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Aquaporin-4-binding autoantibodies in patients with neuromyelitis optica impair glutamate transport by down-regulating EAAT2.

Hinson SR, Roemer SF, Lucchinetti CF, Fryer JP, Kryzer TJ, Chamberlain JL, Howe CL, Pittock SJ, Lennon VA - J. Exp. Med. (2008)

Bottom Line: The effect of NMO-IgG on astrocytes has not been studied.Marked reduction of EAAT2 in AQP4-deficient regions of NMO patient spinal cord lesions supports our immunocytochemical and immunoprecipitation data.Thus, binding of NMO-IgG to astrocytic AQP4 initiates several potentially neuropathogenic mechanisms: complement activation, AQP4 and EAAT2 down-regulation, and disruption of glutamate homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine and Pathology, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.

ABSTRACT
Neuromyelitis optica (NMO)-immunoglobulin G (IgG) is a clinically validated serum biomarker that distinguishes relapsing central nervous system (CNS) inflammatory demyelinating disorders related to NMO from multiple sclerosis. This autoantibody targets astrocytic aquaporin-4 (AQP4) water channels. Clinical, radiological, and immunopathological data suggest that NMO-IgG might be pathogenic. Characteristic CNS lesions exhibit selective depletion of AQP4, with and without associated myelin loss; focal vasculocentric deposits of IgG, IgM, and complement; prominent edema; and inflammation. The effect of NMO-IgG on astrocytes has not been studied. In this study, we demonstrate that exposure to NMO patient serum and active complement compromises the membrane integrity of CNS-derived astrocytes. Without complement, astrocytic membranes remain intact, but AQP4 is endocytosed with concomitant loss of Na(+)-dependent glutamate transport and loss of the excitatory amino acid transporter 2 (EAAT2) . Our data suggest that EAAT2 and AQP4 exist in astrocytic membranes as a macromolecular complex. Transport-competent EAAT2 protein is up-regulated in differentiating astrocyte progenitors and in nonneural cells expressing AQP4 transgenically. Marked reduction of EAAT2 in AQP4-deficient regions of NMO patient spinal cord lesions supports our immunocytochemical and immunoprecipitation data. Thus, binding of NMO-IgG to astrocytic AQP4 initiates several potentially neuropathogenic mechanisms: complement activation, AQP4 and EAAT2 down-regulation, and disruption of glutamate homeostasis.

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Glutamate transporter expression in human tissue. (A) Normal spinal cord from a single subject. Duplicate sections were dual stained for AQP4 (red) and EAAT2 or EAAT1 (green), i.e., four sections. Merged images show AQP4 and EAAT2 colocalization (top, yellow), but no colocalization of AQP4 and EAAT1 (bottom); mn, motor neuron. (B) Spinal cord tissue from a single NMO-IgG–seropositive patient. Three sections of nonlesioned lumbar region (top) serve as staining control for lesioned cord (bottom). The lack of complement deposition (C9neo, brick red in lesioned cord, bottom) and high expression of AQP4 in both white and gray matter are typical of normal cord tissue; EAAT2 is highly enriched in gray matter. Prominent deposition of C9neo in gray matter of lesioned thoracic cord (bottom, same patient) corresponds to focal regions of AQP4 and EAAT2 loss in adjacent sections. AQP4 is partially retained in the white matter. Asterisk, central canal. Bars: (A) 20 μm; (B) 200 μm.
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fig5: Glutamate transporter expression in human tissue. (A) Normal spinal cord from a single subject. Duplicate sections were dual stained for AQP4 (red) and EAAT2 or EAAT1 (green), i.e., four sections. Merged images show AQP4 and EAAT2 colocalization (top, yellow), but no colocalization of AQP4 and EAAT1 (bottom); mn, motor neuron. (B) Spinal cord tissue from a single NMO-IgG–seropositive patient. Three sections of nonlesioned lumbar region (top) serve as staining control for lesioned cord (bottom). The lack of complement deposition (C9neo, brick red in lesioned cord, bottom) and high expression of AQP4 in both white and gray matter are typical of normal cord tissue; EAAT2 is highly enriched in gray matter. Prominent deposition of C9neo in gray matter of lesioned thoracic cord (bottom, same patient) corresponds to focal regions of AQP4 and EAAT2 loss in adjacent sections. AQP4 is partially retained in the white matter. Asterisk, central canal. Bars: (A) 20 μm; (B) 200 μm.

Mentions: Our observations in primary astrocytes, type 2 differentiated CG-4 cells, and transfected nonneural HEK-293 cells indicate that the interaction of NMO-IgG with AQP4 induces at least three possible outcomes, each potentially pathogenic: (a) complement activation, (b) down-regulation of AQP4, and (c) coupled down-regulation of the EAAT2 glutamate transporter. Our immunohistochemical analysis of nonpathologic human CNS tissue (both cortical and spinal cord) reveal that EAAT2, but not EAAT1, normally colocalizes with AQP4 in gray matter astrocytes (Fig. 5 A) and that EAAT2 is most enriched in spinal cord gray matter (Fig. 5 B, top). These findings were reproducible and consistent with published rodent cord findings (19, 29).


Aquaporin-4-binding autoantibodies in patients with neuromyelitis optica impair glutamate transport by down-regulating EAAT2.

Hinson SR, Roemer SF, Lucchinetti CF, Fryer JP, Kryzer TJ, Chamberlain JL, Howe CL, Pittock SJ, Lennon VA - J. Exp. Med. (2008)

Glutamate transporter expression in human tissue. (A) Normal spinal cord from a single subject. Duplicate sections were dual stained for AQP4 (red) and EAAT2 or EAAT1 (green), i.e., four sections. Merged images show AQP4 and EAAT2 colocalization (top, yellow), but no colocalization of AQP4 and EAAT1 (bottom); mn, motor neuron. (B) Spinal cord tissue from a single NMO-IgG–seropositive patient. Three sections of nonlesioned lumbar region (top) serve as staining control for lesioned cord (bottom). The lack of complement deposition (C9neo, brick red in lesioned cord, bottom) and high expression of AQP4 in both white and gray matter are typical of normal cord tissue; EAAT2 is highly enriched in gray matter. Prominent deposition of C9neo in gray matter of lesioned thoracic cord (bottom, same patient) corresponds to focal regions of AQP4 and EAAT2 loss in adjacent sections. AQP4 is partially retained in the white matter. Asterisk, central canal. Bars: (A) 20 μm; (B) 200 μm.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2571922&req=5

fig5: Glutamate transporter expression in human tissue. (A) Normal spinal cord from a single subject. Duplicate sections were dual stained for AQP4 (red) and EAAT2 or EAAT1 (green), i.e., four sections. Merged images show AQP4 and EAAT2 colocalization (top, yellow), but no colocalization of AQP4 and EAAT1 (bottom); mn, motor neuron. (B) Spinal cord tissue from a single NMO-IgG–seropositive patient. Three sections of nonlesioned lumbar region (top) serve as staining control for lesioned cord (bottom). The lack of complement deposition (C9neo, brick red in lesioned cord, bottom) and high expression of AQP4 in both white and gray matter are typical of normal cord tissue; EAAT2 is highly enriched in gray matter. Prominent deposition of C9neo in gray matter of lesioned thoracic cord (bottom, same patient) corresponds to focal regions of AQP4 and EAAT2 loss in adjacent sections. AQP4 is partially retained in the white matter. Asterisk, central canal. Bars: (A) 20 μm; (B) 200 μm.
Mentions: Our observations in primary astrocytes, type 2 differentiated CG-4 cells, and transfected nonneural HEK-293 cells indicate that the interaction of NMO-IgG with AQP4 induces at least three possible outcomes, each potentially pathogenic: (a) complement activation, (b) down-regulation of AQP4, and (c) coupled down-regulation of the EAAT2 glutamate transporter. Our immunohistochemical analysis of nonpathologic human CNS tissue (both cortical and spinal cord) reveal that EAAT2, but not EAAT1, normally colocalizes with AQP4 in gray matter astrocytes (Fig. 5 A) and that EAAT2 is most enriched in spinal cord gray matter (Fig. 5 B, top). These findings were reproducible and consistent with published rodent cord findings (19, 29).

Bottom Line: The effect of NMO-IgG on astrocytes has not been studied.Marked reduction of EAAT2 in AQP4-deficient regions of NMO patient spinal cord lesions supports our immunocytochemical and immunoprecipitation data.Thus, binding of NMO-IgG to astrocytic AQP4 initiates several potentially neuropathogenic mechanisms: complement activation, AQP4 and EAAT2 down-regulation, and disruption of glutamate homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine and Pathology, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.

ABSTRACT
Neuromyelitis optica (NMO)-immunoglobulin G (IgG) is a clinically validated serum biomarker that distinguishes relapsing central nervous system (CNS) inflammatory demyelinating disorders related to NMO from multiple sclerosis. This autoantibody targets astrocytic aquaporin-4 (AQP4) water channels. Clinical, radiological, and immunopathological data suggest that NMO-IgG might be pathogenic. Characteristic CNS lesions exhibit selective depletion of AQP4, with and without associated myelin loss; focal vasculocentric deposits of IgG, IgM, and complement; prominent edema; and inflammation. The effect of NMO-IgG on astrocytes has not been studied. In this study, we demonstrate that exposure to NMO patient serum and active complement compromises the membrane integrity of CNS-derived astrocytes. Without complement, astrocytic membranes remain intact, but AQP4 is endocytosed with concomitant loss of Na(+)-dependent glutamate transport and loss of the excitatory amino acid transporter 2 (EAAT2) . Our data suggest that EAAT2 and AQP4 exist in astrocytic membranes as a macromolecular complex. Transport-competent EAAT2 protein is up-regulated in differentiating astrocyte progenitors and in nonneural cells expressing AQP4 transgenically. Marked reduction of EAAT2 in AQP4-deficient regions of NMO patient spinal cord lesions supports our immunocytochemical and immunoprecipitation data. Thus, binding of NMO-IgG to astrocytic AQP4 initiates several potentially neuropathogenic mechanisms: complement activation, AQP4 and EAAT2 down-regulation, and disruption of glutamate homeostasis.

Show MeSH
Related in: MedlinePlus