The effect of cigarette smoke extract on the differentiation of CD4^＋ T cells into Th17 cells and regulatory Tcells

Objective To evaluate the effect of cigarette smoke extract （CSE） on the differentiationof CD4^＋ T cells into CD4^＋ IL-17 ＋ T（Th17） cells and CD4^＋ CD25 ^＋ CFoxp^3＋ regulatory T（Treg） cells stimulation in vitro, and to provide experimental basis for exploring the chronic airway inflammation caused by cigarette smoke exposure. Methods Blood samples were obtained from 11 healthy volunteers, （5 male,6 female）. CD4^＋ T cells were purified by immune magnetic beads and cultured in 96-well plates. The experiments comprised of 8 groups..a blank control group, a T cell stimulating agent （anti-CD3/28） group,a CI）3/28 and CSE group, a cytokines group, a cytokines and CSE group, an aryl hydrocarbon receptor （AHR） agonist （FICZ） group, an AHR antagonist （resveratrol） group, and a solvent control group. （;ells were collected at the 5th day, and flow cytometry was used to evaluate the proportions of Th17 and Treg cells. Results @The effect of CSE on the differentiation of CD4^＋T cells into Thl7 cells the ratio of Thl7 cells in the blank control group was （0.69＋ 0.12）±,and after the addition of T cell stimulating agent the ratio increased to （1.32 ＋ 0.12）%, while CSE further increased the ratio of Th17 cells to （2.17＋0.24）± ,（ t =3.21, P 〈0.01）. Similarly,in the cytokines group,the ratio of Th17 cells was （1.35＋0.08） %,and the addition of CSE,increased the ratio to （2.58±0.39）%（ t =3. 13, P 〈 0.01）. （2）The effect of CSE on the differentiation of CD4^＋ T cells into Treg cells In the blank control group,Treg cells accounted for （0.21±0.19）% of the CD4^＋ T cells,and increased to （3.59± 0.37）% after T cell stimulation. But in the cytokines group, the ratio of Treg cells was dramatically increased （5.85± 0.76） % compared with the T cell stimulating agent group,however,after the addition of CSE, Treg cells markedly decreased （3.07＋0.33）% （t =3.74, P〈0.01）. Likewise,CSE also reduced the ratio of Treg cells （2.19±0.19）% （t =2.71, P 〈0.05） after T cell stimulating agents. （3）AHR activation on CD4^＋ T cell differentiatieon into Treg cells and Th17 cells： In the AHR antagonist （FICZ） group, as compared with the cytokine control group, the ratio of Treg cells remarkably decreased [ （2.60±0.40）%,（5.85± 0.76）% （ t =4.18, P 〈0.01） ,but on the contrary,the ratio of Th17 ceils increased [（2.86± 0.43）.%, （1.35 ± 0.08）%] （t = ,3.65, P 〈0.01）. However, in the AHR antagonist （resveratrol） group, the ratio of Th17 and Treg cells were significantly decreased compared to the cytokine control group [（0.42±0.07）%,（1.35±0.08）% （t=7.71, P〈0.01）,[（0.33±0.14）%, （ 5.85 ± 0.76 ） % ] （ t = 8.87, P 〈0.001 ）, suggesting that probably the proliferation and differentiation uf CD4 ^＋ T cell were inhibited by resveratrol. Conclusions The results showed that CSE increased Th17 cell but decreased Treg cell differentiation from CD4^＋T cells in vitro,possibley through the AHR pathway. Key words: Cigarette smoke extract ;  Th17 ;  Regulatory T cells Aryl hydrocarbon receptor ;  CD4 ^＋ T lymphocytes