To build an Functional Electrical Stimulation (FES) system that assists individuals with spinal cord injury (SCI) to improve sitting balance, one needs to determine multidirectional stiffness of the trunk during sitting and demonstrates that this stiffness can be influenced by FES. Therefore, the purposes of this study were: (1) to develop a novel protocol to quantify and identify the multidirectional stiffness of the trunk during sitting; and (2) to test the effect of FES on the multidirectional stiffness of the trunk. 15 able-bodied subjects were recruited. Each subject was perturbed in 8 different directions using 8 linear motors during quiet sitting with and without FES applied to his front/back trunk muscles. Stiffness in the lateral directions was the highest and FES increased stiffness of the trunk in the anterior/posterior direction. These results can be used to develop an FES protocol for individuals with SCI to improve their sitting balance.

To build an Functional Electrical Stimulation (FES) system that assists individuals with spinal cord injury (SCI) to improve sitting balance, one needs to determine multidirectional stiffness of the trunk during sitting and demonstrates that this stiffness can be influenced by FES. Therefore, the purposes of this study were: (1) to develop a novel protocol to quantify and identify the multidirectional stiffness of the trunk during sitting; and (2) to test the effect of FES on the multidirectional stiffness of the trunk. 15 able-bodied subjects were recruited. Each subject was perturbed in 8 different directions using 8 linear motors during quiet sitting with and without FES applied to his front/back trunk muscles. Stiffness in the lateral directions was the highest and FES increased stiffness of the trunk in the anterior/posterior direction. These results can be used to develop an FES protocol for individuals with SCI to improve their sitting balance.

Loss of the superficial zone of articular cartilage is an early change in osteoarthritis and with disease progression the deep zone (DZ) of cartilage shows progressive mineralization. To date, the mechanism(s) regulating post-natal articular cartilage mineralization is poorly understood. Previously, we have shown that inorganic polyphosphate inhibits mineralization of in vitro-formed DZ cartilage. We developed an indirect co-culture method to investigate the effect of superficial zone chondrocytes (SZC) on mineralization in DZ cartilage. Our findings suggest that SZC suppress mineralization by modulating polyphosphate levels in DZ cartilage via FGF-18. Furthermore, SZC promote glycosaminoglycan and collagen accumulation in the extracellular matrix of cartilage formed by DZ chondrocytes. This study provides insight into the interaction between chondrocyte subpopulations and possible mechanism(s) controlling post-natal articular cartilage mineralization. Moreover, the results of this study establish polyphosphate and FGF-18, separately or in combination, as therapeutic candidates for articular cartilage repair and osteoarthritis prevention.

Recent evidence suggests that “mesenchymal stem cells” (MSCs) are resident in the perivascular compartment of connective tissues. However, since the definition of a stem cell assumes that these progenitors have clonal self-renewal and multi-lineage differentiation potential, the term “MSC” has been criticised, as it has been impossible to isolate definitive clonally derived “MSCs.” To test for this most basic definition of a stem cell, here it is shown that human umbilical cord perivascular cells (HUCPVCs) are capable of multilineage differentiation in vitro and, more importantly, in vivo, displaying the ability to differentiate into functionally synthetic cells that direct and contribute to rapid connective tissue healing by producing bone, cartilage and fibrous stroma in a mouse injury model. Uniquely, these cells can be enriched to >1:3 clonogenic frequency in early passage culture, making it possible to isolate clones and daughter sub-clones from mixed gender suspensions, determined to be definitively single-cell-derived by Y-chromosome fluorescent in situ hybridization (FISH) analysis. Each clone was assayed for multi-lineage differentiation capacity into the five mesenchymal lineages: myogenic, adipogenic, chondrogenic, osteogenic and fibroblastic (stroma). The observation that daughter sub-clones possess equal or lesser differentiative potential to their respective parent clones demonstrated the two intrinsic properties of stem cells in vitro: clonal self-renewal and multi-lineage differentiation. This evidence provides a new hierarchical structure of robust MSCs self-renewing to produce more restricted progenitors that gradually lose differentiation potential until a state of complete restriction to the fibroblast is reached. The methods described herein combined with recognition of this lineage hierarchy provides a significant advance to the understanding of MSC biology, and will enable interrogation of the properties of robust self-renewal and differentiation of MSCs in serially transplanted living recipients.

The emergence of bionanotechnology has allowed the design of novel technological tools for a variety of biomedical applications. Despite the incompatibility of native semiconductor quantum dots with biological environment, this class of nanocrystals was among the first nanomaterials to demonstrate their use in biological labeling and imaging applications. In this thesis, new surface modification chemistry and synthetic strategies were developed to produce high quality biocompatible and bioconjugatable near-infrared emitting quantum dots for deep tissue in vivo imaging and detection applications. By carefully selecting a specific mixture of semiconductor elements to obtain a desired bandgap energy and optimizing the procedure for surface coating, successful synthesis of high quality, watersoluble, near-infrared emitting quantum dots were demonstrated. These developments allows for the use of quantum dots as alternative contrast agents for sophisticated biological imaging applications that are currently unachievable using conventional uorophores. In addition, using metallic nanoparticles, it was found that cells possess the ability to differentiate nanoparticles of various sizes upon their binding with specific membrane receptors. These receptors undergo rapid cellular internalization which altered their trafficking dynamics and down-stream signaling processes. The amplitude of such alteration was highly dependent on the size of the nanoparticle with most efficient internalization occurring at 40 nm - 50 nm size range. These observations raise important questions regarding the mechanisms governing similar processes and cell behaviours documented during viral infections. Whether such biological phenomenon are evolutionarily conserved as natural defense mechanisms to counter foreign invasion, or whether many of the known viruses are naturally selected to breach the primary defense of cells - the plasma membrane, remains to be elucidated. In summary, nanotechnology offers great promises for biological research and medicine. This thesis demonstrates the use of semiconductor and metallic nanostructures for imaging, detecting and administrating therapeutics in cancerous cells, tissues and animal models. Although the results presented in this thesis are preliminary, and the technologies demonstrated are still years away from practical use, these studies nevertheless, pave the way for future experimental researches within the field of nanomedicine, and provide insights into the understanding of the most fundamental yet highly complex processes in cell biology.

Recent advances in the field of bionanotechnology have enabled researchers to design a variety of tools to detect, image and monitor biological process in cells. Despite this progress, the limited understanding of nanomaterial-cellular interactions has hindered the widespread use of these nanomaterials in biological systems. In this thesis, we examined the potential effects of metallic nanoparticle geometry on important cellular processes such as membrane trafficking, intracellular transport and subcellular signalling. We found that the size of nanoparticles plays an important role on their ability to interact with the cell surface receptors thus dictating their subsequent ability to activate intracellular signalling cascades. Interestingly, trafficking of these nanoparticles was dependent on their size due to biochemical and thermodynamical constraints. These findings suggest that nanomaterials actively interact with biological systems, thus, directly modulating vital cellular processes. In addition, by utilizing various physical and chemical properties of nanomaterials, we developed a novel class of hybrid nanoscaled carrier systems capable of delivering semiconductor quantum dots (QDs) into live cells without inducing membrane damage. Using biodegradable polymeric nanoparticles, bioconjugated QDs were encapsulated and delivered into trafficking vesicles of live cells. The environmentally sensitive surface charge of the polymeric nanoparticles exhibited positive zeta potential inside acidic endo-lysosomes, thus enabling their escape from the vesicular sequestration into the cytosol. Hydrolytic-induced degradation then releases the bioconjugate QDs for active labelling of subcellular structures for real-time studies. Unlike previously described intracellular QD delivery methods, the proposed system offers an efficient way to non-invasively deliver bioconjugated QDs without inducing cell damage, enabling researchers to accurately monitor cellular processes in real-time. The understanding of both physical and chemical properties of nanomaterials is crucial to the design of biocompatible nanosystems to study fundamental processes in biological systems. Here, we demonstrated that both the size and surface chemistry of nanoparticles can be modified to obtain desired biological responses. Future experimental efforts to study other physical and chemical properties could allow the development of more sophisticated and effective platforms for biological applications.

In clinical oncology and experimental therapeutics, changes in tumour growth rate or volume have been traditionally the first indication of treatment response. These changes typically occur late in the course of therapy. Currently, no routinely available imaging modality is capable of assessing tumour response to cancer treatment within hours or days after delivery of radiation treatment. Therefore, the goal of this thesis is to develop the use of ultrasound imaging and ultrasound characterization methods with frequencies of 10 to 30 MHz to assess non-invasively tumour response to radiotherapy, early, within hours to days after delivery of radiotherapy. Responses to radiotherapy were characterized initially in vitro in a well-controlled environment using cell samples. It was demonstrated experimentally that the changes in ultrasound integrated backscatter and spectral slopes were the direct consequences of cell and nuclear morphological changes associated with cell death. The research in vitro provided a basis for the in vivo research that characterized responses to radiotherapy in cancer mouse models. The results from mouse tumour models indicated that quantitative ultrasound could detect the regions in a tumour that corresponded in histology to areas of cell death. In order to understand the cellular morphological changes responsible for ultrasound scattering at these frequencies and assist in the interpretation of experimental data, numerical simulations of ultrasound scattering from four different cell lines exposed to radiotherapy were conducted and compared to experimental results. It was concluded that the increases measured in ultrasound backscatter could in part be explained by the increase in the randomization of cell nuclei resulting from the increase in the variance of cell sizes following cell death. In this thesis, it is demonstrated that ultrasound imaging and quantitative ultrasound methods were able to detect non-invasively early responses to radiotherapy in vitro and in vivo. The mechanism behind this detection was linked to changes in the acoustic properties of nuclei and changes in the spatial organization of cells and nuclei following cell death. This provides the groundwork for future investigations regarding the use of ultrasound in cancer patients to individualize treatments non-invasively based on their responses to specific interventions.

This thesis presents the development of a digital microfluidic system to achieve automated sample preparation for the vitrification of mammalian embryos for clinical in vitro fertilization (IVF) applications. This platform included micro devices fabrication, an imaging system, a high voltage control system, and a LabVIEW interface. Individual micro droplets manipulated on the digital microfluidic device were used as micro-vessels to transport a single embryo through a complete vitrification procedure. The device showed cell survival and development rates of 77% and 90%, respectively, which are comparable to the control groups that were manually processed. Technical advantages of this approach, compared to manual operation and channel-based microfluidic vitrification, include automated operation, cryoprotectant concentration gradient generation, and feasibility of loading and retrieval of embryos.

Topographical orientation is the ability to orient oneself within the environment and to navigate through it to specific destinations. Topographical disorientation (TD) refers to deficits in orientation and navigation in the real environment, and is a common sequela of brain injuries. People with TD often have difficulties interacting with and perceiving the surrounding environment. The literature suggests that patients with TD are likely to benefit from research leading to clinical standards of practice and technology to facilitate topographical orientation. In the light of the above, the objectives of this thesis were to investigate methods of realizing a context-aware, wearable mediated environment system for indoor navigation, and to develop a standard method of quantifying the impact of such a system on indoor navigation task performance. In realizing these objectives, we first conducted an extensive literature review of in-door localization systems. This review served to identify potential technologies for an indoor, in-situ wayfinding assistive device. Subsequently, an automated navigation algorithm was designed. Our algorithm reduced the navigational effort of simulated patients with topographical disorientation while accounting for the physical abilities of the patient, environmental barriers and dynamic building changes. We introduced and demonstrated a novel energy-based wayfinding metric, which is independent of route complexity. An experiment was conducted to identify preferred graphical navigation tools for mediated reality wayfinding guidance. Different combinations of spatial knowledge, graphical presentations and reference frames were considered in the experiment. The data suggested that the locator and minimap are the preferred navigational tools. Two unique optical-inertial localization systems for real-time indoor human tracking were created. The first localization system was oriented to pedestrians, while the second was implemented on a wheelchair. Empirical tests produced localization accuracies comparable to those reported in literature. Finally, a fully operational mediated reality location aware system for indoor navigation was realized. Tests with human participants indicated a significant reduction in physical effort in comparison to the no-tool condition, during wayfinding tasks in an unfamiliar indoor environment. Collectively, the findings and developments of this thesis lay the foundation for future research on wearable, location-based navigational assistance for individuals with wayfinding difficulties.

The spine is the most common location of metastatic disease in the skeleton. The occurrence of bone metastasis can lead to severe clinical consequences and a significant decline in quality of life. The evaluation of metastatic disease in the spine has to date been mainly qualitative. More widespread access to multiple imaging modalities has motivated the development of 3D methods to quantitatively evaluate metastatic disease in the spine. Quantitative evaluation is important both in assessing stability of the metastatic spine and the progression/ response of the tumour and bone to treatment over time. Previous studies quantifying stability in the metastatic spine have focused primarily on osteolytic tumours. Local and systemic treatments have impacted the nature of vertebral metastasis, increasing the occurrence of mixed osteolytic and osteoblastic disease. Thus, it is important to focus analyses on models able to accurately represent diverse distribution patterns found in bony metastasis. Preclinical models are widely used in studying the process of metastasis and are able to represent both osteolytic and osteoblastic disease. This proposal aims to establish the biomechanical implications of metastatic disease in the spine through the evaluation of stability and tumour burden in a preclinical model using a multifaceted engineering-based approach. It is hypothesized that the use of automated analysis techniques applied to multimodality imaging will allow quantification of the impact of metastasis on biomechanical stability, tumour burden and bony architecture in the spine, and motivate prediction models that accurately reflect vertebral integrity in both osteolytic and mixed osteolytic/osteoblastic models of spinal metastasis. Specifically, this work aims to: 1) Utilize and compare μMR and μCT based radiologic methods to quantify tumour involvement and vertebral architecture in a rat model of spinal metastasis; and 2) Evaluate the ability of 2D, 3D, and continuum based methods to quantify structural integrity in vertebral metastasis. Overall, this work will focus on developing automated methods to quantify stereologic parameters, and quality in the metastatic spine and the evaluation of stability measures from 2D structural rigidity, Finite Element analysis, image registration and experimental methods. Ultimately this work will yield automated analysis techniques and evaluate the abilities of these methods to predict failure in metastatic vertebrae.