【摘要】 探索小鼠骨髓源性肥大细胞能否在体外诱生CD4+CD25+Foxp3+调节性T细胞。从小鼠股骨获取骨髓细胞,加入完全RPMI 1640培养基(含IL-3和SCF各10 ng/ml)诱导4周。用甲苯胺蓝染色法观察肥大细胞异染颗粒;流式检测CD117和FcεRIα双阳性细胞比例。将肥大细胞与同基因来源的T细胞按不同比例(1∶2、1∶1、2∶1)置于48孔培养板中培养,作为三个实验组;未加肥大细胞的单独T细胞组作对照组。四个组均加入CD3抗体和CD28抗体各2μg/ml,IL-2 1 000U/ml,5 d后流式检测Foxp3表达情况。RT-PCR,免疫组化法检测肥大细胞中TGF-β1的表达。与对照组相比,实验组Foxp3表达均升高(对照组:3.37%±0.40%;实验组为:8.23%±0.80%、10.87%±1.25%、13.63%±0.55%)。RT-PCR和免疫组化均检出TGF-β1表达。肥大细胞能在体外诱导T细胞转化为CD4+CD25+Foxp3+调节性T细胞,可能与肥大细胞表达TGF-β1有关。更多还原

【Abstract】 To detect whether mouse mast cells can induce CD4+CD25+Foxp3+regulatory T cells(Treg) in vitro.Bone marrow cells obtained from C57BL/6(H-2b) mice were cultured with IL-3(10 ng/ml) and SCF(10 ng/ml) for 4 weeks.The purity of bone marrow mast cells(BMMCs) was tested by flow cytometry.The expression of TGF-β1 in BMMCs was tested by RT-PCR and immunohistochemistry.Then the BMMCs were co-cultured with T cells of C57BL/6 mice at 1∶2,1∶1 and 2∶1 ratios in the presence of anti-CD3 antibody(2 μg/ml),anti-CD28 antibody(2 μg/ml) and IL-2(1 000 U/ml).The percentages of CD4+CD25+Foxp3+ T cells in the co-cultured system were compared among different groups by flow cytometry on day 5.It was found that the percentages of CD4+CD25+Foxp3+ cells were significantly higher in the ratio of BMMC/T 2∶1 group(13.63% ± 0.55%),the 1∶1 group(10.87% ± 1.25%)and 1∶2 group(8.23% ± 0.80%) than in the control group(3.37% ± 0.40%).The expression of TGF-β1 was determined in the mouse BMMCs by RT-PCR and immunohistochemistry.Our findings indicate that CD4+ T cells can be induced to become CD4+CD25+Foxp3+ Treg by BMMCs in vitro and the secretion of TGF-β1 by BMMCs might be involved in this process.更多还原