism of fats, proteins, and carbohydrates. Defects in insulin production or action can lead to diabetes characterized by derangements in glucose handling and metabolic disease. Diabetes affects 420 million people worldwide, increasing morbidity, mortality and placing a burden on healthcare of nations. There is a need for rapid and accurate monitoring of insulin levels to optimize diabetes management and facilitate early diagnosis of insulin related chronic diseases. Conventional strategies such as HPLC, MALDI-TOF, ELISA, etc. used for insulin detection are not suitable for point-of-care testing (POCT) as they are expensive, and require sample preparation, sophisticated instruments, and skilled personnel. Our goal was to develop techniques to allow POCT for insulin in real time. In this study we developed two lateral flow assays (LFAs) based POCT platforms using aptamers as the biorecognition molecules for colorimetric and fluorescent detection of insulin. A range of conditions were tested such as concentrations of aptamers, reporter molecules used, volume of sample required, and assay time to obtain quantify insulin levels using a standard LFA reader. The colorimetric LFAs had linear detection range of 0.01-1 ng.mL-1 and LOD of 0.01 ng.mL-1. The fluorescent LFAs exhibited a linear detection range of 0-4 ng.mL-1 and 0.1 ng.mL-1 LOD. Various signal amplification strategies were incorporated, ie., gold-silver amplification technique and rolling circular amplification (RCA) to further enhance the signal. The developed colorimetric LFAs were successfully used for insulin quantification in rat blood, human blood, and human saliva samples. Although insulin levels were quantified within 12 min, some issues arose such as coagulation, need for dilution, and non-uniform flow through the test strips. Further work is required to optimize blood handling to progress an insulin POCT in real time. Future work could develop a multiplexed strip for detection of different analytes such as HbA1c, glucose, and C-peptide for better management of diabetes, along with a smartphone reader App. This research goes some way to addressing the challenge of providing a reliable and rapid approach for highly sensitive and specific detection of insulin for POCT applications.

  • (2021) Muralidharan, Madhuvanthi
    Thesis
    Retinal implants aim to provide artificial vision to those profoundly blind by stimulating the residual network to elicit visual percepts. While human clinical trials have demonstrated encouraging results including the presence of visual percepts as well as partial visual restoration, the vision quality provided remains limited. One potential cause of this poor performance has been attributed to the indiscriminate activation of functionally-different retinal ganglion cell (RGC) types. To combat this problem, a promising strategy has been to design stimulation strategies that are capable of selectively, or preferentially, activating different cell types. One such approach to realise this goal has been through the use of high frequency stimulation (HFS) which was shown to be effective in preferentially activating two major retinal ganglion cell types– ON and OFF. While encouraging, the utility of the technique to target a broader range of cell types, and under different stimulation conditions and environments was still unclear. The studies presented in this thesis were designed to improve the understanding of HFS based preferential activation. Using in vitro whole-cell patch clamp of RGCs in mice (C57BL/6J and rd1), an investigation into whether HFS could be used to preferentially activate four major RGC types namely, ON-sustained (ONS), ON-transient (ONT), OFF-sustained (OFFS), and OFF-transient (OFFT), was undertaken. Results suggested that three of the four targeted cell types could be preferentially activated against the remaining population. A subsequent study documented the responses and the preferential activation capabilities of the aforementioned cell types when the high frequencies were modulated with short stimulation bursts, varying sequence orders and in a continuous waveform. It was shown that the ON (sustained and transient) RGCs typically exhibited more consistent responses and preferential activation regions irrespective of the frequency order, or when presented as a continuous waveform. A final study examined the responses of rd1 ON and OFF RGCs to HFS both with and without the presynaptic degenerate network. The network did appear to have an effect on the HFS evoked responses, and particularly increased the variability of the responses which in turn affected the preferential activation of the cell types. Additionally, a comparison into the specific intrinsic properties between the rd1 and healthy RGCs found that these properties may differ between the cell groups. Overall, this thesis investigated the usefulness of HFS to preferentially activate different cell types and across various stimulation conditions and environments and found that HFS remains a viable stimulation technique to reduce indiscriminate activation of functionally-distinct cell types.