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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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Histoimmunoprecipitation with a reference patient serum revealed a band at around 300 kDa (arrow).
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Fig8: Histoimmunoprecipitation with a reference patient serum revealed a band at around 300 kDa (arrow).

Mentions: Histoimmunoprecipitates from either rat or porcine cerebellum contained high amounts of IgG when one of the reference sera was used, whereas they were generally low after incubation of sera from healthy controls. Next to the immunoglobulins, the immunoprecipitated PC antibody-positive reference serum showed a protein band corresponding to a molecular mass of approximately 300 kDa in SDS-PAGE stained with colloidal Coomassie (Figure 8). The band was absent in the control samples. The proteins precipitating from rat and porcine cerebellum were identified as ITPR1 from the corresponding organisms by mass-spectrometric analysis. Western blot analysis with the polyclonal rabbit anti-ITPR1 antibody showed a strong reaction at 300 kDa of the immunoprecipitate obtained with the patient serum, while there were no reactions with fractions obtained with control sera. When used for double staining in IFA, the polyclonal anti-ITPR1 antibody produced an overlay with the reference serum used in the MALDI-TOF experiments.Figure 8


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)

Histoimmunoprecipitation with a reference patient serum revealed a band at around 300 kDa (arrow).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig8: Histoimmunoprecipitation with a reference patient serum revealed a band at around 300 kDa (arrow).
Mentions: Histoimmunoprecipitates from either rat or porcine cerebellum contained high amounts of IgG when one of the reference sera was used, whereas they were generally low after incubation of sera from healthy controls. Next to the immunoglobulins, the immunoprecipitated PC antibody-positive reference serum showed a protein band corresponding to a molecular mass of approximately 300 kDa in SDS-PAGE stained with colloidal Coomassie (Figure 8). The band was absent in the control samples. The proteins precipitating from rat and porcine cerebellum were identified as ITPR1 from the corresponding organisms by mass-spectrometric analysis. Western blot analysis with the polyclonal rabbit anti-ITPR1 antibody showed a strong reaction at 300 kDa of the immunoprecipitate obtained with the patient serum, while there were no reactions with fractions obtained with control sera. When used for double staining in IFA, the polyclonal anti-ITPR1 antibody produced an overlay with the reference serum used in the MALDI-TOF experiments.Figure 8

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