Serum autoantibodies against α7-nicotinic receptors in subgroups of patients with bipolar disorder or schizophrenia: clinical features and link with peripheral inflammation

Abstract

There is growing evidence that autoantibodies (AAbs) against proteins expressed in the brain are playing an important role in neurological and psychiatric disorders. Here, we explore the presence and the role of peripheral AAbs to the α7-nicotinic acetylcholine receptor (nAChR) in inflammatory subgroups of psychiatric patients with bipolar disorder (BD) or schizophrenia (SCZ) and healthy controls. We have identified a continuum of AAb levels in serum when employing a novel ELISA technique, with a significant elevation in patients compared to controls. Using unsupervised two-step clustering to stratify all the subjects according to their immuno-inflammatory background, we delineate one subgroup consisting solely of psychiatric patients with severe symptoms, high inflammatory profile, and significantly increased levels of anti-nAChR AAbs. In this context, we have used monoclonal mouse anti-human α7-nAChR antibodies (α7-nAChR-mAbs) and shown that TNF-α release was enhanced upon LPS stimulation in macrophages pre-incubated with α7-nAChR-mAbs compared to the use of an isotype control. These findings provide a basis for further study of circulating nicotinic AAbs, and the inflammatory profile observed in patients with major mood and psychotic disorders.

Fig. 1. Anti-α7-nAChR IgGs in blood samples.

a Serological reactivity of human serum on the α7-nAChR extracellular domain measured by ELISA, the values equal to the background noise are indicated by the orange square (PSY psychiatric patients, HC healthy controls, neg ctrl rabbit goat anti-rabbit secondary antibody with HRP, neg ctrl human = goat anti-human secondary antibody HRP). Sample ODs ranged from 0.12 to 1.79. b Immunostaining of 1 µg/ml of monoclonal anti-human α7-nAChR on CHO-K1 cells expressing or not the human α7-nAChR labeled with secondary goat anti-mouse IgG-AlexaFluor 647 (scale bar = 10 µm).

Fig. 2. Principal component analyses (PCA) on serum immune-inflammatory markers.

a Left panel: correlation circle including all the serum cytokines and anti-α7-nAChR AAbs; right panel: spatial distribution of diagnoses on the correlation circle (blue = HC; orange = BD; red = SZ). b Decomposition of explained variance by anti-a7-nAChR AAs (Table 2) principal components, the five first dimensions explained 53.5% of the variance. c Variables contribution for each dimension.

Fig. 3. Hierarchical clustering defines 5 clusters based on immune-inflammatory profile.

a Heatmap representation of inflammatory profiles within all the population (diagnoses are highlighted as follows: blue = HC, dark orange = hospitalized BD, light orange = ambulatory BD, dark red = hospitalized SZ, light red = ambulatory SZ). b Selection of the optimal number of clusters according to inertia. c Dendrogram illustrating cluster slicing and composition.

Fig. 4. Anti-α7-nAChR AAbs and tumor necrosis factor in clusters.

a Boxplot of anti-α7-nAChR AAb OD between clusters in healthy controls (striped bar) and psychiatric patients. b Levels of TNF-α (left panel) and TNF-β (right panel).

Fig. 5. TNF-α release assay upon LPS challenge in macrophages incubated with nAChR drugs or Ab.

a No effect of nAChR agonist PNU-282987 (F = 1173, P = 0,3774)* and b nAChR PAM (F = 0,6635, P = 0,6581)* compared to control. c Anti-inflammatory effect of the nAChR partial agonist NS6740 (F = 84,48, P < 0,0001)* after 6 h of stimulation with LPS E. coli. d Anti-α7-nAChR-mAb increases the LPS-induced TNF-α release in human macrophages compared to isotype control (IC-mAb, 53.80; P < 0,0001)*. *One-way ANOVA followed by Holm–Sidak’s post hoc test for multiple comparisons. Dashed line= release of TNF-α after 6 h of LPS challenge without any pharmacological agents (100%). For each condition, N = 3 independent experiments.