The role of cytokines in the pathogenesis and immunity of Marek's disease (MD), a herpesvirus-induced T-cell lymphoma in chickens, is poorly understood. Two different experiments were used to examine the potential role of particular cytokines in the pathogenesis and immune responses of MD. First, chicken embryo fibroblasts (CEF) were stimulated with lipopolysaccharide (LPS) and/or recombinant chicken interferon-γ (rChIFN-γ) and used to develop techniques for examining transcription of IFN-α, IFN-γ, inducible nitric oxide synthase (iNOS), interleukin (IL)-1β, IL-2, IL-6 and IL-8 by reverse transcription polymerase chain reaction (RT-PCR). Addition of LPS and/or rChIFN-γ resulted in the up-regulation of mRNA for iNOS, IL-1β and IL-6, while IFN-γ was up-regulated by LPS alone. IL-2 was down-regulated by the treatments. Second, to determine the effects of Marek's disease herpesvirus (MDV) infection on cytokine transcription in vivo, chickens were infected with MDV at 21 days of age and examined at 7 days post-infection (p.i.) (exp. 1) or were infected with MDV at 1 day of age and examined from 3 to 15 days p.i. (exp. 2). In MDV-infected chickens, IFN-γ transcription was up-regulated as early as 3 days p.i. until the termination of the experiment at 15 days p.i., while iNOS and IL-1β were up-regulated between 6 and 15 days p.i. Infection of 1-day-old chicks increased levels of mRNA for IFN-γ and iNOS between 16- and 64-fold at 9 days p.i. These results suggest that IFN-γ and iNOS may play an important role in the pathogenesis of MD.