NFM Cross-Reactivity to MOG Does Not Expand a Critical Threshold Level of High-Affinity T Cells Necessary for Onset of Demyelinating Disease

Of interest to the etiology of demyelinating autoimmune disease is the potential to aberrantly activate CD4(+) T cells due to crossrecognition of multiple self-epitopes such as has been suggested for myelin oligodendrocyte glycoprotein epitope 35-55 (MOG(35-55)) and neurofilament medium protein epitope 15-35 (NFM15-35). NFM15-35 is immunogenic in C57BL/6 mice but fails to induce demyelinating disease by polyclonal T cells despite having the same TCR contact residues as MOG(35-55), a known encephalitogenic Ag. Despite reported cross-reactivity with MOG-specific T cells, the polyclonal response to NFM15-35 did not expand threshold numbers of MOG(38-49) tetramer-positive T cells. Furthermore, NFM lacked functional synergy with MOG to promote experimental autoimmune encephalomyelitis because NFM-deficient synonymous with knockout mice developed an identical disease course to wild-type mice after challenge with MOG(35-55). Single-cell analysis of encephalitogenic T cells using the peptide: MHC monomer-based two-dimensional micropipette adhesion frequency assay confirmed that NFM was not a critical Ag driving demyelinating disease because NFM18-30-specific T cells in the CNS were predominantly reactive to MOG(38-49). The absence of NFM contribution to disease allowed mapping of the amino acids required for encephalitogenicity and expansion of high-affinity, MOG-specific T cells that defined the polyclonal response. Alterations of N-terminal residues outside of the NFM15-35 core nonamer promoted expansion of high-affinity, MOG(38-49) tetramer-positive T cells and promoted consistent experimental autoimmune encephalomyelitis induction, unlike mice challenged with NFM15-35. Although NFM15-35 is immunogenic and crossreactive with MOG at the polyclonal level, it fails to expand a threshold level of encephalitogenic, high-affinity MOG-specific T cells.