Necrotic enteritis toxin B (NetB)-producing Clostridium perfringens (CP) type A is the etiological agent of necrotic enteritis (NE) - an economically significant disease in broiler chickens. Understanding the immune response to CP infection in broiler chickens is becoming important to develop effective vaccines against NE. An experiment was conducted to determine the expression levels of selected cytokine genes in the intestine and cecal tonsil of CP-challenged broiler chickens. In a floor-pen housing, broiler chickens were randomly assigned to the following treatment groups: 1) bacitracin methylene disalicylate (BMD)-free control diet with no CP challenge (CX), 2) BMD-supplemented diet with no CP challenge (CM), 3) BMD-free control diet with CP challenge (PCX), or 4) BMD-supplemented diet with CP challenge (PCM). The establishment of CP infection was confirmed, with the treatment groups exposed to CP having a 1.5 to 2-fold higher CP levels (P < 0.05) compared to the non-exposed groups. On day 1 and 7 post-challenge, jejunal segments and cecal tonsils were collected from experimental chickens for quantitative real-time RT-PCR analysis to determine the expression levels of interleukin (IL)-1β, interferon-γ (IFN-γ), IL-2, IL-13, IL-17, IL-10, and transforming growth factor (TGF)-β genes. Levels of antibodies to CP recombinant proteins were also determined in the plasma of experimental chickens. Results indicated that on day 7 post-challenge, IL-1β (proinflammatory cytokine), IL-13 (Th2 cytokine), and IL-17 (Th17 cytokine) were upregulated (P < 0.05) in CP-challenged PCX and PCM treatments, compared to the unchallenged (control) CX and CM treatments. A reverse trend was observed for TGF-β (anti-inflammatory cytokine), while no change was observed in IFN-γ (Th1 cytokine). Levels of plasma antibodies (IgY) to CP recombinant proteins were higher in CP-challenged treatments (PCX and PCM; P < 0.05), compared to their corresponding controls (CX and CM). It was concluded that CP infection induced inflammatory response in the intestine of broiler chickens, and the mechanisms of inflammation are probably mediated via Th2 and Th17 cells.