College of Science, Engineering and Food Science - Doctoral Theses
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Item Reconstruction of milk fat globules/membrane (MFGs/MFGM) and characterization of its physicochemical properties and techno-functionalities(University College Cork, 2024) Sun, Yanjun; Miao, Song; Roos, Yrjo; O'Sullivan, Maurice; Teagasc; China Scholarship Council; State Key Laboratory of Dairy BiotechnologyMost commercially available infant formulas (IFs) lack phospholipids and milk fat globule membrane (MFGM) proteins. To mimic human milk's lipid profile, MFGs/MFGM ingredients from sweet whey or buttermilk are fortified in IF. Understanding the structural and compositional, and techno-functionalities changes of MFGs/MFGM is crucial for their application. The MFGs/MFGM has been reconstructed by the mostly used processing methods in dairy industry, acidification (pH 6.30 and 5.30, 25℃), and novel non-thermal treatment, ultrasonic treatments with different intensities. Additionally, anionic polysaccharide- high-methoxyl pectin (HMP) was incorporated to the raw milk before the cream separation, examining the resulting physicochemical properties and techno-functionalities. Results indicated that pH adjustment affected the MFGs structure and MFGM protein compositions, particularly at pH 5.30, causing protein loss such as butyrophilin (BTN) and Periodic Acid Schiff 6/7 (PAS 6/7) and particle size changes. Fourier Transform Infrared Spectroscopy (FTIR) spectra revealed more proteins compositions in MFGs/MFGM prepared from sweet whey (MFGM-C) compared to bovine raw milk, showing lower interfacial tension and ζ- potential. In addition, there were no apparent polymorphs in MFGM-C, and lower exothermic or endothermic enthalpy was needed during the heating and quenching procedures. Acidification, especially pH 6.30 conditions, led to the adsorption of non-membrane milk proteins on the MFGs or MFGM fragments and formed more types of conjugate detected by FTIR. Consequently, this induced changes in the crystal form proportion and thermal behaviour of MFGs/MFGM at pH 6.30. Different acoustic powers generated by the 20 kHz and 40 kHz independently or synchronously working conditions, affected the interfacial properties and thermal behaviour of MFGs/MFGM samples. 20 kHz treatments led to a relatively even particle distribution in MFGs/MFGM compared to 40 kHz and 20 + 40 kHz treated MFGs/MFGM. Ultrasound treatment significantly resulted in the decrease of interfacial tension (π) in all MFGs/MFGM (p0.05). Three typical crystal forms, α-crystal, β-form and β'-form, were observed in control and ultrasonicated 5 min MFGs/MFGM, but α-crystal disappeared in all the 15 min treated MFGs/MFGM. Ultrasonic duration has a more profound effect on the increase of β'- crystal than the intensity. Ultrasonic treatments promoted the crystallisation of MFGs/MFGM fat at higher temperature but increased the melting temperature of MFGs/MFGM. Additionally, the 15 min treatments caused more pronounced changes in the intramolecular β-sheet, random coil, and α-helix structures compared to the 5 min treatments. 20 + 40 kHz/ 15 min treatment led to aggregated membrane proteins indicated by the confocal laser scanning microscopy (CLSM) images. All the MFGs/MFGM exhibited shear-thinning behaviour, except for the 40 kHz treated MFGs/MFGM, which displayed non-Newtonian behaviour at the lower shear rate. The preliminary investigation on the effects of HMP incorporation on the MFGs/MFGM materials showed the enhanced absorbance at the Amide I and Amide II bands and altered fingerprint regions among all the MFGs/MFGM. Larger particle sizes ranging from 10 to 100 μm presented in the 0.30% HMP MFGs/MFGM, suggesting the formation of conjugates between HMP and milk proteins. These structural and compositional changes induced by HMP affected the physical stability and rheological properties of MFGs/MFGM. Notably, MFGs/MFGM with 0.30% HMP exhibited the poorest stability, a significant increase in apparent viscosity, and shear-thinning behavior throughout the analysis. In summary, acidification, ultrasonic treatments, and HMP incorporation significantly impact the structural, compositional, and functional properties of MFGs/MFGM ingredients.Item New tools and methods for analysis of the sources and spatial distribution of air pollution(University College Cork, 2024) Byrne, Rósín; Hellebust, Stig; Wenger, John; Environmental Protection Agency; European CommissionRecent improvements in air quality sensors (AQSs) have presented an affordable and scalable way to enhance air quality monitoring. However, fully exploiting these new data sources requires novel data-driven methodologies to address the inherent uncertainties in the measurements reported by AQSs. This work goes beyond calibration exercises to highlight the potential of AQS networks for air quality assessments, particularly after demonstrating good sensor-reference correlations and minimal inter-sensor variation. Using three distinct datasets from AQS networks in Cork City and Dungarvan, Ireland, a range of analytical approaches have been developed and applied to understand the local spatiotemporal variability of PM2.5 in these urban areas. A novel method leveraging high temporal resolution data from PurpleAir sensors linked rapid, short-lived concentration fluctuations to local emissions. The analysis revealed stark spatial contrasts in the proportion of local emissions across Cork City. On average, half of the total PM2.5 exposure originated mainly from local emissions and occurred in just 30% of the time. Discrete Fourier transform spectral analysis was also used to estimate local PM2.5 contributions, yielding comparable results when applied to the same dataset. Analysis of the temporal changes to the contribution of local emissions in Cork City between 2021 and 2023 suggested that legislative changes may be positively influencing the proportion of locally emitted PM2.5 in winter, however, this effect is not uniform across the city. A new metric, the concentration similarity index (CSI), was developed and optimised for AQS network data. The CSI was used to assess the spatial representativeness of Environmental Protection Agency (EPA) monitoring locations in both Dungarvan and Cork City, revealing moderate exposure representation. However, significant spatial variability within each sensor network indicated strong differences in the degree of representation by the central monitoring locations and emphasised that location type, rather than geographical proximity, plays a key role in air quality representation. Additionally, a frequency-based method was integrated with the analysis of winter EPA PM2.5 data and aethalometer-derived equivalent black carbon (eBC) data in a small Irish town, to investigate the changing impact of local emissions over time. Analysis of data from 2018 to 2024 showed considerable reductions in winter pollution levels, suggesting an overall improvement in air quality in the area. Analysis of the Ångström absorption exponent indicated an evolving emissions profile, with an increase in wood or peat/turf burning in recent years, while spectral analysis showed general reductions in both regional/background and local particulate pollution. This research offers insight into cost-effective, spatially dense air quality monitoring solutions that can inform vital pollution mitigation strategies. The new methodologies developed across all datasets, demonstrate the potential for integrating AQS networks and data-driven techniques into air quality monitoring and assessment.Item Statistical methods for mapping kinetics together with associated uncertainties in long field of view dynamic PET studies(University College Cork, 2024) Wu, Qi; O'Sullivan, Finbarr; Huang, Jian; University College Cork; Science Foundation Ireland; National Cancer InstitutePositron Emission Tomography (PET) is an essential diagnostic imaging technique in clinical care settings, as well as in medical research. It plays a crucial role in diagnosis, prognosis, treatment planning, and clinical decision-making. PET imaging, a well-established radio-tracer imaging technique, involves injecting a radio-tracer to analyze in-vivo metabolic processes. Dynamic PET scanning provides multiple time frames, offering more detailed metabolic information. However, traditional methods like Patlak and compartmental modeling are commonly used in data obtained from conventional scanners. The use of constant or exponential residue functions may be limited in complex environments, such as diverse tissues or multiple organs. This thesis aims to develop statistical approaches for enhancing and assessing parametric imaging from dynamic PET scans. The Non-Parametric Residue Mapping (NPRM) procedure is established as an entirely automatic process that integrates data-driven segmentation, non-parametric residue analysis, and voxel-level kinetic mapping. A model-based image-domain bootstrapping method is developed with the objective to generate reliable uncertainty estimates, which are crucial for accurate data interpretation and subsequent treatment decisions. This method uses an empirical distribution of re-scaled data and a non-parametric approach for analysis of the spatial correlation structure. Numerical simulations using both direct Filtered Backprojection (FBP) and iterative Maximum Likelihood (ML) reconstructions are considered. Illustrative examples on conventional scanners and Long Axial Field-of-View (LAFOV) PET scanners are conducted. A short-duration dynamic scanning protocol is proposed to enhance the quantitation of a shortened dataset specifically. This protocol utilizes NPRM and machine learning techniques to aim at making short dynamic acquisition protocols clinically feasible.Item Defining gut mediated metabolism for health and disease(University College Cork, 2024) Quilter, Karina; Joyce, Susan; Melgar Villeda, Silvia; Science Foundation IrelandWe investigated the overall concept that human produced and microbially modified bile acids could act as an indicator of health. In chapter 3 we verified disruptions to bile acid metabolism in a porcine model of metabolic syndrome towards cardiovascular disease, induced by diet. In chapter 4 we applied dietary means to address mild hypercholesterolemia and followed BA readouts to indicate health parameters, we determined that BAs could be used as indicators of return to health but only when acute fed conditions were initiated and tracked over a 6 hour time period. Furthermore we noted that baseline fold change in BAs could be applied to distinguish responder and non-responder directions in a clinical setting. In chapter 5 we examined the metabolic outputs relating to elite athletes through 3 intervention studies. Our indications were that a key subset of BAs -significant in westernized MetS and CVD induction- were reduced in elite athletes. We further noted that different sports elicited different correlations with both microbes and metabolites in these small cohorts. In chapter 6, the key BAs associated with MetS-CVD and their interchangeable intermediates were examined at the cellular level and shown to have different outcomes in cell organelle functionality and related gene expression systems depending on dietary lipid interventions. In all, certain BAs are convergent indicators of disease, they diversify in health, just like microbiome composition. A number of key BAs were identified that can indicate health and the push towards disease in this study.Item Gut-heart axis in a large animal model of metabolic syndrome and heart failure(University College Cork, 2024) Cluzel, Gaston; Caplice, Noel M.; Stanton, Catherine; SFI ManufacturingBackground The metabolic syndrome (MetS) is a pathological condition diagnosed as the combination of obesity with either hypertension, dyslipidaemia, or hyperglycaemia. MetS constitutes a deadly cocktail of cardiovascular risk factors that greatly increases patient mortality. Among the cardiovascular complications of MetS, heart failure with preserved ejection fraction (HFpEF) represents one of the greatest unmet clinical needs of the 21st century. Indeed, as HFpEF prevalence increases along with soaring MetS cases, current therapeutic strategies fail to prevent disease complications. Therefore, novel approaches are required. MetS and HFpEF are accompanied by a low-grade inflammation (LGI) state. LGI is characterised as a steady but lingering increase in circulating inflammatory factors. Inflammatory signalling is known for promoting structural and functional changes in the myocardium that may contribute to HFpEF. Thus, decreasing LGI may reduce HFpEF progression. While the precise origin of LGI is uncertain, the gut microbiome has recently emerged as a hidden organ with critical immune regulatory functions. Crucially, the gut microbiome is tightly connected to the intestinal barrier. In MetS and HFpEF, patients show alteration of the gut microbiome and of the intestinal barrier, a phenomenon called gut permeability. Gut permeability results in the translocation of bacterial antigens from the gut lumen to circulation. Circulating bacterial antigens are pro-inflammatory, that contributes to LGI and, indirectly, to cardiac structural changes and HFpEF. Therefore, therapeutic strategies aimed at the gut microbiome may effectively prevent HFpEF via reducing gut permeability and LGI. This concept is described as the “gut-heart” axis. The gut-heart axis constitutes a novel field of investigation in cardiometabolic disorders and may answer the urgent need for novel therapeutic strategies directed against HFpEF. However, more research is needed to characterise the mechanisms involved in gut-heart signalling. Aims of the project This thesis aimed at characterising the cardiac pathological mechanisms involved in gut-heart signalling, and determining whether they can be modulated by a microbiota-targeted treatment. Methodology In this project, gut-heart axis pathological signalling was characterised using a porcine model of MetS and HFpEF induced by Western diet (WD) and hypertensive corticosteroid salts (desoxycorticosterone acetate, DOCA). Then, to investigate the effects of a gut microbiome-targeted intervention on MetS and HFpEF, this model was supplemented with a synbiotic product combining soluble corn fibre and Lactobacillus mucosae. Inflammatory signalling associated with HFpEF structural changes was investigated in the four cardiac chambers. In particular, the project focused on the roles of tumour necrosis factor (TNF)-α, lipopolysaccharide (LPS) and NOD-like receptor family, pyrin domain containing 3 (NLRP3). These central inflammatory pathways may be key in transducing gut-originating LGI into cardiac pathological signalling in HFpEF. Results Upon WD and DOCA challenge, the porcine model constituted a clinically-relevant reproduction of MetS and HFpEF. MetS was characterised by increased body weight, severe hypertension, hypercholesterolemia, and hypertriglyceridemia. HFpEF was characterised by left atrium enlargement (LAE) and left ventricle hypertrophy (LVH). LAE was associated with tissue apoptosis, and LVH was accompanied by cardiomyocyte hypertrophy. Left atrium (LA) and left ventricle (LV) also had increased inflammatory activity with cardiac macrophage (Mφ) expansion, and activation of TNF receptor 1 (TNFR1), toll-like receptor 4 (TLR4), and NLRP3 pathways. Moreover, the increase in TNFR1, TLR4, and NLRP3 activity was colocalised with cardiac Mφ, microvascular endothelial cells, and cardiomyocytes. While no structural or pressure-induced changes were observed in right heart chambers, the right atrium and the right ventricle also exhibited prominent inflammatory signalling. Data not reported in this thesis indicated that the model exhibited LGI and features of gut permeability. Overall, the porcine model of MetS and HFpEF was characterised by inflammatory cardiac changes along with systemic and intestinal alterations. Synbiotic treatment of MetS pigs reduced LAE, LA cardiomyocyte apoptosis, and LVH, but did not affect MetS core parameters. These improvements in cardiac structural changes were associated with a reduction in cardiac Mφ expansion and in TNFR1, TLR4, and NLRP3 activity in all four cardiac chambers. Reductions in TNFR1, TLR4, and NLRP3 activity were colocalised within the cardiac Mφ, microvascular endothelial cells, and cardiomyocytes populations. Data not reported in this thesis also indicated that synbiotic treatment reduced LGI and gut permeability. Therefore, synbiotic treatment targeted at the gut microbiome reduced pathological signalling along the gut-heart axis, and effectively reduced cardiac structural changes associated with HFpEF. Discussion The porcine model of MetS and HFpEF stood out as a robust model for investigating gut-heart axis inflammatory signalling. The study also highlighted the central role of TNFR1, TLR4, and NLRP3 in driving structural changes in HFpEF through pro-apoptotic and pro-hypertrophic signalling. Crucially, synbiotic treatment targeted at the gut microbiota effectively reduced HFpEF-associated structural changes via reducing cardiac inflammatory signalling. Finally, while exempt of structural changes, the right heart reflected accurately and dynamically the systemic changes in gut-heart axis pathology and treatment. Conclusions Synbiotic targeting of the gut microbiome resulted in cardiac structural improvements in a clinically-relevant porcine model of MetS and HFpEF. This study demonstrates the critical role of gut-heart inflammatory signalling cardiometabolic disease progression.