We have developed assays to detect antibodies to human NF155 and NF186, and show that such autoantibodies are found in 4% of patients with CIDP and AIDP. Using an EAN model, we have demonstrated that antibody targeting of NF in vivo enhances and prolongs neuritis. Our findings are in harmony with a recent report showing nodal reactivity in a proportion of neuropathy patients, although in that article rodent but not human NF was used as an antigen.10
When comparing our 2 assays, ELISA is most suitable for identifying subjects with anti-NF antibodies among patients with inflammatory neuropathy and distinguishing them from controls. Antibodies that are reactive to NF by ELISA but not by flow cytometry can bind NF in a physiologic setting and may be pathogenic, as exemplified by NF155-specific rabbit antibodies (figure e-1C) that bound to paranodes on tissue sections and caused complement-dependent demyelination in an in vitro myelination culture model (Chris Linington, unpublished data, 2012).
Unexpectedly, almost all the serum reactivities observed were directed against only one isoform. This is surprising because NF155 and NF186 are splice variants such that the 8 extracellular domains from the N
-terminus are virtually identical, differing only in the 2 domains closest to the transmembrane region.22
We expressed truncated variants of NF155 and NF186, and identified the Fn3-Fn4 domains as the target for NF155-specific reactivity and Mucin-Fn5 domains for NF186 specificity.
Would the antibodies to NF, which we detect in a minority of patients with CIDP and patients with AIDP, be expected to contribute to the pathology? We investigated the principal pathogenic potential of NF-reactive antibodies with an EAN model, and this yielded 2 main findings. First, antibodies to NF can enhance and prolong an ongoing neuritis. Second, antibodies to NF alone are not pathogenic. Previous studies of this P2 peptide–induced EAN model have demonstrated nerve infiltration by autoreactive T cells and macrophages, yet autoantibodies were not implicated.9
We added anti-pan-NF mAbs into this disease setting and found that NF-targeting exacerbated disease. A similar observation was made in studies using a T-cell transfer EAN model in which addition of antimyelin antibodies greatly exacerbated disease.23
For human antibodies to be pathogenic, the target should be accessible. Inflammatory neuropathies are characterized by pronounced immune cell infiltration24,25
that can open blood–nerve barrier and provide access to autoantibodies.
Would the isoform-specific recognition of NF by human autoantibodies be compatible with pathogenicity? NF186 is displayed on the nodes of Ranvier, and it has been shown that antibody targeting of NF186 in the presence of complement disrupts nerve conduction.12
The anti-NF186 antibodies that we found in 2 patients with AIDP were of complement-activating isotypes IgG1 and IgG3. Furthermore, antibody binding to NF186 might disrupt its ability to perform its normal functions; for example, binding to gliomedin26
or other extracellular ligands.
Antibodies to NF155 found in some patients with AIDP and patients with CIDP could be pathogenic only if their target at the paranodes becomes accessible. Since a disruption of paranodal architecture is a feature of different nerve pathologies such as ischemia and inflammation,27,28
these antibodies could contribute to the pathology. Anti-NF155 antibodies were reported to inhibit myelination by blocking the formation of the Caspr/contactin/NF155 complex, the core structure at paranodal loops for adhesion between axon and glial cell.29
Such a disruption of the paranodal junctions can result in severe reduction of conduction velocity even in the absence of obvious demyelination.30
Thus, it is tempting to speculate that such antibodies interfere with remyelination in patients with AIDP and CIDP.
We made the unexpected observation that in 2 patients with CIDP with remarkably high anti-NF reactivity the anti-NF155 autoantibodies were largely IgG4 with an additional contribution of IgG1, IgG2, IgG3, IgM, and IgA. Anti-NF155 IgG4 in these patients with CIDP may have an antigen-blocking function to impair myelination and thereby nerve conduction, as IgG4 is generally not complement activating nor does it bind Fc receptors on effector cells.31
This blocking effect of IgG4 antibodies may be pathogenic. For example, in endemic pemphigus foliaceus, a blistering skin disease, IgG4 antibodies against desmoglein-1 cause a direct disruption of the epithelial layer, leading to blister formation.32
Apart from IgG4, low levels of complement-activating and immune cell–activating isotypes were also observed in these 2 patients with CIDP. Two other patients with CIDP who benefited from PE had antibodies to NF155. The pathogenic activity of these patient-derived anti-NF antibodies may be shown in further studies.