| Food allergies are a growing global health concern and can lead to serious health complications Iike anaphylaxis,food-protein-induced enterocolitis syndrome,food-induced proctocolitis and atopic dermatitis.Probiotic supplementation has been used as a preventive or therapeutic strategy for allergic disorders.However,the inmmune mechanism was needed to clarify urgently.In this study,the effects of probiotics on Th1/Th2 balance and the number of regulatory T(Treg)cells in murine cow's milk allergy were analyzed.The shift in dendritic cells(DCs)subsets and their maturation status in insestinal tissues and spleen were further investigated.Moreover,we clarified the cellular and molecular pathways in probiotic-specific modulation of DCs.These results provided the first evidence of the signalling mechanisms of probiotics's effect on cow's milk allergy.The main results of this study are as followed:The benefical effects of Lactobacillus paracasei L9(L9)on allergic reactions to cow' milk allergen in mice was assessed.The results showed that L9 significantly decreased serum levels of histamine,mouse mast cell protease-1(mMCP-1)and total IgE by 25.4%,31.9%and 27.6%receptively.The Th2 related cytokine IL-4 level in splenocyte and serum was reduced by 30.6%and 36.6%.Thl related cytokine IFN-? production was increased by L9 supplement(p<0.05).The results indicated that L9 treatment prevented allergic sensitization of cow' milk allergen through shifting Th2-dominated trends to Thl-dominated responses.Treg cells can regulate Th1/Th2 balance by cytokines production,such as TGF-? and IL-10.In this study,lymphocytes from the spleen,MLN and PP were isolated and cultured in vitro.The results showed that L9 significantly induced the number of Foxp3+Treg and IL-10+Treg in spleen,MLN and PP.The increament of these two kinds of Tregs were more than 50%in MLN.Besides,L9 increased the levels of TGF-? and IL-10 in serum and lymphocytes culture supernatants(p<0.05).Compared with BLG group,the IL-4 level in lymphocytes of PP,MLN and spleen in L9 group was decreased by 45.8%,47.2%and 32.6%,respectively.The IFN-? production was also induced by L9 supplement(p<0.05).The results suggested that The L9 supplement suppressed the aberrant balance of Th1/Th2 responses in mice,via upregulation of the Foxp3+Treg and IL-10+Treg responses.To evaluate the possible involvement of DCs in L9-induced Treg cells,the number of DCs subsets in different groups was analyzed.CD103+DC and CX3CR1+DC were isolated from intestine tiusse of L9-treated mice,and co-cultured with CD4+T from allergic mice.The results showed that the percentage of CD103+DC in PP,MLN and spleen from L9 treated mice were increased by 44.7%?34.7%and 26.4%,while,the percentage of CX3CR1+DC in PP,MLN and spleen were improved by 62.7%,1 18.4%and 28.9%respectively.Moreover,L9 induced production of TGF-? and IL-10 in CD103+DC,which increased the number of Foxp3+Treg and IL-10+Treg in vitro byl 18.1%and 56.4%respectively.L9 also induced IL-10 level in CX3CRI+DC,which improved the percentage of IL-10+Treg in vitro by 108.4%.The results demonstrated that L9-modulated CD103+DC and CX3CR1+DC induced Treg cells.To clarify the cellular and molecular pathways in L9-specific modulation of DCs,bone marrow-derived dendritic cells(BMDCs)was stimulated by L9 in vitro.The results showed that the L9-induced CD103 and CX3CR1 expression in BMDCs was associated with increased toll-like receptor 2(TLR2)expression and mitogen-activated protein kinase(MAPK)phosphorylation.However,TLR2 siRNA transfection and inhibitors of MAPK pathways reduced the expressions of CD 103 and CX3CRland their immunosuppressive cytokines(TGF-? and IL-10)in L9-stimulated BMDCs.Cumulatively,our data demonstrated L9-induece CD103+DC and CX3CR1+DC differentiation through TLR2-MAPK pathways.Taken together,the present work indicated that L9-modulated CD103+DC and CX3CR1+DC suppressed murine allergic responses to BLG trough TLR2-MAPK pathways. |
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