Figure 3.

Gene expression profiles for immunological activities in growing feathers of vitiligo-expressing Smyth chickens. Growing feather samples were taken from Smyth chickens beginning at 1-day post-hatch and continuing twice a week for 113 days. cDNA from samples collected from 39 days before- through 13 days post-vitiligo onset (i.e. -39 d to 13 d) were analyzed for relative gene expression of: (A) potential early markers of immunological activities, (B) markers of pro-inflammatory and recruitment activities and (C) markers of cell-mediated immunity and anti-inflammatory activities. Relative gene expression was determined by the efficiency-calibrated ΔΔCt method and is expressed as fold change relative to the calibrator sample. Gene expression profiles were obtained for individual chickens, using individual-specific day 29 samples as calibrators and 28S as a reference gene. Data were aligned and averaged by time (days) relative to age of vitiligo onset (set to 0 d; indicated by vertical dashed line). Smyth chickens were 43 to 57 days of age at time of vitiligo onset. The overall effect of time was determined using a mixed effects regression model setting time and individual bird as a fixed and random effect respectively. When a significant time effect was found, post-hoc multiple means comparisons were made against the -39 d samples using the Tukey-Kramer p-value adjustment. Differences were considered significant at P < 0.05 (*). Data are plotted as mean ± SEM; n = 3 to 8 chickens per time point.