FIGURE 6.

Effect of rearing environment and source of nutrients during lactation on absolute CD3+CD4+ T helper cells (A), CD3+CD8+ cytotoxic T cells (B), and CD3+CD4+Foxp3+ regulatory T cells (C) in blood [×10⁶/ml] in 4-, 11-, 20-, 32-, and 55-day-old pigs (LS means). One pig from each litter (10 litter per treatments) was selected for blood sampling. The same pigs were sampled throughout the entire study. The whole blood samples (3 mL) were obtained at day 4 (initial), 11, 20 (weanling), 32, 55, and 103 of ages. PBMCs were isolated using Histopaque^®1077 (Sigma-Aldrich, St. Louis, MO, United States) density gradient centrifugation, counted by hemocytometer, and expressed as blood concentration based on the volume of whole blood used to isolate the PBMCs, the total number of PBMCs isolated, and the proportions of lymphocyte subsets determined by immunofluorescent staining and flow cytometry. Results on day 103 are presented in Supplementary Table S4. Two stars refer to ages having significant differences between treatments when compared to other ages at probability value <0.01.