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Inducible nitric oxide synthase in Theiler's murine encephalomyelitis virus infection
E L Oleszak1, C D Katsetos, J Kuzmak
1Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania 19106, USA.
Abstract:
We investigated the role of inducible nitric oxide synthase (iNOS) in Theiler's murine encephalomyelitis virus (TMEV) infection of susceptible (SJL) and resistant (C57BL/6 [B6]) strains of mice. TMEV is an excellent model of virus-induced demyelinating disease, such as multiple sclerosis (MS). Previous studies of others have suggested that NO may play a role in the pathogenesis of demyelinating disease. The presence and level of iNOS were determined in the brains and spinal cords of SJL and B6 TMEV-infected mice by the following methods: (i) PCR amplification of iNOS transcripts, followed by Southern blotting with an iNOS-specific probe, and (ii) immunohistochemical staining with an anti-iNOS-specific affinity-purified rabbit antibody. iNOS-specific transcripts were determined in the brains and spinal cord of both SJL and B6 TMEV-infected mice on days 0 (control), days 3, 6, and 10 (encephalitic stage of disease), and days 39 to 42, 66, and 180 (demyelinating phase) postinfection (p.i.). iNOS-specific transcripts were found in the brains and spinal cords of both SJL and B6 TMEV-infected mice at 6, 10, and 39 (SJL) days p.i., but they were absent in mock-infected mice and in TMEV-infected SJL and B6 mice at 0, 3, 66, and 180 days p.i. Immunohistochemical staining confirmed the presence of iNOS protein in both TMEV-infected SJL and B6 mice at days 6 and 10 p.i., but not at days 0, 3, 66, and 180 days p.i. Weak iNOS staining was also observed in TMEV-infected SJL mice at 42 days p.i. iNOS-positive staining was found in reactive astrocytes surrounding areas of necrotizing inflammation, particularly in the midbrain. Weak iNOS staining was also observed in cells of the monocyte/macrophage lineage in areas of parenchymal inflammation and necrosis (mesencephalon) and in leptomeningeal and white matter perivascular infiltrates of the spinal cord. Rod-shaped microglia-like cells and foamy macrophages (myelin-laden) were iNOS negative. These results suggest that NO does not play a direct role in the late phase of demyelinating disease in TMEV-infected mice.
Insights
Inducible nitric oxide synthase (iNOS) was investigated in Theiler's murine encephalomyelitis virus (TMEV) infection. Results indicate iNOS is not directly involved in the late demyelinating phase of this viral disease model.
Area of Science:
- Neuroscience
- Immunology
- Virology
Background:
- Theiler's murine encephalomyelitis virus (TMEV) infection serves as a model for virus-induced demyelinating diseases like multiple sclerosis.
- Nitric oxide (NO) has been implicated in the pathogenesis of demyelinating conditions.
Purpose of the Study:
- To investigate the role of inducible nitric oxide synthase (iNOS) in TMEV infection in susceptible (SJL) and resistant (C57BL/6) mice.
- To determine the presence and levels of iNOS during the acute and chronic phases of TMEV infection.
Main Methods:
- Quantitative PCR (qPCR) and Southern blotting to detect iNOS transcripts.
- Immunohistochemical staining to identify iNOS protein expression in brain and spinal cord tissues.
- Analysis of iNOS expression in SJL and C57BL/6 mice at various time points post-TMEV infection.
Main Results:
- iNOS transcripts and protein were detected in both mouse strains at acute (6-10 days post-infection) and early chronic (39 days post-infection in SJL mice) stages.
- iNOS expression was localized to reactive astrocytes and cells of the monocyte/macrophage lineage in inflamed areas.
- iNOS was absent during the late chronic (66-180 days post-infection) and mock-infected stages.
Conclusions:
- Nitric oxide (NO) does not appear to play a direct role in the late demyelinating phase of TMEV infection.
- iNOS expression is associated with the inflammatory response during the acute and early chronic phases of TMEV infection.