Heart failure (HF) is a complex, debilitating disease that affects approximately 6.5 million Americans (Ades et al., 2013). HF is a large reason for hospital readmissions, and subsequently, a major contributor to rising health care costs. Unfortunately, there is no cure for HF, but various interventions such as cardiac rehabilitation (CR) have been employed to help patients manage the symptoms. However, the lack of patients ever being referred to cardiac rehabilitation is disturbing. Healthcare providers play an essential role in providing education about heart failure and CR, and thus should be knowledgeable about these principles themselves. Therefore, the aim of this project is to evaluate data from a survey sent to nurse practitioners (NPs) regarding whether HF patients are being referred to CR appropriately, and if barriers are limiting use of CR programs. This was a descriptive, nonexperimental study with a survey design seeking to understand if NP providers are following evidence-based guidelines when treating HF patients and if providers consider CR programs to be an appropriate treatment tool. A survey questionnaire was distributed to NPs who are members of Coalition of Arizona Nurses in Advanced Practice (CAZNAP). Data from 27 surveys were used for analysis. Results showed that nurse practitioner respondents felt they had a good understanding of heart failure education and diagnosis. A majority also considered CR to be a useful tool for HF patients, and all agreed that health care providers affect CR enrollment and participation rates. A mere nine respondents (33%) reported being introduced to the outcomes and benefits of CR in their graduate education. Findings also confirmed previous work suggesting that providers are not adequately referring HF patients to CR, as 33% of NPs reported they have never referred a patient to CR. With respect to these findings, it is important to identify methods to assist providers with proper education about CR and its referral methods. As supported by the literature review, improved referral rates to CR can lead to better management and health outcomes for HF patients. Therefore, further research is needed to identify interventions that promote increased CR referral rates.

Both hypertrophic (HCM) and dilated cardiomyopathy (DCM), though differing in their aetiologies, share features of impaired resting energetics. The aim of this thesis was to determine if cardiac high energy phosphate metabolism, measured as the phosphocreatine (PCr)/ATP ratio using 31Phosphorus magnetic resonance spectroscopy (31P MRS), is further impaired during exercise in these pathologies. This would provide a possible explanation for the high incidence of exercise related death in HCM and DCM as well as the blunted inotropic response to exercise in DCM. Furthermore, this thesis investigates the role of stress perfusion and stress tissue oxygenation in HCM (as these are hypothesized to exacerbate the primary defect in energetics) and exercise training in DCM (which is hypothesized to improve function though the mechanisms are uncertain). This work developed a novel protocol for measuring 31P MRS in a clinically acceptable time frame. The traditional acquisition is at least 20 minutes (as much as 40 minutes in subjects with lower pulse rates). This is a particularly long time to allow for exercise in the magnet particularly in the symptomatic DCM cohort. Hence this work meticulously developed a shorter 8 minute protocol. Its validity, reproducibility and application to exercise were confirmed. The post processing of the MRS data was further improved for calculating blood contamination and tested with both simulated and patient data, including normal, hypertrophied and thinned myocardium. Applying this new method, this thesis is the first to report a further decrease in exercise energetics in HCM. The relationship between perfusion, tissue de-oxygenation and energetic compromise during exercise was then explored in HCM. Athletes, with physiological hypertrophy, were used as an additional control group in these experiments. These results demonstrated a strikingly blunted oxygenation response of the HCM heart to stress even in the pre-hypertrophy HCM mutation carriers. However, as a group, the data did not show a correlation between the blunted oxygenation response and the percentage change in PCr/ATP during exercise. None-the-less, these results can potentially be useful for distinguishing between hypertrophy in the athletes and pathological hypertrophy in HCM and for distinguishing HCM mutation carriers’ pre hypertrophy and the normal heart. In the DCM cohort, this thesis explored the impact of exercise training on cardiac metabolism and function. The results showed no change in cardiac energetics and left ventricular ejection fraction during 8 minutes of exercise. In addition, an eight week home exercise programme did not alter resting or exercise cardiac PCr/ATP, but improved cardiac function during rest and exercise, and increased exercise tolerance and quality of life scores. In conclusion, this thesis reports further insights into cardiac exercise energetics in HCM and DCM and its relationship to perfusion and oxygenation in HCM and to exercise training in DCM. Therapies that decrease the energy cost of cardiac work during exercise may prove beneficial targets to explore further in these conditions.

The principal substrate used by the normal adult human heart is free fatty acids, the remainder being, predominantly,carbohydrate. During failure, the heart becomes less reliant on fatty acid metabolism, possibly as a result of tissue hypoxia. Therefore, understanding hypoxic adaptation may explain the metabolic changes that occur during the development of heart failure.As peroxisome proliferator activated receptor alpha (PPARα) modulates cardiac fatty acid metabolism, the work in this thesis focused on the role of PPARα in cardiac metabolic adaptation to chronic hypoxia. It was found that isolated hearts from chronically hypoxic (11% O₂ for 3 weeks)mice were more glycolytic, had reduced PPARα expression and decreased fatty acid metabolism,but had normal function, determined using in vivocine-MRI. ³¹P MRS of isolated perfused mouse hearts showed a drop in PCr with hypoxia, but ΔG_ATP was not altered, indicating that metabolic reprogramming was sufficient to maintain ATP production and contractile function. Increased or decreasedPPARα expression, using a high fat diet or PPARα null mice, respectively, prevented metabolic adaptation to hypoxia and caused cardiac dysfunction. Hypoxia with high fat feeding was particularly deleterious, reducing ejection fraction by 9%,possibly due to increased mitochondrial uncoupling. PPARβ/δ and γ were not involved in the adaptation to hypoxia, and none were modified by PPARα stimulation or ablation. Cardiac VEGF and PDK1, prominent hypoxia-inducible factor (HIF) targets, were increased by hypoxia, indicating that HIF may have been involved in metabolic adaption. However, high fat feeding prevented VEGF accumulation during hypoxia, suggesting that impaired HIF signalling may have contributed to the maladaptive response to hypoxia. In order to determine the relationship between HIF and PPARα, HIFwas stabilised pharmacologically using FG2216/BIC in HL-1 cardiomyocytes, to show decreased PPARα expression and caused similar metabolic changes to those seen in the in vivo hypoxic heart. In conclusion, this study demonstrated that HIF downregulation of PPARα is crucial for metabolic adaptation and maintenance of cardiac function during chronic hypoxia. Similar metabolic changes that occur in end-stage heart failure may also be a response to increasing hypoxia.

The aim of this thesis was to describe and explore how the partner relationship of patient–partner dyads isaffected following cardiac disease and, in particular, atrial fibrillation (AF) in one of the spouses. The thesis is based on four individual studies with different designs: descriptive (I), explorative (II, IV), and cross-sectional (III). Applied methods comprised a systematic review (I) and qualitative (II, IV) and quantitative methods (III). Participants in the studies were couples in which one of the spouses was afflicted with AF. Coherent with a systemic perspective, the research focused on the dyad as the unit of analysis. To identify and describe the current research position and knowledge base, the data for the systematic review were analyzed using an integrative approach. To explore couples’ main concern, interview data (n=12 couples) in study II were analyzed using classical grounded theory. Associations between patients and partners (n=91 couples) where analyzed through the Actor–Partner Interdependence Model using structural equation modelling (III). To explore couples’ illness beliefs, interview data (n=9 couples) in study IV were analyzed using Gadamerian hermeneutics. Study I revealed five themes of how the partner relationship is affected following cardiac disease: overprotection, communication deficiency, sexual concerns, changes in domestic roles, and adjustment to illness. Study II showed that couples living with AF experienced uncertainty as the common main concern, rooted in causation of AF and apprehension about AF episodes. The theory of Managing Uncertainty revealed the strategies of explicit sharing (mutual collaboration and finding resemblance) and implicit sharing (keeping distance and tacit understanding). Patients and spouses showed significant differences in terms of self-reported physical and mental health where patients rated themselves lower than spouses did (III). Several actor effects were identified, suggesting that emotional distress affects and is associated with perceived health. Patient partner effects and spouse partner effects were observed for vitality, indicating that higher levels of symptoms of depression in patients and spouses were associated with lower vitality in their partners. In study IV, couples’ core and secondary illness beliefs were revealed. From the core illness belief that “the heart is a representation of life,” two secondary illness beliefs were derived: AF is a threat to life, and AF can and must be explained. From the core illness belief that “change is an integral part of life,” two secondary illness beliefs were derived: AF is a disruption in our lives, and AF will not interfere with our lives. Finally, from the core illness belief that “adaptation is fundamental in life,” two secondary illness beliefs were derived: AF entails adjustment in daily life, and AF entails confidence in and adherence to professional care. In conclusion, the thesis result suggests that illness, in terms of cardiac disease and AF, affected and influenced the couple on aspects such as making sense of AF, responding to AF, and mutually incorporating and dealing with AF in their daily lives. In the light of this, the thesis results suggest that clinicians working with persons with AF and their partners should employ a systemic view with consideration of couple’s reciprocity and interdependence, but also have knowledge regarding AF, in terms of pathophysiology, the nature of AF (i.e., cause, consequences, and trajectory), and treatments. A possible approach to achieve this is a clinical utilization of an FSN based framework, such as the FamHC. Even if a formalized FSN framework is not utilized, partners should not be neglected but, rather, be considered a resource and be a part of clinical caring activities. This could be met by inviting partners to take part in rounds, treatment decisions, discharge calls or follow-up visits or other clinical caring activities. Likewise, interventional studies should include the couple as a unit of analysis as well as the target of interventions.

The purposes of this study were (1) to determine the relationship between recitation in the classroom and changes in the cardiac rate, (2) to determine the effects on cardiac rate of anticipation of recitation and tests, (3) to determine the effects on cardiac rate of compliments and assurance directed toward students by the teacher, and (4) to determine the effects on the cardiac rate of verbal threats and ridicule.

Atrial fibrillation (AF) is the most common arrhythmia encountered in clinical practice, occurring in 1% of the adult population of North America. Although AF does not typically lead to risk of immediate mortality, it is a potent risk factor for ischemic stroke. When left untreated AF reduces quality of life, functional status, cardiac performance and is associated with higher medical costs and an increased risk of death. Catheter ablation is a commonly used treatment method for those who suffer from drugrefractory AF. Prior to ablation, intra-cardiac mapping can be used to determine the activation sequence of cardiac tissue, which may be useful in deciding where to place ablation lesions. However, the electrical potential that is recorded during mapping is not a direct reflection of the current density across the tissue because the potential recorded at each point above the heart tissue is influenced by every cell in the tissue. This causes the recorded potential to be a blurred version of the true tissue current density. The potential that is observed can be described as the convolution of the true current density with a point spread function. Accordingly, deconvolution can, in principle, be used in order to improve the resolution of potential maps. However, because the number of electrodes which can be deployed transvenously is limited by practical restrictions, the recorded potential field is a sparsely sampled version of the actual potential field. Further, an electrode array cannot sample over the entire atrial surface, so the potential map that is observed is a truncated version of the global electrical activity. Here, we investigate the effects of electrode sampling density and edge extension on the ability of deconvolution to improve the resolution of measured electrical potentials within the atria of the heart. In particular, we identify the density of sensing electrodes that are required to allow deconvolution to provide improved estimation of the true current density when compared to the observed potential field.

M.A. / The study investigated the effects of cardiac arrest and myocardial infarction on long-term memory function. Given that anoxia has more serious neuropsychological ramifications than hypoxia, it was hypothesized that the cardiac arrest group would perform poorer than the myocardial infarction group in visuo-spatial and auditory-verbal recall and recognition memory. When brain insult prevails, affective changes may occur and may reflect the trauma related to the illness and partly to the cognitive dysfunction. Thus it was hypothesized that the Beck Depression Inventory scores would be significantly elevated in the cardiac arrest group. Each group consisted of 15 participants. The mean age for the cardiac arrest group and myocardial infarction group was 59.47 years (SD = 9.24) and 58.87 years (SD = 7.22), respectively. Sex, age, education, hypertension, diabetes mellitus, and smoking were controlled. However, the analysis did not reveal any significant between-group differences. There was no significant difference on the BDI, and both groups were moderately depressed, the cardiac arrest (BDI: mean score = 17.07, SD = 16.97) and myocardial infarction (BDI: mean score = 18.33, SD = 18.35). The researchers acknowledged the potential effects that beta-adrenoceptor antagonists and diuretics, and angiotensin-converting enzyme have on memory and cognitive performance, respectively. However, the analysis did not reveal a significant between-group difference for these variables. The neuropsychological test battery comprised: Rey-Auditory Verbal Learning Test (RAVLT), Rey-Osterreith Complex Figure Test (ROCFT), Wechsler Memory Scale-Revised (WMS-R), Wechsler Adult Intelligence Test (WAIS) Symbol Search and Digit Symbol Substitution Test, Raven’s Standard Progressive Matrices, and the Oral Word Controlled Test (FAS). The memory function of the cardiac arrest group was characterized by deficits in visuo-spatial and auditory-verbal recall and recognition memory. In addition, the retention intervals were not mediating factors. This group was also impaired in visuo-spatial perception, constructional and organizational ability, and psychomotor speed. The impairment that characterized the myocardial infarction group converged on all auditory-verbal attentional tasks, indicating that this group has a selective impairment in auditory-verbal attention. Moreover, both groups exhibited equal levels of impairment in orientation, and uniform performance in executive function and verbal fluency. The memory function after cardiac arrest is characterized by deficits in visuo-spatial and auditory-verbal deficits in recall and recognition memory as well as impairment in visual perception, constructional ability, and psychomotor speed. By contrast, myocardial infarction patients are specifically impaired in auditory-verbal attention.

"Diabetic cardiovascular autonomic neuropathy (DCAN) is common in patients with diabetes mellitus, and causes abnormalities in heart rate control as well as central and peripheral nervous system dynamics. A good understanding of DCAN is not established yet. An effective way to detect diabetes mellitus at an early stage is still undiscovered, which method is highly desired by researchers and patients. One reason why the pathogenesis of DCAN is unclear is that non- invasive assessment of DCAN in humans and animals has been problematic. The non-stationary and non- linear natures of the interested physiological signals have placed great limitation on traditionally algorithms. To overcome this limitation, this work proposes a series of time- varying, nonlinear and non-invasive methods to assess cardiac autonomic dysregulation from ECG and PPG records. Including a non-stationary method called PDM, which is based on principal dynamic mode (PDM) analysis of heart rate variability (HRV), nonstationary power spectral density called TVOPS-VFCDM and also standard spectrum analysis method of HRV. We are also able to study and analyze a series of long term and short term ECG and PPG data. In a pilot study, ECG was measured via telemetry in conscious 4 month old C57/Bl6 controls and in Akita mice, a model of insulin- dependent type I diabetes, while PPG was measured via tail pulse oximetry system from 2 month old to 4 month old. The results indicate significant cardiac autonomic impairment in the diabetic mice in comparison to the controls at 4 month old and such impairment start to present at 3 month old. Further, both immunohistochemistry and Western blot analyses show a reduction in nerve density in Akita mice as compared to the control mice, thus, corroborating our data analysis records."

There is significant variation between individual patients in the magnitude and pattern of their systemic response to cardiac surgery. Poor outcomes in these patients have been associated with a dysfunctional host response. This thesis seeks to define such variability at the level of gene expression by sequential analysis of transcription before and after surgery for a low risk group of patients undergoing elective cardiac surgery and cardiopulmonary bypass (CPB) patients using expression microarray profiling. To that aim, we analysed sequential global gene expression patterns in circulating peripheral blood leukocytes. We also investigated the role of DNA sequence variation in modulating the observed changes in gene expression. This approach allowed us to identify important genetic modulators and novel biological pathways and gain new insights into the mechanisms that regulate the host response to surgery.

Fluorescence microscopy techniques, restricted by the diffraction limit of light, have seen a remarkable advancement in recent years. An approach called dSTORM (direct stochastic optical reconstruction microscopy) utilises the photoswitching capabilities of organic fluorophores when in the presence of special mounting media, the solution within which the sample is placed, to detect single molecule fluorescing events over time. The image that can be reconstructed from these events is not diffraction limited, but instead is limited by how well each event can be precisely localised. In Chapter 3 the importance of using a suitable mounting buffer in order to achieve super-resolution dSTORM is discussed in detail. A quantitative method for determining the reactivity of thiol dSTORM switching mountants was developed for use within the lab. Every fluorescent probe has different photophysical properties which can be manipulated by varying the composition of the switching buffer to enhance desirable qualities, such as; increased photon counts, faster switching rates, and longer survivability. In addition to investigating the effects of buffer composition the use of a near UV light-source was also explored as a means of manipulating the same properties to improve overall resolution and quality of the resulting images. A range of photoswitchable fluorescent dyes were tested including Alexa Fluor 660 which is a dye that to my knowledge has not been greatly tested for use in single molecule localisation microscopy by others to date. This dye performed strongly alongside the traditional Alexa Fluor 647 used for dSTORM imaging in optimal conditions. A relatively new approach to single molecule imaging which does not require the fluorophore to photoswitch, called DNA-PAINT (point accumulation for imaging in nanoscale topography), has been investigated throughout this thesis. This approach relies on the transient binding of small oligonucleotide sequences, called “Imagers”, to target docking strands anchored in positions of interest. These imagers have a photostable and bright fluorophore conjugated to the oligonucleotide. It is the transient immobilisation of the imager strand, as it binds to a fixed docking strand, which appears as stochastic blinks. The duration of these events, which can be extended by increasing the number of overlapping base pairs, is primarily responsible for improved localisation precision and therefore potentially overall resolution. At the end of Chapter 3 I compare this new pointillism microscopy approach, DNA-PAINT, with dSTORM using a set of custom-designed oligonucleotide sequences that allow both formats to be employed on the same target. The transient binding of small strands of oligonucleotides offers a far more controllable system for stochastic imaging. In Chapter 4 I use this superior approach to achieve greater resolution than other fluorescence techniques in biological samples, sufficient to visualise single ryanodine receptors (RyR). The RyR are extremely important in the contraction of muscle cells as they are capable of detecting transient changes to calcium concentration and are responsible for releasing large stores of calcium from the sarcoplasmic reticulum. With DNA-PAINT I observed that RyRs cluster into irregular arrays which contain significant gaps that are occupied by other proteins, including junctophilin (JPH). The stoichiometry of JPH with RyR varied cluster to cluster, exposing a new complexity in the regulation of RyRs. In Chapter 5, quantitative super-resolution is reliably achieved through the implementation of quantitative DNA-PAINT (qPAINT) within the Python Microscopy Environment (PYME) software. Quantitative measurements are possible because of the statistical predictability of DNA hybridisation and the near constant influx of fresh imager strands by diffusion. This results in limited photobleaching, a permanent dark state. The frequency with which a region of interest blinks is proportional to the number of binding sites available, and therefore the mean dark time between detected events is also inversely proportional. I validate my approach to qPAINT, which maintains the spatial information of individual structures, by using a DNA-origami test slide. Two distinguishable structures were present and an estimate for the ratio of available docking sites between them was satisfactorily established. I conclude that with this tool, molecule densities can be inferred and information about biological samples can be probed to new levels. The results of the full methodological approach to accomplish dual-colour super-resolution imaging of optically thick cardiac tissue, using both dSTORM and DNA PAINT techniques, is discussed in detail in Chapter 6. The current range of photoswitchable fluorophores limits the possible combination of molecular dyes for use with dSTORM and some compromise is made in their selection. For DNA-PAINT, the prospect of chromatic aberration is removed by imaging the same dye in subsequent rounds of imaging. The process, called Exchange-PAINT, allows the user to remove previously imaged imager strands, through a series of washes, and replace them with a complementary sequence for another target. I introduce the concept of using quencher strands to eliminate signal from unwanted imager sequences, accelerating their removal in samples of reduced diffusion and decreasing the risk of sample disturbance, in a process we termed Quencher Exchange-PAINT. Using this technique, I achieve superior super resolution results in optically thick samples. The results presented in this thesis are expected to (1) lead to a better understanding of the variables associated with single molecule localisation microscopy, (2) further reveal the complexity in cardiac protein distribution, (3) quantify relationships between co-localising proteins and other targets, and (4) apply DNA-PAINT to imaging in optically thick biological samples. This study shows promise for the future applications of the DNA-PAINT pointillism super-resolution method and its ability to investigate a multitude of biological questions.