Figure 4.

TLR2, TLR3, and TLR9 surface protein expression in monocytes after toll-like receptor agonists stimulation. Peripheral blood mononuclear cells from AITD patients and healthy controls were treated with a TLR2, TLR3, or TLR9 agonist or PBS as basal for 24 h and stained with anti-CD14 and anti-TLR2, TLR3, and TLR9 mAbs for flow cytometry. We first selected singlets using forward scatter area Vs forward scatter height parameters. From the singlet population, we removed dead cells from total events by gating on LIVE/DEAD events. Then, we selected total monocytes from the living using the pan-monocyte marker CD14, then TLR2 positive, TLR3 positive, and TLR9 positive, respectively. (A) Flow cytometry dot plots from AITD and HC depicting TLR2 expression by CD14 enriched monocyte cells (left panels). The number in the right quadrants represents the percentage of CD14⁺TLR2⁺ between AITD and healthy controls. (B) Flow cytometry dot plots from AITD and HC depicting TLR3 expression by CD14 enriched monocyte cells (left panels). Percentages of CD14⁺TLR3⁺ cells were compared between HC and AITD patients (right panels). (C) Flow cytometry dot plots from AITD and HC depicting TLR9 expression by CD14 enriched monocyte cells (left panels). Percentages of CD14⁺TLR9⁺ cells were compared between HC and AITD patients (right panels). Each bar represents the mean ± SD of three groups. Level of significant is *P < 0.05 and **P < 0.01. AITD, autoimmune thyroid disease; HT, Hashimoto’s thyroiditis; GD, Graves’ disease; HC, healthy controls. Statistical analyses were conducted using one-way ANOVA. Pam3CSK4, Poly I:C, and CpG ODN were used as TLR2, TLR3, and TLR9 agonists, respectively.