Journal
INTERNATIONAL IMMUNOPHARMACOLOGY
Volume 9, Issue 10, Pages 1209-1214Publisher
ELSEVIER
DOI: 10.1016/j.intimp.2009.07.001
Keywords
Inflammatory myopathy; Duchenne Muscular Dystrophy; NF-kappa B inhibitor; High-throughput screening (HTS) assay; Luciferase assay; C2C12 muscle cells
Categories
Funding
- National Institute of Health [RO1-AR050478, 5U54HD053177, RO1-NS029525]
- Foundation to Eradicate Dystrophy, Muscular Dystrophy Association
- Myositis Association
- US Department of Defense [W81XWH-05-1-0616]
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Specific therapies are not available for inflammatory muscle diseases. We and others have shown that the pro-inflammatory NF-kappa B pathway is highly activated in these conditions. Since NF-kappa B is an important therapeutic target, we decided to utilize an in vitro screening assay to identify potential inhibitors that block TNF-alpha induced NF-kappa B activation in a C2C12 muscle line stably expressing an NF-kappa B luciferase reporter gene. Upon evaluation of multiple anti-inflammatory agents in undifferentiated myoblasts as well as differentiated myotubes, we found different levels of inhibition depending on the state of differentiation. Interestingly, we found that some drugs that are known to inhibit NF-kappa B in immune cells were not effective in muscle cells. Drug toxicity was assessed for using an MTT cell viability assay, and the validity of the luciferase assay was verified by immunostaining for NF-kappa B nuclear translocation in myoblasts. In conclusion, we have determined the optimal assay conditions for detecting potentially valuable NF-kappa B inhibitors for the first time in a muscle cell line that may have significant therapeutic potential for inflammatory muscle diseases. (C) 2009 Elsevier B.V. All rights reserved.
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