1,721,110 research outputs found
Modulation of regulatory T cell function through specific molecular pathways in transplantation
Full text linkRegulatory T cells (Tregs) are crucial mediators of immune homeostasis that have shown immense promise as a cellular therapy for immune-mediated pathologies such as transplant rejection. Methods to optimise current Treg-based therapies are critical for its successful and widespread translation to the clinical setting. The aim of this study was to investigate the role of two molecular pathways for the modulation of Treg biology and function.
In the first part of this study, the role of the interleukin-33 (IL-33)/ST2 axis was interrogated in Tregs. This highly pleiotropic pathway has recently been demonstrated to have an important role in Treg biology. Here, it is shown that IL-33-expanded Tregs have enhanced in vivo suppressive function in a transplantation model. An in-depth characterization of IL-33-expanded Tregs is presented, providing a detailed quantitative transcriptomic and immunophenotypic analysis. Interestingly, this reveals IL-33-expanded Tregs to harbour a phenotypic signature of enhanced potential for graft-homing.
In the second part of the study, Treg plasticity and the environmental factors which govern their phenotype and function were examined, with a particular focus on the role of hypoxia and the principal regulators of oxygen homeostasis, the HIF/PHD pathway, in modulating Treg biology. Using an array of HIF/PHD2 transgenic mouse models, the study demonstrates that silencing of the critical prolyl hydroxylase domain-containing enzyme, Phd2, induces an autoimmune-like phenotype in mice. This is driven by upregulated HIF-2a signalling, in which the suppressive function of Treg populations is significantly impaired with loss of ability to control graft rejection. Additionally, evidence is presented to demonstrate that HIF-2a may be inhibited to enhance suppressive function of both mouse and human Tregs. These findings are replicated, at least in part, in mice subjected to chronic hypoxic.
While significant progress has been made to unlock the therapeutic potential of Tregs, there remains a great deal more to be learned regarding their behaviour and function for their effective and safe implementation within the clinic. The findings presented in this thesis uncover methods for the molecular manipulation of Tregs to help exploit their versatility for therapeutic use
Social Regulation of the Escape and Swim Motor Circuits in Dopamine Receptor Type 1 Mutant Zebrafish (Danio rerio)
No full textDominance hierarchies are an evolutionary mechanism to ensure the stability of animal groups by enabling the division of resources like food and mates according to social rank. Once dominance is established, the behavior pattern of individual group members shifts according to their social rank. We are motivated to better understand the neural bases of social behavior and how social activity influence nervous system function particularly the role of neuromodulators in regulating spinal motor circuits. Using zebrafish as a model organism, we tested the importance of the dopamine type 1 receptor (DRD1) in regulating social activity and spinal motor circuits in socially dominant and submissive fish by genetically knocking the DRD1 receptor.
Our motivation stems out of the fact that in many animal species dopamine plays an important role in regulating aggression, motivation and spinal motor activity. However, it remains poorly understood how social factors can influence dopaminergic signaling and its impact on motor function. We focused out attention on two easily quantifiable behaviors: the startle escape response and swimming behaviors.
In wildtype zebrafish, we found that subordinate animals display an elevated startle response sensitivity and reduced swimming activity compared to dominant animals. However, preliminary results show that although the DRD1 KO animals display similar status-dependent behavior patterns compared to wildtype pairs, these differences are less distinct. DRD1 KO subordinates continue to display heightened escape response sensitivity and reduced swimming activity compared to dominants, but those differences are less discernable. We aim to verify these results with a larger sample size. If confirmed, our results would suggest that the DRD1 receptor potentially plays an important role in regulating motor activity in a socially status-dependent manner
Characterization of Social Status-Dependent Neuromodulation in Zebrafish (Danio rerio)
No full textIn zebrafish (Danio rerio), social interactions between adult males consist of a series of aggressive encounters that ultimately lead to the formation of stable hierarchies of either socially dominant or subordinate animals. Although it has been shown that social status leads to neurophysiological changes in brain structure and function, our understanding of the underlying mechanisms that control behavioral function remains limited. We show that socially dominant animals display increased swimming activity. Conversely, social Subordinates display decreased swimming activity, but an enhanced sensitivity of the C-start escape circuit. We also show that whole brain expression of dopamine transporter (DAT) was significantly up-regulated in Dominants compared to Subordinates. In addition, Dopamine 1 receptor (D1R) expression was down-regulated in Subordinates compared to Dominants, suggesting that there is a social-status dependent regulation of the dopaminergic (DA) system. Finally, we show that visual cues play an important role in regulating zebrafish dominance relationships and the prioritization of different motor outputs by using a zebrafish line lacking pigmentation. Taken together, our results reveal that neuromodulation by DA signaling and visual information provides a mechanism for the nervous system to adapt to changes in social conditions by permitting the animal to prioritize a socially appropriate behavioral response
The Effect of Visual Queues on Social Status Formation and Mauthner Sensitivity in Zebrafish
No full textZebrafish of the TLN strain, a genetic variant of the Tüpfel Long-fin (TL) wild-type zebrafish that does not display the trademark striped scale pattern of wild-type zebrafish, but are instead pale, were tested through a series of behavioral observations to determine the role of visual queues in dominant/subordinate relationships. TLN zebrafish did not show significant differences in swimming behavior, escape response, or escape response latency, suggesting that it is possible that visual queues play a role in the formation of dominant/subordinate relationships, and in turn would affect the neurological sensitivity of the escape pathway when exposed to a threatening auditory stimulus
THE EFFECT OF SOCIAL EXPERIENCE ON BRAIN MORPHOLOGY IN ZEBRAFISH (DANIO RERIO)
No full textAdult male Zebrafish form stable dominance relationships that consist of dominant and subordinate animals. The long-term effects of social dominance on nervous system function remain poorly understood. Here, we investigated how social stress affects the morphological architecture of the hypothalamic dopaminergic nuclei implicated in social regulation and aggression. The dopaminergic system is a prime target of social factors given dopamine’s (DA) involvement in aggression, depression, stress and anxiety. We tested the hypothesis that social dominance induces morphological reorganization of the hypothalamic dopaminergic Posterior Tubercular rostral (PTar) and Caudal (PTac) nuclei. Both nuclei receive visual and olfactory social cues and relay the integrated information to spinal cord locomotor circuits to modulate locomotor activity. To test the effect of social dominance on the number of DA neurons we used the transgenic zebrafish line Tg(dat:EGFP) that expresses EGFP in all CNS dopaminergic neurons. Using confocal imaging of brain slices coupled with digital analysis, we compared the number of DA neurons in the PT nuclei between dominant and subordinate animals after two weeks of continuous social interactions; while group-housed (6 communally housed males) served as a control group. Our results show a significant increase in the number of DA neurons in dominants compared subordinate animals, while the number of DA neurons in communals was not statistically significant from neither dominants nor subordinates. Our results demonstrate that social dominance induces morphological reconfiguration of the hypothalamic DA system in a social status-dependent manner, and likely to have broader implications on other vertebrate social species
THE EFFECT OF SOCIAL EXPERIENCE ON THE SYNERGISTIC NEUROMODULATION OF MOTOR CIRCUITS
No full textForming social groups is a critical adaptive strategy for the survival of many animal species, where conflict between members can result in dominance relationships. Dominance relationships are often formed through aggressive interactions and influence the cognitive and physiological properties of individuals in a status-dependent manner. The aim of this dissertation is to discern how zebrafish utilize aggression and visual cues to reinforce a stable dominance relationship, and what effect stable dominance has on the neurobiological processes underlying motor behavior, with a focus on dopamine signaling. Dopamine has been implicated in aggression, social regulation, and modulation of motor circuits, making it a central point of study regarding the relationship between social dominance and motor behavior. Using behavioral, pharmacological, and genetic approaches, I have found that (1) in addition to physical aggression, zebrafish alter their stripe coloration intensity to reinforce dominance, (2) stable dominance alters dopamine signaling through decreased expression of the dopamine transporter and the dopamine receptor type-1b, (3) changes in dopamine signaling manifest in altered escape and swimming locomotor behavior, and (4) dopamine acts synergistically with other neurotransmitters to regulate status-dependent motor circuit activation. Results from this dissertation have provided evidence for how stable dominance impacts modulation of motor circuits, revealing how changes in relative excitability of multiple neuromodulatory inputs provide a mechanism for the nervous system to adapt to changes in social conditions and allow animals to select a socially appropriate behavioral response
THE ROLE OF THE LATERAL LINE IN EARLY LARVAL ZEBRAFISH AND CONTRIBUTION TO MULTISENSORY BEHAVIORS
No full textThe lateral line is a hair cell-based sensory system that is important for multisensory behaviors like schooling, rheotaxis, and predator/prey detection in aquatic vertebrates. This dissertation work uses the first genetic model for the congenital loss of lateral line function in zebrafish. We have found zebrafish ohnologs of a gene required for hair cell function whose mRNA expression patterns are cleanly partitioned between inner ear and lateral line hair cell populations. Genetic disruption of each ohnolog results in specific loss of either auditory/vestibular function or lateral line function. Since lateral line mutants exhibit normal auditory and vestibular behaviors, we can investigate lateral line-mediated behaviors in developing larval zebrafish. In my second chapter, I develop a repeated treatment timeline using neomycin sulfate to continually ablate the lateral line in early larval zebrafish to compare to the genetic lateral line mutant. I examine hair cell regeneration and proliferation rates, as well as associated toxicity of repeated neomycin treatments in larval zebrafish. In conclusion, I find that low doses of neomycin sulfate treatments every 12 hours in 3-4 day old zebrafish are sufficient to continuously disable lateral line function. In my third chapter, I begin my analysis of these "lateral line-less" fish to investigate the role of the lateral line in modulating expression of the social hormone parathyroid hormone 2. Parathyroid hormone 2 (Pth2) is a vertebrate-specific neuropeptide for which thalamic expression is upregulated by social contact with conspecifics. In this project, I measure pth2 levels in zebrafish mutants lacking hair cell function in either the lateral line only, or in both the inner ear and lateral line. Socially raised lateral line mutants express lower levels of pth2 relative to wild-type siblings, but there is no further reduction when all sensory hair cells are nonfunctional. However, social isolation of hair cell mutants causes a further reduction in pth2 expression, pointing to additional unidentified sensory cues that influence pth2 production. Lastly, I report that social context modulates fluorescent transgenes driven by the pth2 promoter. Altogether, these data suggest that lateral line mutants experience a form of isolation, even when raised socially. In my fourth chapter, I characterize a novel swim bladder over-inflation phenotype associated with the lateral line mutants. Larval zebrafish achieve neutral buoyancy between 3-4 days post-fertilization by gulping air from the water's surface to inflate their swim bladders. We define this behavior of swimming to the air-water interface as "surfacing." Little is known about the sensory basis for this underappreciated behavior of larval fish. I observe that approximately half of lateral line mutants over-inflate their swim bladder during initial inflation and become positively buoyant. Thus, I hypothesize that larval zebrafish use their lateral line to sense the air-water interface during the surfacing behavior to regulate swim bladder inflation. I report that (i) over-inflation is caused by abnormal surfacing behaviors in lateral line mutants, (ii) lateral line defects are responsible for swim bladder over-inflation, and (iii) the lateral line is specifically responsible for surface detection during initial inflation. In summary, I reveal a novel sensory basis for achieving neutral buoyancy where larval zebrafish use their lateral line to sense the air-water interface and regulate initial swim bladder inflation. In my fifth chapter, I characterize surfacing as a multi-sensory behavior that involves photosensory and chemosensory cues in addition to mechanosensory cues. Only half of lateral line mutants over-inflate their swim bladder, so I hypothesize that other sensory systems, like photosensory cues, help larvae to properly perform the surfacing behavior for initial swim bladder inflation. In this chapter, I use a combination of genetic and environmental manipulations to investigate the roles of ocular and non-ocular photosensation, social isolation, and chemosensory cues on initial swim bladder inflation in the context of genetic loss of lateral line function. Overall, I find that initial swim bladder inflation in larval zebrafish is affected by non-ocular photosensory and chemosensory cues when larvae are genetically deprived of both visual and lateral line sensation
SOCIAL REGULATION OF THE ENDOCANNABINOID SYSTEM IN ZEBRAFISH MOTOR CIRCUITS
No full textSocial status-dependent modulation of neural circuits has been investigated extensively in vertebrate and invertebrate systems. However, the effect of social status on shifting the balance in activation between competing neural circuits is poorly understood. Zebrafish (Danio rerio) form stable social relationships that consist of socially dominant and subordinate animals. Once the social hierarchy is formed, social status-dependent differences in behavior patterns emerge. Subordinate animals startle more readily in response to auditory stimuli, while dominants swim at a higher frequency than subordinates. Here, we investigated the role of the endocannabinoid system (ECS) in regulating the activation of the swim and escape circuits based on social status. Our aim was to investigate how the ECS facilitates the transition between swim and escape circuits in socially dominant and subordinate animals. Endocannabinoids act as retrograde signaling molecules between neurons and are implicated in inhibition of both excitatory and inhibitory neurotransmission via retrograde binding of the cannabinoid 1 (CB1) or cannabinoid 2 (CB2) receptor. A previous study revealed a novel role for the endocannabinoid 2-Arachidonoylglycerol (2-AG) in modulating the switch in activation between the swim and startle circuits in zebrafish. The ECS can be up- or down-regulated by altering levels of 2-AG or targeting CB1 receptor function. To better understand how social status regulates the ECS and its effects on circuit activation, we studied the effects of two drugs, AM-251 and JZL184, on the regulation of status-dependent differences in swim and escape behavior. AM-251 competitively blocks endocannabinoid signaling by binding to CB1 receptor, while JZL184 increases 2-AG concentration by inhibiting monoacylglycerol lipase (MAGL), the degradative enzyme for 2-AG. First, we show that increasing ECS activity via intramuscular injection of JZL184 differentially affects swim and escape behavior according to social status. Secondly, we show that block of CB1 function with AM-251 reduces startle sensitivity and swimming frequency, and that its effects are concentration dependent. Thirdly, we utilize a dopamine receptor 1 knockout fish (D1KO) to demonstrate that the effects of ECS modulation on startle involves the dopamine D1 receptor system. Collectively, these findings support the notion that the ECS, as reflected by changes in swimming and escape behavior in response to treatment with JZL184 and AM-251, is socially regulated and involved in the social status-dependent shift in the balance of motor circuit activation, and that these effects are mediated in part via dopaminergic pathways. Our results represent an important step forward in the field of social neuroscience and better define the path toward a comprehensive understanding of the molecular factors that control social behavior
SOCIAL REGULATION OF THE A11 DOPAMINERGIC NUCLEUS IN ZEBRAFISH (Danio rerio)
No full textAggression is a universal behavioral feature that is present in majority of social animals. In nature, aggression facilities the formation and stabilization of social relationships and allow animals to divide resources according to rank. In zebrafish, aggressive interactions lead to stable dominance formation that consist of dominants and subordinates. Although social-status-dependent differences in behavior must arise due to neural plasticity, mechanisms of how neural circuits are reconfigured to cope with aggression and social stress are poorly described. Here, I describe how the hypothalamic dopaminergic A11 nucleus, is morphologically influenced by social status in male zebrafish. The A11 integrates and relays sensory social information to spinal motor circuits to modulate behavior; thus, the A11 is potentially prone to socially induced plasticity. I hypothesized that morphological plasticity of the A11 is differentially regulated in animals of different social ranks. I tested my hypothesis by combining non-invasive behavioral observations and histological approaches. Specifically, I used Tg(dat:eGFP) zebrafish, that specifically express eGFP in the dopaminergic system to investigate how dyadic agonistic interactions affect the A11 morphology and axosomatic network connectivity, then correlated morphological plasticity with changes in motor activity. I show that A11 cell number and synaptic connectivity are socially regulated. The number of A11 neurons increases in socially dominant animals as social relationships mature, while synaptic connectivity is increased in subordinates. Secondly, I show that these morphological differences are reversable as social environment evolves and correlate with adaptations in motor activity. Results from this project improve our understanding of how social dominance induces morphological plasticity of social decision-making networks and their influence on behavior
An examination of scar modelling and assessment methods for the evaluation of treatments
Full text linkScarring is the final common pathway for healing within the skin irrespective of age, gender and race. Scars can be itchy, painful, tight and above all, cosmetically disfiguring. Despite advances in surgery and trauma management, there is currently no reliably effective treatment for reducing or preventing scarring. The primary aim of this thesis is to assess the currently available models for scarring and evaluate/further develop the utility of current assessment tools, in an effort to design a pilot randomised control trial for silicone gel treatment of scars.
Review of scar models in humans and animals demonstrated the limitations and drawbacks of many existing methods to assess scar treatments. Examination of currently used subjective and objective scar assessment tools in a plastic surgery clinic helped to develop protocols and methodology for a scar treatment research trial. Long-term scar outcomes assessed using a novel subjective patient reported outcome measure for paediatric burn patients demonstrated no statistically significant difference between those treated surgically, and those treated conservatively. A pilot randomised control trial to produce high quality evidence for silicone gel sheeting in the treatment of scars was set up and successful in recruitment.
Scarring remains a difficult condition for clinicians to manage, with many treatments utilised on a poor evidence basis. Here, we have demonstrated difficulties in establishing a scientific scar treatment model; and created a pilot study that will help to provide high quality evidence for the efficacy of silicone gel sheeting as a treatment for scars
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