The Mechanism Of The Schistosoma Japonicum Infected Dendritic Cells Inhibit Allergic Reaction

Objective To study the inhibitory effect on allergic asthma of dendritic cells(DC)from Schistosoma japonicum-infected mice.Methods In the first part, CD8α+CD11c+and CD8α- CD11c+ DCs from SJ/SEA mice and na?ve mice were respectively isolated and purified using CD8α+ and CD11c+ microbeads. Firstly, we detected cytokine expressed in these DC subsets by using qRT-PCR. And then 5 ×105 DC subset cells of SJ/SEA infected mice were adoptively transferred to each mouse by tail vein and induced asthma with OVA after 1h. We observed immunol response in lung and collected serum for measurement of OVA-specific IgE; Bronchoalveolar lavage fluids(BALF) for measurement of various cytokines(IL-4, IL-5, IL-10, IL-12 and TGF-β);Spleen cells culture supernatant were collected for determination of IL-4, IL-5, IL-10,IL-12 and TGF-β. In the second part, we studied how SJDC can induce differentiation of CD4+T cells by cultured DC in vitro. We cultured bone marrow DC and stimulated them by soluble egg antigen(SEA) for 18 hours, and used DC stimulated by LPS as positive control, DC stimulated by medium as negative control. We detected the surface marker and cytokine of DC by FASC, IHC and qRT-PCR. And then we co-cultured DC with CD4+T cells from OVA infected mice for 48 h and 72 h and detected cytokine secreted by CD4+T cells using qRT-PCR. In the third part, we found the mechanism of SJ infected mice inhibit airway mucus overproduction from IL-13 point. We infected mice with SJ/SEA and induces asthma with OVA. We observed immunol response and mucus in lung by HE and PAS staining and detected MUC5 ACmRNA and IL-13 mRNA expressed in lung by qRT-PCR. We also examined the expression area of IL-13 and CD4+T in lung by IHC. Meanwhile, We cultured DC in vitro and detected IL-13 mRNA by qRT-PCR after stimulated by SEA. And then co-cultured them with CD4+T cells and detected IL-13 mRNA by qRT-PCR of CD4+T cells. Results In the first part we found CD11 bmRNA in CD8α-DC was increased than CD8α+DC both in SJ/SEA infected mice and non-infected mice by qRT-PCR(P< 0.05). And the CD80 mRNA and CD86 mRNA in SJCD8α-DC was decreased compared with SJCD8α+DC(P<0.05), IL-10 mRNA was increased(P<0.05), on the contrary, TGF-β mRNA(P<0.001), IL-12 mRNA was decreased(P<0.05). Thepulmonary inflammation of mice in adoptively transferred SJ/SEA DC Group was diminished markedly(P<0.005). Moreover, the levels of OVA-specific IgE in serum was also decreased(P<0.05). IL-4, IL-5 in BALF or Spleen culture supernatant of mice in adoptively transferred SJ/SEA DC Group was decreased significantly(P<0.05), while the levels of IL-10 、 IL-12 、 TGF-β in BALF or Spleen culture supernatant of mice in adoptively transferred SJ/SEA DC Group was increased significantly(P<0.05). In the second part compared with DC stimulated by LPS using FACS, the surface marker like CD80, CD86 and CD40 of SEADC was decreased(P<0.05). The result of IHC shown that CD11 c expressed on SEADC was decrease compared with LPSDC. And the CD80 mRNA, CD86 mRNA of SEADC was decrease(P<0.05), on the contrary, IL-1βm RNA, IL-10 mRNA, IL-12 mRNA, IL-6mRNA, IL-23 mRNA, TGF-βmRNA was increase compared with MediumDC by qRT-PCR(P<0.05). Forthmore, SEADC could inhibited OVA CD4+T cell to express IFN-γmRNA(P<0.05)and IL-17mRNA(P<0.05), but could induced them to express more IL-4mRNA and IL-10mRNA(P<0.05). In the third part, the HE and PAS staining results shown that the secretion of mucus in SJ/SEA+OVA group was less than OVA mice. And the expression of IL-13mRNA( P < 0.05) and MUC5ACmRNA( P < 0.01) in SJ+OVA group was less than OVA group by qRT-PCR. Furthermore, the expression area of IL-13 was decreased by IHC and the expression of IL-13 was decreased both in mRNA level and protein level(P<0.05).Forthmore, We cultured bone marrow DC in vitro and stimulated them with SEA, the result found IL-13 was increase compared with MediumDC by qRT-PCR(P<0.05).SEADC could inhibited OVA CD4+T cell to express Il-13mRNA(P<0.05) by co-cultured them with CD4+T from OVA induced mice. Conclusion SEADC maybe immature and could induced CD4+T cells differentiate to regulatory T cells(Tregs).And CD8α-CD11c+ are tolerance DC, they could secrete IL-10 to induce Tregs and inhibit asthma. Furthermore, SJ infected mice could inhibit CD4+T to secret IL-13.