title: Environmental Influencers of Health: Investigating Interactions Between Psychological Stress and the Western Diet on Cardiometabolic Risk Factors
Creators: name: Hatton-Jones, Kyle
Publisher: name: Griffith University
Year: 2022
Subjects: subject: Environmental Influencers
subject: Psychological Stress
subject: Western Diet
subject: Cardiometabolic

description: This thesis aimed to investigate the interaction of chronic dietary and psychological stress on health and disease, specifically, the impacts on the biological pathways implicated in altered cardiometabolic and mood states. As will be discussed in Chapter 1: Literature Review, numerous pathophysiological mechanisms and pathways associated with dietinduced obesity (DIO) can be modulated by chronic psychological stressors (CPS) to ultimately alter disease outcomes. However, our understanding of these interactions is incomplete due to the complex relationship between the stress-response and metabolic, immunological, behavioural, and microbial systems across the human lifespan. Furthermore, while being driven primarily by chronic overnutrition and obesity, the development of cardiometabolic disorders is multifactorial, with dysfunction of several tissues and homeostatic mechanisms contributing to disease outcomes. This complexity in the aetiology of these disease states requires a systems-based approach to properly understand the interactions between dietary and psychological stimuli across multiple organ systems and communication networks. This thesis aims to contribute to our understanding of the pathophysiology of metabolic and mood disorders by investigating the interactions that could occur between metabolic, cardiovascular, neuronal, and gastrointestinal systems to influence cardiometabolic risk, behaviour, and myocardial tolerance to injury. In this thesis, data from clinical research trials and data generated using animal models and ex vivo experiments were used to assess the impact of DIO and CPS on health and disease. The main research areas discussed within this thesis are cardiometabolic risk, colonic health, mood disorders, and myocardial ischaemic/reperfusion (I/R) injury. The contents of this thesis is organised into seven chapters and an appendix. Chapter 1 is a literature review that is broken down into four distinct sections. The first section focuses on the impact of dietary and stress-related disturbances on Australian and global communities and aims to highlight the significance of this research. The second section describes biological pathways relevant to obesity and the development of metabolic diseases. In the third section, some significant effects of chronic stress on pathways that impact obesity are described. The fourth section will detail the cardiovascular and cognitive consequences of the interactions described in section three. Chapter 2 outlines the findings of a study investigating whether there is an association between biological correlates of abnormal mood and cardiovascular disease (CVD) and depressive-like symptoms (DLS) in a human cohort. Number of DLS was determined using the Centre for Epidemiological Studies Depression scale (CES-D). Biological associations with DLS were assessed within a cross-sectional human cohort (n=516). Traditional biomarkers (anthropometric, metabolic, cardiovascular) were compared between participants experiencing a high number of depressive-like symptoms and those experiencing few DLS (DLS cohorts; n=77 per group). In addition, novel biomarkers associated with abnormal mood states in obesity (Chapter 1: Section 6) were assessed within a subgroup of the DLS cohorts (n=23 per group). Within the heterogenous population, pulse rate, systolic blood pressure, hip circumference, and insulin were found to be significantly different between low and high DLS participants. However, no significant alterations to the remaining traditional and novel circulating biomarkers were observed within participants experiencing a high number of DLS. Furthermore, only pulse rate, alcohol consumption, and BMI could significantly predict CES-D score. The most likely explanation for this outcome is the heterogeneity of the sample population, the non-specific DLS questionnaire utilised, and the complex relationship between stress, biology, and behaviour. Chapter 3 describes an animal model (n=48) used to investigate the impact of CPS and DIO's alone and when combined. The chapter focused on the impact on metabolism, anxiety-like behaviour, and myocardial tolerance to an I/R insult. Recognising the need for accessible and automated tools to facilitate the collection and analysis of animal behaviour data; an automated tracking tool for behavioural analysis using machine learning algorithms was developed and the work was subsequently published in Computers in Biology and Medicine (Appendix B). For the animal model, half the mice were fed a western diet (WD) for 18 weeks to develop phenotypical characteristics of DIO (weight gain, glucose intolerance). Diets were maintained for a further four weeks while half the mice from each dietary group were subjected to daily chronic restraint stress (CRS). Body weight, glucose tolerance, and behaviour reflective of anxiety were assessed at multiple time points. At the endpoint, hearts were excised and perfused using a Langendorff perfusion system. Myocardial functional recovery was evaluated after 25 minutes of normothermic global ischaemia and 45 minutes of reperfusion. The WD resulted in an increased body weight, worsened blood lipids, and a loss of glucose tolerance in mice. Furthermore, the study noted that WD-fed mice exhibited modest anxiety-like behaviour after 15-weeks of feeding. CRS temporarily reduce body weight and body weight gain in both dietary groups, however a differential effect on glucose tolerance was noted. WD feeding and CRS did not promote robust changes to ex vivo cardiovascular function, however, the study did demonstrate minor differences in how each intervention impacted cardiovascular function. Furthermore, the obesogenic diet combined with chronic stress did not act synergistically to worsen cardiovascular function. This lack of change may have been due to the type of stressor and chronicity that may have resulted in habituation and consequently a reduced stressresponse. This paper was subsequently accepted for publication on 17/12/2021, doi: 10.14814/phy2.15170 Chapter 4 utilised tissue from the animal model described in Chapter 3 to investigate the individual and combined effect of the WD and CRS on microbiome composition and distal colonic gene expression. Microbiome composition of caecal contents was determined using 16s RNA sequencing. RNA was extracted from colonic tissue, and the expression of genes related to inflammation, tight junction proteins (TJP), neuroendocrine, stress hormones, and microbial metabolite receptors was quantified using multiplex technology and real-time qPCR. CRS improved the Firmicutes/Bacteroidetes ratio in WD-fed animals while expanding the pathological Proteobacteria phyla suggesting a complex microbial response in WD mice exposed to CRS. WD-fed mice exhibited lower expression levels of colonic TJP and Tlr4, suggesting alterations to the expression of proteins regulating intestinal permeability and colonic immunity. CRS did not alter these genes, nor any additional regulatory genes, and no synergistic effect with WD was observed; however, CRS exposure altered colonic bile acid receptor expression. The reduced stress-response due to habituation discussed in Chapter 3 potentially impacted the gastrointestinal results that were assessed in this chapter. A paper outlining the findings of this study has been accepted for publication by Neurogastroenterology &amp; Motility (10/11/2021) and is presented here in its accepted format, doi: 10.1111/nmo.14300 Chapter 5 investigated the effects of DIO and CRS on adipose tissue (AT) and hippocampal tissue gene expression using the animal model described in Chapter 3 and 4. The genes quantified were ones that have been implicated in insulin resistance, mood disorders, and cognitive decline. Associations between behavioural indices, biomarkers of metabolism, and expression of regulatory pathway genes across multiple tissues were also investigated. Neither WD feeding nor CRS were found to promote robust changes in either AT or hippocampal tissue gene expression. The findings from this study suggest that the WD and CRS intervention did not induce a robust pathological DIO or stressed phenotype to drive tissue-specific gene expression changes. Hippocampal and AT gene expression results could not explain the anxiety-like behaviour observed in WD mice in Chapter 3. Interestingly, it was noted that a subpopulation of mice exposed to CRS exhibited reduced gene expression of AT adiponectin which is consistent with other studies in stress-susceptible mice. Chapter 6 aimed to investigate the impact of kynurenine (KYN), a tryptophan (TRP) metabolite, on myocardial tolerance to I/R. Dysregulation of KYN metabolism has been noted in obesity, stress-related mood disorders, and cardiovascular disease; however, the molecular mechanisms of action are still poorly understood. In this chapter, mouse hearts were Langendorff perfused, exposed to low and high concentrations of KYN and subjected to an I/R insult. Functional recovery and cell death were analysed to determine the impact of the two KYN concentrations on myocardial tolerance to the I/R insult. Mitochondrial respiration, antioxidant capacity, and changes to gene expression were assessed to identify possible mechanisms of action for KYN in the myocardium. Exposure of hearts to both KYN and an I/R insult worsened myocardial ischaemic tolerance, however, a paradoxical concentration-dependent response was noted. The loss of myocardial ischaemic tolerance could not be explained by changes in myocardial respiration ortotal antioxidant capacity (TAC). An aryl hydrocarbon receptor (AHR)- dependent mechanism for the different outcomes observed in KYN exposed hearts is proposed that warrants further research. Chapter 7 discusses findings and outlines the general conclusions of each study presented in this thesis and the contributions of this work to the literature regarding the combined impact of CPS and DIO on the gastrointestinal tract, metabolism, mood states and behaviour, and cardiovascular disease. Overall, the thesis documented several changes within the WD-fed mice and observed several complex biological responses to stress that are highly context-dependent. The type, duration, and frequency of the stressor, the resilience/susceptibility of the individual animal, and the historical context all play an important role in determining the stress-response and its relationship with WD-feeding and DIO and therefore should be taken into consideration in future research.