Intrathecal Pathogenic Anti-Aquaporin-4 Antibodies in Early Neuromyelitis Optica (1)
Neuromyelitis optica (NMO) is a severe demyelinating disorder that primarily affects the optic nerve and spinal cord resulting in vision loss and paralysis. Serum autoantibodies (NMO-IgGs) directed against the aquaporin-4 (AQP4) water channel has been shown to be a disease-specific marker of NMO pathology.
The VH family germline distribution of the CSF CD138+ plasma cell repertoire was significantly skewed from the expected germline distribution.17. VH2 family germline sequences dominated the plasma cell VH repertoire and accounted for 37% of the total sequences. Sequence analysis of plasma cell clone 13 revealed a common set of somatic mutations punctuated by a large number of sequence-specific replacement mutations.
Patient serum and CSF, but not control serum, bound to the surface of glioblastoma cells (LN18AQP4) transfected with human AQP4, human fetal astrocytes14 (HFAs), and mouse cerebellar sections3. Six of 11 NMO CSF rAbs bound with high affinity to LN18AQP4 cells and HFAs. The binding of CSF rAbs to HFAs and AQP4-transfected LN18AQP4 cells was quite similar.
While 5 of 6 AQP4- specific rAbs produced the characteristic staining of CNS microvessels, pia, and subpia on mouse cerebellar sections, rAb-43 did not stain mouse cerebellar sections, indicating that this plasma cell rAb recognized a species-specific epitope on the human AQP4 protein. A selective reduction in the viability of LN18AQP4 glial cells exposed to serum complement and NMO patient serum IgG or CSF IgG but not in glial cells transfected with an empty vector (LN18CTR). NMO patient serum IgG and CSF IgG directed selective ADCC of LN18AQP4 but not LN18CTR target cells.
In the presence of serum complement, AQP4-specific CSF rAbs resulted in a marked and specific reduction in cell viability in the LN18AQP4 cell line but not in LN18CTR cells. Similarly, in an in vitro ADCC assay, AQP4-specific CSF rAbs specifically reduced the viability of LN18AQP4 but not LN18CTR target cells after prolonged incubation with human NK cells. dose-dependent. Serial dilution of the AQP4-specific rAb-10 demonstrated a clear lytic dose response from 5 μg/ml (5.9% survival) to 0.05 μg/ml (74% survival). There was a significant increase in the percentage of NK cells expressing surface CD107a after contact with HFAs preincubated with patient serum or CSF.
Four NMO CSF rAbs (rAb-10, rAb-168, rAb-51, rAb-43), a monoclonal antibody against myelin oligodendrocyte glycoprotein (mAb 8-18C5), and rAb-2B4, a control rAb against measles virus nucleocapsid, were transfused by retrobulbar venous plexus injection into rats previously immunized with guinea pig MBP72–85. Intravenous transfer of the AQP4-specific rAb-10, however, resulted in novel pathologic changes prototypic for NMO such as perivascular astrocyte depletion and complement deposition. No loss of GFAP immunoreactivity was observed in rats transferred with rAb-2B4, rAb-168, rAb-51, rAb-43, mAb 8-18C5 or PBS. However, between 3 and 7% of total spinal cord area were devoid of astrocytes in rats transferred with rAb-10. a human AQP4-specific rAb derived from a clonally expanded intrathecal plasma cell produces NMO-related immunopathology, perivascular astrocyte destruction and complement deposition, after intravenous transfer into EAE animals.
Serum autoantibodies against AQP4 (NMO-IgG) are a specific biomarker that distinguishes patients with NMO from those with MS. Serum AQP4 antibody could enter the CNS by endothelial transcytosis or through regions of increased BBB permeability, but AQP4-specific autoantibodies are only a minor component of the serum IgG fraction and the initial NMO lesions do not demonstrate any regional predilection.