Figure 5
Interference with ChTLR15 downregulated the expression levels of molecules in the ChTLR15/ChNF-κB-ChNLRP3/ChIL-1β signaling pathway. A Monolayer DF1 cells (1.5 × 106 cells per well) were treated with siChTLR15#1 for 2 h and then stimulated with purified E. tenella sporozoites (4 × 105 sporozoites per well). At 0, 2, 4, 6, and 8 h post-stimulation, cells in each group were collected to quantify the mRNA levels of ChTLR15, ChMyD88, ChNF-κB, ChNLRP3, ChCaspase-1, ChIL-18 and ChIL-1β by quantitative real-time PCR (qPCR). Chicken β-actin was used as a reference gene. B DF1 cells transfected with negative control (NC) siRNA and stimulated with (NC + Stim) or without (NC + Unstim) E. tenella sporozoites were designed as controls. Protein expression levels of ChTLR15 and ChNLRP3 in cells were determined by Western blot at 0, 2, 4, 6, and 8 h post-stimulation. C Protein expression levels of ChIL-1β in culture supernatant were analyzed using an ELISA kit (Jiancheng Bioengineering Institute, Nanjing, China) at 0, 2, 4, 6, and 8 h post-stimulation. *Indicates a significant difference (*p < 0.05, **p < 0.01, ***p < 0.001).