Bettina Kuenz

Antibody to aquaporin-4 in the long-term course of neuromyelitis optica.

Neuromyelitis optica (NMO) is a severe inflammatory CNS disorder of putative autoimmune aetiology, which predominantly affects the spinal cord and optic nerves. Recently, a highly specific serum reactivity to CNS microvessels, subpia and Virchow–Robin spaces was described in patients with NMO [called NMO–IgG (NMO–immunoglobulin G)]. Subsequently, aquaporin-4 (AQP4), the most abundant water channel in the CNS, was identified as its target antigen. Strong support for a pathogenic role of the antibody would come from studies demonstrating a correlation between AQP4-Ab (AQP4-antibody) titres and the clinical course of disease. In this study, we determined AQP4-Ab serum levels in 96 samples from eight NMO–IgG positive patients (median follow-up 62 months) in a newly developed fluorescence-based immunoprecipitation assay employing recombinant human AQP4. We found that AQP4-Ab serum levels correlate with clinical disease activity, with relapses being preceded by an up to 3-fold increase in AQP4-Ab titres, which was not paralleled by a rise in other serum autoantibodies in one patient. Moreover, AQP4-Ab titres were found to correlate with CD19 cell counts during therapy with rituximab. Treatment with immunosuppressants such as rituximab, azathioprine and cyclophosphamide resulted in a marked reduction in antibody levels and relapse rates. Our results demonstrate a strong relationship between AQP4-Abs and clinical state, and support the hypothesis that these antibodies are involved in the pathogenesis of NMO.

Complement activating antibodies to myelin oligodendrocyte glycoprotein in neuromyelitis optica and related disorders

BackgroundSerum autoantibodies against the water channel aquaporin-4 (AQP4) are important diagnostic biomarkers and pathogenic factors for neuromyelitis optica (NMO). However, AQP4-IgG are absent in 5-40% of all NMO patients and the target of the autoimmune response in these patients is unknown. Since recent studies indicate that autoimmune responses to myelin oligodendrocyte glycoprotein (MOG) can induce an NMO-like disease in experimental animal models, we speculate that MOG might be an autoantigen in AQP4-IgG seronegative NMO. Although high-titer autoantibodies to human native MOG were mainly detected in a subgroup of pediatric acute disseminated encephalomyelitis (ADEM) and multiple sclerosis (MS) patients, their role in NMO and High-risk NMO (HR-NMO; recurrent optic neuritis-rON or longitudinally extensive transverse myelitis-LETM) remains unresolved.ResultsWe analyzed patients with definite NMO (n = 45), HR-NMO (n = 53), ADEM (n = 33), clinically isolated syndromes presenting with myelitis or optic neuritis (CIS, n = 32), MS (n = 71) and controls (n = 101; 24 other neurological diseases-OND, 27 systemic lupus erythematosus-SLE and 50 healthy subjects) for serum IgG to MOG and AQP4. Furthermore, we investigated whether these antibodies can mediate complement dependent cytotoxicity (CDC). AQP4-IgG was found in patients with NMO (n = 43, 96%), HR-NMO (n = 32, 60%) and in one CIS patient (3%), but was absent in ADEM, MS and controls. High-titer MOG-IgG was found in patients with ADEM (n = 14, 42%), NMO (n = 3, 7%), HR-NMO (n = 7, 13%, 5 rON and 2 LETM), CIS (n = 2, 6%), MS (n = 2, 3%) and controls (n = 3, 3%, two SLE and one OND). Two of the three MOG-IgG positive NMO patients and all seven MOG-IgG positive HR-NMO patients were negative for AQP4-IgG. Thus, MOG-IgG were found in both AQP4-IgG seronegative NMO patients and seven of 21 (33%) AQP4-IgG negative HR-NMO patients. Antibodies to MOG and AQP4 were predominantly of the IgG1 subtype, and were able to mediate CDC at high-titer levels.ConclusionsWe could show for the first time that a subset of AQP4-IgG seronegative patients with NMO and HR-NMO exhibit a MOG-IgG mediated immune response, whereas MOG is not a target antigen in cases with an AQP4-directed humoral immune response.

Frequency and prognostic impact of antibodies to aquaporin-4 in patients with optic neuritis.

BACKGROUND Antibodies to aquaporin-4 (AQP4-Ab) are found in 60-80% of patients with neuromyelitis optica (NMO), a severely disabling inflammatory CNS disorder of putative autoimmune aetiology, which predominantly affects the optic nerves and spinal cord. OBJECTIVE To assess the frequency of AQP4-Ab in patients with optic neuritis (ON), and to investigate the prognostic implications of AQP4-Ab seropositivity in such patients. PATIENTS AND METHODS AQP4-Ab serum levels were determined in 224 individuals from Austria, Denmark, France, Germany, Italy, and Turkey using a newly developed fluorescence immunoprecipitation assay employing recombinant human AQP4. RESULTS AQP4-Ab were detectable in 8/139 (5.8%) patients with acute monosymptomatic optic neuritis (AMON) and in 10/17 (58.8%) patients with established NMO and a last relapse of acute ON (NMO/ON), but not in 32 patients with multiple sclerosis or in 36 healthy controls. At last examination, 4/8 (50%) seropositive AMON patients had met the criteria for NMO but 0/128 seronegative AMON patients. Disease severity differed significantly between seropositive and seronegative AMON. Complete bilateral or unilateral blindness occurred in six AQP4-Ab positive patients, but only in one AQP4-Ab negative patient. AQP4-Ab levels did not vary between seropositive AMON and NMO/ON and did not correlate with disease severity. Female gender, a relapsing course, and concomitant autoimmunity were associated with AQP4-Ab seropositive status and risk of developing NMO. CONCLUSION AQP4-Ab is relatively rare among patients with AMON, but if present it predicts a high rate of conversion to NMO within one year.

Patterns of antibody binding to aquaporin-4 isoforms in neuromyelitis optica.

Background Neuromyelitis optica (NMO), a severe demyelinating disease, represents itself with optic neuritis and longitudinally extensive transverse myelitis. Serum NMO-IgG autoantibodies (Abs), a specific finding in NMO patients, target the water channel protein aquaporin-4 (AQP4), which is expressed as a long (M-1) or a short (M-23) isoform. Methodology/Principal Findings The aim of this study was to analyze serum samples from patients with NMO and controls for the presence and epitope specificity of IgG and IgM anti-AQP4 Abs using an immunofluorescence assay with HEK293 cells expressing M-1 or M-23 human AQP4. We included 56 patients with definite NMO (n = 30) and high risk NMO (n = 26), 101 patients with multiple sclerosis, 27 patients with clinically isolated syndromes (CIS), 30 patients with systemic lupus erythematosus (SLE) or Sjögrens syndrome, 29 patients with other neurological diseases and 47 healthy controls. Serum anti-AQP4 M-23 IgG Abs were specifically detected in 29 NMO patients, 17 patients with high risk NMO and two patients with myelitis due to demyelination (CIS) and SLE. In contrast, IgM anti-AQP4 Abs were not only found in some NMO and high risk patients, but also in controls. The sensitivity of the M-23 AQP4 IgG assay was 97% for NMO and 65% for high risk NMO, with a specificity of 100% compared to the controls. Sensitivity with M-1 AQP4 transfected cells was lower for NMO (70%) and high risk NMO (39%). The conformational epitopes of M-23 AQP4 are the primary targets of NMO-IgG Abs, whereas M-1 AQP4 Abs are developed with increasing disease duration and number of relapses. Conclusions Our results confirm M-23 AQP4-IgG Abs as reliable biomarkers in patients with NMO and high risk syndromes. M-1 and M-23 AQP4-IgG Abs are significantly associated with a higher number of relapses and longer disease duration.

Cerebrospinal fluid B cells correlate with early brain inflammation in multiple sclerosis.

Background There is accumulating evidence from immunological, pathological and therapeutic studies that B cells are key components in the pathophysiology of multiple sclerosis (MS). Methodology/Principal Findings In this prospective study we have for the first time investigated the differences in the inflammatory response between relapsing and progressive MS by comparing cerebrospinal fluid (CSF) cell profiles from patients at the onset of the disease (clinically isolated syndrome, CIS), relapsing-remitting (RR) and chronic progressive (CP) MS by flow cytometry. As controls we have used patients with other neurological diseases. We have found a statistically significant accumulation of CSF mature B cells (CD19+CD138−) and plasma blasts (CD19+CD138+) in CIS and RRMS. Both B cell populations were, however, not significantly increased in CPMS. Further, this accumulation of B cells correlated with acute brain inflammation measured by magnetic resonance imaging and with inflammatory CSF parameters such as the number of CSF leukocytes, intrathecal immunoglobulin M and G synthesis and intrathecal production of matrix metalloproteinase (MMP)-9 and the B cell chemokine CxCL-13. Conclusions Our data support an important role of CSF B cells in acute brain inflammation in CIS and RRMS.

Plasma levels of soluble adhesion molecules sPECAM-1, sP-selectin and sE-selectin are associated with relapsing-remitting disease course of multiple sclerosis

Adhesion molecule mediated leukocyte migration into the central nervous system is considered to be a critical step in the pathogenesis of multiple sclerosis (MS). We measured plasma levels of the soluble adhesion molecules sPECAM-1, sP-selectin and sE-selectin in 166 MS patients and in 36 healthy blood donors with ELISA. sPECAM-1, sP-selectin and sE-selectin plasma concentrations showed a significant increase in the relapsing-remitting disease course of MS and were elevated during relapse. These findings indicate that sPECAM-1, sP-selectin and sE-selectin might be implemented as paraclinical markers of disease activity in MS with restriction to the clinical course of the disease.

Olfactory threshold is impaired in early, active multiple sclerosis

Background: Olfactory dysfunction has been reported in multiple sclerosis (MS). However, to date no data are available on different qualities of olfactory function, namely odour identification, odour discrimination and odour perception threshold. Objective: To assess different qualities of olfactory function in patients with MS and correlate these with demographic data, clinical data, depression, quality of life and cognitive functions. Methods: In this cross-sectional study, 50 patients with MS or clinically isolated syndrome and 30 healthy controls were included. Olfactory function was measured using the Sniffin’ Sticks test. Results: The scores for odour identification (p = 0.001), odour perception threshold (p = 0.037) and the combined score of odour identification, discrimination and perception threshold (TDI, p = 0.002) were significantly lower in MS. Hyposmia for identification (p = 0.0017), threshold (p = 0.017) and TDI score (p = 0.0014) was more frequent in MS. Olfactory threshold was impaired in patients who were clinically active in the previous year (p = 0.026) and in patients with a disease duration less than 2 years (p = 0.0093). Identification score was negatively correlated with disease duration (p = 0.0017). Olfactory function was not associated with disability, depression or quality of life. Conclusions: We report evidence for qualitatively distinct hyposmia in MS, with increased smell threshold in the early inflammatory phases of the disease and impaired identification with a more widespread chronic disease.

Differing immunogenic potentials of interferon beta preparations in multiple sclerosis patients

Interferon beta (IFNβ) is a first-line therapy for multiple sclerosis (MS). However, some patients experience a decline in efficacy with continued therapy due to the development of anti-IFNβ neutralizing antibodies (NAb). We investigated the frequency of NAb cross-sectionally in 846 MS patients who were receiving IFNβ-1b, IFNβ-1a im, or IFNβ-1a sc. The frequency of NAb in patients receiving IFNβ-1a im was lower (5%) than in patients treated with any other form of IFNβ (22-35%) (P < 0.0001). Binding antibodies (BAb) were measured in 808 patients. The frequency differed significantly between treatment groups, ranging from 45% (IFNβ-1a im) to 88% (IFNβ-1b). The proportion of NAb-positive patients within the BAb-positive group differed significantly among treatment groups, ranging between 12% (IFNβ-1a im) and 51% (IFNβ-1a sc). The median NAb titer from all IFNβ-1a-treated patients was higher than from IFNβ-1b-treated patients (446 versus 171 NU/mL, P = 0.04). Among NAb-positive patients, the frequency of NAb titers > 100 NU/mL was 71% for IFNβ-1a compared with 58% for IFNβ-1b (P = 0.04). Except for conflicting data regarding IFNβ-1a sc, the results are generally consistent with the literature and together with the differing proportion of NAb-positive patients within the BAb-positive group, provide further insight into the immunogeni-city of the IFNβ preparations.

Persistency of neutralizing antibodies depends on titer and interferon-beta preparation

Background: Neutralizing antibodies (NAbs) affect the efficacy of interferon-beta (IFNβ) treatment in multiple sclerosis (MS) patients, particularly if NAbs persist. Persistency depends on NAb titers, which differ between IFNβ preparations. Objective: This study evaluated IFNβ preparation-specific NAb cut-off titers during early treatment for prediction of NAb persistency. Methods: Patients who had at least one NAb test between 12 and 30 months (baseline) as well as after more than 48 months (follow-up) on IFNβ treatment were included in this longitudinal study. Results: At baseline 1064 patients had a NAb test. Of those, 203 had a follow-up test. In the follow-up group 23.2% of patients were NAb positive during baseline. NAb frequency significantly decreased by 40.7% in the IFNβ-1a and by 60% in the IFNβ-1b group at follow-up after a mean time of 75.4 months on treatment, and median NAb titers decreased significantly in both groups. During baseline, NAb titers of >258 neutralizing units (NU) had a sensitivity of 81.3% and a specificity of 90.9% in the IFNβ-1a group, whereas titers of >460 NU had a sensitivity of 100% and a specificity of 91.7% in the IFNβ-1b group to predict persistency at follow-up. When these cut-off titers are applied, 10.2% of all treated patients developed persistent NAbs. Conclusion: IFNβ preparation-specific NAb cut-off titers for prediction of NAb persistency, which may be useful in individual treatment decision making, are provided.

Neuromyelitis Optica in Austria in 2011: To Bridge the Gap between Neuroepidemiological Research and Practice in a Study Population of 8.4 Million People

Background In 2008 the Austrian Task Force for Neuromyelitis Optica (NMO) started a nation-wide network for information exchange and multi-centre collaboration. Their aim was to detect all patients with NMO or NMO spectrum disorders (NMO-SD) in Austria and to analyse their disease courses and response to treatment. Methods (1) As of March 2008, 1957 serum samples (of 1557 patients) have been tested with an established cell based immunofluorescence aquaporin-4 antibody (AQP4-ab) assay with a high sensitivity and specificity (both >95%). All tests were performed in a single reference laboratory (Clinical Dept. of Neurology of the Innsbruck Medical University). (2) A nation-wide survey with several calls for participation (via email newsletters, articles in the official journal of the Austrian Society of Neurology, and workshops) was initiated in 2008. All collected data will be presented in a way that allows that every individual patient can be traced back in order to ensure transparency and to avoid any data distortion in future meta-analyses. The careful and detailed presentation allows the visualization and comparison of the different disease courses in real time span. Failure and response to treatment are made visible at one glance. Database closure was 31 December 2011. All co-operators were offered co-authorship. Results All 71 NMO- or NMO-SD patients with AQP4-ab positivity (age range 12.3 to 79.6 years) were analysed in detail. Sex ratio (m:f = 1:7) and the proportion of patients without oligoclonal bands in cerebrospinal fluid (86.6%) were in line with previously published results. All identified patients were Caucasians. Conclusions A nationwide collaboration amongst Austrian neurologists with good network communications made it possible to establish a database of 71 AQP4-ab positive patients with NMO/NMO-SD. This database is presented in detail and provides the basis for further studies and international cooperation in order to investigate this rare disease.