To Explore The Inhibitory Effect Of Dendritic Cells(DC)from Mice Infected With S.japanicun On Allergic Asthma And Their Mechanism Actions By Regulating IL-13 Production

Objective: To explore the underlying mechanism that adoptive transfer of dendritic cells(DCs)in mice infected by Schistosoma japonicum(SJ)can regulate the expression of interleukin-13(IL-13)to inhibit mucus production in allergic asthma.Methods:First,10 normal control mice and 10 SJ infected mice were used to separate DCs.The mice in the experiment were divided into 4 groups(6 mice/per group): control group,SJ group,OVA group,and.SJ +OVA group.1.SJ infected mice model in vivo was established.We extracted DCs from spleen in control mice and SJ infected mice separately by MACS and adoptive transferred DCs to BALB/C mice through caudal vein.Allergic asthma was induced by the OVA.Mice were executed after 4 weeks.2.The lung of the mice was put in to 4% PFA to make frozen sections.Histopathological analysis was performed on O.C.T.(optimal cutting temperature)embedded sections by HE staining and PAS staining.3.Total RNA was isolated from the lung tissue and cDNA synthesis was performed.Then,mRNA levels of IL-13 and related cytokines were analyzed by qRT-PCR4.The spleen cells were extracted and cultured in 37 ? incubator.The protein level of IL-13 in the supernatant fluid was detected by ELISA.5.The femur of mice was gotten after executed to collect DCs.Prepare a single spleen cell suspension from the mice spleen and extract CD4~+ T cells with MS column.The DCs and CD4~+ T cells were co-cultured,the total RNA of which was isolated.Next,mRNA levels of IL-13 was analyzed by qRT-PCR.Results:1.In the in vivo experiment,we observed simple OVA and SJ + OVA-induced asthma group had inflammatory cell infiltration,indicating asthma was successfully induced in OVA and SJ + OVA group.Compared with the simple OVA-induced asthma group,the bronchial wall of the SJ + OVA group was intact,the infiltration of pulmonary interstitial inflammatory cells was reduced,and the bronchial wall thickening was not obvious.The production of mucous fluid in the lumen also decreased.It suggest that adoptive transfer of DCs in mice infected by SJ could inhibit inflammatory cell infiltration and mucus production.2.We analyzed the level of IL-13 and mucin in the adoptive transfer group and simple asthma group.Compared with the healthy control group and SJ group,the expression levels of IL-13 and mucin protein in SJ + OVA group were significantly higher than those in the control group and SJ group.However,the gene expression levels of IL-13 and mucin in SJ + OVA group were significantly inhibited compared to OVA-induced group(P <0.05).The method of immunofluorescence staining was used to detect IL-13 and airway mucin MUC(MUC for mucin,we determined one of the important mucin-MUC5AC).The results showed that compared with simple OVA group,IL-13 and airway mucin expression was inhibited in SJ DCs adoptive transfer group.3.In vitro experiments,use SEA to stimulate cells(simulated SJ infection in vivo stimulation).The results showed that LPS stimulation group compared with SEA stimulation group,IL-13 mRNA expression increased,but was too low relative to the expression of GAPDH,suggesting that DCs was not the most important inducing cells of IL-13.Immunofluorescence staining was used to further identify IL-13 expression cells(lung tissue in OVA simple asthma group).IL-13 and CD4~+ T cells had co-expression region,suggesting that CD4~+ T cells in this model was the main expression cell of IL-13.4.The levels of interleukin-13 in spleen cell culture supernatant showed that the levels of interleukin-13 in OVA group and SJ + OVA group were significantly higher than those in control group.Compared with OVA group,the level of interleukin-13 in SJ + OVA group was significantly inhibited,and there was significant difference,which was consistent with the result of gene expression test.5.DCs treated by SEA was co-cultured with CD4~+ T cells in mice induced by OVA.The vitro cell co-culture results showed that the expression of interleukin-13 was inhibited in SEA treated group compared with the control group(untreated DCs)and LPS stimulation group at 48 h and 72 h after co-culture,suggesting that DCsstreated with SEA could inhibit the expression of interleukin-13 of CD4~+ T cells Conclusion:1.Adoptive transfer of SJDCs not only inhibit the inflammatory cell infiltration but also has a certain inhibitory effect on mucus secretion.2.DCs are not the most important inducing cells of interleukin-13,However,the main expression cells of interleukin-13 are CD4~+ T cells.3.SEA-treated DCs can inhibit the expression of interleukin-13 in CD4~+ T cells.