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Department of Immunopathology, SA Pathology, Adelaide, SA, AustraliaDepartment of Clinical Immunology and Allergy, Royal Adelaide Hospital, Adelaide, SA, Australia
Department of Immunopathology, SA Pathology, Adelaide, SA, AustraliaDepartment of Clinical Immunology and Allergy, Royal Adelaide Hospital, Adelaide, SA, Australia
Department of Immunopathology, SA Pathology, Adelaide, SA, AustraliaDepartment of Clinical Immunology and Allergy, Royal Adelaide Hospital, Adelaide, SA, AustraliaDepartment of Immunology, Flinders University, Bedford Park, SA, Australia
Autoantibodies against intracellular antigen, glial fibrillary acidic protein (GFAP), were recently described in 16 patients presenting with meningoencephalomyelitis; a disease entity termed GFAP astrocytopathy.
Clinical, neuroradiological, diagnostic and prognostic profile of autoimmune glial fibrillary acidic protein astrocytopathy: a pooled analysis of 324 cases from published data and a single-center retrospective study.
Currently, no specific clinical or histological diagnostic criteria exist for GFAP astrocytopathy. Therefore, the presence of antibodies directed against GFAP are critical for diagnosis of this rare and emerging clinical entity.
Eight GFAP isomers have been described, with GFAP-α being the most common autoantibody target.
Laboratory methods for detection of GFAP-specific IgG include staining of neuronal tissues by either indirect immunofluorescence (IIF) or immunohistochemical methods while transfected human embryonic kidney 293 (HEK293) cell-based assay (CBA) or Western blot (WB) are used as confirmatory assays. The striking fibrillary staining pattern in the cerebellar tissue is characteristic for these autoantibodies, and when identified should raise suspicion of GFAP antibodies. Neuronal IIF is routinely available in immunopathology laboratories and can be used to assess cerebrospinal fluid (CSF) and serum, providing a useful initial screen whilst antigen-specific confirmatory assays are not currently available in the routine diagnostic setting.
From the immunopathology laboratory perspective, there are still a few issues that need to be addressed. Firstly, the reporting of GFAP antibody pattern is not consistent among immunopathology laboratories in Australia and the neuronal IIF is currently available in 15 of the 85 centres. Secondly, the GFAP pattern is not represented in frequently used atlases of immunofluorescence patterns to date. Thirdly, a consensus on the requirement for a confirmatory laboratory method has not been reached. As such we aimed to raise awareness of this spectacular immunofluorescence pattern and its association with GFAP astrocytopathy and highlight the importance of neuronal autoantibody testing in the diagnosis of this neurological disorder.
We present three cases of GFAP astrocytopathy, in whom a diagnosis was based on the presence of suggestive staining using IIF on commercial neuronal slides according to the manufacturer's instructions (Inova Diagnostics, USA) and clinical correlation. Subsequently, antigen-specific testing by CBA (Euroimmun, Germany) confirmed this antibody specificity (Fig. 1). The clinical and laboratory findings are summarised in Table 1.
Fig. 1Indirect immunofluorescence (IIF) on rat cerebellum and cell based assay (CBA) findings for the three cases. (A) Case 1. (i) Strong linear fibrillary staining in molecular and granular layers. (ii) Staining in granular layer and white matter. (iii) Staining in granular layer and along the vessel sparing Virchow–Robin space. (iv) Moderate positive ++ 1:1. (v) Strong positive +++ 1:100. (B) Case 2. (i) Moderate fibrillary staining in molecular and granular layer. (ii) Moderate staining in granular layer. (iii) Strong staining in granular and molecular layer along vessels sparing Virchow–Robin space. (iv) Trace positive (+) 1:100. (v) Strongly positive +++ 1:32. (C) Case 3. (i) Moderate fibrillary staining in granular and molecular layers. (ii) Moderate staining in granular and molecular layer. (iii) Negative. (iv) Weak positive + 1:320. (v) Weak positive + 1:1 (image not available). ∗Low power 10× magnification ∗∗High power 20× magnification. (iv/v) CBA dilutions noted in lower right corner. GL, granular layer; ML, molecular layer; WM, white matter.
At present, IIF on neuronal tissue is a simple, cost effective and widely available means of screening for neuronal antibodies of multiple specificities. Patterns of staining can be identified by experienced readers, which in some instances are sufficiently specific to be reported (for example Hu, Ri, Yo), and in others require confirmatory testing. GFAP antibodies display a characteristic Bergmann radial pattern seen in pial, subpial, subventricular and perivascular locations of the cerebellum tissue substrate (Fig. 1).
Whilst there is some overlap with other neuronal antibodies, such as aquaporin 4 antibodies seen in neuromyelitis optica, the presence of staining in the cell body and end process throughout all layers of the cerebellum is specific for GFAP.
Ultimately, the specificity of the pattern needs to be confirmed by an antigen-specific confirmatory test which as yet is not available in the routine diagnostic setting.
We propose that the diagnostic laboratory algorithm should include this step, especially when less typical patterns of staining are seen on IIF. Until fast, reliable and cost effective confirmatory assays are available in diagnostic laboratories, this pattern cannot be reported with certainty, ultimately limiting the number of laboratories routinely reporting this pattern.
Controversy still exists regarding the clinical relevance of staining seen in serum and CSF samples, for establishing a diagnosis of GFAP astrocytopathy. Recent reports suggest that GFAP antibody positivity in CSF may represent a more important clinical finding
Clinical, neuroradiological, diagnostic and prognostic profile of autoimmune glial fibrillary acidic protein astrocytopathy: a pooled analysis of 324 cases from published data and a single-center retrospective study.
Whilst the lack of autoantibodies in the CSF usually goes against their pathogenic role in CNS disease, it is proposed by Castillo-Gomez et al. that the CNS can act as a substrate binding pathological antibodies and preventing their breach of the extracellular matrix-CSF barrier.
Whilst our small sample size limits conclusions in this area, considering the typical diagnostic laboratory algorithm for neurological autoantibody testing,
we propose the importance of testing on both serum and CSF samples to improve diagnostic yield.
In conclusion, GFAP astrocytopathy is a newly described neuroinflammatory disorder, the diagnosis of which can be challenging and is strongly supported by neuronal autoantibody testing. IIF on both CSF and serum is an ideal screening modality but requires experienced readers familiar with the specific staining pattern, whilst confirmatory tests such as CBA are likely to play an important and emerging role in this condition.
Conflicts of interest and sources of funding
The authors state that there are no conflicts of interest to disclose.
References
Fang B.
McKeon A.
Hinson S.R.
et al.
Autoimmune glial fibrillary acidic protein astrocytopathy: a novel meningoencephalomyelitis.
Clinical, neuroradiological, diagnostic and prognostic profile of autoimmune glial fibrillary acidic protein astrocytopathy: a pooled analysis of 324 cases from published data and a single-center retrospective study.
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