The Impact of Weight Fluctuation on Atrial Substrate and the Prevention of Atrial Remodelling With the Use of Anti-Fibrotics

Atrial fibrillation (AF) is the commonest sustained arrhythmia in humans and is responsible for a significant socioeconomic burden. Affected individuals can suffer significant symptoms and are at risk of potentially life-threatening complications. Obesity is increasingly recognised as risk factor for the development of this arrhythmia. Weight fluctuation is common during attempted weight loss and has detrimental cardiovascular effects in human cohort studies, including patients with AF. However, the pathophysiological mechanisms by which this occurs are unclear. The first aim of this thesis is to characterise the electrophysiological effects of weight fluctuation using an obese ovine model. Previous studies have demonstrated that obesity promotes the development of atrial fibrosis as well as the upregulation of profibrotic factors in atrial tissue. The second major aim of this thesis is to investigate the effect of blockade of these profibrotic receptors on obesity-related atrial remodelling. Chapter 2 describes the use a fluctuating weight model in order to study the electrophysiological changes over time. Weight fluctuation was associated with progressive changes in atrial electrophysiology. This group demonstrated reduction in conduction velocity when compared to a lean control group, particularly following a second cycle of weight gain followed by weight loss. These changes were less severe when compared to an obese group. Additionally, the changes in conduction were more heterogeneous than in animals with persistent obesity. This resulted in an increased propensity to AF when compared with lean controls. Chapter 3 investigates the role of endothelin receptor blockade in the prevention of atrial substrate in obesity. Obesity was again induced in ovine subjects and two groups were compared. One was treated with the endothelin receptor antagonist (ERA) bosentan whilst the other acted as a control group. Animals treated with bosentan had attenuation of obesity-related conduction slowing. This was seen on both endocardial and epicardial surfaces. Importantly, there was no effect on either haemodynamics or refractory periods. AF inducibility was also reduced by ERA treatment. Examination of atrial demonstrated reduced fibrosis and downregulation of pro-fibrotic factors with ERA treatment. Importantly, this effect was independent of the TGF-β pathway. Chapter 4 examines the effect of the TGF-β receptor antagonist tranilast on the obese ovine atrium. A similar model of induced obesity was used to compare tranilast treatment with a control group. Animals receiving tranilast demonstrated attenuation of conduction slowing. Endo and epicardial mapping showed this slowing was heterogeneous across atrial sites, perhaps suggesting a predominantly local mechanism in the development of these electrophysiological changes.Thesis (Ph.D.) -- University of Adelaide, Adelaide Medical School, 201

Atherosclerosis

The first national conference with international participation, “Fundamental aspects of atherosclerosis: scientific research for improving the technologies of personalized medicine”, was held in Novosibirsk on 15 October, 2021. The purpose of this conference was to disseminate the latest basic and clinical findings in the fields of etiology, clinical characteristics, and modern diagnostics and treatments of atherosclerosis among various relevant specialists. The conference was intended for practicing cardiologists, primary care physicians, medical geneticists, and physician–scientists. The conference included plenary sessions, specialty sessions, satellite symposia, an open competition for young scientists

Subclinical and clinical atherosclerosis in Non-alcoholic Fatty Liver Disease is associated with the presence of hypertension

Background and aims Non-alcoholic fatty liver disease (NAFLD) is associated with increased cardiovascular risk. However, whether NAFLD contributes independently to the development of cardiovascular disease is not fully understood. Our study aimed at assessing the differences in several indices of atherosclerosis, arterial stiffness and cardiac morphology among patients with isolated NAFLD, isolated hypertension (HT) or with combination of the two conditions. Methods and results One hundred and sixty-nine participants (mean age=50.4±10.2 yrs; males=73.6 %) were divided according to the presence of NAFLD and HT in three groups: only-NAFLD (55 patients), only-HT (49 patients) and NAFLD+HT (65 patients). Exclusion criteria were BMI≥35Kg/m2 and presence of diabetes mellitus. Carotid ultrasonography was performed to measure markers of atherosclerosis and arterial stiffness. Cardiac remodeling was analyzed using echocardiography. Prevalence of subclinical and overt atherosclerosis was significantly higher in the NAFLD+HT patients as compared to the other two groups (atherosclerotic plaques: 43.1%, 10.9%, 22.4% (p<0.001), in NAFLD+HT, NAFLD and HT groups). No differences were found among indices of arterial stiffening and cardiac remodeling across the three groups. In multivariate regression analysis the coexistence of NAFLD and HT was an independent risk factor for overt atherosclerosis (OR=4.88; p=0.03), while no association was found when either NAFLD or HT was considered alone. Conclusion Overt atherosclerosis was significantly present only in NAFLD+HT patients, but not in patients presenting with isolated NAFLD. This implies that the impact of NAFLD on vascular structure and function could depend on the coexistence of other major cardiovascular risk factors, such as HT

Cultivate Quantitative Magnetic Resonance Imaging Methods to Measure Markers of Health and Translate to Large Scale Cohort Studies

Magnetic Resonance Imaging (MRI) is an indispensable tool in healthcare and research, with a growing demand for its services. The appeal of MRI stems from its non-ionizing radiation nature, ability to generate high-resolution images of internal organs and structures without invasive procedures, and capacity to provide quantitative assessments of tissue properties such as ectopic fat, body composition, and organ volume. All without long term side effects. Nine published papers are submitted which show the cultivation of quantitative measures of ectopic fat within the liver and pancreas using MRI, and the process of validating whole-body composition and organ volume measurements. All these techniques have been translated into large-scale studies to improve health measurements in large population cohorts. Translating this work into large-scale studies, including the use of artificial intelligence, is included. Additionally, an evaluation accompanies these published studies, appraising the evolution of these quantitative MRI techniques from the conception to their application in large cohort studies. Finally, this appraisal provides a summary of future work on crowdsourcing of ground truth training data to facilitate its use in wider applications of artificial intelligence.In conclusion, this body of work presents a portfolio of evidence to fulfil the requirements of a PhD by published works at the University of Salford

Investigating the delivery of IGF-1 with in vitro and in vivo model systems of myocardial infarction

Myocardial infarction (MI) is characterised by the irreversible death of cardiac muscle with loss of up to 1 billion cardiomyocytes (CM). Despite survival post-MI dramatically improving in the last two decades, more than 20% of patients suffering MI will still develop heart failure (HF), an incurable condition where the heart is no longer able to meet the body’s needs for blood supply. Amongst novel therapeutic avenues currently being explored, intramyocardial delivery of cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) holds great promise to replace the lost functional tissue. However, the effects of the ischemic microenvironment on these cells still need to be investigated, and protective strategies need to be developed. This thesis examines the delivery of the pro-survival growth factor Insulin like Growth Factor-1 (IGF-1) in the settings of hiPSC-CMs exposed to acidic pH and through a hydrogel-based approach in an in vivo model of MI. Following MI, the heart switches from aerobic metabolism to anaerobic glycolysis, causing a pH drop to 6.5-6.8. The aim of the first part of this thesis was to mitigate the effects of acidic pH on hiPSC-CMs using the pro-survival growth factor IGF-1. It was shown that acidic pH negatively affects hiPSC-CMs in terms of viability, metabolic activity, cardiac gene expression and CMs yield obtained through differentiation. IGF-1 was able to recover the effects of acidic pH, and it could, therefore, be used as a protective strategy for in vivo cell therapy approaches. Another promising strategy for preventing HF progression following MI is the minimally invasive delivery of injectable hydrogels, which can provide mechanical support to damaged tissue and deliver bioactive factors with pro-survival actions. Here, a thermoresponsive injectable hydrogel composed of a triblock copolymer of polyethylene glycol (PEG) and polycaprolactone (PCL) was synthesised and characterised in vitro and in vivo. The hydrogel was prepared with or without insulin-like growth factor-1 (IGF-1) and injected intramyocardially in a mouse MI model. Echocardiography, strain analysis and histological assessments showed that the injection of the biodegradable thermoresponsive hydrogel was effective in ameliorating pathological remodelling, improving overall cardiac function and myocardial mechanics. In the future, implementing novel therapeutic approaches like the ones presented in this thesis could prevent the progression to HF, improving the quality of life of patients affected by myocardial infarction and limiting the socio-economic burden of the disease.Open Acces

Optimization of Feline Adipose-derived Multipotent Stromal Cell Isolation and Canine Cranial Cruciate Ligament Regeneration with Intra-articular Adipose-derived Multipotent Stromal Cells

Adult multipotent stromal cells (MSCs) have become popular within veterinary sciences for direct administration and tissue regeneration due to their differentiation, trophic and immunosuppressive properties. However, current isolation and expansion techniques provide a heterogeneous population with other types of cells, which may affect the efficiency and efficacy of treatments. The progenitor properties and function need to be further evaluated in vitro prior to in vivo application. The first part of this dissertation was to identify the optimal MSC source for general canine orthopedic applications such as intra-articular injection and joint tissue regeneration. Based on side-by-side comparisons of cell doublings, colony forming unit frequencies, target gene expression and immunophenotype, the infrapatellar fat pad (IFP) had MSCs with higher yield, expansion, multipotentiality and progenitor immunophenotypes than joint capsular and cranial cruciate ligament (CrCL) synovium. The second part of this dissertation was designed for optimization of isolation and in vitro expansion techniques for feline adipose-derived MSCs (ASCs). A new method was designed with which a therapeutic dose of ASCs (7 x 106 cells/kg) was available within 3 cell passages from epididymal adipose excised during castration. The isolated ASCs maintained their progenitor properties after cryopreservation. Feline-specific culture medium to induce adipogenesis, osteogenesis and chondrogenesis was also developed in the study for later application in feline tissue regeneration studies. The third part of this dissertation to apply canine IFP ASCs for CrCL regeneration. A custom perfusion bioreactor was designed and built to accommodate canine CrCL templates on which to grow viable grafts with IFP ASCs. Culture medium and template composition and design were optimized to induce the ASCs into fibroblasts to form a collagenous ligament. Compared to ASC templates cultured in stromal (basal) medium those cultured in induction medium had enhanced collagen deposition, procollagen synthesis and ligament specific gene mRNA levels. The results from this investigation confirm the feasibility of growing new canine CrCLs from ASCs in the laboratory for implantation to treat dogs with CrCL disruption. Overall, the investigations in this dissertation provide vital translational information for canine and feline MSC therapies, and may contribute to better efficiency and efficacy of in vivo treatment and tissue regeneration

MR assessment of end-organ damage in the metabolic syndrome and diabetes mellitus

The first part of this thesis focuses on assessing end-organ damage in individuals with the metabolic syndrome and diabetes mellitus using magnetic resonance imaging (MRI) and spectroscopy (1H-MRS). We performed cross sectional and intervention studies to investigate the contribution of obesity, dietary conditions, metabolic environment and exogenous disruptors (chemotherapy) to cardiovascular end-organ damage and the reversibility of this damage with a low caloric diet. The effects of interventions, dietary as well as non-dietary, were explored using imaging technology. As ethnicity is an important aspect of the pathophysiology in diabetes mellitus, we also examined the influence of ethnic factors on diabetes mellitus and its complications. The second part of this thesis focuses on safety, feasibility and implementation of innovative MR techniques at higher field strengths for assessment of cardiovascular disease.Nederlandse Hartstichting, Novo Nordisk B.V., Boehringer Ingelheim B.V., Servier Nederland Farma B.V., Ipsen Farmaceutica B.V., Guerbet Nederland B.V. and Sanofi Nederland B.V.UBL - phd migration 201

The distribution and characteristics of endogenous cardiac stem cells in the adult human heart

The human myocardium harbours resident multi-potent cardiac progenitor cells (CPCs). We investigated the distribution, properties, differentiation potential and effect of LV function on CPCs in all chambers of the human heart. Biopsies from all chambers of the heart from the same patient with good (EF>45; n=5) and impaired LV function (EF<45; n=5) was analysed for c-kitpos and MDR-1pos CPCs. CPCs were isolated using MACS from ten patients (Good and Impaired LV, n=5/group) and was characterised. CPCs were identified in all chambers of the heart in both groups. The RA from good LV group had significantly (p<0.05) less c-kitpos (6± 0CPCs/mm2) and MDR-1pos CPCs (5± 1 CPCs/mm2). In the impaired LV group, the LV (38± 2 CPCs /mm2) had significantly more c-kitpos CPCs. Overall, the impaired LV group had significantly (p<0.05) more c-kitpos (32± 1CPCs /mm2) and MDR-1pos (47± 1 CPCs /mm2). Irrespective of LV function both c-kitpos and MDR-1pos CPCs were significantly higher (p<0.05) in ventricle than atria. CPCs from the LV (80±2%) are significantly (p<0.05) more proliferative than RV (64±4%) and RA (64±6%) in good and impaired LV group, respectively. Regardless of LV function the atria and ventricle showed no difference in proliferation. Cardiosphereogenesis was significantly (p<0.05) higher in the good LV group. Irrespective of the LV function, cardiosphereogenesis, α-sarcomeric actin and calponin expression were significantly increased (p<0.05) in the LV chamber. In impaired LV group, the LV showed significant (p<0.05) expression for Nkx2.5. Overall, the cardiomyogenic and calponin expression were significantly (p<0.05) increased in impaired LV patients. The vWF expression was significantly (p<0.05) increased in LA and the atria of the good LV group. In conclusion, there is a variation in the distribution, stem cell properties and differentiation potential of CPCs across all 4 chambers of the human heart. These variations are also affected by the LV function.Open Acces