Single-Cell Repertoire Analysis Demonstrates that Clonal Expansion Is a Prominent Feature of the B Cell Response in Multiple Sclerosis Cerebrospinal Fluid

Abstract

Single-cell RT-PCR was used to sample CD19+ B cell repertoires in cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS) or viral meningitis. Analysis of amplified Ab H and L chain products served to identify the rearranged germline segment and J segment, and to determine the degree of homology for the H and L chain sequence of individual B cells. The B cell repertoire of viral meningitis CSF was predominately polyclonal, whereas B cell clonal expansion was a prominent feature of the IgG repertoire in three of four MS patients. Two dominant clonal populations in one MS CSF accounted for ∼70% of the IgG H chain V regions sequenced, while the corresponding IgM repertoires were more heterogeneous. One clonal B cell population revealed multiple L chain rearrangements, raising the possibility of a role for receptor editing in shaping the B cell response in some MS patients. The most immediate implications of identifying rearranged Ig sequences in MS B cells is the potential to accurately recreate recombinant Abs from these overrepresented H and L chains that can be used to discover the relevant Ag(s) in MS.

CNS Aquaporin‐4‐specific B cells connect with multiple B‐cell compartments in neuromyelitis optica spectrum disorder

“…CSF CD19 + CD138 + plasmablast heavy‐ (VH) and light‐chain (VL) variable region sequences were recovered by RT‐PCR and DNA sequencing as described previously 1, 7. Recombinant antibodies were produced in HEK293 cells (Invitrogen, Carlsbad, CA, R620‐07)7 and reactivity tested via immunofluorescence for binding to a permanent cell line expressing M23‐AQP4 8…”

“…Recombinant antibodies were produced in HEK293 cells (Invitrogen, Carlsbad, CA, R620‐07)7 and reactivity tested via immunofluorescence for binding to a permanent cell line expressing M23‐AQP4 8…”

High throughput analysis of TCR-β rearrangement and gene expression in single T cells

“…The efficacy of the approach at the single cell level is comparable to other methods which focus on the amplification of less complex rearranged BCR genes. 28,29 T cells isolated from diseased tissues cannot only be characterized for their TCR-b rearrangement but also for the expression of additional genes.…”

Elucidating the Molecular Basis of Multiple Sclerosis and Understanding the Disease Pathophysiology

“…More so, Owens et al found a significant over-representation of the H and L chains of Ig receptors on B-cells of MS patients. Effectively, this may increase the possibility of Ig recombination and antigen identification [100].…”