Angiotensin type-1 receptor and ACE2 autoantibodies in Parkinson´s disease

Figures

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  Fig. 1. Autoantibody serum levels and their correlations with inflammatory cytokines and 27-hydroxycholesterol in controls and PD patients. Parkinson´s disease (PD) patients had significantly higher serum levels of AT1-AA (a; Wilcoxon test, W = 4708, P = 0.0019) and ACE2-AA (b; Wilcoxon test, W = 3709; P < 0.0001) than controls. Results of Spearman’s correlation matrix show variables (AT1-AA, ACE2-AA, 27-OHC, cytokines) correlating at varying levels of significance (*P < 0.05; **P < 0.01; ***P < 0.001) in PD patients (c) and controls (d). In a and b, data distribution is shown using a box plot with boxes representing the IQR and the median (black line) and whiskers representing ± 1.5 IQR. IQR Interquartile range. **P < 0.01; ***P < 0.001. 27-OHC, 27-Hydroxycholesterol; AT1-AA: Autoantibodies for AT1 receptors; ACE2-AA: ACE2 Autoantibodies.

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  Fig. 2. Increase in serum and CSF AT1-AA and ACE2-AA levels induced by dopaminergic degeneration in rats. Serum levels of AT1-AA (a) and ACE2-AA (c) 10 days and 4 weeks after 6-OHDA injection (i.e. in the acute period of degeneration and neuroinflammation and when the lesion is stabilized and neuroinflammation has decreased, respectively). A significant increase in AT1-AA (b) and ACE2-AA (d) is observed in CSF 10 days and 4 weeks after 6-OHDA injection. A significant increase in levels of LIGHT cytokine (f) and transglutaminase activity (e) is observed in the nigral region 10 days and 4 weeks after 6-OHDA lesion, which may contribute to the increase in autoantibody production. Data are given as means ± SEM. *P < 0 .05 compared to the control group. Kruskal–Wallis One Way Analysis of Variance on Ranks with Student-Newman-Keuls Method post hoc test (a–c, e, f) and one-way ANOVA followed by the Student–Newman–Keuls Method for multiple comparisons (d). AT1-AA: Autoantibodies for AT1 receptors; ACE2-AA: ACE2 Autoantibodies.

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  Fig. 3. Primary mesencephalic cultures showing that AT1-AA enhance the deleterious effects of 6-OHDA (10 µM). In 6-OHDA-treated cultures, deleterious effects on cell viability are not significant with a dose of 10 ng/µl AT1-AA (i.e. 10 µM 6-OHDA + 10 ng/µl AT1-AA) (a) but are significant at 100 ng/µl (i.e. 10 µM 6-OHDA + 100 ng/µl AT1-AA), as revealed by the MTT assay (b) and TH-positive neuron counting (d), which was inhibited by the AT1 receptor blocker candesartan. AT1-AA alone did not induce significant decrease in cell viability (a, b, d). Interestingly, AT1-AA administration (100 ng/µl) also induced a marked increase in levels of TNF-α in cultures, which is significantly reduced by treatment with candesartan (c). Photomicrographs of dopaminergic (TH-ir) neurons in control cultures (e), cultures treated with AT1-AA plus 6-OHDA (f) and cultures treated with AT1-AA plus 6-OHDA plus candesartan (g). Scale bar 25 µm. Data are given as means ± SEM. *P < 0 .05 compared to the corresponding control group, #P < 0.05 compared to AT1-AA group; &P < 0.05 compared to 6-OHDA group; $P < 0.05 compared to 6-OHDA + AT1-AA group. Kruskal–Wallis One Way Analysis of Variance on Ranks with…

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  Fig. 4. Dopaminergic neurons, astrocytes and microglia bearing fluorescent AT1-AA. Triple fluorescent labelling for (green) the cell marker anti-α-tubulin (a, e, i), the dopaminergic neuron marker Tyrosine hydroxylase (TH; m), the microglial marker CD11b (OX42; q), the astroglial marker GFAP(u), and the nuclear marker Hoechst 33342 (blue), and AT1-AA (red dots) in cell lines of dopaminergic neurons (N27; a–d), microglia (HMC3; e–h), astroglia (C6; i–l), and primary mesencephalic cultures (PMC; m–x) containing dopaminergic neurons (m–p), microglia (q–t) and astrocytes (u–x). Labelling for AT1-AA (red dots) is observed at the cell surface, with practically no colocalization (yellow) with the cytoplasmatic markers. Scale bars: 15 µm.

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  Fig. 5. Proposed model for AT1-AA and ACE2-AA generation and effects in PD. Dopaminergic degeneration/neuroinflammation induces an increase in modified AT1 receptors, possibly mediated by inflammation-related cytokines and transglutaminases (TG2), acting as neoantigens that reach the CSF. From the CSF, via the lymphatic system, neoantigens can drain into cervical lymph nodes. In the cervical nodes, B cells can respond against neural autoantigens, inducing antigen-specific B cells and circulating autoantibodies. Activated B cells and/or autoantibodies may then migrate into the intrathecal compartment through the BBB, as circulating AT1-AA induce BBB disruption acting on endothelial AT1 receptors. In the brain, AT1-AA enhance AT1 receptor activation and the proinflammatory arm (Ang II/AT1) of the nigral RAS. Furthermore, AT1 activation promote shedding of transmembrane ACE2 into a soluble circulating form by activating proteases such as ADAM17, which decreases local Angiotensin 1–7 and the anti-inflammatory RAS arm (Ang 1–7/MasR) function. The increase in soluble ACE2 levels may induce ACE2-AA, which act as ACE2 antagonists further reducing the activity of transmembrane ACE2 and the…