The effect of genetic susceptibility and immunosuppressant treatment on mononuclear cell phenotype in multiple sclerosis

Abstract

Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS). The immune system plays a significant role in the pathophysiology of the disease and research into the mechanisms that may be potentially contributing to the disease is essential to help advance knowledge in the field and ultimately seek better outcomes for MS patients. A key set of cells that are believed to play a role in MS immunopathology are mononuclear cells which consist of B-cells, T-cells, monocytes and their derivatives. The aims of this thesis was to assess the effects of 1) genetic susceptibility and 2) immunosuppressant treatment, on mononuclear cell phenotype in MS. The first aim involved interrogating a key co-stimulatory molecule, cluster of differentiation 40 (CD40), expressed on mononuclear cells which is known to play a fundamental role in the regulation of the humoral immune response. CD40 is a risk gene for the development of MS and I sought to assess the effects of the single nucleotide polymorphism (SNP) rs1883832 on the phenotype of CD40-expressing mononuclear cells. Utilising a range of bimolecular methods and samples taken from MS individuals and healthy matched-controls (HC), I showed that SNP rs1883832, particularly the ‘risk’ genotype (homozygous TT) was correlated with reduced soluble CD40 (sCD40) levels in HC, but not MS patients. I did not observe any phenotype-dependent differences in sCD40 levels between MS (in quiescent state) and during active demyelination compared to HC. Upon assessment of key cytokines produced by B-cell and monocytes, I observed elevated gene expression of the pro-inflammatory cytokine interleukin-1 beta (IL-1β) in the MS population linked to the CD40 risk TT-genotype. In the second component, I showed that the immunosuppressant drug Cladribine, is highly toxic to mononuclear cells, and that it down-regulates the expression of the key phagocytic marker MERTK at the protein level (preliminary data, study is ongoing). In summary, the data presented in this thesis suggests that the CD40 risk gene and the immunosuppressant Cladribine play roles in altering the phenotype in mononuclear cells. The understanding from this body of work provides some insights into how genetic susceptibility can drive changes in mononuclear cell phenotype which may impact on the modulation of co-stimulatory mechanisms in humoral immunity. Moreover, the findings from the Cladribine study may assist the field in the discovery of alternative mechanisms of the drug on mononuclear cells and therefore inform a refined approach to treatment of MS.