目的 探讨1,25-二羟维生素D,[1,25(OH)2D3]处理的树突细胞(DC)在变应性气道炎症中的免疫调节作用及其机制.方法 小鼠骨髓来源DC分2组,分别用1,25(OH):D,和磷酸盐缓冲液(PBS)处理,RT-PCR和蛋白质印迹法检测2组Dc Notch配体Mrna和蛋白表达.将2组DC和Notch配体中和抗体封闭的1,25(OH)2D3处理DC分别与小鼠脾脏来源CIM+T细胞共培养,流式细胞仪检测CD4+T细胞中CD4+CD25+Foxp3+T细胞百分比.将卵清蛋白(OVA)致敏小鼠分2组,每组5只,分别过继转移1,25(OH)2D3组DC[1,25(OH)2D3-DC组]和PBS组DC(PBS-DC组),以OVA激发气道炎症后行肺脏病理、支气管肺泡灌洗液(BALF)中自细胞介素4(IL-4)、IL-5、IL-13和干扰素γ(IFN-γ)水平以及脾脏CD4+T细胞中CD4+CD25+Foxp3+T细胞百分比检查.结果 1,25(OH)2D3组DC Notch配体Jaggedl和Jagged2 Mrna表达(0.376±0.029、0.564±0.018)和蛋白表达(0.786±0.034、0.632±0.026)均明显高于PBS组(分别为0.146±0.032、0.267±0.012和0.124±0.025、0.098±0.012,均P＜0.01).与CIM+T细胞共培养后,1,25(OH)2D3组CD4+T细胞中CD4+C1)25+Foxp3+T细胞百分比(22.49%±0.56%)明显高于PBS组(6.67%±0.60%,P＜0.01);经Jagged2中和抗体封闭组CIM4CD25+Foxp3+T细胞百分比(6.56%±1.89%)明显低于未封闭组(20.37%±1.64%,P＜0.01),1,25(OH)2D3-DC组小鼠气道炎症明显轻于PBS-DC组;BALF中IL-4、IL-5、IL-13和IFN-γ水平(pg/ml)分别为33±5、134±23、91±11和未检测到(＜12.5),均明显低于PBS-DC组(分别为55±7、332±49、152±19、23±6,均P＜0.01);脾脏CD4+T细胞中CIM+CD25+Foxp3+T细胞百分比(14.69%±1.14%)明显高于PBS-DC组(2.38%±0.14%,P＜0.01).结论 1,25(OH)2D3处理DC对变应性气道炎症有抑制作用,这种作用可能与1,25(OH)2D3处理DC通过Jaggect2介导的Notch信号途径诱导CD4+CD25+Foxp3+T细胞生成有关.%Objective To explore the mechanism and immunoregulatory role of 1,25-dihydroxy vitamin D3[1,25(OH)2D3]-treated dendritic cells(DCs)in allergic airway inflammation.Methods Mouse bone maiTow-derived DCs were treated by 1,25(OH)2D3 for72 h.The expression levels ofdifferent Notch ligands:Jaggedl,Jtmoged2,Deltal,Delta3,and Delta4 in these DCs were detected by RT-PCR and westem blotting.Mouse spleen CIM+T cells were cultured with 1,25(OH)2D3-treated DCs or 1,25(OH)2D3-treated DCs Mocked by polyclone antibody of Jaggedl or Jagged2(control group)for48 h.The pementage of CIM4CD25+Foxp3+T cells in CD4+T cells was detected by flow cytometry(FCM).Ten mice underwent intraperitoneal injection of ovalbumin(OVA)to be sensitized and tIIen divided into 2 groups to inhale 1,25(OH)2 D3-treated DC suspension and PBS-DC suspension respectively for 6 successive days.Then the micc were killed.Bronchoalveolar lavage fluid was obtained to detect tlle eosinophil count and the levels ofinterleukin(IL)-4,IL-6,IL-13,and interferon(IFN)-γ,and pathological examination of lung was conducted.Spleens were taken out to isolate the CD4+T cells,and immunolabeling and FCM were used to detect the percentage of CD4+CD25+Foxp3+T ceils.Results The mRNA and protein expression levels of Jaggedl and Jagged2 in the 1,25(OH)2D3-treated DCs were(0.376±0.029)and(0.786±0.034),and (0.564±0.018)and(0.632±0.026)respectively,all significantly higher than those of the control group [(0.146.±0.032)and(0.124±0.025),and(0.267±0.012)and(0.098±0.012)respectively,all P＜0.01)].The percentage of CD4+CD25+Foxp3+T cells in the CD4+T cells cultured with 1,25 (OH)2D3-treated DCs was(22.49%±0.56%),significantly higher than that of the a PBS control group [(6.67%±0.50%),P＜0.01].The percentage of CD4+cD25+Foxp3+T cells in CD4+T cells after cultured with 1,25(OH)2D3.treated DC blocked by polyclone antibody of Jagged2 was(6.56%±1.89%),significantly lower than that of the un.blocked control group[(20.37%±1.64%),P＜0.01].The levels of IL-4,IL-5,IL-13,and IFN-γ,and eosinophil count in the BALF of the 1,25(OH)2D3-treated DC group were(33±5)pg/ml,(134±23)pg/ml,(91±11)pg/ml,and undeteetable(＜12.5 pg/m1),and(236±29)×103/ml.all significantly lower than those of the PBS-Dc group[(55±7)pg/ml,(332±49)pg/ml,(152±19)pg/ml,and(23±6)pg/ml,and(588±56)×103/ml,all P＜0.01].The percentage of CD4+CD25+Foxp3+T cells in the spleens of the 1,25(OH)2D3-treated DC group was(14.69%±1.14%),signifieantly higher than that of the PBS-treated DC group[(2.38%±0.14%,P＜0.01).Conclusion Treatment of the DCs with 1,25(OH)2D3 inhibits the allersic inflammation in tlle airway,maybe via the induction of CD4+CD25+Foxp3+regulatory T cells by 1,25(OH)2D3-treated DCs through Jagged2-mediated Notch signal pathway.
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