Epigenetic dysregulation of adaptive immunity in multiple sclerosis

<jats:p>&lt;p dir="ltr"&gt;Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS), in which immune cells contribute to neuroinflammation, demyelination, and neurological disability. Although current therapies reduce inflammatory activity, the molecular mechanisms underlying immune dysregulation and treatment response remain incompletely understood. This thesis investigated epigenetic and transcriptional regulation of immune cells in MS, with a focus on cerebrospinal fluid (CSF) cells as a CNS-proximal immune compartment.&lt;/p&gt;&lt;p dir="ltr"&gt;Two low-input DNA methylation methods, enzymatic methyl sequencing (EM- seq) and post-bisulfite adaptor tagging (PBAT), were first compared. Both methods quantified DNA methylation from limited DNA input, but EM-seq showed better library quality and coverage, supporting its use for clinical samples with limited material. Genome-wide DNA methylation profiling of CSF cells from people with relapsing-remitting MS and controls then identified disease-associated epigenetic alterations enriched in immune activation, cell adhesion, migration, and transcriptional regulation pathways. Integration with gene expression data linked these changes to altered immune regulation and suggested a connection between immune dysregulation and CNS-related molecular pathways, including protocadherin genes.&lt;/p&gt;&lt;p dir="ltr"&gt;The effects of rituximab, a B cell-depleting anti-CD20 monoclonal antibody, were investigated in paired CSF samples collected before and after treatment. Rituximab was associated with reduced inflammatory activity and coordinated changes in DNA methylation, gene expression, and immune cell composition, indicating that B cell depletion reshapes the CNS-proximal immune environment beyond direct removal of B cells. Finally, paired transcriptomic and DNA- methylation profiling of auto-proliferating memory B cells identified an inflammatory, ABC-like state characterized by IFN-y- and NF-KB-related transcriptional activation and widespread hypomethylation, including at genes related to the STAT1-T-bet-CXCR3 axis.&lt;/p&gt;&lt;p dir="ltr"&gt;Together, this thesis shows that epigenetic and transcriptional profiling of rare immune cell populations can provide important insights into MS pathogenesis and treatment response and highlights regulatory pathways relevant for future biomarker and therapeutic development.&lt;/p&gt;&lt;h3 dir="ltr"&gt;List of scientific papers&lt;/h3&gt;&lt;p dir="ltr"&gt;I. &lt;b&gt;Han, Y.&lt;/b&gt;, Zheleznyakova, G.Y., Liang, H., Sorini, C., Pahlevan Kakhki, M., Ruffin, N., Hallén, N., Rao Prakash, C., Beckers, V., Ivanova, E., Khademi, M., Olsson, T., Karlsson, M.C.I., Piehl, F., Kelsey, G., Kular, L., Needhamsen, M. and Jagodic, M., 2026. DNA methylation landscape of cerebrospinal fluid cells in multiple sclerosis: an epigenome-wide association study. bioRxiv, 2026.02.25.708054. [Accepted]&lt;/p&gt;&lt;p dir="ltr"&gt;II. &lt;b&gt;Han, Y.&lt;/b&gt;, Zheleznyakova, G.Y., Marincevic-Zuniga, Y., Kakhki, M.P., Raine, A., Needhamsen, M. and Jagodic, M., 2022. Comparison of EM-seq and PBAT methylome library methods for low-input DNA. Epigenetics, 17(10), pp.1195-1204. &lt;a href="https://doi.org/10.1080/15592294.2021.1997406" target="_blank" rel="noreferrer"&gt;https://doi.org/10.1080/15592294.2021.1997406&lt;/a&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;III. &lt;b&gt;Han, Y.&lt;/b&gt;, Starvaggi Cucuzza, C., Dethlefsen, O., Ewing, E., Noori, P., Khademi, M., Al Nimer, F., Piehl, F., Needhamsen, M. and Jagodic, M. Rituximab induces epigenetic remodeling of cerebrospinal fluid immunity in multiple sclerosis. [Manuscript]&lt;/p&gt;&lt;p dir="ltr"&gt;IV. Jelcic, I., Naghavian, R., Fanaswala, I., Macnair, W., Esposito, C., Calini, D., &lt;b&gt;Han, Y.&lt;/b&gt;, Marti, Z., Raposo, C., Sarabia Del Castillo, J., Oldrati, P., Erny, D., Kana, V., Zheleznyakova, G., Al Nimer, F., Tackenberg, B., Reichen, I., Khademi, M., Piehl, F., Robinson, M.D., Jelcic, I., Sospedra, M., Pelkmans, L., Malhotra, D., Reynolds, R., Jagodic, M. and Martin, R., 2025. T-bet+ CXCR3+ B cells drive hyperreactive B-T cell interactions in multiple sclerosis. Cell Reports Medicine, 6(3), 102027. &lt;a href="https://doi.org/10.1016/j.xcrm.2025.102027" target="_blank" rel="noreferrer"&gt;https://doi.org/10.1016/j.xcrm.2025.102027&lt;/a&gt;&lt;/p&gt;</jats:p>