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Extensive dysregulations of oligodendrocytic and astrocytic connexins are associated with disease progression in an amyotrophic lateral sclerosis mouse model.

Cui Y, Masaki K, Yamasaki R, Imamura S, Suzuki SO, Hayashi S, Sato S, Nagara Y, Kawamura MF, Kira J - J Neuroinflammation (2014)

Bottom Line: Non-cell-autonomous motor neuronal death is suggested in a mutant Cu/Zn superoxide dismutase 1 (mSOD1)-mediated amyotrophic lateral sclerosis (ALS) model, in which glial cells play significant roles in disease progression.The levels of Cx47 and Cx32 mRNAs were also decreased at these stages.Our findings indicate that oligodendrocytic and astrocytic GJ proteins in the anterior horns of spinal cord in mSOD1-Tg mice are profoundly affected at the disease-progressive and end stages, where disruption of GJs among glial cells may exacerbate motor neuronal death.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurology, Neurological Institute, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. kira@neuro.med.kyushu-u.ac.jp.

ABSTRACT

Background: Non-cell-autonomous motor neuronal death is suggested in a mutant Cu/Zn superoxide dismutase 1 (mSOD1)-mediated amyotrophic lateral sclerosis (ALS) model, in which glial cells play significant roles in disease progression. Connexins (Cxs) form homotypic or heterotypic gap junctions (GJs) and allow direct intercellular communications among nervous tissue cells. The role of Cxs in motor neuron disease has never been investigated; therefore, we aimed to evaluate alterations of Cxs in mSOD1-transgenic (mSOD1-Tg) mice in comparison with their non-transgenic (non-Tg) littermates at the same ages.

Methods: We pathologically evaluated temporal changes to astrocytic Cx43/Cx30 and oligodendrocytic Cx47/Cx32 immunoreactivities at presymptomatic, disease-progressive, and end stages, relative to aquaporin-4 (AQP4), glial fibrillary acidic protein (GFAP), excitatory amino acid transporter-2 (EAAT2), myelin-oligodendrocyte glycoprotein (MOG), and Nogo-A immunoreactivities, and observed neuronal loss by NeuN and neurofilament immunostaining, and microglial response by Iba-1 immunostaining. We also performed quantitative immunoblotting and real-time PCR analyses for Cxs.

Results: The mSOD1-Tg mice showed neuronal and axonal loss in the anterior horns of the lumbar spinal cord accompanied by increased activation of microglia compared with non-Tg mice at the disease-progressive and end stages. Expression patterns of Cxs were not different between mSOD1-Tg and non-Tg mice at the presymptomatic stage, but immunoreactivities for GFAP, Cx43, Cx30 and AQP4 were increased in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages. By contrast, Cx47 and Cx32 immunoreactivities were markedly diminished in Nogo-A-positive oligodendrocytes in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages, especially in oligodendrocytes showing SOD1 accumulation. EAAT2 immunoreactivity was also diminished in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages. Quantitative immunoblotting revealed a significant reduction in Cx47 and Cx32 protein levels in mSOD1-Tg mice at the disease-progressive and end stages. The levels of Cx47 and Cx32 mRNAs were also decreased at these stages.

Conclusions: Our findings indicate that oligodendrocytic and astrocytic GJ proteins in the anterior horns of spinal cord in mSOD1-Tg mice are profoundly affected at the disease-progressive and end stages, where disruption of GJs among glial cells may exacerbate motor neuronal death.

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Overexpression of SOD1 in oligodendrocytes of mSOD1-Tg mice. (A-C) Immunoreactivity for SOD1 is observed in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice at 20 weeks of age and immunoreactivity for Cx47 is preserved at the surface of oligodendrocytes. (D-F) However, SOD1 is markedly accumulated in neurons and astrocytes in the anterior horns of mSOD1-Tg mice at 20 weeks of age and accumulation of SOD1 was also observed in oligodendrocytes with Cx47-immunopositivity in their cytoplasm (arrows). Membranous staining of Cx47 is not visible in abnormal-shaped oligodendrocytes. The nucleus is stained with DAPI (A-F blue). Scale bar; 5 μm (A-F).
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Figure 7: Overexpression of SOD1 in oligodendrocytes of mSOD1-Tg mice. (A-C) Immunoreactivity for SOD1 is observed in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice at 20 weeks of age and immunoreactivity for Cx47 is preserved at the surface of oligodendrocytes. (D-F) However, SOD1 is markedly accumulated in neurons and astrocytes in the anterior horns of mSOD1-Tg mice at 20 weeks of age and accumulation of SOD1 was also observed in oligodendrocytes with Cx47-immunopositivity in their cytoplasm (arrows). Membranous staining of Cx47 is not visible in abnormal-shaped oligodendrocytes. The nucleus is stained with DAPI (A-F blue). Scale bar; 5 μm (A-F).

Mentions: To elucidate the mechanism by which mSOD1 affects expression of Cx47 and Cx32, we performed double immunofluorescence staining for mSOD1 and Cx47/Cx32 in non-Tg and mSOD1-Tg mice. At 20 weeks of age, immunopositivity for SOD1 was subtle in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice (Figure 7A-C). By contrast, expression of SOD1 was markedly upregulated in the anterior horns of mSOD1-Tg mice. The cytoplasmic accumulation of SOD1 was also observed in dysmorphic Cx47-positive oligodendrocytes in mSOD1-Tg mice, and membranous staining of Cx47 was no longer observed on these abnormal oligodendrocytes (Figure 7D-F).


Extensive dysregulations of oligodendrocytic and astrocytic connexins are associated with disease progression in an amyotrophic lateral sclerosis mouse model.

Cui Y, Masaki K, Yamasaki R, Imamura S, Suzuki SO, Hayashi S, Sato S, Nagara Y, Kawamura MF, Kira J - J Neuroinflammation (2014)

Overexpression of SOD1 in oligodendrocytes of mSOD1-Tg mice. (A-C) Immunoreactivity for SOD1 is observed in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice at 20 weeks of age and immunoreactivity for Cx47 is preserved at the surface of oligodendrocytes. (D-F) However, SOD1 is markedly accumulated in neurons and astrocytes in the anterior horns of mSOD1-Tg mice at 20 weeks of age and accumulation of SOD1 was also observed in oligodendrocytes with Cx47-immunopositivity in their cytoplasm (arrows). Membranous staining of Cx47 is not visible in abnormal-shaped oligodendrocytes. The nucleus is stained with DAPI (A-F blue). Scale bar; 5 μm (A-F).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4016493&req=5

Figure 7: Overexpression of SOD1 in oligodendrocytes of mSOD1-Tg mice. (A-C) Immunoreactivity for SOD1 is observed in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice at 20 weeks of age and immunoreactivity for Cx47 is preserved at the surface of oligodendrocytes. (D-F) However, SOD1 is markedly accumulated in neurons and astrocytes in the anterior horns of mSOD1-Tg mice at 20 weeks of age and accumulation of SOD1 was also observed in oligodendrocytes with Cx47-immunopositivity in their cytoplasm (arrows). Membranous staining of Cx47 is not visible in abnormal-shaped oligodendrocytes. The nucleus is stained with DAPI (A-F blue). Scale bar; 5 μm (A-F).
Mentions: To elucidate the mechanism by which mSOD1 affects expression of Cx47 and Cx32, we performed double immunofluorescence staining for mSOD1 and Cx47/Cx32 in non-Tg and mSOD1-Tg mice. At 20 weeks of age, immunopositivity for SOD1 was subtle in the nuclei and cytoplasm of oligodendrocytes in the anterior horns of non-Tg mice (Figure 7A-C). By contrast, expression of SOD1 was markedly upregulated in the anterior horns of mSOD1-Tg mice. The cytoplasmic accumulation of SOD1 was also observed in dysmorphic Cx47-positive oligodendrocytes in mSOD1-Tg mice, and membranous staining of Cx47 was no longer observed on these abnormal oligodendrocytes (Figure 7D-F).

Bottom Line: Non-cell-autonomous motor neuronal death is suggested in a mutant Cu/Zn superoxide dismutase 1 (mSOD1)-mediated amyotrophic lateral sclerosis (ALS) model, in which glial cells play significant roles in disease progression.The levels of Cx47 and Cx32 mRNAs were also decreased at these stages.Our findings indicate that oligodendrocytic and astrocytic GJ proteins in the anterior horns of spinal cord in mSOD1-Tg mice are profoundly affected at the disease-progressive and end stages, where disruption of GJs among glial cells may exacerbate motor neuronal death.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurology, Neurological Institute, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. kira@neuro.med.kyushu-u.ac.jp.

ABSTRACT

Background: Non-cell-autonomous motor neuronal death is suggested in a mutant Cu/Zn superoxide dismutase 1 (mSOD1)-mediated amyotrophic lateral sclerosis (ALS) model, in which glial cells play significant roles in disease progression. Connexins (Cxs) form homotypic or heterotypic gap junctions (GJs) and allow direct intercellular communications among nervous tissue cells. The role of Cxs in motor neuron disease has never been investigated; therefore, we aimed to evaluate alterations of Cxs in mSOD1-transgenic (mSOD1-Tg) mice in comparison with their non-transgenic (non-Tg) littermates at the same ages.

Methods: We pathologically evaluated temporal changes to astrocytic Cx43/Cx30 and oligodendrocytic Cx47/Cx32 immunoreactivities at presymptomatic, disease-progressive, and end stages, relative to aquaporin-4 (AQP4), glial fibrillary acidic protein (GFAP), excitatory amino acid transporter-2 (EAAT2), myelin-oligodendrocyte glycoprotein (MOG), and Nogo-A immunoreactivities, and observed neuronal loss by NeuN and neurofilament immunostaining, and microglial response by Iba-1 immunostaining. We also performed quantitative immunoblotting and real-time PCR analyses for Cxs.

Results: The mSOD1-Tg mice showed neuronal and axonal loss in the anterior horns of the lumbar spinal cord accompanied by increased activation of microglia compared with non-Tg mice at the disease-progressive and end stages. Expression patterns of Cxs were not different between mSOD1-Tg and non-Tg mice at the presymptomatic stage, but immunoreactivities for GFAP, Cx43, Cx30 and AQP4 were increased in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages. By contrast, Cx47 and Cx32 immunoreactivities were markedly diminished in Nogo-A-positive oligodendrocytes in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages, especially in oligodendrocytes showing SOD1 accumulation. EAAT2 immunoreactivity was also diminished in the anterior horns of mSOD1-Tg mice at the disease-progressive and end stages. Quantitative immunoblotting revealed a significant reduction in Cx47 and Cx32 protein levels in mSOD1-Tg mice at the disease-progressive and end stages. The levels of Cx47 and Cx32 mRNAs were also decreased at these stages.

Conclusions: Our findings indicate that oligodendrocytic and astrocytic GJ proteins in the anterior horns of spinal cord in mSOD1-Tg mice are profoundly affected at the disease-progressive and end stages, where disruption of GJs among glial cells may exacerbate motor neuronal death.

Show MeSH
Related in: MedlinePlus