New mathematical models of inert gas transport through biological tissue in hyperbaric environments

The thesis is concerned with a fundamental mathematical analysis of inert gas transport through biological tissue at a raised ambient partial pressure. Three basic time-scales of transport in tissue are defined and their relationship examined and compared with existing models, which e.re shown to be usually inadequate in one or more ways. As a result three new mathematical models are proposed and solved both asymptotically and numerically. The first is applied to experimental data for non-perfused tissue which yields an improved value of the intracellular diffusion coefficient for nitrogen. An expression is also derived which should be useful in evaluating this constant and the volume fraction of extracellular fluid. The second embraces a number of current models and is applicable to perfused tissue. It should be useful in interpreting inert gas uptake curves. The model is applied to experimental data, and a source of possible error is discovered in using experimental non-asymptotic time constants. The third is a model which claims to resolve the controversy between the diffusion and perfusion theories of gas transport in tissue. The result is that in the large, diffusion is more important than perfusion, except in muscle tissue where they interact. Three different methods of numerical inversion of the Laplace Transform are compared and one is shown to be the most useful for solving gas uptake problems. The main result of the thesis is a contribution to the establishment of a mathematical basis for gas transport in various situations in the biological sphere

PET/MR imaging of hypoxic atherosclerotic plaque using 64Cu-ATSM

ABSTRACT OF THE DISSERTATION PET/MR Imaging of Hypoxic Atherosclerotic Plaque Using 64Cu-ATSM by Xingyu Nie Doctor of Philosophy in Biomedical Engineering Washington University in St. Louis, 2017 Professor Pamela K. Woodard, Chair Professor Suzanne Lapi, Co-Chair It is important to accurately identify the factors involved in the progression of atherosclerosis because advanced atherosclerotic lesions are prone to rupture, leading to disability or death. Hypoxic areas have been known to be present in human atherosclerotic lesions, and lesion progression is associated with the formation of lipid-loaded macrophages and increased local inflammation which are potential major factors in the formation of vulnerable plaque. This dissertation work represents a comprehensive investigation of non-invasive identification of hypoxic atherosclerotic plaque in animal models and human subjects using the PET hypoxia imaging agent 64Cu-ATSM. We first demonstrated the feasibility of 64Cu-ATSM for the identification of hypoxic atherosclerotic plaque and evaluated the relative effects of diet and genetics on hypoxia progression in atherosclerotic plaque in a genetically-altered mouse model. We then fully validated the feasibility of using 64Cu-ATSM to image the extent of hypoxia in a rabbit model with atherosclerotic-like plaque using a simultaneous PET-MR system. We also proceeded with a pilot clinical trial to determine whether 64Cu-ATSM MR/PET scanning is capable of detecting hypoxic carotid atherosclerosis in human subjects. In order to improve the 64Cu-ATSM PET image quality, we investigated the Siemens HD (high-definition) PET software and 4 partial volume correction methods to correct for partial volume effects. In addition, we incorporated the attenuation effect of the carotid surface coil into the MR attenuation correction _-map to correct for photon attention. In the long term, this imaging strategy has the potential to help identify patients at risk for cardiovascular events, guide therapy, and add to the understanding of plaque biology in human patients

Modeling Vascular Diffusion of Oxygen in Breast Cancer

Oxygen is a vital nutrient necessary for tumor cells to survive and proliferate. Oxygen is diffused from our blood vessels into the tissue, where it is consumed by our cells. This process can be modeled by partial differential equations with sinks and sources. This project focuses on adding an oxygen diffusion module to an existing 3D agent-based model of breast cancer developed in Dr. Norton’s lab. The mathematical diffusion module added to an existing agent-based model (ABM) includes deriving the 1-dimensional and multi-dimensional diffusion equations, implementing 2D and 3D oxygen diffusion models into the ABM, and numerically evaluating those equations using the Finite Difference Method. I started by diffusing a point source in a 2D grid, then diffusing a line in a 2D grid, then a cubic patch in a 3D grid, and finally, diffusing oxygen from blood vessels into the tissue. Then, I programmed the supply function to represent the continuous oxygen supply from vasculature, and the uptake function to represent the oxygen uptake by cancer cells

The ontogeny of cardiorespiratory support of metabolism

The formation of the cardiovascular system and its role in gas exchange has long been speculated to occur concomitantly. Although this premise has been suggested and quoted for more than a century, there are few studies to date which have attempted to validate these claims. Furthermore, the few which do exist have been primarily concerned with two things: first, how environment may affect the functional morphology; and second, how these changes may affect hemodynamics directly. As a result, our understanding of how gas exchange is coupled with to cardiovascular function is seriously lacking; The goal of this dissertation was to understand how, or if, the cardiovascular and respiratory systems coordinate function during development. Populations of amphibians were reared in various environments which included carbon monoxide (CO). This allowed for direct assessment of the efficacy of Hb in bulk O{dollar}\sb2{dollar} transport. The results indicated that CO, and the subsequent elimination of Hb function, had few ill effects on either aerobic or anaerobic metabolism. Further, the data indicated that cardiovascular function was mildly elevated. A separate study, set out to determine what factors limit overall O{dollar}\sb2{dollar} transport in developing embryos by limiting the available quantities of gas and eliminating Hb function. The results indicated that aerobic metabolism was unaffected in all populations. Cardiovascular function was mildly elevated, but only in populations of animals exposed to CO. Finally, gas exchange was modeled in developing embryos to determine the role of diffusion and plasma transport in overall O{dollar}\sb2{dollar} uptake. Calculations of maximal O{dollar}\sb2{dollar} flux indicate that diffusion would allow for enough gas to be exchanged to support aerobic metabolism in early life. Moreover, the role of plasma transport was considered in addition to diffusion, it be came clear that their combined transport would be adequate to support metabolism for animals in late life; A separate study evaluated how body composition changes with progressive development. From this it was determined that amphibians are unlike their fish counterparts in composition. When the total energy pool available for growth and development was calculated and compared with aerobic metabolism, it was shown that energy was not a limiting resource during development; Collectively these data indicated that Hb was not essential of O{dollar}\sb2{dollar} uptake and that the cardiovascular system as a whole may play a reduced role in total O{dollar}\sb2{dollar} turnover. Furthermore, the data indicates that neither diffusion nor perfusion limits total gas exchange. In addition, the model indicate that diffusion may be a viable mean to obtain O{dollar}\sb2{dollar} early in development, and that late in development convection of plasma coupled with diffusion could support metabolism. Finally, the body composition work indicate that resource limitations were not set by available substrate

Oxygen consumption of bone marrow and comb tissue of capons and normal chickens and the in vitro effect of androgens

Call number: LD2668 .T4 1954 A4Master of Scienc

Myocardial slices as an in vitro platform to study cardiac disease

In vitro models are the pillars of fundamental research and drug discovery, offering reductionist methods to better understand cellular responses in isolation. Often these methods are oversimplified, which makes their relevance to human biology and clinical translation ambiguous. Living myocardial slices (LMSLMSs) are viable thin (200-400μm) cardiac tissue slices, with preserved native multicellularity, architecture, mechanical and electrophysiological responses. Recent development in their culture, by us and others, paved the way for long-term preservation of adult mammalian heart tissue in vitro, without significant changes in its function and structure. This model has been extensively used in healthy tissue; however, to date, there are no established pathological models to study disease progression in vitro. Here we hypothesised that LMSLMSs can be used as an informative in vitro disease model to study temporal and spatial changes in cardiac function/structure in response to local cardiac damage. Before inducing cardiac damage, we further improved and characterised the cultured LMS model by designing robust tissue holders, optimising the oxygenation of the media, and establishing the best slice thickness (300μ) for oxygen diffusion and tissue stability in culture. We found that the LMSLMSs were adequately oxygenated in the inner layers and responded to mechanical stimuli with an increase in their contraction and hyperpolarisation of the mitochondrial membrane. We then developed a cryoinjury model, by applying a cooled probe on the LMSLMSs. We found that injury created a distinct necrotic area, surrounded by a border zone (BZ). The injury resulted in preserved force but electrical instability, with the presence of spontaneous contractions. Microscopic analysis of the BZ showed the presence of high numbers of spontaneous Ca2+ sparks, which could be affected by inhibiting the activation of Ca2+/calmodulin-dependent protein kinase II (CamKII). The inhibitory effect was more pronounced in endocardial LMSLMSs, showing transmural differences of CamKII under pathological conditions. Structural analysis of the BZ also showed an acute increase of the sarcomere length and loss of t-tubule density upon culture, that could also account for the arrhythmogenicity of the injured LMSLMSs. One application of therapeutic interventions on the model, by using extracellular vesicles (EVs), did not show any functional or molecular improvements. This thesis demonstrates the significance of using diseased LMSLMSs to study the way that local injury affects tissue stability, function, and structure. Further work is required to better understand the link between spontaneous Ca2+ and contraction events, as well as finding successful therapeutic interventions.Open Acces

Cardiopulmonary exercise testing in the assessment and treatment of young people with cystic fibrosis

Cystic fibrosis (CF) is the most common, genetically inherited, life-shortening condition in the Caucasian population, with ~11,000 people in the United Kingdom having the disease. The genetic defect responsible for CF results in accumulation of thick, sticky mucus that blocks the airways and digestive systems. As there is currently no cure for CF, it is a disease that is managed using antibiotics, nutrition, physiotherapy and exercise. Exercise capacity, as measured by peak oxygen uptake (V̇O2peak), and where possible, maximal oxygen uptake (V̇O2max), is reduced in patients with CF and a low V̇O2peak is associated with increased risk of hospitalisation, mortality and low quality of life. As a result, regular exercise testing is recommended, with cardiopulmonary exercise testing (CPET) considered the ‘gold standard’ procedure by leading international clinical organisations. The purpose of this thesis was to further our understanding surrounding the use of CPET in the assessment and treatment of children and adolescents with CF. The first component of this thesis sought to identify and evaluate submaximal parameters of aerobic function derived from CPET, namely the oxygen uptake efficiency slope (OUES) and plateau (OUEP). Findings revealed that allometric scaling for body surface area (BSA) was necessary when evaluating OUES, and a power function of 1.40 (i.e. OUES/BSA1.40) removed residual effects of body size (Chapter 4). Subsequently, results identified that the OUES was not a valid surrogate of aerobic fitness in CF, despite a significant correlation (r = 0.47, p = 0.004) with V̇O2max when expressed relative to body mass, as it was unable to discriminate aerobic fitness within a CF group, nor against a control group (Chapter 5). As OUES was not a valid surrogate of aerobic fitness, the utility of OUEP as an independent marker of aerobic fitness was explored. Whilst the OUEP was correlated with V̇O2peak in CF, when expressed as an absolute value (r = 0.43, p = 0.010) and when allometrically scaled for body mass (r = 0.52, p = 0.001), it was unable to discriminate aerobic fitness to the same extent as V̇O2peak. However, the OUEP was associated with disease status and severity, being significantly (p < 0.001) lower in the CF group, but also significantly and positively correlated with lung function (forced expiratory volume in one-second [FEV1]) in the CF group (r = 0.43, p = 0.010), a finding that warrants further, longitudinal investigation (Chapter 6). The second component of this thesis utilised CPET to investigate musculoskeletal limitations to the reduced V̇O2max that has previously been reported in CF. Parameters of muscle size (thigh cross-sectional area, muscle cross-sectional area and thigh muscle volume) were first quantified using magnetic resonance imaging, alongside the error associated with estimating muscle volume using alternative calculation techniques (Chapter 7). These parameters were then allometrically scaled for, which successfully removes residual effects of muscle size (i.e. muscle ‘quantity’) from V̇O2max. When this scaling is undertaken, V̇O2max is lower in children with CF relative to age- and sex-matched controls, indicating that exercise capacity is not size-dependent in CF and that intrinsic muscular factors (i.e. muscle ‘quality’) are likely responsible for the reduced V̇O2max observed in CF (Chapter 8). Finally, the third component identified applications of CPET for both patients with CF and staff responsible for care. CPET, using a case-study approach, was utilised to describe exercise-related changes in an 11 year old female with CF following surgical insertion of a percutaneous endoscopic gastrostomy and overnight nutritional supplementation. This evaluation identified a maintenance of V̇O2max over one year, in contrast to a fluctuation in FEV1, and increase in body mass index (BMI), therefore highlighting the independent prognostic information afforded by use of CPET (Chapter 9). Following this patient-centred application of CPET, two meetings were held with NHS staff, to provide a platform for exchange of ideas and best practice, but to also survey roles, responsibilities, prevalence of CPET and resources needed for effective implementation of exercise testing and training (Chapter 10). In conclusion, this thesis has further highlighted the utility of CPET in the management of CF. Moreover, it has explored the prognostic and diagnostic properties of CPET, as well as its implementation for patients and staff alike

Engineering oxygen transport for improving cell performance in hepatic devices

The bioartificial liver (BAL) advancing medical technology aims to provide temporary liver substitution for patients in dire need of liver transplants and also for drug development. Yet despite nearly fifty years of development, the approval for this medical device from U.S. regulatory agencies (e.g., Food and Drug Administration) is still pending. The reasons for this have both clinical and fundamental aspects. Current clinical trial data does not convincingly demonstrate a higher survival rate of patients that received BAL treatment than non-BAL treated control. This issue may be attributed to the fundamental fact that, as a medical bioreactor, BALs still lack the effectiveness at the level of the natural liver. As oxygen (O2) is a key substance to determine efficiency of the hepatocytes housed in the BAL, intensifying the O2 conditions within the BAL cellular space will elevate overall BAL performance. This proposition has been substantiated by several studies. With higher efficiency the BAL may increase the liver patients' survival rate, benefit new drug development, and may ultimately attain the government approval in the U.S. The work of this doctoral study focuses on methods of enhancing O 2 transport into three-dimensional (3D) customized hepatic devices. Firstly, enriched O2 conditions were established within customized hepatic systems by applying an inert organic compound - perfluorocarbon (PFC). The PFC-treated hepatic cells demonstrated high cytochrome P450 (CYP 450) function performance especially when 3D gelatin sponge were used as the scaffold. They also exhibited less glucose consumption. Next the 3D iv gelatin sponge scaffold was then characterized in a computational fluid dynamics (CFD)-based simulation to clarify the reasons for the performance improvement. The results of this simulation also suggest that using the new 3D cellular scaffold is an effective method for addressing the O2 delivery problem previously reported for a novel BAL design, the four quadrant bioreactor (4QB) when using the 4QB for the support of larger cell numbers. Lastly, the effects of a previously custom designed flow device and the gelatin sponge scaffold, on the drug metabolism of rat primary hepatocytes (RPHs) were evaluated. The key results from the drug metabolism tests were confirmation of the benefits of combining the 3D gelatin sponge scaffold and flow condition in increasing the hepatocytes drug metabolism enzyme performance. Surprisingly, the results also demonstrated the suppression of the RPHs drug metabolizing ability in flow devices and relevant analysis to this phenomenon was also conducted. This doctoral study has thus provided valuable information on experimental and numerical approaches for improving the fundamental performance of future BAL designs. It mainly highlights that in BALs, the 3D cell cultures and efficient flow perfusion are key to O2 delivery for the scaffold interstitial region (extracellular space). The work thus helps moving toward their development one step closer to establish the future clinical trial and industrial application of BAL devices

Magnetic Resonance Imaging of the Rat Retina

The retina is a thin layer of tissue lining the back of the eye and is primarily responsible for sight in vertebrates. The neural retina has a distinct layered structure with three dense nuclear layers, separated by plexiform layers comprising of axons and dendrites, and a layer of photoreceptor segments. The retinal and choroidal vasculatures nourish the retina from either side, with an avascular layer comprised largely of photoreceptor cells. Diseases that directly affect the neural retina like retinal degeneration as well as those of vascular origin like diabetic retinopathy can lead to partial or total blindness. Early detection of these diseases can potentially pave the way for a timely intervention and improve patient prognosis. Current techniques of retinal imaging rely mainly on optical techniques, which have limited depth resolution and depend mainly on the clarity of visual pathway. Magnetic resonance imaging is a versatile tool that has long been used for anatomical and functional imaging in humans and animals, and can potentially be used for retinal imaging without the limitations of optical methods. The work reported in this thesis involves the development of high resolution magnetic resonance imaging techniques for anatomical and functional imaging of the retina in rats. The rats were anesthetized using isoflurane, mechanically ventilated and paralyzed using pancuronium bromide to reduce eye motion during retinal MRI. The retina was imaged using a small, single-turn surface coil placed directly over the eye. The several physiological parameters, like rectal temperature, fraction of inspired oxygen, end-tidal CO2, were continuously monitored in all rats. MRI parameters like T1, T2, and the apparent diffusion coefficient of water molecules were determined from the rat retina at high spatial resolution and found to be similar to those obtained from the brain at the same field strength. High-resolution MRI of the retina detected the three layers in wild-type rats, which were identified as the retinal vasculature, the avascular layer and the choroidal vasculature. Anatomical MRI performed 24 hours post intravitreal injection of MnCl2, an MRI contrast agent, revealed seven distinct layers within the retina. These layers were identified as the various nuclear and plexiform layers, the photoreceptor segment layer and the choroidal vasculature using Mn54Cl2 emulsion autoradiography. Blood-oxygenlevel dependent (BOLD) functional MRI (fMRI) revealed layer-specific vascular responses to hyperoxic and hypercapnic challenges. Relative blood volume of the retina calculated by using microcrystalline iron oxide nano-colloid, an intravascular contrast agent, revealed high blood-volume in the choroidal vasculature. Fractional changes to blood volume during systemic challenges revealed a higher degree of autoregulation in the retinal vasculature compared to the choroidal vasculature, corroborating the BOLD fMRI data. Finally, the retinal MRI techniques developed were applied to detect structural and vascular changes in a rat model of retinal dystrophy. We conclude that retinal MRI is a powerful investigative tool to resolve layer-specific structure and function in the retina and to probe for changes in retinal diseases. We expect the anatomical and functional retinal MRI techniques developed herein to contribute towards the early detection of diseases and longitudinal evaluation of treatment options without interference from overlying tissue or opacity of the visual pathway