The Role Played By Tregs In The Inhibitory Effect On Asthma By DCs Of Schistosoma Japonicum-infected Mice

Objective To examine whether the DCs of Schistosoma japonicum (SJ)-infected mice develop the inhibitory effect on asthma through inducing Tregs by using an adoptive transfer approach. Methods DCs from SJ-infected mice and naive mice (designated as SJDCs and NDCs) were respectively isolated and purified using CDllc+ microbeads. Some of the freshly isolated DCs were cultured with complete medium using 96-well culture plates at 5×105cells/well in the presence of 0.1 mg/ml OVA for 72 h. The levels of IL-12 and IL-10 in culture supernatants were measured by ELISA. In addition, some of the freshly isolated DCs were used for adoptive transfer. Concretely, fifteen BALB/c mice were divided into three groups randomly.106 SJDCs were adoptively transferred to each mouse in Group A by tail vein, while 106 NDCs were adoptively transferred to each mouse in Group B in the same way. The mice in Group C were without DCs transfer.2 hours later, mice in Group A, B and C were sensitized and challenged with OVA to induce asthma.4 weeks later, all the mice were killed. The single spleen cell suspensions were collected for analyzing the level of CD4+CD25+Foxp3+regulatory T cells (Foxp3+Tregs), CD4+CD25+IL-10+ regulatory T cells (IL-10+Tregs) and the intracellular expression of IL-4, IL-5 and IFN-y; Bronchoalveolar lavage fluids (BALF) were collected for eosinophils cell counting; Lungs were collected for histological and immunohistochemical analysis. Results The levels of IL-10 in SJDCs and NDCs culture supernatants were (64.80±3.6050)ng/ml, (43.45±0.7720) ng/ml; the levels of IL-12 in SJDCs and NDCs culture supernatants were (5.88±0.0429)ng/ml, (5.91±0.0330)ng/ml respectively. SJDCs produced significantly higher levels of IL-10 than NDCs (P＜0.001), while the levels of IL-12 were comparable between SJDCs and NDCs (P>0.05); For the mice in Group A, B and C, the percentage of foxp3+Tregs in CD4+T cells were (8.44±0.3209)%, (4.34±0.6465)%, (3.62±1.2834)%respectively; the percentage of IL-10+Tregs in CD4+T cells were (4.86±0.8112)%, (2.80±0.6205)%, (2.30±0.9138)% respectively; the percentage of IL-4+CD4+T cells in CD4+T cells were (3.54±0.8444)%, (5.68±0.7328)%, (5.96±1.992)% respectively; the percentage of IL-5+CD4+T cells in CD4+T cells were (4.20±0.6745)%, (6.70±0.6285)%, (6.82±1.9955)% respectively; the percentage of IFN-γ+CD4+T cells in CD4+T cells were (7.82±0.9550)%, (7.06±2.1733)%, (7.98±3.9392)% respectively. Compared with mice in Group B and C, the levels of foxp3+Tregs and IL-10+Tregs were increased significantly (P＜0.01, P＜0.001) in Group A, the levels of IL-4+CD4+and IL-5+CD4+T cells decreased significantly in Group A (P<0.01), whereas the levels of IFN-y+CD4+T cells were comparable (P>0.05). Moreover, the number of eosinophils (104) in BALF for the mice in Group A, B and C were 6.083±1.4043,26.100±4.7647, 35.733±7.3730 respectively. Compared with mice in Group B and C, the numbers of eosinophils in Group A decreased significantly (P<0.05). Immunohistochemical analysis showed that, for the mice in Group A, B and C, the percentage of the positive area of CCL-11 in the blood vessels and tracheas in lung tissue were successively 0.2737±0.0218,0.4818±0.0199,0.5339±0.0322; the average optical density (AOD) of the positive area of CCL-11 in the blood vessels and tracheas in lung tissue were successively 0.2546±0.0172,0.3160±0.0235,0.3874±0.0236. Compared with the mice in Group B and C, AOD and the percentage of the positive area of CCL-11 in Group A significantly decreased (P＜0.05). Histological analysis showed that pulmonary inflammation of the mice in Group A diminished markedly, compared with the mice in Group B and C (P<0.05). Conclusion DCs of helminth-infected mice played a critical role in inhibition of allergic asthma through enhancing Tregs responses.