FIGURE 5.

In vitro morphine withdrawal and corticosterone treatment decreased IL-12p40 promoter activity and message levels in LPS-stimulated CRL2019 macrophages. CRL2019 macrophages were transfected with IL-12p40 promoter construct followed by morphine withdrawal paradigm and stimulated for 6 h with 100 ng/ml LPS. Promoter activity (A) is presented as standardized luciferase activity. Total RNA was extracted, and message levels were analyzed using real-time PCR (B) and gel-based PCR (C) for IL-12p40. Housekeeping gene β-actin was amplified as an internal control. The graph was plotted against data normalized to β-actin expression. Each treatment group was tested in triplicate, and results are representative of at least three independent experiments ± S.D. (error bars). #, p < 0.01 versus control samples; *, p < 0.01; **, p < 0.05; ***, p < 0.001. RLU, Relative Luciferase Units.