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Antibodies to the inositol 1,4,5-trisphosphate receptor type 1 (ITPR1) in cerebellar ataxia.

Jarius S, Scharf M, Begemann N, Stöcker W, Probst C, Serysheva II, Nagel S, Graus F, Psimaras D, Wildemann B, Komorowski L - J Neuroinflammation (2014)

Bottom Line: The antibody bound to PC somata, dendrites, and axons, resulting in a binding pattern similar to that reported for anti-Ca/anti-ARHGAP26, but did not react with recombinant ARHGAP26.By contrast, anti-ARHGAP26-positive sera did not bind to ITPR1.Our findings suggest a role of autoimmunity against ITPR1 in the pathogenesis of autoimmune cerebellitis and extend the panel of diagnostic markers for this disease.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neuroimmunology, Department of Neurology, University of Heidelberg, Im Neuenheimer Feld 400, 69120, Heidelberg, Germany. sven.jarius@med.uni-heidelberg.de.

ABSTRACT
We report on a serum autoantibody associated with cerebellar ataxia. Immunohistochemical studies of sera from four patients referred for autoantibody testing revealed binding of high-titer (up to 1:5,000) IgG antibodies, mainly IgG1, to the molecular layer, Purkinje cell layer, and white matter on mouse, rat, porcine, and monkey cerebellum sections. The antibody bound to PC somata, dendrites, and axons, resulting in a binding pattern similar to that reported for anti-Ca/anti-ARHGAP26, but did not react with recombinant ARHGAP26. Extensive control studies were performed to rule out a broad panel of previously described paraneoplastic and non-paraneoplastic anti-neural autoantibodies. The characteristic binding pattern as well as double staining experiments suggested inositol 1,4,5-trisphosphate receptor type 1 (ITPR1) as the target antigen. Verification of the antigen included specific neutralization of the tissue reaction following preadsorption with ITPR1 (but not ARHGAP26) and a dot-blot assay with purified ITPR1 protein. By contrast, anti-ARHGAP26-positive sera did not bind to ITPR1. In a parallel approach, a combination of histoimmunoprecipitation and mass spectrometry also identified ITPR1 as the target antigen. Finally, a recombinant cell-based immunofluorescence assay using HEK293 cells expressing ITPR1 and ARHGAP26, respectively, confirmed the identification of ITPR1. Mutations of ITPR1 have previously been implicated in spinocerebellar ataxia with and without cognitive decline. Our findings suggest a role of autoimmunity against ITPR1 in the pathogenesis of autoimmune cerebellitis and extend the panel of diagnostic markers for this disease.

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Confirmation of ITPR1 as the target antigen by a recombinant cell-based indirect immunofluorescence assay employing HEK293 cells transfected with full-length human ITPR1 and mock-transfected HEK293 cells as control substrates.
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Fig9: Confirmation of ITPR1 as the target antigen by a recombinant cell-based indirect immunofluorescence assay employing HEK293 cells transfected with full-length human ITPR1 and mock-transfected HEK293 cells as control substrates.

Mentions: As further confirmation of correct antigen identification, the PC antibody-positive sera and controls were then analyzed by an RC-IFA using HEK293 expressing murine ITPR1 and mock-transfected HEK293 (Figure 9, upper panel). All reference sera but none of the controls reacted with the ITPR1-expressing cells (Figure 9, middle panel). In contrast, mock transfection did not result in any antibody binding (Figure 9, lower panel). The congruence of the patients’ autoantibody target and ITPR1 was further demonstrated by the dose-dependent competitive abolition of antibody binding to PCs by HEK293 lysates containing ITPR1 (Figure 7E). Antibody binding was unaffected when comparable lysates from mock-transfected HEK293 were used (Figure 7F).Figure 9


Antibodies to the inositol 1,4,5-trisphosphate receptor type 1 (ITPR1) in cerebellar ataxia.

Jarius S, Scharf M, Begemann N, Stöcker W, Probst C, Serysheva II, Nagel S, Graus F, Psimaras D, Wildemann B, Komorowski L - J Neuroinflammation (2014)

Confirmation of ITPR1 as the target antigen by a recombinant cell-based indirect immunofluorescence assay employing HEK293 cells transfected with full-length human ITPR1 and mock-transfected HEK293 cells as control substrates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4300617&req=5

Fig9: Confirmation of ITPR1 as the target antigen by a recombinant cell-based indirect immunofluorescence assay employing HEK293 cells transfected with full-length human ITPR1 and mock-transfected HEK293 cells as control substrates.
Mentions: As further confirmation of correct antigen identification, the PC antibody-positive sera and controls were then analyzed by an RC-IFA using HEK293 expressing murine ITPR1 and mock-transfected HEK293 (Figure 9, upper panel). All reference sera but none of the controls reacted with the ITPR1-expressing cells (Figure 9, middle panel). In contrast, mock transfection did not result in any antibody binding (Figure 9, lower panel). The congruence of the patients’ autoantibody target and ITPR1 was further demonstrated by the dose-dependent competitive abolition of antibody binding to PCs by HEK293 lysates containing ITPR1 (Figure 7E). Antibody binding was unaffected when comparable lysates from mock-transfected HEK293 were used (Figure 7F).Figure 9

Bottom Line: The antibody bound to PC somata, dendrites, and axons, resulting in a binding pattern similar to that reported for anti-Ca/anti-ARHGAP26, but did not react with recombinant ARHGAP26.By contrast, anti-ARHGAP26-positive sera did not bind to ITPR1.Our findings suggest a role of autoimmunity against ITPR1 in the pathogenesis of autoimmune cerebellitis and extend the panel of diagnostic markers for this disease.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neuroimmunology, Department of Neurology, University of Heidelberg, Im Neuenheimer Feld 400, 69120, Heidelberg, Germany. sven.jarius@med.uni-heidelberg.de.

ABSTRACT
We report on a serum autoantibody associated with cerebellar ataxia. Immunohistochemical studies of sera from four patients referred for autoantibody testing revealed binding of high-titer (up to 1:5,000) IgG antibodies, mainly IgG1, to the molecular layer, Purkinje cell layer, and white matter on mouse, rat, porcine, and monkey cerebellum sections. The antibody bound to PC somata, dendrites, and axons, resulting in a binding pattern similar to that reported for anti-Ca/anti-ARHGAP26, but did not react with recombinant ARHGAP26. Extensive control studies were performed to rule out a broad panel of previously described paraneoplastic and non-paraneoplastic anti-neural autoantibodies. The characteristic binding pattern as well as double staining experiments suggested inositol 1,4,5-trisphosphate receptor type 1 (ITPR1) as the target antigen. Verification of the antigen included specific neutralization of the tissue reaction following preadsorption with ITPR1 (but not ARHGAP26) and a dot-blot assay with purified ITPR1 protein. By contrast, anti-ARHGAP26-positive sera did not bind to ITPR1. In a parallel approach, a combination of histoimmunoprecipitation and mass spectrometry also identified ITPR1 as the target antigen. Finally, a recombinant cell-based immunofluorescence assay using HEK293 cells expressing ITPR1 and ARHGAP26, respectively, confirmed the identification of ITPR1. Mutations of ITPR1 have previously been implicated in spinocerebellar ataxia with and without cognitive decline. Our findings suggest a role of autoimmunity against ITPR1 in the pathogenesis of autoimmune cerebellitis and extend the panel of diagnostic markers for this disease.

Show MeSH
Related in: MedlinePlus