Chickens from broiler, Leghorn, and Fayoumi lines were fed diets with ingredients to affect immune function: β-glucans, ascorbic acid, or corticosterone. Spleens were tested for expression of genes involved in immune response: interleukin-4 (IL-4), IL-6, IL-18, and macrophage inflammatory protein-1β (MIP-1β). Birds from the broiler line did not show any change in splenic gene expression associated with the dietary immunomodulators, perhaps due to the stringent selection of these birds for growth. The corticosterone diet was associated with increased expression of IL-4, indicative of an immune response relying primarily on humoral defenses. The Leghorn and Fayoumi lines showed opposing changes in expression of IL-4, IL-6, and IL-18 in response to the ascorbic acid and β-glucans enhanced diets, suggesting that processing of these immunomodulators and/or immune signaling in these lines are different. Our findings emphasize the need to further evaluate the effects of dietary immunomodulators before applying them in commercial settings.