Patient-derived lymphocytes drive smoldering lesion pathology in a chimeric multiple sclerosis mouse model

Abstract Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system characterized by demyelination, axonal injury, and neurodegeneration. Smoldering lesions— defined by an inactive core, poor remyelination, and a rim of chronically activated microglia— are hallmarks of a progressive disease course and correlate with irreversible disability. The mechanisms driving their formation remain poorly understood. Using a chimeric mouse model, we investigated the long-term impact of healthy donors (HD) and MS patient-derived lymphocytes (LY) graft on the evolution of spinal cord demyelinated lesion. Three months post-grafting, only MS-derived LY persisted as perivascular cuffs interacting with vascular and murine immune cells, mirroring MS smoldering lesion pathology. MS LY-grafted mice exhibited impaired functional recovery and slower somatosensory evoked potential (SSEP) conduction compared to controls. Electron microscopy confirmed glial scar formation, persistent demyelination and a lesion architecture with an inflammatory inactive center but active rim. Single-nucleus RNA sequencing revealed an imbalance in CNS cell populations, with MS LY-grafted mice showing reduced neuronal abundance, enriched activated microglia, and immature oligodendroglial profiles, correlating with slower SSEP conduction speeds. Immune cell subclustering identified an enrichment of disease-associated microglia and interferon-responsive microglia in MS LY-grafted mice, marked by elevated pro-inflammatory markers and active myelin phagocytosis. Oligodendroglial cells displayed downregulated myelination and stress-response genes, with disrupted myelin ultrastructure confirmed by electron microscopy. Multiblock analysis revealed donor-specific variability, with some MS LY inducing outcomes akin to HD, while others drove severe pathology. Our model enables mechanistic dissection of the transition from focal to diffuse CNS pathology in MS and captures patient-specific pathophysiological signatures, providing a platform for personalized mechanistic studies and targeted therapeutic strategies. Competing Interest Statement The authors have declared no competing interest. Abbreviation - CNS - Central Nervous System - COP - Committed oligodendrocyte precursor cell - DAM - Disease associated microglia - DEG - Differential gene expression - EAE - Experimental Autoimmune Encephalomyelitis - EM - Electron microscopy - HD - Healthy donor - IHC - Immunohistochemistry - IRM - Interferon responsive microglia - LPC - Lysophosphatidylcholine - LY - Lymphocytes - MCOA - Multiple Co-inertia analysis - MFOL - Myelin forming oligodendrocyte - MS - Multiple Sclerosis - NG - Non-grafted - OL - Oligodendrocytes - OPC - Oligodendrocytes Precursors Cell - PBMC - Peripheral Blood Mononuclear cell - PIRA - Progression Independent of Relapse Activity - PPMS - Primary Progressive Multiple Sclerosis - RAW - Relapse Associated Worsening - RGCCA - Regularized generalized canonical correlation analysis - RRMS - Relapsing Remitting Multiple Sclerosis - SnRNAseq - Single nuclei RNA sequencing - SPMS - Secondary Progressive Multiple Sclerosis - SSEP - Somatosensory evoked potential.