Anthriscus sylvestris-derived extract induces Th1 and Th17 cell differentiation via the upregulation of IL12 and IL23 production

Figures & data

Figure 1. AS-derived extract induces the expression of MHC class II, IL12, and IL23 in macrophages rather than peripheral DCs.Note: DO11.10 Balb/c mice were intraperitoneally injected with OVA peptide_323–339 (100 µg) plus either vehicle or AS extract (4 µg) three times a week for two weeks. Splenocytes were prepared from both vehicle- or AS extract-treated mice after immunization. Both the MHC class II cell surface and intracellular IL12p40/IL23p19 production levels were measured on macrophages (gated on F4/80⁺CD11b⁺CD11c^- population) or DCs (gated on CD11c⁺ population) by flow cytometry. Data are shown as the means ± SD (n = 3; *P ≤ 0.05; **P ≤ 0.01).

Figure 2. In vivo treatment with AS extracts induces Th1 and Th17 differentiation.Note: After two weeks of immunization, purified splenic CD4⁺ T cells were activated with plate-bound anti-CD3 (10 µg/ml) and anti-CD28 (1 µg/ml) mAbs for 16 hrs and subsequently restimulated with PMA (50 ng/ml) and ionomycin (1 µg/ml) in the presence of brefeldin A (10 µg/ml) for an additional 2 hrs. (A) The production levels of intracellular IFNγ and IL4 by CD4⁺ T cells gated on CD4⁺KJ1-26⁺ population were measured by flow cytometry. (B) The intracellular IL17 production level by CD4⁺KJ1-26⁺ T cells was assessed by flow cytometric analysis. (C) The mRNA levels of T helper cell transcription factors in purified splenic CD4⁺ T cells from either vehicle- or AS extract-treated mice were analyzed by RT-PCR. Data are shown as the mean ± SD (n = 3; *P ≤ 0.05; **P ≤ 0.01).

Figure 3. In vivo treatment with AS extracts upregulates IL17 production by NKT cells and neutrophils.Note: After two weeks of immunization, the intracellular expression levels of IFNγ (A) and IL17 (B) by splenocytes from vehicle- or AS extract-treated mice were measured by flow cytometry gated on DX5⁺TCRβ⁻ (NK cells), DX5⁺TCRβ⁺ (NKT cells), or non-B/non-T (NBNT) Gr-1⁺CD11b⁺ (neutrophils) populations. Data are shown as the means ± SD (n = 3; *P ≤ 0.05; **P ≤ 0.01).

Figure 4. Addition of AS extracts results in a dramatic reduction in the generation of OVA-specific Foxp3⁺ Treg cells.Note: After two weeks of immunization, total splenocytes from either vehicle- or AS extract-treated mice were prepared and permeabilized for intracellular Foxp3 staining. The intracellular expression of Foxp3 was analyzed by flow cytometry gated on CD4⁺CD25⁺KJ1-26⁺ population. Data are shown as the means ± SD (n = 3; *P ≤ 0.05).