title: The Politics of Child Health Technologies: Social Values and Public Policy on Drug Funding for Children in Canada

contributor:
Abelson, Julia
Health Policy

subject:
health policy
child health
social values
drug policy
health technology assessment
political science

abstract:
Health technology assessment (HTA) frameworks appraise the value of technologies – be they drugs, devices, procedures or services – to inform policy decision-making and resource allocation amongst alternatives within publicly funded health systems. The prevailing principles and metrics by which HTA is conducted were designed with adult health conditions and treatments in mind.  The evidentiary and normative dimensions of HTA frameworks may have unique repercussions for drug policy and coverage decisions in children, but their relevance to child health has received almost no critical scrutiny in either academic or policy circles. Approaches to paediatric drug coverage approval and access currently lack child-specific data on social values and priorities, a core component of HTA in most countries with public drug funding programs, including Canada.
This thesis presents a mixed methods study of social values relevant to child HTA and drug policymaking in publicly funded health systems, comprised of three original scientific contributions. The first of these is a critical interpretive synthesis (CIS) of the academic literature on the moral dimensions of child health and social policymaking across a range of disciplines and policy domains. The second is a grounded theory analysis of qualitative interviews with diverse health system stakeholders on the social values and health system factors relevant to child HTA and drug funding policy in Canada. The third is a stated preference survey of the general public that assesses societal preferences for health resource allocation to children as compared to adults, to generate evidence for priority setting on health technologies within Canada’s publicly funded health system. Together, these studies yield specific knowledge about the policy landscape for child health technologies in Canada, broad conceptual insights into the normative and methodological dimensions of child HTA, and a foundational understanding of the social values relevant to drug policy decisions for children.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)

abstract: Drug research, development and policy have historically excluded children. One area of persistent neglect is public policy on funding for paediatric medicines. In most publicly funded health systems, including Canada’s, decisions about which drugs to cover are made through a formal process called health technology assessment (HTA). This dissertation examines the role and challenges of HTA as applied to child health technologies, with a focus on the social values that inform drug policy for children. It addresses existing gaps in knowledge through the integration of insights from: 1) a comprehensive review of the academic literature on the moral dimensions of child health and social policymaking; 2) in-depth qualitative analysis of the HTA and drug policy environments for children in Canada, employing Ontario as a case study; and 3) a survey of the Canadian public on health system resource allocation for children. Together, these studies generate a detailed picture of the Canadian policy landscape for child health technologies, insights into the fit of current HTA approaches to the realities of child health and illness, and a foundational understanding of the social values relevant to drug policy decisions for children.


title: INVESTIGATION OF LATTICE PHYSICS PHENOMENA WITH UNCERTAINTY ANALYSIS AND SENSITIVITY STUDY OF ENERGY GROUP DISCRETIZATION FOR THE CANADIAN PRESSURE TUBE SUPERCRITICAL WATER-COOLED REACTOR

contributor:
Novog, David
Pencer, Jeremy
Buijs, Adriaan
Engineering Physics

subject:
Generation IV nuclear reactor
Canadian PT-SCWR
Advanced nuclear Energy system
Lattice physics phenomena
Energy group structure optimization
SCALE code
Uncertainty Analysis
Sensitivity Analysis
Transport Calcuations
Optimization and algorithm

abstract:
The Generation IV International Forum (GIF) has initiated an international collaboration for the research and development of the Generation IV future nuclear energy systems. The Canadian PT-SCWR is Canada’s contribution to the GIF as a GEN-IV advanced energy system. The PT-SCWR is a pressure tube reactor type and considered as an evolution of the conventional CANDU reactor. The PT-SCWR is characterized by bi-directional coolant flow through the High Efficiency Re-entrant Channel (HERC). The Canadian SCWR is a unique design involving high pressure and temperature coolant, a light water moderator, and a thorium-plutonium fuel, and is unlike any operating or conceptual reactor at this time. The SCWR does share some features in common with the BWR configuration (direct cycle, control blades etc…), CANDU (separate low temperature moderator), and the HTGR/HTR (coolant with high propensity to up-scatter), and so it represents a hybrid of many concepts. Because of its hybrid nature there have been subtle feedback effects reported in the literature which have not been fully analyzed and are highly dependent on these unique characteristics in the core. Also given the significant isotopic changes in the fuel it is necessary to understand how the feedback mechanisms evolve with fuel depletion. Finally, given the spectral differences from both CANDU and HTR reactors further study on the few-energy group homogenization is needed. The three papers in this thesis address each one of these issues identified in literature. Models were created using the SCALE (Standardized Computer Analysis for Licensing Evaluation) code package.
Through this work, it was found that the lattice is affected by more than one large individual phenomenon but that these phenomena cancel one another to have a small net final change. These phenomena are highly affected by the coolant properties which have major roles in neutron thermalization process since the PT-SCWR is characterized by a tight lattice pitch. It was observed that fresh and depleted fuel have almost similar behaviour with small differences due to the Pu depletion and the production of minor actinides, 233U and xenon.
It was also found that a higher thermal energy barrier is recommended for the two-energy-group structure since the PT-SCWR is characterized by a large coolant temperature compared to the conventional water thermal reactors. Two, three and four optimum energy group structure homogenizations were determined based on the behaviour of the neutron multiplication factor and other reactivity feedback coefficients. Robust numerical computations and experience in the physics of the problem were used in the few-energy group optimization methodology. The results show that the accuracy of the expected solution becomes highly independent of the number of energy groups with more than four energy groups used.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)


title: Reciprocal Influence of Learning a New Bimanual Coordination Pattern and Performing Existing Coordination Patterns

contributor:
Lee, Tim
Human Biodynamics

subject: bimanual coordination learning reciprocal

abstract: Popular theories of motor learning (e.g., Adams, 1971; Schmidt, 1975) rely heavily on formation of new skills through refinements of pre-existing ones. Dynamic Pattern Theory has the advantage of being able to assess initial individual differences on the required task so that the subject becomes the important unit of measure. The general purpose was to identify the reciprocal influence of intrinsic patterns and learning a new pattern. In two experiments subjects were required to practice a rhythmic bimanual coordination task of the forearms using linear sliding devices. In the first experiment, 7 subjects practiced a 90° relative phase pattern for 45, 15 s trials on each of 6 practice days. In-phase and anti-phase trials were performed pre-and post-practice. Subjects were provided terminal feedback with a Lissajou figure after each practice trial and augmented feedback was provided after every 5th practice trial. Mean constant error (CE) for individual subject data and absolute CE (|CE|) for group data were used as measures of accuracy. Standard deviation of relative phase was used as a measure of stability (VE). Subjects were able to learn the 90° pattern and performance plateaued by the fourth practice day. Neither intrinsic pattern showed any destabilization, although a temporary decrease in accuracy (CE) within days was found. The four week retention test revealed no change for any pattern. Experiment 2 compared two groups practicing either 45° or 135° relative phase. It was predicted that the 135° relative phase pattern would be easier to learn because of the reduced competition from the less stable 180° intrinsic pattern. The procedure was similar to Experiment 1 except that subjects practiced for only four days. Performance of the practiced patterns was never as accurate as 0° and 180° but variability of performance was not different for both practiced and intrinsic patterns by Day 4. There was no difference in either accuracy or stability between the two groups on the practiced patterns. As in Experiment 1, there was no change from the last day of practice to the four week retention test. Individual subject data revealed numerous different paths to learning the required pattern. Constant error and VE values for intrinsic patterns were not particularly good predictors of ability to learn the practiced pattern. Additionally, a low VE was not indicative of a low CE or vice-versa when practicing the required pattern. The results from both experiments show that early in learning, competition biases performance away from the intrinsic attractors. Later in practice, subjects stabilize their performance of the new pattern and the intrinsic patterns do not destabilize. Differences in performance of the required pattern may have depended, not only on dynamic principles, but also on motivation, handedness, and conceptualization of the task.

abstract: Thesis

abstract: Master of Science (MSc)


title: Predator Mediated Competition: Predator Feeding on Two Different Trophic Levels

contributor:
Wolkowicz, G. S. K.
Mathematics

subject: predator, mediated competition, feeding, trophic levels, growth-limiting nutrient

abstract:
 We consider a model of the chemostat in which three competitor populations compete
for a single, essential, growth-limiting nutrient. As well, the least efficient competitor population also acts as a predator on the most efficient competitor population. Bifurcation
methods are used to obtain information about the qualitative behaviour of the model. A complete description of the global stability is given for the case when Lotka-Volterra response functions describe both competitor-nutrient and predator-prey interactions. For certain parameter values, the model predicts coexistence of the three species. The model also shows that the elimination of the predator population or the elimination of a competitor population can cause the system to collapse from three species to one.

abstract: Thesis

abstract: Master of Science (MSc)


title: The Effect of Velocity on Muscle Morphology Following Eccentric High-Resistance Training in Young Males

contributor:
Phillips, Stuart M.
Kinesiology

subject: velocity, muscle morphology, eccentric high resistance, training, young males, hypertrophic

abstract:
 It is known that high-resistance training induces morphological changes in skeletal muscle. Following a resistance training program, increases in maximum torque generating capacity are observed due to both neural adaptations and hypertrophic gains within the trained muscle. Although it has been established that a muscle hypertrophies due to the addition of myofibrillar proteins through increased protein synthesis, the exact mechanism which stimulates the hypertrophic response is unknown.

 Previous reports have shown that training in the absence of eccentric contractions generally produces less muscle growth and strength gains, as well as inflicting less damage to the muscle ultrastructure. Likewise, fast eccentric contractions have been shown to increase muscular strength to a greater extent than slow contractions. It has been hypothesized that fast eccentric contractions may maximize muscular damage, thus invoking a greater response of repair mechanisms, including satellite cell recruitment, which would allow an increased addition of contractile proteins to be added to the injured muscle, increasing muscle size and strength to a greater degree.

 The effect of fast and slow eccentric training was investigated using a bilateral, within subject model. Twelve men trained one arm fast (3.66 rad/s) and one arm slow (0.52 rad/s) for 8 weeks on an isokinetic training apparatus. Type I muscle fibre size
increased with training by an average of 9.3±12.0% (P<0.05, main effect for time). Type II muscle fibres increased more in the subjects' fast trained arm when compared to the slow trained arm according to ATPase histochemical analysis (P<0.05, time x condition interaction). Likewise, whole arm cross-sectional area showed that the fast trained arms had an average increase of 6.8±5.5 % whereas the slow arms only had an average increase CSA of 5.1±5.7% (P=0.065, time x condition interaction). Maximum torque generating capacity was also increased to a greater degree (P<0.05, time x condition interaction) in the fast trained arm with an average of 10.3±16.4 Nm, whereas the slow trained arm increased only 7.3±15.0 Nm, across testing speeds. A decrease in the percentage of type IIx fibres was seen in both arms after training according to both ATPase histochemical staining and MHC gel electrophoresis; however, the percentage of type IIa fibre area increased in the fast trained arms (8.4±8.6%) more significantly (P<0.05, time x condition interaction) than the slow trained arms (1.7±10.9%).

 Seven males were trained in a similar manner to determine the extent of muscle damage which was evaluated by both Z-band streaming and force production decrements. After a single exercise bout of fast eccentric training in one arm and slow eccentric training in the other, it was determined that a 1.97±0.74 areas of moderate Z-band streaming per mm^2 of muscle in the fast exercised arm compared to 0.89±0.79 areas of moderate Z-band streaming per mm^2 of muscle in the slow trained arm (P<0.05). In conclusion, training using fast (3.66 rad/s) eccentric contractions causes a greater degree of muscle damage, hypertrophy, and strength gains than does training with slow (0.52 rad/s) eccentric contractions.

abstract: Thesis

abstract: Master of Science (MSc)


title: Characterization of the Ionic Currents In Cultured Small Intensely Fluorescent Cells from Superior Cervical Ganglia of Neonatal Rats

contributor:
Nurse, C.A
Biology

subject: ganglion, autonomic cardiovascular reflexes, postganglionic nerve trunk

abstract:
The superior cervical ganglion (SCG) is the largest of the sympathetic chain
ganglia which control a number of autonomic cardiovascular reflexes via neural activity
in the postganglionic nerve trunk. In addition to the large principal neurons, these
ganglia contain a minority population of smaller cells, the small intensely fluorescent or
SIF cells, so named because of their intense fluorescence following treatments which
reveal the presence of endogenous catecholamines (mainly dopamine in the rat). The
physiological functions of the SIF cells are largely unknown and various roles have been
proposed including (i) dopaminergic interneuron, which modulates ganglionic
transmission, (ii) endocrine function, since many of them have a close association with
the vasculature and (iii) chemosensory function, similar to that of the arterial
chemoreceptors which sense blood gases and pH. Understanding the physiological role
of SIF cells has been hampered by their small size, sparse distribution and relative
inaccessibility, all of which render microelectrode electrophysiological studies difficult.
In this thesis these limitations were overcome by use of (i) dissociated cell cultures of the
rat SCG, in which growth conditions favoured SIF cell survival but not that of the
principal neurons, and (ii) the novel high resolution patch clamp/whole cell recording
technique which is ideal for the study of the electrophysiology of small cells.
The ionic currents, which underlie many basic electrophysiological processes,
were characterized in 5-16 day old cultures of SIF cells obtained from the SCG of
neonatal rats. The main methodology consisted of whole cell recording under voltage
clamp conditions, which permit the study of membrane ionic currents. Five main ionic
currents were identified in all of the SIF cells ( > 100) studied: (i) a fast transient inward
Na+ current, sensitive to the well-known blocker of voltage-gated Na+ channels i.e.
tetrodotoxin or TTX; (ii) the delayed rectifier outward K+ current that is found in a
variety of cell types; (iii) a Ca2+- activated outward K+ current, sensitive to Ca2+ channel
blockers; (iv) a transient inward Ca2+ current which appears to be carried by N-type
Ca2+ channels and (v) a slower, sustained inward Ca2+ current which appears to be
carried by L-type Ca2+ channels. In addition a third type of outward K+ current, the fast
transient K+ current or lA, was found in SIF cells obtained from 3-7 day old rats, but not
from 1 day old rats. It therefore appears that this lA current, which is known to
modulate firing frequency in neurons develops rapidly in vivo during the first postnatal
week. This broad repertoire of ion channels in SIF cells suggests several possible sites
for modulation by various agents including neurotransmitters, neuromodulators, or other
chemosensitive agents. Since SIF cells were recently proposed to have arterial
chemoreceptor function similar to glomus cells, the effect of one such stimulus, i.e. an
acidic (intracellular) pH, was tested. It has recently been suggested that a decrease in
intracellular pH is part of the pathway responding to extracellular stimuli in the glomus
cell (Stea, Alexander and Nurse, in press). Acidification of the SIF cell's cytoplasm with
the K+/H+ ionophore nigericin resulted in a suppression of both the fast inward Na+
current as well as the outward K+ current. However, these effects do not appear to beunique to SIF cells and therefore the possibility of a chemoreceptor role m the
cardiovascular system requires further study.
In summary, the characterization of the various ionic currents in SIF cells
resulting from this thesis provides the necessary background which should eventually
resolve not only the question of the physiological role of SIF cells in autonomic ganglia,
but also help to understand the underlying mechanisms responsible for SIF cell function.

abstract: Thesis

abstract: Master of Science (MS)


title: Micromechanical Analysis of Induced Anisotropy in Granular Materials

contributor:
Guo, Peijun
Civil Engineering

subject:
Granular material
Micro-mechanics
Fabric anisotropy
strong and weak sub-network

abstract:
Granular materials, such as sand, are systems consisting of huge numbers of particles that interact with each other through inter-particle contacts. Different from continuum materials, a granular material displays distinctive features due to the discrete nature of the microstructure, characterized by a spatial arrangement of inter-particle connection as well as a force-chain network. With a consideration of the contact force, the overall contact network is divided into a strong sub-network and a weak sub-network that carry contacts with normal contact forces larger and lower than the average normal contact force, respectively. Thus, the fabric anisotropy for different contact networks, are employed to characterize the microstructure of the granular material.
	In this research, the behavior of granular materials subjected to quasi-static shear was extensively investigated in terms of the fabric evolution including the magnitude and direction of anisotropy for different contact networks. Both statistical and micromechanical approaches were adopted to obtain the macroscopic properties, such as the fabric tensor, Cauchy stress tensor and the second-order work, in terms of the micro-scale variables. The discrete element method (DEM) was employed to simulate laboratory tests along fixed loading paths; for example, 2D tests along proportional strain paths, 2D simple shear tests and 3D tests along radial stress paths on the π-plane.
	Results demonstrated that the induced fabric anisotropy for the overall contact network can be related to the deviatoric stress ratio for both two-dimensional and three-dimensional conditions. The relation was found to be not unique, depending on the loading paths as well as the stress state. Nevertheless, a unique linear fabric-stress relation was presented between the stress tensor and fabric tensor for the strong sub-network. Specifically, the obliquity of this linear relation was found to be a function of the mean stress. This description held true for initially isotropic specimens subjected to proportional and non-proportional loading paths. On the other hand, for the initially anisotropic specimen, this correspondence only worked at the critical stress state.
	According to Nicot and Darve (2006), the macro second-order work cannot be interpreted as a summation of the local second-order work from the contact plane. The second-order work induced by the fabric evolution and the volumetric change must also be taken into account. The second-order work induced by the fabric evolution cannot be neglected in 2D analysis along proportional strain paths. Moreover, the vanishing of the second-order work is related to the fabric anisotropy in contact sub-networks that the decrease of fabric anisotropy for the weak sub-network or the degradation of weak sub-network was observed to be an indicator of deformation instability even though the strong sub-network dominants the shear resistance. The degradation of strong sub-network was a necessary but not a sufficient condition of instability.
	The direction of the fabric anisotropy for the strong sub-network was observed to be coaxial with the orientation of the principal stress. The principal direction of fabric anisotropy for the weak sub-network was always perpendicular to that of the strong sub-network, regardless of whether the principal stress rotated or not. For the overall contact network, however, the direction of the fabric anisotropy was not necessarily in line with the major principal stress direction, even for an initial isotropic granular assembly. Therefore, the finding by Radjaï et al.(1998) that the direction of the fabric anisotropy for the weak sub-network is perpendicular to that for the overall contact network only held true for the loadings in which the critical stress could be approached no matter if the principal stress orientation rotated or not. Under this circumstance, the fabric anisotropy for the overall contact network could be interpreted as a function of sub-networks’ anisotropy weighted by the ratio of contact number in each sub-network over the total number of contacts.
	At critical state, both the strong sub-network and the overall contact network developed high fabric anisotropy with the weak sub-network being mostly isotropic. When plotted on the π-plane, both the fabric anisotropy for the strong sub-network and the fabric anisotropy for the overall contact network depended on the stress paths but were independent of the mean stress level. The response surface of the former could be expressed as a Lade’s surface. The response envelope of the latter was an inverted Lade’s surface.

abstract: Dissertation

abstract: Doctor of Philosophy (PhD)

abstract: In civil engineering, granular materials are ubiquitous, such as sand, gravel, rock, and concrete. Due to the discrete nature of microstructure, this type of material usually displays exceedingly complicated behaviours under shear, for example, dilatancy, non-coaxiality, critical state, instability, and anisotropy. These mechanical responses are notoriously difficult to model and most existing models are phenomenological and lack a clear physical meaning. To provide a clear physical meaning for the constitutive model of granular material, the current study explored the evolution of the microstructure within the granular material subjected to quasi-static shear and the micromechanical origins of those macroscopic behaviours such as critical state, non-coaxiality, and instability. Both micromechanical analysis and discrete element method were applied. Results showed that the evolution of the whole microstructure depended on the loading condition. However, the evolution of the microstructure joined by the ‘strong contacts’ was independent of the loading path. At critical state, the microstructure was highly anisotropic, not unique and depended on the stress paths. The rearrangement of the microstructure helped to maintain the stability of a granular material. The instability of the granular material was triggered by the failure of the microstructure joined by the ‘weak contacts’.


title: Ultratrace Lithium and Boron Analysis by Neutron Activation and Helium Isotope Mass Spectrometry

title: Li and B Analysis by NAA and He Isotope Mass Spectrometry

contributor:
Clarke, W.
Physics

subject: lithium boron neutron helium spectrometry isotope

abstract: The binding of the elements lithium and boron to human plasma proteins is investigated through the techniques of thermal neutron activation and helium isotope mass spectrometry. Since normal physiological levels of lithium and boron in blood and plasma are in the ultratrace (ppb) range, lithium in particular is frequently below the detection limit of many instruments. The success of the detection method used in this work is due to the extremely large cross section of ⁶Li for the thermal neutron reaction ⁶Li(n,³H)⁴He, and of ¹⁰B for the thermal neutron reaction ¹⁰B(n,α)⁷Li. The high sensitivity of the mass spectrometer, originally designed for oceanographic studies of helium isotopes and tritium, allows measurement of as little as 2x10⁴ atoms of ³He from the decay of tritium, and 2x10¹⁰ atoms of ⁴He from alpha-particles. It has frequently been stated that lithium does not bind to plasma proteins. However, our results clearly show that lithium does bind to a number of these proteins, at least 𝘪𝘯 𝘷𝘪𝘵𝘳𝘰. Boron is also shown to bind to proteins, with a pattern similar to that of lithium . Although a clear identification of the specific plasma proteins which bind lithium and boron must await further investigation, a number of possibilities are suggested here, based on the data obtained.

abstract: Thesis

abstract: Master of Science (MSc)


title: The Accuracy of Dual Photon Absorptiometry Measurements of Soft Tissue Composition

contributor:
Webber, C.E
Physics

subject: densitometer, dependent error, soft tissue

abstract:
During routine measurements of body composition using a 153Gd
based dual photon densitometer, it was observed that negative
values were being obtained for the body fat fraction in some
adults, in children and in small animals. In these three groups,
there appears to be a body size dependent error whereby the
measured fat fraction becomes increasingly negative as subject size
becomes smaller.
The fat fraction is derived from relating the measured mass
attenuation coefficient of soft tissue to an internal calibration
based on the use of water and lard as substitutes for muscle and
fat. To investigate whether this procedure for instrument
calibration is the cause of the fat fraction errors, soft tissue
phantoms which contained known amounts of fat, water and protein
were prepared. Over the range of fat fractions used, accurate
results were obtained.
By using prepared soft tissue and water phantoms it was
established that the measured fat fraction incorrectly became
progressively smaller as object thickness decreased and incorrectly
increased with object thickness. However, accurate measurements
were obtained if the equivalent tissue thickness is greater than
9 em and less tnan 16 em of water. Equally reproducible
measurements are obtained at all thicknesses investigated.
When dual photon measurements of body composition in 13 adolescent females were compared with measurements obtained from
skinfold thicknesses or bioimpedance, there was good agreement
between techniques but dual photon results demonstrated a broader
range of variation with body size. Comparisons between dual photon
absorptiometry derived body composition measurements of 52 male
athletes with results obtained from under water weighing allowed
for derivation of a simple correction factor for the accuracy
errors due to body size.

abstract: Thesis

abstract: Master of Science (MSc)


title: Analysis of Small Biomolecules by Esi- and Maldi-Mass Spectrometry

contributor:
Terlouw, Johan
Chemistry

subject: biomolecules esi- maldi-mass spectrometry

abstract: This thesis describes the use of mass spectrometric methods based upon electrospray ionization, ESI, and (matrix-assisted) laser desorption/ionization, (MA)LDI, for the quantitative analysis of small biomolecules. Structure analysis when required, was obtained through tandem mass spectrometry (MS/MS). The Girard type reagent, 4-hydrazino-4-oxobutyl tris(2,4,6 trimethoxy)phenyl phosphonium bromide, in combination with the solid phase derivatization technique, is used to selectively prepare a pre-ionized malondialdehyde derivative to be analyzed by MALDI or LDI. The in situ derivatization and isolation minimize interferences from other components in biological samples. The combination of pre-ionization and aromatic functionalities allows for laser induced ionization without the need of matrix. The combination of these techniques provides an avenue for development of automation to produce a high throughput method of analysis. Chapter 3 involves the study of the complexation of diols to the oxovanadium ion. The oxovanadium (IV) complex of ethylene glycol is used as a reference to study the complexation of other diols and amines with the vanadyl ion. The ES spectra of various diols studied produce intense signals for the mixed and the analyte complexes, indicating effective complexation of the analytes with the vanadyl ion. Oxovanadium (IV) is observed to be more selective for complexation to diols than amines. This eliminates the possibility of interference from N-containing ligands to the detection of diols by the reference complex. The electrospray spectrum is used for quantitation and the tandem mass spectrometry spectrum for structure confirmation. The MS/MS spectrum also assists the identification of the diols by the structural differences within their isomers. The equilibrium constant of a set of diols was determined and its calibration curve were constructed. This study produces an alternative method to detect and quantify diols in aqueous solutions and blood samples.

abstract: Thesis

abstract: Master of Science (MSc)


title: The Synthesis Of 11C-Labelled Melatonin Agonists from 11C-Carbon Dioxide

contributor:
McCarry, Brian E.
Chemistry

subject: radioisotope, neurohormone melatonin, Positron Emission Tomography

abstract:
This thesis describes the application of the radioisotope 11C to the synthesis of two
analogues of the neurohormone melatonin. The labelled compounds were intended to be
used as tracers for the medical imaging technology Positron Emission Tomography (PET).
[
11C]Carbon dioxide, produced in a small on-site cyclotron by the nuclear reaction
14N(p,a)11C, was converted into [11C]CH3COC1 by reaction first with CH3MgBr, followed
by reaction with phthaloyl dichloride. The labelled acid chloride was distilled into a
solution of an amine, yielding the corresponding 11C-labelled amide, which was purified
by a simple solid-phase extraction method.
An apparatus was designed and built that allowed the remote synthesis with
several hundred millicuries of [11C]C02• The apparatus was mounted in a hot cell and
operated remotely with a Macintosh Powerbook programmed in Hypercard. The apparatus
and software are generic for these acylation reactions.
The individual reaction steps were optimized in terms of reaction time, solvents
and equipment; radiosyntheses of a number of purified labelled acetamides were
completed in 35 minutes. The radiochemical yields ranged from 15 to 20% with specific
activities in the 500 mCi/J..tmol range at the end of the synthesis.
2-Iodo-[11C-acetyl]melatonin (11) and 7-methoxynaphthylenyl-1-ethyl-N-[11Cacetyl]
acetamide (15) were synthesized for the first time for PET studies. It was shown
that both compounds readily cross the blood-brain-barrier and penetrate into all brain
tissues. Specific binding to the melatonin receptors in the suprachiasmatic nuclei of the
hypothalamus could not be visualized with either one of the 11C-labelled ligands because
of low specific activity and high nonspecific binding.

abstract: Thesis

abstract: Master of Science (MSc)


title: Postabsorptive Glucose Decreases the Excitatory Effects of Taste on Ingestion

contributor:
Weingarten, H.P
Psychology

subject: hypothesis, plasma, sham feeding, IP glucose

abstract:
To test the hypothesis that postprandial rises of plasma
glucose attenuate the motivation derived from positive
tastes, I analyzed the effects of intraperitoneal (ip)
injections of glucose on sham feeding, a preparation in
which food intake is motivated primarily by taste
sensations. IP glucose suppressed sham feeding, with
maximal suppressions approximating 42%, but only when
glucose was administered contiguous with oropharyngeal
stimulation. The food intake inhibition produced by ip
glucose interacted with palatability; smaller doses of
glucose were required to suppress less concentrated sucrose
solutions. Closing the gastric cannula increased the
potency with which ip glucose inhibited eating suggesting
synergy of postabsorptive glucose with other postgastric
satiety signals. The inhibition of eating produced by ip
glucose did not result from malaise. Thus, postabsorptive
rises of plasma glucose decrease the ability of taste cues
to drive ingestion and suggest that this phenomenon may
contribute to spontaneous meal termination.

abstract: Thesis

abstract: Master of Science (MSc)


title: A Lateral-Inhibitory-Network Model of the Central Processing of Auditory Nerve Activity

title: A Model of the Central Processing of Auditory Nerve Activity

contributor:
Bruce, Ian
Electrical Engineering

subject: inhibitory auditory nerve processing

abstract:
Lateral inhibitory networks (LINs) of neurons are thought to be prominent in sensory systems and are known to enhance spatial edges and peaks in their input excitation patterns. It is postulated, based on experimental findings, that lateral inhibition contributes to central, sub-cortical, auditory processing. Previous computational LIN models of the central processing of auditory nerve activity were based on highly simplified, non-spiking models of neurons. A more biologically realistic LIN model of spiking neurons was thus developed to investigate the plausibility of such networks achieving contrast enhancement and speech feature extraction.
The model developed is a single-layer, uniform, recurrent LIN structure. Each neuron in the LIN is described by a leaky integrate-and-fire model with conductance-based synaptic input. Input spike instances were obtained from Bruce and colleagues' [2003] model of the auditory periphery for synthesized speech stimuli or from a Bernoulli approximation of a Poisson process to represent spontaneous activity from the auditory nerve. The effect of neural and network parameters on contrast enhancement exhibited in the mean spike rates was measured. It was found that the spiking LIN is able to achieve contrast enhancement if the values of the neuronal parameters fall within a very specific and narrow range. Furthermore, the spatial edge in the input had to be high and steep. Compared to non-spiking neuron models, it is quite difficult for spiking neurons to achieve contrast enhancement. The spiking LIN was found to be capable of making formant frequencies more distinct in the average rate profiles of speech stimuli presented at high intensities. However, synchrony of the neural activity to the formant frequencies was largely lost. This architecture of spiking neurons is therefore unlikely to be how contrast enhancement and speech feature extraction is realized in the central auditory system.

abstract: Thesis

abstract: Master of Applied Science (MASc)


title: The Effect of the Muller-Lyer Illusion on the Planning and Control of Manual Aiming Movements

title: The Planning and Control of Manual Aiming Movements

contributor:
Elliott, Digby
Kinesiology

subject:
müller-lyer illusion
manual
movements

abstract: Three experiments were conducted to investigate the two visual systems hypothesis (Milner & Goodale, 1995) and the planning-control hypothesis (Glover, 2002). Experiment 1 required the participants to make rapid aiming movements to 25 cm and 35 cm tails-in, no tails, or tails-out Müller-Lyer stimuli following a 0 ms or 5000 ms no-vision delay. In Experiment 2, the participants executed their movements with full vision of the Müller-Lyer vertices that either remained the same or changed to a different configuration upon movement initiation. Vision was occluded either 350 ms or 450 ms after the onset of the movement. Experiment 3 was similar to Experiment 2, except the amount of visual feedback for on-line control was constrained to 200 ms, 400 ms, or 600 ms. The results of these experiments are problematic for both hypotheses. The participants exhibited a similar pattern of illusion-induced bias in both short and long delay conditions. In addition, the magnitude of the aiming bias increased as the movement unfolded (Experiment 1). Furthermore, even though participants were engaging in on-line control the illusion continued to exert its effects on aiming during the latter stages of the movement (Experiment 2). This effect was also observed when participants had sufficient time to process visual feedback in order to modify their movements (Experiment 3). Taken together, the results suggest that on-line control is biased by visual illusions.

abstract: Thesis

abstract: Master of Science (MSc)


title: The Characterization of Baxb, a Splice Variant of the Pro-Apoptotic Bax and Protein

contributor:
Andrews, D.W
Biochemistry

subject: baxa, RNA, isoform, carboxyl-terminal sequence

abstract:
Baxa is a pro-apoptotic member of the Bcl-2 protein family that regulates a
key point in the control of apoptosis. The bax RNA undergoes a complex pattern
of RNA splicing, with eight splice isoforms known to date. The next most
abundant isoform to Baxa is Baxp, which has a unique carboxyl-terminal
sequence and consequently lacks the transmembrane domain of Baxa. This
study characterized Baxp as part of a larger project aimed at deducing the role of
cellular localization in Bax protein function.
A transient transfection assay was designed to determine the cell death
activity of a protein in adherent cells. Baxp induced cell death to a greater extent
than Baxa when transiently expressed in NIH 3T3 cells. The levels of Baxp
expression were always low, regardless of the cell type or transfection method
used. Additionally, Baxp adopted a conformation in which amino acids 13 to 19
are accessible to the monoclonal antibody 6A7. In Baxa this epitope is only
exposed in the membrane-bound activated conformation that is associated with
accessibility of the BH3 domain of Baxa that mediates protein-protein
interactions among Bcl-2 family members. Although the mechanism remains
elusive, Baxp was identified as a potent inducer of cell death.

abstract: Thesis

abstract: Master of Science (MSc)


title: Dimension Reduction and Clustering of High Dimensional Data using a Mixture of Generalized Hyperbolic Distributions

contributor:
McNicholas, Sharon
Statistics

subject: Model-based clustering, dimension reduction, statistical learning

abstract:
Model-based clustering is a probabilistic approach that views each cluster as a component
in an appropriate mixture model. The Gaussian mixture model is one of the
most widely used model-based methods. However, this model tends to perform poorly
when clustering high-dimensional data due to the over-parametrized solutions that
arise in high-dimensional spaces. This work instead considers the approach of combining
dimension reduction techniques with clustering via a mixture of generalized
hyperbolic distributions. The dimension reduction techniques, principal component
analysis and factor analysis along with their extensions were reviewed. Then the aforementioned
dimension reduction techniques were individually paired with the mixture
of generalized hyperbolic distributions in order to demonstrate the clustering performance
achieved under each method using both simulated and real data sets. For a
majority of the data sets, the clustering method utilizing principal component analysis
exhibited better classi cation results compared to the clustering method based
on the extending the factor analysis model.

abstract: Thesis

abstract: Master of Science (MSc)


title: Stochastic Approaches for Estimating the Performance of Low Impact Development Practices

contributor:
Guo, Yiping
Civil Engineering

abstract:
To reduce the impacts of urbanization on the natural hydrologic cycle, low impact development (LID) practices have been increasingly implemented worldwide. This thesis aims at improving the existing analytical models and developing new sets of analytical equations to quantify the performance statistics of structural LID practices. As a starting point, the previously developed analytical probabilistic approach is extended for developing an event-based probabilistic model of infiltration facilities, which is based on the exponential probability distributions of rainfall characteristics and the mathematical representation of the hydrologic processes involved in the operation of infiltration facilities. Analytical equations for the determination of their overflow frequency and stormwater capture efficiency are obtained.
To better understand the antecedent condition and avoid using any simplifying assumptions about it, an analytical stochastic approach is then proposed for the analysis of rainwater harvesting (RWH) systems and permeable pavement systems (PPSs). Using this approach, the forward stochastic differential equations that relate the probabilities of the moisture state involved in the operation of RWH systems and PPSs are established. The steady-state probability distributions of moisture contents are then analytically solved from these stochastic differential equations. Based on these steady-state probability distributions, the stormwater capture and water saving efficiencies of RWH systems are analytically derived. Applying the stochastic approach together with the analytical probabilistic approach to study the operation of PPSs, analytical equations that can be used for evaluating the stormwater capture efficiency of PPSs are also obtained.
Verifications of all the analytical solutions are made for a wide range of cases located in different climate regions by comparison with continuous simulations. It is demonstrated that the analytical equations presented in this thesis provide an easy-to-use tool with higher accuracy and wider application range for the planning and design of LIDs, which is much needed given the increasing implementations of LIDs.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)


title: An Examination of the Mechanisms Underlying Acute and Chronic Hypoxic Regulation of L-Type Ca2+ Channel a 1CSubmits

contributor:
Fearon, Ian M.
Biology

subject: vascular smooth muscle cells, influx, hypertension, hypoxic responses, mitochondral complexes

abstract:
L-type Ca2+ channels, found in vascular smooth muscle cells, function to control
Ca2+ influx, which directly regulates the degree of tension in the vasculature. An influx
of Ca2+ causes these cells to contract while inhibition of this channel causes muscle
relaxation, a major goal in treating hypertension. Acute hypoxia inhibits, and chronic
hypoxia enhances, Ca2+ channel currents. The mechanisms underlying these hypoxic
responses were examined in HEK 293 cells by altering cellular levels of proposed
mediators of 0 2 sensing which have previously been shown to be involved in the redox
model of 02 sensing in various cell types. In these studies I investigated the roles of
mitochondrial complexes and NADPH oxidase function, and changes in cellular ROS
levels, on the acute and chronically hypoxic regulation of recombinant L-type Ca2+
channels. An increase in H202, a form of ROS, by exogenous application was found to
enhance Ca2+ currents. However neither catalase nor H202 affected the acute hypoxic
response. In contrast superoxide dismutase (SOD) abolished hypoxic inhibition of
recombinant L-type Ca2+ channels, suggestive of a role of 02- production in 02 sensing.
Altered production of this ROS during hypoxia may occur within the mitochondria since
acute 02 sensing was abolished in mitochondria-depleted p0 cells. Alterations in
NADPH oxidase activity via application of NADPH oxidase inhibitors such as DPl and
P AO did not mediate the acute hypoxic response. Hypoxic regulation of mitochondrial
complex I may also mediate the response to chronic hypoxia since current enhancement
by this stimulus was abolished by rotenone. These findings support the involvement of
altered mitochondrial function in the 0 2 sensing pathway which mediates the hypoxic
responses of recombinant L-type Ca2+ channel a1c subunits.

abstract: Thesis

abstract: Master of Science (MSc)


title: Structure and Function Studies of FKBP65:A Putative Molecular Chaperone of Tropoelastin

contributor:
Ananthanarayanan, V.S
Biochemistry

subject: immunophilin class, cyclophilins, macrolides, turbidity, tropoelastin

abstract:
FKBP-65 is a member of the immunophilin class of proteins consisting primarily
of the cyclophilins and the FKBP's which bind the immunosuppressant drugs cyclosporin
A and FK506, respectively. Immunophilins possess peptidylprolyl cis-trans isomerase
(PPiase) activity which is inhibited upon binding of their respective macrolides. Specific
cellular targets of most immunophilins and the role of PPiase activity in vivo remain
largely unknown. FKBP-65 has been proposed as a molecular chaperone of tropoelastin
(TE), the soluble precursor of elastin (Davis et al. 1998). TE contains 12% proline
residues, many of which are found in VPGVG repeats. When P2 is in the trans
conformation, these motifs form repeated type-II ~-turns and ~-spirals resulting in selfassociation
of TE via an inverse temperature-dependent transition known as coacervation.
Coacervation can be monitored by turbidity increases at 300 nm. We have used purified
recombinant FKBP-65 in coacervation assays with chick aorta TE to show that FKBP65
specifically affects the coacervation characteristics of TE in a concentration-dependant
manner. The overall extent of coacervation of TE could be increased by more than 2-fold
over controls by inclusion of nM amounts of FKBP-65 in the assay. Also, FKBP-65
decreases the coacervation onset temperature of TE by 5-l 0°C. Structural evidence
suggests that the influence of FKBP65 on tropoelastin coacervation may be due to its
ability to increase the ~ structural content of tropoelastin. These results suggest that
FKBP-65 may be a physiologically relevant, TE-specific molecular chaperone.

abstract: Thesis

abstract: Master of Science (MSc)


title: Fusion and Steric Stabilization of Liposomes Containing Membrane-Anchored Biopolymers

contributor:
Epand, Richard
Biochemistry

subject: fusion, steric stabilization, liposomes, membrane-anchored, bipolymers

abstract:
 Studies examining the characteristics of membranes that facilitate and affect fusion are central to understanding the intricacies of inter- and intra-cellular fusion processes and expanding the existing knowledge of other roles membranes may have. In this thesis a model membrane system, using Sendai Virus and Egg phosphatidylcholine (EggPC) liposomes containing the receptor glycophorin A (proteoliposomes), was used in examining different fusion with proteoliposomes prepared by different methods. For the first time glycophorin A
was incorporated into EggPC liposomes vectorially. This was accomplished separately with two detergents: octylglucoside and CHAPS. Fusion of Sendai Virus with the reconstituted proteoliposomes revealed that octylglucoside reconstituted proteoliposomes exhibited lower fusion compared with CHAPS reconstituted proteoliposomes. Efforts to determine the basis for this difference, using either proteinase K or O-glycosidase digestion and subsequent fragment
analysis using SDS-PAGE and silver-staining, were inconclusive. A separate study examined the ability of large membrane-anchored biopolymers (chosen in virtue of their large hydrophilic domains) to sterically stabilize Egg PC liposomes. Glycophorin A, the lipophosphoglycan (LPG) from Leishmania donovani, and a polyethyleneglycol-conjugated phospholipid (PEG5000-PE) were incorporated into Egg PC liposomes. In each case, binding of a soluble fluorescent probe, NeutrAvidin Oregon Green, to liposomes containing biotin-conjugated lipid was restricted. This supports the idea that large membrane-anchored biopolymers are able to sterically stabilize liposomes.

abstract: Thesis

abstract: Master of Science (MSc)


title: Deformation of Granular Materials under Multi-Directional Loading

subject:
geotechnical engineering
constitutive modelling

abstract:
The deformation and failure properties of granular soils largely affect the stability of upper structures built on or in such soils. Owing to its discrete nature as well as the randomness of particle shape and inter-particle connectivity, the internal structure of a granular material usually exhibits a certain level of anisotropy.  In addition, the microstructure of a granular material evolves following certain patterns, which are influenced by the initial fabric, void ratio, stress level, as well as the stress or deformation history. It has been a major challenge to properly describe the deformation of anisotropic granular materials in constitutive models especially when the materials are subjected to cyclic loading. The existing constitutive models usually have limited capabilities in describing the behaviour of granular materials subjected to repeated loading with principal stress rotation. How to quantify the microstructure change and how to consider the changing microstructure in constitutive models have been two missing links for building a comprehensive model framework.

This research aimed at developing a constitutive model that can properly describe the deformation of granular soils under repeated multi-directional loading. To achieve this goal, a systematic study was performed, including a comprehensive experimental study and a theoretical development of a stress-strain model with proper consideration of the influence of fabric. The developed model was verified with experimental results and then implemented into a finite element code to solve boundary-valued problems.

In the first part of this study, a comprehensive experimental study was carried out to investigate the behaviours of granular materials under both monotonic and cyclic loading to investigate the influence of the intermediate principal stress and the major principal stress direction on soil responses. The results of monotonic loading tests showed that both the strength and dilatancy of sand decreased notably with an increase of either the intermediate principal stress or the inclination angle of the major principal stress direction relative to the major principal fabric direction. The stress states at failure from the tests suggested that the benchmarked Matsuoka-Nakai and Lade-Duncan failure criteria are only valid under certain conditions. From the cyclic loading tests, it was observed that, in addition to the increased intermediate principal stress, varied cyclic loading direction caused a significant increase in accumulative volumetric compaction.

To consider the microstructural dependencies of granular materials, a more general mathematical formulation of stress-dilatancy was developed based on the assumption of the existence of a critical state fabric surface that is expressed as a function of the invariants of the fabric tensor. This assumption was also used to establish the fabric evolution law. The implementation of the resulting stress-dilatancy formulation and the fabric evolution law in elasto-plasticity theory produced interesting modelling results consistent with experimental observations with respect to the microstructural aspects of granular materials. The developed constitutive model was further extended to cyclic loading within the framework of hypo-plasticity with kinematic hardening. The model was capable of describing the behaviour of sand subjected cyclic loading under various conditions including the variation of loading directions.

Finally, the constitutive model was implemented into a commercial software package ABAQUS via the subroutine UMAT.  The capacity of the proposed stress-strain model in solving boundary value problems was examined. Six series of elements tests were designed to examine the proposed model under different initial void ratios, degrees of anisotropy, loading directions, and stress paths. Furthermore, a series of simulations were performed for the settlement of footing on sands with different bedding plane orientations. Results from the simulations were found to be consistent with experimental observations.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)


title: Net Ecosystem CO2 Exchange in Natural, Cutover and Partly Restored Peatlands

contributor:
Waddington, J. M.
Geography

subject: ecosystem, CO2 exchange, natural, cutover, restored peatlands, soil moisture content

abstract:
 Peatlands are an important component of the global carbon cycle, storing 23 g C m-2 yr-1 to comprise a global carbon pool of approximately 455 Pg. Peat drainage and harvesting results in removal of surface vegetation, thereby reducing gross photosynthesis to zero. Moreover, lowering the water table increases carbon oxidation. Consequently, peatland drainage and mining can reduce or eliminate the carbon sink function of the peatland. In the first part of this study, net ecosystem CO2 exchange was studied in a natural (NATURAL), two-year (YOUNG) and seven-year (OLD) post cutover peatland near Ste. Marguerite Marie, Quebec during the summer of 1998. Although the NATURAL site was a source of CO2 during the study season, CO2 emissions were 270 to 300% higher in the cutover sites (138, 363, and 399 g CO2-C m-2; NATURAL, YOUNG and OLD, respectively). Active restoration practices and natural re-vegetation of peatlands have the potential to return these ecosystems to net carbon sinks by increasing net ecosystem production (NEP) and therefore decreasing CO2
emissions to the atmosphere. Net ecosystem CO2 exchange in a natural (NATURAL) peatland and a partly restored peatland (REST) near Ste. Marguerite Marie, Quebec, was compared with a naturally re-vegetated peatland (RVEG) near Riviere-du-Loup, Quebec. Ecophysiological parameters indicate that the REST site was more than twice as productive as the natural LAWNS and three times as the RVEG site (GPmax=18.0, 8.3, and 6.5 g CO2 m-2 d-1, respectively). These results indicate that active restoration improves carbon sequestration over natural re-vegetation but that the net carbon sink function at both sites has not been restored. The presence of Sphagnum cover at the RVEG site resulted in a significant decrease in net ecosystem respiration (NER), indicating the potential for decreasing soil respiration at restored cutover sites through increasing the volumetric soil moisture content.

abstract: Thesis

abstract: Master of Science (MSc)


title: Studies on Genomic Sequences For the Heat Shock Proteins hsp60 and hsp10 From Chinese Hamster Ovary Cells

contributor:
Gupta, R.S
Biochemistry

subject: cDNA sequences, heat shock proteins, Chinese hamster ovary cells, genomic library, regulatory sequences

abstract:
Although the eDNA sequences for the 10 k:Da (hsp 10, hsp 1 0) and the 60 k:Da (hsp60,
cpn60) heat shock proteins have been obtained for a number of mammalian species, until very
recently information was not available on the functional genes encoding these proteins. The
primary objective of this work was to clone and sequence the functional genes for these
proteins from CHO, Chinese hamster ovary cells. Screening of a lambda EMBL3 CHO
genomic library with the CHO hsp 10 eDNA identified a clone containing the putative hsp 10
functional gene. A -5.5 kb fragment was isolated from one of these clones by enzymatic
digestion and -3.3 kb was sequenced. The clone was found to contain consensus regulatory
sequences upstream of the putative transcription initiation site, + 1, including two Sp 1 binding
sites, a CAAT box, and a single heat shock element, HSE, but lacked a TATA box. The
coding region consists of four exons, identical to the hsp10 CHO eDNA sequence, separated
by three introns, of 200 bp, 600 bp and 1600 bp in size, containing conserved splice sites.
Screening of the same EMBL3 CHO genomic library with the CHO hsp 10 eDNA also
resulted in isolation of a full length processed pseudogene with -90 % identity to the eDNA.
This pseudogene lacked introns, contained a poly(A) tract, as well as various single bp
changes, additions and deletions. The upstream region of this pseudo gene was found to
contain similarity to the human LINE sequence, a DNA repetitive element. PCR amplification
ofCHO-WT genomic DNA resulted in isolation offive additional processed pseudogenes,
corresponding to the central -270 bp of the CHO hsplO eDNA. All the pseudogenes
displayed a high degree of similarity to the CHO hsp 10 eDNA sequence despite the presence
of numerous mutations. Prior to this report, pseudogenes had not been found associated with
hsp 10. The identification of these pseudogenes suggests the presence of a multi gene family
for this heat shock protein in the CHO genome.
Previously, a semi-processed pseudogene, Gel, was identified for hsp60 from CHO
cells which contained a single -87 bp intron near its 3' end (Venner eta/., 1990). From this
pseudo gene, a fragment containing the -87 bp intron was isolated for use as a probe to screen
a lambda EMBL3 CHO genomic library. This resulted in isolation of several positive clones,
two of which were purified, a -1.0 kb fragment amplified by PCR and then sequenced
revealing two additional semi-processed pseudogenes, designated .A4 and .AS. These
pseudo genes were found to be homologous to the GC 1 clone, containing many similar
mutations as well as the -87 bp intron. Utilizing CHO-WT genomic DNA, a separate PCR
amplification resulted in isolation of a -2.5 kb fragment which was partially sequenced and
found to correspond to the putative hsp60 functional gene. The fragment contained one
exon, which was identical to the CHO hsp60 eDNA in the region sequenced, and two introns
of800 bp and 1500 bp. This fragment can now provide an ideal probe for isolation ofthe
CHO hsp60 functional gene.

abstract: Thesis

abstract: Master of Science (MSc)


title: Prognosis and Management of Patients who had Trauma Necessitating Orthopedic Surgeries

contributor:
Guyatt, Gordon
Bhandari, Mohit
Busse, Jason
Thabane, Lehana
Health Research Methodology

subject:
fracture
antibiotic prophylaxis
predictors
persistent pain

abstract:
The current thesis aims to address the prognosis and management of patients who have injuries necessitating orthopaedic surgery.
	In Chapter 1 I introduce the thesis, and in Chapter 5 I offer conclusions and summarize the contribution of the work. In Chapter 5, I address the scope, rationale, key findings, limitations and implications.
	Chapter 2 is a systematic review and meta-analysis investigating the effectiveness of antibiotic prophylaxis in patients with open fracture of the extremities. The results demonstrate moderate quality evidence of an important reduction in the infection rate in patients receiving, versus not receiving, antibiotic prophylaxis. We found no difference in infection rate with longer (3 to 5 days) versus shorter (1 day) duration of antibiotics – this finding warrants only low confidence.
	Chapter 3 is a systematic survey of current practice and recommendations regarding antibiotic prophylaxis in open fracture management. Authors of publications over the last decade strongly support early systemic antibiotics prophylaxis for patients with open fractures of extremities. In practice, most used systemic antibiotics with both gram-positive and gram-negative coverage, and continued the administration for 2 to 3 days. Most recommendations suggested gram-positive coverage for less severe injuries, and administration duration of no more than 3 days (half suggested 1 day). For more severe injuries, most recommendations suggested broad antimicrobial coverage continued for 2 to 3 days.
	Chapter 4 is a longitudinal study investigating predictors of persistent post-surgical pain after tibia fracture. We found significant independent associations between resolution of pain and male sex, non-smoking and alcohol consumption. Age, obesity, type of fracture (closed versus open), additional injuries, and post-operative weight-bearing status did not predict resolution of pain. Our findings suggest that clinicians should be particularly alert to the possibility of troublesome post-operative pain in female smokers who do not drink alcohol. Clinicians may consider counselling patients to discontinue smoking, inform them that they are at nearly double the risk of incidence of troublesome post-operative pain (in addition to the long-term adverse health consequences of smoking).

abstract: Thesis

abstract: Doctor of Philosophy (PhD)

abstract: Antibiotic prophylaxis reduces infection with 10% fewer event rate than the group without antibiotic prophylaxis (low to moderate confidence in estimates). The optimal antibiotic regimens and duration remain uncertain. There is a higher risk of persistent post-surgical pain in female smokers who do not use alcohol, following tibia fractures.


title: Doubling Albumin: In Vivo Consequences of Reiterating Albumin in a Single Polypeptide Chain

contributor:
Sheffield, W.P.
Biology

subject:
albumin
polypeptide
in vivo

abstract: Objective. Albumin is an abundant and slowly-cleared plasma protein. Our laboratory previously incorporated albumin into recombinant fusion proteins to extend the plasma half-life of small proteins of potential therapeutic utility. We sought to determine if reiteration of albumin in a single polypeptide chain would further extend its half-life. Design. Hexahistidine-tagged rabbit serum albumin (RSA) with Cys 34 altered to Ala to prevent disulphide-bonded dimerization was produced in yeast (H₆-RSA-C34A), and compared to a reiterated forma of H₆-RSA-C34A containing two domains of amino acids 1-584 separated by a hexaglycine spacer. Clearance of these purified iodinated proteins was compared in rabbits. Materials and Methods. Site-directed mutagenesis employing PCR was used to alter the encoding plasmid. Proteins secreted from transformed Pichia pastoris yeast cell lines were purified using nickel-chelated affinity chromatography and radioiodinated by the Iodogen method. Labeled proteins were injected intravenously into rabbits and the residual acid-precipitable protein concentration in serial plasma samples was determined over time. Results. P. 𝘱𝘢𝘴𝘵𝘰𝘳𝘪𝘴 cells transformed with the expression plasmids secreted 140 kDa H₆-diRSA and 70 kDa H₆-RSA-C34A proteins, which were purified to apparent homogeneity. Mean terminal catabolic half-lives (±SD) were 4.9 (±0.7) and 3.0 (±0.3) days for H₆-RSA-C34A and H₆-diRSA, respectively. Then values were 9 for H₆-diRSA and 12 for H₆-RSA-C34A. conclusions. H₆-diRSA was cleared from the circulation more rapidly than H₆RSA-C34A. We hypothesized that increased catabolism of the reiterated molecule could be due to an increased rate of cellular uptake and endocytosis of H₆-diRSA due to increased avidity for cellular binding sites. Non-reiterated albumin therefore appears to be optimal as a carrier protein for small recombinant blood products.

abstract: Thesis

abstract: Master of Science (MSc)


title: PEA3 Subfamily Transcriptional Activation of Osteopontin, A Transformation-Associated Protein

contributor:
Hassell, J. A.
Biology

subject: PEA3 transcription osteopontin protein

abstract: PEA3, ERM, and ER81 comprise a subfamily of ETS transcription factors that upregulate genes correlated with an increased metastastic potential of tumors. In mouse embryo fibroblast (MEF) cells, PEA3 is required for transformation by activated Ras or Neu, but the means by which PEA3 mediates Ras-transformation is not clear. Osteopontin (OPN) expression is induced upon B-ras-transformation and purified PEA3 can bind the OPN promoter by gel-shift analysis. In this study, OPN expressed higher transcript levels in the wildtype MEF 4 cell line than the PEA3 null MEF 1 cell line and was further characterized as a potential PEA3 target gene by Northern blot analyses and transient transfection studies. Northern blot analyses of 4 wildtype MEF (4, 100, 101, 104), 5 FEA3 null MEF (1, 115, B5, B10, B12), and 5 MEF 1 retransformant cell lines that stably reexpress PEA3 showed a good correlation between OPN and ERM transcript levels in 9/11 cell lines although at least 2 PEA3 subfamily members were coexpressed in 8/11 cell lines that expressed high OPN transcript levels. This suggested that the PEA3 subfamily additively regulated OPN and that ERM protein was more abundant than PEA3 and ER81 protein levels in the MEF cell lines. The relative PEA3 subfamily protein levels remain to be clarified. Transient transfection assays in the HEK 293-1 C cell line indicated that the OPN promoter was responsive to PEA3 and that the promoter region between -258 to -88 was required for maximal OPN promoter activity. There are 16 candidate core ETS binding sites in the -777 /+79 OPN promoter which could be responsible for PEA3 subfamily transactivation. The OPN promoter was more active in the MEF 4 cell line than the MEF 1 cell line, corresponding to their relative number of expressed PEA3 subfamily members. Ectopic expression of dominant negative PEA3 suppress ed OPN promoter activity in the MEF 4 cell line. Furthermore, ectopic expression of PEA3, ERM, or ER81 increased OPN promoter activity in the MEF 1 or COS-1 cell line. Thus OPN is transcriptionally regulated by the PEA3 subfamily and represents a target gene that can mediate the progression of tumor cells.

abstract: Thesis

abstract: Master of Science (MSc)


title: Identification andCharacterization of a Zinc Finger Transcription Factor That Interacts with the Cell Cycle Regulator Host Cell Factor-1

title: Identification and Characterization of a Transcription Factor

contributor:
Capone, John
Biochemistry

subject: zinc transcription factor-1

abstract: Host cell factor-1 (HCF-1) is an evolutionarily conserved protein first discovered through its interaction with the herpes simplex viral transactivator VP16. Both the amino-terminal VP 16 interaction domain and the adjacent, less studied basic domain of HCF-1 are essential in cell cycle regulation. However, the mechanism(s) of this regulation is unknown. Our objective was to provide insight into the importance of the basic domain by studying proteins that interact with this region. Using the yeast two-hybrid system with the HCF-1 basic region as bait, we identified a novel protein named HCF-1 interacting zinc finger protein (HIZ). The purpose of this research was to characterize HIZ and determine if its interaction with HCF-1 could reveal a novel cellular target for HCF-1. A putative HIZ full-length sequence was determined and its primary structure was studied in detail. HIZ contains 16 C₂H₂ zinc fingers in its amino terminal. HIZ also contains a novel glutamine-rich motif adjacent to a potent autonomous transactivation domain. The presence of a DNA-bincing domain structure and a strong, functional transactivation domain indicate that HIZ is a novel transcriptional factor. Northern blot analysis showed tissue specific expression of HIZ with highest levels in the testis, skeletal muscle, liver and pancreas, suggesting possible roles in spermatogenesis, differentiation and metabolism. The HIZ-HCF-1 interaction was verified 𝘪𝘯 𝘷𝘪𝘵𝘳𝘰 using glutathione-s-transferase (GST) pull-down assays and the minimal HIZ domain for HCF-1 interaction was mapped to the same region critical for transactivation. Recent studies have identified the transcription factors Miz-1, GABP, LZIP, Zhangfei and PGC-1β as proteins that interact with the two domains of HCF-1 important in cell cycle control. Also, these studies have shown that interaction with HCF -1 is important in regulation of their transactivation potential. Thus, the effect of HCF-1 on HIZ activation was studied using transient transfection assays. Similar to its effect on the known cell cycle inhibitor, Miz-1, our studies showed that cotransfection with HCF-1 significantly inhibited HIZ transactivation. The expression pattern of HIZ in a matched tumour/normal tissue array showed that HIZ is expressed at a significantly lower level in tumours of the lung and uterus in comparison to normal tissues from the same individual. This finding suggests a correlation between HIZ expression and tumour formation, possibly in conjunction with the known cell cycle regulator HCF -1.

abstract: Thesis

abstract: Master of Science (MSc)


title: Changes in Muscle Protein Synthetic Rate and Ultrastructure Following Resistance Exercise

contributor:
MacDougall, J. D.
Human Biodynamics

subject: muscle protein resistance exercise

abstract: The purpose of this study was to correlate the extent of myofibrillar disruption with muscle protein synthetic rate (MPS) following an isolated bout of concentric or eccentric elbow flexor resistance exercise. Six strength-trained males performed 8 unilateral sets of 8 repetitions at 80% concentric 1RM. The absolute amount of work performed by each arm was controlled by having both the concentrically-exercised (CON) and eccentrically-exercised (ECC) arms lift or lower the same weight through the same range and magnitude. Biopsies from biceps brachii of each arm, extracted ~21 h post -exercise, were analyzed electron microscopically to quantify myofibrillar disruption. The severity of disruption was classified as focal (FOC), moderate (MOD), or extreme (EXT). MPS of both arms was calculated from the increment in L- [1,2-¹³C₂] leucine abundance in biopsy samples relative to the mean plasma [1, 2-¹³C₂] -α-KIC enrichment at isotopic plateau using the primed-constant infusion technique over ~10 h so that the midpoint of the assessment period was ~24 h post-exercise. The severity of disruption was significantly (P < 0.001) greater in both the FOC (11.2%) and MOD (12.2%) compared to the EXT (1.6%) rating. Absolute disruption of fibers was significantly greater (P = 0.007) in the ECC (44.7%) as compared to baseline (BASE) samples (3.9%), obtained following 5 d where no arm training had occurred. In addition, ECC samples showed ~40% greater total disruption than CON samples (44.7% vs. 26.7%). Despite this, a positive correlation (r = 0.89) was found between individual values for MPS and the percentage of disrupted fibers in tissue from the ECC but not CON arm. These findings indicate that, in strength-trained males, residual myofibrillar disruption from a previous training session is essentially repaired within 5 d, but that resistance exercise induced muscle damage did not appear to provide the activating signal for elevating MPS between -21-29 h post-exercise.

abstract: Thesis

abstract: Master of Science (MSc)


title: Characterization of the AF-1 Activity of mPPARa and mPPARy2

subject: AF-1

subject: mPPARa

subject: mPPARy2

subject: transcription

abstract:
Peroxisome Proliferator-Activated Receptors (PPARs) belong to the nuclear receptor superfamily of transcription factors. PPARs, like some other nuclear receptors, bind their cognate DNA binding sites as heterodimers with Retinoid X Receptor (RXR). Like most members of this superfamily, PPARs regulate transcription of target genes in response to specific lypophilic ligands. Given the fact, however, that many members of the nuclear receptor superfamily possess a ligand independent activation function (AF -1) in their amino-terminal A/B domains, we identified and characterized the AF-1 activity in the A/B domains of mPPARα and mPPARγ2. When fused to the GAL4 DNA Binding
Domain (DBD), the A/B domains of both mPPARα and mPPARγ2 subtypes are transcriptionally active both in mammalian and yeast cells in the absence of cognate PPAR activators, indicating that both PPAR subtypes do in fact contain AF -1 activities in their respective A/B domains. Our deletion studies localizing the region in the A/B domains of the PPAR subtypes indicate that the amino-terminal region of A/B domains of both PP AR subtypes is necessary, but not sufficient, for the AF -1 activity. After having identified and characterized the AF-1 activities of both mPPARα and mPPARγ subtypes, we proceeded to investigate how this activity is controlled. Generally, nuclear receptors, including PPARs, do not activate transcription when ligand is absent, suggesting that the ligand-independent AF -1 activity is somehow masked. We used the GAL4 DBD to investigate this phenomenon. When fused to GAL4 DBD, the full-length mPPARα receptor is not active in mammalian cells in absence of ligand despite the presence of the AF -1 activity in the A/B domain. Only when the PPAR activator Wy-14 is added, does the receptor become active. Carboxy-terminal deletion
of the fusion at the junction of the DBD (C domain) and the Hinge region (D domain)
results in partially active protein whose activity does not depend on the presence of ligand, suggesting a role of the hinge region in the masking of the AF-1 activity. The transcriptional activity of PPARs is ligand independent when PPARs are expressed as GAL4 DBD fusions or allowed to bind reporter genes as dimers with RXR. Yeast do not possess nuclear receptors or any homologues of known accessory proteins which interact with nuclear receptors. Given our observations and those of others made in yeast and mammalian cells, we suggest that the masking of the AF -1 activity occurs in the inactive state of the receptor, and that mammalian cells contain unidentified factor(s)
which is able to maintain PPARs in the inactive state, thus enabling the masking of the AF-1 activity.
Nuclear receptors including PPARs are known to interact with a host of accessory proteins which modulate their activity and are thought to act as a bridge between the receptors and the basal transcription machinery. One of these proteins is RIP140 whose function has not been determined. Our studies suggest that RIP 140 is a co-activator for the PPAR/RXR heterodimer, as well as for the AF-1 activity of mPPARα and mPP ARγ2.

abstract: Thesis

abstract: Master of Science (MSc)


title: Affinity Chromatography using Concatemeric Functional Nucleic Acids for Biosensing

contributor:
Brennan, John
Chemistry and Chemical Biology

subject:
DNA
Functional nucleic acids
Affinity chromatography
sol-gel

abstract: This thesis describes the use of functional nucleic acid (FNA) superstructures entrapped within monolithic macroporous sol–gel-derived silica for solid-phase flow-based sensing of small molecules and macromolecular proteins. The work described herein overcomes a long-standing issue with entrapment of biomolecule into sol–gel-derived materials; the mesoporous pore morphology required to retain entrapped biomolecules prevents detection of large analytes as these can’t access the entrapped species. It is shown that large DNA superstructures can be produced through rolling circle amplification of a functional nucleic acid, resulting in concatemeric FNA species with dameters of several microns.  Such species can be entrapped within macroporous sol-gel derived materials with micron-sized pores with minimal leaching, thus allowing for detection of a wide range of molecules, including biomolecules. Optimal materials for  entrapment of FNA superstructures was achieved using a high-throughput material screening method, which minimized biomolecule leaching while maintaining FNA activity. Using an optimized material, concatemeric aptamer superstructures were entrapped within macroporous monolithic columns for flow-based detection of small molecules and proteins, extending the range of analytes that can be analyzed using biohybrid monolithic columns. Preliminary studies on the formation and properties of a DNAzyme superstructure for detection of E. coli detection were also performed, which  provided valuable information on factors that must be controlled to allow reproducible fluorescence-based detection of E. coli using the crude intracellular matrix as the target.

abstract: Thesis

abstract: Master of Science (MSc)


title: Additive Manufacturing of Hydrogels for Vascular Tissue Engineering

contributor:
Selvaganapathy, Ponnambalam Ravi
Biomedical Engineering

subject: Tissue Engineering, Hydrogels, 3D Printing, Additive Manufacturing, Vascularization, Microfluidics, Microchannels, Extrusion, Lamination, Adhesive, Coaxial

abstract:
One of the major technical challenges with creating 3D artificial tissue constructs is the lack of simple and effective methods to integrate vascular networks within them. Without these vascular-like networks, the cells embedded within the constructs quickly become necrotic.
 This thesis details the use of a commercially available, low-cost, 3D printer modified with a microfluidic printhead in order to generate instantly perfusable vascular-like networks integrated within gel scaffolds seeded with cells. The printhead featured a coaxial nozzle that allowed the fabrication of hollow, gel tubes (500µm–2mm) that can be easily patterned to create single or multi-layered constructs. Media perfusion of the channels caused a significant increase in cell viability.
This microfluidic nozzle design was further modified to allow for multi-axial extrusion in order to 3D print and pattern bi- and tri-layered hollow channel structures. Most available methodologies lack the ability to create multi-layered concentric conduits inside natural extracellular matrices, which would more accurately replicate the hierarchal architecture of biological blood vessels. The nozzle used in this work allowed, for the first time, for these hierarchal structures to be embedded within layers of gels in a fast, simple and low cost manner. This scalable design allowed for versatility in material incorporation, thereby creating heterogeneous structures that contained distinct concentric layers of different cell types and biomaterials.
This thesis also demonstrates the use of non-extrusion based 3D biofabrication involving planar processing by means of hydrogel adhesion. There remains a lack of effective adhesives capable of composite layer fusion without affecting the integrity of patterned features. Here, silicon carbide was found for the first time to be an effective and cytocompatible adhesive to achieve strong bonding (0.39±0.03kPa) between hybrid hydrogel films. Multi-layered, heterogeneous constructs with embedded high-resolution microchannels (150µm-1mm) were fabricated in this way.
With the new 3D fabrication technology developed in this thesis, gel constructs with embedded arrays of hollow channels can be created and used as potential substitutes for blood vessel networks as well as in applications such as drug discovery models and biological studies.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)

abstract: Additive manufacturing (AM) involves any three-dimensional (3D) fabrication technologies that is used to produce a solid model of a predetermined design. AM techniques have recently been used in tissue engineering applications for fabrication of 3D artificial tissues that resemble architectures and material properties similar to that of the native tissue. Utilizing AM for this purpose presents the advantage of increased control in feature patterning, which leads to the realization of more complex geometries. However, there still remains a lack of simple and effective methods to integrate vascular networks within these 3D artificially engineered scaffolds and tissue constructs. Without these vascular-like networks, the cells embedded within the constructs would quickly die due to a lack of nutrient delivery and waste transport. This remains one of the biggest challenges in true 3D tissue engineering. This thesis presents a number of fast, effective and low-cost AM biofabrication techniques to address this challenge.


title: Microsomal Ferritin Iron Release in Relation to TCDD Toxicity

contributor:
Sweeney, G.
Biochemistry

subject:
iron
TCDD poisoning
ferritin
microsomes

abstract: Previous experiments have demonstrated that the hepatoxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) involves synergism with iron. It has been postulated that it is storage iron that plays a role in toxic effects observed with TCDD poisoning. It was hypothesized that TCDD induction of the Ah gene locus in some way leads to mobilization of storage iron from the iron storage protein ferritin. Microsomal ferritin iron release was investigated in intact microsomes and in a reconstituted system. Evidence is presented which suggests that NADPH-cytochrome c (P450) reductase is capable of effecting the release of iron from flavin (FMN), and that at the same time, the reduced flavin thus generated is capable of effecting the release of iron from ferritin. While no direct evidence was obtained, the results do lend support to the hypothesis that TCDD toxicity could result from the mobilization of iron from ferritin by the TCDD induced microsomal electron transport system.

abstract: Thesis

abstract: Master of Science (MSc)


title: Quantification of the Activities of the Anti-Apoptotic Proteins BCL-2 and BCL-XL

title: Quantification of the Activities of BCL-2 and BCL-XL

contributor:
Andrews, David
Biochemistry

subject: anti-apoptotic proteins BCL-2 BCL-XL

abstract:
Apoptosis is the process by which organisms eliminate excess, damaged or hazardous cells in a controlled manner. This process is controlled by the Bcl-2 family of proteins. Bcl-2 and Bcl-XL are anti-apoptotic paralogues that can replace CED-9, the sole homologue in C. elegans. It has therefore been assumed that Bcl-2 and Bcl-XL are replaceable and functionally identical. However, evidence in some mammalian cells indicates that this may not be the case. The purpose of this project was to exhaustively compare the anti-apoptotic activities of Bcl-2 and Bcl-XL in one cell type. As Bcl-2 and Bcl-XL have been found to localize to the ER and the outer mitochondrial membrane, we also determined whether subcellular location affects the function of these proteins differently. MCF-7 breast cancer cells were stably transfected with Bcl-2 and Bcl-XL alternatively targeted to the ER or mitochondria, and exposed to various doses of
doxorubicin; PARP cleavage was measured using quantitative Western blotting as an indication of apoptosis to obtain EC₅₀ values in the different cell lines. The levels of both Bcl-2 and Bcl-XL affected anti-apoptotic activity; specific degradation of both proteins was noted at higher doses of doxorubicin. Nevertheless, the results indicated clearly that there was a difference between Bcl-2 and Bcl-XL. Using EC₅₀ values Bcl-XL mutants were at least 8 times more protective than Bcl-2 mutants. Furthermore, most of the cleavage products of PARP in Bcl-XL expressing clones were due to non-caspase-7 proteases, a pattern not seen with Bcl-2. Bcl-2 and Bcl-XL localized to mitochondria were most effective, while cytosolic and ER localized Bcl-XL were less effective, and Bcl-2 at these sites did not inhibit apoptosis.

abstract: Thesis

abstract: Master of Science (MSc)


title: High-Resolution Characterization of Nitrogen-Doped Carbon Support Materials Decorated with Noble Metal Atom Catalysts

contributor:
Botton, Gianluigi A
Materials Engineering

subject:
graphene
N-dopants
Single atom catalysts
carbon nanotubes
Electron microscopy
EELS
TEM
STEM

abstract:
Graphene and its functionalized derivatives, such as nitrogen-doped graphene, have recently become a popular substrate material for the proton exchange membrane fuel cell (PEMFC) due to its enhanced electrical conductivity, electrochemical stability, and increased surface area when compared to the conventional, carbon black. In order to further develop the alternative fuel industry, the Pt catalyst within the PEMFC must also be considered. Single Pt atoms have a higher surface area to volume ratio when compared to nanoparticles, thus offering the potential to create a more affordable and efficient PEMFC. In this thesis, electrode materials comprising single Pt atoms and clusters, produced using atomic layer deposition (ALD) on various C derivatives, including graphene, N-doped graphene, carbon nanotubes (CNTs), and N-doped CNTs (NCNTs) are investigated through the utilization of aberration corrected transmission electron microscopy.
Structural and chemical analysis was performed on thermally exfoliated N-doped graphene and CVD-produced graphene that was exposed to N+ ion sources. It was determined that the thermally exfoliated N-doped graphene maintained the short-range order of the graphene lattice; however, local inhomogeneities existed for the total N concentration, and the specific N-dopants within and between graphene sheets. More importantly, Pt atoms and clusters were observed and determined to be primarily stabilized at the edge of the N-doped graphene sheets. The stabilization of the Pt atoms and clusters resulted in a significantly higher mass and specific activity for the hydrogen evolution reaction, when compared to the use of a graphene substrate and Pt nanoparticles on C black. The N+ ion implantation in the CVD graphene showed the incorporation of N-dopants; however, electron energy loss spectroscopy revealed structural damage to thin sheets.
NCNTs were also characterized in this thesis as possible gas containers, and as a substrate material to examine the effects of varying ALD conditions. It was determined that the NCNTs were an effective N2 gas conduit, wherein a decreasing pressure was observed with an increase to the inner diameter of the nanotubes. Using similar NCNTs, the effect of dosing time, temperature, and substrate on the Pt size were analyzed using ALD. While no singular condition resulted in the sole production of single Pt atoms, modifying both the substrate and dosing time were shown to provide the greatest potential for producing individual Pt atom catalysts.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)


title: Issues regarding the sharing of interim results by the Data Safety Monitoring Board of a trial with those responsible for the conduct of the trial.

contributor:
Thabane, Lehana
Buckley, Norman
Paul, James
Mbuagbaw, Lawrence
Health Research Methodology

subject:
Data Safety Monitoring Boards
Data Monitoring Committee
Interim data sharing
Clinical Trial

abstract:
Background and Objectives: Sharing of interim results by the Data Safety Monitoring Board
(DSMB) with non-DSMB members is an important issue that can affect trial integrity. The objective of this dissertation was to determine the views of the stakeholders on what kind of interim results can or should be shared by the DSMB, why, and with whom among those responsible for the conduct of a trial.

Methods:
We first conducted a systematic search of the literature to assess views and current evidence on sharing interim results. Secondly, we conducted two cross-sectional surveys aimed at those involved in trials to solicit their views on what type of interim results should be shared by the DSMB with non-DSMB members, with whom and under what circumstances. Thirdly, we assessed for any potential association of demographic factors with the sharing of certain interim results and their perceived usefulness, using regression analysis.

Results:
Mixed views exist in the literature on interim result sharing practices. Evidence from the surveys conducted resulted in the following findings.

What to share: Based upon the survey results from our cross-sectional survey (Chapter 4), the interim control event rate (IControlER), the adaptive conditional power (ACP) and the unconditional conditional power (UCP) should not be shared. Most respondents from this survey thought the interim combined event rate (ICombinedER) could be shared provided proper conditions and provisions are in place. However, based on our cross-sectional scenario-based survey (Chapter 3), it was demonstrated that the ICombinedER, when shared at interim, is compatible with three possible interim results (Drug X doing better than placebo, worse than placebo or performing the same as placebo).

Why share or not share: Respondents indicate that the ICombinedER can be shared because it does not unmask relative effects between groups, and keeps the steering committee (SC) informed about the trial’s progress; however, with the condition that sharing this type of result should be specified a priori including for what purpose and be at the DSMB’s discretion, especially if the control group rate is known from the literature. However, it is important to note that the ICombinedER, demonstrated with evidence from our cross-sectional scenario-based survey (Chapter 3), is compatible with three possible interim results and should not be shared because it has low usefulness and is flawed due to multiple interpretations. The IControlER and the ACP should not be shared because they are unmasking of interim results. It was mentioned that ICombinedER is usually known by the SC and sponsor making it easy to determine group rates if the IControlER is known. The UCP should not be shared because it is a technical measure that is potentially misleading of interim results.

With whom to share: Survey results from Chapter 4 indicated that the ICombinedER can be
shared with the SC and that the IControlER, the ACP, and the UCP should not be shared with any non-DSMB members by the DSMB. However, evidence from Chapter 3 also indicates that the ICombinedER should not be shared with any non-DSMB member.

Factors associated with sharing: Having experience with greater than 15 trials with private industry sponsorship was found to be associated with not sharing the IControlER and an increase in perceived usefulness in sharing the ACP. Though some other demographic factors were found to be associated with sharing the ICombinedER and the UCP, they were sensitive to missing data upon our sensitivity analysis and will require more validation.

Conclusions:
Though mixed views exist within an extensive literature review on interim result sharing practices, survey evidence from this dissertation suggests that the ICombinedER, IControlER, the ACP and the UCP should not be shared with any non-DSMB member. The IControlER and ACP can be unmasking of interim results and the UCP is a technical measure that is potentially misleading. We agree with this reasoning. The majority of respondents from the survey in Chapter 4 indicated that the ICombinedER can be shared with the SC because it does not unmask relative effects between groups, however it was also stipulated that sharing this measure should be specified a priori and for what purpose and be at the DSMB’s discretion, especially if the control group rate is known from the literature. Even though the majority from our second survey in Chapter 4 indicate sharing the ICombinedER with the SC, we do not recommend sharing the ICombinedER at interim with any non-DSMB member because, as demonstrated with evidence from our cross-sectional scenario-based survey in Chapter 3, this measure is compatible with three possible interim results potentially leading to the introduction of trial bias at interim by those privy this interim measure and their interpretation. Based on the findings from the survey from Chapter 4, there appears to be a lack of awareness in how sharing the ICombinedER is flawed, of low usefulness, and potentially dangerous. The perceived desire to have this measure shared seems misguided. Experience with greater than 15 trials with private industry sponsorship was found to be associated with not endorsing the sharing the IControlER and an increase in perceived usefulness in sharing the ACP by the DSMB at interim. In regards to implications for future research, this characteristic should be
further evaluated to see if this subgroup has insight into interim trial management practices that protect from trial bias.

Results from this research have implications for practice and guidelines concerning trial design and protocols, and DSMB charters. These results can also help assess the need for proper safeguards around sharing an interim result when deemed appropriate by the DSMB and under their discretion, that prevent the introduction of bias that could alter the final trial results generated.

abstract: Thesis

abstract: Doctor of Philosophy (PhD)


title: PEA3 and ER81: Roles in Transformation and Mammary Gland Development

contributor:
Hassell, John
Biology

subject: mammary

subject: development

subject: PEA3

subject: ER81

subject: transformatiion

abstract:
PEA3 is the founding member of a subfamily of closely related Ets transcriptional regulatory
proteins that includes ERM and ER81. The PEA3 subfamily members share greater than 95% identity in their ETS DNA binding domain and 500/o sequence similarity overall, suggesting these genes may serve redundant functions. The overexpression of each member is positively correlated with HER2 mediated breast tumorigenesis in humans and mice, suggesting a role for this subfamily in mammary development and oncogenesis. This study first addresses the role of PEA3 in cellular transformation mediated by oncogenic Ras and Neu. Wildtype and PEA3-null mouse embryo fibroblast cell lines were infected and tested for focus formation. PEA3-null fibroblasts are refractory to transformation as compared to their wildtype counterparts. Ras and Neu transformed foci show elevated PEA3 subfamily mRNA transcripts and PEA3 protein. ERM and ER81 are expressed in PEA3-null fibroblasts and do not appear to compensate for loss of function mutations in the PEA3 gene resulting in the transformation-defective phenotype. Expression of candidate PEA3 target genes (MMP-3 and MMP-9, which have known roles in transformation) is compromised in PEA3-null fibroblasts. Re-expression of PEA3 in these cells rescues the transformation-deficient phenotype and restores expression of MMP-3 and MMP-9. Hence, PEA3 appears to be a crucial effector in Ras and Neu mediated transformation, in addition to serving an important regulatory role of genes involved in cell motility and invasive tumor behaviour. This study also addresses the role of ER81 in normal mammary gland development. PEA3 is required for normal mammary gland development, as displayed by the reduced branching phenotype in PEA3-null female mice. Mice lacking functional ER81 were generated to determine if ER81 serves a similar role in mammary gland development. ER81 is expressed in the epithelial cells of mammary buds at E 1 0.5, when these structures first appear during mouse embryogenesis. ER81 is then differentially expressed during postnatal mammary gland development, with highest expression occurring at times of extensive epithelial branching. During puberty, expression is observed in undifferentiated cap and body cells of terminal end buds, in differentiated luminal and myoepithelial cells of ducts. During pregnancy, expression in luminal epithelial cells is lost, but persists in the myoepithelial cells within the ducts and alveoli. Targeted disruption of both ERSt alleles result in severely runted mice that die by 4 weeks of age, thereby precluding study of mammary gland development in these mice beyond this developmental stage. However, loss of a single ER81 allele results in healthy looking mice, comparable in size and lifespan to wildtype littermates. Studies employing ER81 heterozygous mice reveal a 50% allelic dose is sufficient for normal mammary gland development. Loss of a single ER81 allele did not result in any overt phenotypes in ductal branching, lobulo-alveolar development, or morphology of the surrounding fat pad.

abstract: Thesis

abstract: Master of Science (MSc)