EBV dysregulation is associated with immune imbalance in multiple sclerosis

BACKGROUND AND OBJECTIVES: Epstein-Barr virus (EBV) infection is a prerequisite for the development of multiple sclerosis (MS), yet whether EBV acts merely as a trigger at disease onset or also contributes to immune dysregulation and disease progression remains unclear. To explore potential mechanisms linking EBV to immune alterations, we performed a comprehensive analysis of EBV markers and immune-related gene expression in peripheral blood samples from therapy-naïve persons with MS (PwMS) and healthy donors (HD) and assessed EBV transcripts in CSF cells to explore compartment-specific viral activity. METHODS: Peripheral blood mononuclear cells (PBMCs) and serum from PwMS (n = 77) and HD (n = 40) were analyzed. EBV serology, DNA load, and RNA expression were assessed by ELISA, droplet digital PCR, and preamplified real-time RT-PCR, respectively. EBV RNA was also evaluated in PwMS CSF cells. Gene expression profiling of 47 immune-related genes selected for their relevance to MS was also performed in PBMCs. Data were analyzed using univariate and multivariate statistical approaches also considering demographic, clinical, and radiologic information. Exploratory factor analysis (EFA) was used to identify transcriptional signatures associated with MS. RESULTS: Anti-EBNA1 IgG titers were higher in PwMS. In addition, EBV RNA and DNA were more frequently detected, and viral load was increased compared with HD. Notably, EBV transcripts associated with latency II/III (LMP1, LMP2A, EBNA1, EBNA3A) and lytic reactivation (BZLF1, gp350/220) were more prevalent in PwMS. Although viral RNA was detected in only 7% of CSF samples, all positive cases showed profiles consistent with viral reactivation. Immune gene expression analysis revealed broad upregulation of cytotoxic effectors, type I interferon pathways, and chemokine signaling in PwMS. EFA identified a significantly different gene signature linking BZLF1 expression with inflammatory genes, type I interferon responses, and chemokines involved in immune cell migration, in PwMS. DISCUSSION: Our findings support the hypothesis that EBV latency disruption and lytic reactivation contribute to immune dysregulation in MS. The association between EBV transcriptional activity and immune gene alterations may uncover potential peripheral biomarkers of EBV-driven pathology. These molecular signatures may provide insights into novel therapeutic avenues and peripheral biomarkers for MS monitoring.