Purpose: Morocco is undergoing a nutrition transition, characterised by increasing prevalence non-communicable diseases (NCD), including obesity. In that context, it is crucial to focus on fruit and vegetable (F&V) intake as they may have a preventive effect on weight gain and NCDs. Objectives: The objectives of the present work were: to develop an objective measure of F&V intake and to provide a holistic understanding of factors that may influence F&V consumption, such as socio-demographic and psychosocial factors. Methods: The target population was Moroccan women (20-49 years), living in the urban area of Rabat-Salé. This PhD involved three different studies: the first was based on focus groups that yielded qualitative data of women’s views of F&V; the second study involved validating a quantitative F&V Food Frequency Questionnaire (FFQ); the third a cross sectional population survey-which incorporated findings from studies 1 and 2 to assess dietary intake and the factors influencing F&V consumption. Results: Validation analyses suggested that the quantitative FFQ developed was reliable and valid to measure F&V intake. The mean F&V intake was 213g per day. Women with higher education, higher economic status and better knowledge scores ate significantly larger amounts of F&V than others. Processed food consumption was inversely associated with vegetable intakes. In terms of psychosocial factors, the strongest predictor of intention to eat fruit was control beliefs. Normative beliefs were the strongest predictor of intention to eat vegetables. Intention was the strongest predictor of both fruit and vegetable consumption. Conclusion: The data collected gave an overview of the amount of fruit and vegetables consumed by urban Moroccan women, and enabled a better understanding of the determinants of fruit and vegetable intake. As a consequence, data sheds light on possible avenues for policies and nutrition interventions to focus on in Morocco, in order to increase fruit and vegetable consumption.

The work in this thesis has investigated the potential for a unique HMW glucose polymer (Vitargo, Swecarb AB, Sweden; MW of 500-700 g.mol-1) derived from barley starch to augment short-term post exercise muscle glycogen resynthesis above that of an isoenergetic LMW glucose polymer (Maxijul, SHS International, UK; MW of 900 g.mol-1). The HMW glucose polymer has been previously investigated in comparison to a LMW glucose solution with studies reporting a 70% greater muscle glycogen content after 2 hr recovery from glycogen-depleting exercise (Piehl-Aulin et al., 2000) and an enhanced gastric emptying at rest (Leiper et al., 2000). More recently an improved work output (10%) in a maximal exercise test performed 2 hr after exhaustive exercise was demonstrated after ingestion of the HMW glucose polymer compared to an isoenergetic LMW glucose polymer (Stephens et al., 2008). Key observations in this study were a greater rate of rise in blood glucose and serum insulin concentration during recovery with ingestion of the HMW compared to the LMW glucose polymer. Thus it was suggested that the improvement in performance in the secondary exercise bout could potentially be attributed to greater muscle glycogen availability present at the onset of the test. This hypothesis was subsequently tested initially in this thesis with the quantification of muscle glycogen content after cycling to exhaustion and ingestion of the same HMW and LMW glucose polymers. However, despite undertaking an identical exercise protocol, in contrast with the study by Stephens et al (2008), no differences in the rate of rise in blood glucose or serum insulin were observed. Accordingly muscle glycogen resynthesis measured 2 hrs after exhaustive exercise was similar following ingestion of the HMW and LMW glucose polymers (118 vs. 123 mmol.kg-1). Thus exercise performance in a secondary bout was near identical between both polymers (173 vs. 175 kJ). It was concluded that the LMW and HMW glucose polymers elicited similar post exercise muscle glycogen resynthesis however, since the sampling interval in this study using muscle biopsies was large (2 hr), it may have negated to highlight any early differences in muscle glycogen content. Therefore further investigation was undertaken that focused on more subtle sequential fluctuations in muscle glycogen by using ultra-high field 13C MRS following feeding of the same HMW and LMW glucose polymers. Marginal increases in muscle glycogen during 1 hr of recovery from prolonged exercise were reported after ingestion of the HMW and LMW glucose polymers (6 and 4% respectively). Additionally, increases in muscle glycogen after ingestion of both glucose polymers above that of a zero-energy control were not seen after 1 hr of recovery when a greater magnitude of resynthesis would be expected with the former. It was thus postulated that irrespective of the improved sensitivity of ultra-high field 13C MRS, the technique may not be suited to post exercise muscle glycogen resynthesis determination due to the methodological issue of subject positioning inhibiting typical gastric emptying patterns. When considering the implications of these studies it appears that the HMW glucose polymer does not augment post exercise muscle glycogen resynthesis above that of an isoenergetic glucose polymer with a much lower molecular weight. Nonetheless given that the blood glucose and serum insulin profiles over a 2 hr recovery in the first study of this thesis and the study by Stephens et al (2008) were notably different with the same test solutions, it was considered that there may be a disparity with the HMW glucose polymers utilised. Importantly the production of the HMW glucose has altered such that the manufacturing process has deviated from granulation to agglomeration with the native starch evolving from potato to corn and more recently barley. It was suggested that the most recent HMW glucose polymer used presently had deviated away from its initial characteristics leading to the blood glucose and serum insulin responses observed in the first study of this thesis. Indeed by then comparing post exercise ingestion of a previous granulated version of the HMW glucose polymer with a more soluble agglomerated version in the same experimental protocol as the first study, an initial greater rise in serum insulin was observed in the first 55 min of post exercise recovery. Thus alterations in manufacturing from granulation to agglomeration do appear to have affected properties related to postprandial insulin secretion. However this effect on insulin was not seen overall over the 2 hr recovery period and no differences in blood glucose or exercise performance in a secondary bout were observed suggesting other factors such as the native starch may be influential. It can thus be concluded that the difference in postprandial glucose and insulin responses seen between previous work and the present investigation may be due to altered physical characteristics of the HMW glucose polymer. No differences in intrinsic viscosity, rheology or molecular weight were noted between the agglomerated and granulated versions of the HMW glucose polymer thus the alterations in the origin material may account for more influence on digestibility in vivo. Further investigation would be warranted into effects on post exercise muscle glycogen resynthesis and exercise performance provided that the HMW glucose polymer could be returned to its original formulation.

Peroxisome Proliferators (PPs) are a class of chemicals that cause a programme of augmentative liver growth, however, the mechanism which regulates the induction of hepatic DNA synthesis as a result of exposure to peroxisome proliferators is currently uncharacterized. This study sets out to characterise the induction of DNA synthesis in mouse by peroxisome proliferators, as a prerequisite for investigating and identifying the genes that are responsible for induction of DNA synthesis to control liver growth. Administration of BrdU in drinking water can reduce mouse body weight; an optimized protocol was devised, which does not lead to body weight loss, and which enables reliable measurement of DNA synthesis. Male 129S4/SvJae mice were treated with a single dose of ciprofibrate (100-400 mg kg-1) or methylclofenapate (25 mg kg-1) for two days. Although liver to body weight ratios increased significantly at all doses, no induction in DNA synthesis was observed within 2 days. Subsequent time course studies with ciprofibrate (100 mg kg-1day-1) or methylclofenapate (25 mg kg-1day-1) showed that liver-to-body weight ratio was significantly increased in treated groups by day 2, but that the induction of DNA synthesis was increased significantly only after three days of treatment, for both compounds. No induction of hepatic DNA synthesis was observed in PPARa null mice after treatment with ciprofibrate (100mg kg-1day-1) for 2 or 6 days, showing that the effect required the PPARa. A dose-response study with 0,1,3,10,30,100 or 200 mg kg-1 day-1 ciprofibrate for 3 days, or with 0,10,30,100 mg kg-1 day-1 ciprofibrate for 4 days revealed that liver to body weight ratios were significantly increased in 129S4/SvJae mice treated with 10mg kg-1day-1 and greater ciprofibrate at 3 and 4 days, whereas hepatic labelling index was significantly increased at 100 mg kg-1 day-1 ciprofibrate at 3 days after dosing, with progressive increases at doses of 30 and 100 mg kg-1 day-1 ciprofibrate at 4 days after dosing. In order to explain the early time course of induction of DNA synthesis reported by Styles [113] [164] in Alderley Park mice, a time course study was performed between 1-4 days in Alderley park mice using methylclofenapate (25mg kg-1day-1). The study showed that liver growth was induced by day 2, but DNA synthesis was significantly induced only after 3 days of dosing. To evaluate species differences, the time-course of induction of DNA synthesis was examined in F-344 rats treated with ciprofibrate (50mg kg-1day-1) for 1-4 days. The liver-to-body weight ratio was significantly increased in all time points, but DNA synthesis was significantly increased after 2 days of dosing. These findings demonstrate that there was a delay in induction of DNA synthesis by peroxisome proliferators in mouse by at least 48 hours. This delay in response is not due to strain differences. Moreover, induction of DNA synthesis in rat was earlier than those in mouse, which makes rats a feasible experimental model to study the immediate early genes/ proteins induced by peroxisome proliferators to induce liver growth.

The project set out to use a very precise three dimensional tracking system to identify changes in joint condition for use in clinical assessment. Untried and untested the CODA-3 was brought into the department and put through a six month period of validification in order to evaluate it's capabilities. These are described in detail in the text. Once satisfied that the equipment was capable of measuring minute rapidly changing position of it's prismatic markers, pilot studies were devised to assess it's ability to reproduce the results from well recognised gait-analysis techniques. It gave promising results. The next task was to determine which set of parameters we could derive using CODA-3 that would be of use in describing the kinematics of the diseased and/or prosthetic knee for use as a tool in clinical assessment. Using FORTRAN, subroutines were written and run on a DEC LSI-11 computer, to collect, store and analyse the x, y and z coordinates of the eight CODA landmarks. It was hoped that by appropriate siting of the markers the velocities and accelerations of the segments comprising a joint could be monitored throughout the gait cycle. The resultant patterns of these parameters were plotted out, and the actual data values stored. It was hypothesised that weaknesses in a joint, whether or not detectable by clinical examination would, at points in the gait cycle of maximum joint loading be seen as ectopics in the smooth waveform of the acceleration and velocity of the profiles expected from the normal knee. The results the author presents would suggest that if the limitations of this particular model could be overcome (as it is reported they will be) then the technique has the capability of highlighting abnormalities in a joint. The author is doubtful however that these same weaknesses could not be detected by the clinician. The system may well have other applications related to this area of work and these are discussed.

Interval timing behaviour refers to the ability of animals and humans to adapt their behaviour to temporal regularities in their environments. Two important classes of interval timing behaviour are temporal discrimination (discriminating between the durations of external events) and temporal differentiation (behavioural adaptation during an ongoing interval). It has been known for many years that drugs that affect central dopaminergic function can alter both forms of timing behaviour. More recently, evidence has been accumulated which shows that manipulation of central 5-hydroxytryptaminergic (5-HTergic) function can also influence interval timing behaviour. The experiments described in this thesis examined the effects of drugs acting at some subtypes of 5-HT receptors on temporal discrimination and temporal differentiation in the rat. Chapter 1 contains a review of the relevant literature. First, the anatomy, biochemistry and receptor pharmacology of the 5-HTergic system is outlined, and a selective review of the role of 5-HT in some behaviours relevant to this project is presented. This is followed by an overview of the behavioural methodology that has been used to study timing behaviour in animals, and an account of the major theories of timing behaviour. Finally, the behavioural pharmacology of timing behaviour is reviewed. Chapters 2-7 describe a series of experiments examining the effects of drugs acting at 5-HT1A, 5-HT2A/2C, and 5-HT3 receptors on temporal discrimination and temporal differentiation. Experiment 1 examined the effect of the 5-HT3 receptor agonist m-chlorophenylbiguanide (m-CPBG) and the non-selective agonist quipazine on temporal discrimination performance in the discrete-trials psychophysical procedure. Quipazine produced a dose-dependent disruption of temporal discrimination, consisting of a rightward displacement and flattening of the fitted psychometric function, reflected in a significant increase in the values of the indifference point T50 and the Weber fraction. m-CPBG had no significant effect on either T50 or the Weber fraction. The effects of quipazine were completely abolished by the 5-HT2A receptor antagonist ketanserin, but not by the 5-HT3 receptor antagonist topanyl 3,5-dichlorobenzoate (MDL-72222), indicating that the effect of quipazine was mediated by 5-HT 2A, and not 5-HT 3 receptors. In experiment 2, the effects of quipazine and m-CPBG were examined on temporal differentiation performance in the free-operant psychophysical procedure. Quipazine dose-dependently displaced the psychometric function to the left, reducing the value of T50, and significantly increased the Weber fraction. m-CPBG had no effect on the parameters of the function. The effects of quipazine were reversed by co-administration of ketanserin, but not by co-administration of MD L-72222. These results suggest that while 5-HT 2A receptor stimulation has a robust influence on temporal differentiation, 5-HT3 receptor stimulation does not. Experiment 3 further examined the effect of 5-HT2A receptor stimulation on temporal discrimination. The 5-HT 2A/2C receptor agonist 2,5- dimethoxy-4-iodoamphetamine (DOI) increased the Weber fraction and tended to increase T50. Ketanserin and the highly selective 5-HT2A receptor antagonist (± )2,3-dimethoxyphenyl-1-(2-( 4-piperidine )-methanol) (MDL-I00907) fully antagonized the effects of DOI The results indicate that DOI disrupts temporal discrimination via stimulation of 5-HT2A receptors. Experiment 4 examined whether intra-striatal injection of DOI would affect temporal discrimination, and whether the effect of systemically administered DOI on temporal discrimination would be blocked either by MDL-100907 or by 8-( 5 -(2, 4-dimethoxy-5 -( trifluoromethylphenylsulphonamido )phenyl-5-oxopentyl)-1 ,3,8- riazaspiro( 4.5)decane-2,4-dione RS- 102221: a selective 5-HT2C receptor antagonist), administered directly into the dorsal striatum. Intra-striatal injection of DOI did not affect temporal discrimination. Systemically administered DOI disrupted temporal discrimination; this effect was not attenuated by intra-striatal injection of MDL-100907 or RS102221, suggesting that the 5-HT2 receptors that mediate DOI's effect on temporal discrimination are not located in the dorsal striatum. Experiments 5 and 6 examined the effects of intra-striatally administered DOI, MDL-100907 and RS-102221 on temporal differentiation. In experiment 5, systemic injection of DOI significantly reduced T50. This effect was antagonized by systemically administered MDL-100907. In experiment 6, intra-striatally administered DOI had no significant effect on T50 or the Weber fraction. Intra-striatal injections of MDL-100907 and RS-102221 did not alter temporal differentiation, and failed to reverse the effects of systemically administered DOI. The results suggest that the 5-HT2 receptor population responsible for DOI’s effect on temporal differentiation is not located in the dorsal striatum. Experiment 7 examined the effect of a 5-HT1A and a 5-HT2A receptor agonist on another widely-used temporal differentiation schedule, the fixed interval peak procedure. The 5-HT1A receptor agonist 8-hydroxy-2-( di-n-propylamino) tetralin (8-0H-DPAT) and the 5-HT2A/2C receptor agonist DOI had similar effects on performance: both agonists displaced the peak function to the left and reduced the peak time, tpeak. The effect of 8-0H-DPAT was antagonized by the selective 5-HT1A receptor antagonist N-[2-( 4-[2-methoxyphenyl]- 1-piperazinyl)ethyl]-N-2-pyridinylcyclohexane-carboxamide (WAY- 100635), and the effect of DOI by the 5-HT2A receptor antagonist ketanserin, respectively. These results, taken together with previous findings with the free-operant psychophysical procedure, suggest that 5-HT1A and 5-HT2A receptors mediate similar effects on temporal differentiation. The final chapter (Chapter 8) summarizes the findings from the project, and discusses their implications for the putative role of 5-HT in interval timing and for current theoretical accounts of timing. It is argued that current models of timing behaviour that assume the existence of a unitary 'pacemaker-driven' internal clock may have difficulty accommodating the finding that the same drug can have qualitatively different effects on temporal discrimination and temporal differentiation. Some possible directions for future research in this area are also discussed.

The extracellular matrix (ECM) has emerged as a major regulator of cell behaviours. Changes in extracellular matrix, especially its composition, organization/ dimensionality, and rigidity have been implicated in various aspects of cellular functions including cell growth, migration, and differentiation. In my thesis, I have focused on the effect of two biophysical properties of the extracellular matrix namely dimensionality and rigidity in the inflammatory and fibrotic pathologies of human lung. To study the role of matrix dimensionality, firstly electrospun scaffold based three-dimensional (3D) culture with similar architecture of human lung was developed. By applying this 3D model, inflammatory response was studied in an in vivo like environment by using NF-κB transcription factor activation as a tool for probing inflammatory response in human lung fibroblasts. According to my observations, it was confirmed that the matrix dimensionality together with spatial organisation of cells is crucial in lung inflammatory response, evidenced by the observation of the differences in the level and pattern of inflammatory response between 2D and 3D culture systems. To study the role of matrix rigidity in progression of lung fibrosis, we developed the ECM-based hydrogel platform with tuneable stiffness level relevant to normal and fibrotic lung. By using this disease relevant platform, I have shown that stiff matrix but not soft matrix can induce the myofibroblast differentiation and fibroblast proliferation, the two major features of lung fibrosis. To date, the molecular mechanisms underpinning this cellular mechanosensing process in response to matrix stiffening remains unknown. To achieve this, I further investigated the involvement of two potential mechanosensitive signalling pathways namely, Rho associated coiled coil forming kinase (ROCK) signalling and talin- (focal adhesion adaptor) signalling in this process. Interestingly, my data show that ROCK signalling differentially regulated stiffness induced myofibroblast differentiation between soft normal and stiff fibrotic matrix. Moreover, both ROCK isoforms 1 and 2 are synergistically important in myofibroblast differentiation driven by rigid matrix and the absence of one ROCK isoform can exaggerate myofibroblast differentiation on stiff fibrotic matrix. Regarding talin signalling, my preliminary data confirms that talin1 can control both stiffness induced fibroblast proliferation and myofibroblasts differentiation on stiff matrix. In contrast to talin1, talin2 showed a protective role in controlling myofibroblast differentiation. In conclusion, we have successfully developed two in vitro lung models for studying the effect of matrix dimensionality and rigidity in lung inflammation and fibrosis. Overall my PhD work has elucidated the significant contribution of biophysical cues of external cellular environment in lung inflammation and fibrosis.

Smooth muscle (SM) tissue is found in many parts of the body, primarily in sheets or bundles surrounding hollow organs. The main function of the tissue is the regulation of organ tone via its contractile state. Dysfunction of SM in diseases such as asthma and atherosclerosis affect millions worldwide. Current methods for studying SM primarily rely on ex vivo animal tissues or 2D in vitro models. Animal models do not accurately recreate human disease states and 2D models are cultured on stiff surfaces lacking the elastic properties and 3D morphology found in natural extracellular matrix in vivo. Therefore it is desirable to develop both an in vitro model of SM that possesses the ability to contract and a method in which this contraction can be measured. In order to achieve this, primary rat aortic SM cells and primary human airway SM cells were cultured in collagen hydrogels; both free floating in the form of collagen disks and under uniaxial tension in order to generate aligned SM collagen constructs. When stimulated with contractile agonists, these constructs contract in a uniaxial fashion. The design of the constructs allows them to be attached to a force transducer allowing the physical force of contraction to be measured. The force of contraction was dependant on the agonist concentration and could be antagonised by the presence of an L-type calcium channel blocker. In order to improve the alignment and uniformity of the smooth muscle population a range of aligned electrospun scaffolds were produced from polyethylene terephthalate (PET), cross-linked gelatin and cross-linked gelatin methacrylate (GelMa). The average fibre diameter of the scaffolds ranged from approximately 200 nm to several micrometres. Additionally the Young’s moduli of the scaffolds ranged from around 1x105 to 1x108 Pa. In all cases, scaffolds were highly aligned; alignment was achieved by using a rapidly rotating collector mandrel. Culture of primary SM cells upon these scaffolds showed that the cells readily adhered to and proliferated upon the scaffolds over a 10 day culture period. The cells formed a highly aligned population following the topographical cues of the aligned fibrous scaffolds. Additionally, the cells stained positive for SM markers in all cases, indicative of a contractile phenotype. When stimulated with 100 µM UTP, the SM cells were able to contract the gelatin and GelMa scaffolds but not the PET scaffolds. SM seeded GelMa scaffolds were cultured for 10 days prior to attachment to the previously mentioned force transducer apparatus. Upon stimulation the seeded scaffolds contracted generating forces greater than those achieved by the hydrogel model, and was reproducible over several experiments.

Translocation of wild Asian elephants (Elephas maximus) is used extensively to mitigate human-elephant conflict (HEC) in Peninsular Malaysia since 1974. Very little is known about the fate of translocated elephants after relocation due to challenges in observing elephants in the dense rainforest. Advances in wildlife endocrinology suggest that faecal glucocorticoid metabolites (fGCM) can be used to study adrenal activity remotely, to assess the Hypothalamic-Pituitary-Adrenal (HPA) axis response towards stressors. The aim is to assess the impact of translocation on wild Asian elephants in Peninsular Malaysia using faecal endocrinology and GPS technology. The specific objectives are: (i) adapting hormone sampling methods for use under tropical field conditions, (ii) comparing fGCM concentrations between translocated and local resident elephants using enzyme immunoassay, and (iii) quantifying gastrointestinal parasite eggs and microflora ciliates in faecal samples to detect signs of immunosuppression. We found that Asian elephant’s fGCM (80 dungpiles, 685 subsamples) are stable up to eight hours in the field. From the monitoring of wild elephants at the release sites, between two months up to a year, translocated elephants (N=5) had lower fGCM concentrations in comparison to local resident elephants (N=4; Linear Mixed Models: t=-2.77, df=7.09, P=0.027). There were no differences in gastrointestinal parasite egg counts (P > 0.05) or microflora ciliate counts (P > 0.05) between translocated and local resident elephants. In conclusion, translocation does affect elephant physiology but this is in the opposite direction from that expected – a prolonged decrease rather than increase of adrenal activity. It is unknown if these conditions could cause immunosuppression, but it could adversely affect stress response and health of the elephant (e.g. adrenal insufficiency, chronic fatigue or Post-Traumatic Stress Disorder). When assessing HEC mitigation, conservation authorities and other stakeholders need to consider that translocation may not be the best solution for HEC, as it will have long-term consequences on elephants’ health.

The skin is the most extensive and accessible organ in the human body. It efficiently provides a barrier to an external hostile environment whilst maintaining and regulating fundamental physiological functions. The sophisticated and complex nature of this natural barrier requires continued analytical advancement to offer further insight into both its biological mechanisms and how to target the delivery of compounds through it. This work presents the use of a recently emerging technique in the field of skin research, time of flight - secondary ion mass spectrometry (ToF-SIMS), to investigate the presence of both native and exogenous compounds in skin tissue from samples collected both in vivo and ex vivo. Subtle changes to the stratum corneum lipid composition have been shown to exert significant effects on the barrier properties of the skin and are associated with numerous skin disorders. The analysis of these lipid species and factors affecting their composition, both internal and external, is therefore a vital area of research. Using ToF-SIMS, this work has conducted an examination of changes to this lipid composition that have resulted from aging of the skin. This has been achieved by undertaking extensive development of a recently proposed surface analysis method to analyse sequential tape strips of stratum corneum. This study was unprecedented in demonstrating that ToF-SIMS could obtain information on human skin from samples collected in vivo. Changes in the levels of both cholesterol sulfate and long chain fatty acids were observed as a consequence of both intrinsic and extrinsic aging, offering confirmatory evidence to previously theorised skin aging mechanisms. Research relating to the effective permeation of compounds across this skin barrier is also of upmost importance, both to the pharmaceutical and cosmetic industries, to enable the design of new topical formulations for skin delivery. Currently employed methods to assess the permeation of a compound are heavily focused on dermal delivery, with limited information obtained on the effectiveness of a compound to permeate into the upper layers of the skin. This research has therefore pioneered a dynamic SIMS method to conduct depth profile analysis of ex vivo porcine skin tissue, enabling the permeation of exogenous compounds to be monitored as a function of skin depth. This work is novel in successfully producing 3D spatially resolved chemical profiles of exogenous compounds within biological tissue using ToF-SIMS. The permeation of four different vitamin C related compounds, popular ingredients in anti-aging cosmetic formulations, were assessed using this method, highlighting a significant difference in permeation efficiency between them. An investigation into the delivery of ascorbic acid to the skin from various different formulations was also achieved, highlighting a permeation enhancing effect from delivery via a novel supramolecular gel formulation. The method developed for surface analysis was also successfully applied to monitor the permeation of ascorbic acid through human stratum corneum following in vivo application of an ‘off-the-shelf’ cosmetic product.

Research during the past decade has highlighted the functional role of perivascular adipose tissue (PVAT) in regulating the contractility of the underlying vascular smooth muscle cell layer. However, the mechanisms underlying these observations are poorly understood. As a sexual dimorphism has been identified in the regulation of vascular tone, the aim of this thesis was to determine whether there are sex differences in PVAT-mediated regulation of the porcine coronary arterial (PCA) tone. In the presence of adherent PVAT, which was stored overnight at 4Co, contractions to the thromboxane mimetic U46619 and endothelin-1 were significantly reduced in PCAs from females, but not PCAs from males. In PCAs pre-contracted with U46619, re-addition of PVAT caused relaxation in PCAs from females, but not PCAs from males. This relaxant response in coronary arteries derived from females was inhibited by a combination of both NO synthase inhibitor (L-NAME) and the cyclooxygenase inhibitor indomethacin. Pre-incubation with an anti-adiponectin antibody abolished the relaxant effects of PVAT, indicating that adiponectin is likely to be the mediator released from the fat. There was no difference in either expression or release of adiponectin from PVAT between sexes. On the other hand, the adiponectin receptor agonist adipoRon produced greater relaxation in PCAs from females compared to PCAs derived from males. No adiponectin receptor 1 expression was detected in PCAs whilst adiponectin receptor 2 and adiponectin binding protein (APPL1) were expressed equally in PCAs from both sexes. Pre-incubation with the hydrogen sulphide enzyme inhibitors 4-propargylglycine (PPG) and 2-(aminooxy)-acetic acid (AOAA) did not reduce the anticontractile responses to PVAT. Cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3MPST) are expressed in PVAT from both sexes equally, with a relatively low activity, whilst no cystathionine γ-lyase (CSE) expression has been identified in PVAT from both sexes. These data indicate a clear sex difference in the regulation of coronary artery tone in response to adiponectin receptor stimulation, which may underlie the anticontractile effects of PVAT in females. Although H2S synthesizing enzymes are expressed in PVAT, they have no functional role in PVAT-induced vasorelaxation. In arteries from both male and female pigs, addition of fresh PVAT caused a contraction, which was partially inhibited by the cyclooxygenase inhibitors indomethacin and flurbiprofen. The PGF2α receptor (FP) antagonist AL8810 attenuated the PVAT-induced contractions in arteries from males, whereas the TXA2 receptor (TP) antagonist GR32191B inhibited the PVAT-induced contractions in arteries from females. Although there was no difference in PGF2α levels in PVAT between females and males, PGF2α produced a larger contraction in arteries from males, correlating with a higher FP receptor expression detected by immunoblotting. In contrast, the release of TXB2 from PVAT from females was greater than the release from males PVAT, but there was no difference in the contraction by the TXA2 agonist U46619. Furthermore, there was no difference in the role of extracellular Ca2+ and Rho kinase pathway (intracellular Ca2+) in the action of U46619, or TP receptor expression in arteries from different sexes. Pre-incubation with the reactive oxygen species (ROS) scavenger N-acetyl-L-cysteine (NAC), a non-selective NADPH oxidase (Nox) inhibitor (DPI), a selective Nox1 inhibitor (ML171), a selective Nox2 inhibitor Phox-I2 and selective Nox1 and Nox4 inhibitor (GKT137831) significantly reduced PVAT-induced vasoconstriction in PCAs from both females and males. Nox1, Nox2 and Nox4 were expressed equally in PVAT from male and females. However, in PCAs, Nox1 expression was greater in females whilst Nox4 was higher in males. Nox2 was expressed equally in PCAs from different sexes. ML171 and phox-I2 reduced superoxide anion (O2-) production in PVAT and PCAs from both sexes. GKT137831 inhibited H2O2 production in PCAs, but not PVAT, from both sexes. Both U46619 and PGF2α had similar effects on ROS production in that they did not increase O2- production in PVAT while they enhanced O2- production in PCA from females only. Functional studies showed that Nox inhibitors could inhibit the PCAs response to both U46619 and PGF2α in females only. The adipose-derived compound chemerin-9 caused a significantly higher vasoconstriction in PCAs from females in comparison with males PCAs. Similarly, chemerin-9 enhanced Nox activity in females PCAs but not in males PCAs. Chemerin mRNA was expressed in the PVAT, and the chemerin receptor ChemR23 was expressed in PCAs, although there were no sex differences. The anti-chemerin antibody can abolish the PVAT-induced contractions in both sexes. The sexual dimorphism in the contraction to chemerin could be explained by differences in the signalling downstream of the chemerin receptor in PCAs rather than the expression of chemerin or its receptor in PVAT and PCA, respectively. In conclusion, this thesis has demonstrated clear sex differences in the regulation of coronary artery tone by PVAT, with a dual effect of PVAT on the contractility of the PCA. PVAT stored overnight enhanced vasorelaxation in PCAs from females only due to the increased adiponectin activity compared to the males PCAs. In contrast, fresh PVAT augments vasoconstriction in PCAs from both sexes. Prostanoids have an important role in PVAT-induced vasoconstriction in which PGF2α and TXA2 are responsible for vasoconstriction in male and female PCAs, respectively. In addition, chemerin may have a role as a PVAT-derived contractile agent in female PCA only, while Nox-derived ROS regulates PVAT-induced contraction in the PCAs from both sexes. This study highlighted the importance of considering potential sex differences in the responses to PVAT and that sex-specific drug treatment may represent a potential strategy in the treatment of cardiovascular diseases.