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Modulation of intracellular oxidant activity in C2C12 myotubes by cytokines
Author(s) -
Arbogast Sandrine,
Smith Jeffrey D,
Smith Jackie L,
Reid Michael B
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a1455-d
Subject(s) - myogenesis , tumor necrosis factor alpha , chemistry , cytokine , nitric oxide , intracellular , western blot , reactive oxygen species , endocrinology , medicine , microbiology and biotechnology , biochemistry , biology , myocyte , gene
Nitric oxide (NO) and reactive oxygen species are produced in pathological conditions (sepsis, diabetes, and COPD). These illnesses are associated with an increase in circulating cytokines. This study examines the effect of different cytokines on intracellular oxidant activity. We hypothesized that 1) Cytokines will increase oxidant activity in C2C12 myotubes 2) Interferon gamma (IFNγ) will potentiate the effect of tumor necrosis factor alpha (TNFα) increasing NO production after 48h. We measured the overall oxidant activity using DCFH‐DA and NO derived activity using DAF‐FM. C2C12 myotubes were treated for 1 hour with TNFα or IFNγ. In the second protocol, we examined the effect of repeated treatments with TNFα, IFNγ or TNF + IFN after 48hours. INOS expression was measured using RT‐PCR, and western blot. After 1 hour, TNF α increased DCFH sensitive oxidant activity (+ 117%, P< 0.05) whereas IFN had no effect. After 48h, TNFα or IFNγ increased overall oxidant activity. TNF + IFN was the only treatment that significantly increased DAF signal (+ 190%, P< 0.05). The signal was abolished using the NOS inhibitors L‐NAME and 1400W. This result was corroborated by an increase in INOS mRNA expression (+600 fold, P<0.05) and iNOS protein (+15876%, P<0.05). These data indicate that different oxidant species are involved in cytokine ‐induced response. Supported by NIH grant HL 59878.