Nerve growth factor (NGF) represents a new therapeutic strategy for multiple sclerosis (MS) because of its immunomodulatory and neuroprotective activity. To analyze changes in NGF expression experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats. In the dorsal root ganglia (DRG) of animals with EAE, NGF mRNA and protein increased between 18 - 24 days post induction (dpi) during complete neurological recovery. In the spinal cord (SC) of animals with EAE, NGF mRNA and protein expression increased at 15 dpi and 12 dpi, respectively, to reduce EAE-induced disability. We identified the 25 kDa pro-NGF as a biologically active isoform during EAE. EAE SC axons demonstrate a loss or thinning of myelin which correlated with maximal neurological disability. NGF plays a role in minimizing EAE-induced inflammation and myelin damage to promote neurological recovery. NGF may be an “off switch” for a cytokine-neurotrophin signaling triad to govern the extent of myelin damage.

Encephalitis lethargica affected a large number of people in the pandemic in the early 1900s (von Economo, 1930). Histological and biochemical data suggest that autoimmune mechanisms play an important role in this disorder and recently serum anti-basal ganglia antibodies (ABGA) have been detected in affected sporadic cases associated with evidence of recent streptococcal infection (Dale et al., 2004a). ABGA are also associated with other neuropsychiatric disorders including Sydenhams chorea, paediatric autoimmune neuropsychiatric disorders associated with streptococcal infections, Tourettes syndrome and obsessive compulsive disorder. The precise frequency, presentation, disease course, treatment response and causes of these disorders are still unknown. As ABGA are strongly associated with recent streptococcal infection, these disorders represent a potentially good model for the study of molecular mimicry and autoimmunity. The present study focussed on various aspects of this group of disorders. Profiling of group A streptococcus isolates from both patients with postulated post-streptococcal disorders of the CNS and controls was done which highlighted differences in virulence factors like M protein and superantigens between the two groups. Also in the present study we demonstrated the pathogenicity of anti-neuronal antibodies found in patients in both an in vivo and in vitro setting. An animal model of the disorders was produced by passive transfer of antibodies from patients which resulted in symptoms reminiscent of diseases like encephalitis lethargica and dystonia. An active immunization animal model using GABHS proteins and recombinant proteins (putative autoantigens) was also developed. Furthermore, the autoantibodies from patients and animal models were analysed on both neuronal and non-neuronal cells where they demonstrated to have a functional effect on cytotoxicity, apoptosis, calcium flux and enolase activity. N-methyl D-aspartate glutamate receptor and voltage gated potassium channel have been recently been implicated in a 16 range of neurological disorders, hence we also tested the patient sera for antibodies against these receptors and found a group of patients to be positive. In summary, EL and other ABGA-associated disorders are still an emerging entity, with major implications for neuropsychiatry. As auto-antibody mediated diseases respond to immunomodulatory therapy, identifying and defining the pathogenesis of these disorders is important so that patients can be appropriately treated.

Prion diseases are a group of fatal neurodegenerative protein-misfolding diseases. Microglia, the resident myeloid cells found within the brain, have been shown to demonstrate a reactive morphology during the disease process with conflicting evidence for both a neurotoxic and neuroprotective role. The studies presented here aimed to investigate the role of microglia activation using transcriptomic and morphological analysis of prion disease in mice. Initially, the host immune response to prion disease was explored using a publically available mouse prion disease dataset. Re-analysis of this dataset was performed using BioLayout Express3D; a novel software tool that supports the visualisation and clustering of correlation networks. Disease-associated genes up-regulated during the later stages of infection were present in two main clusters. The cellular origin of these genes was explored by examining their expression in a dataset comprised of pure populations of cells. This demonstrated that the primary cluster of up-regulated transcripts encompassed genes expressed mainly by microglia and to a lesser extent astrocytes and neurons. The secondary cluster comprised almost exclusively of interferon response genes. The conclusions of these analyses were different from those of the original study that suggested disease-associated genes were primarily neuronal in origin. Mouse models of prion disease were established by infecting a novel line of BALB/cJ inbred mice, expressing EGFP under control of a myeloid specific Csf1r promoter, with the 79A prion strain. Quantification of the morphological changes of EGFP expressing microglia suggested the cells accumulated in the medulla at sites of early misfolded protein deposition with minimal change in their overall appearance. An activated microglia morphology was not observed until protein deposition was extensive. Isolation of EGFP expressing microglia was performed for transcriptome analysis. The vast majority of disease associated genes demonstrated increased expression at the onset of clinical symptoms. The gene list was found to be highly enriched for genes associated with an innate immune response regulated by the NFκB signalling cascade. Also highly enriched were processes associated with protein translation, energy production and stress response. These data suggest a high metabolic load is burdened by proliferating microglia; and as part of a response which is strikingly more pro-inflammatory in nature than has previously been attributed to the microglia phenotype within prion disease. As an active contributor to normal homeostasis, microglia are more than just innate immune surveillance and are now considered an integral component in both the healthy and diseased brain. The ramifications of activation toward the microglia phenotype shown here will have direct and potentially cytotoxic influence on neighbouring microglia and other brain cell types implying microglia as major contributors to the neurotoxic environment found within the CNS during prion disease. Furthermore the identification of genes associated with metabolism offer many intriguing possibilities for manipulating the activity of microglia in pre-clinical therapeutic intervention.

The regulation of the immune system is complex, with many factors involved in controlling immune cell development, activation and homeostasis. These factors include neuropeptides as well as classic immunoregulatory molecules such as cytokines, chemokines and hormones. Neuropeptides and tachykinins in particular are known to be involved in immune response modulation through a cascade of events including vasodilation, plasma extravasation, the activation of immune cells, the secretion of pro-inflammatory cytokines and the recruitment of more immune cells. Furthermore, there is growing evidence that tachykinins play a role in hematopoiesis with Substance P as the proposed effector molecule. In 2000, our lab discovered a new tachykinin with remarkable structural similarity to SP and SP-like neurokinin receptor binding affinity. This molecule was designated Hemokinin-1 due to its expression in hematopoietic cells and its function in B cell development. Further gene expression analysis of HK-1 reveals a wide expression pattern although HK-1 transcripts are found predominantly in peripheral tissues while SP is mainly expressed in neuronal tissue. Based on this differential expression pattern, it has been suggested that HK-1 may act as the peripheral tachykinin and may have functions distinct from SP. In addition, given the crossreactivity of the SP antibodies to HK-1, it is important to determine whether HK-1 is the actual mediator of some functions previously attributed to SP. In this thesis, we examine the differential expression pattern of HK-1 to determine molecular mechanisms of regulation of HK-1 transcription and ultimately provide clues to its function in the immune system. In our analysis of the HK-1 promoter, we found a major difference in the basic transcriptional control of HK-1 and SP at the level of transcription initiation and identified several transcription factors including CREB and NFκB involved in regulating TAC4 gene expression in immune cells. Data presented in this thesis also reveal that the HK-1 gene is a direct target of Early B-cell Factor, a transcription factor known to activate B cell-specific genes as well as genes involved in adipogenesis and neuronal development. Our results show EBF regulates HK-1 gene expression in differentiating B cells as well as a monocytic cell line. Our data indicate EBF may also be responsible for the high levels of HK-1 transcript in the olfactory epithelium, suggesting a bridge between the nervous system and the immune system.

Multiple sclerosis (MS) is a leading cause of neurological disability in young adults. Although current treatments can reduce symptomology and relapse rate, they are unable to prevent the chronic neurodegeneration that occurs at later stages. MS pathology is mediated by complex interactions between invading immune cells, neurons, glia, and endogenous stores of neural progenitor cells (NPCs). Factors critical to NPC/immune cell communication as well as the survival, differentiation, and proliferation of NPCs are not well defined. Elucidation of these factors will allow for the advancement of NPC transplantation therapies as well as the identification of novel pharmacological targets. Fas – a member of the tumor necrosis superfamily of death receptors – has diverse, cell-specific functions and is a major modulator of autoregulation within the immune system. Although Fas is expressed by NPCs, its exact role in this cell type was previously unknown. To contribute to this body of knowledge, the experiments in this dissertation examined the role of the Fas receptor (Fas) and Fas ligand (FasL) in NPC survival, differentiation, and T-cell cross-talk in vitro and in vivo in experimental autoimmune encephalomyelitis (EAE; a well-established animal model of MS). Activation of Fas via FasL increased NPC survival by decreasing apoptosis (as opposed to increasing proliferation) in vitro. This decreased apoptosis correlates with upregulation of the inhibitor of apoptosis protein (IAP) Birc3. Further investigation into the importance of Fas in NPCs was accomplished by comparing wild-type and Fas-deficient (lpr) NPCs. Lpr NPCs exhibited decreased apoptosis, decreased proliferation, and increased differentiation to oligoprogenitor and neuronal lineages. These studies suggest the Fas system plays multifaceted roles in NPCs and that its exact functions are dependent on both functional Fas expression and presence or absence of FasL. To determine the role of Fas/FasL in neuroimmune cross-talk, co-cultures of wild-type or lpr NPCs with different T-cell subtypes (Th1, Th2, and Th17 cells) were performed. Th1 cells were the only subtype capable of inducing NPC apoptosis. Th1-mediated death was dose-dependent and was not mediated via Fas. On the other hand, NPCs were able to induce significant apoptosis in pro-inflammatory Th1 and Th17 cells without affecting anti-inflammatory Th2 cells. NPC-induced Th17 cell death was mediated via Fas. These data suggest NPCs can specifically target pro-inflammatory T-cells and can promote neuroprotection by inducing death of these proencephalogenic cells. Finally, intravenous injection of wild-type or lpr NPCs into EAE mice reduced clinical symptoms and CNS immune infiltrate to the same extent. Few NPCs enter the CNS, where they remain undifferentiated. This suggests the main mechanism through which NPCs produce beneficial results in EAE is via peripheral immunoregulation, which is not dependent on Fas expression. Overall, this dissertation elucidates the Fas system as an important modulator of NPC cell-fate and immunoregulatory capacity.

Neuromodulation of the immune system has been described to be influenced by hemispheric lateralisation (HL), the stable tendency to relatively utilise one hemisphere or its functions over another. To date there has not been a systematic review of research in this phenomenon conducted, and only one study has examined the effects of HL on the progression of a disease – Human Immunodeficiency Virus (HIV). That research was conducted on a small sample with little control for confounders. The present work sought to compile a systematic review of literature concerning HL and immunity in humans, using effect size analysis. Further, the present work also describes an empirical advancement of this earlier HIV study with stricter control over confounds in a larger sample. The findings corroborated the theory of asymmetrical immune influence by HL via the systematic review showing clear, relatively consistent and strong relationships between left-HL and immunopotentiation. The empirical prospective study extended current knowledge of this relationship in HIV to identify a moderator – HAART treatment. Specifically, left-HL predicted better immunity in HIV-1 patients independent of confounders, with further findings of the same pattern in untreated patients, but not in HAART-treated patients. Further observations were made between HL and HIV-relevant behaviours, again adding to current knowledge. The finding of left-HL being associated with fewer sexual partners in Europeans presents new information of relevance to public health. The combined findings of the present work suggest that left-HL has predictive value in illness (HIV-1) and in general immunity. The present work adds to the existing knowledge new information concerning a moderating factor of the HLimmunity relationship in HIV, and behavioural implications of HL which impact upon HIV disease. Potential explanations for moderation, proposals for neurobiological mechanisms and direction towards future, more rigourous study in the field, both in HIV and immunity, are discussed.

Interactions between the immune system and the nervous system are currently underappreciated, assumed to play minor mechanistic roles in disease pathogenesis. In contrast, our laboratory has demonstrated the importance of this relationship with significant impact, initially in Type 1 Diabetes (T1D). The experiments presented here build on our previous work to provide insights into the etiology of Multiple Sclerosis (MS) and Type 2 Diabetes (T2D). Transient Receptor Potential Vanilloid-1 (TRPV1) is an ion channel expressed on peripheral sensory afferent neurons that fundamentally control T1D pathogenesis. Here we show that mice with genetic ablation of TRPV1 are protected from EAE progression, attributable to reduced central nervous system (CNS) leukocyte passage. The pathogenic role of TRPV1 in permeabilizing the blood-CNS barriers may also translate to MS, as patients with progressive disease show a significant mutation bias within the TRPV1 gene. We were simultaneously intrigued by the growing worldwide obesity epidemic, and we observed that obese mice develop more severe EAE compared to lean mice. This was mechanistically linked to an expansion of TH17 cells, driven by sustained rises of IL-6 in obese mice. This research implies new therapeutic opportunities for the many obese patients with diverse autoimmune diseases. Finally, the immune system, obesity, and T2D are functionally linked, and we contributed to research that uncovered a large presence of immune cells in adipose tissue that drive insulin resistance. Manipulation of T cells and B cells affects local inflammation as well as whole-body insulin resistance and glucose homeostasis. Intriguingly, auto-antibodies in insulin resistant individuals are specific for a number of unique proteins, including glial fibrillary acidic protein (GFAP), initially shown by our laboratory to play a key role in T1D progression. We further characterized the autoimmune and neuronal progression elements that steer disease pathogenesis, and observed that administration of a vaccine containing GFAP is able to dramatically reduce weight gain and insulin resistance in mice. The data presented in this thesis provide a number of novel, mechanistic observations linking the immune and nervous systems in disease, and implies several potential avenues for treatment.

Interactions between the immune system and the nervous system are currently underappreciated, assumed to play minor mechanistic roles in disease pathogenesis. In contrast, our laboratory has demonstrated the importance of this relationship with significant impact, initially in Type 1 Diabetes (T1D). The experiments presented here build on our previous work to provide insights into the etiology of Multiple Sclerosis (MS) and Type 2 Diabetes (T2D). Transient Receptor Potential Vanilloid-1 (TRPV1) is an ion channel expressed on peripheral sensory afferent neurons that fundamentally control T1D pathogenesis. Here we show that mice with genetic ablation of TRPV1 are protected from EAE progression, attributable to reduced central nervous system (CNS) leukocyte passage. The pathogenic role of TRPV1 in permeabilizing the blood-CNS barriers may also translate to MS, as patients with progressive disease show a significant mutation bias within the TRPV1 gene. We were simultaneously intrigued by the growing worldwide obesity epidemic, and we observed that obese mice develop more severe EAE compared to lean mice. This was mechanistically linked to an expansion of TH17 cells, driven by sustained rises of IL-6 in obese mice. This research implies new therapeutic opportunities for the many obese patients with diverse autoimmune diseases. Finally, the immune system, obesity, and T2D are functionally linked, and we contributed to research that uncovered a large presence of immune cells in adipose tissue that drive insulin resistance. Manipulation of T cells and B cells affects local inflammation as well as whole-body insulin resistance and glucose homeostasis. Intriguingly, auto-antibodies in insulin resistant individuals are specific for a number of unique proteins, including glial fibrillary acidic protein (GFAP), initially shown by our laboratory to play a key role in T1D progression. We further characterized the autoimmune and neuronal progression elements that steer disease pathogenesis, and observed that administration of a vaccine containing GFAP is able to dramatically reduce weight gain and insulin resistance in mice. The data presented in this thesis provide a number of novel, mechanistic observations linking the immune and nervous systems in disease, and implies several potential avenues for treatment.

The regulation of the immune system is complex, with many factors involved in controlling immune cell development, activation and homeostasis. These factors include neuropeptides as well as classic immunoregulatory molecules such as cytokines, chemokines and hormones. Neuropeptides and tachykinins in particular are known to be involved in immune response modulation through a cascade of events including vasodilation, plasma extravasation, the activation of immune cells, the secretion of pro-inflammatory cytokines and the recruitment of more immune cells. Furthermore, there is growing evidence that tachykinins play a role in hematopoiesis with Substance P as the proposed effector molecule. In 2000, our lab discovered a new tachykinin with remarkable structural similarity to SP and SP-like neurokinin receptor binding affinity. This molecule was designated Hemokinin-1 due to its expression in hematopoietic cells and its function in B cell development. Further gene expression analysis of HK-1 reveals a wide expression pattern although HK-1 transcripts are found predominantly in peripheral tissues while SP is mainly expressed in neuronal tissue. Based on this differential expression pattern, it has been suggested that HK-1 may act as the peripheral tachykinin and may have functions distinct from SP. In addition, given the crossreactivity of the SP antibodies to HK-1, it is important to determine whether HK-1 is the actual mediator of some functions previously attributed to SP. In this thesis, we examine the differential expression pattern of HK-1 to determine molecular mechanisms of regulation of HK-1 transcription and ultimately provide clues to its function in the immune system. In our analysis of the HK-1 promoter, we found a major difference in the basic transcriptional control of HK-1 and SP at the level of transcription initiation and identified several transcription factors including CREB and NFκB involved in regulating TAC4 gene expression in immune cells. Data presented in this thesis also reveal that the HK-1 gene is a direct target of Early B-cell Factor, a transcription factor known to activate B cell-specific genes as well as genes involved in adipogenesis and neuronal development. Our results show EBF regulates HK-1 gene expression in differentiating B cells as well as a monocytic cell line. Our data indicate EBF may also be responsible for the high levels of HK-1 transcript in the olfactory epithelium, suggesting a bridge between the nervous system and the immune system.

Thesis (M.Sc.)--Acadia University, 1998. / Includes bibliographical references (leaves 168-183). Also available on the Internet via the World Wide Web.