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In patients with multiple sclerosis (MS), microglia become activated due to the autoimmune inflammatory response which is directed against the central nervous system (CNS). Following the first disease relapse, microglia remain activated and do not return to a resting state during remissions. Chronically-activated microglia release inflammatory mediators that cause CNS tissue damage, and as such, MS progression has been associated with widespread, chronic microglial activation that correlates with neurodegeneration. To date, only one histone demethylase, Jmjd3, has been described to have a role in inflammation. In agreement with this, up-regulation of Jmjd3 expression was observed following microglial treatment with several pro-inflammatory stimuli, including a range of toll-like receptors ligands and cytokines, suggesting a universal role of Jmjd3 during microglial activation. Subsequent ChIP-qPCR assays revealed that Jmjd3 was recruited to the promoters of Il6, Ccl3, Ccl5 and Nos2 following activation, which, in turn, presented a decrease in their H3K27me3 levels. Using an experimental autoimmune encephalomyelitis (EAE) mouse model of MS, Jmjd3 expression was shown to be increased in activated microglia from mice in the acute and late phases of disease. Immunization with complete Freud’s adjuvant (CFA) alone, also caused microglial activation with Jmjd3 induction, indicating a CFA-mediated TLR2 and TLR4 stimulation of microglia. Further investigation, in which primary microglia were isolated from mice deficient in Jmjd3 (Jmjd3-/-), however demonstrated that the absence of Jmjd3 alone had no resultant effect on the expression of a subset of immune response and inflammation related genes, including the Jmjd3 target genes Il6, Ccl3, Ccl5 and Nos2, before or after activation. This suggested that Jmjd3 acts in concert with a repertoire of other demethylases to facilitate microglia activation, and as such was rendered redundant in this setting. Deciphering the epigenetic profile of microglia in MS and determining whether it is involved in the maintenance of chronic microglial activation in the progressive phase of the disease remains an important line of investigation, and through a clearer understanding of its role in MS pathophysiology, could lead to the development of novel therapeutic interventions in the future.

Bone is a highly dynamic organ system comprised of various cell types that are constantly working to maintain the health and stability of bone. The main cells involved are the osteoblasts that form bone, the osteoclasts that degrade bone, and the osteocytes that act as sensors of the microenvironment and coordinate a response. An imbalance of the interactions between the cell types can potentially result in pathological states in bone at the microscopic level that can then affect the entire skeleton. Moreover, a number of genetic mutations can also lead to pathogenic changes in bone. An example of such is the development of sclerotic bone lesions in patients with the disease tuberous sclerosis complex. Tuberous sclerosis complex, or TSC, is an autosomal dominant disorder affecting approximately 1.5 million people worldwide. It is caused by a mutation in one of the genes encoding either member of the TSC1-TSC2 complex. Molecularly, TSC1-TSC2 negatively regulate the mechanistic target of rapamycin (mTOR) kinase in the mutli-protein complex mTORC1. Activation of mTORC1 leads to an upregulation of protein synthesis and cell growth. Tuberous sclerosis patients are heterozygous for TSC1 or TSC2, and post-natal loss of the second allele results in the development of multiple, benign, tumor-like hamartomas in various organ systems, most notably affecting the brain, kidneys, lungs, skin, and heart. Additionally, CT scans of patients reveal multiple loci of dense, compact bone termed sclerotic bone lesions. The bone lesions were most commonly seen in the posterior elements of the vertebrae and while they are asymptomatic, a remarkably high frequency of patients express them. To further investigate and better understand the mechanisms of tuberous sclerosis complex in bone, we analyzed a mouse model with heterozygous deletion in Tsc2. Initial examination showed the Tsc2+/- mice recapitulated tumors in various organ systems, most notably the kidney, and presented bone lesions in the pelvis and elements of the vertebrae. To further investigate the mechanism driving the disease state, we used a Cre driver thought to be specific for osteoclast (Cathepsin K-Cre, or Ctsk-Cre) to selectively delete Tsc2. Cathepsin K-Cre; Tsc2fl/fl mice exhibit a remarkably high bone mass. This study examined three specific aspects of this high bone mass phenotype. First, we sought to verify that the increased bone mass caused by Ctsk-Cre driven Tsc2 deletion was dependent on mTORC1 upregulation. This was done by generating Ctsk-Cre;Tsc2fl/fl mice lacking Raptor, a mTORC1 component essential for function. Next, we investigated the cell of origin driving the increase bone density by utilizing additional Cre drivers specific for osteoclasts and osteocytes. Additionally, we used radiation chimeras to assess if donated wild type cells could rescue the observed phenotype. We lastly explored the role of a secreted signaling molecule, CTHRC1, that has been proposed as a candidate to mediate osteoclast-osteoblast interaction, in the high bone mass phenotype of Ctsk-Cre;Tsc2fl/fl mice. Selective deletion of Tsc2 in bone cells provides an excellent model to investigate pathways regulating bone mass and strength and may provide new candidate targets for treating diseases of low bone mass, such as osteoporosis.

Doctor of Philosophy / Multiple sclerosis (MS) is the most common neurological disease affecting young adults. MS affects approximately 1 in 1000 people and, like other autoimmune diseases, women are more likely to be affected than men. The illness typically onsets between the ages of 20 and 40, and hence usually affects women of child-bearing age. The course of the MS is often unclear for years after diagnosis and since most women are diagnosed in their child-bearing years, they often have to make reproductive choices before their prognosis is clear and while the future remains uncertain. For women with MS, starting a family is an individual choice that needs to balance the importance of motherhood for the woman and her partner against the risks that she will be unable to care for the infant or child as a result of increasing disability. In other areas of medicine where finely balanced decisions are required, there has been a recent proliferation of decision aids that aim to inform people of the benefits and risks of opposing courses of action. In addition, decision aids help patients to weigh their values against the risks and benefits to make an informed decision. Despite the existence of over 200 decision aids to help patients consider decisions related to their medical conditions, not one exists that deals with the decision of whether or not to have a family for women with a chronic disability, such as MS. This thesis developed and evaluated a decision aid for women with MS to help them decide whether to start, forego or enlarge their families. The study utilised the criteria set out for the development of decision aids, according to the Cochrane Systematic Review of Patient Decision Aids (O'Connor et al., 2003). The first aim was to determine the proportion of women who are undecided about the motherhood choice and for whom a decision aid may be relevant. Results found that the motherhood choice was relevant to 46% of the women who responded to an initial mail-out. The second study aimed to establish women’s current concerns and thoughts regarding pregnancy and motherhood, and their response to the pilot decision aid. Twenty women participated in qualitative interviews and results supported previous findings that the mother’s health concerns, coping with parenting and societal attitudes are significant concerns when considering this decision. This study further identified concerns from different groups that had a direct impact on the decision to have children, including the experience of parenting, the child’s well-being and the timing and pressure of the decision. The main study was a randomised controlled trial of the decision aid aiming to determine whether the decision aid facilitated decision-making in women with MS. The study confirmed that the decision aid presented a balanced view to women, increased knowledge, reduced decisional conflict, increased decisional self-efficacy and certainty of the decision, and was free from adverse effects on psychopathology. The final component of the study was a 12 month follow-up which aimed to explore the long-term effectiveness of the decision aid and what aspects were valued by the women who received it. It was found that over time, women in the intervention group did maintain their certainty, but women in the control group also became more certain of their choice. At follow-up, the difference in certainty was no longer significant between the two groups. However, women did report that the intervention was useful in (a) providing access to information previously unavailable or difficult to obtain, (b) facilitating communication between women, their partners and health care professionals, (c) aiding them in considering and utilising their networks of support, and (d) preparing them for potential difficulties. In summary, this thesis developed and evaluated a decision aid for women with MS who are considering motherhood. The results showed that many women were undecided and, in the absence of good information on the topic, many women had concerns about pregnancy and parenthood. The decision aid was shown to be effective across a range of measures and free from adverse psychological effects. Hence, this is evidence-based resource can now be recommended for those women with MS who are currently contemplating motherhood.

Thesis (Ph.D.)--University of Delaware, 2007. / Principal faculty advisor: Todd D. Royer, Dept. of Health, Nutrition, and Exercise Sciences. Includes bibliographical references.

Thesis (M.S.)--University of Delaware, 2006. / Principal faculty advisor: James G. Richards, College of Health Sciences. Includes bibliographical references.

BACKGROUND: Differences in disease-modifying therapy (DMT) prescribing patterns between different groups of neurologists have not been explored. HYPOTHESIS: MS-specialist neurologists use a broader range of DMTs in contrast to generalist neurologists who preferentially prescribe Avonex. METHODS: Ontario neurologist demographic and geographical characteristics were linked to 2009 DMT prescription data. Lorenz curves and Gini coefficients were constructed to examine prescribing patterns; separating neurologist characteristics dichotomously and separating Avonex from the other DMTs. Gini Coefficients were compared using jack-knife statistical techniques to derive 95% confidence intervals. RESULTS: Prescriptions are highly concentrated with 12% of Ontario neurologists prescribing 80% of DMTs. High-volume prescribers show a broader range of DMT use while low-volume prescribers tend to use a particular DMT. CONCLUSIONS: The majority of DMTs are prescribed by a small subset of neurologists. High-volume prescribers show more variability in DMT use while low-volume prescribers tend to individually focus on a narrower range of DMTs.

BACKGROUND: Differences in disease-modifying therapy (DMT) prescribing patterns between different groups of neurologists have not been explored. HYPOTHESIS: MS-specialist neurologists use a broader range of DMTs in contrast to generalist neurologists who preferentially prescribe Avonex. METHODS: Ontario neurologist demographic and geographical characteristics were linked to 2009 DMT prescription data. Lorenz curves and Gini coefficients were constructed to examine prescribing patterns; separating neurologist characteristics dichotomously and separating Avonex from the other DMTs. Gini Coefficients were compared using jack-knife statistical techniques to derive 95% confidence intervals. RESULTS: Prescriptions are highly concentrated with 12% of Ontario neurologists prescribing 80% of DMTs. High-volume prescribers show a broader range of DMT use while low-volume prescribers tend to use a particular DMT. CONCLUSIONS: The majority of DMTs are prescribed by a small subset of neurologists. High-volume prescribers show more variability in DMT use while low-volume prescribers tend to individually focus on a narrower range of DMTs.

Low vitamin D status has been consistently associated with an increased risk of multiple sclerosis (MS). Further, preclinical and in vitro data demonstrate immune regulatory properties of 1,25-dihydroxyvitamin D that may be beneficial for patients with MS. To date evidence of beneficial in vivo immunomodulation by supplementation with vitamin D3 in humans is lacking. In a one-year, open-label, phase I/II dose-escalation study of vitamin D3 (average ~14,000 IU/d over one year) with calcium (1,200mg/d) in patients with MS, we compared the effects of treatment on safety outcomes, clinical outcomes and selected biomarkers of immune system activity, relative to matched MS patients [age, sex, disease duration, disease modifying therapy, and expanded disability status scale (EDSS)] randomized to receive no supplementation. Mean serum 25(OH)D concentrations were 78.1±27.0 nmol/L at baseline and at one-year were 82.7±34.8 and 179.1±76.1 nmol/L in control and treated groups, respectively. Serum and urinary calcium and all other safety outcomes were unchanged throughout the trial. Compared to controls, treated patients tended to have fewer relapses (McNemar, p=0.09) and a greater proportion had a stable or improved EDSS at study end (p=0.018). We observed significantly reduced lymphocyte proliferative responses to antigenic challenge in the treatment group at one year, compared to baseline and control group responses. High serum 25(OH)D concentrations were not associated with short-term adverse effects in patients with MS, but with evidence of clinical improvement and beneficial immunomodulation.

Low vitamin D status has been consistently associated with an increased risk of multiple sclerosis (MS). Further, preclinical and in vitro data demonstrate immune regulatory properties of 1,25-dihydroxyvitamin D that may be beneficial for patients with MS. To date evidence of beneficial in vivo immunomodulation by supplementation with vitamin D3 in humans is lacking. In a one-year, open-label, phase I/II dose-escalation study of vitamin D3 (average ~14,000 IU/d over one year) with calcium (1,200mg/d) in patients with MS, we compared the effects of treatment on safety outcomes, clinical outcomes and selected biomarkers of immune system activity, relative to matched MS patients [age, sex, disease duration, disease modifying therapy, and expanded disability status scale (EDSS)] randomized to receive no supplementation. Mean serum 25(OH)D concentrations were 78.1±27.0 nmol/L at baseline and at one-year were 82.7±34.8 and 179.1±76.1 nmol/L in control and treated groups, respectively. Serum and urinary calcium and all other safety outcomes were unchanged throughout the trial. Compared to controls, treated patients tended to have fewer relapses (McNemar, p=0.09) and a greater proportion had a stable or improved EDSS at study end (p=0.018). We observed significantly reduced lymphocyte proliferative responses to antigenic challenge in the treatment group at one year, compared to baseline and control group responses. High serum 25(OH)D concentrations were not associated with short-term adverse effects in patients with MS, but with evidence of clinical improvement and beneficial immunomodulation.