BACKGROUND: There are only limited data about peri-interventional pain during cardiac electrophysiological procedures without analgosedation. In this study, peri-interventional pain and recollection of it after the intervention were evaluated. METHODS: A total of 101 patients (43 electrophysiological/ablation procedures and 58 device surgeries) reported pain on a numerical rating scale (NRS; 0-10) before (pre), during (peri), and after (post) the intervention. Maximum pain (maxNRS) and the average of pain (meanNRS) were used for statistical analysis. Peri-interventional pain was compared with postinterventional data of the recollection of peri-interventional pain (peri-post). Patients were allocated into 2 groups (with 51 and 50 patients, respectively) to evaluate the mode of patient-staff interaction on pain recollection. Depressive, anxiety, and somatic symptom scales (Patient Health Questionnaire-15, Generalized Anxiety Disorder-7, and Patient Health Questionnaire-15) were used to analyze their influence on pain recollection. RESULTS: In total, 49.6% of patients (n = 50) complained of moderate to severe pain (maxNRS) at least once during the procedure. The comparison between peri and peri-post data revealed the following (median (range)-maxNRS, peri: 3 (0-10) versus peri-post: 4 (0-9) (ns), and meanNRS, peri: 1.4 (0-7) versus peri-post: 2.0 (0-6) (ns). The mode of patient-staff interaction had no influence on pain. No effect was found for psychosocial factor concerning pain and the recollection of pain. The results of the linear regression showed no influence of low-dose midazolam on recollection of pain. CONCLUSION: Half of the patients reported moderate to severe pain at least once during cardiac electrophysiological procedures without analgosedation. However, on average, patients reported only low pain levels. Postinterventional derived data on discomfort reflect the peri-interventional situation.:Inhaltsverzeichnis 1 Einführung in die Thematik 1.1 Hintergrund zur Thematik 1.2 Inhaltlicher Gegenstand 1.3 Fragestellung 2 Publikation 3 Zusammenfassung der Arbeit 4 Literatur I Darstellung des eigenen Beitrages II Selbstständigkeitserklärung III Lebenslauf IV Publikationen V Danksagung

<p>  </p> <p>Cardiovascular disease has been the leading cause of death in the United States for over 70 years. To evaluate the extent and progression of cardiovascular disease, non-invasive imaging techniques are frequently used clinically and pre-clinically. Current echocardiographic and cine magnetic resonance approaches rely on measurements that are typically obtained from two-dimensional images, which assumes uniformity of the structure being evaluated. To explore methods to potentially address these shortcomings, our group has developed and validated high frequency four-dimensional ultrasound techniques as well as created a software toolbox that allows for measurement of myocardial kinematics. In this thesis, I assisted in the application of these methods to two murine models of disease states: myocardial infarction and aortic aneurysm. Another study I aided in focused on cardiac magnetic resonance imaging data from patients with Duchenne muscular dystrophy. From our software, we are able to obtain various strain and strain rate estimates that reveal significant functional changes in infarction and Duchenne muscular dystrophy earlier than standard measurement techniques. Furthermore, we are able to identify vascular expansion, transmural thickening, and changes in hemodynamics prior to aneurysm development. Earlier detection and localization allows for more targeted surveillance and interventions, which ultimately may result in improved clinical outcomes. Ideally, these findings can be used to expand the capabilities of cardiac research and the development of clinically applicable imaging techniques and treatments to better address underlying cardiovascular pathophysiology. </p>

No description available.

Intra-aortic balloonpump (IABP) counterpulsation therapy is a volume displacement device designed to provide partial assistance to the left ventricle of the heart. Critical care nurses are expected to manage IABP therapy. It is therefore important that the critical care nurse has the knowledge to manage IABP therapy in a safe and therapeutic manner. The question arises: does the critical care nurse have the knowledge to manage IABP therapy? The purpose of this research study is to explore and describe the knowledge of the critical care nurse regarding the management of IABP therapy. The design of this research study is a quantitative, descriptive and contextual study, in which a sample survey was performed, using a questionnaire (based on a literature study) under controlled conditions. The knowledge of the majority of critical care nurses tested was found to be insufficient. Safe management guidelines and in-service training have been proposed to improve the situation. / lntra-aortiese ballonpomp (IABP) teenpulsasie terapie is 'n volume verplasings apparaat, antwerp om gedeeltelike ondersteuning aan die linker ventrikel van die hart te bied. Kritiekesorgverpleegkundiges is verantwoordelik vir die hantering van rASP terapie. Die vraag ontstaan: beskik die kritiekesorgverpleegkundige oor voldoende kennis rakende die hantering van IABP terapie? Die doel van hierdie studie is om die kennis van kritiekesorgverpleegkundiges te ondersoek en te beskryf rakende die hantering van IABP terapie. Die resultate van hierdie navorsingstudie dui daarop dat die meerderheid kritiekesorgverpleegkundiges wat getoets was oor onvoldoende kennis beskik ten opsigte van IABP terapie. Formulering van riglyne en indiensopleiding is aanbeveel om hierdie situasie te verbeter. Die navorsingsontwerp is kwantitatief, beskrywend en kontekstueel van aard, waartydens 'n gerieflikheidsteekproeftrekking gedoen is, met gebruik van 'n vraelys (gebasseer op 'n literatuurstudie) onder gekontrolleerde toestande. / Health Studies / M.A. (Nursing Science)

Thesis (MSc)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: Diabetes is considered a major threat to human health in both developed and developing nations. Cardiovascular disease which is common in diabetic patients has increased the overall disease affliction. Moreover, stress-induced hyperglycemia has led to increased mortality and morbidity in patients with an acute myocardial infarction (MI), whether the patient has diabetes or not. In addition, acute MI might stem from stress-induced hyperglycemia capability to increase inflammation and oxidative stress resulting in a worse functional cardiac outcome. Hyperglycemia-induced oxidative stress can similarly result in the formation of miss folded or damaged proteins that may be eliminated by the ubiquitin-proteasome system (UPS). Futhermore, hyperglycemia-induced oxidative stress can also result in dysregulation of the UPS that removes these misfolded proteins. Additionally, an increasing body of evidence implicates UPS dysfunction in cardiac diseases and hyperglycemia which has been associated with increased inflammation and blunted cardiac function in response to ischemia-reperfusion. Literature however is blurred whether a reduction or a rise in the UPS is damaging with hyperglycemia and in response to ischemia-reperfusion. In light of this, we hypothesized that UPS inhibitors such as Z-Leu-Leu-Leu-al (MG-132) and lactacystin, protects the rat heart against ischemia-reperfusion under hyperglycemic perfusion conditions. Isolated rat hearts were perfused ex vivo with Krebs-Henseleit buffer containing 33 mM glucose vs. controls (11 mM glucose) for 60 min, followed by 20 min global ischemia and 60 minutes reperfusion ± PI treatment (MG-132 and lactacystin), anti-inflammatory (Ibuprofen) and anti-oxidant (NAC). Infarct size was determined using Evans Blue dye and 1% 2,3,5-triphenyl tetrazolium chloride (TTC) staining with 20 minutes regional ischemia and 2 hours reperfusion ± PI’s treatments. Tissues were collected at the end of the global ischemia experiments and analyzed for UPS activity, oxidative stress, apoptosis and inflammation. Our data expressed a reduced cardiac contractile function in response to ischemia and reperfusion under hyperglycemic conditions as well as an increase in UPS activity. PI treatment resulted in cardio-protection for ex vivo rat heart model exposed to ischemia and reperfusion under hyperglycemic conditions as well as ibuprofen and NAC. In parallel lactacystin treatment significantly decreased myocardial oxidative stress, apoptosis, and inflammation which provided cardio-protection in response to ischemia and reperfusion under hyperglycemic conditions This study shows that acute hyperglycemia elicits myocardial oxidative stress, apoptosis and inflammation that in time results in an increase in contractile dysfunction following ischemia and reperfusion. However, we found that PI treatment with both MG-132 and lactacystin blunted high glucose-induced damaging effects which resulted in a robust cardio-protection in response to ischemia and reperfusion under hyperglycemic conditions, by reducing oxidative stress, decreasing apoptosis and limiting inflammation. A parallel outcome was observed at baseline although the underlying mechanisms driving this process still need to be clarified. Our findings indicate that the UPS may be a unique therapeutic target to treat ischemic heart disease in diabetic patients, and non-diabetic individuals that present with stress-induced hyperglycemia. In summary, this thesis established that PIs act as a novel cardio-protective intervention to treat acute hyperglycemia with associated cardiovascular complications. / AFRIKAANSE OPSOMMING: Diabeties word beskou as ‘n baie groot problem vir menslieke gesondhied vir biede die ontwikkel en onontwikkelende lande. Kardiovaskulêre siekte wat normaal met diabetiese pasiente geassoseerd word veroorsaak ‘n toeneemende druk, wat hierdie siekte laat toeneem. Verder meer vergroot stresgeïnduseerde hiperglukemie die mortaliteit van pasiente met of sonder diabeties wat akute miokardiale infarksie onder lede het. Akute miokardiale infarksie kan ook ontstaan van stresgeïnduseerde hiperglukemie se bekwaamheid om meer inflamasie en oksidante stress te veroorsaak wat in ‘n meer swakker funksionele kardiale toestand. Hiperglukemiegeïnduseerde oksidatiewe stres ook tot wanregulering van die ubikwitien-proteosoomsisteem (UPS) wat wangevoude protïene verwyder, aanleiding gee. Kontrasterende data bestaan van verhoogde/verlaagde UPS aktiwietiet, sowel as met hiperglukemie en/of in reaksie tot isgemie-reperfussie. As gewolg hiervan,, hipotetiseer ons dat Z-Leu-Leu-Leu-al (MG-132) and lactacystin as ‘n nuwe kardiobeskermingsmiddel kan optree deur miokardiale oksidatiewe stress, inflamasie en UPS aktiwiteit te verlaag in reaksie op isgemie-reperfussie tydens akute hiperglukemiese toestande kan verlaag. Geïsoleerde rotharte is ex vivo met Krebs-Henseleit buffer, wat, 33 mM glukose vs. kontrole (11 mM glukose) bevat, vir 60 min geperfuseer, daarna is dit deur 20 min globale isgemie gevolg en 60 min reperfussie ± PI behandeling (MG-132 and lactacystin), antiflammatoriese behandeling (Ibuprofen) en antioxidant behandeling (NAC). Infarkgrootte is bepaal deur Evans bou kleursel en 1% 2. 3-5 tripfeniel tetrazoloimcholierd (TTC) kleuring met 20 minute regionale ischemie, en 2 uur reprefussie ± PI’s behandeling. Weefsels is aan die einde van die globale isgemie eksperimente versamel, en vir oksidatewe stres, apoptose en inflammasie ontleed. Ons data toon aan dat kardiale kontraktiele funksie in reaksie op isgemie-reperfussie onder hiperglukemiese toestande verlaag het asook ‘n toename in UPS aktiwitiet veroorsaak. PI behandeling het gelei tot kardiale beskerming vir ex vivo rotharte wat aan isgemie-reperfussie onder hiperglukemiese toestande blootgestel was sowel as ibuprofen en NAC. Parallel hiermee het lactacystin oksidatiewe stres, apoptose, inflmasie, en UPS aktiwiteit na isgemie-reperfussie, verlaag in reaksie isgemie-reperfussie onder hiperglukemiese toestande. Hierdie studie het bevind dat akute hiperglukemie, miokardiale oksidatiewe stres lei tot oksidante stress, apoptose, en inflamasie na kontraktiele wanfunksionering na isgemie-reperfussie lei. Ons het bevind dat beide MG-132 en lactacystin behandeling, hoë glukose-geïnduseerde skadelike effekte onderdruk, en kardiale-beskerming in reaksie op isgemie-reperfussie onder hiperglukemiese toestande ondervind was deur oksidante stress, apoptose, en inflamasie te verlaag. ‘n Soorgelyke effek is tydens die basislyn waargeneem, alhoewel die onderliggende meganisme wat hierdie proses meer ondersoek instel. Ons bevinding dei dat die UPS ‘n nuwe behandeling teiken kan word in sgemie-geïnduseerde reperfussie onder aktute en chroniese hoë glukose toestande. In opsomming, het die tesis belowend bevindinge gevind wat ‘n nuwe terapeutiese intervensie vir die behandeling van akute hiperglukemie met geassosieërde kardiovaskulêre komplikasies gebruik kan word.

Thesis (PhD)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Left ventricular hypertrophy (LVH) is a strong independent predictor of cardiovascular morbidity and mortality, while its regression is associated with an improved clinical prognosis. It is, therefore, vital to elucidate and fully comprehend the mechanisms that contribute to LVH development and to identify markers that indicate a strong predisposition to the development of severe cardiac hypertrophy, before its occurrence. Hypertrophic cardiomyopathy (HCM) serves as a model to investigate LVH development. This primary cardiac disease is characterised by LVH in the absence of increased external loading conditions and is caused by defective sarcomeric proteins, as a result of mutations within the genes encoding these proteins. However, the hypertrophic phenotype of HCM is largely complex, as we see strong variability in the extent and distribution of LVH in HCM, even in individuals with the same disease-causing mutation from the same family; this points toward the involvement of additional genetic and environmental modifiers. Components of the renin-angiotensin-aldosterone system (RAAS) influence LVH indirectly, through their key role in blood pressure regulation, but also directly, due to the direct cellular hypertrophic effects of some RAAS components. Previous genetic association studies aimed at investigating the contribution of RAAS variants to LVH were largely centred on a subset of polymorphisms within the genes encoding the angiotensin converting enzyme (ACE) and angiotensin II type 1 receptor genes, while the renin section and RAAS components downstream from ACE remained largely neglected. In addition, most previous studies have reported relatively small individual effects for a small subset of RAAS variants on LVH. In the present study we, therefore, employ a family-based genetic association analysis approach to investigate the contribution of the entire RAAS to this complex hypertrophic phenotype by exploring both the individual as well as the compound effects of 84 variants within 22 RAAS genes, in a cohort of 388 individuals from 27 HCM families, in which either of three HCM-founder mutations segregate. During the course of this explorative study, we identified a number of RAAS variants that had significant effects on hypertrophy in HCM, whether alone or within the context of a multi-variant haplotype. Through single variant association analyses, we identified variants within the genes encoding angiotensinogen, renin-binding protein, the mannose-6-phosphate receptor, ACE, ACE2, angiotensin receptors 1 and 2, the mineralocorticoid receptor, as well as the epithelial sodium channel and the Na+/K+-ATPase β-subunits, that contribute to hypertrophy in HCM. Using haplotype-based association analyses, we were able to identify haplotypes within the genes encoding for renin, the mannose-6-phosphate receptor, angiotensin receptor 1, the mineralocorticoid receptor, epithelial sodium channel and Na+/K+-ATPase α- and β subunits, as well as the CYP11B1/B2 locus, that contribute significantly to LVH. In addition, we found that some RAAS variants and haplotypes had statistically significantly different effects in the three HCM founder mutation groups. Finally, we used stepwise selection to identify a set of nine risk-alleles that together predicted a 127.80 g increase in left ventricular mass, as well as a 13.97 mm increase in maximum interventricular septal thickness and a 14.67 mm increase in maximum left ventricular wall thickness in the present cohort. In contrast, we show that a set of previously identified “pro-LVH” polymorphisms rather poorly predicted LVH in the present South African cohort. This is the first RAAS investigation, to our knowledge, to provide clear quantitative effects for a subset of RAAS variants indicative of a risk for LVH development that are representative of the entire pathway. Our findings suggest that the eventual hypertrophic phenotype of HCM is modulated by the compound effect of a number of RAAS modifier loci, where each polymorphism makes a modest contribution towards the eventual phenotype. Research such as that presented here provides a basis on which future studies can build improved risk profiles for LVH development within the context of HCM, and ultimately in all patients with a risk of cardiac hypertrophy. / AFRIKAANSE OPSOMMING: Linker ventrikulêre hipertrofie (LVH) is 'n sterk onafhanklike voorspeller van kardiovaskulêre morbiditeit en mortaliteit, terwyl LVH regressie verband hou met ‘n verbeterde kliniese voorspelling. Dit is dus noodsaaklik om die meganismes wat bydra to LVH ontwikkeling ten volle te verstaan en merkers wat 'n sterk geneigdheid tot die ontwikkeling van ernstige kardiale hipertrofie te identifiseer, voordat dit voorkom. Hipertrofiese kardiomiopatie (HKM) dien as 'n model om LVH ontwikkeling te ondersoek. Hierdie primêre hartsiekte word gekenmerk deur LVH en word meestal veroorsaak deur foutiewe sarkomeer proteïene as gevolg van mutasies binne die gene wat kodeer vir hierdie proteïene. Die hipertrofiese fenotipe van HKM is egter grootliks kompleks; ons sien, by voorbeeld, sterk veranderlikheid in die omvang en die verspreiding van LVH in HKM, selfs in individue met dieselfde siekte-veroorsakende mutasie binne dieselfde gesin, wat dui op die betrokkenheid van addisionele genetiese en omgewing modifiseerders. Komponente van die renien-angiotensien-aldosteroon sisteem (RAAS) beïnvloed LVH indirek, deur middel van hul belangrike rol in bloeddruk regulasie, maar ook direk, as gevolg van die direkte sellulêre hipertrofiese gevolge van sommige RAAS komponente. Vorige genetiese assosiasie studies wat daarop gemik was om die bydrae van RAAS variante LVH te ondersoek, was hoofsaaklik gesentreer op 'n groepie polimorfismes binne die gene wat kodeer vir die “angiotensin converting enzyme” (ACE) en angiotensien II tipe 1-reseptor gene, terwyl die renien gedeelte en RAAS komponente stroomaf van ACE meestal nie ondersoek was nie. Daarbenewens het die meeste vorige studies relatief klein individuele gevolge gerapporteer vir 'n klein groepie RAAS variante op LVH. In die huidige studie het ons dus 'n familie-gebaseerde genetiese assosiasie-analise benadering gebruik om die bydrae van die hele RAAS tot hierdie komplekse hipertrofiese fenotipe te ondersoek deur 'n studie van die individuele-, sowel as die saamgestelde effekte van 84 variante binne 22 RAAS gene, in 'n groep van 388 individue vanaf 27 HKM families, waarin een van drie HCM-stigter mutasies seggregeer. Gedurende die loop van hierdie studie het ons 'n aantal RAAS variante wat ‘n beduidende uitwerking op HKM hipertrofie geïdentifiseer, hetsy alleen of binne die konteks van' n multi-variant haplotipe. Deur middel van enkele variant assosiasie toetsing het ons variante geïdentifiseer binne die gene wat kodeer vir angiotensinogen, renien-bindende proteïen, die mannose-6-fosfaat reseptor, ACE, ACE2, angiotensien reseptore 1 en 2, die mineralokortikoïd reseptor, sowel as die epiteel natrium kanaal en Na+/ K+-ATPase β-subeenhede, wat bydra tot HKM hipertrofie. Deur die gebruik van haplotipe-gebaseerde assosiasie ontleding was ons in staat om haplotipes te identifiseer binne die gene wat kodeer vir renien, die mannose-6-fosfaat reseptor angiotensien reseptor 1, die mineralokortikoïd reseptor, epiteel natrium kanaal en die Na+/ K+-ATPase α-en β subeenhede, sowel as die CYP11B1/B2 lokus, wat aansienlik bydra tot LVH. Verder het ons bevind dat sommige RAAS variante en haplotipes statisties beduidende verskillende effekte gehad het in die drie HKM stigter mutasie groepe. Laastens, het ons stapsgewyse seleksie gebruik om 'n stel van nege risiko-allele wat saam' n toename van 127.80 g in linker ventrikulêre massa, sowel as 'n 13.97 mm toename in maksimum ventrikulêre septale dikte, en' n 14.67 mm verhoging in maksimum linker ventrikulêre wanddikte voorspel, te identifiseer in die huidige kohort. In teenstelling hiermee wys ons dat 'n stel van voorheen geïdentifiseerde "pro-LVH" polimorfismes swakker gevaar het as LVH-voorspellers in die huidige Suid-Afrikaanse kohort. Hierdie is die eerste RAAS ondersoek, tot ons kennis, wat ‘n duidelike kwantitatiewe gevolge vir 'n stel RAAS variante wat ‘n verhoogde risiko tot LVH ontwikkeling aandui, wat verteenwoordigend is van die hele RAAS. Ons bevindinge dui daarop dat die uiteindelike hipertrofiese fenotipe van HKM gemoduleer word deur die saamgestelde effek van 'n aantal RAAS wysiger loki, waar elke polimorfisme ' n beskeie bydrae maak tot die uiteindelike fenotipe. Navorsing soos dié wat hier aangebied word dien as 'n basis waarop toekomstige studies kan bou vir ‘n verbeterde risiko-profiel vir LVH ontwikkeling binne die konteks van die HKM, en uiteindelik in alle pasiënte met' n verhoogde risiko vir kardiale hipertrofie.

Empirical evidence suggests that complex cognitive processes such as decision-making can be influenced by incidental affect (i.e. emotional states unrelated to the decision), which may have importance implications for furthering our understanding and treatment of mood disorders. Following up on previous behavioral findings suggesting that sad mood leads to biases in social decision-making, the present research first investigated how such biases are implemented in the brain. Nineteen adult participants made decisions that involved accepting or rejecting monetary offers from others in an Ultimatum Game (a well known economic task), while undergoing functional magnetic resonance imaging (fMRI). Prior to each set of decisions, participants watched a short video clip aimed at inducing either sadness or a neutral emotional state. Results indicated that sad participants rejected more "unfair" offers than those in the neutral condition, thereby replicating our previous findings. Neuroimaging analyses revealed that receiving unfair offers while in a sad mood elicited activity in brain areas related to aversive emotional states and somatosensory integration (anterior insula) and to cognitive conflict (anterior cingulate cortex). Sad participants also showed a diminished sensitivity in neural regions associated with reward processing (ventral striatum). Importantly, insular activation uniquely mediated the relationship between sadness and decision bias, demonstrating how subtle mood states can be integrated at the neural level to bias decision-making.In a second study, we assessed to what extent such affect infusion in decision-making may translate to clinical depression, a mood disorder involving chronic sad affect. Fifteen depressed and twenty-three nondepressed individuals made decisions to accept or reject monetary offers from other players in the Ultimatum Game. Like transiently sad, but healthy, individuals, depressed participants reported a more negative emotional reaction to unfair offers. However, unlike sad healthy individuals, they accepted significantly more of these offers than did controls. A positive relationship was observed in the depressed group, but not in controls, between acceptance rates of unfair offers and resting cardiac vagal tone, a physiological index of emotion regulation capacity. These findings suggest distinct biasing processes in depression, which may be related to higher reliance on regulating negative emotion.

This exploratory study used Diagnostic Content Validity (DCV) and the Clinical Diagnostic Validation (CDV) models proposed by Fehring (1986) to clinically identify and validate the defining characteristics for Alteration in Tissue Perfusion: Cardiac. The literature based Kelly Cardiac Assessment Tool (KCAT) was designed as the data collection tool. The diagnostic content validity of the KCAT was 0.70. Twenty subjects, 18 years old and older were selected from a population who were admitted as inpatients in a southwestern university affiliated hospital. Data were collected through patient interviews, independent nurse assessment, and review of laboratory data. Using the steps described in Fehring's CDV model (1986) one major defining characteristic and 13 minor defining characteristics were clinically validated. The tool CDV score was 0.62. The nursing diagnosis Alteration in Tissue Perfusion: Cardiac was clinically validated and one major and 13 minor defining characteristics were identified.

<p>Calcineurin is both necessary and sufficient to induce cardiac hypertrophy, an independent risk factor for arrhythmia, dilated cardiomyopathy, heart failure, and sudden cardiac death. However, current knowledge of the downstream effectors of calcineurin is limited. My study utilizes <italic>Drosophila melanogaster</italic> to 1) establish a reliable model for discovering novel modifiers of calcineurin-induced cardiomyopathy; and 2) discover and characterize novel modifiers of calcineurin-induced cardiomyopathy.</p><p>In this study, I generated sensitized <italic>Drosophila</italic> lines expressing constitutively active calcineurin (CanA<super>act</super>) that was either fused to yellow fluorescent protein (YFP) or a Flag epitope (Flag-tagged) specifically in the heart using the cardiac-specific tinC driver (<italic>tinC-CanA<super>act</super></italic>). These sensitized lines displayed significant cardiac enlargement as assayed via optical coherence tomography (OCT), histology, and confocal microscopy. The feasibility of this method was established by testing <italic>Drosophila</italic> expressing deficiency of a known calcineurin modifier, Mef2. </p><p>Employing a targeted deficiency screen informed by calcineurin modifier screens in the eye and mesoderm, Galactokinase (<italic>Galk</italic>) was discovered as a novel modifier of calcineurin-induced cardiomyopathy in the fly through 1) genetic deficiencies, transposable elements, and RNAi disrupting <italic>Galk</italic> expression rescued <italic>tinC-CanA<super>act</super></italic>-induced cardiomyopathy; and 2) transposable element in <italic>Galk</italic> rescued <italic>tinC-CanA<super>act</super></italic>-induced decreased life span. Further characterization identified that the genetic disruption of <italic>Galk</italic> rescued CanA<super>act</super>-induced phenotypes driven in the posterior wing, but not ectodermaly, mesodermaly, or ubiquitously driven phenotypes. In a separate region, genetic disruption of the galactoside-binding lectin, galectin, was also found to rescue <italic>tinC-CanA<super>act</super></italic>-induced cardiac enlargement.</p><p>Together, these results characterize <italic>tinC-CanA<super>act</super></italic>-induced cardiac enlargement in the fly, establish that the <italic>tinC-CanA<super>act</super></italic> sensitized line is a reliable model for discovering novel calcineurin regulators and suggest that galactokinase and galectin-regulated glycosylation is important for calcineurin-induced cardiomyopathy. These results have the potential to provide insight into new treatments for cardiac hypertrophy.</p> / Dissertation

Cardiac and orthopaedic diseases are significant causes of morbidity and mortality in dogs and are therefore critical areas for veterinary research. More information regarding the pathophysiology of these diseases, and the development of novel therapeutics are sorely required and adult stem cells (ASCs) are a promising source of cells for both investigation of these diseases in vitro and also potentially therapeutics in the longer term. ASCs are a readily available source of multipotent cells which bypass the ethical issues surrounding embryonic stem (ES) cells. ASCs have been described in several tissues of the body, and typically differentiate along specific cellular routes related to original source location. This thesis investigates whether ASCs can be isolated and cultured from the dog from two specific locations; cardiac, producing cardiac stem cells (CSCs); and the bone marrow, producing mesenchymal stem cells (MSCs). These cell sources will be extensively characterised at their baseline for morphology, culture behaviour and gene marker expression. Following characterisation each cell source will be subjected to differentiation techniques to examine canine ASC multipotent differentiation potential. CSCs were isolated from cultured atrial cardiac explant tissue taken from dogs post-mortem, with owners’ consent. These cells were able to survive successive passages in serum free media and formed large spherical cell clusters, termed ‘cardiospheres’. CSCs were capable of clonal expansion under controlled culture conditions, demonstrating their ability for self-renewal. Characterisation of these cells demonstrated the expression of CSC markers; c-Kit, GATA 4 and Flk-1 and no expression of cardiac lineage markers including cardiac troponin T and I, Nkx2.5, the cardiac ryanodine receptor and the β1-adrenergic receptor. Primary canine MSCs were isolated from bone marrow aspirates using ficoll separation and cultured on tissue culture plastic. Canine MSCs closely resembled MSCs described from other species, such as the human and mouse, and were found to express CD44 and STRO-1 and were negative for CD34 and CD45. CSCs and MSCs were exposed to published cardiac directed differentiation protocols and differentiation then analysed using cellular morphology and gene expression. Canine CSCs appeared to differentiate partially along cardiac lineages with upregulation of cardiac troponin T and Nkx2.5, and down regulation of c-Kit and endothelial lineage markers. Canine MSCs demonstrated some morphological changes during cardiac differentiation, and demonstrated up-regulation of Nkx2.5 and Flk-1 but no significant alteration in other markers examined. This suggested that cardiac directed differentiation was not as successful with canine MSCs compared to CSCs and conflicting with published data using rodent MSC models. Murine MSCs were used as a positive control cell line for cardiac directed differentiation, based upon published literature. Critically there were key marker expression differences between baseline murine and canine MSCs, including the expression of cardiac markers such as cardiac troponin T and I, and the Ryanodine receptor. Furthermore, expression analysis of cardiac genes changed with time in culture and passage number and no significant alteration was seen when cells were subjected to the cardiac differentiation protocol; thereby bringing into question the data regarding successful cardiac differentiation using murine MSCs. Canine MSCs were further differentiated toward a chondrocyte lineage to investigate the use of MSCs for orthopaedic research. Canine MSCs were successfully differentiated toward articular type cartilage, with demonstration of extracellular matrix secretions, an upregulation of collagen type II with downregulation of collagen type I and the development of SOX9 expression in differentiated cells. This thesis builds the groundwork for future ASC research in the dog. Successful isolation and culture of two ASC sources from the dog is demonstrated. Cardiac and cartilage directed differentiation was successful using primary sourced cells, but differentiation was found to be limited to highly specific routes for each stem cell source. The results presented here highlight the importance of analysing baseline stem cells extensively prior to differentiation and in particular, before making comparisons between cell populations isolated from different locations and species.