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Inducible nitric oxide synthase gene expression and enzyme activity correlate with disease activity in murine
A H Cross1, R M Keeling, S Goorha
1Department of Neurology and Neurosurgery, Washington University School of Medicine, St. Louis, MO 63110, USA. crossa@neuro.wustl.edu
Abstract:
Messenger RNA encoding inducible NO synthase (iNOS) was measured by competitive reverse transcriptase polymerase chain reaction (cRT-PCR) and ribonuclease protection assays in spinal cords from mice at varying stages of experimental allergic encephalomyelitis (EAE) and from control mice. iNOS mRNA was increased in spinal cords from mice with acute EAE. cRT-PCR assays revealed a 10-20-fold increase in iNOS mRNA in spinal cords during acute EAE compared with the level observed in normal mouse spinal cords. Functional iNOS activity, as assessed by assay of calcium-independent citrulline production, was also significantly increased in spinal cords from mice with acute EAE in comparison to normal controls. The correlation of functional iNOS expression with active disease in EAE in consistent with a pathogenic role for excess NO in this model of cell-mediated central nervous system autoimmunity.
Insights
This study shows increased inducible nitric oxide synthase (iNOS) mRNA and activity in the spinal cord during experimental allergic encephalomyelitis (EAE) in mice. These findings suggest a role for excess nitric oxide (NO) in this autoimmune disease model.
Area of Science:
- Neuroimmunology
- Molecular Biology
- Pathology
Background:
- Experimental allergic encephalomyelitis (EAE) is a mouse model for central nervous system (CNS) autoimmune diseases.
- Inducible nitric oxide synthase (iNOS) produces nitric oxide (NO), a molecule implicated in various physiological and pathological processes.
Purpose of the Study:
- To investigate the expression and activity of iNOS in the spinal cord during EAE.
- To determine the potential role of iNOS-derived NO in the pathogenesis of EAE.
Main Methods:
- Messenger RNA (mRNA) encoding iNOS was quantified using competitive reverse transcriptase polymerase chain reaction (cRT-PCR) and ribonuclease protection assays.
- Functional iNOS activity was assessed by measuring calcium-independent citrulline production.
Main Results:
- iNOS mRNA levels were significantly elevated (10-20 fold) in the spinal cords of mice with acute EAE compared to controls.
- Functional iNOS activity was also markedly increased in spinal cords during active EAE.
Conclusions:
- The heightened expression and activity of iNOS in EAE spinal cords correlate with active disease.
- These results support a pathogenic role for excess nitric oxide (NO) in this model of CNS autoimmunity.